Ann N Y Acad Sci, 1987, 495, 169 - 84 Functional activity of raphe neurons transplanted to the hippocampus and caudate-putamen . An immunohistochemical and neurochemical analysis in adult and aged rats; Steinbusch HW et al.; Adult (3-month-old) and aged (28-month-old) rats that had been pretreated with 5,7-DHT in both lateral ventricles received grafts of cell suspensions taken from the RR or MR regions taken from the embryonic stages E12-21 . These cell suspensions were implanted unilaterally into the rostral part of the hippocampus or the caudate-putamen for immunohistochemical and neurochemical studies . MR and RR cell suspensions have the potency to regenerate serotonergic fibers in the previously denervated adult and aged hippocampus and caudate-putamen . The RR cell suspension, however, also showed outgrowth of other transmitter-specific neuronal systems, specifically noradrenaline and substance P . To evaluate the functional activity of the serotonergic reinnervation, we have combined immunohistochemistry and neurotransmitter release studies on adjacent hippocampus slices of adult rats . Results showed that after a survival time of 10 weeks, the serotonergic innervation of the hippocampus was greatly restored and, moreover, that the K+-induced Ca2+-dependent release of 5-HT amounted to about 80% of normal values . There appeared to be a striking similarity between the immunohistochemical and neurochemical data regarding the increase in the number of newly formed serotonergic fibers, the increase of the release of radiolabeled 5-HT, and the extent of the outgrowth in the hippocampus. J Periodontol, 1987 Jan, 58(1), 30 - 3 Biologic preparation of diseased root surfaces . An in vitro study; Assad DA et al.; The study evaluated the effect of 2% sodium desoxycholate combined with whole plasma from a single donor on gingival fibroblast attachment to diseased root surfaces . Twenty extracted periodontally involved teeth were cut into halves buccolingually and sterilized by moist heat under high pressure . The diseased root surface of each control half was rubbed with a sterile cotton pellet saturated with Dulbecco's phosphate-buffered saline (PBS) . The diseased root surface of each experimental half was rubbed with a sterile cotton pellet saturated with 2% sodium desoxycholate and then rubbed with a pellet soaked in human plasma . The control and experimental halves were placed in separate petri dishes, and a fibroblast cell suspension was added to each dish . The mean number of attached cells per half tooth was calculated for each group . Tooth surfaces treated with PBS (controls) showed a mean of 307 +/- 63 attached cells for 17 tooth halves; the experimental treated surfaces exhibited a mean of 650 +/- 130 attached cells for 16 halves . The difference between these numbers was statistically significant (P less than 0.01) . These findings suggest that the desoxycholate/plasma combination enhanced in vitro fibroblast attachment to diseased root surfaces. Arkh Patol, 1987, 49(12), 31 - 7 {Morphological changes in splenic autotransplants following splenectomy: clinical and experimental studies}; Sapozhnikova MA et al.; Reports results of clinical, morphologic, and laboratory studies carried out in 18 patients allografted with spleen fragments into the greater omentum following splenectomy for a traumatic injury of the spleen, and in 20 splenectomized dogs allografted with spleen fragments or a spleen cell suspension into various parts of the abdominal cavity . Spleen tissue regeneration in the allografts was found to be nearing completion 6 to 8 weeks after the transplantation . The regeneration took less time to be completed in dogs grafted with spleen fragments than in those grafted with a cell suspension, but occurred at similar rates whatever the site in the abdominal cavity into which the fragments or suspension had been grafted . In the allografted patients, the incidence of posttransplantation suppurations was lower than in a similar group of patients who had undergone splenectomy without allografting. Urol Int, 1987, 42(5), 326 - 9 Local bone resorption induced by serially transplantable human renal cell carcinoma in nude mice; Nemoto R et al.; The high incidence of metastatic bone disease in urological cancer makes it necessary for clinicians to look for a valid experimental model to investigate the basic interactions between cancer cells and bone in order to improve the treatment . A new model of bone resorption was defined, namely the subcutaneous injection of tumor cell suspensions of serially transplanted renal cell carcinoma in nude mice after disruption of the periosteum of the calvaria . The tumor induced osteolysis associated with osteoclast proliferation with reactive bone formation . The X-p film from nude mice calvaria showed the same multiple osteolytic punched-out lesions as those of the patient's skull. Symp Soc Exp Biol, 1987, 41, 363 - 78 Mechanisms of freezing damage; Pegg DE; Freezing of aqueous systems involves numerous simultaneous changes but this review concentrates on direct effects of the formation of ice and the consequent concentration of solutes in the remaining liquid phase . It is generally believed that cell injury at low cooling rates is principally due to the concentration of both intracellular and extracellular electrolytes and that cryoprotectants act by reducing this build-up . New experimental data are presented to support this explanation; we find that the extent of damage to human red blood cells during freezing in solutions of sodium chloride/glycerol/water can be quantitatively accounted for by the increase in solute concentration . However, we also show that a given degree of damage occurs at lower concentrations of solute in the presence of higher concentrations of glycerol; it appears that glycerol contributes an element of damage itself . Recently published studies from Mazur's laboratory have suggested that the dominant damaging factor at low cooling rates is actually the reduction of the quantity of unfrozen water rather than the corresponding increase in salt concentration that accompanies freezing . These data are re-evaluated, and it is argued that the experimental results could equally well be explained by a susceptibility of cells to shrinkage and re-expansion as the concentration of external impermeant solutes first increases during freezing and then decreases during thawing . It is concluded that external ice probably has no directly damaging effect upon dilute suspensions of cells . However, it is also argued that ice is directly damaging whenever it forms intracellularly, and also when it forms extracellularly in densely packed cell suspensions . In the latter case the damage is probably due to recrystallization of the ice masses during thawing . Extracellular ice also has a directly damaging effect when tissues and organs are frozen . The difficulties of designing experimental methods that will yield unequivocal results is emphasized, and consequently the above conclusions must be regarded as tentative at the present time. Cancer Invest, 1987, 5(6), 593 - 611 Applications of immunocytochemistry to clinical cytology; Johnston WW et al.; This article reviews the recent studies reporting the applications of immunocytochemistry to diagnostic problems in clinical cytology . A series of studies with monoclonal antibody (MAb) B72.3 is discussed in detail . MAb B72.3, reactive with a high molecular weight, glycoprotein, tumor-associated antigen, designated TAG-72, has been shown previously to be reactive with formalin-fixed, paraffin-embedded tissue sections of adenocarcinomas of the ovary, colon, and breast, but not a variety of normal adult tissues . It has demonstrated utility as an immunocytochemical adjunct to diagnose carcinoma in cell block and cytocentrifuge preparations of human serous effusions, with selective reactivity for tumor cells (particularly adenocarcinoma) over reactive mesothelium . Using the avidin-biotin complex (ABC) method of immunoperoxidase staining and formalin-fixed, paraffin-embedded cell suspensions, MAb B72.3 detected tumor cells in effusions from the majority of patients with adenocarcinoma of the breast . No reactivity was demonstrated in any cell type in benign effusions . In contrast, MAb B72.3 showed no reactivity to leukemic or lymphomatous effusions, or to mesothelial cells from malignant effusions . MAb B72.3 also detected tumor cells in effusion specimens from most of the patients with "non-small cell" carcinoma of the lung and with carcinoma of the ovary . MAb B72.3 was also used with fine-needle aspiration biopsies (FNABs) and corresponding surgically excised tumors to determine cellular reactivity . Positive staining with MAb B72.3 was observed in needle aspirates of the great majority of "non-small cell" carcinomas of the lung, adenocarcinomas of the breast, adenocarcinomas of the colon, and carcinomas from other body sites . In contrast, small cell carcinomas of the lung, malignant melanomas, lymphomas, sarcomas, and glial tumors stained negatively with the antibody . Most benign lesions from the breast, lung, pancreas, parotid, and thyroid also showed no staining . In many patients, tumor-bearing tissue had also been resected and was available for comparative examination with MAb B723 . In more than 90% of these patients, the staining patterns of tumor cells in the aspirates were found to be predictive of the patterns of antibody reactivity in the comparable surgically resected tumors . From these studies it is concluded that MAb B72.3 defines a tumor-associated antigen that is expressed in neoplastic cells versus benign cells, is most selectively expressed in carcinomas, and may be used as a novel adjunct for the diagnosis of neoplasms in effusions and in FNABs. Graefes Arch Clin Exp Ophthalmol, 1987, 225(5), 351 - 6 Growth and characterization of rabbit corneal cells in vitro; Cook SD et al.; Primary organ cultures of rabbit corneal epithelium, stroma, and endothelium were established by microdissection . Secondary cultures of epithelial cells, keratocytes, and endothelial cells were established by serial passage . The doubling time for epithelial cells and keratocytes was 18 h, and endothelial cells doubled their number in 5 days . Ultrastructural studies demonstrated characteristic morphological, nuclear, and cytoplasmic features of corneal epithelial cells, keratocytes, and endothelial cells and confirmed the identity of the cell lines . The purity of the three distinct cell types was ascertained by indirect immunofluorescence techniques, using antibodies against keratin, which identified epithelial cells, and fibronectin, which identified keratocytes and endothelial cells . The indirect immunofluorescence technique represents a simple method to screen an aliquot of a cell suspension and determine the purity of corneal cells grown in vitro. Boll Ist Sieroter Milan, 1987, 66(2), 153 - 7 Mitotic activity and chromosomal findings in lymph nodes from patients with carcinoma or non-Hodgkin's lymphoma; Ballaban K et al.; We studied the chromosomal constitution of lymph node cells from two patients with non-Hodgkin's lymphoma, one patient with Crohn's disease, and four patients with carcinoma, using regional lymph nodes that microscopically did not contain metastatic tumor . No consistent chromosomal abnormalities were detected and normal metaphases were predominant . We also studied the mitotic activity of lymph node cell suspensions from six patients with carcinomas . Mitoses were always more abundant after 3-4 days in culture, and were consistently higher in cultures to which phytohemagglutinin had not been added . No mitotic activity was seen in lymph nodes that histologically were shown to contain metastatic carcinoma . Possible implications of these findings are discussed. Clin Lab Haematol, 1987, 9(3), 297 - 306 Comparison of immunofluorescence and immunoperoxidase techniques for demonstration of phenotype and monoclonality in lymphoproliferative disorders; Marsden KA et al.; To determine whether a simple immunoperoxidase (IP) technique could replace the traditional immunofluorescence (IF) technique (by fluorescence microscopy) for determination of phenotype and monoclonality in cell suspensions from patients with lymphoproliferative disorders, we tested samples from 48 cases by both methods, using small panels of antibodies designed to determine proportions of B-cells, T-cells and cells bearing immunoglobulin light chains . There were 27 cases of chronic lymphocytic leukaemia (CLL), 6 of prolymphocytic leukaemia (ProLL), and 15 of non-Hodgkin's lymphoma (NHL) . We found that IP was at least as reliable as IF in all cases . In the 47 B-cell cases, the mean proportion of cells apparently belonging to the malignant clone was higher by IP than by IF; conversely, the proportion of B-cells displaying the non-malignant light chain marker and the proportion of residual T-cells was apparently lower by IP . We conclude that IP is a useful and reliable technique for determining phenotype and monoclonality in lymphoproliferative disorders; because it is tolerant of delays to samples in transit and requires only simple facilities, it may be particularly suitable for small laboratories. Arch Immunol Ther Exp (Warsz), 1987, 35(1), 79 - 86 Bone marrow transplantation in Polish conditions . A modified method of marrow collection and preparation for transplantation; Jedrzejczak WW et al.; Marrow is collected from posterior iliac crests using specially developed "Wiktor's" needles and from other sites using commercially available sternal needles into heparinized syringes . Then it is directly injected into collecting flasks through specially designed bone marrow filters . This assures single cell nature of marrow cell suspension which then may be either directly infused into the recipient or processed for red cell or T cell depletion . During collection marrow cell suspension is mixed using magnetic stirrers . The described method allows collection of sufficient number of marrow cells for engraftment