Exp Neurol, 1990 Jan, 107(1), 11 - 22 Synaptic connections formed by grafts of different types of cholinergic neurons in the host hippocampus; Clarke DJ et al.; The present experiment was performed to determine whether different types of grafted central cholinergic neurons are able to form synaptic contacts with host hippocampal neurons . Grafts from the septal-diagonal band area, which contain the neurons that normally innervate the hippocampal formation, were compared to those from the nucleus basalis magnocellularis region (NBM), the striatum, the pontomesencephalic tegmentum of the brain stem, and the spinal cord . The regions were dissected from 14- to 16-day-old rat fetuses, and the same number of viable cells (35 x 10(4} from each of the different regions was stereotaxically injected as a cell suspension into the hippocampus of rats subjected to a complete fimbria-fornix lesion, transecting the intrinsic septohippocampal pathways . At 14 to 17 weeks after transplantation, the brains were processed for choline acetyltransferase (ChAT) immunocytochemistry at the light and electron microscopic levels and acetylcholinesterase (AChE) histochemistry at the light microscopic level . There was a great variation in the number of surviving ChAT-positive cells among the different graft types . The septal grafts contained the highest number of ChAT-positive cells, and the striatal grafts showed the lowest numbers . The NBM, brain stem, and spinal cord grafts were in between . The differences in the number of ChAT-positive neurons between the groups matched, in general, the differences found in the magnitude of graft-derived AChE-positive fiber growth into the host hippocampal formation . At the electron microscopical level, all types of grafts were capable of forming synaptic contacts with host elements, however, with vast differences in the number of synapses found . The septal grafts produced the highest number of contacts, whereas the striatal and spinal cord grafts produced very few contacts . The ultrastructure of the cholinergic fibers from grafts obtained from the forebrain areas, i.e., septum, NBM, and striatum all appeared normal, whereas brain stem and spinal cord grafts produced different types of anomalies . The results show that grafted cholinergic neurons, that normally do not innervate the hippocampus, can send axons and form synaptic contacts in the host hippocampus . The ability to reinnervate the denervated hippocampal target appears to be shared by the embryologically closely related forebrain cholinergic neuron types, i.e., the septal, NBM, and striatal neurons . The marked differences in overall fiber ingrowth and number of synapses observed between these different types of grafts could be explained largely on the basis of differences in survivability of each grafted neuron type . By contrast, the reinnervation obtained from the grafted brain stem and spinal cord neurons were both quantitatively and qualitatively abnormal.(ABSTRACT TRUNCATED AT 400 WORDS) J Clin Endocrinol Metab, 1990 Jan, 70(1), 28 - 34 Steroidogenesis in an estrogen-producing adrenal tumor in a young woman: comparison with steroid profiles associated with cortisol- and androgen-producing tumors; McKenna TJ et al.; There is only one previous report of an estrogen-secreting adrenal tumor occurring in a woman during reproductive years . Our patient presented with mild hirsutism associated with menstrual bleeding every 3-6 weeks . The occurrence of apparently intermenstrual bleeding prompted an evaluation of estrogen levels . Markedly elevated plasma estrone levels were found (860-2305 pmol/L; normal, 50-340) . Lesser relative elevations in 11-deoxycortisol and androstenedione were noted . Computed tomographic scanning of the adrenal glands identified a large tumor, which was subsequently resected . Estrone levels fell to 120 pmol/L, and all other abnormalities were corrected . Eighteen months after adrenalectomy, ovulation occurred regularly, and steroid levels were entirely normal . Steroid production in a cell suspension made from tissue obtained from the 190-g tumor was compared with that occurring in normal human adrenal cells . The production of estrone by the tumor cells was 40-fold greater than that by normal adrenal cells . There was also a mild excess of 11-deoxycortisol produced by tumor cells, but the tumor cells were less than 50% as efficient as normal cells in producing cortisol, dehydroepiandrosterone, androstenedione, testosterone, and dehydroepiandrosterone sulfate . Examination of the steroid profile in plasma occurring in three other patients with adrenal tumors reveals that while elevations in estrone occur frequently, this is usually due to the peripheral conversion of very high levels of androstenedione . Estrone, androstenedione, and 11-deoxycortisol plasma levels were elevated in all four patients; dehydroepiandrosterone sulfate was elevated in only two of four patients . After resection of one of these tumors, all steroid levels remained normal despite the occurrence of extensive metastases . These observations confirm the difficulty of making a diagnosis of estrogen excess in a woman during reproductive years because of the paucity of physical signs . The acquisition of aromatase activity was clearly demonstrated by tumor cells from our patient in vitro . Elevated plasma concentrations of estrone, androstenedione, and 11-deoxycortisol provide useful markers for adrenal tumors, but no one steroid can be relied upon in all tumors, and metastases may lack the steroidogenic capabilities of the primary tumor. Exp Brain Res, 1990, 82(3), 641 - 50 Intrahippocampal cholinergic grafts in aged rats compensate impairments in a radial maze and in a place learning task; Schenk F et al.; Age-related cognitive impairments were studied in rats kept in semi-enriched conditions during their whole life, and tested during ontogeny and adult life in various classical spatial tasks . In addition, the effect of intrahippocampal grafts of fetal septal-diagonal band tissue, rich in cholinergic neurons, was studied in some of these subjects . The rats received bilateral cell suspensions when aged 23-24 months . Starting 4 weeks after grafting, they were trained during 5 weeks in an 8-arm maze made of connected plexiglass tunnels . No age-related impairment was detected during the first eight trials, when the maze shape was that of a classical radial maze in which the rats had already been trained when young . The older rats were impaired when the task was made more difficult by rendering two arms parallel to each other . They developed an important neglect of one of the parallel tunnels resulting in a high amount of errors before completion of the task . In addition, the old rats developed a systematic response pattern of visits to adjacent arms in a sequence, which was not observed in the younger subjects . None of these behaviours were observed in the old rats with a septal transplant . Sixteen weeks after grafting, another experiment was conducted in a homing hole board task . Rats were allowed to escape from a large circular arena through one hole out of many, and to reach home via a flexible tube under the table . The escape hole was at a fixed position according to distant room cues, and olfactory cues were made irrelevant by rotating the table between the trials . An additional cue was placed on the escape position . No age-related difference in escape was observed during training . During a probe trial with no hole connected and no proximal cue present, the old untreated rats were less clearly focussed on the training sector than were either the younger or the grafted old subjects . Taken together, these experiments indicate that enriched housing conditions and spatial training during adult life do not protect against all age-related deterioration in spatial ability . However, it might be that the considerable improvement observed in the grafted subjects results from an interaction between the graft treatment and the housing conditions. Dev Neurosci, 1990, 12(4-5), 326 - 39 The myelin-deficient rat mutant: partial recovery of oligodendrocyte maturation in vitro; Espinosa de los Monteros A et al.; The morphological and immunocytochemical identification and characterization of the myelin-forming cell, the oligodendrocyte, have defined a model system for developmental studies . The myelin-deficient (md) rat mutant lacks myelin in the central nervous system and fails to express the normal developmental increase in oligodendroglial and myelin markers, apparently as a consequence of a point mutation in the proteolipid protein gene . In the present work, we compared the developmental pattern of primary glial cultures derived from newborn md rat brains to those derived from wild-type animals . Brain cell suspensions were prepared from each rat pup and cultured separately . We found by immunocytochemical and enzymatic analyses for the various markers that the developmental cascade of oligodendroglial marker expression is delayed, oligodendrocytes failing to mature compared to normal cultures . However, a partial recovery of marker expression was observed in md-derived cultures as compared to development previously reported in the intact md animals . We suggest that the partial recovery of the sequential expression of oligodendroglial markers may be due to a supportive environment provided to the oligodendrocyte progenitor cell (1) by tissue culture conditions or (2) by the absence of the blood-brain barrier in contrast to its presence in the intact animal. Cancer Immunol Immunother, 1990, 32(3), 173 - 8 Postoperative active specific immunization in curatively resected colorectal cancer patients with a virus-modified autologous tumor cell vaccine; Lehner B et al.; Active specific immunotherapy was performed in a phase I study in 20 colorectal cancer patients after surgical resection of the tumor . An autologous tumor cell vaccine surface modified by Newcastle disease virus (NDV) was used, which showed the following characteristics . After mechanical and enzymatic dissociation of the tumor tissue an average of 5 x 10(7) cells/g tissue was obtained . According to trypan blue dye exclusion assay the average viability was 72% . Following irradiation (200 Gy) the inactivation of proliferative activity of the cells could be demonstrated by the absence of incorporation of 3H-labelled thymidine . The cells were, however, still metabolically active as shown by the incorporation of {3H}-uridine and a mixture of 3H-labelled amino acids . Epithelium-specific antigens (detected by mAb HEA125) were expressed on more than 75% cells of the cell suspension indicating a high amount of (epithelium-derived) tumor cells . In order to increase the immunogenicity of the tumor cells the suspended cells were infected by the nonlytic, apathogenic Ulster strain of NDV . The successful modification of tumor cells with NDV could be shown by electron microscopy . Three weeks postoperatively cells were thawed, virus-modified, and inoculated intradermally in the upper thigh . Several cell and virus concentrations were tested in each patient . As control, tumor cells without NDV, NDV alone and normal colon mucosa were used . The number of tumor cells ranged from 2 x 10(6) up to 2 x 10(7) cells and NDV concentrations from 4 to 64 hemagglutination units (HU) were tested . Sixteen patients responded with a delayed-type hypersensitivity (DTH) skin reaction to the vaccine . The best DTH reaction, measured 24 h following vaccination, was obtained using a vaccine consisting of 1 x 10(7) tumor cells and 32 HU NDV (median induration of 8 mm) . Response to NDV alone was seen in 2 patients only (median induration of 3 mm); 12 patients responded to tumor cells (1 x 10(7) alone (median induration of 4 mm) . Of 10 patients tested with normal colorectal mucosa, 4 responded with a median induration of 3.5 mm . DTH responses to the vaccine of 1 x 10(7) tumor cells and 32 HU NDV increased throughout the repeated vaccinations to a median induration of 9.5 mm at the end of the therapy . No severe side-effects in the course of the immunotherapy, except for mild fever in 4/20 patients, were observed . The results of our phase I study show that this type of autologous colorectal tumor cell vaccine is ready for a large clinical trial to prove its efficacy. Yao Xue Xue Bao, 1990, 25(6), 469 - 72 {The effects of HCG and LHRH-A on progesterone secretion by human placenta villi cells of early and term gestation in vitro}; Zhu BT et al.; In the present study, the method of collagenase-pepsin prepared cell suspension of human placental villi from early (10-12 weeks) and term (38-40 weeks) gestation cultured in vitro combined with progesterone radioimmunoassay was employed . The results indicated that: 1 . In the placenta villi cells from early and term gestation in vitro, both exogenous and endogenous hCG showed a stimulatory effect on progesterone secretion, and the effect was approximately proportional to the concentration of the exogenous hCG added . 2 . In the placenta villi cells from term gestation in vitro, after addition of sufficient rabbit antihCG IgG to the culture medium, LHRH-A exhibited no obvious influence on this hCG-independent progesterone secretion . However, in the absence of rabbit anti-hCG IgG in the culture medium, LHRH-A showed a stimulatory action on progesterone secretion probably via its stimulatory effect on hCG secretion which could subsequently increase the progesterone secretion . 