Am J Physiol, 1991 Oct, 261(4 Pt 2), H1329 - 34 Simultaneous measurement of contraction and oxygen consumption in cardiac myocytes; Rose H et al.; A setup has been developed that simultaneously measures the mechanics and the energetics of electrically induced contractions at physiological frequencies of isolated cardiac myocytes . The core of the setup is a self-manufactured stimulation chamber in which most of the myocytes are in suspension while some are attached to a plastic cover slip prepared from culture Petri dishes . The analysis of the contractile behavior of the attached myocytes is based on an image-processing system with digitized frames of a charge-coupled device camera . Thirty-six frames illuminated by a stroboscope are taken at increasing time intervals between stimulus and flash (snap), allowing one to resolve the contraction cycle with a very high time resolution (down to 1 ms) . The number of pixels that differ between each of these frames and a "reference" frame of the cells in the relaxed state (slack cell length) are used to quantify the contractions . An oxygen electrode in the chamber registers the drop of oxygen tension resulting from the consumption by the myocytes, which exhibit a strictly aerobic metabolism . The resulting data are also stored and analyzed in an IBM-AT-compatible computer. Infect Immun, 1991 Oct, 59(10), 3387 - 92 A role for gamma interferon, tumor necrosis factors, and soluble T-cell receptors in the depressed blastogenic response of spleen cells of Mycobacterium lepraemurium-infected mice; Richard L et al.; Spleen cells of Mycobacterium lepraemurium-infected mice were cultured on petri dishes coated with mycobacterial antigens, and antigen-reactive cells were isolated . Upon incubation in mitogen- or antigen-free culture medium, these cells released mediators capable of depressing the in vitro proliferative response of normal splenocytes to specific antigen and to concanavalin A and lipopolysaccharide . One of these mediators was identified with gamma interferon (IFN-gamma), mainly on the basis that treatment of supernatants with monoclonal anti-IFN-gamma antibodies markedly reduced the suppressive activity contained therein . Detectable levels of tumor necrosis factor alpha (TNF-alpha) and TNF-beta were present in spleen cell culture supernatants of infected mice . Moreover, low doses of recombinant TNF-alpha and TNF-beta were found to potentiate the suppressive activity of exogenous IFN-gamma . Soluble T-cell receptors beta were also detected in the culture supernatants . The elimination of these molecules with monoclonal anti-T-cell receptor beta (F23.1) antibodies immobilized on a plastic surface partially reversed the depression of the response to mycobacterial antigen but did not affect the response to mitogens . These results revealed the complex nature of suppressor mediators that are produced by mycobacterial antigen-reactive cells and that regulate the in vitro proliferative response. J Neurol Sci, 1991 Oct, 105(2), 211 - 6 Enhanced proteolytic activities in cultured fibroblasts of Alzheimer patients are revealed by peculiar transketolase alterations; Paoletti F et al.; Characteristic alterations of transketolase (TK) in extracts from cultured Alzheimer fibroblasts have previously been reported (Paoletti et al . (1990) Biochem . Biophys . Res . Commun., 172: 396-401) . These abnormalities, encountered in 9 out of 13 Alzheimer patients, were revealed following isoelectric focusing and consisted of enzyme forms having unusually high alkaline pI values (alkaline bands) . The present work has shown that immunologically detected alkaline bands were progressively expressed when Alzheimer fibroblasts were incubated for three weeks without medium changes . Full expression of the altered enzyme pattern was not linked to relative cell density in the petri dish; rather, it appeared to be dependent directly on the time elapsed since cell confluence was reached . Alkaline bands could artificially be induced also in both crude and pure TK preparations from normal cells by a treatment with commercial proteases, particularly chymotrypsin . Moreover, specific inhibitors of endogenous cysteine-proteases were capable of abolishing TK alkaline bands in Alzheimer fibroblasts thus turning a pathological into a normal enzyme pattern . Results obtained suggest that Alzheimer fibroblasts contain enhanced Ca(2+)-independent cysteine-proteolytic activities as compared to normal and other pathological cells . These enzymes, exhibiting chymotrypsin-like activity, might exert their degradative effects at the time of cell extraction using TK and probably other cell components as potential substrates . However, peculiar TK abnormalities represent so far an useful biochemical marker detectable in fibroblasts of living Alzheimer patients and closely associated to this neurological disorder. Eur Respir J, 1991 Oct, 4(9), 1066 - 75 Toxic effects of oxygen on cultured alveolar epithelial cells, lung fibroblasts and alveolar macrophages; Housset B et al.; Exposure to hyperoxia results in endothelial necrosis followed by type II cell proliferation . This suggests that type II cells are resistant to hyperoxia . Oxygen-induced lung injury may result from an overproduction of oxygen metabolites normally scavenged by antioxidants such as superoxide dismutase (SOD), glutathione peroxidase, catalase and reduced glutathione (GSH) . Therefore, resistance of type II cells to hyperoxia may be linked to high antioxidant activities . To test this hypothesis we compared in vitro the effects of a 24 h exposure period to 95% O2 on cultured type II cells, lung fibroblasts and alveolar macrophages isolated from rats . We show that type II cells, when compared with other cell types, are highly sensitive to hyperoxia as shown by increased lactate dehydrogenase (LDH) release, decreased deoxyribose nucleic acid (DNA) and protein content of Petri dishes and decreased thymidine incorporation into DNA . Synthesis of dipalmitoylphosphatidylcholine was also significantly reduced . Antioxidant enzyme activities as well as glutathione content were not higher in type II cells than in other cell types . However, hyperoxia results in a decreased SOD activity and glutathione content in type II cells which was not observed in fibroblasts . We conclude that adaptative changes in SOD and glutathione metabolism could be important defence mechanisms in cells exposed to hyperoxia. Endocrinology, 1991 Aug, 129(2), 838 - 42 Rapid augmentation of prolactin cell number and secretory capacity by an estrogen-induced factor released from the neurointermediate lobe; Ellerkmann E et al.; 17 beta-Estradiol (E2) has been shown to exert an acute stimulatory effect on PRL secretion via an indirect action involving the neurointermediate lobe (NIL) . In the present study we used a reverse hemolytic plaque assay to determine whether this effect was manifested as an augmentation of the number of PRL secretors and/or an increase in the amount of hormone released per PRL cell . Cultures of anterior pituitary (AP) and NIL cells from ovariectomized rats were cultured overnight, exposed to the treatment (E2 or vehicle) for 3 h, and then subjected to a reverse hemolytic plaque assay that was carried out in the presence or absence of TRH . Concurrent exposure of AP cells to E2 and NIL cells evoked an 11-12% increase in the overall proportion of PRL-secreting cells . This was true when the AP and NIL cells were incubated as a mixed culture and when the two cell types were maintained in the same petri dish but on separate plastic supports, and TRH did not significantly influence this response . The effect of E2 on the number of PRL secretors was negated when NIL cells were not present throughout the experiment or when they were removed from the cultures just before commencement of E2 treatment . Simultaneous treatment with E2 and NIL cells also significantly augmented the sizes of PRL plaques produced under basal conditions by AP cells . Taken together, these results demonstrate that E2 stimulates NIL cells to release an activity that enhances PRL secretion in two ways: 1) by recruiting additional PRL cells into the secretory pool, and 2) by augmenting the secretory capacity of individual PRL cells. Transfusion, 1991 Jul-Aug, 31(6), 483 - 90 Inactivation of viruses in platelet suspensions that retain their in vitro characteristics: comparison of psoralen-ultraviolet A and merocyanine 540-visible light methods; Dodd RY et al.; The ability of two fundamentally different photochemical procedures to inactivate model viruses in platelet suspensions was compared . Merocyanine 540 (MC 540) with visible light was used as an example of an oxygen-dependent chemical-directed at the viral membrane, and aminomethyl trimethyl psoralen (AMT) with ultraviolet A light (UVA) was used as an example of a nucleic acid-directed system . Antiviral conditions in petri dishes were identified and the effects of these procedures on platelet suspensions in plastic storage containers were studied . Concentrations of photochemicals in the 10 to 150 mumol range with 30 to 60 minutes of visible light (MC 540) or 1 to 2 minutes of UVA (AMT) readily inactivated 5 to 6 log10 of vesicular stomatitis virus (VSV) and other model viruses in platelet suspensions, provided the plasma concentration was reduced to about 15 percent by the use of a synthetic platelet storage medium . Extracellular pH, morphology scores, and aggregation response dropped markedly when platelets were treated with MC 540 and visible light . However, treatment with 136 mumol per L of AMT and 1 to 3 minutes of UVA could inactivate 5 log10 of VSV in platelet suspensions with retention of platelet characteristics for 4 days, particularly if oxygen levels were reduced during treatment . These studies demonstrate that AMT-UVA treatment meets the initial requirements for virus inactivation in platelet suspensions. J Invest Dermatol, 1991 Jun, 96(6), 916 - 20 Pyrimidine dimer induction and repair in cultured human skin keratinocytes or melanocytes after irradiation with monochromatic ultraviolet radiation; Schothorst AA et al.; We compared the susceptibilities of cultured melanocytes and keratinocytes to dimer induction in DNA by monochromatic ultraviolet (UV) radiation . Keratinocytes as well as melanocytes were derived from human foreskin, grown as a monolayer in petri dishes, covered with phosphate-buffered saline containing 0.1% glucose, and irradiated . UV irradiation was carried out at 254, 297, and 302 nm as well as with a light source emitting predominantly 312 nm . The induction of pyrmidine dimers was assessed by determination of the number of T4 endonuclease V-sensitive sites (ESS) . We found a slightly higher response for dimer induction in melanocytes at 254, 297, and 302 nm; this difference was only significant at the 297-nm wavelength . Action spectra for pyrimidine dimer induction were derived from the exposure-response data obtained . The action spectra mimic to a large degree the action spectra for dimer induction in other cultured mammalian cells . The repair rate during a post-irradiation period lasting up to 24 h was substantially the same for the two cell types . The percentage of T4 endonuclease V-sensitive sites (ESS) remaining 9 and 24 h after irradiation was 45% and 30%, respectively. J Invest Dermatol, 1991 Jun, 96(6), 888 - 97 Mediators, initiating the inflammatory response, released in organ culture by full-thickness human skin explants exposed to the irritant, sulfur mustard; Rikimaru T et al.; Mediators released from injured human skin that initiate the inflammatory response have not been adequately identified . Organ culture of full-thickness skin explants enables us to do so, because injury to the skin can be made in vitro, eliminating the rapid leakage of serum and infiltration of leukocytes that occur in vivo . In our studies, the military vesicant sulfur mustard (SM) (10 microliters of a 0.01 to 1.0% dilution) was topically applied to injure the epidermis of the explant . Then, the explants were cultured in small Petri dishes, usually for 18 h at 36 degrees C, and the organ-culture fluids were assayed for various inflammatory mediators . We found that the culture fluids from SM-exposed and control explants contained similar amounts of angiotensin-converting enzyme, trypsin-like and chymotrypsin-like proteases, acid phosphatase, beta-glucuronidase, beta-galactosidase, lysozyme, deoxyribonuclease, ribonuclease, interleukin 1, and lactic dehydrogenase . However, the culture fluids from SM-exposed explants contained increased amounts of histamine and plasminogen-activating activity, and often prostaglandin E2, when compared to culture fluids from control explants . After 3 to 4 d in culture, full-thickness human skin explants, when exposed to 0.2% SM (but not when exposed to 1.0% SM), sometimes showed separation of the epidermis and increased