at the expense of acceptable risk for the donor . Data for 10 consecutive marrow collections are reported. Exp Brain Res, 1987, 69(1), 183 - 94 Dopamine neurons grafted unilaterally to the nucleus accumbens affect drug-induced circling and locomotion; Brundin P et al.; The purpose of the present study was to investigate the amplifying function of the nucleus accumbens septi region (NAS) in 6-hydroxydopamine (6-OHDA)-induced rotational behaviour by implanting fetal dopamine (DA)-rich mesencephalic cell suspensions unilaterally in the NAS of rats previously subjected to combined mesostriatal (MS) and NAS 6-OHDA lesions . First, all the rats received a unilateral 6-OHDA lesion of the ascending MS DA pathway, which produced an amphetamine-induced rotational asymmetry towards the lesioned side . In a second step, the rats received a local bilateral 6-OHDA lesion of the NAS which, as previously shown, caused a significant attenuation of the amphetamine-induced locomotor (1.5 mg/kg) and rotational (5 mg/kg) behaviour . Finally, some of these MS + NAS lesioned rats received a unilateral mesencephalic DA graft into the NAS ipsilateral to the original MS lesion . The unilateral DA-rich grafts in the NAS significantly elevated the amphetamine-induced locomotion and ipsilateral circling (opposite to the direction of rotation produced when a graft is placed in the ipsilateral caudate-putamen), suggesting that the NAS plays only an amplifier role in locomotor behaviour and not a directional role . In addition, these grafts significantly attenuated the supersensitive locomotor response observed in lesioned rats when given apomorphine (0.05 mg/kg) . The findings emphasize the amplifying role of the NAS in locomotion and circling behaviour and they extend previous findings demonstrating the functional heterogeneity of the striatal complex as well as the regional specificity of the graft-derived functional effects . Moreover, the results argue against the notion that DA grafts can function through a diffusion of transmitter over large distances since, despite the large size of the grafts, the functional graft effects were well localized to the reinnervated NAS and ventromedial striatal regions . We conclude, therefore, that graft-induced amelioration of postural and locomotor deficits are affected through different parts of the striatal complex, and that multiple graft placements are required to produce more complete recovery of motoric behaviour in the DA-depleted brain. Digestion, 1987, 36(4), 201 - 12 Prostaglandin protection against ethanol-induced gastric injury: regulatory effect on the mucus glycoprotein metabolism; Tsukada H et al.; Effects of ethanol and prostaglandin on the metabolic processes of gastric mucosal cells have been evaluated by studying the biosynthesis and intra- and extracellular distribution of mucus glycoprotein . The rat gastric mucosal cell suspensions were subjected to a short-term tissue culture in the presence of 0-1.5 M ethanol and 0-100 ng/ml 16,16-dimethyl prostaglandin E2 (DMPGE2), using {3H}proline and {3H}palmitic acid as markers of mucin apoprotein synthesis and modification . Ethanol at concentrations of 0.02-0.1 M stimulated the glycoprotein synthesis, but the product was 2-3 times more extensively and rapidly degraded by pepsin than the glycoprotein synthesized in the absence of ethanol . Higher concentrations of ethanol (0.1-1.5 M) caused a marked reduction in the glycoprotein synthesis and a depletion of the intracellular mucus glycoprotein stores, and at 1.5 M ethanol the synthetic processes ceased to function . When the incubated tissue cultures were challenged with DMPGE2, the highest synthetic and secretory stimulation was achieved at 10 ng/ml . The peptic susceptibility of the glycoprotein synthesized in the prostaglandin-enriched culture was somewhat lower than that of controls, and the proportion of the undegraded glycoprotein remaining after 22 h of digestion was significantly higher . Furthermore, the intracellular compartments were not depleted of mucus glycoprotein and the amount of the mucin apoprotein precursor increased to about 30% . Addition of DMPGE2 prior to ethanol treatment resulted in the stabilization of cellular processes and the cells responded as if ethanol was not present in the medium . Moreover, the intracellular stores of mucus glycoprotein were not depleted and the secretory rate was moderately elevated . The regulatory effect of DMPGE2 on the glycoprotein synthesis and distribution was significant, but diminished with increasing amounts of ethanol in the medium . The results suggest that DMPGE2 imposes a stabilizing effect on the secretory processes and controls the mucus glycoprotein synthesis, but the extent of this protective action against ethanol is limited and depends upon the concentration of ethanol penetrating the mucosal cells. Int Arch Allergy Appl Immunol, 1987, 82(3-4), 411 - 3 In vitro IgE synthesis induced by human T cell clones in normal B cells and its suppression by heterogenous T cell populations; Romagnani S et al.; Thirty-four out of 119 human T cell clones established from tonsillar or peripheral blood T cell suspensions of 3 nonallergic individuals were able to induce normal B cells to synthesize remarkable amounts of IgE in vitro . The activity of these clones was apparently mediated by triggering of the monomorphic molecular complex CD3 immediately before or during their incubation with the target B cells . The addition to the cultures of mitogen-stimulated autologous unfractionated T cells inhibited in a dose-dependent fashion, the T cell clone induced IgE, but not IgG, synthesis. Virchows Arch A Pathol Anat Histopathol, 1987, 410(5), 443 - 7 Evidence for neoplastic cell differentiation in mediastinal T lymphoblastic lymphoma; Ruco LP et al.; Immunostaining of frozen sections from a mediastinal T lymphoblastic lymphoma (T-LL) revealed the existence of two neoplastic cell populations characterized by different degrees of maturation . Several large nodules of 3A1+/T11+/T9+ T6-/T4-/T8-/T3- lymphoid cells, resembling normal early thymocytes, were surrounded by 3A1+/T11+/T9+/T6+/T4+/T8+/T3- cells resembling normal cortical thymocytes . The junctional area between early and cortical lymphocytes was occupied by numerous Leu-M3+/PAM-1+/DR+ reticular macrophages which were also characterized by J5 reactivity . Cytokeratin+/keratin+ epithelial cells were absent . Immunostaining of paraffin sections and of cytocentrifuge smears obtained from tumour cell suspensions revealed that a consistent percentage (8%) of neoplastic lymphoblasts were S-100+ . Our findings are consistent with a cortical T-LL presenting areas of dedifferentiated cells or, alternatively, with an early T-LL whose cells were able to differentiate into cortical thymocytes, perhaps through the interaction with a specialized subset of J5+ macrophages. J Am Acad Dermatol, 1987 Jan, 16(1 Pt 1), 61 - 74 Electron microscopic and immunolabeling studies of the lesional and normal skin of patients with mycosis fungoides treated by total body electron beam irradiation; Braverman IM et al.; Biopsy specimens were taken from lesional and normal skin of nine patients with mycosis fungoides before and after total body electron beam therapy . By electron microscopy, lesional skin had one and one-half to ten times as many epidermal Langerhans cells and indeterminate cells as did the normal skin . In successfully treated lesional skin 1 month after the end of electron beam therapy, the density of epidermal Langerhans cells and indeterminate cells had decreased markedly . In incompletely resolved lesions, Langerhans cells and indeterminate cells were still at pretreatment levels . Epidermal T6 and Ia antigens showed the same pattern of response . Epidermal cell suspensions from lesional and normal skin before and after electron beam therapy were assayed for epidermal thymocyte activating factor . The values of production of this factor did not correlate with the source of the epidermal cells, response to therapy, or the patient's disease course . Skin lesions resembling xerosis and parapsoriasis and histologically lacking the criteria for mycosis fungoides appeared during clinical remissions . These nonspecific skin lesions had densities of epidermal Langerhans cells, indeterminate cells, and T6-positive and Ia-positive cells comparable to levels found in pretreatment lesional skin. Folia Histochem Cytobiol, 1987, 25(1), 3 - 15 Cytochemical and functional evaluation of ACTH responsive isolated rat adrenocortical cells . The maintenance of sex differences; Fichna P et al.; The critical evaluation of isolation methods for obtaining the adrenocortical cell suspension due to trypsin or collagenase digestion was done . Some collagenase advantages were indicated by morphological observations on the staining smears as well as by ACTH stimulation test . The cytochemical reactions for enzyme activities had the limited applications for those purposes . It also appeared that commonly applied dye exclusion tests were inadequate for characterization of cell suspension . The possible role of the adrenocortical cell debris in the basal corticosterone production was pointed out . The maintenance of the sex dimorphism and the functional differences in the adrenocortical cells isolated from male and female rats have been observed. Histochemistry, 1987, 87(1), 27 - 38 Skeletal muscle cell populations . Separation and partial characterization of fibroblast-like cells from embryonic tissue using density centrifugation; Yablonka-Reuveni Z et al.; Cell suspensions from the breast muscles of 10-day old chicken embryos were separated into non-myogenic, fibroblast-like cell fractions and a mononucleated, myogenic cell fraction by Percoll density centrifugation . Isolated populations were characterized by their morphology in both mass cultures and individual macroscopic clones and by the immunocytochemical detection of skeletal muscle- and smooth muscle-specific proteins in individual cells . Cell populations were also characterized by their protein patterns using sodium dodecyl sulfate/polyacrylamide gel electrophoresis . The less dense, non-myogenic cells comprised 16% of the cells . In culture they were predominantly flattened, stellate cells and gave rise to clones lacking myotubes . These fibroblast-like cells were negative for skeletal muscle myosin or muscle type creatine phosphokinase . Less than 0.1% of these cells demonstrated strong fluorescence when stained with anti-desmin or anti-smooth muscle specific actin . This observation suggested that the vast majority of these cells were not related to vascular smooth muscle cells . Also, over 99% of the non-myogenic cells did not display characteristic properties of endothelial cells . The denser myogenic cell fraction comprised over 80% of the cells and in clonal cultures gave rise to about 70% myogenic clones . An additional 30% of clones from this fraction were non-myogenic indicating heterogeneity in this population . We conclude that Percoll centrifugation can be employed for the isolation of myogenic and non-myogenic cell populations directly from the embryonic muscle . Moreover, this procedure allows the direct analysis of cell-specific proteins (e.g., by gel electrophoresis) without the need for cell culturing . The results thus obtained closely reflect the status of the cells in the intact muscle. Cytometry, 1987 Jan, 8(1), 14 - 9 Comparison of tissue disaggregation techniques of transitional cell bladder carcinomas for flow cytometry and chromosomal analysis; Smeets AW et al.; DNA index (DI) measurements and chromosomal analysis of 42 transitional cell carcinomas were done after mechanical and enzymatical disaggregation of the tumor specimens . The results obtained with these different disaggregation techniques were compared in the 33 cases (79%) that showed recognizable chromosomes . The enzymatically obtained cell suspensions could not be used for chromosomal analysis after short-term culture of 24 hours . In four cases, the DI after enzymatical treatment could not be estimated . In most cases, the DI obtained from the tumor cells was similar for both aggregation techniques, with the exception of four cases of enzymatically treated cell suspensions in which the DI could not be estimated . The average DI of the aneuploid tumors was 13% higher than the corresponding chromosome count . In 19% of the aneuploid tumors the proportion of aneuploid cells could not be measured after enzymatical treatment . In the remaining suspensions the proportion of diploid cells was higher after enzymatical disaggregation than after mechanical treatment . It is concluded that for flow cytometric and direct chromosomal analysis of bladder tumors, the mechanical disaggregation technique is most suitable. Leuk Res, 1987, 11(11), 987 - 94 Preclinical studies of a panel of 12 monoclonal antibodies in view of bone marrow purging in acute lymphoblastic leukemia; Racadot E et al.; We analysed the optimal conditions for autologous bone marrow purging using complement binding monoclonal antibodies (mAbs) . Twelve mAbs belonging to four clusters (CD9, CD10, CD19, CD24), alone or combined were evaluated by using direct cytotoxicity and clonogenic assays . We observed the following data: (1) optimal cytotoxicity was reached with doses of 1-10 micrograms mAbs for 10(7) cells, (2) the concentration of the cell suspension had to be below 3 X 10(7)/ml, (3) combinations of mAbs were more effective than a single mAb treatment, (4) in the case of an IgM isotype, there seems to be a clear dissociation between the amount needed for optimal toxicity and that for antigenic saturation measured by cytofluorometry. Experientia Suppl, 1987, 52, 317 - 22 Phytochelatins, the heavy metal binding peptides of plants: characterization and sequence determination; Grill E; The isolation of a homologous series of five novel heavy metal complexing peptides from plant cell suspension cultures is reported . Their structures were established as (gamma-Glu-Cys)n Gly, n = 3-7 . These peptides appear upon induction with heavy metals and represent the principal metal binding activity in induced cells . The name phytochelatin (PC) is proposed for this new class of natural products in higher plants . The hitherto postulated metallothioneins do not exist in plants but their function of chelating heavy metals via metal-thiolate coordination is fulfilled by the simpler phytochelatins . They are not primary gene products because of their repetition of gamma-glutamyl linkages. Arch Dermatol Res, 1987, 279 Suppl, S97 - 103 Electrophoretic mobility of corneocytes measured by laser Doppler spectroscopy; Plewig G et al.; A new approach studying the characteristics of the stratum corneum is presented: the electrophoretic mobility of corneocytes by laser Doppler spectroscopy . The detergent scrub technique was used for harvesting corneocytes from three body regions (forehead, palm, and sole) of normal persons (n = 20) under casual conditions and after thorough defattening of the skin with 70% isopropyl alcohol or petrol . Similarly, cells from the forehead, shoulder, and palm were obtained from 22 acne patients treated with isotretinoin (13-cis retinoic acid) 0.5-0.7 mg/kg body weight (b.wt.)