3 . In the placenta villi cells from early gestation, LHRH-A exhibited an inhibitory effect on both hCG-independent and hCG-dependent progesterone secretion. Clin Lab Haematol, 1990, 12 Suppl 1, 13 - 21 Recommended methods for the assignment of assay values to stabilized cell suspensions; England JM; Stabilized blood suspensions are required for the transfer of data from accurately calibrated reference instruments to service laboratory instruments . However, the more the cells are stabilized to increase shelf life, the less like fresh blood they become . Fixation is quite satisfactory for producing red count standards but it affects the flexibility, shape factor and the assigned MCV of the erythrocytes . Because of these problems values must be assigned indirectly by assigning values to fresh blood by reference methods and subsequently comparing fresh bloods with stabilized suspensions on a range of user instruments . The details for these methods are given and the reasons for these requirements are discussed. Arch Dermatol Res, 1990, 282(6), 402 - 7 Electron microscopic study of cultured cells from the murine hair tissues: cell growth and differentiation; Tanigaki N et al.; The cultured hair cells from 4-day-old C3H mice were studied by electron microscopy . The hair roots isolated from the skin by collagenase digestion were dispersed into a cell suspension by treatment with a mixture of trypsin and ethylenediaminetetraacetate . The cells were cultured in MCDB-153 (a medium containing seven growth factors) for 1, 3, 6 or 13 days . The number of cultured cells on day 3 was twice that on day 1, and stayed at the same level until day 13 . By electron microscopy, some of the cells cultured for 1 day were seen to be undifferentiated and others already showed differentiation into various hair structures . Such differentiated cells disappeared on day 3 and most of the cells cultured for 3 days were undifferentiated . Cell cultured for 6 days were differentiated showing inner root sheath cell, hair cortical cell and medulla cell structures . The characteristics of these cultured cells corresponded well to those of in vivo cells of the hair tissues from the back skin of 7-day-old C3H mice . On day 13 degeneration occurred in the cultured cells . In none of these cultures were mesenchymal cells, such as fibroblasts, found . The present electron microscopic study reveals that immature cells obtained from mouse hair tissues proliferate in vitro and differentiate into several subpopulations corresponding to those of in vivo cell layers of hair tissues . The present culture technique may be useful for studies of hair cell growth and differentiation. Tsitologiia, 1990, 32(7), 712 - 9 {The relationship of damage to and death of ascitic tumor cells during starvation to the ATP and free calcium content in the cells}; Gabai VL et al.; Ascite tumor cells EL-4 were incubated in conditions of energy starvation (Hanks salt solution with rothenone and without glucose) at 37 degrees C for 3 hours . Under these conditions, some structural cell damages appeared within the first hours: enlarging and flattening of the cells, blebbing, vacuolization of the cytoplasm, nuclear chromatin condensation . Later on, a share of cells with obvious damage decreased, whereas that of the cells stained with trypan blue (dead cells) much increased (up to 90% after a 3 hour incubation) . The cellular ATP decreased abruptly (up to 10% of the control) during the first 10 minutes of starvation . Free Ca2+ concentration increased within 1 hour of incubation more than two-fold . The conditions promoting Ca2+ influx (ionophore A23187 + Ca2+ in medium) accelerated the damage and cell death . However, the increase in free Ca2+ concentration did not trigger any damage in the energy-starved cells, since in the Ca2(+)-depleted medium (no increase in free Ca2(+)-concentration) the development of damages was not prevented . The damage initiation was irreversible: the addition of glucose to cell suspensions after 0.5-1 hour of their incubation in energy-starved condition did not prevent the development of damage, while ATP content in these cells was much increased. Ann N Y Acad Sci, 1990, 600, 384 - 402; discussion 402-4 Aging and regenerative capacity of the rat serotonergic system . A morphological, neurochemical and behavioral analysis after transplantation of fetal raphe cells; Steinbusch HW et al.; Morphological dissimilarities between the brains of young (3 months) and aged (28 months and older) rats were demonstrated using serotonin-immunocytochemistry . A degeneration of the serotonergic system, noted as a decreased innervation and the appearance of enlarged or swollen varicosities, was observed particularly in the frontoparietal cortex, and the neostriatum of the aged rat brain . No direct relationship between this aberrant morphology and decrease in density of serotonin-innervation was found as we demonstrated a decline in fiber density without the appearance of aberrant serotonergic fibers in the hippocampus . HPLC analysis revealed that serotonin (5-HT) and 5-hydroxyindolacetic acid (5-HIAA) levels in the frontoparietal cortex, hippocampus and raphe area were increased in the aged rat, while the 5-HT level in the caudate-putamen complex was not different from the young adult rat . The ratio 5-HIAA/5-HT, indicative of 5-HT turnover, appeared increased in the frontoparietal cortex, sensoric part, the caudate-putamen and the raphe area, while this ration in the frontoparietal cortex, motoric part and the hippocampus was not altered in the aged rat . Behavioral screening revealed a decrease spatial performance of aged males in a Morris Water-Maze task . To investigate whether the age of the host recipient was of influence on the regenerative capacity, a fetal raphe cell suspension of embryonic day E 15 was implanted in the caudate-putamen of young adult as well as aged rats . Neither differences in survival of the serotonergic cells nor in fiber outgrowth between both groups appeared five weeks after transplantation . Subsequently, transplantation of raphe cells in the hippocampus of young adult rats, after lesioning the hippocampal serotonergic innervation with 5,7-DHT, was performed to compare behavioral, morphological and neurochemical effects of the implants . It appeared that 11 months after transplantation the serotonergic innervation of the previously denervated hippocampus was greatly restored . There was a striking resemblance between the immunohistochemical and neurochemical data with respect to the increase in the amount of newly formed serotonergic fibers, the increase in uptake of {3H}-5-HT and in 5-HT and 5-HIAA levels . Also the behavior of lesioned and lesioned + transplanted males was rather similar to controls . In the behavioral tests we were mainly interested in hippocampal functioning, therefore orientation was of our prime interest . The other behavioral tests were only to confirm that the possible changes were linked to hypothalamic or extra-hypothalamic functions.(ABSTRACT TRUNCATED AT 400 WORDS) Am J Pediatr Hematol Oncol, 1990 Fall, 12(3), 245 - 56 Ex vivo chemopurging of autologous bone marrow with 4-hydroperoxycyclophosphamide to eliminate occult leukemic cells . Laboratory and clinical observations; Yeager AM et al.; The application of autologous bone marrow transplantation (ABMT) in treating acute leukemias in children has been limited by the presence of residual occult viable leukemic cells in the marrow cell suspension . One approach to this problem is the ex vivo treatment ("purging") of the autograft to eradicate these tumor cells yet spare the normal lymphohematopoietic stem cells . Initial studies of acute myeloid leukemia (AML) in a rodent model demonstrated that incubation with 4-hydroperoxycyclophosphamide (4HC), a congener of cyclophosphamide and an active alkylating agent in aqueous solution, could effectively eliminate viable AML cells from marrow cell suspensions without apparent toxicity to normal stem cells . We have conducted clinical trials of ABMT with 4HC-treated marrow in children with acute leukemia in remission; marrow was collected, treated ex vivo with 4HC (100 micrograms/ml), and cryopreserved in liquid nitrogen until reinfusion . Children received pre-ABMT conditioning with either high-dose cyclophosphamide and total body irradiation (CY-TBI) for acute lymphocytic leukemia (ALL) or high-dose busulfan and cyclophosphamide (BU-CY) for AML . Of nine children who underwent ABMT with 4HC-treated marrow for ALL in second complete remission (CR2), all relapsed (eight in the marrow, one in the central nervous system) at a median of 5 months (range, 2-17) after ABMT and all have died with relapsed ALL or as a consequence of its treatment . Twenty-nine children with AML (five in CR1, 24 in CR2) received autografts with chemopurged marrow at a median remission duration of 3 months (range, 2-15) . Three patients died from sepsis during aplasia; 10 children (one in CR1 and nine in CR2) relapsed with AML at a median of 7 months (range, 2-23) after ABMT, for an actuarial relapse rate of 47% . Sixteen patients with AML (four in CR1, 12 in CR2) are in unmaintained remission at a median of 16 months (range, 6-102) after ABMT, for an actuarial disease-free survival of 49% . Although ABMT with 4HC-treated marrow appears to have a limited role in the treatment of children with ALL who lack a suitable related donor, the results in AML are encouraging and compare favorably with both syngeneic and allogeneic BMT in similar groups of patients. Invasion Metastasis, 1990, 10(5), 281 - 8 Lymphatic metastases from the peritoneal cavity are increased in the postinflammatory state; Levine S et al.; Cell suspensions of chemically induced tumors (rhabdomyosarcoma) were transplanted into the peritoneal cavities of Lewis rats . In normal animals, the greater omentum was the main site of tumor growth, and transdiaphragmatic metastases to regional lymph nodes in the mediastinum were few and small . In animals during the healing phase of a chemical peritonitis, the greater omentum was fibrotic, shrunken, and inactivated . The loss of the scavenging function of the omentum was associated with wide dissemination of the tumor in the peritoneal cavity and increased access of the tumor to the lymphatic stomata on the peritoneal surface of the diaphragm . Number and size of transdiaphragmatic metastases in draining lymph nodes were greatly increased in this postinflammatory state. Int J Syst Bacteriol, 1990 Jan, 40(1), 66 - 70 Arylsulfatase activity of Mycobacterium avium, M . intracellulare, and M . scrofulaceum; Falkinham JO 3rd; A rapid (3-h) arylsulfatase assay for cell suspensions of mycobacteria, in which p-nitrophenyl sulfate is used as the substrate, was developed . Arylsulfatase activity was found in cell suspensions of representative strains of Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium scrofulaceum grown without the substrate in either Middlebrook 7H9 medium containing 0.2% (wt/vol) glucose and 0.05% (vol/vol) Tween 80 or Dubos broth medium, but was absent in cells grown in a low-pH, minimal medium containing 1% (vol/vol) Tween 80 as the sole carbon source . The levels of arylsulfatase activity of representatives of all three species were equal whether the activity was measured at pH 5.5, 6.5, or 7.5 and whether the cells were suspended in phosphate or Tris buffer . The addition of high levels of sulfate (present in the low-pH, Tween 80-containing medium) to Middlebrook 7H9 medium resulted in significantly lower levels of arylsulfatase activity in strains of M . scrofulaceum, but did not affect the levels in either M . avium or M . intracellulare . The levels of arylsulfatase activity were highest in M . avium, intermediate in M . intracellulare, and lowest in M . scrofulaceum strains . Polyacrylamide gel electrophoresis of crude extracts from late-log-phase cells of representatives of each species produced activity bands of unique mobility (one in M . avium, three in M . intracellulare {82, 5, and 13%}, and two in M . scrofulaceum {60 and 40%}). Acta Leprol, 1990, 7(2), 157 - 61 Isolation and characterization of infiltrates in the nerves of patients with neuritic leprosy; Kumar V et al.; A study was done on the characteristics of infiltrating cells in the nerves of 9 patients with pure neuritic leprosy, by preparing a single cell suspension . The patients had no skin lesion . Histopathological examination revealed that 2 of the 9 nerves showed granulomas characteristics of tuberculoid leprosy, while the remaining 7 had features of lepromatous granulomas . In the nerves showing tuberculoid granulomas, a high proportion of lymphocytes were T cells as they formed rosettes with sheep erythrocytes and only a few percent were EAC rosette forming cells . On the other hand, the nerves showing lepromatous granulomas contained only occasional lymphocytes which formed E and EAC rosettes . Macrophages from the granulomas of all the nerves were esterase positive, peroxidase negative, contained M . leprae and did not exhibit C3 surface receptors. J Clin Gastroenterol, 1990, 12 Suppl 1, S177 - 80 Protective effect of sofalcone and 16,16-dimethyl-PGE2 on isolated rat gastric cells; Kobayashi K et al.; The effects of 16,16-dimethylprostaglandin E2 (dm-PGE2) and sofalcone on damage caused by ethanol in surface epithelial cells isolated from rat stomach were examined . The surface epithelial cells (SECs) accounted for 83% of the isolated gastric mucosal cells, mucus neck cells for 5%, parietal cells for 9%, and chief cells for 3% . To suspensions of SEC, either dm-PGE2 at concentrations of 10(-7), 10(-6), or 10(-5) M or sofalcone at concentrations from 5 X 10(-6) to 10(-4) M was added; some cells were treated with neither drug . Ten minutes later, ethanol was added to each cell suspension to a final concentration of 15%, and 5 min later the viability was evaluated by trypan blue exclusion . At 10(-6) M, dm-PGE2 reduced ethanol-induced cell damage most strongly (p less than 0.001) . Sofalcone also helped to prevent cell damage caused by ethanol in a dose-dependent manner . These results suggest that dm-PGE2 and sofalcone protect gastric mucosa not only from gross visible damage but they directly protect gastric cells from damage. Folia Med Cracov, 1990, 31(1-2), 73 - 9 {Binding of various polycyclic aromatic hydrocarbons (PAH) by plasma albumin and blood cells}; Lipniak M et al.; We have studied in vivo and in vitro partition of pyrene, fluoranthene and benz(a)anthracene (BaA) between blood cells and plasma or phosphate buffer pH 7.4 and their binding to bovine serum albumin . The level of PAH in blood and plasma was determined in rats after i.v . administration of pyrene, fluoranthene and BaA in dose 20 mg/kg by gas chromatography method . In vitro 20 micrograms of PAH was added to blood samples or blood cell suspension . These samples were incubated at 37 degrees C . After 1 hr the concentration of PAH in blood cell and plasma or buffer was determined . The data indicate that whole pyrene and fluoranthene are distributed to blood cells . 