Transplantation, 1992 Jul, 54(1), 32 - 7 The role of ultraviolet B-irradiated leukocyte transfusions and cyclosporine in intestinal transplantation; Xiao F et al.; To explore the efficacy of ultraviolet B-irradiated donor-specific leukocyte transfusions (UV-DSLT) with short-term cyclosporine to control intestinal allograft rejection, 75 adult Lewis (RT1l) rats underwent total small-intestinal transplantation from Brown-Norway (RT1n) donors . Recipients were randomly divided into ten treatment and control groups utilizing various combinations of donor-specific and third-party (Wistar-Furth, RT1u) leukocyte transfusions (TPLT), with or without transfusion UVB irradiation, and either alone or in combination with short-term cyclosporine administration (5 mg/kg intramuscularly on days -7, 0, 1, and 2 relative to transplantation) . Leukocytes (10(8) cells) separated from a spleen cell suspension were infused on day -7 . Certain transfused leukocytes were treated with 12,000 joules/m2 of UVB irradiation . Groups were monitored for mean survival time (MST) and cause of death . UV-DSLT alone (MST = 19.8 +/- 4.6) or in combination with cyclosporine (UV-DSLT+CsA, MST = 53.1 +/- 22.5) significantly (P less than 0.003-0.0002, Mantel-Cox) prolonged recipient survival when compared with appropriate controls (i.e., no treatment, MST = 11.2 +/- 3.4; CsA, MST = 17.2 +/- 9.0; UV-TPLT, MST = 12.4 +/- 4.0; and UV-TPLT+CsA, MST = 25.1 +/- 9.7) No significant increase in graft-versus-host disease occurred in any group, with 85% (64/75) of the recipients dying of acute rejection . Conversely, the UV-DSLT+CsA group had a significant increase (9/11; chi-square, P less than 0.0001) in chronic rejection . Because UV-DSLT+CsA improved survival as compared with third-party controls, a limited donor-specific unresponsiveness may have been induced . Furthermore, this treatment produces a consistent, chronic rejection rodent intestinal allograft model. Eur J Biochem, 1992 Jul 1, 207(1), 117 - 23 Malonate decarboxylase of Malonomonas rubra, a novel type of biotin-containing acetyl enzyme; Hilbi H et al.; Cell suspensions or crude extracts of Malonomonas rubra grown anaerobically on malonate catalyze the decarboxylation of this substrate at a rate of 1.7-2.5 mumol.min-1.mg protein-1 which is consistent with the malonate degradation rate during growth . After fractionation of the cell extract by ultracentrifugation, neither the soluble nor the particulate fraction alone catalyzed the decarboxylation of malonate, but on recombination of the two fractions 87% of the activity of the unfractionated extract was restored . The decarboxylation pathway did not involve the intermediate formation of malonyl-CoA, but decarboxylation proceeded directly with free malonate . The catalytic activity of the enzyme was completely abolished on incubation with hydroxylamine or NaSCN . Approximately 50-65% of the original decarboxylase activity was restored by incubation of the extract with ATP in the presence of acetate, and the extent of reactivation increased after incubation with dithioerythritol . Reactivation of the enzyme was also obtained by chemical acetylation with acetic anhydride . These results indicate modification of the decarboxylase by deacetylation leading to inactivation and by acetylation of the inactivated enzyme specimens leading to reactivation . It is suggested that the catalytic mechanism involves exchange of the enzyme-bound acetyl residues by malonyl residues and subsequent decarboxylation releasing CO2 and regenerating the acetyl-enzyme . The decarboxylase was inhibited by avidin but not by an avidin-biotin complex indicating that biotin is involved in catalysis . A single biotin-containing 120-kDa polypeptide was present in the extract and is a likely component of malonate decarboxylase. Gan To Kagaku Ryoho, 1992 Jul, 19(7), 981 - 6 {Combination chemotherapy of solid tumor--effects of hyaluronidase on doxorubicin (DXR) penetration into multicellular tumor spheroids (MTS)}; Kohno N et al.; We have studied the effects of hyaluronidase (HYD) on the penetration and cell kill effect of doxorubicin (DXR) using multicellular tumor spheroids (MTS) . MTS of approximately 500 microns in diameter were prepared by liquid over lay culture technique from PC-10 lung and HEp-2 laryngeal squamous carcinoma cell lines . Cells in MTS and monolayer were exposed for various durations to HYD, followed by 1 hr, rest interval, and by 1 hr . exposure to DXR . MTS and monolayer cells were then trypsinized to a single cell suspension and subjected to clonogenic assay . For PC-10 MTS, pretreatment with HYD for 24 hr . resulted in approximately 10-fold increases in DXR cell kill effects as compared to DXR alone . HEp-2 MTS were more sensitive to HYD pretreatment . Thus, 1 hr . exposure to HYD produced approximately 4-fold increases in DXR-induced cell lethality . Fluorescent microscopic study revealed that 1hr . exposure of MTS to DXR produced DXR fluorescence only 1-2 outer layer of MTS . When MTS were pretreated with HYD, there was an enhanced penetration of DXR fluorescence into the MTS core . HYD-induced enhancement of DXR penetration and its cell kill effect was dependent on the exposure time and tumor cell origin. Exp Neurol, 1992 Jul, 117(1), 59 - 70 Fate of B1-B2 and B3 rhombencephalic cells transplanted into the transected spinal cord of adult rats: light and electron microscopic studies; Rajaofetra N et al.; Embryonic cell suspensions (14-day embryos) containing either B3 or B1-B2 serotonergic cell groups were obtained by microdissection of specific rhombencephalic regions and transplanted into the transected spinal cord of adult male Sprague-Dawley rats . After 3 months of survival, the animals were sacrificed and the spinal cords processed for the immunocytochemical detection of serotonin (5-HT) . 5-HT-immunoreactive fibers from B1-B2-grafted cells were selectively distributed in the ventral horn and the intermediolateral cell column (IML) where they established conventional synaptic contacts . However, B3 5-HT cells grew and extended their processes into the dorsal horn where in addition we observed scarce synaptic contacts as in the normal spinal cord . These results suggest that the specificity of the 5-HT innervation of the spinal cord by grafted neurons is due, at least partly, to the presence of local mechanisms mediating guidance and cell recognition, possibly operating in conjunction with preexisting substrate pathways. Proc Soc Exp Biol Med, 1992 Jul, 200(3), 431 - 41 Stromal cells derived from spleen or bone marrow support the proliferation of rat natural killer cells in long-term culture; Tjota A et al.; Rat nylon wool nonadherent bone marrow cells were propagated for up to 75 days in co-culture with stromal cells derived from either spleen or bone marrow . Interleukin (IL) 1 enhanced the ability of spleen stroma to support the long-term culture of natural killer (NK) cells, ostensibly by inducing these support cells to synthesize other cytokines . Flow cytometry studies indicated that the nylon wool separation procedure enriched the concentrations of mature NK cells from 7.9% to 38.1% for splenocytes and from 3.8% to 19.5% for bone marrow cells . Analyses of the adherent zones of suspended nylon screen NK cell cultures revealed substantial numbers of large granular lymphocytes that expressed NK 323+/MOM/3F12/F2- phenotypes . The presence of both mature and immature cells of the NK lineage in this matrix was inferred by the presence of both IL-2 receptor (IL-2R) positive and IL-2R negative, and OX-8+ and OX-8- NK 323+ cells over the greater than 4-month experimental period . Suspended nylon screen cultures displayed a greater potential for producing cytolytic cells than either co-cultures of bone marrow nonadherent cells on stroma monolayers or suspension cultures . The large granular lymphocytes produced in suspended nylon screen cultures could be transformed into active killers of YAC-1 targets by IL-2 . In contrast to bone marrow nonadherent cells, more splenic nylon-wool-passed cells displayed a mature NK phenotype, but their proliferative potential and ability to be transformed into cytolytic cells by IL-2 decreased rapidly in culture . In the suspended nylon screen culture system, NK cells migrate from the underlying stroma in stages as they mature, retain their cytolytic potential, and manifest a capacity for self-renewal . Cultured cells were routinely dissociated into single cell suspensions via enzyme treatment and were reinoculated onto "fresh" nylon screen/stromal cell templates after passage through nylon wool columns . These co-cultures continued to generate cytolytic cells in numbers greater than those of the initial inoculum. Exp Lung Res, 1992 Jul-Aug, 18(4), 435 - 46 Alveolar and interstitial macrophage populations in the murine lung; Crowell RE et al.; Pulmonary macrophages (PM) exist in two general anatomical compartments in the lower respiratory tract: the alveolar space (alveolar macrophages, AM) and the interstitium (interstitial macrophages, IM) . We determined the relative contribution that macrophages in each of these compartments make to the size of the total PM population found in the lungs of C3H/OUJ mice, while also evaluating how efficiently bronchoalveolar lavage (BAL) removes AM from the murine lung . These objectives were accomplished by combining extensive BAL with subsequent mechanical and enzymatic dissociation of the lungs in conjunction with in situ and in vitro phagocytic assays involving opsonized erythrocytes (EA) to identify mononuclear phagocytes . On average, 2.5 x 10(6) cells were recovered by extensive BAL, and approximately 78% of these cells ingested EA in vitro . To determine the efficiency of BAL in removing PM from the alveolar space, EA were instilled intratracheally into intact lungs, which had been removed from the chest cavity, and allowed to incubate for 60 min; this was followed by exhaustive BAL and subsequent lung digestion . After these procedures, approximately 4% of the dissociated lung cells contained EA, indicating that these cells were alveolar in origin but had not been removed despite extensive BAL . Subtraction of these AM from the total EA+ cells in lung cell suspensions following a second in vitro incubation with EA indicated that approximately 37% of all PM were within the interstitium . These results suggest that, while AM comprise the majority of lung macrophages, IM constitute a larger component of the total PM population in murine lungs than previously reported . In addition, this study, like several previous investigations using other species, indicates that a significant proportion of AM remain in the lung despite attempts to remove them with BAL . Accordingly, residual AM significantly contaminate the IM population present in murine lung cell suspensions even after extensive lavage. J Endocrinol, 1992 Jul, 134(1), 97 - 106 Effects of gonadotrophin-releasing hormone, recombinant human activin-A and sex steroid hormones upon the follicle-stimulating isohormones secreted by rat anterior pituitary cells in culture; Ulloa-Aguirre A et al.; FSH is produced and secreted from the anterior pituitary gland of rats in multiple molecular forms . At times of high gonadotrophin-releasing hormone (GnRH) and oestrogen output (e.g . the morning of the day of pro-oestrus) the pituitary increases the production of FSH isoforms with isoelectric point (pI) values greater than 5.0, whilst sex steroid deprivation leads to the production of strongly acidic and less in-vitro biologically active FSH molecules . It is not known, however, whether sex steroids modulate the production of specific FSH isoforms by a direct action at the pituitary level or indirectly through altering the rate of synthesis and/or secretion of GnRH . In order to obtain some insight on this issue, we examined the charge heterogeneity of FSH secreted by cultured pituitary cells exposed to different FSH-releasing factors, oestradiol-17 beta and progesterone, alone or in different time-sequenced combinations . Anterior pituitary glands from 21-day-old female rats were enzymatically dispersed into a single cell suspension and cultured for 5 days . During days 1 to 3, cells were incubated in the absence of factors or steroid hormones; on days 3 to 4, cells were incubated in the absence (controls) or presence of either oestradiol-17 beta (3.67 nmol/l) or oestradiol-17 beta plus progesterone (3.67 and 31.8 nmol/l respectively) . Finally, during days 4 to 6, GnRH (10 nmol/l) or recombinant human activin-A (2 nmol/l) were added to half of all culture wells . Media from each cell group were concentrated and the several forms of secreted FSH were then separated by polyacrylamide gel isoelectric focusing (pH range 6.5-4.0) and quantitated . All media concentrates contained several forms of immunoactive secreted FSH focusing within a pH range of 6.44-4.23 . A large amount (51-76%) of total FSH recovered focused within a pI range of 4.9-4.0 (area 3), whilst 20-43% and 4-8% of the total were identified within pI range of 5.9-5.0 (area 2) and 6.5-6.0 (area 1) respectively.