/day for 12-16 weeks, and in patients receiving arotinoid (Ro 15-0778) 192 mg (n = 5) or 500 mg (n = 5) per kg/b.wt./day for 6 weeks (forehead and shoulder) . In another experiment, cell suspensions with a pH ranging from 5.0-7.3 were evaluated . Measurements were performed by dynamic laser light scattering . This laser application allows exact electrophoretic mobility measurements in a short time (3 min) . When cells pass the laser beam, the scattered light is frequency-shifted due to the optical Doppler effect . These frequency shifts are analyzed by the heterodyne light beating technique . The analog signal of the photodetector is converted into a power spectrum by Fourier analysis . This power spectrum represents the spectrum of electrophoretic cell mobility distribution . Results showed different electrophoretic mobility values for corneocytes dependent on the topographic region: forehead 1.18 +/- 0.16, palm 1.10 +/- 0.14, and sole 0.83 +/- 0.10 (means +/- SEM) micron cm/Vs . Defattening with isopropyl alcohol decreased the mobility values to 0.90 +/- 0.09 (p less than or equal to 0.01), 0.95 +/- 0.10, and 0.77 +/- 0.10 micron cm/Vs respectively.(ABSTRACT TRUNCATED AT 250 WORDS) Thymus, 1987, 10(1-2), 5 - 12 Survey of experimental data on fetal liver transplantation; Royo C et al.; Fetal liver transplantation has been shown to induce hematological and immunological reconstitution in irradiated rodents, dogs, horses, and sheep . Engraftment and reconstitution without GvHD has been readily obtained using histocompatible donors . When mismatched fetal donors were used, a comparatively larger number of donor cells was required, in addition to pre-treatment of host with higher doses of irradiation or irradiation plus chemotherapy . Stem cell suspensions devoid of any T lymphocyte can be transplanted across major histocompatibility barrier without inducing overt GvHD . The transplanted animals become tolerant to both donor and host grafts. Comp Biochem Physiol A, 1987, 88(4), 613 - 8 Blood oxygen equilibria and theoretical models . II . A critical estimation; Vorger P; 1 . Theoretical models can fit the oxygen equilibria of trout and human red cell suspensions, and describe the apparent oxygenation process in the red cell . 2 . The assumption that full oxygenation is attained at atmospheric O2 pressures can result in biphasic Hill plots and high Hill coefficients for high O2 saturations . This phenomenon must not be confused with aggregation of hemoglobin . 3 . Problems specific to measurements of red cell suspensions, regarding the ionic cellular composition and its stability with time, are approached . Changes of buffer osmolarity, and--for trout--addition of adrenaline, within physiological proportions, have no impact on the results . 4 . This tends to validate the general significance of equilibrium data obtained on this material, regarding the effects of protons and organic phosphates, although complex ionic movements across the red cell membrane are known to occur in the animal under certain circumstances. Arch Immunol Ther Exp (Warsz), 1987, 35(1), 71 - 8 Technique of preparation of hemopoietic cells of human fetal liver for transplantation; Jedrzejczak WW et al.; The preparation of single cell suspension from human fetal liver is described . The technique is based on pressing the liver through wire mesh followed by repeated aspiration into a syringe through needles of decreasing internal diameter . Subsequently, the cell suspension is depleted of cell debris by a "triple medium sedimentation procedure" without net loss of hemopoietic cells . Following centrifugation and resuspension the preparation is ready for either transplantation or storage . About 2 and 11 X 10(9) cells were obtained per liver depending on the age of fetus in the range of 16 and 24 weeks of gestation in three representative preparations . The cell suspension contained comparable numbers of hemopoietic progenitors to the adult bone marrow suspension as assayed using plasma clot diffusion chamber technique. J Neurochem, 1987 Jan, 48(1), 29 - 39 Cellular origins of cyclic GMP responses to excitatory amino acid receptor agonists in rat cerebellum in vitro; Garthwaite J et al.; Incubated slices and freshly dissociated cells from 8-day-old rat cerebellum were used to try to identify the cells that participate in the large increases in cyclic GMP levels that follow activation of excitatory amino acid receptors in this tissue . In the slices, cyclic GMP responses to L-glutamate and related excitants were unaffected by tetrodotoxin and could be replicated by the guanylate cyclase activator nitroprusside . Nitroprusside and the receptor agonists appeared to activate the same pool of the enzyme . Prior destruction of neuroblasts, deep nuclei, or Golgi neurones did not cause loss of responses to L-glutamate . If granule cells were rendered necrotic, however, the cyclic GMP responses to all excitants tested were reduced by greater than or equal to 90% . Substantial losses of responses to veratridine and high K+ levels also occurred, but the nitroprusside-induced elevations were unaffected . In dissociated cell suspensions, the magnitude of responses to receptor agonists, but not those to nitroprusside, was markedly dependent on cell concentration . Responses to L-glutamate were the same in cell suspensions that were Purkinje cell depleted and Purkinje cell enriched . It is concluded that granule cells are primarily involved in the cyclic GMP responses to excitatory amino acids but that the cyclic GMP accumulations occur elsewhere, probably in glial cells. Membr Biochem, 1987-88, 7(3), 143 - 52 Inhibition of stimulated cell responses by phorbol esters and other activators of protein kinase C: sites of action; Sha'afi RI et al.; The addition of the protein kinase C activator phorbol esters to cell suspension a few minutes prior to stimulation inhibits the agonists-induced biochemical changes and cell responses . This inhibition is prevented by protein kinase C inhibitors . Activation of protein kinase C down regulates the stimulated responses by affecting one or more of the steps in the exitation-response coupling . This includes the receptors, the quanine-nucleotide-binding protein, the activity or distribution of phospholipase C, and other steps. Vet Med Nauki, 1987, 24(9), 31 - 5 {Experimental infection of sheep with the virus of bovine enzootic leukemia}; Sandev N; The intraperitoneal, the muscular, and the nasal infection with a virus, containing material (blood of a cow, affected by leukosis, with 48100 leucocytes in a cubic mm and 92% of lymphocytosis) and cell suspension FLK, producing leukosis virus, provokes leukosis infection . It was established that the peroral and the intraconjunctival application of virus material does not provoke leukosis till the 6th month (term of observation) . With saliva and semen, obtained from positive serologic reagents with persistent lymphocytosis also does not provoke infection . By the immunofluorescent test, in the infected sheep were observed antibodies against the summary glycoprotein antigen, which appear after 1-2 months with continuous seroconversion till 18th month and precipitation titer 1:2 till 1:64 . Among 30% of the sheep, after the 5th and 8th month, was established leucocytosis, up to 18800 leucocytes, and persistent lymphocytosis--91-93%. Adv Exp Med Biol, 1987, 219, 489 - 513 Demonstration of hCG binding sites and hCG stimulated steroidogenesis in different populations of interstitial cells; Bhalla VK et al.; The mechanism by which luteinizing hormone (LH) promotes the production of testosterone in Leydig cells by binding to its high affinity sites was reinvestigated . Collagenase dispersed interstitial cells when purified by the application of a variety of techniques such as unit gravity sedimentation, gradient centrifugation, and a combination of the two procedures, were separated into two LH/hCG responsive cell fractions . The two types of interstitial cells displayed distinct biochemical and morphological characteristics . One cell type (the light cell) bound 125I-labeled human chorionic gonadotropin (125I-labeled hCG) with high affinity (Ka approximately equal to 3.33 x 10(9) M-1) but testosterone was not produced by this cell type as a result of hCG target cell receptor interaction . On the other hand, hCG stimulated the production of testosterone in another cell type (the dark/heavier cell) . Steroidogenesis was maximally stimulated (700-800 percent over basal) by concentrations of hCG in the range of 3 x 10(-10) M, but high affinity binding sites for 125I-labeled hCG were not detectable . The residual binding that occurred did not obey saturation kinetics and was predominantly nonspecific . The stimulation of steroidogenesis by hCG in dark/heavier cells was dose and time dependent . Addition of dibutyryl or bromo cAMP (1 mM) to the cell suspension resulted in production of testosterone demonstrating the involvement of an hCG sensitive adenylate cyclase system in the transfer signaling process . These observations suggest the lack of a direct association between the occupancy of high affinity binding sites by hCG and testosterone production in rat Leydig cells . The stimulation of a biological response by a pathway independent of hCG occupancy of high affinity binding sites on Leydig cell is discussed and morphology of light and dark/heavier cells is presented . Autoradiographic evidence substantiates the conclusions. J Steroid Biochem, 1987, 27(4-6), 929 - 34 Control of aldosterone secretion in zona glomerulosa cell suspensions and in the perfused adrenal gland of the rat; Vinson GP et al.; The secretion of aldosterone and its responses to stimulation have been studied in rat adrenal zona glomerulosa tissue incubated as intact capsules or as collagenase-dispersed cell suspensions, and in intact perfused rat adrenal glands . Several differences are apparent in the functions of the various preparations . Aldosterone secretion rates are similar in incubated intact capsules and in the perfused gland . Relative to corticosterone, lower yields of aldosterone are obtained in dispersed glomerulosa cell in vitro . This may be related to the loss in the dispersed cells of a pool of tissue steroid (aldosterone or a precursor) which is revealed only in intact tissue incubations by trypsin stimulation of aldosterone secretion . Trypsin-released aldosterone is increased by prior dietary sodium restriction . In addition, differences occur in the responses of dispersed cells and perfused glands to stimulation . Perfused glands from animals on a normal diet are less sensitive to stimulation by ACTH or alpha-MSH, but more sensitive than dispersed cells to angiotensin II amide . In the perfused gland, sensitivity of response (lowest effective concentration) to all three stimulants is increased by prior dietary sodium restriction, in contrast to dispersed cells in which increased sensitivity has been reported only to alpha-MSH . The perfused gland is particularly sensitive to angiotensin II amide, and a bolus administration of 1 amol gives significant stimulation in glands from animals on low sodium intake . Electrical (field) stimulation or dopamine administration at 10(-6) mol/l (which is ineffective in dispersed cells) both depress aldosterone secretion by the perfused gland . The data suggest that the sequestered pool of steroid is utilized in the perfused gland for aldosterone secretion . They furthermore suggest that in the intact gland there are mechanisms, which involve neural components, for intraglandular regulation of aldosterone secretion, which are lost in dispersed cells in vitro . Such mechanisms may be involved in sensitivity increases in sodium depletion. J Steroid Biochem, 1987, 27(4-6), 1017 - 22 The corticostatic (anti-ACTH) and cytotoxic activity of peptides isolated from fetal, adult and tumor-bearing lung; Zhu QZ et al.; The possibility that corticosteroids influence lung maturation was suggested in 1968 by Buckingham et al . and supported by studies mounted in humans and in rabbits by Liggins and Howie and by a collaborative study from the N.I.H . To our knowledge no reverse process, i.e . the regulation of adrenal function by the lung, has been postulated . In attempting to isolate ACTH from fetal lung we found peptides which depressed corticosterone production . Eventually, we isolated a group of peptides from rabbit fetal lung which showed potent inhibition of corticosterone production in rat adrenal cell suspensions . Using high-resolution reverse-phase HPLC and high-sensitivity gas-phase sequencing techniques the primary structure of one of these peptides has been determined and it was called corticostatin . The adult rabbit lung contains a large quantity of a very similar peptide, which elutes close to corticostatin but whose amino acid composition differs by a