39% of BaA in whole blood is present in blood cells and 56.9% in plasma . Plasma protein binding of PAH were determined by gel filtration 0.4 cm3 of PAH-protein solution was given on a Sephadex column . PAH binding parameters were obtained from simple linear regression of Scatchard plots of the data . In a albumin solution (0.4%) the number of binding sites was 2 for BaA, 3 for pyrene and 11 for fluoranthene . The binding constants for BaA, fluoranthene and pyrene were 0.17; 0.71 and 1.11 mol-1 X 10(4), respectively. Pathology, 1990 Jan, 22(1), 5 - 9 Comparison of flow cytometry and retrospectively applied static cytometry on lymphoid tissue; Jones A et al.; Twenty cases of non-Hodgkin's malignant lymphoma were examined using both flow cytometry (FCM) and static cytometry (SCM) DNA analysis to detect aneuploidic cell populations . FCM was performed on fresh cell suspensions whilst SCM was performed retrospectively on formalin-fixed, paraffin-embedded samples of the same tissue . A total of 34 aneuploidic cell lines were detected by FCM compared to a total of 41 detected by SCM . Of the 20 cases 14 showed DNA indexes within 10% of each other for each method, indicating a reasonable degree of comparability between the two methods . Of the 6 cases which did not show comparable DNA indexes, 2 displayed similar cell lines for both methods, but the major abnormal cell populations were of different ploidy by SCM as compared to FCM . A qualitative comparison is thus possible between these two cases . The results suggest that retrospectively applied SCM ploidy analysis of lymphoid tissue is comparable to FCM on fresh tissue samples and can thus be used in retrospective studies of both prognostic and diagnostic significance. Biorheology, 1990, 27(1), 47 - 65 Further investigations of red cell deformability with nickel mesh; Arai K et al.; Although the filtration method has been widely employed in red cell deformability studies, the structural irregularity of the pores of a Nuclepore polycarbonate membrane has always been a major problem . Anegawa, T . et al . (Clin . Hemorheol., 7, 1987) obtained a higher reproducibility with the filtration method using a newly designed thin metal film with pores engraved by the photofabrication technique . We further studied the pressure - flow rate relationship of red cell suspension employing this nickel mesh . The filtration of red cell suspensions through the nickel mesh was not influenced by leukocytes contamination or added leukocytes up to a leukocyte count of 250 cells/mm3 within an experimental limitation . On the other hand, the flow was greatly influenced by leukocytes contamination when the polycarbonate membrane was used . The nickel mesh was found to be useful in detecting major determinants of red cell deformability, such as cell geometry and internal cellular viscosity, and in detecting abnormalities of red cell deformability in a patient with microangiopathic hemolytic anemia . In conclusion, the present study clearly shows that the nickel mesh is preferable for investigating red cell deformability to the polycarbonate membrane from a quantitative point of view . This material should contribute to the physiologic and clinical investigation of red cell deformability. Haematologica, 1990 Jan-Feb, 75 Suppl 1, 1 - 5 Processing and manipulation of human hemopoietic stem cells . The Italian Cooperative Study Group on Cell Manipulation in Hematology; Iacone A; In this paper, we reviewed the basic principles and methods of collection, processing and manipulation of marrow and peripheral haemopoietic progenitor cells . The advances in cell separation are due to the progress in blood bank technology and to the introduction of more sophisticated devices . This allows to process larger amounts of blood or marrow and to obtain a final single cell suspension . For this purpose, several procedures have been employed, including the unit gravity sedimentation, the centrifugal elutriation, the isopyknic sedimentation and the positive selection on columns of CD34+ cells . As expected, the emerging problems are the extreme heterogeneity of the methods employed and the unreliability of the quality control . The proposal for standardized protocols for each step of the various procedures is urgent, and the institution of multicenter studies is recommended. Cancer Chemother Pharmacol, 1990, 26(2), 122 - 6 Hepatic tissue distribution of doxorubicin-loaded nanoparticles after i.v . administration in reticulosarcoma M 5076 metastasis-bearing mice; Chiannilkulchai N et al.; In our previous studies, doxorubicin-loaded polyisohexylcyanoacrylate nanoparticles have been proven to increase dramatically the antitumoral activity of the cytotoxic agent in metastasis-bearing mice . The experimental model consisted of metastases induced by i.v . inoculation of reticulosarcoma M 5076 cell suspension to C57BL/6 mice . The improved efficacy of the drug was noted in terms of either metastasis count or survival . Therefore, tissue-distribution studies of this drug delivery system within the metastatic liver after i.v . administration were undertaken to gain more insight into the mechanism of action . Doxorubicin measurements in healthy hepatic or neoplastic tissue were carried out together with histological examinations using transmission electron microscopy . These results demonstrated the hepatic tissue to be an efficient reservoir of the drug when it was injected associated with nanoparticles . Accumulation of biodegradable nanoparticles with associated doxorubicin in Kupffer cells created a gradient of drug concentration for a massive and prolonged diffusion of the free drug towards the neoplastic tissue. Annu Rev Neurosci, 1990, 13, 415 - 40 Dopamine cell replacement: Parkinson's disease; Yurek DM et al.; Significant progress in neural transplantation has been observed over the last several decades . As a neuroanatomical tool, neural transplantation studies are able to examine the mechanisms involved in the development and integration of neurons into the complex neural circuitries of the brain . Today, embryonic neural tissue can be successfully transplanted as solid tissue chunks or as dissociated cell suspensions . Within the parenchyma of the brain, transplanted embryonic neurons develop mature morphology and do not appear to invoke an immunological response by the lost immune system . Not only do these neurons exhibit robust development but there is also evidence that transplanted neurons restore some degree of function to neurologically damaged circuitry; however, the extent of reintegration into the host neural circuitry still remains unclear . Moreover, the long-term survival and functioning of transplanted nerve cells also remains an unanswered question . Advances in the emerging field of genetic engineering may eventually lead to genetically modified neurons that are capable of synthesizing neurotrophic factors or missing neurotransmitters and restoring function in brain-damaged areas . The use of neural transplantation to replace damaged nerve cells in neurodegenerative disorders, such as Alzheimer's or Parkinson's disease, is promising based on our current knowledge . However, our basic scientific knowledge of neural transplants is incomplete and warrants a prudent approach toward application of neural transplantation techniques in clinical research. Acta Obstet Gynecol Scand, 1990, 69(2), 153 - 9 Detection of human papilloma virus in women referred for colposcopy . A comparison between different diagnostic methods; Boden E et al.; Various methods presently available for the diagnosis of genital Human Papilloma Virus (HPV) were compared regarding their sensitivity in women referred for specific diagnosis and treatment because of atypical Pap smears or clinically suspected neoplasia . Colposcopic examination was performed in all cases . In addition to taking a second Pap smear, cell suspensions were made from 105 women and analysed by the Filter In Situ Hybridization (FISH) technique and tested for HPV 6 + 11 and HPV 16 + 18 + 31 . The FISH technique was also used for the possible detection of HPV-DNA in a reference material comprising 119 apparently healthy women with normal Pap smears . Colposcopically directed cervical biopsies in altogether 196 specimens were obtained from 155 women for histopathological examination and also for the detection of HPV-DNA by the Southern blot hybridization technique . These specimens were tested for the presence of HPV 6, 11, 16, 18, 31 and 33 . Three per cent of the women with and 34% without cytological signs of HPV in Pap smears had cervical intraepithelial neoplasia (CIN) III according to histopatology . CIN III was present in 35% of biopsies with and 59% of biopsies without histological signs of HPV in the biopsies . Altogether 46% of the women were HPV-DNA positive . Of the women analysed by Southern blot, 39% were HPV-DNA positive . Of the samples analysed by FISH, 27% with atypical cells were HPV-DNA positive, compared with 11% of the samples from reference women with normal cytology.(ABSTRACT TRUNCATED AT 250 WORDS) Zh Nevropatol Psikhiatr Im S S Korsakova, 1990, 90(2), 12 - 5 {Prognostic value of NA K ATPase activity in multiple sclerosis}; Ierusalimskii AP et al.; The paper is concerned with the results of measuring the activity of Na+, K+, ATPase in red blood cell suspension in patients with different courses of multiple sclerosis (MS) within the range of physiological temperatures (34 to 42 degrees C) . Account was taken of the interrelation of structural phasic transitions in cell membranes to the changes in enzymatic activity . There was a spasmodic increase of the activity of Na+, K+, ATPase at 35 degrees C and 40 degrees C at the moment of the clinical signs of disease exacerbation together with a twofold lowering of the enzymatic activity at a temperature of 35 degrees C during remission . The rise of the enzymatic activity at a temperature of 35 degrees C anticipates clinical exacerbation and thus may be of prognostic importance in the determination of the type of the disease course. J Steroid Biochem, 1990 Jan, 35(1), 127 - 32 Pituitary and gonadal function during physical exercise in the male rat; Harkonen M et al.; The effects of training and acute exercise on serum testosterone, luteinizing hormone (LH) and corticosterone levels and on testicular endocrine function in male rats were studied . In the first part of the study, the rats were trained progressively on a treadmill, over 8 weeks . Training did not change the basal levels of serum testosterone, LH and corticosterone, or the testicular concentrations of testosterone and its precursors progesterone and androstenedione . The levels of testicular LH (30.3 +/- 2.6 ng/g wet wt, mean +/- SEM) and lactogen (150 +/- 14 pg/g) receptors were unchanged after training . However, the capacity of testicular interstitial cell suspensions to produce cAMP and testosterone increased by 20-30% during in vitro gonadotropin stimulation . In the second part, the trained and untrained control animals underwent acute exhaustive exercise . Serum testosterone levels decreased by 74 and 42% in trained and untrained rats, respectively (P less than 0.02), and corticosterone rose by 182% in trained and 146% in untrained rats (P less than 0.01), whereas the LH level was unchanged . Testicular levels of testosterone and its precursors decreased, with the exception of unchanged androstenedione, in trained rats; the cAMP concentration was unchanged . In both trained and untrained rats, acute exercise decreased the capacity of interstitial cell suspensions to produce cAMP, whereas there were no consistent effects on testosterone production . Acute exercise had no effect on LH or lactogen receptors in testis tissue . In conclusion, training had no effect on serum or testicular androgen concentrations, but increased Leydig cell capacity to produce testosterone and cAMP . Acute exercise decreased serum and testicular testosterone concentrations without affecting serum LH . A direct inhibitory effect of the increased serum corticosterone level on the hypothalamic-pituitary level and/or testis may be the explanation for this finding. Am J Physiol, 1990 Jan, 258(1 Pt 2), F52 - 60 Influence of Na+ intake on dopamine-induced inhibition of renal cortical Na(+)-K(+)-ATPase; Seri I et al.; The enzyme L-amino acid decarboxylase (L-AADC), found in abundance in rat proximal tubule cell cytosol, converts L-dopa to dopamine . Dopamine, in turn, suppresses proximal tubule sodium transport by inhibiting Na(+)-K(+)-ATPase activity . We sought to determine whether changes in dietary sodium intake in rats lead to adaptation of dopamine formation and dopamine-induced Na(+)-K(+)-ATPase inhibition . In rats on a high-salt (HS) diet, the maximal velocity (Vmax) of renal cortical L-AADC was 78 +/- 19% higher than that in rats on a low-salt (LS) diet . The Michaelis constant (Km) of the enzyme remained unchanged . In renal cortical tubule cell suspensions the L-dopa-induced inhibition of ouabain-sensitive oxygen consumption (QO2) was significantly greater in rats on HS diet than in rats on LS diet . Furthermore, L-dopa completely inhibited the nystatin-induced rise in QO2 in the HS but not in the LS group . Carbidopa, an inhibitor of L-AADC, abolished the L-dopa-induced inhibition of nystatin-stimulated QO2 in cells from HS rats and was without significant effect in cells from LS rats . L-Dopa-stimulated K+ efflux was greater in cells from HS rats at 28 +/- 1 nmol.min-1.mg protein-1, compared with 7 +/- 6 nmol.min-1.ng protein-1 in cells from LS rats . By contrast, ouabain-stimulated K+ efflux did not differ between the groups.