collagenase activity (i.e., hydroxyproline release) . Thus, histamine (from local mast cells), and prostaglandin E2 and plasminogen-activating activity (probably from both mast cells and epidermal cells) are apparently involved in early mediation of the inflammatory response. Endocrinology, 1991 Jun, 128(6), 3299 - 309 Hormonal control of cell to cell communication: regulation by thyrotropin of the gap junction-mediated dye transfer between thyroid cells; Munari-Silem Y et al.; By microinjection of Lucifer yellow (LY) and analysis of the cell to cell transfer of the fluorescent probe, we have examined 1) the ability of thyroid cells in primary culture to reconstitute gap junctions and 2) the effects of extracellular signals on the functional activity of these junctions . Isolated thyrocytes cultured in tissue culture-treated petri dishes either formed monolayers or reorganized in follicular structures in the presence of the glycoprotein hormone TSH . In both culture conditions, LY-coupled cells were evident after 24-36 h . The communication between cells forming a reconstituted thyroid follicle was maintained for up to 9 days . In contrast, the dye coupling between cells in monolayer progressively decreased with time . The cell to cell communication, i.e., the number of dye-coupled cells in thyroid cell monolayer, was increased by TSH in a time- and concentration-dependent manner . The TSH action was not related to de novo protein synthesis . (Bu)2cAMP exhibited stimulatory effects similar, in terms of time course and amplitude of action, to those of TSH . The phorbol ester 12-O-tetradecanoyl phorbol 13-acetate rapidly inhibited both basal and TSH- or (Bu)2 cAMP-activated cell to cell communication . The dye coupling of cells in reconstituted follicles was also blocked by a short 12-O-tetradecanoyl phorbol 13-acetate treatment in both the presence and absence of TSH . Our data show that thyroid cells in culture, regardless of the full expression of the differentiated phenotype, rapidly reestablish intercellular gap junctions . The functional activity of gap junctions appears to be regulated 1) positively by a hormone, TSH, probably acting via the cAMP and protein kinase-A pathway, and 2) negatively by phorbol esters through the activation of protein kinase-C, the two regulatory pathways being interdependent. Invest New Drugs, 1991 May, 9(2), 149 - 57 Comparative toxicity of fostriecin, hepsulfam and pyrazine diazohydroxide to human and murine hematopoietic progenitor cells in vitro; Du DL et al.; The in vitro myelotoxic potentials of three investigational antitumor agents, Fostriecin, Hepsulfam and pyrazine diazohydroxide (PZDH), were evaluated utilizing clonogenic assays . Human and murine marrow cells were exposed to each drug for 1 hr prior to culture in microcapillary (human) or Petri dish (murine) assays . Fostriecin (0.22-220 microM), Hepsulfam (0.34-340 microM) and PZDH (0.68-680 microM) inhibited myeloid (CFU-gm), erythroid (BFU-e, CFU-e) and megakaryocytic (CFU-meg) colony formation in a concentration-dependent manner . CFU-e from both species were more sensitive to Fostriecin than the other progenitors and murine cells more sensitive overall to Fostriecin than their human counterparts . Murine CFU-e were also more sensitive to Hepsulfam than human CFU-e, with CFU-gm and BFU-e being similarly affected in both species . Human BFU-e were greatly inhibited by PZDH, whereas murine BFU-e were relatively resistant to its toxic effects . Fostriecin was the most toxic of the three antitumor agents, with PZDH the least toxic. J Clin Periodontol, 1991 May, 18(5), 305 - 11 Light and electron microscopic evaluation of biocompatibility, resorption and penetration characteristics of human collagen graft material; Quteish D et al.; This study was initiated to test the biocompatibility, resorption and penetration characteristics of human collagen graft material in vitro and in vivo using light (LM) and electron microscopy (EM) . To study this relationship, pieces of glutaraldehyde cross-linked collagen sponges (1 x 1 x 0.5 cm), were: (1) cultured in sterile Petri dishes with human gingival fibroblasts and human periodontal ligament fibroblasts for 2 weeks; (2) implanted in subcutaneous pockets made in both thighs (total 20 sites) of 10 Sprague-Dawley rats for 7-56 days . The behaviour of the growth of the fibroblasts was studied by inverted light microscopy (LM), then tissue culture specimens were studied from without and within using low-temperature scanning electron microscopy (LTSEM) . Blocks obtained from the graft sites of the rat were processed for LM and transmission EM . Long-term LM observations showed attachment and random orientation of cells on and around the collagen sponge in culture during the first 48 h . Between 7 and 14 days, the majority of the cells adjacent to the sponge were orientated at right angles to its margin with their long axes approximately parallel to each other . The LTSEM revealed that large numbers of HGF and HPLF grew onto the collagen sponges, but no cellular penetration to the middle of the sponge was seen . LM and TEM of the rat specimens showed a cellular reaction to the collagen graft, as well as slow resorption, and fibroblast invasion of the graft at 6-8 weeks . It was concluded that the human collagen graft was biocompatible with HGF and HPLF, with penetration first observed at 42 days post-implantation . In the in vivo study, the collagen underwent slow resorption over a period of 8 weeks. Am J Respir Cell Mol Biol, 1991 May, 4(5), 440 - 8 Attachment characteristics of bovine bronchial epithelial cells to extracellular matrix components; Rickard KA et al.; Attachment of cells to extracellular matrix (ECM) plays an important role in the regulation of cell growth and differentiated function . We hypothesized that bronchial epithelial cells preferentially attach to ECM proteins and utilize specific receptors for ECM proteins . Bronchial epithelial cells were obtained from bovine lung by protease digestion . Both freshly isolated and cultured bronchial epithelial cells were plated onto plastic petri dishes coated with bovine serum albumin, type I collagen, type IV collagen, fibronectin, laminin, ECM synthesized by cultured bronchial epithelial cells, or uncoated . Freshly isolated cells demonstrated significant attachment to ECM but weak attachment to other matrix proteins . Cultured bronchial epithelial cells attached well to ECM; however, they had relatively increased attachment to type I collagen, type IV collagen, fibronectin, and laminin compared to freshly isolated cells . To determine whether the attachment of bronchial epithelial cells is arginine-glycine-aspartic acid (RGD)-mediated, an RGD-containing peptide known to block attachment mediated by many integrin receptors was added to the media (400 micrograms/ml) . There was no inhibition of attachment of freshly isolated cells; however, there was significant but not complete inhibition of the attachment of the cultured cells to type IV collagen, laminin, and fibronectin, but not to type I collagen or ECM . Thus, freshly isolated bronchial epithelial cells readily adhere to ECM, and the attachment does not appear to be mediated by RGD-dependent receptors . Cultured bronchial epithelial cells demonstrate increased attachment to component proteins of ECM, and this attachment is, in part, to RGD-dependent receptors. Appl Microbiol Biotechnol, 1991 May, 35(2), 159 - 64 High density culture of anchorage-dependent animal cells by polyurethane foam packed-bed culture systems; Matsushita T et al.; Monkey kidney cells (Vero) and Chinese hamster ovary cells (CHO-K1) attached to the internal surface of polyurethane foam (PUF) and grew to a high cell density (1.1 x 10(8) cells/cm3 PUF and 4.2 x 10(7) cells/cm3 PUF, respectively) in a PUF-plates packed-bed culture system . This density of Vero cells was twice that obtained previously with a PUF-particles packed-bed culture system . A maximum cell density of 6.7 x 10(7) cells/cm3 culture vessel volume was obtained in a PUF-disc packed-bed culture of Vero cells . From the cell density of CHO-K1, growing in a monolayer on the surface of PUF and a petri dish, per bulk volume of PUF, we estimated that a surface area to volume ratio of PUF plates effective for cell growth was about 109 cm2/cm3. Exp Parasitol, 1991 Apr, 72(3), 311 - 20 Schistosoma mansoni: possible involvement of protein kinase C in linoleic acid-induced proteolytic enzyme release from cercariae; Matsumura K et al.; The possible involvement of protein kinase C and Ca2+ metabolism in the proteolytic enzyme release from schistosome cercariae was studied . Cercariae were placed in dechlorinated tap water containing 0.37 mM calcium in the small glass petri dish and exposed to the stimuli (linoleic acid, phorbol esters, and Ca2+ ionophore) with or without inhibitors of protein kinase C or Ca2+ metabolism . The proteolytic activity of incubation medium of cercariae thus treated was measured by the azocoll assay . The penetration response of cercariae induced by linoleic acid, a physiological stimulus, was mimicked by phorbol esters . When exposed to phorbol esters, 0.02 to 2 microM of 12-O-tetradecanoylphorbol-13-acetate (TPA) and 0.2 to 2 microM of phorbol-12,13-dibutyrate (PDBu), cercariae ceased the swimming movement, began a rhythmic thrusting of the anterior tip of the parasite, and released the proteolytic enzyme, but they did not shed the tails . Lowering Ca2+ in water by addition of 5 mM ethylene glycol-bis(beta-aminoethyl ether) N,N,N',N'-tetraacetic acid (EGTA), phorbol ester-induced release of enzyme was completely inhibited . Phorbol ester-induced release of enzyme was partially inhibited by 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), an inhibitor of protein kinase C, at a concentration of 100 microM . H-7 alone, at a concentration of 100 microM, did not affect the swimming movement of cercariae . The cercariae were stimulated to release the enzyme by high concentrations (10 and 100 microM) of the Ca2+ ionophore, A23187, but enzyme was not released by low concentrations (0.5 and 1 microM) of this drug . Cercariae exposed to A23187 behaved differently from those exposed to phorbol esters . They ceased swimming, showed strong muscle contraction, and shed their tail . A23187 stimulated cercariae to release the enzyme in the water containing 5 mM EGTA . A23187-induced enzyme release was not inhibited by N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), a calmodulin antagonist, trifluoperazine (TFP), a better calmodulin antagonist on schistosome, or by verapamil, a Ca2+ channel blocker . Linoleic acid-induced release of enzyme was partially inhibited by 0.5 and 5 mM of EGTA and by 1 to 100 microM of H-7 . While it was not inhibited by N-{2-(methylamino)ethyl}-5-isoquinolinesulfonamide (H-8) and N-(2-guanidinoethyl)-5-isoquinolinesulfonamide (HA-1004), inhibitors of cyclic nucleotide-dependent protein kinase which were used as negative controls of H-7, W-7, TFP, 8-(N,N-diethylamino)octyl 3,4,5-trimethoxybenzoate (TMB-8), an intracellular Ca2+ antagonist, and verapamil.