single glycine and possibly a serine . This peptide is also potently corticostatic . We have isolated homologous human peptides but these show no apparent corticostatic activity in rat adrenal cell suspensions . They do, however, show interesting effects on in vitro growth of lung cell lines. Cancer Chemother Pharmacol, 1987, 20(2), 109 - 14 A comparison of adriamycin and mAMSA . II . Studies with V79 and human tumour multicellular spheroids; West CM et al.; Multicellular spheroids were used to compare the two chemotherapeutic agents adriamycin (ADM) and 4'{(9-acridinyl)-amino} methanesulphon-m-anisidide (mAMSA) . Chinese hamster cells, V79 379A, a human small cell lung carcinoma, designated ME/MAR, and a human melanoma xenograft, HX117, were grown as spheroids (200 or 400 micron in diameter) and treated with either drug for 1 h, at 37 degrees C, in air . Cytotoxicity was assayed using both cell survival and growth delay . Both drugs were highly toxic towards V79 but showed less activity toward the human tumour single cell suspensions; ADM was more effective towards HX117 and ME/MAR than mAMSA . When grown as spheroids, the cells developed marked resistance to both drugs . In all cases, cytotoxicity was drug dose and spheroid size dependent . The response of HX117 spheroids to both drugs was similar . In contrast, ADM was more effective toward 200 micron diameter ME/MAR spheroids, and mAMSA showed greater activity than ADM against V79 spheroids . Both endpoints gave qualitatively equivalent results, and a comparison of the two showed relatively long growth delays for a given level of cell kill, for both drugs and with all three cell lines . The greater cytotoxicity of ADM toward ME/MAR spheroids is consistent with the clinical finding that ADM has a use in the treatment of small cell carcinoma of the lung, while mAMSA has not demonstrated any activity in the treatment of lung cancer. Hematol Pathol, 1987, 1(4), 209 - 15 Leu-M1 antigen as a marker of acute nonlymphoid leukemia; Chan J et al.; The monoclonal antibody Leu-M1 was originally described as a marker of myeloid/monocytic cells and their precursors . Its usefulness as a myeloid marker in bone marrow specimens was further supported by a limited number of studies using flow cytometry and cell suspensions . Recent reports on its utility in paraffin sections for the diagnosis of acute leukemia have not shown it to be an effective marker of myeloid differentiation . To further evaluate the diagnostic usefulness of Leu-M1 in acute leukemia, we investigated the immunoreactivity of Leu-M1 antigen in a series of 100 plastic-embedded bone marrow biopsies from patients with acute leukemia and compared these results to those obtained in a series of 30 paraffin-embedded specimens . We also investigated the usefulness of neuraminidase pretreatment of sections to enhance the sensitivity of Leu-M1 . In plastic sections, we were able to demonstrate positive Leu-M1 staining in 48 of 50 (96%) cases of acute nonlymphoid leukemia (ANL) (FAB classification M1-M5) and 0 of 50 cases of acute lymphocytic leukemia (ALL) . Comparatively, Leu-M1 stained positively in 10 of 20 (50%) cases of ANL and 0 of 10 ALL embedded in paraffin . In both plastic and paraffin sections, pretreatment with neuraminidase enhanced the sensitivity of Leu-M1 reactivity, but markedly decreased its specificity . Our study suggests Leu-M1 is both a reliable marker of myelomonocytic differentiation and a useful diagnostic tool in the differentiation of ANL from ALL in plastic sections, and confirms its limited usefulness in paraffin-embedded material. Curr Genet, 1987, 12(8), 617 - 23 Nucleotide sequence and molecular characterization of a maize mitochondrial plasmid-like DNA; Smith AG et al.; The mitochondrial genome of Black Mexican Sweet maize consists of the principal genome, a 2.3 kb minilinear DNA, a 1,913 bp (1.9 kb) and a 1,445 bp (1.4 kb) minicircular DNA . The three extrachromosomal DNAs exhibit characteristics of autonomous replication in cell suspension culture . The complete sequence of the 1.4 kb minicircle was determined . It has 61 bp of near perfect sequence homology to the 1.9 kb minicircle . Both minicircular DNAs are transcriptionally active; the longest open reading frame of the 1.4 kb minicircle was 231 bp . A putative origin of replication was identified as a high A + T sequence . These minicircles were present in some but not all of 20 maize lines surveyed . None of the lines examined carried the 1.4 kb minicircle without the 1.9 kb minicircle . Nuclear DNA of one line of the seven examined carried homology to both DNAs. Biomed Pharmacother, 1987, 41(9-10), 477 - 80 I-J+ suppressor cells are probably involved in regulation of spleen colony formation by hematopoietic stem cells (CFUs); Sanin AV; Treatment of murine bone marrow and spleen cell suspensions with anti-I-Jk alloantiserum and complement resulted in an increased number of CFUs (revealed by Till and McCulloch exocolonization technique) compared with controls . The stimulatory effect was observed with 7-8 day CFUs and was negligible or absent with 11-12 day CFUs . Normal thymus cells treated with anti-I-Jk antiserum plus complement would augment the colony-forming potential of normal marrow cells. Comp Immunol Microbiol Infect Dis, 1987, 10(3-4), 187 - 204 {Lymphocyte markers in domestic animals . I . Mitogenic lectins and non-mitogenic lectins}; Djilali S et al.; The authors reviewed the lymphocyte markers in domestic animals . The first part is devoted to lectins . The general and methodologic aspects of lectins as mitogens are studied . The main mitogen lectins are emphasized in regard to their field of utilisation and the technical aspect of their use . The non-mitogen lectins are presented for cell suspension or tissue section staining. Int Arch Allergy Appl Immunol, 1987, 84(2), 212 - 6 Kidney mast cells, IgE and release of inflammatory mediators capable of altering renal haemodynamics; Assem ES et al.; Collagenase-dispersed human, guinea pig and rat kidney cells, actively or passively sensitized with IgE or IgG subclasses, released histamine, leukotriene C4 and thromboxane B2 on challenge with the specific antigen or antisera to mast-cell-sensitizing immunoglobulins . A similar reaction occurred in the isolated perfused kidney of sensitized guinea pigs, which additionally showed vasoconstriction . Mast cells were identified in cell suspensions and tissue sections . These findings provide support for the possible role of IgE and mast cells in renal pathophysiology. Basic Appl Histochem, 1987, 31(2), 183 - 90 DNA-protein flow cytometric analysis in non-Hodgkin lymphoma; Del Bino G et al.; A flow cytometric study of DNA and protein contents was performed on cell suspensions obtained from 73 adult patients with non-Hodgkin lymphoma . Bivariate analysis identified a second subpopulation, not revealed by DNA determination, in 25% of the tumors . Protein heterogeneity was more frequently observed in diffuse than in nodular histology according the Rappaport classification and in high-grade than in low-grade malignancy tumors by the Kiel classification and the Working Formulation, but it was not related to ploidy or cell proliferative rate . The presence of an additional subpopulation, detected by protein analysis, defined as monoclonal by DNA analysis, could adversely affect clinical outcome in terms of response to treatment and overall survival. J Membr Biol, 1987, 97(3), 223 - 40 Patch-clamp study of isolated taste receptor cells of the frog; Avenet P et al.; Taste discs were dissected from the tongue of R . ridibunda and their cells dissociated by a collagenase/low Ca/mechanical agitation protocol . The resulting cell suspension contained globular epithelial cells and, in smaller number, taste receptor cells . These were identified by staining properties and by their preserved apical process, the tip of which often remained attached to an epithelial (associated) cell . When the patch pipette contained 110 mM KCl and the cells were superfused with NaCl Ringer's during whole-cell recording, the mean zero-current potential of 22 taste receptor cells was -65.2 mV and the slope resistance 150 to 750 M omega . Pulse-depolarization from a holding voltage of -80 mV activated a transient TTX-blockable inward Na current . Activation became noticeable at -25 mV and was half-maximal at -8 mV . Steady-state inactivation was half-maximal at -67 mV and complete at -50 mV . Peak Na current averaged -0.5 nA/cell . The Ca-ionophore A23187 shifted the activation and inactivation curve to more negative voltages . Similar shifts occurred when the pipette Ca was raised . External Ni (5 mM) shifted the activation curve towards positive voltages by 10 mV . Pulse depolarization also activated outward K currents . Activation was slower than that of Na current and inactivation slower still . External TEA (7.5 mM) and 4-amino-pyridine (1 mM) did not block, but 5 mM Ba blocked the K currents . K-tail currents were seen on termination of depolarizing voltage pulses . A23187 shifted the IK(V)-curve to more negative voltages . Action potentials were recorded when passing pulses of depolarizing outward current . Of the frog gustatory stimulants, 10 mM Ca caused a reversible 5- to 10-mV depolarization in the current-clamp mode . Quinine (0.1 mM, bitter) produced a reversible depolarization accompanied by a full block of Na current and, with slower time-course, a partial block of K currents . Cyclic AMP (5 mM in the external solution or 0.5 microM in the pipette) caused reversible depolarization (to -40 to -20 mV) due to partial blockage of K currents, but only if ATP was added to the pipette solution . Similar responses were elicited by stimulating the adenylate cyclase with forskolin . Blockage of cAMP-phosphodiesterase enhanced the response to cAMP . These results suggest that cAMP may be one of the cytosolic messengers in taste receptor cells . Replacement of ATP by AMP-PNP in the pipette abolished the depolarizing response to cAMP.(ABSTRACT TRUNCATED AT 400 WORDS) Folia Haematol Int Mag Klin Morphol Blutforsch, 1987, 114(2), 273 - 81 {Hyperacidification-induced, clear appearance of venules caused by the swelling of erythrocytes in microcirculation}; von Ardenne M et al.; In the mesenterium of rats it was found that an overacidity of the blood flowing in the area of microcirculation and caused by irrigation with acid media may be objectively represented in the clearing of venols measurable by means of video technique . The cause of this clearing process is the swelling of erythrocytes setting in at lower pH-values . According to in-vitro findings this swelling of erythrocytes will lead to an increase of the apparent viscosity of the blood fluid or blood cell suspension respectively in conformity with the increase of hematocrit connected with it. Folia Haematol Int Mag Klin Morphol Blutforsch, 1987, 114(2), 264 - 72 {Use of human albumin as an additional cryoprotective agent in freeze preservation of hematopoietic stem cells}; Reiners B et al.; 14 bone-marrow samples of healthy donors were cryopreserved with 5% of DMSO in combination with different volume percentages of 20% human albumin, with the protective impact of the respective freezing mixture on the proliferating capacity of early hemotopoietic precursor cells being determined by means of CFU-c-technique . A combination of 5% of DMSO with 20% of human albumin with an addition of 20% of autologous serum proved to be the most favourable freezing protection . CFU-c-recovery which was achieved in this way amounted to 78 +/- 7% . A protective impact of human albumin for CFU-c during incubation with the freezing medium at room temperature was statistically not significant . Thus, the method presented is suitable for cryopreservation of bone marrow for autologous transplantation . The advantages over the traditional procedure results from the slight degree of toxicity by reducing the DMSO percentage from the common 10% to 5% as well as from the fact that the thawed cell suspension may be directly infused with the substance for freezing protection, thus avoiding an additional loss of proliferative material . The procedure may be used especially for autologous transplantation in childhood. Folia Haematol Int Mag Klin Morphol Blutforsch, 1987, 114(2), 203 - 11 Splenic monocyte-macrophage dependent suppression of autologous granulocyte-progenitors growth in Hodgkin's disease; Hansz J et al.; The studies described compare the effect of spleen cell suspensions from 11 patients with Hodgkin's disease (HD) and 5 healthy subjects on the clonal growth of autologous marrow granulopoietic progenitors in diffusion chamber culture (CFU-G/D) . Adherent monocyte/macrophage fraction of splenocytes from HD suppresses the proliferation of autologous CFU-G/D . This inhibition was mediated by an indomethacin-sensitive humoral factor(s) . Non-adherent lymphoid cells stimulated myeloid colony formation . Dose response curves demonstrated a markedly increased inhibitory-activity production already by low numbers of splenic monocytes/macrophages from HD whereas a comparable counts of monocytes/macrophages from the spleens of healthy subjects stimulated the CFU-G/D growth . These results may suggest a possible activation of splenic monocytes/macrophages with an enhanced prostaglandin-mediated suppressor activity release for local granulocytopoiesis in the spleens of patients with HD. Biomed Biochim Acta, 1987, 46(2-3), S280 - 4 Accumulation of inosine 5'-monophosphate in human erythrocytes incubated with inosine; Tomoda A et al.; The changes in inosine 5'-monophosphate (IMP) and adenine nucleotides were studied in human erythrocytes incubated with glucose alone, or with glucose plus inosine in the presence or absence of monoiodoacetate, a strong inhibitor of the glycolytic pathways of the cells . HPLC was useful in analyzing the changes in these nucleotides . When the cells were incubated with glucose and inosine for several hours at pH 7.0, 37 degrees C, they produced much IMP, while the cells incubated with glucose and monoiodoacetate accumulated AMP rapidly within 2 hours, accompanied with the decomposition of ADP and ATP; but IMP was not accumulated . When the cells were incubated with glucose and inosine in the presence of monoiodoacetate, AMP increased more rapidly to about 1400 mu mols/ml of cells within 1 hour and then gradually decreased . ATP was consumed completely and ADP decreased correspondingly . However, IMP was produced in this case, though its accumulation was not so high as in the case with glucose plus inosine without monoiodoacetate . The production of IMP depended on the pH of the cell suspension, i.e., more IMP was produced at higher pHs. Oncology, 1987, 44(2), 102 - 7 Ploidy and proliferative activity of human brain tumors . A flow cytofluorometric study; Danova M et al.; Ploidy and proliferative characteristics were estimated by flow cytometry of the nuclear DNA content of 92 human brain tumors . Samples were frozen at -20 degrees C immediately after surgery and single cell suspensions were obtained with a mechanical dissociation technique . Propidium iodide was employed for nuclear DNA staining . Human normal brain tissue was used as internal diploid reference standard . 