(ABSTRACT TRUNCATED AT 250 WORDS) Br J Dermatol, 1990 Jan, 122(1), 33 - 42 Antigen presentation in patients with recrudescent orofacial herpes simplex virus infections; Vestey JP et al.; Recovery from epidermal herpes simplex virus (HSV) infection depends primarily on development of an effective cell-mediated immune response, possibly generated following antigen (Ag) presentation by epidermal cells (EC) . The ability of EC to present HSV Ag was investigated in 12 subjects with occasional recrudescent facial HSV infections . All had circulating HSV specific antibodies and cell-mediated immunity to the virus . Peripheral blood mononuclear cell suspensions, depleted of antigen presenting cells (APC) by glass adherence and then enriched for T cells by adsorption on nylon wool columns, did not proliferate in response to HSV Ag . Both EC suspensions, prepared from suction blister roofs, and glass-adherent peripheral blood mononuclear cells (AC) preincubated with ultraviolet-inactivated HSV, reconstituted the T-cell proliferative response to HSV . EC were more efficient than AC at presenting HSV Ag to T cells . Depletion of CD1+ cells from EC suspensions by cell sorting reduced their ability to present HSV Ag and augmentation of CD1+ cell numbers supplemented it . Preincubation of EC or AC with monoclonal antibodies to major histocompatibility complex class II antigens DP, DQ or DR, blocked the lymphoproliferative response to HSV Ag . Evidence was obtained that cells co-ordinately expressing products of the DP, DQ and DR loci are involved in presentation of HSV Ag by both EC and AC. Arch Biochem Biophys, 1990 Jan, 276(1), 160 - 71 Stimulation of human erythrocyte 2,3-bisphosphoglycerate phosphatase by vanadate; Mendz GL et al.; The rates of vanadate-stimulated hydrolysis of 2,3-bisphosphoglycerate in metabolically competent erythrocytes and in hemolysates were determined from data on time courses up to 35 min employing 31P nuclear magnetic resonance spectroscopy . The enhanced rate of hydrolysis of the bisphosphate was attributed principally to the activation of the phosphatase activity of 2,3-bisphosphoglycerate synthase both in cell suspensions and in hemolysates . Information on the concentrations of vanadate and vanadyl present in the preparations was obtained employing 51V nuclear magnetic resonance spectroscopy and electron paramagnetic resonance spectroscopy . Redox reactions involving vanadium ions appeared to be important in establishing the final equilibrium concentrations of the oxy- and oxo-ions (vanadate and vanadyl, respectively), but the data suggested that the activation of the enzyme resulted from direct action of the vanadium ions on the enzyme and not as a consequence of the alteration in the equilibrium of intracellular oxidants and reductants. Membr Biochem, 1990 Jan-Mar, 9(1), 47 - 60 An Escherichia coli mutant conditionally altered in respiratory chain components; Cox JC et al.; Nitrosoguanidine mutagenesis was employed to isolate an Escherichia coli mutant conditionally altered in respiratory chain components . Mutant R25 was able to grow on glucose, fructose, and glycerol but failed to grow on succinate and acetate (suc-) . Also, R25 exhibited leaky growth on DL-lactate, fumarate, and malate (lct*) . The lct* mutation pleiotropically affected a number of respiratory chain components and its expression was conditional with the growth substrate . Glucose-grown R25 resting cell suspensions oxidized DL-lactate and formate; however, these two substrates were not oxidized by fructose- or glycerol-grown cell suspensions . The same conditional pattern was observed for the concentration of cytochrome components, the membrane-associated oxidation of NADH and formate, and formate phenazine methosulfate (PMS) reductase activity; succinate oxidase and PMS reductase activities were not exhibited by membranes under any growth condition due to the suc- mutation . R25 membrane-associated H(+)-translocating ATPase activity was not conditional with the growth substrate . R25PC, a spontaneous lct+ suc- partial revertant of R25, did not exhibit the conditional pattern of R25 . The lct* mutation was found to map in the 27-30-min region and the suc- mutation in the 15-17-min region of the E . coli genome . Two distinct classes of R25 P1kc transductants were isolated that differed in both their growth response on succinate and DL-lactate and their oxidase activities. Am J Med Genet Suppl, 1990, 7, 219 - 24 Intrathymic deficient expansion of T cell precursors in Down syndrome; Musiani P et al.; To correlate the intrinsic cellular immunodeficiency, which is a major cause of increased susceptibility to polytopic infections in Down syndrome (DS) patients, with the histologic abnormalities observed in the thymus of these patients, we have studied thymus fragments and thymocyte cell suspensions from 15 non-institutionalized DS subjects . Comparing to the control age-matched samples, a reduced thymic cortex and a distinct depletion of CD1-positive (+) cells was observed by immuno-histologic examination . The phenotypic analysis of unselected thymocytes showed a significant reduction of CD3+, CD1+, CD4+, and CD8+ cells . When the total thymocyte population was separated into 10 fractions, using a continuous Percoll density gradient, a difference in cell distribution was observed . DS thymuses are almost devoid of high-density thymocytes (fractions 6-9) while more than 75% of the cells were recovered in the lightest 3 fractions (Frs) . In addition, these thymuses were characterized by a marked depletion of CD1+ cells and by a conspicuous reduction of CD3+ cells normally present in the high-density Frs . On the other hand, the lightest 3 Frs of DS subjects were enriched in low-density CD1+ cells . Although enriched in these cells, normally characterized by a high mitotic activity, Fr1 DS thymocytes showed a reduced spontaneous proliferative capacity . When the expression of T cell receptor alpha- and beta-subunits was studied, the percentages of cells stained with anti-alpha and anti-beta antisera were found to be reduced in DS unfractionated thymocytes . The reduced number of high-density CD1+ thymocytes associated with a reduced spontaneous proliferative activity of low-density CD1+ thymocytes suggests that in DS thymuses there is a deficient expansion of immature T cells, resulting in a reduction of the various thymocyte subpopulations, including the thymocyte pool which differentiates into functionally mature T cells expressing the alpha-beta T cell receptor. Cytometry, 1990, 11(8), 883 - 7 Simultaneous cytofluorometric analysis for the expression of cytoplasmic antigens and DNA content in CD3- human thymocytes; Zocchi MR et al.; We describe a method of two-color immunofluorescence staining which allows the simultaneous analysis of both cytoplasmic antigens and cell entry into the S/G2/M cell cycle phases . This analysis was performed on CD3(-)-activated thymocytes obtained from either highly purified CD1-CD3-CD4-CD8- cells or fresh thymus cell suspensions, stimulated with low doses of phorbol-12 myristate-13 acetate (0.5 ng/ml) and interleukin-2 . On the 14th day under these culture conditions about 90% of thymocytes did not express CD3 antigen on the cell surface . CD3- cells were further purified by cell sorting, fixed in paraformaldehyde, and permeabilized with Nonidet-P40 . Then these thymocytes were stained by indirect immunofluorescence with monoclonal antibodies identifying T cell-specific molecules (CD3, CD2, CD28, TCR alpha/beta, and TCR gamma/delta) and analyzed for DNA content . Interestingly, both CD3 and CD28 antigens were detectable in the cytoplasm of most cells (greater than 80%) . Further, the majority of the thymocytes which had entered the S/G2/M phases of the cell cycle (20%) expressed intracellular CD3 and CD28 molecules and reacted with the anti-beta framework beta F1 monoclonal antibody . The relationship between the appearance of CD3 and other T cell markers in the cytoplasm, the cell cycle entry, and the thymocyte development is discussed. Cell Immunol, 1990 Jan, 125(1), 254 - 60 Tissue distribution and appearance in ontogeny of alpha/beta T cell receptor (TCR2) in chicken; Vainio O et al.; We have performed immunoperoxidase staining on cryostat tissue sections and immunofluorescence analysis on cell suspensions to identify cells expressing the alpha/beta T cell antigen receptor during ontogeny and adult life in chickens . We used the mouse monoclonal antibody, TCR2, which was previously shown to recognize the alpha/beta TCR in chickens . TCR2+ cells were observed in thymic cortex and medulla and in T-dependent areas of spleen, intestine, and cecal tonsils of young adult chickens . Some TCR2+ cells were found in the cortex of bursal follicles and in liver . The first TCR2+ cells appear in thymus on Day 13 of the embryonic life and it is only after hatching that TCR2+ cells begin to migrate to the periphery. Dev Immunol, 1990, 1(1), 59 - 66 Are the IL-2 receptors expressed in the murine fetal thymus functional? Zuniga-Pflucker JC, Smith KA, Tentori L, Pardoll DM, Longo DL, Kruisbeek AM. It is well established that the majority of murine fetal thymocytes (day 15 of gestation) express receptors for interleukin 2 (IL-2), but the functional significance of these IL-2 receptors (IL-2Rs) is not clear . In situ hybridization data show a developmentally regulated expression of IL-2 and IL-2R mRNA . IL-2 binding studies were performed on fetal thymocytes and the results show the presence of both high (kD approximately equal to 20 pM) and low (kD approximately equal to 10 nM) affinity IL-2Rs . These IL-2Rs are indeed functional: intact fetal thymic lobes (but not cell suspensions) cultured in IL-2 exhibited an in vitro proliferative response at 20 pM of IL-2, corresponding with the presence of a functional high-affinity IL-2R on fetal thymocytes . The IL-2-dependent growth was primarily observed in the IL-2R+ thymic subset, which contains the CD3-/CD4-/CD8- precursor thymocytes . Furthermore, in vitro blocking of IL-2 in intact fetal thymic lobes resulted in a reduction in the cell yield, which predominantly affected the expansion of the immature CD3-/CD4-/CD8- thymocytes . Our findings strongly support the concept that the IL-2/IL-2R pathway is responsible for the proliferation of precursor cells within the fetal thymus. Neuroscience, 1990, 37(2), 301 - 15 Fetal striatal neurons grafted into the ibotenate lesioned adult striatum: efferent projections and synaptic contacts in the host globus pallidus; Wictorin K et al.; The efferent projections to the host brain from intrastriatal grafts have been examined at the ultrastructural level . Cell suspensions of E14 rat fetal striatal tissue were implanted into the ibotenic acid lesioned caudate-putamen of adult rats . After survival times of at least five months, the anterograde neuronal tracer Phaseolus vulgaris leucoagglutinin was injected into the grafts . Consistent with our previous light microscopical analysis, anterogradely labelled fibres could be followed from the grafts into the host globus pallidus along the normal trajectory of the striatopallidal pathway in the internal capsule, and a few fibres also reached the entopeduncular nucleus and the substantia nigra . In the electron microscope, the graft-derived efferents were seen to be myelinated as they projected caudally along the fibre bundles of the internal capsule, and they were thus similar to the neostriatal projection in normal control animals . In the host globus pallidus, the graft-derived fibres ramified into a terminal network forming morphologically normal synaptic contacts with neuronal elements in the host globus pallidus . A total of 118 synaptic contacts were identified, all of them formed symmetric membrane specializations . The major postsynaptic targets were dendritic shafts (90%), with only a few contacts with spines or small shafts (8%) and perikarya (2%) . The morphology of the synaptic contacts and their distribution on different postsynaptic targets were similar to that which was found in the globus pallidus after tracer-injections into the caudate-putamen of normal control animals . The results show that grafted fetal striatal neurons can grow along the myelinated territory of the internal capsule to reinstate a seemingly normal synaptic input to the previously denervated neurons in the host globus pallidus. Braz J Med Biol Res, 1990, 23(6-7), 567 - 71 Depressed adjuvant arthritis in rats transferred with spleen cells from Trypanosoma cruzi-infected syngeneic donors; Revelli SS et al.; In the present study we investigated whether the attenuating effect of chronic Trypanosoma cruzi (Tc) infection on adjuvant arthritis (AA) in the rat could be transferred to naive recipients . Transfer of whole spleen cells, but not of serum, from Tc-infected rats reduced AA (means +/- SEM: 11 +/- 0.5) in recipient animals (control values, means +/- SEM: 19 +/- 0.7) . Transfer of a T-cell-enriched subpopulation from spleen cells of Tc-infected rats (obtained by filtration through a nylon wool column) resulted in a similar attenuation of AA (means +/- SEM: 7.5 +/- 2.2) . The arthritic response of rats intraperitoneally inoculated with 2 x 10(5) Tc 48 h before induction did not differ from that observed in controls . Neither parasites nor specific antibodies were observed in suckling mice inoculated with serum or cell suspensions employed in transfer experiments . Consequently, the depressive effect on AA could not be directly attributed to Tc per se . We hypothesize that a homeostatic immunosuppressor mechanism may be responsible for this phenomenon. Folia Biol (Praha), 1990, 36(2), 102 - 7 Ontogeny of MHC class II antigens in pigs; Trebichavsky I et al.; Cells constitutively expressing MHC class II antigens have been studied in the course of prenatal development in the Minnesota miniature pig . Frozen sections, cell suspensions and peripheral blood mononuclear cells were examined by using the HL-40 monoclonal antibody cross-reactive with a light