(ABSTRACT TRUNCATED AT 400 WORDS) J Neurosci Methods, 1991 Apr, 37(2), 173 - 82 A simple method for organotypic cultures of nervous tissue; Stoppini L et al.; Hippocampal slices prepared from 2-23-day-old neonates were maintained in culture at the interface between air and a culture medium . They were placed on a sterile, transparent and porous membrane and kept in petri dishes in an incubator . No plasma clot or roller drum were used . This method yields thin slices which remain 1-4 cell layers thick and are characterized by a well preserved organotypic organization . Pyramidal neurons labelled by extra- and intracellular application of horse radish peroxidase resemble by the organization and complexity of their dendritic processes those observed in situ at a comparable developmental stage . Excitatory and inhibitory synaptic potentials can easily be analysed using extra- or intracellular recording techniques . After a few days in culture, long-term potentiation of synaptic responses can reproducibly be induced . Evidence for a sprouting response during the first days in culture or following sections is illustrated . This technique may represent an interesting alternative to roller tube cultures for studies of the developmental changes occurring during the first days or weeks in culture. Clin Rheumatol, 1991 Mar, 10(1), 28 - 30 Mononuclear cells are not involved in BGP synthesis and secretion; Cantatore FP et al.; Osteocalcin or bone GLA protein (BGP) is found at high levels in only two tissues, the extracellular matrix of bone and dentine . Tissue culture experiments have demonstrated that BGP is synthesized by two osteoblastic osteosarcoma cell lines (ROS 2/3 and 17/2) and by normal osteoblastic cells in primary culture . BGP was not found in rat cartilage nor in liver, kidney, lung, spleen, brain, heart, thymus, skeletal muscle . In this study secretion of BGP was assayed by RIA in the supernatants of 48-hour cultures of peripheral blood lymphocytes or monocytes . Lymphocyte cultures were carried out using RPMI-1640 supplemented with L-glutamine and antibiotics at the concentration of 1 x 10(6) cells/ml and activated by PHA (10 ng/ml) . Peripheral blood monocytes were purified by adherence to plastic Petri dishes and treated with cold PBS supplemented with EDTA . Monocytes were cultured as previously described and stimulated with LPS (50 micrograms/ml) . Cell-free supernatants were obtained by centrifugation and stored at -20 degrees C, until the BGP assay was performed . The authors did not observe secretion of detectable amounts of BGP in the supernatants of short-term lymphocyte or monocyte cultures . These data indicate that circulating mononuclear cells are not involved in BGP synthesis and secretion. Radiology, 1991 Mar, 178(3), 677 - 9 US-assisted aspiration thrombectomy: in vitro investigations; Schmitz-Rode T et al.; The authors describe the use of a new ultrasound (US)-aspiration thrombectomy technique . An oscillating US probe was inserted into a thin-walled, large-bore aspiration catheter . Experiments tested the ability of the new device and other catheter combinations to remove clot material from a Petri dish, as well as from small and large vessel models made of silicone and glass tubes, respectively . Results of the experiments demonstrated that an oscillating 1.0-mm US probe inserted into an aspiration catheter (7-9 F in diameter) promoted clot fragmentation and allowed continuous aspiration of thrombi of any size . When compared with simple large-bore catheter aspiration and with mechanical fragmentation by means of a US probe within a catheter that was flushed to cool the probe, US-assisted aspiration thrombectomy demonstrated significantly better results for percutaneous transcatheter removal of fresh thrombi. AJNR Am J Neuroradiol, 1991 Mar-Apr, 12(2), 279 - 81 Metallic otologic implants: in vitro assessment of ferromagnetism at 1.5 T; Shellock FG et al.; MR imaging is contraindicated for patients with certain ferromagnetic implants because of potential risks related to movement or dislodgement . This is especially true for metallic implants located in sensitive areas of the body, such as those placed in and around the ear . Therefore, the ferromagnetic qualities of 35 different metallic otologic implants were assessed by placing them individually on a millimeter scale in a plastic petri dish that was slowly moved into the center of a 1.5-T MR imaging system . None of the metallic otologic implants moved during this procedure . The results demonstrate that each of these implants are made from nonferromagnetic materials and do not pose a risk to patients undergoing high-field-strength MR imaging . These data effectively expand the list of metallic implants that appear to be safe for MR imaging. Prostaglandins, 1991 Mar, 41(3), 263 - 81 Metabolism of arachidonic acid by guinea pig Clara cells; Laporte J et al.; A method for the isolation of non-ciliated bronchiolar epithelial (Clara) cells from the guinea pig is described . Following digestion of the lung tissue with Type XXIV protease, the isolated lung cells showed a viability greater than 90% and contained 3% of Clara cells . Several cell populations were then separated on the basis of size using 2 centrifugal elutriations . The macrophages and endothelial cells were removed from the Clara cells enriched fractions by differential adherence on Petri dishes . The Clara cell-rich suspension was then further purified by centrifugation on Percoll non-continuous density gradients consisting of 48-52-55% Percoll solution . The lower interface and the pellet of the non-continuous gradient consisted of approximately 80% Clara cells . Identification of isolated Clara cells was confirmed by light microscopic observations after nitroblue tetrazolium staining and by ultrastructural characteristic features as observed by electron microscopy . The metabolism of arachidonic acid into prostaglandins and TxB2 by purified Clara cells was examined by enzyme immunoassay (EIA) and leukotriene formation was investigated by reverse phase high performance liquid chromatography (RP-HPLC) . Enriched guinea pig Clara cells incubated with arachidonic acid released TxB2, PGE2 and 6-keto PGF1 alpha, but did not produce leukotrienes . These cells could however transform exogenous leukotriene A4 into leukotriene B4 . These results suggest that guinea pig Clara cells possess the enzymes of the cyclooxygenase pathway required for TxB2, PGE2 and 6-keto-PGF1 alpha synthesis . Clara cells do not possess the 5-lipoxygenase enzyme but show some leukotriene A4 hydrolase activity since they can produce leukotriene B4 upon incubation with leukotriene A4. Lancet, 1991 Feb 2, 337(8736), 269 - 70 Survival of scrapie virus after 3 years' interment; Brown P et al.; Supernatant fluid from a scrapie-infected hamster brain homogenate was mixed with soil, packed into perforated petri dishes that were then embedded within soil-containing pots, and buried in a garden for 3 years . Between 2 and 3 log units of the input infectivity of nearly 5 log units survived this exposure, with little leaching of virus into deeper soil layers . These results have implications for environmental contamination by scrapie and by similar agents, including those of bovine spongiform encephalopathy and Creutzfeldt-Jakob disease. Anat Rec, 1991 Jan, 229(1), 125 - 8 A new technique for explantation and in vitro cultivation of chicken embryos; Dugan JD Jr et al.; A technique is described for explanting and cultivating chicken embryos in plastic drinking cups which have been modified with plastic wrap to reproduce the geometry and dimensions of the egg shell . Successful explantation rates of 97% are possible with a double-window technique, and survivability in cups exceeds that achievable in other in vitro systems (i.e., petri dishes) . Long-term survival to the 21st day of incubation is seen routinely . This system with cups is less expensive than that with petri dishes, and simpler than that with plastic wrap/tripods . Thus, this new method of in vitro cultivation of chicken embryos improves upon explantation rate, survivability and system design, and has a wide range of applications in developmental biology, angiogenesis, cancer, and pharmacology research. J Clin Gastroenterol, 1991, 13 Suppl 1, S58 - 64 Gastric mucosal repair in vitro; Giebel J et al.; We tested whether cultured gastric mucosal cells would be suitable to study the two major steps of repair: restitution and proliferation . Preparations of freshly isolated epithelial cells of guinea pig gastric mucosa were used for the studies . The cells attached to Petri dishes within 10 h and formed monolayers after 48-72 h . Electron microscopy showed that each cell type was able to form lamellipodia (i.e., cell protrusions) during restitution in vivo . When monolayers were wounded with a razor blade, most cells at the edge of damage died within a few minutes, but some recovered from injury . Later, intact cells migrated from the edge into the denuded zone and restored the monolayer within 24-48 h . An increased number of cells near the edge started to synthesize DNA . In conclusion, this model allows one to study in vitro both aspects of mucosal repair. Basic Life Sci, 1991, 58, 3 - 19; discussion 19-25 The molecular biology of radiation carcinogenesis; Hall EJ et al.; Major new insights into carcinogenesis have come from recent advances in cellular and molecular biology . The concept of oncogenes provides a simple explanation for how agents as diverse as radiation, chemicals or retroviruses can induce tumors that are indistinguishable one from another . Oncogenes may be activated by a point mutation, by a chromosome translocation, or by amplification . Ionizing radiations are efficient at the first two mechanisms . While oncogenes are frequently associated with leukemias and lymphomas, they are associated with only 10 to 15% of human solid cancers . The importance of the loss of suppressor genes was suggested first from studies with human-hamster hybrid cells, but has since been shown to be of importance in an increasing number of human solid tumors, from rare tumors such as retinoblastoma to more common tumors such as small cell lung cancer and colorectal cancer . The mechanism of somatic homozygosity clearly involves several steps, some of which, such as a deletion, could be readily produced by ionizing radiation . The multi-step nature of carcinogenesis can be demonstrated in the petri dish, where the transfection of multiple oncogenes is required to transform normal cells from short-term explants . It can be shown, too, in colorectal cancer in the human, where the activation of an oncogene and the loss of more than one suppressor gene may be involved in the progression from normal epithelium to a frank malignancy. Nephrol Dial Transplant, 1991, 6 Suppl 3, 53 - 6 Suppression of beta 2-microglobulin release from lymphocytes by dialysis membranes; Schaefer RM et al.; As lymphocytes are one of the main sources of circulating beta 2-microglobulin (beta 2M), the direct effect of different dialysis membranes on beta 2M release from those cells was studied in vitro . Lymphocytes were isolated from 11 long-term haemodialysis patients and nine healthy controls . Cells were cultured on flat sheet membranes made from either Cuprophan, Hemophan, or polyacrylonitrile . Polystyrole petri dishes were used as controls . Beta 2M concentrations in the supernatant were measured after 3 and 7 days of culture by ELISA techniques . Beta 2M release from lymphocytes obtained from uraemic patients was almost identical to the release from healthy subjects . In the presence of all three membranes the release of beta 2M was less than that produced on polystyrole . This held true for lymphocytes isolated from both healthy and uraemic subjects . As for the three membranes the release of beta 2M into the supernatants was statistically the same when the adsorptive capacity of polyacrylonitrile was taken into account . However, there was a tendency for Cuprophan to exert the strongest inhibition, while Hemophan and polyacrylonitrile reduced beta 2M release to a lesser degree . Based on these data it seems that prolonged interaction between dialysis membranes and lymphocytes does lead to a reduction in beta 2M release. Bioelectromagnetics, 1991, 12(5), 299 - 314 Morphological and electrophysiological changes produced by electrical stimulation in cultured neuroblastoma cells; Krauthamer V et al.; Electric fields, which were equivalent to those generated by medical devices, were applied to cultured neuroblastoma cells (mouse and human) to test for morphological damage and to establish damage thresholds . Each of two methods of applying fields permitted flow of electrical current and minimized exposure of cells to electrode-breakdown products . One method consisted of a pair of parallel wires in a Petri dish by which current was delivered within a fixed volume of flowing tissue-culture media . With the other method, the cells were held in a confined geometrical chamber and current was applied via agar bridges . Under a given set of stimulation parameters, damage was found to be variable from cell to cell . By changing the strength of the electric field (frequency and duration of stimulation held constant), thresholds of several V/cm were found above which cell damage could be reliably produced . Depending on the intensity of the field, damage took the form of cell lysis or damage to neurites . Intracellular recordings from the mouse neuroblastoma cells revealed a correlation between a decline in resting transmembrane potential and stimulus intensity . Human neuroblastoma cells were less susceptible to damage than were the mouse neuroblastoma cells, given the same strength of applied electric fields. Lens Eye Toxic Res, 1991, 8(2-3), 281 - 309 Cell-substratum interactions and the cytoskeleton in cell shape-mediated growth regulation of lens epithelial cells; Iwig M et al.; Cell attachment to a suitable substratum is a precondition for the mitotic growth of nontransformed lens epithelial cells . Cultering of cells in suspension results in a strong decline of the DNA synthetic rate, whereas reattachment induces the reentrance into the cell cycle . Further studies revealed that not anchorage itself but cell flattening is prerequisite for the entrance of cells into the cycle . Flattened cells exert tension to the substratum via numerous filopodia . If the rigidity of the substratum is reduced by loosening of the collagen gel from the bottom of the petri dish, the gel becomes contracted by the traction forces of the cells and the cell shape becomes transformed from a flattened shape into a more spheroidal or longstretched one . This