(ABSTRACT TRUNCATED AT 400 WORDS) Biull Eksp Biol Med, 1992 Jul, 114(8), 187 - 9 {Natural killer activity in man under different experimental conditions}; Cheknev SB; The cytotoxic activity of natural killer (NK) cells against labelled with 3H-uridine target cells of standard human erythromyeloblast line K562 in 63 healthy donors was studied in 14-hr cytotoxic test . The cytotoxic reaction was realized in complete medium supplemented with different types of serum such as foetal calf serum (FCS), autologous or homologous sera in different schemes of incubation . It has been shown that NK cell activity was augmented by 50 per cent (p < 0.05) in the presence of homologous serum added to mononuclear cell suspension for 1 hr at 37 degrees C in comparison with the effect of FCS presence . Thus, the serum of healthy donors contains some factors which not only reflect an individual genotypic information of a donor, but also can be recognized by NK cells and significantly change the cytotoxic NK cell activity . The data obtained exclude possibility of using a homologous serum when positive controls for natural cytotoxic reactions are planned. Chin Med J (Engl), 1992 Jul, 105(7), 544 - 8 Myeloid and erythroid hemopoiesis supported by human bone marrow fibroblasts in vitro; Cao WH et al.; The passages 0 and 3 purified human bone marrow fibroblast layers (FLs) were established by long-term liquid cultures . After 12-day coculture of stromal . cell-depleted marrow cell suspensions with passage 0 FLs, 68.33 +/- 4.04% of the hemopoietic colonies adhered to FLs was myeloid in nature and the other 31.67 +/- 4.04% was erythroid . There were still CFU-E (colony forming unit-erythroid), BFU-E (burst forming unit- erythroid), and CFU-GM (colony forming unit-granulocyte/macrophage) among the nonadherent cells . The media conditioned by passage 0 (F0-CM) and passage 3(F3-CM) FLs stimulated the growth of myeloid and erythroid (BFU-E) colonies . From these data, it is concluded that both FLs and the media conditioned by fibroblasts can stimulate myeloid and erythroid hemopoiesis. Clin Exp Dermatol, 1992 Jul, 17(4), 240 - 5 Flow cytometry analysis of adhesion molecules on human Langerhans cells; Vedel J et al.; The Langerhans cell (LC) migrates between the epidermis and the regional lymph nodes to present antigens . This migration pattern requires the expression of a changing repertoire of cell-surface molecules . In this work, we have investigated the expression of the adhesion molecules CD 11/CD 18 and CD 58 on LCs . Human epidermal cell suspensions were enriched in LCs (mean enrichment 75%) using a two-step technique including a Ficoll-Hypaque gradient followed by Fc receptor panning with IgG-coated sheep erythrocytes . The number of cells obtained per experiment was 750,000 (extremes 280,000-1,800,000), and the following antibodies were tested on fresh suspensions and/or after 48 hours in culture: BB3 (antithyroglobulin negative control IgG2a), OKT6 (anti CD1a, Ortho), anti HLA-DR (Becton-Dickinson), MHM 24 (anti CD 11a, leukocyte typing workshop n(0)3), MO1 and 44 (anti CD 11b, leukocyte typing workshop n(0)3), anti CD 11c (Immunotech), 60.3 and MHM 23 (anti CD 18, leukocyte typing workshop n(0)2), TS2/9.1.1 (anti CD 58, leukocyte typing workshop n(0)3) . We found that amongst CD 11 subunits, only CD 11c was expressed in fresh suspensions, but was weaker than CD 18, and disappeared with culture . CD 58 was not detected in fresh suspensions but appeared after 2 days of culture, confirming earlier work . Thus the LC exhibits cell surface characteristics similar to tissue macrophages (CD 18 and CD 11c) prior to culture . The expression of CD 58 after culture is in accordance with the interaction of LC with CD2 bearing T-lymphocytes during antigen presentation in peripheral lymph-nodes. Neuroscience, 1992 Jul, 49(1), 33 - 44 Behaviour-dependent changes in acetylcholine release in normal and graft-reinnervated hippocampus: evidence for host regulation of grafted cholinergic neurons; Nilsson OG et al.; Grafted neurons obtained from the fetal basal forebrain can provide a functional cholinergic reinnervation of the hippocampal formation in rats with a lesion of the intrinsic septal cholinergic afferents . In the present experiments graft-derived acetylcholine release in the hippocampus was studied by microdialysis in awake rats during different types of behaviours which are known to activate the innate septohippocampal cholinergic system and during different activity periods of the day-night cycle . Two types of basal forebrain grafts were studied: cell suspensions implanted into the hippocampus in rats with an aspirative lesion of the fimbria-fornix, and grafts of solid tissue implanted as a tissue bridge into the fimbria-fornix lesion cavity . Increased acetylcholine overflow was seen in both groups with grafts during sensory stimulation (by handling) . The strongest response (50% increase in acetylcholine release) was seen in rats with solid basal forebrain grafts (equivalent to two-thirds of that seen in intact rats) . Immobilization stress and motor activity (swimming) also resulted in increased, but more variable, acetylcholine release (+ 30%; about one-third of the normal response) . None of these effects was seen in the control rats with fimbria-fornix lesion only . The two-fold difference in hippocampal acetylcholine release in normal animals between day and night was absent in both types of grafted rats . An acute knife-cut, transecting the connections between the solid basal forebrain graft and the host hippocampus, caused an immediate 75% reduction in acetylcholine release (similar to the effect of an acute fimbria-fornix transection in the normal rats) and the response to swimming was no longer evident . The results show that grafted cholinergic neurons can be functionally integrated into the host brain, allowing the grafted neurons to be activated in the correct behavioural contexts, although the changes in acetylcholine overflow were overall smaller and more variable than normal . The ability of the host to influence cholinergic graft activity, most probably mediated via activation of afferent host-graft connections, may contribute to the efficacy of basal forebrain grafts in the amelioration of behavioural impairments in animals with lesions of the forebrain cholinergic system. Environ Health Perspect, 1992 Jul, 97, 115 - 20 Intraphagolysosomal pH in canine and rat alveolar macrophages: flow cytometric measurements; Heilmann P et al.; Intracellular dissolution of inhaled inorganic particles is an important clearance mechanism of the lung and occurs in phagolysosomal vacuoles of phagocytes . Flow cytometric measurements of intraphagolysosomal pH in alveolar macrophages (AM) obtained from beagle dogs, Wistar rats, and from a baboon were made using fluorescein isothiocyanate-labeled amorphous silica particles (FSP) . AM were obtained by bronchoalveolar lavage . FSP were phagocytized by AM in cell suspensions incubated in full media for 24 hr up to 6 days . Dual laser flow cytometry was performed and six-parameter list mode data were recorded from forward scatter, side scatter, and fluorescence intensities at 530 nm excited at 457 nm and 488 nm as well as logarithmic fluorescence intensity at wavelengths 630 nm excited at 488 nm . In this way it was possible to discriminate viable AM with phagocytized FSP from lysing AM with phagocytized FSP and from cells without FSP and from free FSP . Viable cells were distinguished from lysing cells by staining with propidium iodide immediately before the flow cytometric measurement . A calibration curve for the pH value was determined from FSP suspended in buffered media at pH values ranging from 3.5 to 7.5 . First flow cytometrical results indicated that after an incubation time of 24 hr, the mean intraphagolysosomal pH of viable AM was 4.7 +/- 0.3 for dogs and 5.1 +/- 0.5 for rats . The intraphagolysosomal pH of the baboon AM was 4.5. Clin Investig, 1992 Jul, 70(7), 539 - 44 Distribution of lymphocyte subsets and natural killer cells in the human body; Westermann J et al.; The frequency and distribution of B and T lymphocyte subsets have been determined in many body tissues and fluids by preparing cell suspensions and tissue sections from lymphoid and nonlymphoid organs . In humans these studies often concentrate on the blood or on one particular cell source for obvious reasons . However, such data can only be interpreted correctly if the whole immune system is taken into consideration {64} . To facilitate this, reports on the frequencies and the absolute numbers of B and T lymphocyte subsets within various human tissues and fluids have been collected from a wide variety of journals and are briefly summarized here . Since the size of lymphoid organs varies with age (e.g . thymus, tonsils), only the data of adult individuals were included, unless otherwise stated . Natural killer (NK) cells are morphologically quite similar to lymphocytes {59}, but very different functionally . For example, they are not able to recirculate from the blood via the lymph nodes and the thoracic duct back to the blood as lymphocytes do {19} . Thus, human NK cells have been compared with lymphocytes with respect to number and distribution. Plant Mol Biol, 1992 Jul, 19(4), 577 - 88 Analysis of structure and transcriptional activation of an osmotin gene; Nelson DE et al.; A Nicotiana tabacum gene encoding the basic PR-like protein osmotin was isolated and characterized . The gene is derived from the N . sylvestris parent of N . tabacum . In cell suspension cultures of tobacco, the osmotin gene was shown to be transcriptionally activated by treatment with ABA . Transcriptional activation of the osmotin promoter was further investigated in transformed plants carrying copies of a fusion of the cloned promoter to the beta-glucuronidase reporter gene . In these plants, the osmotin promoter is transcriptionally activated by the hormones ABA and ethylene . The sensitivity of the osmotin promoter to ABA applied exogenously decreased with age in both roots and shoots of young seedlings . NaCl shock also activated the promoter in plant tissues . The osmotin promoter is much more active in root tissues than in shoot tissues. J Histochem Cytochem, 1992 Jul, 40(7), 1059 - 65 Direct flow cytometric quantification of alkaline phosphatase activity in rat bone marrow stromal cells; Kamalia N et al.; Cell preparations in cytochemistry are conventionally analyzed with transmitted light after fixation and reaction with agents such as azo-coupling dyes . With cell suspensions stained with fluorescent cytochemical dyes, cells can also be analyzed and sorted by flow cytometry . We have exploited the intense red fluorescence of Fast Red Violet LB generated in cytochemical reactions to perform flow cytometric analyses of alkaline phosphatase (AP) expression in rat bone marrow stromal cells . By modifying staining protocols of single-cell suspensions, we demonstrate that in comparison to staining with Fast Red TR, the method is specific, can distinguish among various levels of enzyme expression within the whole population, and permits enzyme kinetic studies of heterogeneous cell populations . The method was applied to study the effect of the glucocorticoid dexamethasone (Dx) on cell proliferation and AP expression . In low AP-expressing cells, Dx treatment at 10(-8) M increased the {3H}-thymidine labeling index from 3.85% to 5.24% (p less than 0.01) . In contrast, high AP-expressing cells were unlabeled by {3H}-thymidine . The staining and analytical methods reported here facilitate the detection, isolation, and quantification of subpopulations of bone marrow stromal cells that express alkaline phosphatase activity . These experiments demonstrate the value of flow cytometry as an adjunct to conventional cytochemical methods. Immunology, 1992 Jul, 76(3), 446 - 51 The functional activity of Fc gamma RII and Fc gamma RIII on subsets of human lymphocytes; Hadley AG et al.; Subsets of human lymphocytes were isolated from peripheral blood using magnetic beads coated with anti-CD4, -CD8, -CD19 or -CD56 antibodies to yield T4, T8, B and natural killer (NK) cell suspensions with greater than 95% purity . The functional activity of Fc gamma receptor II (Fc gamma RII) and Fc gamma receptor III (Fc gamma RIII) on these subsets was assessed by measuring rosette formation with red cells sensitized with known levels of either rabbit IgG or human (monoclonal or polyclonal) IgG1 anti-D, IgG3 anti-D or IgG3 anti-c (E-IgG) . Lysis of red cells by K cells (mediated by Fc gamma RIII) in antibody-dependent cell-mediated cytotoxicity (ADCC) assays was promoted by polyclonal and some monoclonal antibodies . Using these 'ADCC+' antibodies, minimum red cell sensitization levels required to promote rosette formation with NK cells were 2000 IgG1 or IgG3 molecules/red cell compared to 15,000 IgG1 or 4000 IgG3 molecules/red cell with 'ADCC-' monoclonal antibodies . The greater efficiency of ADCC+ antibodies is consistent with their previously reported ability to bind Fc gamma RIII via CH2 and CH3 domains whereas ADCC- antibodies bind only via CH3 domains . B cells formed rosettes only at high levels of sensitization: approximately 60,000 IgG1 or 20,000 IgG3 anti-D molecules/cell . These data reflect the low affinity of Fc gamma RII for monomeric human IgG . Although over 90% of NK cells bound anti-CD16, and 70% formed rosettes with red cells sensitized with rabbit IgG (30,000 molecules/cell), only 25% of NK cells formed rosettes with E-IgG3 at 100,000 IgG molecules/cell . Approximately 35% of B cells, 10% of T8 cells but no T4 cells formed rosettes with E-IgG (100,000 IgG3 molecules/cell) . With T8, B and NK cells, IgG3 anti-D promoted greater rosette formation than IgG1 anti-D at comparable levels of sensitization . Presumably the longer hinge region of IgG3 enabled it to bridge the gap between negatively charged lymphocytes and red cells more efficiently than IgG1. Cell Tissue Res, 1992 Jul, 269(1), 1 - 11 Regeneration of autotransplanted splenic tissue at different implantation sites; Liaunigg A et al.; Inbred animals (Lewis rats) were used to investigate the regeneration of autologously implanted splenic tissue at intra-omental and subcutaneous sites . Quantitative immunohistology with monoclonal antibodies against lymphocytes and macrophages was performed to analyse the cell density of red pulp (RP), periarteriolar lymphoid sheath (PALS), marginal zone (MZ) and follicle, 7-180 days after transplantation . Antigenic, allogeneic and mitogenic stimulation and Northern blotting were also performed . Transplant groups differed from spleen only in the reduced size of PALS; however, quantitative analysis demonstrated subtle differences between spleen and transplants . The cell density of B-cells and ED-1+ macrophages was reduced in the RP, Tsupp/cyt-cells were decreased and B-cells increased in PALS, and B-cells and Thelper-cells reduced in the MZ . No differences could be detected between the transplant groups . Flow-cytometric analysis of cell suspensions from spleen and transplants revealed a reduction of T-cells (OX-19+), MHC-I and transferrin-receptor-bearing cells in both transplant groups, and a decrease