chain determinant of the SLA-D molecule (MHC class II porcine antigen) . It could be demonstrated that the yolk sac contained cells expressing SLA-D antigens as early as the 24th day, the liver and the spleen on the 39th day of gestation . Splenic cells bearing SLA-D molecules formed periarteriolar structures . Except the spleen, peripheral blood, lymph nodes, bone marrow and intestinal wall were the main sites of SLA-D expression in the perinatal period of the pig. J Endocrinol Invest, 1990 Jan, 13(1), 13 - 8 TRH raises cytosolic Ca2+ in human adenomatous lactotrophs; Spada A et al.; The effect of TRH on cytosolic free calcium concentrations, {Ca2+}i, was evaluated on cell suspensions obtained from 6 human PRL secreting pituitary adenomas . In these cells resting {Ca2+}i levels were variable (mean +/- SE; 103.8 +/- 6.5, n = 25); the addition of 100 nM TRH caused a marked {Ca2+}i rise within 20 sec., the peak values ranging from 200 to 437 nM (285 +/- 10.8 nM, n = 10) . The transients induced by TRH were composed by a rapid increase, due to mobilization of calcium from intracellular stores, followed within a few seconds by a lower plateau which was due to stimulated influx from the extracellular space . In fact, when EGTA and verapamil were applied after TRH they caused the Ca2+ plateau to dissipate rapidly . The addition of 1 microM dopamine (DA) caused a substantial decrease of resting {Ca2+}i (about 10-30%) as well as an inhibition of the plateau phase induced by TRH . The effect of DA completely depended on extracellular Ca2+ . The TRH-induced transients observed in adenomatous cells were quite similar in size and time course to those recorded in normal rat lactotrophs . As previously observed in rat lactotrophs, in adenomatous cells treatment with pertussis toxin (PTx, 1 microgram/ml for 4 h) was unable to affect the {Ca2+}i transients induced by TRH while completely abolished the effect of DA . The effects of TRH on in vivo and in vitro PRL secretion were also evaluated . Before surgery, no patient showed a positive response to the iv administration of 200 micrograms TRH (serum PRL levels: 95 +/- 62 ng/ml in basal conditions vs 124 +/- 92 after TRH, P = NS).(ABSTRACT TRUNCATED AT 250 WORDS) Mod Pathol, 1990 Jan, 3(1), 54 - 60 Comparison of histologic nodal reactive patterns, cell suspension immunophenotypic data, and HIV status; Westermann CD et al.; While cell suspension immunophenotypic studies are widely used as an aid in the diagnosis and classification of lymphomas and leukemias, much less attention has been directed toward interpretation of the results in reactive lymphoid proliferations . Cell suspension immunophenotypic data were therefore analyzed for 119 lymph nodes with reactive lymphoid proliferations which were divided into five major histologic categories: follicular hyperplasia, marked (FH,M), or moderate (FH); dermatopathic lymphadenopathy (DL); diffuse hyperplasia (DH); or "other." With the aid of a computer-assisted morphometer, the following were also measured and calculated: proportion of node occupied by follicles, mean relative follicle size, and mean follicle shape factor . Finally, in 57 cases, the influence of human immunodeficiency (HIV) status on the findings was analyzed . Although individual cases varied widely, cases of DL had significantly more CD3+ (T) cells, higher CD4:CD8 ratios, and fewer CD19+ (B) cells than other categories . Cases of FH,M had significantly lower CD4:CD8 ratios and more CD19+, CD10+, and transferrin receptor positive cells . Cases of FH,M and FH known to be HIV-negative had higher CD4:CD8 ratios than the HIV-positive cases . Peripheral blood CD4:CD8 ratios performed in 38 patients showed a strong correlation with nodal ratios . Morphometric data supported the correlation between follicular hyperplasia and increased proportions of CD19+, CD10+, and transferrin receptor-positive cells . Rare cases had CD5:CD2 or CD3 ratios of greater than 1 or "monoclonal" kappa to lambda ratios . CD4:CD8 ratios varied widely, but aberrant T cell phenotypes were not identified . These studies demonstrate that, although great variation exists, there are certain associations between types of reactive lymphoid hyperplasia and cell suspension immunophenotypic findings.(ABSTRACT TRUNCATED AT 250 WORDS) Chin J Biotechnol, 1990, 6(2), 119 - 23 Plant regeneration from protoplasts of a super Chinese rice (Oryza sativa L.) cultivar--Zhonghua NO.8; Li JX et al.; Green rice plants were regenerated from protoplasts, which were derived from cell suspension of Oryza sativa L . cut . Zhonghua No . 8, a super Chinese rice cultivar with high productivity, good quality and high resistance to both bacterial blight and blast . The embryogenic calli were initiated from mature embroys . It took about 4 months to establish the cell suspenion . The regeneration plants from protoplasts were obtained in 2 months after the isolation of protoplasts. Int Arch Allergy Appl Immunol, 1990, 93(2-3), 205 - 11 Increased expression of high-affinity low-density lipoprotein receptors on human T-blasts; Huber LA et al.; Like all cells, lymphocytes need cholesterol for proper function, a requirement met by a finely tuned homeostasis between intracellular synthesis and uptake from the environment via low-density lipoproteins (LDL) . We used flow cytometry to analyze the receptor activity of resting cells and T blasts incubated/activated in serum-free culture medium, or in medium supplemented with 25-5,000 micrograms/ml LDL . Dioctadecyl-indocarbocyanine has proved to be a useful fluorescent probe for investigating the LDL receptor activity of lymphocytes . The results show the receptor activity of day-3 resting T cells to be reduced more than 50% by 50 microgram LDL/ml, whereas 100-fold higher concentrations are necessary to achieve the same level of reduction in day-3 PHA blasts . The LDL receptor activities of individual blood donors' resting T cells, in vitro cholesterol-deprived resting T cells, and activated T blasts, were compared using two analytical techniques: spectrofluorometric analysis of detergent-solubilized cell suspensions and flow cytometric analysis of single living cells . Receptor affinity was determined by Scatchard analysis of spectrofluorometric binding curves, and by Line-weaver-Burke plots of flow cytometric data . Both methods yielded essentially identical dissociation constants (Kd) for cholesterol-deprived resting T cells and mitogen-activated T blasts, which fell in the expected range for the high-affinity LDL receptor (4.1-8.9 nM) . In addition, spectrofluorometric analysis, but not flow cytometry, permitted quantification of LDL uptake.(ABSTRACT TRUNCATED AT 250 WORDS) Biomed Biochim Acta, 1990, 49(12), 1157 - 64 Liposome mediated in vitro transfection of pancreatic islet cells; Welsh N et al.; The aim of this study was to evaluate the suitability of the liposome technique for transfection of pancreatic islet cells in vitro . For this purpose, fetal islets were isolated and cultured free floating for two days after which they were further cultured, either intact or dispersed into islet cell suspensions, with different DNA-liposome preparations . The DNA-construct used were the control plasmid (pSP65) and the viral oncogene v-src contained in the plasmid pSPRIsrc . A previous study showed that islet cells transfected by means of electroporation with pSPRIsrc displayed an increased thymidine incorporation rate, making this plasmid suitable for further transfection studies . The DNA was associated with the liposomes by means of surface adhesion . The liposomes used were either conventional phosphatidylcholine-containing liposomes, phosphatidylethanolamine/oleic acid containing liposomes (pH-sensitive liposomes) or Lipofectin . After the exposure of islet cells to the DNA-liposome preparations, the transfection efficiency was assessed by determination of the uptake of the DNA-constructs (Southern blot analysis) and expression of the gene construct into an mRNA (Northern blot analysis) . In addition, the impact of the different DNA-liposome preparations on islet DNA replication (thymidine incorporation rates) was determined . It was found that two days after exposure to the DNA-liposomes, the v-src construct was located in islet cell nuclei and that v-src derived transcripts were transiently expressed in the islet cells . The Lipofectin liposomes were more efficient in transfecting islet cells than the pH-sensitive liposomes as assessed by the blotting techniques.(ABSTRACT TRUNCATED AT 250 WORDS) Acta Biol Hung, 1990, 41(1-3), 257 - 65 Continuous induction of unscheduled DNA synthesis by gamma irradiation; Weniger P et al.; The induction of DNA-synthesis in non-S-phase cells is a very sensitive measure of a preceding damage of DNA . Usually, in an in vivo-in vitro test (treatment of an animal, incorporation of H3-thymidine in a cell suspension) the damaging of DNA takes place hours to days before the evaluation . In this case, the time course of the UDS-induction after a single dose of 1 Gy gamma irradiation was observed over a long period of time (21 months) . C57 black mice served as test animals . In an age of about 80 days they were irradiated and the induction of unscheduled DNA synthesis was measured at ten time intervals during the whole life-span of the animals . Although the repair in this gamma radiation damage in DNA is a very quick process--with centrifugation in alkaline sucrose a half-life of some minutes is found--an induction of unscheduled DNA synthesis could be seen at the irradiated animals until the end of their life (640 days) . The reason for this could be permanent disorders in cellular regulation caused by the gamma irradiation. Horm Metab Res Suppl, 1990, 25, 82 - 7 A fluorometric viability assay for single human and rat islets; London NJ et al.; A microfluorometric viability assay for isolated human and rat islets of Langerhans has been developed using the fluorochromes fluorescein diacetate and propidium iodide . Fluorescein diacetate causes live cells to fluoresce green under blue light excitation (490 nm); propidium iodide causes dead cells to fluoresce red under green light excitation (454 nm) . The fluorescence intensity from the live and dead cells within a single islet was selectively measured by photometry using 520 nm and 610 nm barrier filters with blue and green light excitation respectively . All measurements were corrected for background fluorescence . The proportion of dead cells within single human or rat islets measured by microfluorometry was found to correlate highly significantly (r = 0.99, P less than 0.001) with the proportion of dead cells measured by dissociating the same islet into a single cell suspension and counting the actual proportion of dead cells . This assay therefore provides a rapid, accurate and objective measurement of the proportion of dead cells within isolated human and rat islets. Stereotact Funct Neurosurg, 1990, 54-55, 328 - 36 Grafting of embryonic motoneurons into spinal cord and striatum of adult mice; Demierre B et al.; The aim of the present work was to determine whether embryonic motoneurons could survive in the adult CNS and whether they display different survival and growth characteristics in their natural site (the spinal cord) as compared to an ectopic site (the striatal region of the brain) . Specific labelling of embryonic motoneurons was obtained by retrograde transport of a fluorescent tracer (carbocyanine) followed by partial purification of the dissociated motoneurons on a density gradient . This technique offers the advantage that only the motoneurons in the cell suspension used for the grafts contain the fluorescent tracer . This study demonstrates that these motoneurons can survive and differentiate in the white and grey matters, not only in the adult spinal cord, but also in the brain . Furthermore, these motoneurons can migrate approximately 2 mm in the spinal cord and 4 mm in the brain. Haematologia (Budap), 1990, 23(3), 151 - 9 Fibronectin and the adhesive properties of rat lymphocytes obtained from different peripheral lymphoid tissues; Altankov G et al.; A comparative investigation has been carried out on the effect of plasma fibronectin (Fn) on the adhesive properties of normal rat lymphocytes obtained from different lymphoid tissues: blood, spleen, mesenteric and tonsillar lymph nodes . Fn was immobilized on the basis of its ability to bind to gelatin . We established that concentrations of 40-50 micrograms/ml are sufficient for a saturation effect on Fn coating . For spleen cells an adhesion of 55.7 +/- 9.3%, for mesenteric lymph nodes 34.5 +/- 8.7% and for tonsillar cells 33.8 +/- 3.2% was observed . Blood lymphocytes showed the lowest adhesion, 21.3 +/- 4.2% . Compared to the other lymphoid tissues, the spleen cells exhibited a "basal" adherence to surfaces coated with gelatin only: 19.2 +/- 4.1% . T lymphocytes participate to a greater extent in the process, since their number was significantly reduced in cell suspensions after adhesion to both gelatin and gelatin-Fn coated surfaces . The addition of soluble Fn leads to a competitive inhibition of the lymphocyte adhesion to gelatin-Fn coated surfaces . The data demonstrated the important role of Fn for the adhesive interactions of lymphocytes during their functional distribution in the tissues. Pathobiology, 1990, 58(5), 241 - 8 Reactivity of human tissues with monoclonal antibodies to myeloid activation and differentiation antigens . An immunohistochemical study; Koch AE et al.; We have characterized antimyeloid monoclonal antibodies (mAbs) produced to human rheumatoid arthritis (RA) synovial tissue macrophages (MPs) (8D7) and to lipopolysaccharide (LPS)-treated U937 cells (3D8) . The 3D8 antigen is upregulated with LPS stimulation of monocytes/MPs and during monocyte maturation . The 8D7 antigen is upregulated on functionally distinct subpopulations of RA synovial tissue MPs . We used immunohistochemistry to determine the spectrum of reactivity of these unique mAbs on myeloid cell suspensions, monocytes, and mature tissue