cell shape transition is connected with a decrease in RNA- and protein synthesis and a stop of DNA synthesis . During further experiments it was demonstrated that microfilaments are involved in gel contraction and cell shape alteration, respectively . Furthermore, intact microfilaments are needed for G0-G1-S-transition . Desintegration of microfilaments by cytochalasin is without influence on ongoing DNA synthesis but hinders strongly the entrance of cells into the S-phase . The survey gives some recent results on the molecular basis of cell substratum interactions as well as the structure and function of the cytoskeleton . The role of the cytoskeleton in cell shape-mediated growth regulation is discussed. J Physiol (Paris), 1991, 85(3), 158 - 70 Behavioural effects of genetically engineered cells releasing dopa and dopamine after intracerebral grafting in a rat model of Parkinson's disease; Horellou P et al.; The relative importance of synaptic versus paracrine dopamine transmission for the occurrence of functional effects following intrastriatal grafting is not fully established . In the present study we grafted cell lines, expressing the form I of human tyrosine hydroxylase after infection with a recombinant retrovirus and selection in tyrosine-free-medium, to the denervated striatum in order to analyse the extent to which extracellular dopamine levels can be restored and the effect of a diffuse release of dopamine on motor impairement in a rat model of Parkinson's disease . In petri dish, the modified fibroblast cells (NIH.3T3) release DOPA constitutively whereas the modified endocrine cells (RIN) store and release dopamine in a regulated way . Interestingly, in denervated striatum, grafts of modified fibroblast cells produce DOPA which was efficiently converted into dopamine by the host striatal tissue . In the grafted striatum, both fibroblast and endocrine cells restore subnormal levels of diffuse release of dopamine which is notably unaffected and stimulated, respectively, by high concentration of potassium, in connection with the in vitro properties of the grafted cells . The intrastriatal grafts of modified cells partially reversed the apomorphine-induced but not the amphetamine-induced motor asymmetry . We discuss the implications of these results in the context of Parkinson disease. Med Radiol (Mosk), 1991, 36(11), 22 - 3 {A comparative evaluation of 2 methods for cloning human lung tumors}; Sviridova IK et al.; Clonogenic capacity of tumor cells from 59 human lung cancer specimens was investigated in Petri dishes (24 specimens) and in capillaries (40 specimens), and parallel cloning was done in 5 cases . The results were as follows: (a) bacterial contamination in capillaries was noted less frequently than in Petri dishes (12.5 vs . 25%); (b) cells did not form colonies in capillaries in 42.5% and in Petri dishes in 25%; (c) colony-forming effectiveness in capillaries was 3.46-fold higher than that in Petri dishes; (d) the amount of tumor cells for analysis in capillaries is 10-fold less than that for analysis in Petri dishes . However, the absolute number of colonies, indispensable for assessment of tumor sensitivity to antitumor action (not less than 15 colonies per capillary or Petri dish) was obtained in 67% in Petri dishes and in 33% only in capillaries. Differentiation, 1990 Dec, 45(3), 221 - 9 Filaggrin production by cultured human epidermal keratinocytes and its regulation by retinoic acid; Asselineau D et al.; Filaggrin is a basic protein normally present in the stratum corneum of epidermis . It derives from a high-molecular-weight precursor synthesized in the stratum granulosum of epidermis . This precursor, called profilaggrin, is thought to be associated with the keratohyaline granules of granular cells . It is known that profilaggrin, but not filaggrin, is present in conventional cultures of human keratinocytes grown on plastic petri dishes . In this study, we show that cultured human keratinocytes can convert profilaggrin into filaggrin, when they are grown on a collagen matrix and raised at the liquid-air interface in order to induce terminal differentiation . Moreover, the presence of terminally differentiating keratinocytes above the granular layer is necessary, but not sufficient, for the accumulation of filaggrin . Finally, we show that the accumulation of filaggrin in the outermost layers of submerged cultured human keratinocytes can be triggered by extensive removal (double delipidization) of retinoids from the serum supplement and inhibited when small concentrations (10(-11)-10(-10) M) of retinoic acid are readded to the culture medium . Altogether, the data reported suggest that not only the synthesis of profilaggrin, but also the conversion of profilaggrin into filaggrin are negatively controlled by retinoic acid . Further, it seems that retinoic acid acts directly on the conversion of profilaggrin into filaggrin rather than on the production of terminally differentiating cells capable of accumulating this protein. Boll Soc Ital Biol Sper, 1990 Nov, 66(11), 1043 - 50 {Survival of bone tissue in organotypic culture under intermittent mechanical loading . Preliminary results}; Lozupone E et al.; With the aim to study the mechanism of transduction of mechanical stimuli in biological ones we have realized an experimental device for the application of intermittent mechanical forces on bone specimens in vitro . The scheme of the device is reported in Fig . 1 . It is constituted by a drive shaft which rotates on eccentric axis (1) supporting a longitudinal bar (2) with the load (3) . The latter rests on a piston (4) only during a limited period of every shaft revolution, so that the load becomes intermittent . The bone specimen (5) is placed under the piston and the two are placed in a tube containing the culture medium . This latter is BGJ mod . Fitton-Jackson (Gibco), enriched with fetal calf serum (10%) and ascorbic acid (70 microliters/ml) . Right metatarsi from 18-day-old rats were removed aseptically and placed under the piston for 2-6 days after resection of both ends . The homotypic ones, unloaded, were placed in 30 mm Petri dishes, and used as a control . The incubator environment was 5% CO2 in air (A group), or enriched with O2 (25-35%) (B group) . At the end of the experimental period the bone specimens were fixed in 4% formalin buffered and treated for conventional histologic methods . In the A group most of the osteocytic lacunae were empty . The osteoblasts disappeared already at the 2nd day; the periosteal fibroblast dedifferentiated and multiplied . The deposition or calcification of osteoid were completely lacking . The application of mechanical load promoted deposition of granular degenerative material around the bone, and the periosteal cells, well differentiated, were surrounded by metachromatic material, which resembles cartilage matrix.(ABSTRACT TRUNCATED AT 250 WORDS) Mol Gen Genet, 1990 Nov, 224(2), 177 - 82 Localization by restriction fragment length polymorphism mapping in potato of a major dominant gene conferring resistance to the potato cyst nematode Globodera rostochiensis; Barone A et al.; A major dominant locus conferring resistance against several pathotypes of the root cyst nematode Globodera rostochiensis was mapped on the linkage map of potato using restriction fragment length polymorphism (RFLP) markers . The assessment of resistance versus susceptibility of the plants in the experimental population considered was based on an in vivo (pot) and an in vitro (petri dish) test . By linkage to nine RFLP markers the resistance locus Gro1 was assigned to the potato linkage group IX which is homologous to the tomato linkage group 7 . Deviations from the additivity of recombination frequencies between Gro1 and its neighbouring markers in the pot test led to the detection of a few phenotypic misclassifications of small plants with poor root systems that limited the observation of cysts on susceptible roots . Pooled data from both tests provided better estimates of recombination frequencies in the linkage interval defined by the markers flanking the resistance locus. Neurosci Lett, 1990 Oct 16, 118(2), 172 - 6 Carbon filaments provide support and directionality to growing rat fetal spinal cord explants; Khan T et al.; Spinal cord explants obtained from 15 to 17-day-old fetal rats were cultured on bundles of 5-7 microns diameter carbon filaments attached to the bottom of Petri dishes . After a 3 week incubation period, the cultures were fixed and observed by light, scanning, and transmission electron microscopy . Neurites and glial processes were found to be growing both on and between the carbon filaments . The carbon filaments appeared to provide a biocompatible scaffold which promoted adhesion and gave directionality to the growing cell processes . These properties may make carbon filaments a suitable substrate for in vivo implantation into the damaged spinal cord. Immunol Lett, 1990 Oct, 26(1), 11 - 5 MHC class II antigens on canine bronchoalveolar cells; Chang SC et al.; To evaluate the expression of MHC (major histocompatibility complex) antigens on canine bronchoalveolar cells (BAC), bronchoalveolar lavages (BAL) were performed in mongrel and German shepherd dogs . MHC class II antigens on canine BAC and peripheral blood mononuclear cells (PBMC) were detected by monoclonal antibodies (mAbs) B1F6, 7.5.10.1 and Q5/13 recognising canine MHC class II antigens, using cytofluorometry . These mAbs reacted with more than 20% of BAC and PBMC in both breeds of dog . The percentage of MHC class II positive cells in BAC were lower than those in PBMC . There was no significant difference in the percentages of MHC class II positive BAC and PBMC in mongrel and German shepherd dogs . To further identify the expression of MHC class II antigens on BAC, the cells were separated into adherent and nonadherent cells by petri dish adherence . The percentages of MHC class II positive cells in adherent and non-adherent cell populations were similar . Nearly half the lymphocytes in normal BAC were T cells detected by mAbs F3-20-7 and 1A1; B cells were scarce and represented less than 10% of nonadherent cells . Immunoprecipitation by anti-MHC class II mAbs, and SDS-polyacrylamide gel electrophoresis (SDS-PAGE) revealed MHC class II-like molecules on canine BAC and PBMC . After stimulation with phytohaemagglutinin (PHA), the percentages of class II positive cells in BAC and PBMC were significantly increased . Thus, these anti-MHC class II mAbs may prove to be of advantage in experiments designed to evaluate the changes in class II antigen expression on canine BAC during the course of immune response in the lung, as in pulmonary allograft rejection. J Biol Chem, 1990 Sep 5, 265(25), 15261 - 6 Parathyroid hormone-induced alterations of protein content and phosphorylation in enriched apical membranes of opossum kidney cells; Reshkin SJ et al.; Parathyroid hormone (PTH) reduces Na/Pi co-transport activity in opossum kidney (OK) cells in a process mediated by protein kinases A and C . Further, inactivation of Na/Pi transport involves irreversible inhibition, possibly via internalization, of the transport system . This study analyzed alterations of concentration and phosphorylation of membrane proteins of an apically enriched preparation induced by short (10 min) and long (3 h) term incubation with 10(-10) M PTH of monolayer cultures of the OK-cell line . To this end, an apically enriched membrane fraction was isolated from cells grown on Petri dishes and analyzed by two-dimensional gel electrophoresis . Long term exposure of the cells to PTH induced changes in apical protein concentration . Four proteins were found to be decreased and one protein was found to be increased in its concentration . Addition of 10(-10) M PTH to the cells led to transient phosphorylation of five proteins . In contrast to transient phosphorylation, phosphorylation of one protein increased over the time period of 3 h . Combined analysis of silver staining and autoradiography led to the detection of an acidic 35-kDa protein in which specific phosphorylation increased over a time period of hours . The results document for the first time alterations in apical membrane protein content and phosphorylation state mediated by PTH when added to an intact cellular system . It is concluded that the identified proteins represent possible candidates for being involved directly or indirectly in PTH alterations of membrane transport. J Invest Dermatol, 1990 Sep, 95(3), 325 - 32 Full-thickness human skin explants for testing the toxicity of topically applied chemicals; Nakamura M et al.; This report describes a model organ-culture system for testing the toxicity of chemical substances that are topically applied to human skin . In this system, the viable keratinocytes in the full-thickness skin explants are protected by the same keratinized layer as skin remaining on the