in the number of Thelper-cells and ED-3+ macrophages in subcutaneous transplants . Both transplant groups were defective regarding the allogeneic and pokeweed mitogen response . Aberration of the lipopolysaccharide response was restricted to subcutaneous transplants, which additionally showed abnormal expression of interferon-gamma, interleukin-5 and interleukin-6 mRNA . Thus, subtle alterations of the newly developed microenvironment and/or lymphocyte-homing may influence the regeneration of splenic tissue; the implantation site may represent an important parameter in functional reorganisation. Immunology, 1992 Jul, 76(3), 485 - 90 Expression of nerve growth factor receptor immunoreactivity on follicular dendritic cells from human mucosa associated lymphoid tissues; Pezzati P et al.; Nerve growth factor (NGF) was originally considered as a trophic factor for peripheral sympathetic and sensory neurones; however, recent reports indicate that NGF may induce proliferation of immune and haematopoietic cells . Histochemical studies conducted in human spleen and lymph nodes have suggested the presence of NGF receptor (NGF-R) immunoreactive elements in secondary follicles; however the nature of the cells bearing the NGF-R in lymphoid tissue has not been determined . In this paper we report the results of an immunohistochemical study conducted on mucosa associated lymphoid tissue . Using a specific monoclonal antibody to human NGF-R (mAb 20.4) we observed an NGF-R-immunoreactive population in all secondary lymphoid follicles examined . Double immunostaining revealed that this population was composed of follicular dendritic cells (FDC); lymphoid cells within the germinal centres did not appear to be 20.4 immunoreactive . Cell suspensions from tonsillar follicles also contained NGF-R immunopositive dendritic cells which were enriched by a 20.4 labelled magnetic bead procedure, revealing cells with the morphological characteristics of FDC . Mononuclear cells from human peripheral blood did not contain any NGF-R-immunoreactive elements using our techniques. Eur J Pharmacol, 1992 Jun 17, 216(3), 401 - 5 Inhibition by heparin of platelet activation induced by neutrophil-derived cathepsin G; Evangelista V et al.; Heparin is the most widely used anticoagulant drug for prevention and treatment of thrombosis . However, inhibition of blood coagulation might not fully explain the antithrombotic activity of this drug . The present study shows that different heparin preparations (50 nM) completely prevent human platelet aggregation, serotonin release and thromboxane B2 production induced by purified neutrophil-derived cathepsin G (E.C . No . 3.4.21.20) . This inhibitory effect was not related to the anticoagulant property of the compounds, since a heparin preparation with an inactivated active for antithrombin III was also effective . Heparins inhibited the protease activity of the enzyme over the same range of concentrations . Since the effect of cathepsin G on platelets requires an intact proteolytic active site, the inhibitory effect of the drugs on cathepsin G-induced platelet activation may be explained by a blockade of protease activity . Heparins were also shown to reduce platelet activation induced by cathepsin G released from activated polymorphonuclear leucocytes in mixed cell suspensions . As polymorphonuclear leucocytes might contribute to both arterial and venous thrombosis through platelet activation induced by the release of cathepsin G, this novel property of heparin could be used to optimize its antithrombotic efficacy. Brain Res, 1992 Jun 12, 582(2), 299 - 311 Autoradiographic study of striatal D1 and D2 dopamine receptors in 6-OHDA-lesioned rats receiving foetal ventral mesencephalic grafts and chronic treatment with L-dopa and carbidopa; Blunt SB et al.; Foetal dopamine cell suspensions or sham preparations were implanted into the denervated striatum of rats with a unilateral 6-hydroxy-dopamine (6-OHDA) lesion of the medial forebrain bundle . Some animals were also treated with L-DOPA (200 mg/kg/24 h) and carbidopa (25 mg/kg/24 h) in the drinking water for 5 weeks, followed by a 3-week drug-free period . Rotational responses to apomorphine and (+)-amphetamine were assessed, and the density of D1 and D2 dopamine receptors was evaluated autoradiographically in striatal slices exposed to {3H}SCH 23390 or {3H}spiperone . Foetal grafts reduces apomorphine-induced contralateral rotation and prevented the development of apomorphine-induced stereotypy . Foetal grafts abolished (+)-amphetamine-induced ipsilateral rotation . These effects of the grafts were not altered by treatment with L-DOPA . A unilateral 6-OHDA lesion of the nigrostriatal pathway resulted in an ipsilateral increase in D2 receptor density most marked in the lateral and dorsomedial quadrants of the striatum compared with the contralateral side . Foetal ventral mesencephalic grafts implanted into the lesioned striatum decreased D2 receptor density to levels found in the contralateral intact striatum . Chronic L-DOPA and carbidopa treatment did not alter the effect of the grafts . A 6-OHDA lesion resulted in a reduction of D1 receptor density in the lateral areas of the lesioned striatum at Level 2 . The presence of a foetal ventral mesencephalic graft either alone or together with L-DOPA treatment did not alter the lesion-induced changes in D1 binding density. Biochim Biophys Acta, 1992 Jun 10, 1135(2), 226 - 8 Receptor binding on whole cells can oscillate; Sukhomlin TK et al.; The study of the kinetics of binding of the opiate receptor agonist {3H}DADLE with NG108-15 cell suspensions has revealed a new periodic biological phenomenon, i.e., oscillations of the cellular receptor activity . The absence of oscillations for binding of the receptor antagonist shows that oscillations occur as a result of the transformation of the receptor signal only. Invest Ophthalmol Vis Sci, 1992 Jun, 33(7), 2304 - 15 Effect of intraocular gamma-interferon on immunoregulatory properties of iris and ciliary body cells; Streilein JW et al.; Resistance of the anterior chamber (AC) of mouse eyes to expression of cell-mediated immunity can be overcome by pre-treating the eye with a dose of recombinant rat gamma-interferon (gamma IFN) that is of itself noninflammatory . To study the mechanism of this form of intraocular inflammation, cells of the tissues surrounding the AC (iris, ciliary body, cornea) were studied in vivo for alterations in phenotype and in vitro regarding their effects on antigen-driven T cell activation . The results indicate that gamma IFN: (1) induced class II major histocompatibility complex (MHC) expression on resident bone marrow-derived cells of iris and ciliary body (I/CB), but not the cornea; (2) led to recruitment of bone marrow-derived cells into the I/CB stroma; and (3) failed to induce class II MHC expression on ocular epithelial cells . Cell suspensions prepared from gamma IFN-treated I/CB superficially resembled normal I/CB cells in that neither were able to activate allogeneic T cells and both were able to suppress antigen-driven T cell activation in vitro . However, unlike cells from normal eyes, I/CB cells from gamma IFN-treated eyes suppressed T cell activation primarily through the secretion of prostaglandins . These results indicate that the ability of gamma IFN-treated eyes to display immunogenic inflammation probably does not result merely from the restoration of conventional antigen presenting cells to this environment, but appears to correlate with a critical change in the molecular mediators of immunosuppression . The findings are discussed in terms of the possibility that the eye may be able to respond to abrogation of its primary immunosuppressive microenvironment by erecting a secondary microenvironment that also is capable of suppressing immunogenic inflammation with a different set of antiinflammatory mediators. Leukemia, 1992 Jun, 6(6), 588 - 94 Acute megakaryoblastic leukemia: establishment of a new cell line (MKPL-1) in vitro and in vivo; Takeuchi S et al.; A megakaryoblastic cell line (MKPL-1) was newly established from the bone marrow of an adult patient with acute megakaryoblastic leukemia . This cell line grew in single cell suspension with a doubling time of 30 h and consisted of large primitive blasts with persistent development of giant cells carrying multilobed nuclei . MKPL-1 cells were positive for platelet GPIIb/IIIa (CD41) and GPIIIa (CD61), and expressed OKM5 (CD36), MY7 (CD13), and MY9 (CD33) antigens in the absence of erythroid and lymphoid markers . The cytochemical and morphologic characteristics of MKPL-1 were also consistent with those of megakaryoblasts . The cells did not, however, express ultrastructural platelet peroxidase which is considered to be another marker of the megakaryocytic lineage . Cytogenetic analysis of MKPL-1 revealed a model chromosome number of 92 with abnormal chromosomes including those found in the patient's bone marrow cells . Furthermore, MKPL-1 cells were serially transplanted into nude mice for nine passages with production of lethal tumors and leukemic manifestation . Thus, our megakaryoblastic cell line which can be maintained both in vitro and in vivo would be useful for further studies of the biology of megakaryopoiesis and megakaryoblastic leukemia. J Immunol, 1992 Jun 1, 148(11), 3674 - 8 Mechanisms of experimental cancer cachexia . Interaction between mononuclear phagocytes and colon-26 carcinoma and its relevance to IL-6-mediated cancer cachexia; Strassmann G et al.; In a recent report we showed that IL-6 is an important mediator of experimental cancer cachexia in the colon-26 (C-26) tumor system . In culture, on a per cell basis, C-26.IVX cell line (which develops tumors and induces severe cachexia of syngeneic hosts) produces up to 60-fold less IL-6 than single cell suspensions prepared from freshly excised tumors . In this study, the mechanism behind this observation was investigated . Analysis of the cellular composition of progressing C-26 tumors indicated they contained up to 6% of macrophages . T cells, B cells, and granulocytes were not detected in the tumors . Because C-26.IVX line grown in vitro contained no macrophages, the possibility that macrophage products may augment IL-6 synthesis by the tumor cells was tested . Indeed, IL-1 beta in a dose-dependent manner and at picogram amounts could potentiate IL-6 production by the C-26 cell line . The presence of high affinity receptors for IL-1 on the C-26.IVX cell line was established . These cells expressed approximately 1500 IL-1 sites per cell with a dissociation constant of approximately 20 pM . Next, we attempted to mimic the situation in vivo by coculture of C-26.IVX cells with syngeneic peritoneal macrophages and found that this condition gives rise to an augmented IL-6 production similar to that observed with in vivo derived tumor cells or rIL-1 beta-treated C-26.IVX cells . Furthermore, anti-IL-1 type I receptor antibody completely blocked C-26.IVX IL-6 production induced by either rIL-1 beta or by peritoneal macrophages . Taken together, these data suggest a pathway of IL-6 production by C-26 tumors that involves a cellular interaction between IL-1R-expressing tumor cells and host-derived macrophages . The results also suggest that this interaction significantly contributes to cachectic events endured by the tumor-bearing host. Zentralbl Bakteriol, 1992 Jun, 277(1), 49 - 53 Detection of Mycoplasma contamination in primary calf kidney cell cultures; Pfutzner H et al.; A total of 31 primary cell culture preparations of calf kidney including their processing stages (kidney, washing fluid, cell suspension, cell culture monolayer) were investigated for mycoplasmas by cultural methods . Mycoplasma (M.) arginini, was isolated from 8 out of 27 investigated samples of calf kidney from 3 out of 26 washing fluids, 5 out of 20 cell suspensions, and from 9 out of 21 cell culture monolayers . Furthermore, M . arginini was repeatedly found in throat swabs of the cell laboratory technicians . The results give indications as to the source and route of mycoplasma infections of primary cell cultures. Pharm Res, 1992 Jun, 9(6), 782 - 7 Macrophage activation by polymeric nanoparticles of polyalkylcyanoacrylates: activity against intracellular Leishmania donovani associated with hydrogen peroxide production; Gaspar R et al.; Nanoparticles of polyalkylcyanoacrylates (PACA) can be useful carrier for the targeting of antileishmanial drugs into macrophages and also possess significant antileishmanial activity by themselves . No significant difference in antileishmanial activity could be detected between nanoparticles of five PACAs with differing alkyl side chains, suggesting that the main degradation products of PACA are not involved in their antileishmanial action . The effect of polyisohexylcyanoacrylate (PIHCA) on the induction of the respiratory burst in a macrophage-like cell line (J774G8) was assessed in non-infected macrophages and in macrophages infected with amastigotes of Leishmania donovani infantum, by measuring nitroblue tetrazolium (NBT) reduction and hydrogen peroxide production . Phagocytosis of PIHCA nanoparticles led to a respiratory burst, which was more pronounced in infected than in uninfected macrophages . The production of reactive oxygen intermediates associated with the respiratory burst was inhibited by addition of superoxide dismutase and catalase to the cell suspensions . The addition of catalase to the culture medium together with PIHCA nanoparticles significantly reduced the antileishmanial activity of PIHCA . Moreover PIHCA nanoparticles did not induce interleukin-1 release by macrophages . It is suggested that the antileishmanial action of PIHCA and other PACA nanoparticles results from the activation of respiratory burst in macrophages. Am J Physiol, 1992 Jun, 262(6 Pt 1), G1021 - 6 Effect of osmotic swelling on K+ conductance in jejunal crypt epithelial cells; MacLeod RJ et al.; To further elucidate differences in ion transport properties between jejunal crypt and villus cells, we compared the responses of purified cell suspensions to hypotonic stress using electronic cell sizing to