inflammatory and noninflammatory MPs . The antigens identified by the mAbs were characterized biochemically, by immunoprecipitation of solubilized 125I-labelled antigens from cell surfaces, and immunohistochemically by enzymatic digestion of myeloid cells followed by a cellular ELISA . MAb 3D8, characterized as an anti-CD13 antibody, recognizes a 150-170 kd antigen, has almost exclusive myeloid reactivity, but reacts with Langerhans' cells of the skin and thymus, pointing to shared antigens between these cells and MPs . Unlike 3D8 antigen, 8D7 antigen is strongly expressed in inflammatory states, being present on MPs in granulomata as well as in sarcoid lymph nodes . Both mAbs react with frozen and methanol-Carnoy's fixed, paraffin-embedded tissues and detect antigenic differences among human mononuclear phagocytes present in different anatomical sites and in varying stages of differentiation and activation . These mAbs should prove to be a valuable tool for studying heterogenous populations of myeloid cells. Virchows Arch B Cell Pathol Incl Mol Pathol, 1990, 59(5), 281 - 9 Collagenase of hepatocytes and sinusoidal liver cells in the reversibility of experimental cirrhosis of the liver; Montfort I et al.; In order to explore the cellular source(s) and the behaviour of the collagenolytic activity previously described in rat liver homogenates, in the reversibility of experimental cirrhosis of the liver, enriched suspensions of hepatocytes and of sinusoidal liver cells were obtained by a procedure which employs low EDTA concentrations and no bacterial collagenase . Cell suspensions were prepared from three different groups of animals: 1) normal controls, 2) rats with CCl4-induced cirrhosis of the liver, and 3) rats with swine serum-induced cirrhosis of the liver . Animals were sacrificed in each group upon completion of treatment and also after 3, 6 and 12 months . In each liver wet weight and collagen concentration were determined, and collagenolytic activity of both enriched cell suspensions was measured separately . In addition, histological studies of liver tissue and ultrastructural examination of cell suspensions were performed by standard procedures . Enriched suspensions of both normal hepatocytes and sinusoidal liver cells display Ca2(+)-dependent collagenolytic activities . Both cell suspensions obtained from each of the two types of cirrhotic livers show normal or slightly increased average levels of collagenase activity at the time of treatment discontinuation, when average liver collagen content ranges from 6 to 10-fold over normal, suggesting that the normal collagenase/collagen ratio is disturbed and that collagenolytic activity is deeply decreased in relation to the actual liver collagen load.(ABSTRACT TRUNCATED AT 250 WORDS) Virchows Arch B Cell Pathol Incl Mol Pathol, 1990, 59(5), 263 - 70 Lymphocytes from hepatic inflammatory infiltrate kill rat hepatocytes in primary culture . Comparison with peripheral blood lymphocytes; Ramadori G et al.; In the last few years it has become possible in the liver to isolate lymphocytes from inflammatory infiltrates and to culture them in vitro . Most of the lymphocyte clones obtained are CD 8+ cytotoxic cells, but interactions between these lymphocytes and hepatocytes in primary culture have not been analysed previously . In this study, cloned human T lymphocytes from liver biopsies and from the peripheral blood of patients with chronic hepatitis B or primary biliary cirrhosis, after phenotypical and functional characterization into CD 8+ or CD 4+ cytotoxic lymphocytes, were activated in an antigen-independent fashion by adding either anti CD 3 or anti CD 2/R-3 monoclonal antibodies to the cell suspension . The activated cells were then coincubated with rat hepatocytes in primary culture . The killing capacity of the activated lymphocytes was monitored by light and electron microscopy and by measurement of lactic dehydrogenase (LDH)-release into the culture medium . It was found that cytotoxic CD 8+, but not CD 4+ helper lymphocytes very effectively killed hepatocytes . The killing effect was dependent on the time of cocultivation and on effector-target (E/T) ratio . Total breakdown of the hepatocyte monolayer was achieved after 10-20 h coculture and at an E/T ratio of 10 to 1 . As LDH-release in the culture medium reached about 80% of the total LDH-content, most of the hepatocytes were lysed by activated lymphocytes . Cytotoxic activity of clones obtained from different biopsies was comparable with that of clones from peripheral blood . Hepatocytes in primary culture seem to be very sensitive to the killing capacity of activated cytotoxic lymphocytes. Exp Brain Res, 1990, 81(2), 433 - 7 Seizure suppression in kindling epilepsy by intrahippocampal locus coeruleus grafts: evidence for an alpha-2-adrenoreceptor mediated mechanism; Bengzon J et al.; Intrahippocampal cell suspension grafts, prepared from the locus coeruleus region of rat fetuses, have previously been shown to retard seizure development in rats made hypersensitive to hippocampal kindling by a lesion of the forebrain noradrenergic system . The objective of the present study was to provide evidence that the seizure-suppressant effect elicited by the grafts is mediated via noradrenergic mechanisms . Two groups of rats received 6-hydroxydopamine in the lateral ventricle and then bilateral intrahippocampal locus coeruleus grafts . After 3 months, the grafted animals and a group of normal rats were subjected to hippocampal kindling . One group of grafted animals and the normal rats were injected intraperitoneally with the alpha-2 adrenergic receptor blocker idazoxan before each kindling stimulation . The other grafted rats received vehicle injections . The development of seizures was significantly faster in the grafted and normal rats that had been given idazoxan than in the grafted rats that had not been subjected to alpha-2 receptor blockade . Our data suggest that the seizure-suppressant action exerted by grafts of fetal locus coeruleus in hippocampal kindling is mediated via noradrenergic mechanisms, most likely via activation of postsynaptic alpha-2 adrenoreceptors. Vrach Delo, 1990 Jan, (1), 90 - 3 {Cryopreservation of a suspension of fetal liver cells for clinical use}; Lavrik SS et al.; The authors evaluate a method of procuring and preserving fetal liver cells in a medium containing low-molecular polyvynilpyrrolidon, glucose, lactose, sodium phosphate and sodiumhydrocitrate not requiring washing off . The biomaterial is stored at a temperature of -196 degrees C . The efficiency of this method of cryopreservation of fetal liver cell suspensions was confirmed by high morphological intactness, functional activity and proliferative capacity . Patients with hypoplastic anemia subjected to transfusion of cryopreserved fetal liver cells showed a normalization of bone marrow hemopoiesis, remissions. Exp Brain Res, 1990, 79(1), 3 - 17 Intrastriatal implants of mesencephalic cell suspensions in weaver mutant mice: ultrastructural relationships of dopaminergic dendrites and axons issued from the graft; Triarhou LC et al.; Dissociated cell suspensions were prepared from the ventral midbrain of normal mouse foetuses and stereotaxically implanted into the neostriatum of 2-3 months old homozygous weaver mutant mice, which are severely deficient in dopamine . In tests of amphetamine-induced turning behaviour 60 days after grafting, recipient animals displayed a rotational bias opposite to the grafted side . Prior to perfusion, which was carried out at 80 days after transplantation surgery, the grafted striata of the weaver recipients were deprived of their intrinsic mesostriatal dopamine input by local injections of 6-hydroxydopamine into the ipsilateral substantia nigra in order to selectively study the innervation derived from the graft . Grafts were found to contain an estimated 100-700 tyrosine hydroxylase immunoreactive neurones . An ultrastructural analysis demonstrated that both axons and dendrites immunoreactive for tyrosine hydroxylase extended from the graft into the recipient striatum . In the host striatum proximal to the graft (i.e . at a distance of 0.0-0.5 mm from the graft) the proportion of dendrites to axons was about 1:2, whereas distal to the graft (i.e . at a distance of 0.5-1.0 mm) it was 1:20 . Graft-derived tyrosine hydroxylase immunoreactive axons were primarily found in apposition with unlabelled dendrites or spines of the recipient striatum (greater than 90%) . Graft-derived dopaminergic dendrites received synaptic input from unlabelled axon terminals and were opposed to the unlabelled somata of striatal neurones in a few instances . In conclusion, this study shows that mesencephalic cell suspensions survive in the weaver striatum and provide a functional dopamine innervation which comprises both axonal and dendritic processes. Cancer Immunol Immunother, 1990, 31(1), 19 - 27 Expression of the adhesion molecule ICAM-1 and major histocompatibility complex class I antigens on human tumor cells is required for their interaction with autologous lymphocytes in vitro; Vanky F et al.; In a group of 30 human tumors, comprising 12 lung, 14 ovarian, 2 breast carcinomas, 1 hypernephroma and 1 mid-gut carcinoid, the expression of major histocompatibility complex (MHC) class I molecules and the intercellular adhesion molecule 1 (CAM-1, CD54) was found to vary independently . Some tumors expressed both or neither of these molecules . Among 9/13 ICAM-1+ tumors, in which greater than 50% cells reacted with the anti-ICAM-1 monoclonal antibody (mAb) (LB-2), the class I antigen was also detected on greater than 50% of the cells . Only 2 ICAM-1+ tumors were class-I- . In 5/17 cases the tumors were MHC-class-I+ and ICAM-1- . Lymphocytes collected from the blood or from the tumor site were assayed for recognition on the tumor cells in the auto-tumor cytotoxicity test and in mixed lymphocyte tumor cell culture (MLTC) . Positive results were obtained only with the MHC-class-I+/ICAM-1+ tumors . In vitro treatment of the tumor cell suspensions with interferon gamma and tumor necrosis factor alpha (TNF alpha) induced or enhanced the ICAM-1 and/or class I antigen expression in 8/12 cases . Of the tumor samples treatged, 8/9 acquired stimulatory capacity and 3/10 became susceptible to lysis by the lymphocytes . In 6/6 MLTC performed with the cytokine-treated tumor cells, cytotoxicity against the autologous tumor was generated . Three of these MLTC lymphocytes also lysed the untreated targets . mAb directed to class I antigens or to ICAM-1 inhibited both the stimulation by and the lysis of tumor cells when confronted with fresh lymphocytes . The cytotoxicity generated in the MLTC was also inhibited . If, however, the cytotoxic function was induced in MTLC containing interleukin-2 (5 U/ml), inhibition was obtained only by pretreatment of the targets with mAb against ICAM-1 . The results show thus (a) that the lymphocytes react in vitro with tumor cells only if these express both MHC class I molecules and ICAM-1; (b) that expression of these molecules can be induced by interferon alpha and TNF alpha; (c) that cytotoxic effectors generated in the MLTC with cytokine-treated tumors can also act on the untreated tumor cells . The requirement of the two surface moieties for the interaction with lymphocytes was also substantiated by blockade with relevant mAb. Biochem J, 1990 Jan 1, 265(1), 39 - 44 Identification and characterization of somatostatin receptors in neonatal rat long bones; Bruns C et al.; Somatostatin (somatotropin release inhibiting factor; SRIF) has widespread functions as a modulator of neural activity as well as of endocrine and exocrine secretion . In the present paper, the binding characteristics of somatostatin receptors have been investigated in rat long bones using the stable analogue, 125I-SDZ 204-090, as a ligand . Binding studies revealed the presence of a single class of high-affinity binding sites for 125I-SDZ 204-090 on cells prepared from neonatal rat long bones with an equilibrium dissociation constant (KD) of 70.1 +/- 8.2 pM (n = 3) . An excellent correlation was found between the ability of various somatostatin analogues to inhibit growth hormone in pituitary cells and to displace the binding of 125I-SDZ 204-090 to the bone cell preparation, indicating that the receptors are very similar, if not identical . The localization of the somatostatin-binding sites was examined by autoradiography after labelling in vitro and in vivo . The binding sites were shown by both procedures to be selectively localized to the metaphysis of rat long bones . The labelling experiments in vivo indicate that these receptors can be reached in the living animal by circulating somatostatin analogues . In addition, the analogue SMS 201-995 inhibited the forskolin-stimulated adenylate cyclase activity in bone cell suspensions . These results suggest that somatostatin could be an important regulatory factor in bone metabolism. Int J Radiat Biol, 1990 Jan, 57(1), 103 - 15 Bone marrow from Balb/c mice radiocontaminated with 241Am in utero shows a deficient in vitro haemopoiesis; Van Den Heuvel RL; Radiation damage from 241Am to bone marrow cells was manifest in long-term bone marrow cultures (LTC) from offspring of mice radiocontaminated at the 14th day of gestation (119, 479, 803, 1754 kBq 241Am/kg) . Offspring were reared by their own contaminated mother for 3 weeks postnatal . LTC from these offspring were less able to support in vitro CFC proliferation than control LTC from non-contaminated offspring . This radiation damage persisted 71 weeks after radiocontamination in utero . Using this in vitro culture system, damage was observed at lower doses if 241Am contamination occurred at foetal than at adult ages . Radiation damage was observed only using LTC, while the haemopoietic stem cell concentration (CFU-S, in vitro CFC) and the stromal stem cell concentration (CFU-F) from marrow in situ were not impaired after 241Am radiocontamination in utero . After culturing LTC in 25 per cent FCS and recharging the stromal