86% of benign tumors had unimodal DNA distribution with a DNA index (DNA I = modal channel of the G0/1 peak of the studied population/modal channel of the G0/1 peak of the normal brain) usually within the diploid or near-diploid range . 14.0% had aneuploidy, with an additional cell peak having a median DNA I of 1.60 . Among malignant tumors, these figures were 61.2 and 38.8% (p less than 0.001) . The percentage of S phase cells was higher in malignant (median = 3.6) than in benign tumors (median = 2.0, p less than 0.01), without correlation to histological tumor subtype . Flow cytometry appears to be a useful method for evaluating differences in DNA distribution in tumors of the central nervous system. Histochemistry, 1987, 86(3), 269 - 73 Lectins as markers for eosinophilic leukocytes; Lee MC et al.; Lectin from Griffonia simplicifolia (GSA-I) and soybean (SBA) are reliable markers for human eosinophils . In this study we have shown that fluorochrome labeled GSA-I and SBA can be used for specific labeling of eosinophils in paraffin embedded tissue sections, in peripheral blood smears and in cell suspensions prepared for flow cytometry . These two lectins are useful diagnostic reagents which could be applied for further characterization of cytoplasmic components selectively found in human eosinophils. Int Arch Allergy Appl Immunol, 1987, 82(3-4), 507 - 12 Some studies on human pulmonary mast cells obtained by bronchoalveolar lavage and by enzymic dissociation of whole lung tissue; Pearce FL et al.; Human pulmonary mast cells were obtained by bronchoalveolar lavage (BAL) and by enzymic dissociation of whole lung . The cells released histamine on immunological stimulation or on exposure to a hyperosmolar environment . Cell suspensions similarly released newly generated products of arachidonic acid metabolism . Increased numbers of mast cells were recovered by BAL of asthmatic subjects and patients suffering from sarcoidosis and these cells were hyperresponsive to immunological challenge . Mast cells recovered by BAL and enzymic dissociation were differentially inhibited by antiasthmatic drugs . These data emphasize the potential role of BAL mast cells in pulmonary diseases of diverse origin. Biull Eksp Biol Med, 1987 Jan, 103(1), 95 - 7 {Comparative evaluation of the action of serum proteins on tissue regeneration}; Miroshnikov VM et al.; Processes of skin and long tubular bones' reparative regeneration under the action of serum albumin and alpha-fetoprotein on the body were studied with the use of EPR and histological methods . Relations of spin probe rotatory mobility in cell suspensions and morphological change of examined tissues are shown . It is established that the biological activity of alpha-fetoprotein is more pronounced than that of serum albumin in the effect on the young connective tissue . The stimulating effect of the embryonic protein is demonstrated only in the first week of reparative process while that of serum albumin appeared much more late. J Immunol Methods, 1986 Dec 4, 95(1), 1 - 7 An immunofluorescent method for identifying individual IFN-gamma-producing lymphocytes; Laskay T et al.; An immunofluorescent staining method was developed for detecting human IFN-gamma-producing cells in single cell suspensions . Mononuclear leukocytes, stimulated in vitro to produce IFN-gamma, were fixed and made permeable . The cytoplasmic presence of IFN-gamma was visualized by indirect immunofluorescence using IFN-gamma-specific mouse monoclonal antibodies . The staining was found to be specific for IFN-gamma and allowed the detection of newly synthesized rather than internalized IFN-gamma molecules . The cytoplasmic fluorescence appeared locally in a polar, juxtanuclear position, which overlapped the Golgi apparatus, probably reflecting the glycosylation site of the newly formed IFN-gamma molecules . Two-colour staining experiments showed that the method is useful not only for the detection and enumeration but also for the phenotypic characterization of IFN-gamma-producing cells. J Neurocytol, 1986 Dec, 15(6), 799 - 815 Distribution of the adhesion molecules N-CAM and L1 on peripheral neurons and glia in adult rats; Mirsky R et al.; There is considerable evidence that the cell surface glycoproteins N-CAM and L1 are important mediators of cell-cell adhesion in the nervous system, at least during development . Numerous studies have been devoted to the molecular properties of these proteins and their adhesion role in embryonic and early postnatal development . Much less is known about their importance in mature tissues . A rigorous and comprehensive description of the cell distribution of these molecules in the adult nervous system would clearly form a useful baseline for functional and biochemical studies . In the present work we have addressed this issue and studied the distribution of N-CAM and L1 throughout adult, as opposed to developing, rat peripheral nervous tissue . Particular attention was paid to the ganglia of the enteric nervous system, since adhesion mechanisms within these ganglia are likely to be placed under unusual demands . We report, for the first time, the presence of N-CAM and L1 on mature sensory, sympathetic and enteric neurons in adult rats . Thus, immunostaining of cell suspensions or short-term cultures showed N-CAM and L1 surface labelling on sympathetic and both large and small dorsal root sensory neurons . Both antigens were also present on the surface of enteric neurons in cultures prepared from 10-day-old rats and neonatal guinea pigs . Immunostaining of sections of enteric ganglia from adults indicated that both molecules were also expressed by mature enteric neurons . In sections of mature sciatic nerve neither N-CAM nor L1 immunoreactivity were detected at the site where the plasma membrane of myelinated axons meets the ad-axonal plasma membrane of the myelin-forming Schwann cell . Thus, both N-CAM and L1 were detected on all major classes of peripheral neurons, while their levels in the plasma membrane of myelinated axons may be significantly down-regulated . Similarly, both N-CAM and L1 were present on all major classes of non-myelin-forming peripheral glia in adult rats . This includes the enteric glial cells of the myenteric ganglia, non-myelin-forming Schwann cells in the sciatic nerve, sympathetic trunk and fine autonomic nerves in the gut wall, and the satellite glial cells of sympathetic and dorsal root sensory ganglia . In contrast, myelin-forming Schwann cells did not express detectable levels of N-CAM and only very low levels of L1, which was mainly located near the nodes of Ranvier.(ABSTRACT TRUNCATED AT 400 WORDS) J Am Vet Med Assoc, 1986 Dec 1, 189(11), 1478 - 80 Erythrocyte agglutination associated with heparin treatment in three horses; Mahaffey EA et al.; In vitro erythrocyte agglutination developed in 3 hospitalized horses receiving heparin treatment . The agglutination caused artifactual decreases in erythrocyte counts and increases in mean corpuscular volume (MCV) values . Treatment of cell suspensions with trypsin eliminated the agglutination and the changes in erythrocyte count and MCV . Similar abnormalities in erythrocyte counts and MCV have been reported in healthy horses treated with heparin and have been cited as evidence of hemolysis and regenerative anemia. Gen Comp Endocrinol, 1986 Dec, 64(3), 355 - 61 Urotensin II lowers cytoplasmic free calcium concentration in goby enterocytes: measurements using quin2; Loretz CA et al.; A technique is described for the measurement of cytoplasmic free calcium concentration ({Ca2+}c) in a suspension of goby intestinal epithelial cells using the fluorescent probe quin2 . For 18 cell suspensions, {Ca2+}c was determined to be 142 +/- 14 nM . The caudal neurosecretory peptide urotensin II at 10(-7) to 10(-6) M concentration reduced {Ca2+}c by 20-50% . Together with previous findings linking the calcium-calmodulin system with the stimulation of Na+ and Cl- absorption in this tissue, it appears that alterations in cytoplasmic Ca2+ may mediate urotensin II-induced stimulation of ion transport. Med Radiol (Mosk), 1986 Dec, 31(12), 68 - 70 {Radiation sensitivity of aerated and hypoxic clonogenic cells of Ehrlich ascites carcinoma}; Konopliannikov AG et al.; The authors presented comparative results on the survival of ascitic Ehrlich's carcinoma (AEC) clonogenic cells during exposure to 60Co gamma-beam radiation under the hypoxic and oxygenated conditions in vitro . A well-known method of hypoxia induction in dense cell suspensions by way of forced cell precipitation into a dense precipitate by a short-term suspension centrifugation was modified . Radiosensitivity of such hypoxic cells was characterized by a DO value of 6.59 +/- 0.29 Gy, and that of well oxygenated cells by 2.06 +/- 0.05 Gy . The oxygen enhancement ratio, determined as a ratio of DO values for hypoxic and oxygenated cells, was 3.2. Blood, 1986 Dec, 68(6), 1196 - 200 Lysis of human fibroblast colony-forming cells and endothelial cells by monoclonal antibody (6-19) and complement; Abboud CN et al.; The murine IgG2a monoclonal antibody 6-19 binds to a wide variety of nonhematopoietic cells including human marrow-derived stromal cells but does not bind to marrow or peripheral blood cells . We studied the effects of this antibody and rabbit complement on marrow cells . Fibroblast colonies were eliminated from light density marrow cells by a single incubation with monoclonal antibody 6-19 and complement . The growth and composition of granulocytic and erythroid colonies were unaffected . Specific complement mediated cytotoxicity of the antibody was confirmed on passaged human fibroblasts derived from marrow (more than 99.6% of fibroblasts are killed by a single treatment) . Similar results were obtained with human umbilical cord endothelial cells . In addition, such treatment abolished the initiation of Dexter culture stroma . Incubation of bone marrow cell suspensions with this antibody and complement will allow the study of stroma-free marrow cells in long-term liquid cultures. J Cell Sci, 1986 Dec, 86, 25 - 33 Conditions for fibroblast adhesion without fibronectin; Curtis AS et al.; Conditions that permit the adhesion of BHK fibroblasts to a variety of surfaces after inhibition of protein synthesis and competition of any adsorbed fibronectin or vitronectin with the fibronectin cell-binding tetrapeptide, Arg-Gly-Asp-Ser (RGDS), are defined . Exposure of the cells to serum components at any stage in the preparation prevents cell attachment if cycloheximide or fibronectin tetrapeptide is present . If leupeptin is used cell adhesion and spreading occur even when all fibronectin synthesis is suppressed by cycloheximide inhibition, or fibronectin binding by tetrapeptide competition . The adhesions formed under these conditions appear by interference-reflection microscopy and by general properties to be identical to those formed by cells under normal culture conditions . The cell suspensions produced in the presence of leupeptin rather than other trypsin inhibitors show good adhesion at low temperatures, though the cells hardly spread at all . The results suggest that the role of fibronectin in cell adhesion should be reinterpreted in terms of its possible action as an activator rather than as a bonding molecule. Jpn J Exp Med, 1986 Dec, 56(6), 265 - 9 Lidocaine inhibits macrophage mediated cytotoxicity and enhances neutrophil mediated cytotoxicity; Cameron DJ; Peripheral blood monocyte derived macrophages and polymorphonuclear leukocytes (PMNs) obtained from normal donors kill tumor cells in vitro . However, if lidocaine is added to the macrophage tumor cell suspensions in microgram concentrations (0.1 micrograms-0.1 mg), there is marked inhibition of tumor cell killing . The inhibitory effect for the macrophages resulted from an effect of lidocaine on the effector cells . When the effector cells were preincubated with lidocaine, inhibition in macrophage mediated cytotoxicity was observed . Cytotoxicity was also inhibited when 0.01 mg of lidocaine was added to the macrophage monolayers either at the time of addition of the tumor cells or 15-60 min after addition of the tumor cells whereas no inhibition of cytotoxicity occurred when lidocaine was added more than 60 min after the initiation of the cytotoxic reaction . For the neutrophils it was observed that the lidocaine actually had an enhancing effect rather than inhibitory effect on their ability to kill tumor cells. Clin Exp Immunol, 1986 Dec, 66(3), 654 - 60 Studies in cobra venom factor treated rats of antibody coated erythrocyte clearance by the spleen: differential influence of red blood cell antigen number on the inhibitory effects of immune complexes on Fc dependent clearance; Yousaf N et al.; The splenic component of the mononuclear phagocyte system (MPS) was investigated in decomplemented