evaluate volume changes and 86Rb and 36Cl efflux . After hypotonic swelling, villus enterocytes undergo a regulatory volume decrease (RVD) due to the loss of K+ and Cl- through volume-activated conductances . After 0.6x isotonic challenge in Na(+)-free medium, crypt cells exhibited only partial RVD, with t1/2 congruent to 15 min . The addition of a cation ionophore, gramicidin (0.25 microM), to hypotonically swollen crypt cells caused an accelerated RVD, which was complete with t1/2 congruent to 5 min . Crypt epithelial cells showed no volume-activated 86Rb efflux, but villus enterocytes had an increased rate of 86Rb efflux after hypotonic dilution (P less than 0.001) . Gramicidin added to hypotonically diluted crypt cells greatly increased the rate of 86Rb efflux compared with controls . Both villus (30 s; P less than 0.005) and crypt (2 min; P less than 0.001) cells exhibited volume-activated 36Cl efflux in absence of gramicidin . Cl- channel blockers anthracene-9-carboxylate (9-AC, 300 microM) and indanyloxyacetic acid (IAA-94, 100 microM) prevented crypt RVD (P less than 0.001) in the presence of gramicidin . Ouabain (P less than 0.001) or K(+)-free Na(+)-containing medium, but not Ba2+ (5 mM) or quinine (100 microM), prevented crypt partial RVD . We conclude that crypt cells lack volume-activated K+ conductance . The RVD exhibited by crypt cells, although partial, was due to Cl- loss through a volume-activated Cl- conductance and Na+ loss via Na(+)-K(+)-ATPase. Environ Res, 1992 Jun, 58(1), 117 - 23 Enhanced histamine release from lung mast cells of guinea pigs exposed to sulfuric acid aerosols; Fujimaki H et al.; To clarify the relationship between air pollution and mast cell response, the effects of sulfuric acid aerosols on histamine release from lung mast cells of guinea pigs were investigated . Guinea pigs were exposed to 0.3, 1.0 and 3.2 mg/m3 sulfuric acid (H2SO4) aerosols or 4 ppm nitrogen dioxide (NO2) for 2 and 4 weeks . After the exposure, lung mast cell suspensions were isolated by collagenase treatment and antigen- or A23187-induced histamine release was measured . Antigen-induced histamine release from mast cells was significantly enhanced by the exposure to 1.0 and 3.2 mg/m3 H2SO4 for 2 weeks, but exposure to H2SO4 for 4 weeks did not show the enhancement of antigen-induced histamine release . A23187-induced histamine release was significantly enhanced by the exposure to 1.0 mg/m3 H2SO4 or 4 ppm NO2 for 2 weeks, but suppression of histamine release from lung mast cells stimulated with A23187 was observed by the exposure to 3.2 mg/m3 H2SO4 for 4 weeks . The exposure to 0.3 mg/m3 H2SO4 showed no changes in antigen- and A23187-induced histamine release . The combination of 1.0 mg/m3 H2SO4 with 4 ppm NO2 for 2 weeks resulted in no changes in antigen- and A23187-induced histamine release . These results suggested that functional properties of lung mast cells may be altered by a low concentration of H2SO4 aerosol exposure. Biotechnology (N Y), 1992 Jun, 10(6), 691 - 6 Transgenic plants of tall fescue (Festuca arundinacea Schreb.) obtained by direct gene transfer to protoplasts; Wang ZY et al.; Chimeric hygromycin phosphotransferase (hph) and phosphinothricin acetyltransferase (bar) genes were introduced, using polyethylene glycol treatment, into protoplasts isolated from embryogenic cell suspension cultures of tall fescue (Festuca arundinacea Schreb.), a graminaceous plant that is an important forage crop in temperate pastures . Colonies resistant to either 200 mg/l hygromycin or 100 mg/l phosphinothricin, respectively, were recovered upon selection using bead-type culture systems . Stable integration of the transgenes in the genomes of plants regenerated from resistant callus clones was shown by Southern hybridization analysis . In situ hybridization of a labeled transgene-probe to metaphase chromosomes is shown for one transgenic primary regenerant . Expression of the transgenes in mature plants was demonstrated by HPH enzyme assay or by phosphinothricin-herbicide spraying. Neuroscience, 1992 Jun, 48(4), 857 - 69 Host dopaminergic afferents affect the development of DARPP-32 immunoreactivity in transplanted embryonic striatal neurons; Defontaines B et al.; Homotopic transplantation provides an interesting way to observe the relationships between developing cells and ingrowing host afferents . We have performed a complete and selective elimination of the mesostriatal dopaminergic system in adult rats to observe the influence of its absence on the development and chemical differentiation of embryonic striatal cells . Cell suspensions from striatal primordia of 14-15-day-old embryos were transplanted into host striata that were (i) neuron-depleted by kainic acid (control group) or (ii) deprived of dopamine by 6-hydroxydopamine prior to the neuronal depletion by kainic acid (experimental group) . The expression of dopamine- and adenosine 3',5'-monophosphate-regulated phosphoprotein (DARPP-32) by transplanted cells was observed in correlation with their innervation by host dopaminergic afferents which in turn were identified by tyrosine hydroxylase immunohistochemistry . Observations were made between four days and three months after transplantation . Four days after transplantation, no immunoreactivity for DARPP-32 was observed in transplants of control animals despite the presence of tyrosine hydroxylase-immunopositive fibers growing from the host to discrete cell clusters in the transplant . DARPP-32-labeled cells appeared soon afterwards . Six days after transplantation they displayed varying intensities of immunoreaction, ranging from just detectable to normal levels and were specifically targeted by developing tyrosine hydroxylase-immunopositive fibers . The number of DARPP-32-labeled cells increased rapidly and they formed increasingly compact clusters . Fourteen days after transplantation and afterwards, all the DARPP-32-labeled cells displayed an intensity of immunoreaction and a distribution comparable to that observed in long-term transplants . Transplants in the experimental hosts displayed the same organization and developmental features as the control transplants with the exception of DARPP-32 labeling which was not detected before eight days after transplantation . Ten days after transplantation, the distribution and intensity of DARPP-32 labeling was similar to that observed at six days in the control group . The evolution of DARPP-32 labeling after 10 days in the experimental group paralleled that observed six days post-transplantation and beyond in the control group . Dopaminergic mesostriatal host afferents are able to provide developing cells in grafted striatal tissues with normal innervation very rapidly . Despite this rapidity, the innervation does not seem to have any trophic influence on the general development of the transplant but does affect the onset time of the expression of neurochemical markers that are directly related to its synaptic function.(ABSTRACT TRUNCATED AT 400 WORDS) Int J Radiat Biol, 1992 Jun, 61(6), 767 - 72 The effect of fluoride on photodynamic-induced fluorescence changes of aluminium phthalocyanine in Chinese hamster cells; Ben-Hur E et al.; Fluence-dependent changes in the fluorescence of aluminium phthalocyanine (AlPc) were measured in Chinese hamster ovary (CHO) cells using digital fluorescence microscopy of single cells and spectrofluorimetry of cell suspensions . During illumination the fluorescence initially increased and later progressively decreased . In the presence of fluoride, which protects against phototoxicity of AlPc by forming a fluoroaluminium complex, there was no initial increase in fluorescence: it decreased about 10 times faster than in the absence of fluoride . Qualitatively similar results were observed using single-cell fluorescence microscopy, which also showed the dye to be mostly localized in cytoplasmic organelles and membranes . The pattern of localization did not change during illumination . Concomitant assays of dye extracted from cells revealed little photodegradation that could not account for the fluorescence changes . The absorption spectra of AlPc-loaded cells showed some aggregation of the dye prior to light exposure . During illumination the dye was initially monomerized and subsequently progressively reaggregated . In the presence of fluoride no monomerization was seen, and the aggregation proceeded at a much faster rate . It is concluded that the fluorescence changes are not due to major relocalization of AlPc in the cells, but to light-induced monomerization followed by reaggregation . The protective effect of fluoride may be due to the enhanced aggregation rate, because aggregated dye molecules are photochemically inactive . Because D2(0) affects neither the initial enhanced fluorescence in the absence of fluoride nor the rapid decrease in its presence it appears that 1O2 is not involved in the photodynamic reactions leading to these changes. J Immunol, 1992 Jun 1, 148(11), 3583 - 7 Fibroblast stimulation in schistosomiasis . XII . Identification of CD4+ lymphocytes within schistosomal egg granulomas as a source of an apparently novel fibroblast growth factor (FsF-1); Prakash S et al.; Granulomas that form around Schistosoma mansoni eggs deposited in the liver secrete a variety of fibrogenic factors that may provide a molecular link between chronic inflammation and hepatic fibrogenesis in schistosomiasis . We recently isolated from conditioned medium of egg granuloma cultures a approximately equal to 60-kDa heparin-binding growth factor for fibroblasts . Because this protein is distinct from other defined heparin-binding growth factors, we designated it "fibroblast stimulating factor-1" (FsF-1) . We now report that FsF-1 is a lymphokine . We prepared IgG antibody against purified FsF-1 and determined that it did not cross-react with a variety of growth factors or recombinant interleukins . Using two-color flow cytometry of dissociated granuloma cell suspensions, we observed that approximately 20% to 25% of granuloma CD4+ lymphocytes express surface FsF-1 . We isolated CD4+ granuloma lymphocytes by FACS and observed that these cells spontaneously secrete into culture supernatant a fibroblast mitogen that is neutralized by anti-FsF-1 antibody . Furthermore, anti-FsF-1 can specifically immunoprecipitate a metabolically labeled protein produced by the granuloma CD4+ lymphocytes . The labeled protein has the same apparent molecular mass (approximately equal to 60 kDa) as FsF-1 purified from granuloma culture supernatants . These findings define CD4+ lymphocytes as a source of FsF-1 . Because FsF-1 has biologic and chemical features distinct from most other defined lymphokines and from other heparin-binding growth factors, FsF-1 appears to be a novel lymphokine. Diagn Mol Pathol, 1992 Jun, 1(2), 85 - 97 In situ polymerase chain reaction detection of viral DNA, single-copy genes, and gene rearrangements in cell suspensions and cytospins; Komminoth P et al.; The study of low-copy viral or genomic DNA sequences by in situ hybridization (ISH) is often limited by sensitivity . Using the polymerase chain reaction (PCR) for the amplification of target DNA sequences in fixed cells {in situ PCR} (ISPCR) before ISH, we have been able to greatly improve the sensitivity of ISH . Viral DNA present in low copy number, single-copy genes, as well as immunoglobulin gene rearrangements (VH3 family genes), were successfully amplified in cells in suspension or on glass slides (cytospins) . Single primer pairs were used in the in situ amplification step and 35S- or digoxigenin-11-dUTP-labeled region specific oligonucleotide probes were used for detection of amplificants by ISH . Artifacts, presumably resulting from leakage of in situ amplificants out of cells, may be a significant problem in selected instances . By optimal fixation and permeabilization of cells, limiting PCR cycle number, amplification of long DNA sequences, and/or incorporation of biotinylated dNTPs to produce bulkier amplificants together with washing of cells after ISPCR, diffusion artifacts were significantly reduced . Probe hybridization to single-stranded long PCR fragments or messenger RNA were excluded as a source for false-positive ISPCR results . The techniques reported dramatically increase the sensitivity of ISH in the detection of low-copy viral infection as well as in the study of gene rearrangements, and provide unique opportunities to study chromosomal translocations and point mutations at the cellular level. Cell Biol Int Rep, 1992 Jun, 16(6), 557 - 65 Changes in surface relief of suspended cells are morphological signs of the initial stage of neoplastic transformation in fibroblastic monolayer cultures; Rovensky YA et al.; The percentages of cells with different types of cell surface relief were determined in cell suspensions derived from monolayer cultures . Primary cultures of rat embryo fibroblasts (REF) and cell lines REF (LT) and REF-1, immortalized cells of which preserved normal phenotypic characteristics of the initial primary culture REF, as well as morphologically transformed tumorigenic lines REF (LT) ras and REF-2EJ were studied . In REF suspensions the cells with the blebbed type of surface relief were shown to be predominant as compared with those with microvillus relief whereas cell suspensions derived from both immortalized and fully transformed cultures display the reverse ratio of cells with those types of surface relief . Therefore, the pattern of cell surface relief in cell suspensions derived from fibroblastic monolayer cultures may serve as a morphological marker of the initial stage of neoplastic transformation-immortalization when typical morphological signs of cell transformation are not yet manifested in monolayer cultures. J Med Virol, 1992 Jun, 37(2), 143 - 8 Detection of cytomegalovirus-infected cells by flow cytometry and fluorescence in suspension hybridisation (FLASH) using DNA probes labeled with biotin by polymerase chain reaction; Link H et al.; A biotin-labeled DNA probe specific for the immediate early gene of the cytomegalovirus (CMV) strain AD 169 was generated with the polymerase chain reaction . Nucleic