adherent layer with bone marrow cell suspensions originating either from control offspring or from offspring contaminated with 241Am in utero, some evidence was found that the proliferation capacity of the haemopoietic cells was diminished . However, the nature of effects on the stromal elements is currently somewhat equivocal . Following in utero contamination the stromal adherent cells appeared to support better the production of in vitro CFC. J Immunol, 1990 Jan 1, 144(1), 7 - 15 Antibodies against the CD44 p80, lymphocyte homing receptor molecule augment human peripheral blood T cell activation; Denning SM et al.; The CD44 inhibitor Lutheran {In(Lu)}-related p80 molecule has recently been shown to be identical to the Hermes-1 lymphocyte homing receptor and to the human Pgp-1 molecule . We have determined the effect of addition of CD44 antibodies to in vitro activation assays of PBMC . CD44 antibodies did not induce PBMC proliferation alone, but markedly enhanced PBMC proliferation induced by a mitogenic CD2 antibody pair or by CD3 antibody . CD44 antibody addition had no effect upon PBMC activation induced by PHA or tetanus toxoid . CD44 antibody enhancement of CD2 antibody-induced T cell activation was specific for mature T cells as thymocytes could not be activated in the presence of combinations of CD2 and CD44 antibodies . CD44 antibody enhancement of CD2-mediated T cell triggering occurred if CD44 antibody was placed either on monocytes or on T cells . In experiments with purified monocyte and T cell suspensions, CD44 antibodies A3D8 and A1G3 augmented CD2-mediated T cell activation by three mechanisms . First, CD44 antibody binding to monocytes induced monocyte IL-1 release, second, CD44 antibodies enhanced the adhesion of T cells and monocytes in CD2 antibody-stimulated cultures, and third, CD44 antibodies augmented T cell IL-2 production in response to CD2 antibodies . Thus, ligand binding to CD44 molecules on T cells and monocytes may regulate numerous events on both cell types that are important for T cell activation . Given that recent data suggest that the CD44 molecule may bind to specific ligands on endothelial cells (vascular addressin) and within the extracellular matrix (collagen, fibronectin), these data raise the possibility that binding of T cells to endothelial cells or extracellular matrix proteins may induce or up-regulate T cell activation in inflammatory sites. J Clin Lab Immunol, 1990 Jan, 31(1), 27 - 31 Inhibition of in vitro plaque formation by large granular lymphocyte leukemia cells from F344 rats; Stromberg PC et al.; The effect of large granular lymphocyte leukemia on B lymphocyte function was studied by determining the number of plaques formed in an in vitro hemolytic plaque assay . Leukemia cells inhibited plaque formation by normal splenic lymphocytes in a logarithmic, dose-dependent manner . At the highest leukemia cell concentrations, spleen cell suspensions made 50% fewer plaques . Plaque forming responses were very sensitive to duration of preincubation time in all assays . The number of plaques formed decreased markedly if incubated 2 hr before the assay was performed . Incubation of the cells at 56 degrees C for 8 min did not alter the inhibitory activity but pretreatment with 0.01% trypsin did . Supernatant fluids from leukemia cell suspensions did not inhibit plaque formation . These data suggest that diffuse infiltration of lymphoid tissues by leukemia cells may interfere with some normal lymphocyte functions . Although leukemia cells inhibited splenic B lymphocyte function, leukemic rats did not have hypogammaglobulinemia. Symp Soc Exp Biol, 1990, 44, 193 - 205 Phosphoinositides and plant growth substance action; Hanke DE et al.; Synergism between 0.5 mM inositol and cytokinin in the stimulation of plant tissue culture growth suggests a role for inositol in the mediation of cytokinin action . Investigation of the effects of cytokinin on the pattern of labelling of lipids from radioactive precursors by cytokinin-responsive cells of a cytokinin-dependent soybean cell suspension culture did not detect any reproducible link between cytokinin action and lipid labelling after 10 min . Evidence for links between auxin action and phosphoinositide metabolism in other systems would benefit from confirmation of long term repeatability and more rigorous chemical characterization of the compounds involved . For plant growth substance action to be mediated by release of inositol trisphosphate from phosphatidylinositol bisphosphate, activation of phospholipase C, possibly requiring potentiation by a GTP-binding protein, would be expected . Reports of G-protein effects on plant phospholipase C activity are conflicting . There is evidence for G-protein stimulation of activity from the results of assays using endogenous substrates, although the products released have not been fully characterized . Other results, from assays using exogenous substrates, have shown no effect of GTP analogues on the enzymic breakdown of phosphatidylinositol bisphosphate . Using endogenously labelled membranes from soybean cells, we were unable to detect effects attributable to G-protein potentiation . None of a range of growth substances at physiologically active concentrations proved able to alter detectably the lack of response of polyphosphoinositidase activity to GTP-analogues . The activity was, however, strongly stimulated by Ca2+ at micromolar levels, a characteristic widely reported . In consequence, the possibility that transient increases in the labelling of inositol phosphate fractions may be a result of increases in cytosolic Ca2+ levels needs to be addressed . If there is a role for inositol in soybean cell activation by cytokinin, we have no evidence that it involves polyphosphoinositide cleavage . That there is a special role for inositol in the mitotic cycle of soybean cells and in addition to the maintenance of viability is shown by the results of experiments in which endogenous inositol synthesis was inhibited . Further research aims to identify the inositol-requiring steps and their relationship, if any, with the auxin-requiring and cytokinin-requiring steps in the mitotic cycle of cultured soybean cells. Neuroscience, 1990, 37(2), 353 - 66 Dissimilar responses of adult thalamic monoaminergic and somatosensory afferent fibers to implantation of thalamic fetal cells; Nothias F et al.; It is generally accepted that transplanted fetal neurons can, after several weeks to months, establish connections with the host CNS . Host afferent systems seem, however, to show different types of responses to the presence of grafted fetal neurons . The present study is a preliminary step to identify mechanisms involved in the reactions of adult axons to transplanted fetal neurons . The right ventrobasal thalamus of adult rats was depleted of neurons by in-situ injection of kainic acid and cell suspensions from homotopic thalamic embryonic primordia which were injected into the lesioned area . After various post-implantation delays, ranging from five to 30 days, two types of experiments were performed: (i) noradrenaline and serotonin immunohistochemistry with specific antibodies on alternate sections; and (ii) anterograde tracing using wheat germ agglutinin conjugated to horseradish peroxidase from the dorsal column nuclei and the principal sensory trigeminal nucleus . Five days after transplantation, host monoaminergic fibers (either noradrenergic or serotoninergic) had already grown into the transplants . Ingrowing fibers were thin and poorly varicose, exhibiting endings morphologically similar to the growth cones observed during axogenesis . Seven days after grafting, growth cones were no longer visible and monoaminergic fibers exhibited either normal-sized or very large varicosities . Large varicosities progressively decreased in number and, after three weeks, the fibers displayed a normal adult morphology, forming a dense network all over the transplants . In contrast, host somatosensory afferents, labeled by anterograde transport of wheat germ agglutinin conjugated to horseradish peroxidase, did not grow into the transplants . Intermingling of somatosensory afferents and transplanted cells was observed only after 10 days, when grafted neurons extended outside the original transplantation site into the neuron-depleted area containing the somatosensory afferents . The present results demonstrate that adult monoaminergic and somatosensory afferents, when deprived of their usual target, do not react in a similar way to the addition of fetal neurons . It is proposed that adult monaminergic fibers have the ability to regain morphological (and probably functional) immature forms which were considered to be restricted to the period of axogenesis or to lesion-induced regeneration . In contrast, fetal transplants do not seem to induce, by themselves, a similar alteration of genetic expression in adult somatosensory neurons . It has been proposed that "diffuse" and "point-to-point" axonal systems may be differentiated in the CNS on anatomical bases . The present results add to the identification of two different systems by demonstrating that, in the thalamus, they present dissimilar responses to the implantation of fetal cells. Verh Dtsch Ges Pathol, 1990, 74, 432 - 6 {Induction of primitive neuroectodermal tumors by oncogene complementation}; von Deimling A et al.; Primitive neuroectodermal tumors (PNET) represent a family of undifferentiated neural neoplasms which predominantly occur in children . PNETs are likely to originate from precursor cells and exhibit a marked potential for neuronal, glial and ependymal differentiation . A prominent example is the medulloblastoma of the cerebellum . In a model system using a novel transgenic CNS transplantation model, we have introduced a combination of ras and myc oncogenes into cell suspensions from fetal forebrain (E14) and postnatal cerebellum (P2) of the rat . Oncogene transfer into fetal forebrain grafts resulted in a high incidence of anaplastic neural tumors predominantly derived from glial precursors . Cell lines established from these neoplasms expressed high levels of both oncogenes . In a second experiment, the ras/myc vector was introduced into cell suspensions from neonatal cerebellum . The transformed cells were cultured for 3 weeks . Following stereotaxic transplantation, tumors of a similar morphology were observed . However, one animal developed a neoplasm with features of a cerebellar medulloblastoma . A cell line which exhibits a marked capacity for neurite extension and synaptogenesis was established from this tumor . Since these cells neither express ras nor myc, an insertion mutagenesis event appears to be responsible . Experiments to characterize this mutation are in progress . Our results indicate a potent transforming effect of ras and myc on neural precursor cells in vivo. Arkh Patol, 1990, 52(11), 62 - 3 {Method of bronchoalveolar lavage for performing cytochemical examination in chronic nonspecific pulmonary inflammation}; Grobova OM et al.; The method suggested includes the use of homogenizer terrilytin for removing excessive mucus from broncho-alveolar lavage (BAL) . Cytochemical investigation of BAL neutrophils is reasonable to be performed if the viability of the cell suspension is not lower than 60% . Cytospectrophotometric examination is recommended for an unbiased evaluation of the neutrophil functional state. Stereotact Funct Neurosurg, 1990, 54-55, 364 - 7 Effects of interstitial edema on brain cell transplantation; Miyazaki S et al.; Fetal raphe cells were transplanted into the anterior part of the corpus callosum of serotonin denervated hydrocephalic rats using a cell suspension method . Hydrocephalus was induced by intracisternal injection of kaolin . The survival of the transplanted cells and fiber outgrowth were evaluated according to the level of serotonin and its metabolite, hydroxyindoleacetic acid, using high-performance liquid chromatography with electrochemical detection in the anterior and posterior parts of the corpus callosum 1-2, 5-6, and 7-8 weeks after transplantation . The results suggest favorable effects of interstitial edema associated with hydrocephalus on the survival of transplanted raphe cells and fiber outgrowth. Lab Delo, 1990, (8), 31 - 2 {Determination of the erythrocyte count using a microcolorimeter}; Storozhuk PG et al.; Red cell count is estimated with the use of a MKMF-1 microcolorimeter . 