rats by determining the clearance from the blood of erythrocytes coated with a monoclonal antibody (R3/13) . The infusion of immune complexes (IC), prepared at 10-fold antigen excess, at an appropriate time during the erythrocyte clearance produced a significant increase in the T1/2 of the antibody coated cells . Immune complexes formed with the F(ab')2 fragment of the rabbit antibody did not have any significant effect . A positive correlation was seen between the dose of immune complex infused and the degree of inhibition of erythrocyte clearance . The influence of red cell antigen number on the behaviour of erythrocytes sensitized with R3/13 was studied by comparing the clearance of DA and (DA X PVG) F1 erythrocytes . F1 erythrocytes, with only half the number of specific antigens on their surface that bind R3/13 antibody were cleared much more slowly (82 +/- 2.6 min, mean +/- s.e.) by the spleen than the DA erythrocytes (44 +/- 1.5 min P less than 0.001) . Both cell suspensions were equally susceptible to inhibition by soluble IC . These studies show that the number of specific antigens on the red cell surface influences the rate at which sensitized cells are removed by splenic macrophage Fc receptors but not their susceptibility to inhibition by IC . Our results draw attention to a major defect in the use of autologous erythrocytes coated with anti-rhesus (D) immunoglobulin to assess macrophage Fc receptor function in man. Clin Exp Immunol, 1986 Dec, 66(3), 599 - 605 Relationship of macrophages to defective delayed-type hypersensitivity in B6/lpr mice; Okuyama H et al.; These experiments were undertaken to clarify whether or not suppressor macrophages contribute to deficiency of delayed-type hypersensitivity induced by BCG CW immunization in B6/lpr mice . Addition of indomethacin in vitro did not increase MIF production by immunized lpr lymph node cells . Adherent cells in lymph node cell suspensions from lpr mice did not interfere with MIF production by nonadherent lymph node cells from immunized B6 mice . On the other hand, MIF production by nonadherent lymph node cells from immunized B6/lpr mice was not restored even if they were cultured with adherent lymph node cells from immunized B6 mice . In addition, macrophages from nonimmunized B6/lpr mice showed normal reactivity to MIF when lymphocytes coexisting with the macrophages in large proportions were depleted from PEC . These results suggest that macrophages make no contribution to the deficiency of DTH expression of B6/lpr mice. Am J Vet Res, 1986 Dec, 47(12), 2577 - 83 Dome epithelial M cells dissociated from rabbit gut-associated lymphoid tissues; Roy MJ et al.; Dome and dome epithelial cells were selectively dissociated from gut-associated lymphoid tissues of rabbits . Sequential tissue washes in dithiothreitol, EDTA, and collagenase removed the dome epithelium, without disrupting the follicles or villi, and provided a cell suspension containing 74 +/- 6% lymphocytes, 9 +/- 4% columnar epithelial cells, 10 +/- 7% tangible-body macrophages, and 4 +/- 2% M cells (follicle-associated epithelial cells) . The last mentioned cells were characterized by transmission electron microscopy as large (20 to 55 microns diameter) cuboidal, round, or oval cells with eccentric nuclei and thin membranous processes surrounding empty vacuoles . The M cells were occasionally joined together by tight junctions . Histochemical and immunocytochemical analyses of M cells with the light microscope showed that they were devoid of immunoglobulins and negative for T-cell antigen and secretory component and had no detectable alkaline phosphatase or endogenous peroxidase activity . The M cells had few vacuoles with faint acid phosphatase activity; nonspecific neutral esterase was abundant . Possible uses for dome and dome epithelial cells are discussed. Endocrinology, 1986 Dec, 119(6), 2670 - 82 Flow cytometric analysis and sorting of live male rat anterior pituitary cell types by forward angle and perpendicular light scatter; Hatfield JM et al.; Light scatter patterns produced by living cells in the flow cytometer are known to provide useful information with regard to their size and internal structure . The purpose of this study was to determine if light scatter signals produced by live male rat anterior pituitary cells could be used as markers to aid in the identification and separation of different hormone-containing cell types . The typical light scatter pattern (4 X 10(5) cells/sample X 15 min) had three ridges in the forward angle light scatter (FALS) perpendicular light scatter (PLS) bivariate cell distribution . FALS signals could be correlated with the size of different cell types and PLS signals with their content of cytoplasmic secretory granules . Agranular cells dominated the low PLS ridge while moderately granulated PRL cells and heavily granulated GH cells dominated the medium and high PLS ridges, respectively . These light scatter patterns were reproducible both within and between different cell suspensions . Inclusion of dopamine in the pituitary gland dissociation medium, a treatment known to increase intracellular PRL contents of mammotrophs, increased the intensity of the PLS signals of a large population of cells, presumably PRL cells . Pituitary cells prepared from different aged male rats also showed changes in light scatter . Cell sorting on the basis of FALS-PLS signals established the relationship between cell type and light scatter pattern. Cancer Res, 1986 Dec, 46(12 Pt 1), 6471 - 4 Cancer procoagulant in human tumor cells: evidence from melanoma patients; Donati MB et al.; It has repeatedly been proposed that fibrin plays a role in tumor growth and metastasis . Among tumor cell products or activities which may promote clot formation, cancer procoagulant (CP), a direct activator of coagulation factor X, has been suggested to be selectively associated with the malignant phenotype . We report here the enzymatic and immunological identification of this cysteine proteinase procoagulant in extracts and cells from human melanoma . CP activity was independent of both the intrinsic and extrinsic pathways of blood coagulation, using factor IX and factor VII deficient plasmas, and was inhibited by the cysteine proteinase inhibitors iodoacetamide and HgCl2 . CP activity was detectable in extracts and cell suspensions from all 32 patients studied and was higher in extracts from metastases (14.8 +/- 3.9 units/mg protein) than from the primary tumors (3.7 +/- 1.0 units/mg protein) . CP activity was not affected by an anti-apoprotein III antibody or by concanavalin A, a known inhibitor of thromboplastin . In contrast, no CP activity or antigen was detected in extracts from six benign melanocytic lesions . The procoagulant activity was dependent on factor VII and was inhibited by anti-apoprotein III antibody and by concanavalin A, properties that suggest that the procoagulant was tissue thromboplastin . These data indicate that CP can be expressed by human tumor cells and that, among melanotic lesions, its presence is associated with the malignant phenotype and its activity is particularly high in metastatic cells. Blood, 1986 Dec, 68(6), 1316 - 21 Interleukin-1 (22-K factor) induces release of granulocyte-macrophage colony-stimulating activity from human mononuclear phagocytes; Fibbe WE et al.; An electrophoretically pure preparation of natural human interleukin-1 (IL-1) was shown to stimulate in vitro colony formation in human bone marrow cultures . Day 4 myeloid cluster-forming cells (CFC), as well as early (day 7) and late (day 10) granulocyte-macrophage colony-forming units (CFU-GM) were stimulated in a dose-dependent fashion . At optimal concentrations of IL-1, the number of day 4 CFC reached 72%, the number of day 7 CFU-GM reached 32%, and the number of day 10 CFU-GM reached 80% of the respective numbers of colonies obtained by addition of crude leukocyte-conditioned medium (LCM) . The IL-1-induced stimulatory effect on CFU-GM growth could be completely neutralized by a rabbit anti-IL-1 antiserum . Colony growth was abrogated by depleting the marrow cell suspensions of phagocytic cells prior to IL-1 addition . Conversely, the effect could be reintroduced by addition of marrow-derived adherent cells to bone marrow cell suspensions that had been depleted of both phagocytic and E rosetting T cells . Furthermore, media conditioned by bone marrow-derived adherent cells or by peripheral blood mononuclear phagocytes in the presence but not in the absence of IL-1, stimulated in vitro colony growth of phagocyte-depleted bone marrow cell suspensions . These results indicate that IL-1 induces release of granulocyte-macrophage colony-stimulating activity (GM-CSA) from human mononuclear phagocytes. J Biol Buccale, 1986 Dec, 14(4), 273 - 80 A method for producing monoclonal antibodies to human oral epithelial cells by the hybridoma technique; Gazi MI; Female BALB/c mice 6-8 weeks old were immunized intravenously (IV) with two injections of 30,000 human oral epithelial cells two weeks apart . The sera of these mice, and none of the controls, were positive by indirect immunofluorescence to cell suspensions of human epithelial cells of gingivae, tonsils and skin, but were negative to sections of human spleen, kidney and liver . The fluorescence was apparent only in the prickle cell layer, not at the basement membrane region nor in the connective tissue . The spleen cells from one of the selected immunized mice were fused to mouse myeloma cells, using polyethylene glycol (1500 MW), three days after a third IV booster injection of 30,000 epithelial cells . Three of the monoclonal cell lines produced have shown different staining patterns with various sizes of epithelial cells in suspension, results that differ from the staining pattern obtained with epithelial cells using polyclonal sera . These monoclonal antibodies were directed against antigens that differ from blood group A, B and HLA antigens on the surface of epithelial cells. Cancer Res, 1986 Dec, 46(12 Pt 1), 6400 - 5 Establishment of a novel acute monoblastic leukemia cell line (YK-M2) having a near-triploid karyotype; Ohno H et al.; The YK-M2 cell line was established from the peripheral blood of a patient with acute monoblastic leukemia in whom an anterior mediastinal tumor preceded the peripheral blood manifestation . The established cells grew in a single cell suspension with a doubling time of 60 h and consisted of primitive monoblastic cells . The cells were 52% positive for peroxidase staining and manifested strongly positive activity of alpha-naphthyl acetate esterase, which was completely inhibited by sodium fluoride . The cells showed strong expression of Fc gamma receptors and phagocytosed sensitized ox erythrocytes . When the cells were incubated with 1 alpha,25-dihydroxy-vitamin D3, they were induced to differentiate into mature monocyte-macrophage-like cells, which reduced the nitroblue tetrazolium dye and released a small amount of the superoxide anion . Cytogenetic studies revealed that the cells had a near-triploid karyotype with a modal chromosome number of 68, and the short arm of one No . 17 chromosome was deleted {del(17)(p11)} . The YK-M2 cell line is particularly unique in that the cells retained the polyploid karyotype that may be an initial cytogenetic change in the malignant transformation of the parent leukemia cells. Acta Neurol Scand, 1986 Dec, 74(6), 417 - 24 B cell activation in multiple sclerosis; Sandberg M et al.; A microtechnique was established for the study of the limited numbers of cells available in CSF . The method allowed for the determination of the number of immunoglobulin-secreting cells (IgSC) as well as the quantitation of immunoglobulin or specific antibody secreted into the culture medium . Dose-response curves and kinetic profiles for the IgSC responses induced by pokeweed mitogen (PWM), a polyclonal B cell activator, were similar for CSF cells (CSFC) and peripheral blood mononuclear cells (PBMC) . When equal numbers of unstimulated CSFC and PBMC from patients with multiple sclerosis (MS) were cultured, both the number of IgSC and the amount of secreted IgG were significantly greater in CSFC cultures . The addition of PWM resulted in the differentiation of B cells among both CSFC and PBMC, as shown by an increase of both the number of IgSC and the amount of secreted IgG . Results with cultures of unstimulated cell suspensions from MS patients suggested that CSF cells from these patients may be activated in vivo . The addition of mitomycin-C treated autologous peripheral blood mononuclear cells (PBMCM) to cultures of small numbers of CSFC or PBMC resulted in an augmentation of the number of IgSC in both, whether or not they were stimulated with PWM, and also in an increased secretion of IgG into the culture supernatants . This culture system should prove useful in functional studies when limited numbers of cells are available. J Endocrinol, 1986 Dec, 111(3), 415 - 23 Prostaglandin F2 alpha-induced functional luteolysis: interactions of LH, prostaglandin F2 alpha and forskolin in cyclic AMP and progesterone synthesis in isolated rat luteal cells; Kenny N et al.; The acute antigonadotrophic action of prostaglandin F2 alpha (PGF2 alpha) was examined in dispersed luteal cell preparations from immature superluteinized rat ovaries . Cell suspensions prepared by collagenase digestion and purification over a Percoll density gradient were incubated for 1 h in Eagle's minimum essential medium in the presence and/or absence of LH, PGF2 alpha, N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate (dbcAMP) and forskolin . Medium was assayed for total progesterone and adenosine 3',5'-cyclic monophosphate (cAMP) . Luteal cell preparations showed typical steroidogenic (progesterone) responses to LH, mimicked by both dbcAMP and forskolin . Whilst the threshold LH dose to increase cAMP synthesis was greater than that for progesterone (100 micrograms/l compared with 1 microgram/l), 24 mumol forskolin/l was the threshold dose for both cAMP and progesterone responses . Furthermore, combined doses of LH and forskolin synergistically raised cAMP yet produced less than additive increases in progesterone . Similarly, combinations of dbcAMP plus forskolin produced less than additive progesterone increases . These data suggest that forskolin may not act as a simple mimic of LH . Prostaglandin F2 alpha dose-dependently inhibited forskolin-induced cAMP and progesterone synthesis and also inhibited progesterone synthesis induced by dbcAMP . These data suggest that the antigonadotrophic effect of PGF2 alpha has more than one locus of action, i.e . it both inhibits an adenylate cyclase event associated with cAMP generation and blunts the cellular response to cAMP . The present uncertainty over the exact locus of forskolin's action within the adenylate cyclase complex limits further delineation of the inhibitory action of PGF2 alpha on LH-responsive adenylate cyclase. Eur J Biochem, 1986 Dec 1, 161(2), 391 - 8 Differential response of cultured parsley cells to elicitors from two non-pathogenic strains of fungi . Microsomal conversion of (+)marmesin into psoralen; Wendorff H et al.; Microsomal fractions isolated from parsley cell suspension cultures, which had been challenged with an elicitor from either Alternaria carthami or Phytophthora megasperma f . sp . glycinea, catalyzed the formation of psoralen from synthetic {3-14C}(+)marmesin . Whereas psoralen was the only product formed in incubations with Alternaria-induced microsomes, another unidentified product was isolated from incubations with Phytophthora-induced microsomes . The latter product is neither a precursor nor a product of psoralen . In contrast, microsomes isolated from non-induced parsley cells lacked both of these catalytic activities . The formation of psoralen depends on NADPH as a cofactor and molecular oxygen . Blue-light-reversible CO inhibition and inhibition by various synthetic chemicals known to bind to cytochromes P450 indicated that the reaction is catalyzed by an elicitor-inducible cytochrome P450-dependent psoralen synthase . Fractionation of microsomal preparations by centrifugation revealed that psoralen synthase is associated with the endoplasmic reticulum . Our results suggest that the endoplasmic reticulum of cultured parsley cells is the primary target in the previously reported differential induction by elicitors from these two non-pathogenic strains of fungi. J Endocrinol, 1986 Dec, 111(3), 449 - 54 Effects of a single injection of a new depot formulation of an LH-releasing hormone agonist on spermatogenesis in adult rats; van Kroonenburgh MJ et al.; Adult male Wistar rats were treated with a single injection (500 micrograms s.c.) of a new biodegradable depot formulation of the LH-releasing hormone (LHRH) analogue {D-Ser(But)6}AzGly10-LH-RH (Zoladex; ICI 118,630) to evaluate its potential for inhibiting spermatogenesis . The drug produced a marked (P less than or equal to 0.05) decrease in serum concentrations of FSH, LH and testosterone with a maximum effect 14 days after treatment . Since striking focal histological changes were seen in the testis after only 1 week, at a time when changes in serum gonadotrophins were minimal, there may be a direct effect of the LHRH analogue on spermatogenesis . Degenerative changes in germ cells as well as Sertoli cells could be observed . Flow-cytometric analysis of testicular cell suspensions showed a significant decline in the absolute numbers of haploid cells (spermatids), tetraploid cells (mainly pachytene spermatocytes) and of the numbers of cells in the S-phase of the cell cycle . This suggests that the drug also inhibits proliferation of spermatogonia and/or primary spermatocytes . Testis weight, serum hormone concentrations, and histological and cytological parameters returned to essentially normal values 52 days after the injection . It is concluded that this new method of administration may have practical and pharmacokinetic advantages for the purpose of reversible inhibition of spermatogenesis. Anal Quant Cytol Histol, 1986 Dec, 8(4), 271 - 80 Application of antibodies to intermediate filament proteins as tissue-specific probes in the flow cytometric analysis of complex tumors; Ramaekers FC et al.; The flow cytometric (FCM) analysis of carcinomas is often hampered by the presence of stromal and inflammatory cells in the cell suspensions obtained from such neoplasms . Therefore, an FCM method was developed to distinguish epithelial from nonepithelial cells by using polyclonal and monoclonal antibodies to (cyto)keratins, the epithelial type of intermediate filament proteins . Using a model system of cultured bladder carcinoma (T24) and leukemia (MOLT-4) cells, we tested our hypothesis and procedures by labeling cell mixtures with these antibodies . After incubation with an appropriate intermediate filament antibody and propidium iodide staining, the DNA content and distribution of T24 cells could be analyzed separately from MOLT-4 cells . When applied to cell suspensions of endometrial carcinomas, bladder carcinomas and Grawitz tumors, only the epithelial (primarily carcinoma) cells were stained for cytokeratin; these cells could thus be analyzed separately from stromal, inflammatory and other nonepithelial cells . In this way, a more accurate FCM analysis of the malignant fraction within a tumor can be achieved. J Invest Dermatol, 1986 Dec, 87(6), 792 - 4 Flow cytometric sorting of human epidermal pure basal cell suspensions using a specific antikeratin monoclonal antibody; Staquet MJ et al.; A method is described for flow cytometric sorting of epidermal basal cells, based on the cellular keratin content . KL1, a monoclonal antikeratin antibody specific for the 56.5 kD polypeptide present in suprabasal cells was used to distinguish suprabasal from basal cells . After the action of Triton X-100, epidermal cells were stained in suspensions with KL1 antibody, and KL1-positive cells were separated by flow cytometry from KL1-negative cells (basal cells) . This makes it possible to sort and analyze pure fractions of human epidermal basal cells since no less than 99% of the KL1 sorted cells were bearing bullous pemphigoid antigen . Therefore, the use of specific antibodies to cytoplasmic antigens can be of help in sorting pure fractions of cells of a particular tissue for further studies. Biochem Biophys Res Commun, 1986 Nov 14, 140(3), 1121 - 6 Activation of NADPH oxidase and phosphorylation of membrane proteins in human neutrophils: coordinate inhibition by a surface antigen-directed monoclonal antibody; Pontremoli S et al.; Exposure of human neutrophils to low concentrations of phorbol myristate acetate (PMA) results, after a brief lag, in the production of superoxide anion and the phosphorylation of membrane proteins . Evidence that these responses are linked has now been obtained using a monoclonal antibody directed against an undefined macrophage surface antigen . The addition of this antibody, which recognizes a 90 kDa neutrophil membrane protein, caused dose-dependent delays in the onset of both phosphorylation of neutrophil membrane proteins and in the appearance of superoxide anion, following addition of PMA to the cell suspensions . For each response the lag period increased with increasing concentrations of antibody, but the onset of phosphorylation always preceded by a few minutes the initial appearance of superoxide anion. Biochim Biophys Acta, 1986 Nov 5, 852(1), 25 - 9 Decrease of NADH in yeast cells by external ferricyanide reduction; Yamashoji S et al.; Ferricyanide reduction catalyzed by vitamin K-3 was accompanied by the decrease in intracellular (NAD(P)H concentration of yeast cells, and the rate of ferricyanide reduction depended on intracellular concentration of NADH rather than NADPH . The addition of glucose to the cell suspensions enhanced both ferricyanide reduction and intracellular NADH concentration . The catalytic action of vitamin K-3 on ferricyanide reduction was observed in the presence of NADH and plasma membrane preparations . As the toxic action of vitamin K-3 on cell growth of yeast was enhanced by addition of ferricyanide, ferricyanide reduction catalyzed by vitamin K-3 may inhibit cell growth by decreasing intracellular NADH concentration. J Reprod Immunol, 1986 Nov, 9(3), 187 - 94 Assessment of immune responses to H-Y antigen in naturally inseminated and sperm-injected mice using cell-mediated cytotoxicity assays; Hancock RJ et al.; An in vitro cell-mediated cytotoxicity assay has been used to assess immunity to the male-specific H-Y antigen in female mice after either insemination with male cells at syngeneic natural mating or after injection with syngeneic sperm cell suspensions . Lymphocytes showing specific cytotoxicity for the H-Y antigen could be recovered from both spleen and lymph nodes of female mice injected with sperm . However, insemination of male cells at natural mating did not apparently prime cytotoxic cells against the H-Y antigen in either the spleen or para-aortic lymph nodes draining the uterus of female mice mated once or repeatedly (3-10 X) in the absence of pregnancy . These results are discussed in relation to the factors regulating the immune responsiveness of the female to inseminated antigens. Brain Res Bull, 1986 Nov, 17(5), 697 - 704 GABA and dopamine act directly on melanotropes of Xenopus to inhibit MSH secretion; Verburg-Van Kemenade BM et al.; The release of melanophore stimulating hormone (MSH) from the pars intermedia of the amphibian Xenopus laevis is regulated by multiple factors of hypothalamic origin . The aim of this study was to determine if potential secretagogues function through a direct action on the melanotrope cell . For this purpose an in vitro superfusion system containing isolated melanotropes (cell suspension) was utilized . The viability of the cells in suspension was tested by examining their ability to synthesize, process and release pro-opiomelanocortin (POMC) related peptides . All biosynthetic functions appeared normal, with the exception that the isolated melanotropes are unable to N-terminally acetylate MSH . Release of immunoreactive-MSH from these cells was shown to be Ca2+-dependent, and high K+ stimulated release . Both the neurotransmitters dopamine and gamma-aminobutyric acid (GABA), which are thought to be physiologically important MSH-release inhibiting factors, were shown to inhibit MSH release from isolated melanotropes . Dopamine appeared to function through a dopamine D2 type receptor mechanism while for GABA, both a GABAa and GABAb receptor mechanism are involved. Biofizika, 1986 Nov-Dec, 31(6), 990 - 2 {Effect of anesthetics on porphyrin-sensitized photodamage of cells}; Bolodon VN et al.; The ability of a positively charged local anesthetic dibucain introduced into cell suspension prior to its irradiation to accelerate porphyrin-sensitized photodestruction of different cells is shown . Neutral anesthetics benzocain and ethanol had no such effect . There has been obtained evidence that the dibucain-induced increase in the amount of cell-bound sensitizer plays an essential role in the acceleration of photodestruction. Am J Vet Res, 1986 Nov, 47(11), 2454 - 60 Scanning electron microscopic studies of megakaryocytes and platelet formation in the dog and rat; Handagama P et al.; Megakaryocyte morphology and platelet formation in canine and murine bone marrows were studied by scanning electron microscopy . In situ-fixed bone marrow preparations and cell suspensions of bone marrow provided complementary information for the 2 species (dogs and rats) . Cylindrical processes (proplatelets) of variable length and thickness, originating from the megakaryocyte surface, were in the larger marrow sinusoids and the central vein . Regional constrictions along the length of proplatelets, particularly near their apical region, and the presence of fragments of such processes supported the concept of platelet formation through segmentation of proplatelets . Megakaryocytes presented varied morphology . Surface features resembling platelets were observed on megakaryocytes, indicating that platelets may have been released through surface budding . In conclusion, megakaryocytes formed long proplatelet processes that actively migrated to venous sinusoids to release platelets by fragmentation . Scanning electron microscopy analysis revealed a complex and variable megakaryocyte surface topography . The platelet-like structures on megakaryocyte surfaces may represent platelet release by a budding mechanism . The similarity between murine platelet release and canine platelet release demonstrates that data from rodent models may be applicable to nonrodents. In Vitro Cell Dev Biol, 1986 Nov, 22(11), 632 - 6 Spontaneous cell shedding by tumor cells in monolayer culture; Skehan P et al.; Sarcoma 180 monolayers spontaneously shed single cells and small multicellular aggregates into the surrounding medium to produce a dual population of floating and substratum-attached cells . Shedding was a motility-associated event that occurred when cells attempted to migrate over one another . It resulted from a combination of cell shape change and active motility, which increased sensitivity to fluid shear dislodgement by reducing a cell's surface area of adhesive contact and increasing strain tension at its adhesive contact points . Shedding occurred at all phases of the cell cycle . Extracellular matrix but not conditioned medium enhanced the floating subpopulation by slowing the kinetics of reattachment to plastic and cellular substrate . Although sarcoma 180 cells are anchorage independent in the sense that they grow readily in single cell suspension, they nevertheless exhibited anchorage modulation of their cell cycle . Short periods in suspension produced a mild G1 accumulation, whereas longer periods of anchorage deprivation led to a mild G2 accumulation which appeared to result from an interference with cytokinesis. J Cell Physiol, 1986 Nov, 129(2), 221 - 9 Effect of inositol hexaphosphate on the transient behavior of red cells following a DMSO-induced osmotic