acid hybridisation was carried out with CMV-infected T-lymphoblastoid cells (MOLT-4) after 4 hr to 6 days of culture . Biotin molecules were made visible with streptavidin coupled with fluorescein . The fluorescence signal of the hybridised probe was measured by flow cytometry in the cell suspension . The number of CMV-positive cells was 7% at 4 hr, 8% after 28 hr, 18% after 2 days, 26% after 3 days, 91% after 4 days, 97% after 5 days, and 98% after 6 days . The first detection of CMV antigen (pp65) was possible with immunoenzymatic labeling by day 4, whereas CMV-DNA was detected by PCR after 4 hr . CMV-specific RNA could be detected in a similar way . The analysis of mononuclear peripheral blood leukocytes in a patient with active CMV infection showed 14.7% CMV DNA-positive cells at day 1 and 7% at day 8, as compared to 0.9% and 0.0% cells which were positive for CMV antigen (pp65) by immunoenzymatic labeling at day 1 and day 8, respectively . We conclude that flow cytometry and fluorescence in suspension hybridisation (FLASH) offers a new tool for analysing exactly and quantifying large numbers of cells for specific DNA or RNA and may be useful for other laboratory and clinical applications. Blood, 1992 May 15, 79(10), 2688 - 93 Differential expression of very late activation antigen-3 (VLA-3)/VLA-4 in B-cell non-Hodgkin lymphoma and B-cell chronic lymphocytic leukemia; Baldini L et al.; The expression of beta 1 (very late activation antigens, VLA 1-6) and beta 2 integrins (leukocyte adhesion molecules {Leu-CAM}) in cell suspensions from the peripheral blood of 70 patients with B-cell chronic lymphocytic leukemia (B-CLL), 15 patients with leukemic lymphocytic lymphoma of intermediate differentiation (IDL), as well as from the lymph nodes of 20 patients with low/intermediate-grade non-Hodgkin's lymphoma (NHL) was studied with the aim of characterizing their adhesive phenotype and evaluating its relationship to clinical behavior . CD11a(LFA-1) was more expressed in NHL and IDL than in B-CLL (P = .047), although it was demonstrable in 74.2% of cases; CD11c was more expressed in B-CLL (P less than .0001), and its expression was preserved in almost all of the cases of small lymphocytic lymphoma . In NHL patients, including the cases of IDL, VLA-3 expression was observable in 8 of 35 cases (although always at a low level of intensity), while VLA-4 was almost constantly expressed in a way that was similar to its expression in control normal B cells . On the contrary, in B-CLL patients, VLA-3 was expressed (prevalently at high levels) in 87.1% of cases and VLA-4 only in 37.1% . No correlation was found between adhesion molecule patterns and the clinical features of the diseases . The biofunctional significance of the imbalance of VLA-3 and VLA-4 expression in B-CLL is not easy to explain, but it has undoubted intrinsic value as an additional marker for distinguishing B-CLL from, in particular, those B-cell neoplasms (such as IDL) that share many of the immunocytomorphologic characteristics and the putative normal counterpart (the mantle zone) of B-CLL. Vet Immunol Immunopathol, 1992 May, 32(3-4), 195 - 203 A qualitative assay for beta cell antibodies . Preliminary results in dogs with diabetes mellitus; Hoenig M et al.; Purified beta cells from a radiation-induced transplantable rat insulinoma were used to detect beta cell antibodies in serum from untreated diabetic dogs . Serum from dogs in which anti-beta cell antibodies were induced by injecting a purified beta cell suspension subcutaneously was used as positive control . Following incubation with test sera, fluorescein-labeled anti-dog immunoglobulins were used to visualize binding between the beta cells and dog gamma globulins . Nine of the 23 diabetic dogs showed a strongly positive reaction which was characterized by a ring fluorescence, three showed a weak reaction and 11 were negative, i.e . they showed diffuse fluorescence . In contrast, 14 of the 15 healthy dogs showed diffuse fluorescence and one dog showed a weakly positive reaction . Thyroid, liver and kidney cells did not elicit ring fluorescence . Although females (spayed and intact) represented the majority of the diabetic dogs, there was no correlation between sex and the occurrence of antibodies in the diabetic dogs . There was also no correlation to the age of the dogs . In conclusion, we have developed a specific test for anti-beta cell antibodies . The test is reproducible and economical to perform on a large number of samples. Appl Environ Microbiol, 1992 May, 58(5), 1764 - 7 Dipstick immunoassay to detect enterohemorrhagic Escherichia coli O157:H7 in retail ground beef; Kim MS et al.; A sensitive and easy-to-perform dipstick immunoassay to detect Escherichia coli O157:H7 in retail ground beef was developed by using a sandwich-type assay (with a polyclonal antibody to E . coli O157 as the capture antibody and a monoclonal antibody to E . coli O157:H7 as the detection antibody) on a hydrophobic polyvinylidine difluoride-based membrane . E . coli O157:H7 in ground beef could be detected within 16 h, including incubation for 12 h in enrichment broth and the immunoassay, which takes 4 h . Pure culture cell suspensions of 10(5) or 10(6) E . coli O157:H7 organisms per ml produced intense color reactions in the immunoassay, whereas faint but detectable reactions occurred with 10(3) CFU/ml . The sensitivity of the combined enrichment-immunoassay procedure as determined by using ground beef inoculated with E . coli O157:H7 was 0.1 to 1.3 cells per g, with a false-positive rate of 2.0% . A survey of retail ground beef using this procedure revealed that 1 of 76 samples was contaminated by E . coli O157:H7. Jpn J Cancer Res, 1992 May, 83(5), 483 - 90 Progression of weakly malignant clone cells derived from rat mammary carcinoma by host cells reactive to plastic plates; Hamada J et al.; Tumor progression is the process by which tumor cells acquire more malignant properties, such as invasiveness and metastasis, during tumor development . To elucidate mechanisms of tumor progression, we examined the role of interactions between the tumor and its host by using a cloned cell line, ER-1, which was derived from a rat mammary carcinoma . ER-1 is weakly tumorigenic and non-metastatic when s.c . injected into syngeneic hosts in single cell suspension . However, ER-1 cells show a high incidence of lethal growth when s.c . implanted (5 x 10(2) cells), being attached to a 10 x 5 x 1 mm polystyrene plate . Tumor cell lines (PLT) obtained from tumors which had arisen from the plate-attached ER-1 cells no longer required plates for their growth in normal hosts, and had acquired metastatic ability to the lungs . The malignant phenotypes of PLT were stable under a usual culture condition for at least 6 months . Furthermore, the incidence of tumor development increased when small numbers of ER-1 cells were injected onto plates (or at their periphery) which had previously been implanted s.c . without tumor cells . The tumorigenicity of ER-1 cells increased after they were cocultivated for more than 30 days with host reactive cells obtained from the tissues surrounding the plates . These results suggest that host cells reactive to the foreign body (plastic plate) may not only promote the local growth of ER-1 cells but also convert them into much more malignant tumors. J Exp Zool, 1992 May 1, 262(2), 113 - 21 Cardiac performance of an isolated eel heart: effects of hypoxia and responses to coronary artery perfusion; Davie PS et al.; The pericardial sac containing the heart was removed from large (2.7-6.3 kg) long-finned eels (Anguilla dieffenbachii) . Coronary arteries were cannulated in preparation for perfusion with eel Ringer or red cell suspensions . The hearts were maintained by Ringer perfusion while the performance of the heart was assessed . Responses of the hearts to increases in filling pressure and output pressure were recorded . Maximum cardiac output was 22.3 +/- 1.4 ml/min/kg body mass (mean +/- 1 SEM; N = 9) . The highest cardiac power output was measured at maximum cardiac output and was 3.39 +/- 0.32 mW/g ventricle mass (mean +/- 1 SEM; N = 9) . Eel hearts could sustain output pressures near 80 cm H2O, but cardiac output was reduced and cardiac power output was 1.89 +/- 0.24 mW/g ventricular mass (mean +/- 1 SEM; N = 9) . Maximum cardiac output decreased by 14% when hearts pumped hypoxic Ringer with a PO2 of 11.5 torr . At high input pressures concomitant with high output pressures (80 cm H2O), cardiac power output decreased by 38% upon exposure to hypoxic Ringer . Coronary perfusion of hypoxic hearts with red cell suspensions (mean hematocrit 10.4%) at a rate of 2% of control cardiac output (0.20 ml/min/kg body mass) had no effect on maximum cardiac output . However, coronary perfusion enabled hypoxic hearts to maintain cardiac output when output pressure was raised to 80 cm H2O . Under conditions of high input pressure and high output pressure, power output increased by 20% compared to hypoxic hearts without coronary perfusion.(ABSTRACT TRUNCATED AT 250 WORDS) J Virol, 1992 May, 66(5), 3131 - 9 The full-length transcript of a caulimovirus is a polycistronic mRNA whose genes are trans activated by the product of gene VI; Scholthof HB et al.; Gene expression of figwort mosaic virus (FMV), a caulimovirus, was investigated by electroporation of Nicotiana edwardsonii cell suspension protoplasts with cloned viral constructs in which a reporter gene was inserted at various positions on the genome . The results showed that the genome of FMV contains two promoters; one is used for the production of a full-length RNA and another initiates synthesis of a separate monocistronic RNA for gene VI . Evidence is provided that the full-length transcript, the probable template for reverse transcription, can serve as a polycistronic mRNA for translation of genes I through V and perhaps also gene VI . Expression of all the genes on the polycistronic mRNA is trans activated by the gene VI protein . Reporter gene expression appears most efficient when its start codon is in close proximity to the stop codon of the preceding gene, as for the native genes of caulimoviruses . We propose that the gene VI product enables expression of the polycistronic mRNA by promoting reinitiation of ribosomes to give translational coupling of individual genes. Am J Pathol, 1992 May, 140(5), 1045 - 54 Interleukin-1 receptor antagonist protein inhibits interleukin-8 expression in lipopolysaccharide-stimulated human whole blood; DeForge LE et al.; Interleukin-8 (IL-8) is a neutrophil and lymphocyte chemoattractant and activator that may play an important role in mediating events at sites of inflammation . IL-8 is produced by many cell types in response to a variety of inducers, including interleukin-1 (IL-1) . Studies were conducted to address the question of whether an inhibitor of IL-1 action, IL-1 receptor antagonist protein (IRAP), would suppress IL-8 production . Lipopolysaccharide (LPS)-stimulated human whole blood was used as an ex vivo model of local cytokine production . Preliminary time course studies showed that plasma IL-1 beta levels were fully induced by 6 hours (approximately 15 ng/ml) and persisted at this level over 24 hours . IL-8 production, in contrast, reached a plateau between 6 to 12 hours (5 ng/ml) and then increased rapidly to 17 ng/ml at 24 hours . Addition of IRAP was found to dose-dependently inhibit IL-8 expression at the levels of both protein and mRNA . A 50% reduction in IL-8 protein release occurred at an IRAP dose of 8 micrograms/ml (5 x 10(-7) mol/l) at 24 hours . A 2 hour delay in the addition of IRAP relative to LPS still permitted optimal reduction in IL-8 release, whereas delays of 4-8 hours reduced or eliminated the inhibitory effect . IRAP was found to have no effect on the LPS-stimulated production of IL-1 alpha or IL-1 beta . In addition, experiments performed with isolated peripheral blood cells demonstrated that, whereas monocytes were the major producers of IL-8, IRAP was equally effective in reducing IL-8 production in neutrophil and mononuclear cell suspensions . These studies further document the role of IL-1 in inducing the production of IL-8 and indicate that the ability of IRAP to suppress IL-8 expression may be an important mechanism of the anti-inflammatory properties of this molecule. J Exp Med, 1992 May 1, 175(5), 1353 - 65 Epidermal Langerhans cells from normal human skin bind monomeric IgE via Fc epsilon RI; Wang B et al.; Human epidermal Langerhans cells (LC) bearing IgE are found in disease states associated with hyperimmunoglobulinemia E . When studying the mechanism(s) underlying this phenomenon, immunohistology revealed that a majority of epidermal LC from normal skin of healthy individuals can specifically bind monomeric IgE . IgE binding to LC could neither be prevented by preincubation of the tissue with monoclonal antibodies (mAb) against either Fc epsilon RII/CD23 or Fc gamma RII/CD32, nor by the addition of lactose . However, binding could be entirely abrogated by preincubation with the anti-Fc epsilon RI alpha mAb 15-1, which interferes with IgE binding to Fc epsilon RI alpha gamma transfectants . These observations indicated that IgE binding to epidermal LC is mediated by Fc epsilon RI rather than by CD23, CD32, or the D-galactose-specific IgE-binding protein . This assumption gained support from our additional findings that: (a) the majority of LC exhibited distinct surface immunolabeling with the anti-Fc epsilon RI alpha mAbs 15-1 and 19-1, but not with any of eight different anti-Fc epsilon RII/CD23 mAbs; and (b) transcripts for the alpha, beta, and gamma chains of Fc epsilon RI could be amplified by polymerase chain reaction from RNA preparations of LC-enriched, but not of LC-depleted, epidermal cell suspensions . In view of the preeminent role of Fc epsilon RI crosslinking on mast cells and basophils in triggering the synthesis and release of mediators of allergic reactions, the demonstration of this receptor on epidermal LC may have important implications for our understanding of allergic reactions after epicutaneous contact with allergens. Neurochem Res, 1992 May, 17(5), 475 - 82 Serotonin fiber innervation of cerebellar cell suspensions intraparenchymally grafted to the cerebellum of pcd mutant mice; Triarhou LC et al.; One aspect of integration of implanted neurons into the neuronal circuitry of a defective host brain is the re-establishment of a host-to-graft afferent innervation . We addressed this issue by using the adult cerebellum of 'Purkinje cell degeneration' (pcd) mutant mice, which lack virtually all Purkinje cells after postnatal day (P) 45 . Purkinje cells constitute one of the cerebellar cell types being innervated by axons of raphe serotonin (5-HT) neurons . In normal mice, 5-HT-immunoreactive fibers are distributed to all cerebellar folia . Following Purkinje cell loss in pcd mice, cerebellar 5-HT-immunoreactive fibers persist . Cerebellar cell suspensions were prepared from embryonic day (E) 11-13 normal mouse embryos and were intraparenchymally grafted into the cerebellum of pcd mutants either directly or after pre-treatment with 5,7-dihydroxytryptamine (5,7-DHT) to selectively remove 5-HT cells of donor origin . The state of Purkinje cells and 5-HT axons was monitored in alternate sections by 28-kDa Ca(2+)-binding protein (CaBP) and 5-HT immunocytochemistry, respectively . Serotonin-immunoreactive axons were seen in the grafts from 5 to 32 days after transplantation . In some of the grafts which had not been pre-treated with 5,7-DHT, a small number of 5-HT-immunoreactive cell bodies was found, indicating that part of the 5-HT fiber innervation of the graft could actually derive from donor cells.