donor, and toxicity can be assessed microscopically and/or biochemically . The human skin specimens were discards from a variety of surgical procedures . They were cut into full-thickness 1.0-cm2 explants, and briefly exposed to the military vesicant sulfur mustard (SM), which was used as a model toxicant . The explants were then organ cultured in small Petri dishes for 24 h at 36 degrees C . In the 0.03-1.0% dosage range, a straight-line dose-response relationship occurred between the concentration of SM applied and the number of paranuclear vacuoles seen histologically in the epidermis . Within the same SM dosage range, there was also a proportional decrease in 14C-leucine incorporation by the explants . Thus, the number of paranuclear vacuoles reflected decreases in protein synthesis by the injured epidermal cells . The epidermis of full-thickness untreated (control) human skin explants usually remained viable for 7 d when stored at 4 degrees C in culture medium . During storage, a relatively small number of paranuclear vacuoles developed within the epidermis, but the explants were still quite satisfactory for testing SM toxicity . Incubation (for 4 or 24 h at 36 degrees C) of such control skin explants reduced (often by 50%) the small number of paranuclear vacuoles produced during 4-7 d of storage . This reduction was probably caused by autolysis of many of the vacuolated cells . Two types of paranuclear vacuoles could be identified by both light and electron microscopy: a storage type and a toxicant type . The storage type seemed to be caused by autolysis of cell components . The toxicant type seemed to be caused by an invagination of the plasma membrane . Only toxicant-type vacuoles increased appreciably in number when skin explants were exposed to mustard, and to other toxicants. J Am Mosq Control Assoc, 1990 Sep, 6(3), 406 - 10 Mosquito attraction to substances from the skin of different humans; Schreck CE et al.; Mosquito attraction responses to substances collected from human skin and placed on glass petri dishes were studied . Mosquito response varied according to the source of the substance . Substances removed from the head and hands elicited the greatest attraction response in laboratory-reared mosquitoes . Mosquito response lasted up to 6 h when the substance was aged and was increased by warming the samples from ca . 25 degrees C to 37 degrees C . Of the 12 mosquito species studied, attraction response was greatest in Aedes aegypti . It is cautioned that residues deposited by handling traps or other apparatus used in mosquito studies may influence test results. Am J Respir Cell Mol Biol, 1990 Sep, 3(3), 217 - 26 Induction of surfactant protein (SP-A) biosynthesis and SP-A mRNA in activated type II cells during acute silicosis in rats; Miller BE et al.; The synthesis of the major surfactant protein, SP-A, was studied in activated alveolar type II cells isolated from the lungs of rats exposed to silica by intratracheal instillation . Exposure of rats to silica resulted in large increases in the levels of disaturated phosphatidylcholine and SP-A in the extracellular and intracellular surfactant compartments . Isolated type II cells were used to determine if the observed increases in SP-A were associated with increased SP-A synthesis . Type II cells were isolated by a combination of elastase digestion, centrifugal elutriation, and differential adherence on IgG-coated petri dishes . Type II cells from silica-treated lungs were separated into two populations, designated type IIA and type IIB . The type IIB, or activated population, consisted of type II cells that were larger than normal type II cells and, in addition, contained larger and more numerous lamellar bodies than normal type II cells . Type IIB cells contained 4.3-fold higher levels of SP-A compared to normal type II cells . SP-A synthesis was measured by incubating freshly isolated cells with {35S}Translabel (70% {35S}methionine, 15% {35S}cysteine) for up to 4 h in methionine-free medium, followed by immunoprecipitation of newly synthesized protein . The rate of SP-A synthesis was increased approximately 6.7-fold in the activated type II cells . Analysis of the newly synthesized protein by one-dimensional SDS-PAGE indicated three intracellular forms of SP-A with molecular weights of approximately 26,000, 30,000, and 34,000 . In type II cells from control rats, the 34-kD protein accounted for approximately 93% of the newly synthesized SP-A after 4 h of incubation; only a small amount of radioactivity was associated with the lower molecular weight species . The increased biosynthesis of SP-A in the activated type II cells was associated with a 7.3-fold increase in the level of SP-A mRNA . These results indicate that the content and synthesis of SP-A are both highly elevated in activated type II cells and that these increases may be due to increased levels of SP-A mRNA. Can J Physiol Pharmacol, 1990 Sep, 68(9), 1226 - 30 Reexamination of dopamine as the prolactin-release inhibiting factor (PIF): supplementary agent may be required for dopamine to function as the physiological PIF; Shin SH et al.; A large number of studies have been performed concerning dopamine's inhibitory effect on prolactin release, but many of these studies have examined the effect of dopamine dissolved in a solution containing ascorbic acid . Ascorbic acid, routinely used to protect dopamine from oxidation, alone does not stimulate or inhibit prolactin release, but it can potentiate the inhibitory effect of dopamine in a static monolayer culture system by approximately 100 times . We have closely examined the inhibitory effect of dopamine on prolactin release in the absence of ascorbic acid using a perifusion system . Male rat adenohypophyses were dispersed with trypsin and cultured in a Petri dish to form cell clusters . Inhibition of prolactin release by dopamine (1 mumol/L) in the absence of ascorbic acid was sustained for only 63 min during the 2-h perifusion period . Following a 2-h period of incubation of dopamine in the same experimental solution, the dopamine concentration was reduced from 1 to 0.18 mumol/L, yet this "2-h-old dopamine" was still effective in inhibiting prolactin release (approximately 30 min) . This result suggests that the lactotrophs may be desensitized by chronic exposure to a high concentration of dopamine in the absence of ascorbic acid . In contrast, when a low concentration of dopamine (3 nmol/L) containing ascorbic acid (0.1 mmol/L) was perifused, inhibition of prolactin release was sustained for the entire 2-h perifusion period . Although there may be a large number of explanations for dopamine's transient inhibitory effect on prolactin release, the present results suggest that dopamine may require supplementary agent(s) to effectively inhibit prolactin release and thus function as the prolactin release inhibitory factor (PIF).(ABSTRACT TRUNCATED AT 250 WORDS) Dtsch Zahnarztl Z, 1990 Aug, 45(8), 449 - 51 {In vitro growth of periodontal ligament fibroblasts between bone and root slices}; Sharaf MN et al.; In a first experiment the proliferation and growth kinetics of periodontal ligament fibroblast monolayers generated from human healthy (HPDL) and diseased (DPDL) periodontia were compared . In a second experimental series a periodontal ligament-like space was developed by laying a root slice inside a bony ring slice on the bottom of a tissue culture petri dish and leaving a space of about 0.5 mm between both structures . HPDL cells showed a higher proliferation rate than DPDL cells . Growing into the space between the slices, the PDL cells built up a matrix which was composed of cells and fibrils . HPDL cells built up the matrix more rapidly than DPDL cells. Biochim Biophys Acta, 1990 Jul 12, 1053(2-3), 135 - 43 Proteoglycans synthesized in vitro by nude and normal mouse peritoneal macrophages; Petricevich VL et al.; Peritoneal macrophages from nude mice were found to be functionally similar to 'activated' macrophages from normal mice . The objective of the present study was to characterize the proteoglycans synthesized and secreted in vitro by peritoneal macrophages isolated from nude and normal Balb/c mice and to investigate the relationship between macrophage 'activation' and changes in the proteoglycan patterns . Macrophages obtained by peritoneal lavage were seeded in Petri dishes . After 2 h incubation at 37 degrees C, the adherent cells (macrophages) were exposed to {35S}sulphate for the biosynthetic labelling of proteoglycans . After incubation, the cell and medium fractions were collected and analysed for proteoglycans and glycosaminoglycans . The glycosaminoglycans were identified and characterized by a combination of agarose gel electrophoresis and enzymatic degradation with specific mucopolysaccharidases . It was shown that 3/4 of the total 35S-labelled glycosaminoglycans were in the extracellular compartment after 24-48 h . The macrophages synthesized dermatan sulphate (68%), chondroitin sulphate (7%) and heparan sulphate (25%) . Both cell and medium fractions of normal and nude mouse macrophages contained glycosaminoglycans with the same ratios, although the nude mouse macrophages synthesized 2-fold less glycosaminoglycans than the normal mouse macrophages . Lower levels of 35S-proteoglycans were also obtained from in vitro 'activated' macrophages, but the ratios of dermatan sulphate:chondroitin sulphate: heparan sulphate were altered in these cells as compared to the control . Furthermore, all the 35S-macromolecules found in the extracellular compartment of nude and normal control cells were of proteoglycan nature, in contrast to the medium fractions of 'activated' macrophages, which contain both intact proteoglycans and 'free' glycosaminoglycan chains . These results indicate that, at least as regards the proteoglycans and glycosaminoglycans, the nude mouse macrophages are not identical to the 'activated' macrophages from normal mice. J Steroid Biochem, 1990 Jul 4, 36(4), 377 - 81 Testosterone micromilieu in staged rat seminiferous tubules; Parvinen M et al.; Endogenous testosterone concentrations in rat seminiferous tubules were measured in relation to different stages of the cycle of the seminiferous epithelium . For this purpose, the seminiferous tubules were mechanically separated from the interstitial tissue on a cooled (1 degree C) petri dish under a stereomicroscope without added medium . After recognition of the stages of the cycle by transillumination, the specimens were rapidly transferred by dry forceps into test tubes for testosterone radioimmunoassay . The results of the dry dissection method were compared with measurements on tubules that were kept after separation in phosphate buffered saline (PBS, pH 7.4), in order to reveal the possible leakage of testosterone from the tubules . The maximal concentration of testosterone per unit length of seminiferous tubule was found in stages VII and VIII of the cycle (288 +/- 60 fmol/cm, mean +/- SEM, n = 12), and the minimal in stages IX-XII (219 +/- 57 fmol/cm, P less than 0.01) . If the levels were correlated with unit volumes of the seminiferous tubules, identical concentrations of testosterone (521-542 fmol/mm3, approx . 500 nmol/l) were found in the different stages of the cycle . Despite the similarity of testosterone concentrations in the different parts of the seminiferous tubules the local concentrations of biologically active (i.e . free) testosterone may be modulated by extracellular and intracellular androgen binding components. Peptides, 1990 Jul-Aug, 11(4), 625 - 32 The osteogenic stimulating effect of neuroactive calcitonin gene-related peptide; Bernard GW et al.; Silverman and Kruger (Somatosens . Res . 5(2):157-175; 1987) reported that sensory nerve fibers of the dental pulp secrete calcitonin gene-related peptide (CGRP) . These are localized exactly where secondary or tertiary dentin calcification occurs . Recently we found that CGRP has an osteogenic stimulating effect by increasing the number and size of bone colonies in vitro . The purpose of this study is to test whether there is a relationship between the effects of different doses of CGRP and bone colony numbers and/or size . Rat CGRP in different dosages (0.4, 4 and 40 micrograms/ml in BGJb medium) was added daily to 3 million light density (LD) bone marrow white cells which were harvested from adult Sprague-Dawley rats with the Ficoll-Paque density gradient separation method, then seeded onto a previously prepared feeder layer of fibroblasts in Petri dishes . Seven days after adding CGRP, in the controls without CGRP there were 2 bone colonies; with 0.4 microgram of CGRP there were 4 colonies; with 4 micrograms of CGRP there were 6 colonies; with 40 micrograms there were 9 colonies, indicating there was a significant increase in number of bone colonies with an increase in dose of CGRP between individual groups, respectively (p less than 0.0005 and p less than 0.0001) . In another experiment, intravenous injection of 10 micrograms of rCGRP/kg body weight was performed two hours before surgery . LD bone marrow white cells were collected and seeded onto a feeder layer in Petri dishes exactly as described above.