pulse; Franco RS et al.; An osmotic pulse can be used to incorporate inositol hexaphosphate (IHP) into red cells . The pulse is induced by equilibrating a red cell suspension with DMSO and then rapidly diluting with an isotonic IHP solution . Since IHP binds to hemoglobin and lowers the affinity for oxygen, this method may find application in the preparation of low-affinity cells for experimental and clinical use . The experiments reported here examined the dynamic changes of several red cell variables immediately following the osmotic pulse . The effect of IHP, which has been shown to dissociate red cell cytoskeletons, was evaluated by comparison with a matched phosphate-buffered saline (PBS) diluent . Red cell morphology, volume, and hemoglobin permeability were studied by fixing the cells at times ranging from 0.06 to 300 sec after dilution . Mechanical fragility was measured by subjecting the cells to a short period of shear stress at the same times after dilution . With both diluents, the cells underwent a rapid increase in volume followed by a return towards normal volume with a maximum at less than 250 msec . With IHP diluent, the period of hemoglobin permeability immediately followed the size peak and was completed by about 1 sec after dilution . PBS also induced a second leakage at longer times (10-120 sec), which resulted in a morphological dichotomy with ghosts and intact cells . The choice of diluent also affected sensitivity to shear stress . The IHP-treated cells had a mechanical fragility maximum at about 1 sec . The PBS-treated cells exhibited no enhanced mechanical fragility . An unexpected result was the inhibition of the second phase of lysis in PBS-treated cells by a properly timed shear stress. Mutat Res, 1986 Nov, 163(2), 175 - 80 In vitro SCE discrepancy between embryonic and extraembryonic mouse tissues; Elbling L et al.; In vitro sister-chromatid exchange (SCE) background levels and cytokinetics were compared in embryonic (whole embryo cell suspensions) and extraembryonic (yolk sac and amnion, placenta) cells of inbred and outbred strains at various gestational stages (days 12-17) . Results indicate a tissue origin (embryonal, extraembryonal) related variation in the formation of baseline SCE frequencies and cytokinetics . The significant higher SCE levels in extraembryonic tissues (maximum increase of 2 X the background values of the embryo cells) were independent of mouse strain and gestational stage . An average of 4-5 SCEs/cell in embryo cells is contrasted by 7-9 SCEs/cell in extraembryo cells . Mitotic index was generally lower and average generation time longer (by 2-3 h) in extraembryonic tissue cells . No significant differences in SCE frequencies and no changes in cytokinetics were detected at the BrdU concentrations used (1.2-4.8 micrograms/ml) . The reason for the inter-tissue differences in baseline SCE is still not clear. Cancer Res, 1986 Nov, 46(11), 5662 - 6 Hemostasis following inoculation and during spreading of colon carcinoma in the rat; Zoucas E et al.; Platelet function following inoculation of chemically induced carcinoma was evaluated in the rat . The original line of tumor (NGW1) was obtained using N-methyl-N-nitrosoguanidine . After trypsin homogenation a cell suspension of 0.3 X 10(6) viable tumor cells was injected subserosally in the cecum of each animal . Controls received injections of equal volumes of 0.9% NaCl solution or trypsin . The animals were subjected to laparotomy 2, 4, and 6 weeks after inoculation . Platelet function was assessed in vivo by measuring bleeding time and blood loss during mesenteric vessel transection or liver resection upon laparotomy . Hemoglobin, hematocrit, platelet count, activated partial thromboplastin time, platelet aggregation, thromboxane B2, platelet factor 4, and fibrinogen levels were evaluated after sacrifice by exsanguination . Significant decrease in bleeding time and blood loss was observed in animals with local primary tumors as well as in rats with lymph node metastases . Hemoglobin and hematocrit were decreased in the presence of metastases . Platelet count was not changed . Activated partial thromboplastin time was not affected by the presence of tumor . Platelet aggregation in vitro was accelerated in the presence of primary tumor or lymph node metastases, as well as following addition of tumor cells to platelet suspensions . No changes in thromboxane B2 or platelet factor 4 could be registered . Fibrinogen levels were decreased in the presence of liver metastases . Enhancement of primary hemostasis and platelet function in the presence of colon carcinoma in the rat was demonstrated both in vivo and in vitro . Direct or indirect interaction of the tumor cell with thrombocytes may play a role in determining the metastatic potential of the neoplasm. Cancer Res, 1986 Nov, 46(11), 5602 - 5 Fluorescence anisotropy of cell membranes of doxorubicin-sensitive and -resistant rodent tumoral cells; Montaudon D et al.; We have studied the plasma membrane fluidity of rat C6 glioblastoma cells and simian virus 40-transformed mouse liver cells in culture that had been rendered resistant to doxorubicin . This was done by the evaluation of fluorescence anisotropy of two probes; diphenylhexatriene was used on membrane microsomal fractions, and trimethylammonium-diphenylhexatriene was used on whole cell suspensions as a plasma membrane-specific probe since it does not enter the cells . A higher degree of membrane fluidity was exhibited with both techniques by doxorubicin-resistant glioblastoma cells as compared to the doxorubicin-sensitive strain, but in the transformed liver cells no such alteration was seen in the physical properties of their plasma membranes . A higher degree of acyl group unsaturation was noticed in the glioblastoma cells but not in the transformed liver cells upon acquisition of doxorubicin resistance . A similar simultaneous increase in acyl group unsaturation and membrane fluidity can be obtained easily by growing the sensitive cells with a medium supplemented with exogenous polyunsaturated fatty acids . This alteration does not modify the sensitivity of the cells to doxorubicin . We conclude from our work that the increase in membrane fluidity, which is frequently associated with drug resistance, is neither necessary nor sufficient for the expression of the resistance . The reason for a link between cell resistance to doxorubicin and plasma membrane fluidity remains to be found. Cancer, 1986 Nov 1, 58(9), 1969 - 78 Use of nude mouse xenografts as preclinical drug screens . Further studies on in vitro growth of xenograft tumor colony-forming cells; Taetle R et al.; Previous studies suggested tumor colony-forming cells (CFC) grown from xenografts might be useful as a preclinical, in vitro drug screen . To further evaluate this possibility, eight melanoma and six ovarian carcinoma xenografts were established from untreated patients and tested for in vitro CFC growth . For each tumor, linear relationships between cells plated and colony (30 cells or greater than 75 micron diameter) and cluster (10-30 cells or 50-75 micron) growth were observed . All eight melanomas grew sufficient colonies (greater than or equal to 30) for in vitro drug assessment, although four required hypoxic (pO2 = 40) incubation to reliably attain this level of growth . Only one in six of the ovary xenografts consistently grew enough colonies, and growth was not significantly improved by hypoxic incubation, or addition of luteinizing hormone, follicle-stimulating hormone, or steroid hormones . Cloning efficiencies (colonies + clusters/cells plated) for tumors demonstrating adequate growth ranged from 0.01% to 0.3% . For most tumors, no direct relationship was observed between characteristics of xenograft tumors (size) or their resulting cell suspensions (viabilities, cell yield) and CFC growth . Cell suspensions were incubated with a 3 log concentration of nine established chemotherapeutic agents . Resulting dose-effect curves were linear and showed no plateaus of drug effect . Analyzing 447 in vitro drug trials on six melanomas and one ovarian carcinoma, interexperiment variability was high . Cell lines were established from three xenografts using a low concentration of fetal bovine serum (1%), and also examined for in vitro drug sensitivity . Using both liquid culture isotope incorporation and a colony-forming assay, drug sensitivity profiles for the cell lines were nearly identical to those for parent xenograft CFC . However, assays performed using the cell lines were more reproducible than those using xenograft tissue . The authors conclude that tumor CFC can be reliably grown from melanoma xenografts, but in vitro drug assays using these xenografts are poorly reproducible . The xenografts are a resource for establishing cell lines, and drug assays performed using these lines are highly reproducible . Similarities in drug sensitivity profiles for parent xenograft CFC and derived cell lines suggest that, despite poor reproducibility, repetitive assays using melanoma CFC accurately reflect some properties of cells which sustain tumor cell growth. Transplantation, 1986 Nov, 42(5), 537 - 41 Allogeneic hepatocyte transplantation in the rat spleen under cyclosporine immunosuppression; Makowka L et al.; An innovative approach for stimulating the rapid growth of allogeneic hepatocytes implanted into splenic tissue with maintenance of the structural integrity is described . Single cell suspensions of hepatocytes from normal male ACI-strain rats (RTIa) were injected (2 X 10(6) cells) into the spleen of allogeneic male Fischer (RTI1) recipient rats . A 70% partial hepatectomy (PH) was performed at the same time as hepatocyte transplantation . Animals were treated for 4 days prior to, and 1 day after, transplantation with a feeding regimen containing 0.05% 2-acetylaminofluorene (AAF) to inhibit regeneration of the residual host liver . Animals received cyclosporine (CsA) 3 mg/kg/day s.c . posttransplantation . Histological examination of a standard longitudinal section of the recipient spleen two days posttransplant revealed an approximately 0.54-mm2 area replaced by hepatocytes . By 7 days this had increased to 0.97 +/- .15 mm2 . Without CsA administration, hepatocytes were undetected at 7 days . Both PH and AAF treatment were necessary for successful colonization and sustained proliferation . Withdrawal of CsA treatment at 10 days after transplantation resulted in rapid rejection of established hepatocytes . This study demonstrates that rapid colonization of the rat spleen with allogeneic hepatocytes can be achieved, and that the viability and structural integrity of these transplanted cells can be maintained for at least 14 days using cyclosporine immunosuppression. Mol Cell Endocrinol, 1986 Nov, 48(1), 11 - 20 Islet neoformation in tissue culture; Popiela H et al.; We have devised a tissue culture system that permits de novo formation of islets . Neonatal rat pancreata are enzymatically dissociated into single cells . The cell suspension is filtered through polyester cloth with 20 microns pores to exclude cell aggregates as well as preformed islets and a single cell suspension is then plated into tissue culture dishes at a density precluding reaggregation . Pancreatic cells proliferate forming numerous colonies of epithelioid cells . After a confluent monolayer, cells proliferate into a third dimension, the space occupied by the culture medium . Third dimensional proliferation occurs from basal monolayers of epithelioid colonies . At about 9 days in culture, numerous hillocks are visible that are spaced at about 1 mm from one another . Islets are observed to bud from the hillock surfaces . In 1 micron-thick sections, secretion granules are detected with the light microscope in some islet cells . With the electron microscope three basic cell types are seen . One peripherally located cell type is sparsely granulated and appears to be a precursor cell . The other peripherally located cell type shows a homogeneous population of secretion granules characteristic of A-cells . The third cell type is found in the interior of islets containing granules characteristic of B-cells . Islet cells, but not hillock cells, react immunocytochemically for insulin and glucagon . The cultures secrete 2 to 10-fold the amounts of glucagon present in fresh medium . It is concluded that differentiation of A- and B-cells occurs in neoformed islets. Clin Sci (Lond), 1986 Nov, 71(5), 581 - 7 Development and application of a superfusion technique for the study of renin secretion in rat renal cortical cells; Drury PL et al.; A dynamic column superfusion system has been developed for the study of renin secretion in rat renal cortical cells . Cells were isolated by collagenase digestion and mechanical dispersion, before suspension with polyacrylamide beads and superfusion with oxygenated physiological medium . Renin was detected in the superfusate by incubation of fractions with excess nephrectomized sheep substrate in the presence of angiotensinase inhibitors followed by radioimmunoassay of the angiotensin I generated . Optimized methodology included a purpose-built polytetrafluorethylene flow cell, a 1 h equilibration to achieve a steady state, 5 min eluate collections, a 15 min stimulatory and a 30 min recovery period, and duration of perfusion of up to 270 min . Significant increments above baseline renin release were seen with the stimuli of adrenaline, noradrenaline and isoprenaline . These could be demonstrated with concentrations of 10(-9) mol/l (adrenaline), 5 X 10(-10) mol/l (noradrenaline) and 10(-9) mol/l (isoprenaline) . This technique has significant advantages over previous methods for the study of renin secretion in vitro at the cellular level . It is reproducible and sensitive, and avoids many of the limitations of static cell suspension and kidney slice methods. Am J Reprod Immunol Microbiol, 1986 Nov, 12(3), 70 - 7 Immunoactive products of placenta . V: Soluble factors from murine placenta can block effector stages of maternal ant