(ABSTRACT TRUNCATED AT 250 WORDS) Chem Res Toxicol, 1992 May-Jun, 5(3), 386 - 91 Utilization of glutathione during 1,2-dihaloethane metabolism in rat hepatocytes; Jean PA et al.; The metabolism of 1,2-dihaloethanes (DHEs) to glutathione-containing metabolites by freshly isolated rat hepatocytes was investigated . 1,2-Dichloroethane (DCE), 1,2-dibromoethane (DBE), and 1-bromo-2-chloroethane (BCE) were metabolized to S-(2-hydroxyethyl)glutathione (HEG), S-(carboxymethyl)glutathione (CMG), and S,S'-(1,2-ethanediyl)bis(glutathione) (GEG) . The formation of these glutathione-containing metabolites was concomitant with the depletion of intracellular glutathione (GSH) and accounted for 58%, 84%, and 71% of the DCE-, BCE-, and DBE-induced loss of intracellular GSH, respectively . The covalent binding of {14C}DBE to hepatocyte protein reached 18.7 nmol/mL of cell suspension (7.8 nmol/mg of protein) within 2.0 h of incubation . Half of this covalent binding occurred within 0.5 h of incubation (4.0 nmol/mg of protein) in the presence of high levels of intracellular GSH (30% of initial GSH level at 0.5 h) . Hepatocyte metabolism of 2-chloroacetic acid produced only CMG . 2-Chloroethanol metabolism gave rise to CMG and HEG in a 11.5:1.0 ratio; 2-chloroacetaldehyde produced almost equal amounts of CMG and HEG . GEG formation was increased significantly for DBE and BCE when GSH was added to the medium during treatment, suggesting that the GSH conjugates S-(2-haloethyl)glutathione are exported from the hepatocytes . These results indicate that the glutathione S-transferase-catalyzed conjugation of GSH with the DHEs is responsible for the majority of the DHE-induced GSH depletion . The S-(2-haloethyl)glutathione conjugates appear responsible for the extensive covalent binding to protein observed during {14C}DBE metabolism. Biull Eksp Biol Med, 1992 May, 113(5), 488 - 9 {Role of leukocyte adhesion in filterability of blood cell suspensions}; Redchits EG et al.; We investigated blood cells suspension filterability of 16 donors . The filtration was performed trough 5 microns-pore nuclear filters at constant perfusion pressure 10(5) din/cm2 . We estimated also the adherence of leukocytes and platelets on nylon . The adherence of platelets and mononuclear leukocytes reduced the level of the suspension filterability only by 21% (p > 0.05) . The presence of nonadhesive polynuclear leukocytes in the suspensions did not change practically their filterability . The addition in the suspensions of adhesive polynuclear leukocytes reduced suspension filterability dramatically. Biotechniques, 1992 May, 12(5), 632 - 8 Modifications of the guanidine hydrochloride procedure for the extraction of RNA: isolation from a variety of tissues and adherent/nonadherent cell types; Kamdar SJ et al.; In a previous report dealing with the guanidine hydrochloride protocol for the extraction of RNA from mouse peritoneal macrophages, we identified a major source of RNA-degrading activity and showed that its removal early in the extraction procedure resulted in a more dependable method for the recovery of high-quality RNA . This report extends these findings and demonstrates the general applicability of the technique to a variety of fresh or frozen adherent cell types, cell suspensions and tissues, further highlighting stages at which degradation is most likely to occur and how to avoid a variety of pitfalls associated with the extraction procedure. Phytochemistry, 1992 May, 31(5), 1593 - 601 Anthocyanins from cell suspension cultures of Daucus carota; Glassgen WE et al.; Six anthocyanins were isolated from cell suspension cultures of an Afghan cultivar of Daucus carota by PC or HPLC . The structures of these compounds were elucidated by spectroscopic methods as cyanidin 3-O-lathyroside, cyanidin 3-O-(2''-O-beta-D-xylopyranosyl-6''-O-beta-D-glucopyranosyl-beta-D- galactopyranoside), and the latter acylated with 4-coumaric, ferulic, 4-hydroxybenzoic or sinapic acid . Unusual 1H NMR chemical shifts and 1H NOE data indicate an intramolecular copigmentation of the aglycone with these aromatic residues. Reg Immunol, 1992 May-Jun, 4(3), 119 - 29 Adhesion molecules on the plasma membrane of epidermal cells . IV . Immunolocalization of the intercellular adhesion molecule-1 (ICAM-1, CD54) on the cell surface of a small subpopulation of keratinocytes freshly isolated from normal human epidermis; De Panfilis G et al.; The intercellular adhesion molecule-1 (ICAM-1) is a cell membrane glycoprotein displaying a pivvtal role in cell-cell interactions in the immune system, and is a ligand for LFA-1, which is expressed on leukocytes . ICAM-1 is expressed in different cell types, including epithelial cells in a number of organs; the universal feature on all these cells is ICAM-1 induction from very low ICAM-1 constitutive levels on unstimulated resting cells to very high ICAM-1 levels triggered by mediators released at sites of inflammation . Therefore, since a strong expression of ICAM-1 on keratinocyte (KC) surface was recently demonstrated in various inflammatory skin lesions, in this investigation we asked whether very low ICAM-1 levels might be present on the plasma membrane of unstimulated KC in normal skin . Crude epidermal cell suspensions, freshly isolated from normal human skin, were immunolabeled by anti-ICAM-1 monoclonal antibody and stained by two highly sensitive ultrastructural detection systems, namely, the immunogold (5-nm-sized particles) method and the immunogold-silver-enhancement method . The quantitative analysis of 1000 KC scrutinized under the electron microscope revealed that 17.2% KC were ICAM-1-positive, although a density per KC section (midplane) of merely 18.92 +/- 13.02 5 nm-sized particles was scored (n = 100), indicating that the amounts of ICAM-1 moieties on this KC subset are presumably low . The ICAM-1 expression on a subset of KC in normal skin might account for the trafficking to and from normal epidermis of LFA-1-positive cells, including migrating Langerhans cells and occasional leukocytes. Rev Paul Med, 1992 May-Jun, 110(3), 97 - 101 Simplified method for the analysis of cellular karyotype and phenotype in leukemias; Chauffaille Mde L et al.; The objective of this study was to develop a simplified method for the simultaneous analysis of cellular karyotype and phenotype which would permit the identification of cell origin . We studied 6 patients with AML, 3 with CML (one of which was in blastic transformation) and one ALL . We used a method in which the suspension of bone marrow cells was incubated in TC 199 medium with colchicine and with hypotonic solution formed from glycerol, NaCl, KCl, CaCl2, MgCl2 and sucrose . The slides were prepared from this cell suspension by cytospin and stained for peroxidase, PAS, esterases and iron . The karyotype was studied by direct method and culture . It was possible to relate the cytogenetic marker with cytochemistry characteristics in the same cell in 3 cases, showing the feasibility of cytochemistry techniques in cytogenetical preparations . The best preparations were found through peroxidase . The presence of iron granules allowed identification of erythroblastic lineage in the combined staining . Mitosis with a marker chromosome of leukemic clone in an AML cell with negative peroxidase probably showed a proliferation of more primitive precursor not sufficiently differentiated to show markers. FEBS Lett, 1992 Apr 27, 301(3), 287 - 90 Do submaximal InsP3 concentrations only induce the partial discharge of permeabilized hepatocyte calcium pools because of the concomitant reduction of intraluminal Ca2+ concentration? Combettes L, Claret M, Champeil P. In several types of cells whose cytoplasmic Ca2+ is regulated by inositol phosphate derivatives, low concentrations of InsP3 added to permeabilized cell suspensions induce the rapid discharge of part of the InsPs-sensitive Ca2+ pool instead of slow monophasic release of Ca2+ from the entire pool . As a tentative explanation for this puzzling observation, sometimes called 'quantal release', it was suggested that the reduced intraluminal Ca2+ concentration remaining in the Ca2+ pool after a certain amount of Ca2+ had been released might allosterically reduce the channels' affinity for InsP3 and the corresponding InsP3-dependent Ca2+ efflux, and thus result in partial pool discharge (Irvine, R.F . (1990) FEBS Lett . 263, 5-9) . We have tested this hypothesis by manipulating the Ca2+ pool contents with ionophore, and found that the rate of InsP3-dependent Ca2+ efflux after ionophore-induced partial discharge of the Ca2+ pools was much faster than what was predicted on the basis of this hypothesis . Heterogeneity of the Ca2+ pools appears to be a more likely reason for the 'quantal release' behavior. Biochem J, 1992 Apr 15, 283 ( Pt 2), 347 - 54 Differential pathways (phospholipase C and phospholipase D) of bradykinin-induced biphasic 1,2-diacylglycerol formation in non-transformed and K-ras-transformed NIH-3T3 fibroblasts . Involvement of intracellular Ca2+ oscillations in phosphatidylcholine breakdown; Fu T et al.; Bradykinin (BK) induced a biphasic increase in 1,2-diacylglycerol (DAG) in both K-ras-transformed fibroblasts (DT) and the parent NIH-3T3 cells . The first phase was coincident with the increase in Ins(1,4,5)P3 resulting from PtdIns(4,5)P2 hydrolysis, and the second, sustained, phase was derived from phosphatidylcholine (PtdCho) hydrolysis . In NIH-3T3 cells, stimulation by BK induced greater production of choline than phosphocholine in {3H}choline-labelled cells and appreciable phosphatidylethanol (PtdEtOH) formation in {3H}myristic acid-labelled cells, suggesting that PtdCho was hydrolysed mainly by a phospholipase D (PLD) activity . Pretreatment with propranolol, an inhibitor of phosphatidate phosphohydrolase, markedly diminished the second DAG accumulation, supporting the above notion . In DT cells, BK induced predominantly phosphocholine generation and little PtdEtOH formation, indicating that the PtdCho hydrolysis was due to a phospholipase C (PLC) activity . The BK-induced oscillations in intracellular Ca2+ concentration ({Ca2+}i) observed in single DT cells {Fu, Sugimoto, Oki, Murakami, Okano & Nozawa (1991) FEBS Lett . 281, 263-266} were detected as a sustained {Ca2+}i elevation when assayed in a cell suspension . A receptor-operated Ca2+ channel blocker, SK&F 96365, suppressed both the BK-induced phosphocholine generation and the sustained {Ca2+}i elevation in a similar dose-dependent manner . These results thus suggested that oscillations in {Ca2+}i are involved in the activation of PtdCho-specific PLC in DT cells. Biochem Biophys Res Commun, 1992 Apr 15, 184(1), 36 - 42 Direct measurement of nitric oxide in headspace gas produced by a chicken macrophage cell line in a closed culture system; Sung YJ et al.; A simple and rapid method was applied for direct measurement of nitric oxide (NO) gas produced by cultured macrophages using a modified chemiluminescence detector, the thermal energy analyzer (TEA) . HD11 chicken macrophages (1-3 x 10(6)/ml) were cultured on microcarrier beads (100 mg/ml) in 140 ml air-tight glass jars (5 ml cell suspension per jar) containing 0.5 micrograms/ml of LPS and different concentrations of L-arginine . Headspace gas was sampled at 24 hours of culture via a rubber septum and directly injected into a TEA with a liquid nitrogen trap set at -130 to -140 degrees C . The concentration of NO in the gas sample was quantified using a standard gas mixture of NO (2 microliters/L) in nitrogen . Gas samples from L-arginine-supplemented cultures contained NO (0.028-0.066 pl/microliter), whereas NO was not detected in samples from controls . These results suggest that chicken macrophages synthesize NO gas in a dose-dependent manner relative to L-arginine concentration. Blood, 1992 Apr 15, 79(8), 2141 - 7 Deformability measurements on individual sickle cells using a new system with pO2 and temperature control; Itoh T et al.; Although the rheologic behavior of sickle erythrocytes (SS cells) is highly dependent on oxygen tension (pO2) and temperature, very little data exist regarding the effects of deoxygenation and reoxygenation on the rheology of "individual" SS cells at body temperature . We have devised and assessed a new experiment system, in which micropipette aspiration can be performed on individual cells in a constant-temperature chamber that has ports for changing media with different pO2 (effected in 30 to 120 seconds) and sensing probes for monitoring pO2 and temperature . This system enabled us to simultaneously alter and monitor pO2 at 37 +/- 0.5 degrees