(ABSTRACT TRUNCATED AT 250 WORDS) Cardiovasc Res, 1990 Jul, 24(7), 555 - 9 Oxygen and extracellular fluid restriction in cultured heart cells: electron microscopy studies; Ne'eman Z et al.; STUDY OBJECTIVE--To evaluate the effects of "simulated ischaemia" on the structure of cultured heart cells . DESIGN--Cultured heart cells were subjected for 2 h either to anoxia or to anoxia with simultaneous extracellular volume restriction ("simulated ischaemia") . Cells maintained under normoxic conditions served as controls . The cells were then fixed in situ in Petri dishes with formaldehyde-glutaraldehyde . EXPERIMENTAL MATERIALS--Heart cells from one day old rats on day 5 in culture were used . MEASUREMENTS AND RESULTS--Electron microscope studies were carried out on control and injured cells . "Mildly ischaemic" cells featured raffled and invaginated cell surfaces, reduced matrix density, disorientated mitochondrial cristae due to swelling, and giant mitochondria . Dilatation of rough endoplasmic reticulum and electron dense membrane bound vesicles were observed in the cytoplasm . CONCLUSIONS--The model of simulated ischaemia is in keeping with the classical picture of irreversible cell damage caused by ischaemic injury. J Biolumin Chemilumin, 1990 Jul-Sep, 5(3), 171 - 7 Different influences of cytochalasin B on the activation of human neurophils settled onto petri dishes displayed by simultaneously detected native and luminol-dependent luminescence; Roschger P et al.; Cytochalasin B (CB) is known to interfere reversibly with the cytoplasmic contractile filamental network of mammalian cells . The role of the microfilament system in the mechanism of the reactive oxygen intermediates release of polymorphonuclear leukocytes (PMNL) was studied for different kinds of stimuli . PMNL from fresh human blood were treated with CB and stimulated by adherence on plastic surfaces, by opsonized zymosan, by phorbol myristate acetate and by N-formylmethionyl-phenylalaline . The production of reactive oxygen species were monitored by simultaneous detection of native, luminol-independent, luminescence (NL) and luminol-dependent luminescence (LDL) using a method of spectral discrimination . Different influences of CB on NL with respect to LDL as well stimuli-dependent influences of CB on the luminescence response of PMNL were observed . Especially phagocytosis-associated activation of PMNL was strongly inhibited by CB, whereas LDL was reduced to a much greater extent in comparison with NL . A firm involvement of the microfilament system is indicated, but it depends on the kind of stimulus engaged. Exp Neurol, 1990 Jul, 109(1), 111 - 30 Sulfated proteoglycans in astroglial barriers inhibit neurite outgrowth in vitro; Snow DM et al.; In vivo studies of the roof plate of the spinal cord and midline optic tectum in rodent and the developing subplate in the telencephalon of the chick showed that two glycosaminoglycans, keratin sulfate and chondroitin sulfate, possibly in the proteoglycan form (KS-PG, CS-PG, or KS/CS-PG), were present at times when axons approach closely but do not invade these territories . To address the question of whether KS/CS-PG actively inhibits growth cone elongation and to determine which component(s) of the proteoglycan may be critical to this phenomenon, we used a technique employing nitrocellulose-coated petri dishes onto which stripes of various purified macromolecules were attached . Isolated E9 chick dorsal root ganglia were grown on lanes of KS/CS-PG in alteration with lanes of the growth-promoting molecule laminin (LN) . Neurite outgrowth was abundant along stripes of LN . In contrast, upon encountering a stripe containing KS/CS-PG, neurites either stopped abruptly or turned and traveled along the KS/CS-PG stripe border . The effect was dependent upon the concentration of the proteoglycan with intermediate concentrations producing intermittent patterns of crossing . We mixed LN with the KS/CS-PG, where the LN was in concentrations which alone support outgrowth, and observed that the KS/CS-PG was still inhibitory when such a growth-promoting molecule was present . A 10-fold higher concentration of LN was able to overcome the inhibitory effect of the KS/CS-PG . These results suggest that the interaction of inhibitory and growth-promoting molecules can interact to produce a wide spectrum of neurite patterns ranging from complete inhibition to totally unimpeded outgrowth . Selective enzymatic removal of the KS or CS from the KS/CS-PG permitted various degrees of neurite outgrowth to occur across the previously inhibitory lanes, and digestion of both glycoaminoglycan moieties, leaving only the protein core of the molecule, resulted in a complete lack of inhibition . These assays demonstrated that KS/CS-PG is inhibitory to embryonic dorsal root ganglia neurites in vitro and that complete inhibition requires contributions from both KS and CS moieties. Int J Radiat Biol, 1990 Jul, 58(1), 133 - 44 Respiration-induced oxygen gradients in cultured mammalian cells; Fengler JJ et al.; The effect of cell respiration on the availability of intracellular oxygen was investigated by comparing the radiosensitivities of respiring and non-respiring cells over a range of oxygen tensions . Monolayers of Chinese hamster V79 fibroblasts on glass Petri dishes were irradiated at respiration-inhibiting (4 degrees C) and normal (37 degrees C) cell-culturing temperatures . Desired extracellular oxygen concentrations were achieved by aspirating the culture medium above the cells prior to irradiation, leaving a residual thin film which prevented drying of the cells while allowing rapid equilibration with the overlying gas . Measurement of clonogenic survival revealed that, at equivalent extracellular oxygen concentrations, the cells irradiated at 37 degrees C were less radiosensitive than those irradiated at 4 degrees C . The difference in respiring and non-respiring cell radiosensitivity was dependent on cell shape, and decreased when the cells attached to the Petri dish surface were allowed to assume a flatter configuration . These results imply that at low extracellular oxygen tensions the oxygenation of critical cellular radiation target(s) is dependent on respiration and diffusion distance, as would be expected if oxygen gradients induced by respiration exist within and immediately around actively metabolizing cells. J Leukoc Biol, 1990 Jul, 48(1), 74 - 80 Use of colloidal silica (Sepracell-MN) for enrichment of dendritic cells from human peripheral blood: comparison with other methods; Chehimi J et al.; Three methods are described for enrichment of dendritic cells from human peripheral blood . In method 1, mononuclear cells were incubated in plastic tissue culture flasks for two h . Nonadherent cells were removed . Adherent cells were washed to remove floating cells and incubated for 14 h at 37 degrees C in 5% CO2 . Carbonyl iron was added, and the flasks were incubated for another 2 h . Nonadherent cells were subjected to centrifugation over metrizamide gradient . Phagocytic cells containing ingested carbonyl iron, small lymphocytes, and free carbonyl iron particles passed through the metrizamide, while the interface cell population was enriched for dendritic cells . The purity and yield of enriched dendritic cells were 52.8% and 0.05%, respectively . In method 2, adherent mononuclear cells were cultured overnight, and the released cells (released adherent cells) were centrifuged over metrizamide to separate low-density cells . Monocytes from this low-density cell population were removed by panning over human gamma globulin-coated plastic Petri dishes . In this method the average purity and yield of DC were 63.8% and 0.1%, respectively . In method 3, released adherent cells were treated with anti-CD5 and anti-CD14 monoclonal antibodies plus baby rabbit complement for 15 min, washed, and centrifuged with colloidal silica (Sepracell-MN) . Centrifugation with Sepracell-MN was repeated three times . Low-density cells were panned twice over human gamma globulin-coated plastic Petri dishes . Nonadherent cells were highly enriched for DC . Contamination of T cells, B cells, and NK cells was undetectable by flow cytofluorometry . Contamination of monocytes was less than 2% . This method provided greater than 85.0% purity and 0.4% yield . This method (method 3) combines adherence, complement-dependent lysis, centrifugation with colloidal silica, and panning and provides the best yield and purity; it is therefore recommended for optimal purification of DC. J Parasitol, 1990 Jun, 76(3), 425 - 8 Efficacy of agar-plate culture in detection of Strongyloides stercoralis infection; Arakaki T et al.; Agar-plate culture of feces using a modified petri dish proved to be highly efficient in the detection of Strongyloides stercoralis infection . Furrows left by S . stercoralis on the agar plate were distinguished readily in size from those left by Necator americanus. Mol Cell Biol, 1990 Jun, 10(6), 2669 - 77 Biochemical characterization of the Drosophila dpp protein, a member of the transforming growth factor beta family of growth factors; Panganiban GE et al.; The decapentaplegic (dpp) gene of Drosophila melanogaster is required for pattern formation in the embryo and for viability of the epithelial cells in the imaginal disks . The dpp protein product predicted from the DNA sequence is similar to members of a family of growth factors that includes transforming growth factor beta (TGF-beta) . We have produced polyclonal antibodies to a recombinant dpp protein made in bacteria and used a metallothionein promoter to express a dpp cDNA in Drosophila S2 cells . Similar to other proteins in the TGF-beta family, the dpp protein produced by the Drosophila cells was proteolytically cleaved, and both portions of the protein were secreted from the cells . The amino-terminal 47-kilodalton (kDa) peptide was found in the medium and in the proteins adhering to the plastic petri dish . The carboxy-terminal peptide, the region with sequence similarity to the active ligand portion of TGF-beta, was found extracellularly as a 30-kDa homodimer . Most of the 30-kDa homodimer was in the S2 cell protein adsorbed onto the surface of the plastic dish . The dpp protein could be released into solution by increased salt concentration and nonionic detergent . Under these conditions, the amino-terminal and carboxy-terminal portions of dpp were not associated in a stable complex. Boll Soc Ital Biol Sper, 1990 May, 66(5), 471 - 8 {A model of venous thrombosis induced by stasis and Feiba in rabbits}; Tettamanti R et al.; A model of venous thrombosis, induced by injection of Feiba(R) (Factor Eighth Inhibitor Bypassing Activity) plus stasis, was studied in the jugular veins of anaesthetized rabbits . Right and left jugular vein segments were isolated by surgical technique for a 3 cm length, which included the bifurcation of the vessel, and left "in situ" . Feiba(R) was injected through a marginal ear vein at the dose of 5 U/Kg/0.2 ml; 20 and 25 sec later the contralateral and the homolateral jugular vein segments were ligated respectively . Complete stasis lasted 10 or 20 min, then the vessels were removed, placed in a saline filled Petri dish and visually graded for red thrombus formation on a scale of 0 to 10 . A correlation was found between severity of thrombosis and duration of stasis . This model appears to be suitable for testing heparin or heparin-like substances, in fact a linear correlation was found between log dose of the drug (injected i.v . 