10 ml of Gower's solution are used; 0.02 ml of blood are added to this solution . Photometry of red cell suspension is carried out in a cuvette with a 0.5 cm optic route at lambda 610 nm . Microampermeter data are read from the optic density D scale . The D value is substituted into the formula: red cell count (RC) = K.D.10(6), where K coefficient is found experimentally by measuring the D and by estimating red cell count in microscopy, and is equal to 33.3 . This formula permits estimating red cell count in 1 microliter of blood. Lab Delo, 1990, (8), 27 - 9 {Determination of the rate of utilization of glucose in a suspension of erythrocytes}; Kunitsyn VG et al.; The authors suggest a new method for measuring the rate of glucose utilization in a cell (red cell) suspension, making use of interferometry . This method allows an easy examination of glucose utilization kinetics in time with but 1000 cells . Thermostatic control of the sample in the cuvette of the device permits detecting the temperature effects on glucose metabolism kinetics and thus define the temperature optimal for the process . A correlation was revealed between the point of structure phase transition in the red cell membrane and the optimal temperature for glucose utilization within the range of physiologic temperatures . This method may be useful in medical, biochemical, physiologic, and hematologic studies . The relative error of the technique is 3 percent. Prog Brain Res, 1990, 82, 95 - 101 Angiogenesis and the blood-brain barrier in solid and dissociated cell grafts within the CNS; Broadwell RD et al.; Available evidence suggests that blood vessels indigenous to solid CNS and peripheral tissues grafted to the brain are sustained and maintain the morphological and permeability characteristics they manifest in normal life . Furthermore, these vessels of graft origin anastomose (albeit not rapidly) with vessels of the surrounding host tissue predominantly at the host-graft interface and less so, or not at all, within the graft itself . For these reasons, blood-brain and brain-blood barriers, evident in the late fetal and neonatal CNS, can be expected to exist within CNS grafts placed intracerebrally or extracerebrally, providing the graft remains viable . Peripheral neural and non-neural tissues not possessing cellular barriers to circulating macromolecules do not acquire such barriers subsequent to their transplantation within the CNS . The absence of a blood-brain barrier in the adrenal gland grafted intracerebrally may be relevant for the treatment of Parkinson's disease with blood-borne therapeutics . Compared to solid tissue grafts, cell suspension grafts have the potential of becoming vascularized rapidly . That cell suspensions of neurons and of glia are supplied with BBB vessels of host origin and that the permeability characteristics of host BBB vessels are altered by a tumor cell suspension reaffirm the belief that the type of transplanted cell/tissue indeed determines the permeability characteristics of the blood vessels supplying it . The suspected immunologic privilege of the CNS is not absolute . Eventual host rejection of allografts placed within the third ventricle may be a dual consequence of the absence of a BBB at the level of the host median eminence and involvement of the minor histocompatibility complex. Lab Delo, 1990, (11), 17 - 9 {pH-metric study of ion equilibrium in erythrocyte suspensions exposed to heat}; Vinogradov AP; A method of differentiated measurements of pH is suggested to be used in study of the time course of ionic equilibrium in red cell suspensions exposed to heating . The described method yields more accurate and complete data on the type and distribution of red cells as regards their membrane characteristics. Arch Dermatol Res, 1990, 282(2), 126 - 30 Flow cytometric quantification of human epidermal cells expressing keratin 16 in vivo after standardized trauma; de Mare S et al.; The intermediate filament protein keratin 16 is expressed in hyperproliferative epidermis . The present study aims to clarify the relationship between the expression of this keratin type, hyperproliferation (percentage of cells in SG2M phases), and keratinization (keratin 10 expression) . These three parameters were quantified in biopsy material taken at different time intervals following sellotape stripping--this being a dynamic in vivo model for the induction of hyperproliferation . From the biopsy specimens cell suspensions were prepared, labeled with antibodies KS8.12 (specially directed against keratin 16) and RKSE60 (directed against keratin 10), and analyzed using flow cytometry . Percentages of cells in SG2M phases were assessed by measuring the relative DNA content after propidium iodide staining . Keratin 16 expression in the suprabasal layer anticipated epidermal proliferation, suggesting a role of the suprabasal compartment in the induction of epidermal growth . Keratin 10 expression decreased about 1 day after the onset of keratin 16 expression, indicating that these processes do not depend directly upon each other. Urol Res, 1990, 18(2), 131 - 6 Chemosensitivity testing of primary human renal cell carcinoma by a tetrazolium based microculture assay (MTT); Mickisch G et al.; MTT staining procedures have been used in chemosensitivity testing of established cell lines of human and other sources as well as of human leukaemias, but only limited information on its application in primary solid human tumors is presently available . We have evaluated MTT staining in primary human Renal Cell Carcinomas (RCCs), studied various factors interfering with the optimal use, and finally applied it in subsequent chemosensitivity testing . The method depends on the conversion of a water-soluble tetrazolium salt (MTT) to a purple colored formazan precipitate, a reaction effected by enzymes active only in living cells . Single cell suspensions of RCCs were obtained either by enzymatic dispersion or by mechanical dissagregation, filtered through gauze, and purified by Ficoll density centrifugation . Tests were carried out in 96-well microculture plates . 10(4) viable tumor cells per well at 4 h incubation time with 20 micrograms MTT/100 microliters total medium volume yielded best results . Formazan crystals were dissolved with DMSO, and the plates were immediately measured on a microculture plate reader at 540 nm . Under these criteria, linearity of the system could be demonstrated . For chemosensitivity testing, cells were continuously exposed to a number of drugs prior to the MTT staining procedure . Reproducibility of results was assessed and confirmed by culturing RCCs in flasks additionally, resubmitting them after 1, 2, and 4 weeks to the MTT assay . We conclude that the semiautomated MTT assay offers a valid, rapid, reliable and simple method to determine the degree of chemoresistance in primary human RCCs. Biodegradation, 1990, 1(4), 253 - 61 Reductive dechlorination of 1,2-dichloroethane and chloroethane by cell suspensions of methanogenic bacteria; Holliger C et al.; Concentrated cell suspensions of methanogenic bacteria reductively dechlorinated 1,2-dichloroethane via two reaction-mechanisms: a dihalo-elimination yielding ethylene and two hydrogenolysis reactions yielding chloroethane and ethane, consecutively . The transformation of chloroethane to ethane was inhibited by 1,2-dichloroethane . Stimulation of methanogenesis caused an increase in the amount of dechlorination products formed, whereas the opposite was found when methane formation was inhibited . Cells of Methanosarcina barkeri grown on H2/CO2 converted 1,2-dichloroethane and chloroethane at higher rates than acetate or methanol grown cells. Bioprocess Technol, 1990, 10, 251 - 70 Suspension culture of mammalian cells; Birch JR et al.; Mammalian cell suspension culture systems are being used increasingly in the biotechnology industry . This is due to their many advantages including simplicity and homogeneity of culture . Suspension systems are very adaptable (e.g., for microcarrier, microencapsulation, or other methods of culture) . Their engineering is thoroughly understood and standardized at large scale, and automation and cleaning procedures are well established . Suspension systems offer the possibility of quick implementation of production protocols due to their ability to be scaled easily once the basic culture parameters are understood . The only main disadvantage of the suspension culture systems to date is their inapplicability for the production of human vaccines from either primary cell lines or from normal human diploid cell lines (Hayflick et al., 1987 and references therein) . One of the great advantages of suspension culture is the opportunity it provides to study interactions of metabolic and production phenomena in chemostat or turbidostat steady-state systems . Furthermore, in suspension culture systems from which cell number and cell mass measurements are easy to obtain, rigorous and quantitative estimations of the effects of growth conditions or perturbations of metabolic homeostasis can be made . Such studies can speed up the development of optimal processes . With our increasing understanding of factors influencing expression in mammalian cells (Cohen and Levinson, 1988; Santoro et al., 1988) and the direct application of new methods in suspension culture (Rhodes and Birch, 1988), its usefulness and importance is likely to increase in the future . In this chapter, we have described some of the potential uses of the various suspension culture systems and have covered most of the established technology and literature . Due to the rapid developments and needs in the biotechnology industry and the versatility of suspension culture systems, it is probable that many more variations on this theme will evolve in the near future at both the pilot and production scales. Phytochemistry, 1990, 29(7), 2149 - 52 Biotransformation of 18 beta-glycyrrhetinic acid by cell suspension cultures of Glycyrrhiza glabra; Hayashi H et al.; Two biotransformation products formed from 18 beta-glycyrrhetinic acid by cell suspension cultures of Glycyrrhiza glabra were isolated and their structures determined by chemical and spectral data as 3-O-{alhpa-L-arabinopyranosyl-(1----2)-beta-D-Glucuronopy ranosyl}-24- hydroxy-18 beta-glycyrrhetinic acid and 30-O-beta-D-glycopyrano-syl-18 beta-glycyrrhetinic acid . The formation of glycyrrhizin, the main triterpene glucuronide of the licorice root, was not detected among the biotransformation products . This is the first report of the glucuronylation of an exogenous triterpene in plant cell cultures. Phytochemistry, 1990, 29(4), 1131 - 5 Induction of two prenyltransferases for the accumulation of coumarin phytoalexins in elicitor-treated Ammi majus cell suspension cultures; Hamerski D et al.; Two dimethylallyl diphosphate:umbelliferone dimethylallyltransferase (prenyltransferase) activities, catalysing the 6-prenylation and the 7-O-prenylation, respectively, of umbelliferone in the course of phytoalexin synthesis, increased in Ammi majus cell suspension cultures in response to elicitor treatment . Both enzyme activities were dependent on Mg2+ or Mn2+ with significant preference for Mg2+ in the 6-prenylation reaction . Whereas dark-grown cells did not contain these activities, both prenyltransferase activities were induced rapidly by the addition of elicitor reaching a first maximum after 10-14 hr and a second maximum beyond 30 hr . Other coumarin specific, elicitor-induced enzyme activities of A . majus cells, in contrast, showed only one maximum of activity within the 50 hr experimental period, while the pattern of induction of phenylalanine ammonia-lyase activity resembled that of the prenyltransferases with maxima at ca 8 hr and 20-30 hr . Preliminary data suggest that the apparent biphasic induction of these enzyme activities is due to post-translational enzyme modifications. Tumori, 1989 Dec 31, 75(6), 542 - 6 Detection of the 170 kDa P-glycoprotein in neoplastic and normal tissues; Ronchi E et al.; A membrane purification procedure and an immunoblotting assay have been designed to allow screening of human solid tumors for overexpression of the GP170 glycoprotein without employing a disaggregation method to obtain cell suspensions . The electrophoresed membrane proteins were probed, after Western Blotting, with the C219 monoclonal antibody and iodinated Protein A . The labeling intensity of the bands on the autoradioimmunoblots were quantified by densitometry . To test for the presence of GP170, we used membranes from the UV 2237 fibrosarcoma line and its adriamycin-resistant variant ADMR, grown in vitro or as solid tumor in mice . Membranes of human normal and tumor tissues obtained from previously untreated patients were also tested . An immunoreaction was observed in the adriamycin-resistant UV 2237 lines grown in vitro or in vivo . Quantitatively, the binding of the resistant cell line grown in vitro was higher than that observed in cells grown in mice . Bands in the GP 170 region were observed in 4/7 normal and in 7/7 tumor colon tissues and in the normal medulla from 2 patients with cancer of the renal cortex . No reaction could be found in samples from normal tissue, primary tumor or nodal metastasis from 7 patients with breast cancer. Tumori, 1989 Dec 31, 75(6), 570 - 5 In vivo and in vitro growth of SCLC cells derived from biopsies; Giani S et al.; In order to increase the availability of SCLC cells derived from biopsies, in vivo and in vitro growth methods were investigated . The cells grown in both conditions were periodically monitored for reactivity with 2 monoclonal antibodies (MAbs): MLuC1 directed against SCLC cells and IM1 which recognizes the class II antigen on activated lymphocytes and macrophages . About 50% of the 28 analyzed SCLC specimens were found to proliferate in one or both systems . The in vitro-grown cells exhibited the same heterogeneity found in the original cell suspensions and moreover, in some cases only normal cells were recovered after several in vitro passages . From the subcutaneous transplanted tumors a large number of MLuC1-positive tumor cells could easily be recovered, thus indicating the validity of the in vivo methodology . The MBr1 MAb, directed against an epithelial antigen, was found to react with about 50% of the 26 tested tumors, mainly those which demonstrated in vivo and/or in vitro growth capacity . These data suggest that only some tumors, presumably with peculiar biological characteristics, can efficiently grow in these artificial systems. Biochim Biophys Acta, 1989 Dec 28, 987(2), 239 - 42 Net efflux of chloride from cell suspensions measured with a K+ electrode; Rothstein A et al.; Under appropriate conditions (presence of cation ionophores) net KCl efflux measured with a K+ electrode can be used to estimate conductive Cl- fluxes, a sensitive procedure that allows continuous recording . The procedure was tested in human red cells by demonstrating effects of ionophores and of an anion transport inhibitor, and in dissociated MDCK cells by demonstration of cAMP and volume-activated Cl- fluxes. Biochemistry, 1989 Dec 26, 28(26), 10061 - 5 Coenzyme F430 as a possible catalyst for the reductive dehalogenation of chlorinated C1 hydrocarbons in methanogenic bacteria; Krone UE et al.; Corrinoids, such as aquocobalamin, methylcobalamin, and (cyanoaquo)cobinamide, catalyze the reductive dehalogenation of CCl4 with titanium(III) citrate as the electron donor {Krone et al . (1989) Biochemistry 28, 4908-4914} . We report here that this reaction is also effectively mediated by the nickel-containing porphinoid, coenzyme F430, found in methanogenic bacteria .