C, and to monitor and study a single cell under microscopic observation . The static rigidity (E) and dynamic rigidity (eta) of individual SS cells were determined by repeated aspirations of the same cell under various pO2 . With stepwise reductions in pO2, E and eta showed no significant changes before sickling, but once sickled, their values markedly increased by 10(2)- to 10(3)-fold concomitantly with morphologic alteration of the cell . Thus, the deformability of a single SS cell behaves in an "all or none" manner at a critical pO2, and earlier studies on the effect of deoxygenation on the rheology of SS cell suspensions probably reflect the overall behavior of SS cells with widely distributed critical pO2. Cancer, 1992 Apr 1, 69(7), 1858 - 64 Modulation of cell cycle kinetics in human cancer with total parenteral nutrition; Frank JL et al.; Prior DNA flow cytometric data from the laboratory of the Division of Surgical Oncology, Massey Cancer Center, demonstrated an increase in the hyperdiploid compartment of tumor cells taken from patients with squamous cell carcinoma of the head and neck after a course of total parenteral nutrition (TPN) . To assess a putative increase in the percentage of tumor cells actively synthesizing DNA in this system, the authors administered bromodeoxyuridine (BrdU) intravenously to ten patients before and after the administration of TPN . Cell suspensions prepared from biopsy specimens of normal oral mucosa and tumor tissue were analyzed with flow cytometric study . Before TPN administration, the mean percentage of tumor cells incorporating BrdU was 2.47 +/- 1.11% . After TPN administration, the percentage of S-phase cells increased significantly (P less than 0.05) to a mean of 4.52 +/- 2.67% . Before TPN was given, normal mucosa demonstrated a mean of 7.97 +/- 2.69% of cells incorporating BrdU . After TPN was given, a mean of 8.47 +/- 2.51% was seen (not significant {NS}) . A potential strategy for the use of TPN to enhance tumor cell susceptibility to S-phase-specific chemotherapy is strongly suggested by these data. Nucleic Acids Res, 1992 Apr 11, 20(7), 1517 - 22 cDNA nucleotide sequence and expression of a tobacco cytoplasmic ribosomal protein L2 gene; Marty I et al.; The ribosomal protein L2 is an essential component of the ribosomal large subunit by its relation to the peptidyl transferase reaction, subunit association and elongation factor G-GTP binding . We have isolated a 937 nucleotide long cDNA encoding a cytoplasmic ribosomal L2 protein . Its deduced protein contains 260 amino acid residues and shows 65% identity with eucaryotic RL2 but only 32% identity with the chloroplast homologue . In addition, the protein presents the PROSITE signature which matches all the 50S and 60S L2 proteins and the two residues involved in the peptidyl transferase activity . The corresponding mRNA is accumulated in young plant tissues, in growing cell suspension and in germinating seeds but is not detectable in mature plant tissues, stationary cell suspension and in dry seeds . The mRNA accumulation is correlated with the growth process . Southern blot hybridization shows that cytoplasmic ribosomal protein L2 is encoded by two types of gene which could originate from each parent . highly homologous L2 genes were also detected by Southern blots in the genomes of several monocot and dicot plant species. Plant Mol Biol, 1992 Apr, 18(6), 1121 - 31 Coordinated regulation of two indole alkaloid biosynthetic genes from Catharanthus roseus by auxin and elicitors; Pasquali G et al.; Catharanthus roseus (periwinkle) produces a wide range of terpenoid indole alkaloids, including several pharmaceutically important compounds, from the intermediate strictosidine . The complete mRNA sequence for the enzyme strictosidine synthase (SSS) was determined . Comparison of the primary structure of the encoded protein with the amino-terminal sequence of purified SSS indicated the presence of a signal peptide of 31 amino acids in the putative primary translation product . SSS is encoded by a single-copy gene indicating that isoenzymes reported by others are formed post-translationally from a single precursor . The sss gene and the tryptophan decarboxylase gene (tdc), encoding another enzyme essential for indole alkaloid biosynthesis, are coordinately regulated . In plants steady-state mRNA levels are highest in roots . In cell suspension cultures the genes are rapidly down-regulated by auxin . In contrast, both genes are strongly induced by fungal elicitors such as Pythium aphanidermatum culture filtrate or yeast extract . Induction is a rapid, transcriptional event occurring independent of de novo protein synthesis . These results show that a first important regulatory step in the complex process leading to indole alkaloid accumulation in C . roseus suspension cells is transcription of the biosynthetic genes. Gan To Kagaku Ryoho, 1992 Apr, 19(4), 445 - 50 {Pharmacokinetics and action mechanism of anthracyclines}; Fukushima T et al.; For the purpose of establishing a method for reasonable clinical use of anthracyclines in leukemia chemotherapy, we examined the pharmacokinetics and the mode of action with five anthracyclines such as daunorubicin (DNR), doxorubicin (DOX), aclarubicin (ACR), THP adriamycin (THP), idarubicin (IDA) . In the patients with AML, blood ACR or IDA level increased and then disappeared very rapidly after iv bolus injection . In contrast, their metabolites (M1 or IDAol) increased for up to 2 or 4 hrs and remained much longer than ACR or IDA . The concentration of ACR, IDA or their metabolites were found to be much higher in the leukocyte fraction than in erythrocyte fraction or plasma . In HL60 cell suspension, anthracyclines were rapidly accumulated into the cells, and the uptake of IDA or THP were higher than the other agents . In HL60 cells, anthracyclines accumulated in the nuclear fraction but ACR was accumulated markedly in the cytosol fraction . From the result of DNA binding assay, binding at excess to calf thymus DNA of ACR was suggested to be approximately 2 times higher than that of other agents . DNA strand brakes in HL60 cells treated with anthracyclines were shown by pulse field gel electrophoresis, and IDA was found to have stronger activity to cause the DNA strand breaks . In conclusion, it seemed that anthracyclines showed similar action mechanisms, but in some respects quantitative differences were existed among them . Anthracyclines should be given to patients based on their pharmacological characteristics to obtain higher remission rate and suppress resistant cells. J Histochem Cytochem, 1992 Apr, 40(4), 541 - 53 Production of monoclonal antibodies against a novel glycoprotein synthesized and secreted by dog thyroid C-cells; Kameda Y; A monoclonal antibody (MAb) that reacted only with thyroid C-cells was raised against cell suspensions from dog thyroid glands, to examine a glycoprotein secreted by C-cells . After chronically-induced hypercalcemia and administration of an anti-thyroid drug, reaction products for the antibody markedly decreased in C-cells, coinciding with alterations in calcitonin immunoreactivity . The antigen recognized by the MAb appears to be a secretory protein . The MAb reacted with C-cells from a wide variety of mammalian species, including rats, mice, hamsters, cattle, cats, rabbits, and monkeys . Furthermore, tumor cells of human medullary thyroid carcinoma, which is derived from C-cells, were immunoreactive to the MAb . Exceptionally, C-cells from guinea pigs and pigs were not stained with the MAb . No crossreactivity was observed in any of the dog tissues examined . Immunoblot analysis demonstrated that the MAb recognized a single prominent band at a molecular weight of approximately 79,000 . The 79 KD band reacted with various digoxigenin-labeled lectins, including GNA, DSA, SNA, and MAA; it is a glycoprotein containing mannose, N-acetylglucosamine, and sialic acid . Dog thyroid C-cells were also densely stained with these lectins . The results indicate that thyroid C-cells synthesize and secrete a specific glycoprotein in addition to peptide hormones. Cancer Res, 1992 Apr 1, 52(7), 1933 - 7 Formation of PC12 tumors after transplantation into rat brains: dependence of time course on host age; Hatton JD et al.; Rat pheochromocytoma PC12 cells form tumors when placed into the brains of Sprague-Dawley rats under specific conditions . We now show that tumorigenic potential is regulated by the microenvironment of the developing cerebrum . PC12 cell aggregates were identified in the periventricular or intraventricular spaces within 24 h after injection of cell suspensions into rat brains . In fetal or young neonatal (1-4-day-old) recipient rat brains, these cell aggregates formed large masses within 21 days . The tumor incidence declined in recipient neonates between the ages of 5 and 8 days . In both cases, tumors spread throughout the ventricular system and subarachnoid and Virchow-Robin spaces as they grew . In contrast, tumors were not generated by injections into adult rat brains or by placement of PC12 cell pellets into preformed cavities . Despite the loss of tumorigenicity, surviving cells were present at the injection site . The presence of surviving cells and the ability of another rat cell line (the C6 rat glioma line) to form tumors in adult rat brains suggest that an immune response is not solely responsible for the lack of PC12 tumorigenicity in adult rat brains . We propose that developmentally increasing local concentrations of specific factors (e.g., nerve growth factor of fibroblast growth factor) may also contribute to the suppression of tumor formation in this system. Cancer Res, 1992 Apr 1, 52(7), 1737 - 43 Fate of clonal lineages during neoplasia and metastasis studied with an incorporated genetic marker; Moffett BF et al.; The fate of clonal lineages in tumor formation and metastasis has been studied by genotypic marking of cells from three separate tumor lines of different malignant potential . Marking was accomplished by random incorporation of the neomycin resistance gene and visualized by Southern blot analysis of integration sites . Primary tumors formed by polyclonal cell suspensions of all three cell lines injected s.c . usually remained polyclonal even at late stages of tumor growth and metastatic spread . Lung metastases were often clonal, but it was not unusual to find ones of polyclonal origin . Lymph node metastases were almost always polyclonal and remained so, as they grew large . Sometimes clones present in the original inoculum were absent in the primary . Other times clones visible in the metastases were undetectable in the corresponding primary tumor . Occasionally a single clone became dominant in the primary, and others were eliminated, but this was not a necessary prelude to the onset of invasive or metastatic behavior . It is concluded that there is considerable variation in the results obtained with various cell lines in different circumstances . Even clones which are underrepresented in the original inoculum or the primary tumor can acquire metastatic capability . Hence, progression of malignancy is not uniformly dependent on prior or concurrent extinction of other non- or less metastatic clones in the neoplasm, and the underlying mechanisms of invasion and metastasis can be separated from those which sometimes confer growth supremacy on a clone of tumor cells . The frequent continuing genetic heterogeneity of cells in a neoplasm has substantial implications for clinical treatment protocols. Cancer, 1992 Apr 1, 69(7), 1745 - 9 Detection of a soluble antigen defined by monoclonal antibody 83D4 in serous effusions associated with breast carcinoma; Osinaga E et al.; Monoclonal antibody (MoAb) 83D4 was generated by immunization with cell suspensions obtained from sections of formol-fixed paraffin embedded human breast cancer . It recognized an antigen expressed in breast carcinomas but not in normal breast tissue . Pleural and ascitic fluids from 66 patients were studied by an 83D4 heterologous sandwich radioimmunoassay (SRIA) using solid-phase immobilized wheat germ agglutinin to detect the 83D4 soluble antigen . Using a cutoff level of 5 units/ml of 83D4 antigen, higher values were found in 22 of 27 breast cancer-associated effusions (mean = 10.72 +/- 6.80 units/ml) . The 20 nonmalignant effusion fluids tested showed lower values (mean = 1.16 +/- 1.49 units/ml, P less than 0.001) . The antigen was undetectable or present in low levels in effusions from patients with hematologic malignancies . When SRIA results were compared with conventional cytologic diagnosis in breast-cancer effusions, elevated levels of 83D4 soluble antigen were found in all patients (8 of 8) in whom malignant cells had been detected, in 4 of 8 patients with the diagnosis of "suspected malignancy," and in 10 of 11 patients with negative cytologic findings . Using an immunoglucosidase method on cell smears of various origins, MoAb 83D4 stained metastatic cells of breast and ovary carcinomas but did not reactive with mesothelial cells and other normal or malignant cell types . These results suggest that quantitation of the 83D4 soluble antigen may be used to improve the diagnosis of cancer in serous effusions. Arch Biochem Biophys, 1992 Apr, 294(1), 306 - 13 Catabolism of camphor in tissue cultures and leaf disks of common sage (Salvia officinalis); Funk C et al.; (+)-Camphor constitutes nearly 30% of the monoterpenes accumulated in the leaves of common sage (Salvia officinalis), and as the plant approaches maturity the content of this monoterpene ketone decreases by roughly half . Although the ability to catabolize camphor has been demonstrated previously in sage leaf disks, tissue cultures proved to be a more suitable system for examining the responsible degradative pathway . Cell suspension cultures were shown to convert (+)-{3-3H2}camphor, in sequence, to 6-hydroxycamphor, 6-oxocamphor, alpha-campholonic acid, and 2-hydroxy-alpha-campholonic acid, and each intermediate of the pathway was identified by chromatographic and spectroscopic means . This oxidative ring opening sequence resembles the pathway for camphor degradation