5 min before Feiba(R} and its antithrombotic effect for each duration of stasis tested . In particular the DE50 value calculated for 10 min stasis was 20.5 mcg/Kg i.v . The reproducibility of the model was good even with a small number of animals (n = 6) for each treatment group. Radiat Res, 1990 Apr, 122(1), 72 - 6 Induction of hypoxia in glass versus Permanox petri dishes; Zeman EM et al.; The survival of Chinese hamster ovary cells in culture following graded doses of X rays delivered under aerobic and hypoxic conditions, or treatment with the bioreductive drug SR 4233 under hypoxic conditions, was evaluated as a function of whether cells were plated onto glass or Permanox plastic petri dishes . In the case of treatment with SR 4233, the influence of varying the total volume of medium in the dishes was also studied . While the Permanox petri dishes were sufficient to yield "radiobiological" hypoxia, that is, oxygen enhancement ratios of approximately 3.0 were obtained for X irradiation, they were inferior to glass petri dishes with respect to the hypoxia-selective cytotoxicity of SR 4233 . For a 90-min hypoxic exposure to 40 microM SR 4233, the surviving fraction of cells plated on plastic dishes averaged about 50-fold higher than that of cells plated on glass dishes . Although varying the total medium volume did affect the extent of SR 4233-induced cytotoxicity for glass dishes--drug toxicity decreased slightly with increasing medium volume--this was not the case for the plastic dishes, in which the cell survival following a fixed SR 4233 exposure was essentially constant as a function of medium volume . These results suggest, at least for SR 4233, and under these experimental conditions, that Permanox petri dishes are not satisfactory for such studies. Diabetes Res Clin Pract, 1990 Apr, 9(1), 65 - 73 Extreme insulin resistance due to anti-insulin receptor antibodies: a direct demonstration of autoantibody secretion by peripheral lymphocytes; Di Paolo S et al.; A 59-year-old woman with systemic lupus erythematosus was found to have marked hyperglycemia, extreme insulin resistance and abnormally high plasma immunoreactive insulin . Her circulating erythrocytes displayed a dramatic decrease of 125I-labeled insulin binding . Both the whole serum and purified IgG fraction strongly inhibited the binding of radiolabeled insulin to control erythrocytes . These results suggested, although indirectly, the existence of antibodies to insulin receptors in the serum of the patient . To directly investigate this issue, we used an enzyme-linked solid-phase immunoassay which allows the detection and enumeration of lymphocytes secreting antibodies towards insulin receptors . Peroxidase-conjugated anti-human immunoglobulin is used to reveal the binding of antibodies to insulin receptor-coated dishes . We demonstrated that the patient's mononuclear cells, when briefly incubated in Petri dishes with partially purified insulin receptor, were able to secrete immunoglobulins of G class specifically directed to the antigen . Moreover, only a fraction of the whole population of anti-insulin receptor antibodies was directed towards the insulin binding region of the receptor, seemingly corresponding to the auto-antibodies detected with conventional binding-inhibition assay. Biomaterials, 1990 Mar, 11(2), 133 - 7 Screening of in vitro cytotoxicity by the adhesive film test; Srivastava S et al.; A novel procedure, the adhesive film test, is proposed as a screening method to predict potential cytotoxicity of biomaterials . This in vitro test utilizes a sterile strip of acrylate-based medical adhesive as an anchorage substrate in cytotoxicity studies . The adhesive film allows direct fixation of test samples to the base of the petri dish, ensuring close contact between sample and cells . The test is based on the principle that toxic components present in the test material will readily leach out into the culture medium and adversely affect the local cell population . The main advantage of the adhesive film test is that a viable cell population can be added directly to the test plate and after an incubation period of 24 h, the cellular response can be recorded as either cytotoxic or cytocompatible . Microscopic examination can be followed by quantifying the results using a micrometer to measure cellular attachment areas, migration distances and zones of inhibition . In addition, the adhesive film used to attach the test samples is shown to support extensive fibroblast growth and attachment to its surface and hence can also function as a negative nontoxic control in cytotoxicity studies. Neurol Res, 1990 Mar, 12(1), 41 - 4 Cerebrospinal fluid factors following subarachnoid haemorrhage accelerate collagen lattice contraction by fibroblasts; Smith RR et al.; Proliferation in the intimal layer and medial necrosis are the most consistent findings in the cerebral artery following subarachnoid haemorrhage (SAH) in man . Recently, SEM studies from our laboratory have also shown marked endothelial injury as demonstrated by a profuse platelet carpet . Myofibroblasts proliferate in response to the platelet derived growth factor (PDGF), and abundant collagen is present in the vessel wall . We have employed experiments using fibroblast-populated collagen lattices to study cerebrospinal fluid (CSF) from patients with recent SAH . Isolated rat tail collagen and cultured human dermal fibroblasts are mixed together, placed in 35 mm Petri dishes, and allowed to gel . CSF samples are placed on the surface of the collagen lattice, using 0.2 ml saline for control . The collagen lattices are then incubated and daily measurements recorded . We found that CSF samples from patients with recently ruptured aneurysms significantly accelerate contraction of the collagen lattice . The factor in CSF is heat stable and has a molecular weight of less than 6000. Cytokine, 1990 Mar, 2(2), 142 - 8 Leukocyte inhibitory factor activates human neutrophils and macrophages to release leukotriene B4 and thromboxanes; Conti P et al.; Recent evidence has proved that cytokines can stimulate the production of 5-lipoxygenase products . Leukotriene B4 (LTB4) is a major mediator of leukocyte activation in acute inflammatory reactions, which produce chemotaxis, lysosomal enzyme release, and cell aggregation . Leukocyte inhibitory factor (LIF) also causes biological responses related to inflammation, i.e., LIF directly induces specific granule secretion by polymorphonuclears (PMNs) and potentiates many formyl-methionyl-leucyl-phenylalanine (FMLPs) mediated responses . Since arachidonic acid products are important mediators of inflammation, we have studied the effects of LIF on the arachidonic acid cascade products LTB4 and thromboxane A2 (TxA2) . Resuspended at a final concentration of greater than 95% polymorphonuclear PMNs were isolated and tested with some cytokines on the release of LTB4 and TxA2 . Peripheral blood mononuclear cells were isolated and seeded in Petri dishes and incubated for 60 min . Adherent macrophages were used for the cytokine stimulation study . Both types of leukocytes were treated with LIF, interleukin 6 (IL 6), and granulocyte-monocyte colony stimulating factor (GM-CSF) at different concentrations, and test agents A23187 and FMLP . Radioimmunoassay for LTB4 and TxB2 was determined by the resulting supernatants . Treatment of PMNs and macrophages with LIF at different concentrations proved to generate significant increases in LTB4 and TxA2 production . This was compared with IL 6 and GM-CSF, which had no effects . In these experiments, TxA2 generations could not be attributed to platelet contamination of PMN suspensions . The quantity of platelet contamination was not sufficient to influence how much TxB2 was produced . The similarities of LIF to other arachidonate stimulating cytokines suggest a similar mode of action in producing hematologic changes typical of tissue injury.(ABSTRACT TRUNCATED AT 250 WORDS) Blood, 1990 Feb 15, 75(4), 874 - 80 Inhibition of receptor binding and neutralization of bioactivity by anti-erythropoietin monoclonal antibodies; D'Andrea AD et al.; We have generated four high affinity monoclonal antibodies (MoAbs) to recombinant human erythropoietin (EPO) . All four MoAbs immunoprecipitate radioiodinated native EPO, and the concentrations of MoAbs required for maximum binding range from 10 nmol/L to 100 nmol/L . Two MoAbs, designated Group I MoAbs, bind to an epitope within the N-terminal 20 amino acids of EPO and also immunoprecipitate sodium dodecyl sulfate (SDS)-denatured EPO . Two other MoAbs (Group II MoAbs) do not immunoprecipitate SDS-denatured EPO and do not bind to any of the eight endo C fragments of EPO . We first used murine erythroleukemia (MEL) cells to test the MoAbs for inhibition of EPO-receptor binding . MEL cells, although unresponsive to EPO, express 760 high affinity receptors for EPO per cell (Kd = 0.24 nmol/L) . To assay our MoAbs, MEL cells were grown as monolayers on fibronectin-coated Petri dishes and incubated at 4 degrees C with radioiodinated EPO . Group I MoAbs do not inhibit binding of radioiodinated EPO to the MEL EPO-receptor, but Group II MoAbs do inhibit binding in a dose-dependent manner . We next examined the neutralization of EPO bioactivity by our MoAbs, using EPO-dependent cell line . Only Group II MoAbs inhibit a newly developed EPO-dependent cell growth, demonstrating that inhibition of EPO-receptor binding correlates with neutralization of EPO bioactivity. Wien Klin Wochenschr, 1990 Feb 2, 102(3), 77 - 80 Blood rheology during induced ischaemia of the lower limbs; Ciuffetti G et al.; The filterability rate of whole blood, the red and white blood cell sub-populations, and the erythrocyte and leucocyte count variations were studied during exercise in 20 male non-diabetic smokers, all with Stage II peripheral vascular disease (PVD), and 20 matched controls . A controlled ischaemia was induced using treadmill exercise . Blood samples were taken at rest, at the onset of calf pain and at haemodynamic recovery from peak exercise . Leucocytes were counted, separated using Ficoll-Hypaque into their sub-populations by centrifugation, and adherence to Petri dishes, re-suspended in buffer and filtered through 5 micron pore diameter filters . Whole blood filterability and the leucocyte count were significantly increased at the onset of calf pain . A significant increase was observed in the filterability of the monocyte sub-fraction and this persisted throughout the recovery period. J Anim Sci, 1990 Feb, 68(2), 449 - 53 Incorporation and expression of a neo gene after transfer to caprine hematopoietic cells using a modified retrovirus; Casey DC et al.; The ability of the N2 retrovirus to introduce the selectable neo gene into (transform) caprine hematopoietic cells (CHC) was evaluated . Helper-free amphotropic retrovirus producing cells (RPC) were plated at approximately 1.0 x 10(5) cells per 25 cm2 tissue culture flask, cultured to 80% confluence and irradiated (1,500 rads) prior to CHC incubation . The CHC collected from three donor goats were washed and cultured for 24 h in either the RPC or control flasks . Cells were cultured in Dulbecco's Modified Eagles Medium containing 10% fetal calf serum (FCS) (DMEM) and 4 micrograms polybrene/ml . After 48 h of culture in fresh DMEM, cells were recovered and suspended in Iscove's DMEM supplemented with .3% gar, 12% FCS and 400 micrograms geneticin/ml (G418; neomycin) and transferred to 35-mm petri dishes (7.5 x 10(5) cells) for selection of G418 resistant cells . After 17 d of culture, plates were evaluated for total number of colony forming units (CFU, greater than 10 cells) . Total number of CFU was greater (P less than .01) in treatment samples (means = 175, SEM = 70) than in control cultures (mean = 0, SEM = 0) . The N2 retrovirus appears to be an effective vector for the transformation of CHC and may provide a means to introduce gene(s) into cells of domestic animals. Dev Biol, 1990 Feb, 137(2), 219 - 32 Central nervous system antigen P84 can serve as a substrate for neurite outgrowth; Chuang W et al.; Neurite outgrowth promoting properties of neural cell surface proteins can be assessed by immobilizing isolated membrane proteins on nitrocellulose-coated petri dishes . Using this method, we have identified a unique cell surface antigen, designated P84, as a new neural cell adhesion molecule . Immunoaffinity purified P84 contains three polypeptides with molecular weights of 167, 85, and 66 kDa . When spotted onto nitrocellulose-coated plates, P84 supports adhesion of mouse cerebellar neurons and neurite outgrowth . Glial cell attachment was also observed . Intact monoclonal antibodies directed against P84 inhibit adhesion and outgrowth on a P84 substrate . This antigen is found on the surfaces of neurons in cultures of cerebellar cells . It is also found on a subclass of unidentified flat cells . P84 is not found on oligodendrocytes or GFAP-positive astrocytes . As early as E9, P84 could be detected in the floor plate region of the spinal cord . This pattern persists throughout embryonic development . Postnatally, widespread expression of P84 is observed in a variety of CNS tissues. Gastroenterology, 1990 Feb, 98(2), 322 - 35 Basement membrane components are potent promoters of rat intestinal epithelial cell differentiation in vitro; Hahn U et al.; Basement membranes have been implicated in morphogenesis and cell differentiation . In this study, the effect of basement membrane components on intestinal epithelial cell maturation in a mesenchyme-free environment was investigated . Fetal rat small intestinal epithelial cells (from the 14th-17th day of gestation) were exposed to basement membrane-derived proteins (laminin, collagen type IV, and a complex basement membrane-enriched extract from the Engelbreth-Holm-Swarm sarcoma) and other extracellular matrix proteins (collagen type I and fibronectin) coated onto Petri dishes . The cells attached readily only to fibronectin and basement membrane proteins . For 5 days the developing epithelial