by the soil diphtheroid, Mycobacterium rhodochrous, that ultimately leads to isoketocamphoric as the last defined metabolite that contains all 10 carbons of the original bicyclic nucleus . Studies with both cell cultures and leaf disks also demonstrated that the catabolism of camphor via 1,2-campholide, a metabolite in sage leaves previously described, was a minor degradative pathway . The first step in the metabolism of camphor was demonstrated in cell-free extracts of the cultured sage cells, and several lines of evidence indicated that this microsomal (+)-camphor-6-exo-hydroxylase is a cytochrome P-450-dependent monooxygenase. J Invest Dermatol, 1992 Apr, 98(4), 450 - 8 In situ detection of supernumerary aberrations of chromosome-specific repetitive DNA targets in interphase nuclei in human melanoma cell lines and tissue sections; de Wit PE et al.; The use of non-radioactive in situ hybridization (ISH) with chromosome-specific repetitive DNA probes to study genomic changes, aneuploidy, and heterogeneity during melanocytic tumor progression, relies on its applicability to non-mitotic interphase nuclei, present in cell suspensions and tissue sections . Therefore, we studied the feasibility of detecting numerical aberrations with respect to the (peri-) centromere regions of chromosomes 1 and 7 in intact nuclei of two human melanoma cell lines with different metastatic behavior in nude mice . In addition, we used paraffin sections from xenograft lesions, obtained by inoculation of these cell lines in nude mice (subcutaneous tumors and spontaneous lung metastases) . Paraffin sections from the original primary cutaneous melanoma (with a subepidermal and a dermal part) and two loco-regional metastases were also studied, one of which was the source for the cell lines . These cells and tissues represent examples of materials used in different approaches to the study of melanocytic tumor progression . Regarding the targeted sequences, ISH analysis showed that both cell lines were heterogeneous and aneuploid . The results correlated well with those obtained by ISH on metaphase spreads . Differences between the lines, which could not be detected by flow-cytometric or conventional karyotyping analysis, included data suggestive of a polyploid subpopulation and an extra copy of chromosome 7 in the metastasizing cell line . The polyploid population could be detected also in the paraffin sections of the corresponding subcutaneous xenografts and lung metastases in the mice . Both areas in the patients' primary melanoma could be evaluated separately and showed similar supernumerary aberrations of the chromosome-specific targets . These abnormalities matched those found in both metastases . Our results demonstrate that ISH can be used to visualize genomic abnormalities at the single-cell level in melanocytic nuclei in their natural context, which makes it a promising tool in the histopathology of melanocytic lesions and in the study of melanocytic tumor progression. Biochem J, 1992 Apr 1, 283 ( Pt 1), 265 - 72 Gluconeogenesis stimulated by extracellular ATP is triggered by the initial increase in the intracellular Ca2+ concentration of the periphery of hepatocytes; Koike M et al.; Extracellular ATP, ADP and GTP increased the intracellular free Ca2+ concentration ({Ca2+}i) in a suspension of isolated rat hepatocytes . The {Ca2+}i was determined by measuring fura-2 fluorescence, and its increase was biphasic . The initial transient rise was followed by a longer-lasting plateau . The peak of the early component preceded the plateau level of the second component . A time course of change in {Ca2+}i in single cells at 100 microM-ATP was very similar to that observed in the suspension system . Preincubation of hepatocytes with 40 mM-caffeine, 2 mM-oxalate or 60 microM-dantrolene sodium inhibited the P2 purinergic response . The plateau phase was not observed when measured in the presence of extracellular 100 microM-LaCl3 or in the absence of extracellular Ca2+ . The distribution of {Ca2+}i in single hepatocytes was also determined by fluorescence image analysis . In the initial phase, the increase in {Ca2+}i is greater in the peripheral region than the central region of the cell . Degradation of extracellular ATP by ecto-ATPase in the hepatocyte suspension was measured; the amount of ATP degradation was less than 10-15% of the initial amount (100 microM) during the measurement of the intracellular {Ca2+}i in the cell suspension . Extracellular ATP stimulated glucose synthesis . The rate of glucose production also showed two components, the initial fast component within 1 min and the subsequent slower component . The rate of the initial fast component did not depend on the presence or absence of extracellular Ca2+, whereas the rate of the subsequent component depended on it . The present study shows that the initial transient rise in {Ca2+}i plays an important role in triggering the gluconeogenesis. Scand J Immunol, 1992 Apr, 35(4), 459 - 68 Phenotypical and functional characterization of small intestinal TcR gamma delta + T cells in coeliac disease; Rust C et al.; Increased numbers of TcR gamma delta + T cells are present in the small intestinal epithelium of patients with coeliac disease (CoD) . Their function, however, is unknown . In order to facilitate detailed functional studies, intestinal gamma delta T cells have been isolated from small intestinal biopsies of patients with CoD (n = 18) and controls (n = 14) . As expected, increased numbers of V delta 1+ TcR gamma delta + T cells were detected in freshly isolated intraepithelial cell suspensions (IEL) from CoD patients . Also, in the in vitro expanded IEL T-cell populations from CoD patients the numbers of V delta 1+ TcR gamma delta + T cells were increased compared with similar cell cultures from control patients . From IEL cultures derived from six CoD patients, 107 T-cell clones were generated by limiting dilution and analysed . Sixty of these clones were either CD4 or CD8 positive TcR alpha beta + clones . The remaining 47 clones expressed the TcR gamma delta . Further phenotypical analysis of the gamma delta T-cell clones indicated that the TcR gamma delta + T-cell population in the small intestinal epithelium of CoD patients is heterogeneous: four TcR gamma delta phenotypes could be detected and, although the majority of the TcR gamma delta + T cells were CD4 CD8, gamma delta T-cell clones expressing either a CD8 alpha alpha homodimer, a CD8 alpha beta heterodimer or CD4 were also identified . In contrast to the TCR alpha beta + IEL, most TcR gamma delta + IEL were CD5 negative . Furthermore, biochemical analysis indicated that the increase in V delta 1+ gamma delta T cells in the small intestinal epithelium of CoD patients was not the result of a monoclonal expansion . The small intestinal epithelium-derived gamma delta T-cell clones were functional in vitro since the majority of these clones were able to lyse target cell lines such as K562 . Molt4 and Daudi . These novel findings therefore indicate that the gamma delta T cells in the small intestine of CoD patients represent a heterogeneous population and that such cells are functional in vitro . The isolation and the in vitro propagation and cloning of these cells may open new avenues for the study of the putative immune mechanisms leading to coeliac disease. Photodermatol Photoimmunol Photomed, 1992 Apr, 9(2), 58 - 60 In vitro kinetics of 8-methoxypsoralen penetration into human lymphoid cells; Karolak L et al.; Extracorporeal photochemotherapy (ECPC) requires ex vivo UVA irradiation of blood lymphocytes during the time of the theoretical peak 8-methoxsalen (8-MOP) concentration . The aims of this study were to determine the mechanism of cellular uptake of 8-MOP, its possible saturation and the time needed to reach maximal concentration (Tmax) in lymphoid cells . 8-MOP was measured by liquid chromatography in the supernatant of lymphoid cell suspensions incubated with a known amount of 8-MOP . The kinetics of cellular uptake were determined and showed that equilibrium had already been reached after 2 min and remained constant for at least 60 min . The uptake was independent of temperature (4, 25 and 37 degrees C) and was proportional to the 8-MOP concentration in the supernatant . This indicated that 8-MOP penetrated into lymphoid cells by passive diffusion, rather than by active transport or facilitated diffusion, and was thus a non-saturable process . In addition, intracellular metabolism was negligible . These findings demonstrated that the plasma and lymphocytic Tmax were reached simultaneously and statistical analysis showed them to be significantly correlated, thereby validating the standard ECPC protocol for drug ingestion and lymphocyte irradiation. Boll Soc Ital Biol Sper, 1992 Apr, 68(4), 259 - 62 Serum levels of soluble interleukin-2 receptor (sIL-2R) in B-chronic lymphocytic leukemia; Cacciola E et al.; By using an enzyme-linked immunosorbent assay, the levels of the soluble form of the interleukin-2 receptor (sIL-2R) were evaluated in the peripheral blood of 20 patients with cell chronic lymphocytic leukemia in different stages of disease and in supernatants obtained from enriched B cell suspensions . In either all serum samples or in one out of three supernatants, elevated levels of sIL-2R were found . This could indicate that the B leukemic cells release sIL-2R which in turn, for its potential capacity of binding circulating IL-2, could contribute to the abnormal immunoregulation which characterizes B-CLL . This finding, which needs further investigation, could have prognostic significance. Biosci Rep, 1992 Apr, 12(2), 123 - 33 Neutrophil function in whole blood and after purification: changes in receptor expression, oxidase activity and responsiveness to cytokines; Watson F et al.; Neutrophil function and plasma membrane receptor expression was measured in cell suspensions isolated by two separate procedures and in unfractionated whole blood . When cells were prepared by a combined dextran/ficoll procedure, their ability to generate reactive oxidants in response to fMet-Leu-Phe was greater than in corresponding cells isolated by a one-step procedure on Mono-Poly Resolving Medium (M-PRM) . Cells prepared by both methods could be primed in vitro by rGM-CSF, but the priming ratio was greater in cells prepared by the latter method . The ability of neutrophils in whole blood to generate reactive oxidants in response to fMet-Leu-Phe was extremely low, but this was increased by more than 10 fold if the blood was pre-incubated with rGM-CSF . Similarly, expression of CD 11b and CD 16 was very low (or undetectable) in neutrophils in whole blood, but this was rapidly increased upon priming . Activation by PMA resulted in a down regulation of CD 16 expression as the receptor was shed from the cell surface . Neutrophils isolated by either the dextran/ficoll or the M-PRM method showed increased expression of receptors compared with those in whole blood, although this expression was lower in cells isolated by the latter method . These data indicate that the isolation procedures used to obtain purified neutrophils prime both receptor expression and oxidase function, although these effects are minimalised in isolation procedures using M-PRM . Furthermore, as CD 16 expression on neutrophils in whole blood is rapidly up-regulated during priming, it seems likely that, as for complement receptors, rapidly-mobilisable intracellular stores of this receptor exist. Biull Eksp Biol Med, 1992 Apr, 113(4), 421 - 4 {Obtaining and morphological characteristics of primary monolayer cell cultures of human somatotropinomas}; Komolov IS et al.; This paper describes the development of method of preparing cell suspension obtained from surgical material of patients with pituitary adenomas and acromegaly as well as the procedure of subsequent long-term cultivation in monolayers of the cells isolated . As judged by visual inspection and measurement of growth hormone and prolactin secretion, tumor pituitary cells kept viability and functional activity for at least 6 days of growing in vitro . Immunocytochemical visualization of somato- and lactotrophs of the same histological preparations permitted us to show that vast majority of cultured cells is represented by somatotrophs; however, a small portion of cell population is represented by lactotrophs and lactosomatotrophs . The peculiarities of cytoarchitectonics in two types of cell cultures of human somatotropinomas were studied. Br J Urol, 1992 Apr, 69(4), 392 - 6 Circulating prostate specific antigen-positive cells correlate with metastatic prostate cancer; Hamdy FC et al.; Analytical flow cytometry was used to study circulating prostate specific antigen (PSA)-positive cells in 40 consecutive patients with newly diagnosed, untreated prostate cancer; 25 patients (63%) had metastatic disease confirmed by a positive bone scan . Cell suspensions were prepared for each patient from both the primary tumour and peripheral blood samples . The cells were stained with a monoclonal antibody against PSA, and analysed by flow cytometry; PSA-positive cells were sorted according to their immunofluorescence and light scatter properties . The cellular deoxyribonucleic acid (DNA) content of each specimen was also analysed to establish ploidy status . PSA-positive cells were detected in the peripheral blood of 33 patients (83%) . The presence of these cells in the circulation showed a higher degree of sensitivity and specificity in predicting positive bone scans than did serum PSA levels . Circulating PSA-positive cells may represent either a subpopulation of tumour cells with distinct metastatic properties or, alternatively, host immunocytes which take up PSA in an active or passive manner. Blood, 1992 Apr 1, 79(7), 1695 - 703 Dynamics and localization of early B-lymphocyte precursor cells (pro-B cells) in the bone marrow of scid mice; Osmond DG et al.; Mice homozygous for the scid (severe combined immunodeficiency) mutation are generally unable to produce B lymphocytes, a condition attributed to defective rearrangement of immunoglobulin genes in precursor B cells . Some early B-lineage cell