colonies were monitored in vitro, assessing morphological and functional parameters of cell maturation . Colonies grown on laminin and the basement membrane extract were larger and of greater cell density . An increase in alkaline phosphatase and lactase activity was observed after 3-4 days in these colonies which could be enhanced to yield 90%-100% positive cells by the addition of dexamethasone to the medium while no sucrase-isomaltase activity was elicited . Electron microscopy confirmed a high degree of cellular polarization illustrated by tight junctions and apical microvilli in epithelial cells grown on a basement membrane-like support . In contrast, none of the other proteins stimulated the cells to mature in vitro . The authors conclude that certain basement membrane components actively promote fetal intestinal epithelial cell differentiation. Arch Dermatol, 1990 Feb, 126(2), 175 - 80 Characterization of cellular elements in healed cultured keratinocyte autografts used to cover burn wounds; Petersen MJ et al.; Biopsy specimens from unburned skin were obtained from three severely burned patients and placed into tissue culture . After 2 to 3 weeks, the cultured keratinocytes were released from the Petri dishes and transplanted onto the patient's burn wound, which had been completely excised down to muscle fascia, thereby removing all cutaneous elements . Healing cultured autografts were found to become repopulated with Langerhans cells within 3 to 6 weeks . A neodermis rich in fibronectin rapidly formed between the autografts and muscle fascia . However, using monoclonal antibodies to cytokeratins as markers of differentiation, we found that the autograft keratinocytes expressed an abnormal pattern of differentiation that was similar to the differentiation seen in hyperproliferative states such as psoriasis . In contrast, healed split-thickness graft donor sites and reepithelialized interstices of mesh grafts maintained the basal keratinocyte staining pattern of normal skin with the AE-1 monoclonal antibody. Biomater Artif Cells Artif Organs, 1990, 18(3), 437 - 46 The enhanced attachment and growth of endothelial cells on anhydrous ammonia gaseous plasma modified surfaces of polystyrene and poly(tetrafluoroethylene); Sipehia R; Anhydrous ammonia gaseous plasma technique was used for the surface modification of polystyrene petri dishes and poly(tetrafluoroethylene) (PTFE) membranes . Amino groups were added onto surfaces by exposing them to ammonia plasma . Plasma modified polymeric surfaces and control polymeric surfaces were seeded with bovine pulmonary artery endothelial cells (EC) . It was found that attachment of EC to control polystyrene surface was negligible . On the plasma modified polystyrene surface, there was improved attachment and growth of EC . At 96 hours, plasma modified surfaces yielded an order of 3 magnitudes more cells compared to those on control . Twenty four hours after seeding the cells, the percentage of EC attachment to control PTFE surfaces and modified surfaces were found to be about 36% and 92% respectively. Histochemistry, 1990, 93(4), 359 - 62 Towards microfluorometric quantitation of polyamines in situ . Relationship between cellular polyamine concentration and fluorescence yield of the formaldehyde fluorescamine method; Hougaard DM et al.; Two different fluorescence cytochemical methods, the formaldehyde-fluorescamine (FF) method and the orthophthalaldehyde (OPT) method as well as an immunocytochemical method have been developed for the localization of spermidine and spermine . Of these three methods, the FF-method is the most easy to perform . We have studied the relationship between fluorescence intensity induced by the FF-method and cellular polyamine levels measured by HPLC in MCF-7 cells and HeLa cells . The experiments were designed to obtain different cell concentrations of polyamines . Cells grown on microscope slides in Petri-dishes were partly depleted of spermidine by two days inhibition of their ornithine decarboxylase activity using alpha-difluoromethylornithine . One hr before harvest the cells were exposed to different concentrations (0-30 microM) of spermidine . Microfluorometric results and chemical determinations of spermidine and spermine were obtained from each separate slide . The cellular total polyamine (spermidine + spermine) concentration on the slides varied between 4 and 15 nmol per mg protein (MCF-7 cells) and 5 and 26 nmol per mg protein (HeLa cells) and the corresponding microfluorometric results between 60 and 115 arbitrary units (MCF-7 cells) and 80 and 160 arbitrary units (HeLa cells) . Simple regression analysis showed a good linear relationship between cellular polyamine concentration and FF-fluorescence yield . The correlation coefficient for MCF-7 cells was 0.86 and for HeLa cells 0.82, significance of the correlations was p less than or equal to 0.0001 . Our results add further credence to the specificity of the FF-method and indicate that the method may be useful for microfluorometric quantitation of polyamines in situ. In Vitro Cell Dev Biol, 1990 Jan, 26(1), 51 - 6 Cell proliferation on hydrogels; Nagaoka S et al.; The adhesion and proliferation of mammalian fibroblasts (Flow 7000) on the surface of hydrophilic (copolymer of N-vinyl-2-pyrrolidone and methyl methacrylate) and hydrophobic {polymethylmethacrylate (PMMA) stereocomplex} hydrogels with a wide range in water content were studied morphologically and quantitatively . It was demonstrated that cell proliferation on hydrogels by a static culture method decreased as the water content of the gels increased . However, it is remarkable that the cell proliferation on PMMA hydrogels with a high water content is equivalent to that on glass Petri dishes . The results obtained in the proliferation of cells on the surface of these hydrogels closely correspond to the state of cell adhesion . When fresh medium or air was perfused from the opposite side of the PMMA hydrogel membrane on which the cells were proliferating (perfusion method), the cells continued to grow into a higher density than with the conventional static culture method . In the case of fresh medium perfusion, the amount of proliferated cell was dependent on both the permeability of the membrane and the density of the membrane "scaffolding." Virus multiplication in the cultured cells increased in proportion to the cell density, whereas the cell function was similar in both culture methods. Tsitologiia, 1990, 32(8), 852 - 7 {Benzopyrene hydroxylase induction and the functioning of the immunocompetent cells in human peripheral blood}; Lozovatskii AL et al.; Xenobiotics--inducers of benzo(a)pyrene hydroxylase (BPH)--exert different effects on mitogen-stimulated and mitogen-unstimulated human peripheral blood mononuclear cells (PBC) . In mitogen-stimulated culture xenobiotics highly increase BPH activity and suppress cell blast transformation . The incubation of the unstimulated PBC in the presence of xenobiotics increases insignificantly BPH activity, intensifies T-cell differentiation and concanavalin A-induced proliferation . The BPH activity is mainly associated with the PBC adhered to plastic Petri dishes . However, the control and induced levels of BPH activity depend on the interaction between adhered and nonadhered cells. Growth Factors, 1990, 3(3), 181 - 90 Characterization of 5B12.1, a monoclonal antibody specific for IL-6; Sawamura M et al.; A monoclonal antibody (MAb) specific for interleukin-6 (IL-6) was generated by fusing SP2/0 cells with spleen cells from a mouse immunized with rat spleen cell derived plasmacytoma growth factor (rat PCT-GF) . This MAb inhibited the growth of an IL-6-sensitive murine plasmacytoma clone, MD90, in the presence of the immunogen, rat PCT-GF . More interesting, however, this MAb demonstrated species cross-reactivity by neutralizing murine (recombinant and P388D1 cell line-derived) and human (recombinant) IL-6 . IL-6 neutralization activity was also established in other IL-6 bioassays, such as the proliferation of spleen cells, plasmacytoma T1165, and a B-cell hybridoma 7TD1 . IL-6 neutralization was overcome partially by increasing the concentration of PCT-GF . The MAb had no effect on PCT-GF-independent plasmacytoma KI81 proliferation . Plastic petri dish-bound MAb removed rmIL-6 activity . These results suggest that this MAb specifically binds IL-6 and neutralizes bioactivity of various PCT-GF, rmIL-6, and rhIL-6. Acta Biol Hung, 1990, 41(4), 387 - 97 Limitations of the use of plant material grown in Petri dishes for physiological experiments; Tari I et al.; Roots of plants growing "aeroponically" (AP) on moistened filter paper in Petri dishes for a few days are fairly often used for physiological experiments (e.g . measurement of root growth), for ion or herbicide uptake tests, before the establishment of hydroponic or aseptic cultures although their hormonal status is markedly different from that of the hydroponic (HP) control . On the 4th day of germination the ethylene production of cucumber (Cucumis sativus L . cv . Budai csemege) roots growing in AP under controlled conditions increased considerably and exhibited a maximum curve, HP roots evolved ethylene much more constantly . The morphological changes in AP roots (e.g . inhibited elongation and swelling of primary roots, and increased formation of root hairs), resembling those caused by exogenously applied ethylene, can be prevented with 10(-5) M Ag+, an inhibitor of ethylene action . In roots of one-week-old AP seedlings, the amount of an acidic inhibitor, which as judged from the Rf values is likely to be abscisic acid (ABA), is about twice as high as in HP seedlings . An elevated ethylene or ABA level of AP roots may result in a reduced elongation of the primary roots . Counteraction of this inhibition by Ag+ suggests that the effect of ethylene is the primary event in the reduction of root length . When using plant material grown in Petri dishes the possibility of similar changes in hormonal status of the roots must be taken into consideration. J Lipid Mediat, 1990, 2 Suppl, S93 - 9 Modulation of the priming effects of platelet-activating factor on the release of interleukin-1 from lipopolysaccharide-stimulated rat spleen macrophages; Pignol B et al.; The pharmacologic modulation of the effect of platelet-activating factor (PAF) on the interleukin-1 (IL-1) activity present in the supernatants from lipopolysaccharide (LPS)-stimulated macrophages was investigated . Rat spleen macrophages were isolated by centrifugation on a Ficoll-Hypaque gradient followed by adherence on plastic petri dishes for 60 min at 37 degrees C under a 5% CO2/95% air atmosphere . The IL-1 content in the cell-free supernatants was assessed using the mouse thymocyte proliferation assay . Preincubation of macrophages for 10 min with 10 fM PAF prior to stimulation with 20 micrograms/ml LPS for 24 h markedly increased the IL-1 activity present in the supernatants from macrophages whereas no direct effect of PAF was noted . Although they had no direct effect, addition of L-651,392 (10 microM), a lipoxygenase inhibitor, or the oxygen-derived free radical scavenger mannitol (10 microM) during the 10-min preincubation period with PAF reversed by 105.0% and 79.9%, respectively, the action of the autacoid on IL-1 activity . Pertussis toxin (PT, 1 microgram/ml) decreased by 30% the LPS-induced IL-1 activity . Association of PT with PAF suppressed the enhancing effect of 10 fM PAF on the IL-1 activity present in the supernatants from LPS-stimulated macrophages . Thus, the enhancing effect of PAF on IL-1 release appears to be due to the production of lipoxygenase metabolites, leading to superoxide production and alterations of cAMP levels. Medicina (B Aires), 1990, 50(4), 356 - 60 {Rabies transmission to rodents after ingestion of naturally infected tissues}; Delpietro H et al.; We describe seven trials in which Calomys musculinus and Mus Musculus were fed with naturally rabies-infected tissues extracted from vampire bats, dogs, and bovines . The tissues were not subjected to any kind of previous laboratory handling and were administered directly in Petri dishes; rodents ate them voluntarily . The only infectious tissues were bovine brains taken from outbreaks transmitted by vampire bats (Table 1) . It was possible to infect the two species of tested rodents, and there was no relationship between infection and amount of virus ingested . From the total number of 132 animals that ingested different kinds of rabies-infected tissues, 3 died of rabies infection . From 128 survivors of all the exposed mice, 22 presented seroconversion to rabies . In the infected Calomys musculinus there was evident nervous symptomatology consisting in excitability, aggressiveness, paralysis and isolation of rabies virus from their salivary glands . The possibility that rodents become rabies infected by the ingestion of naturally infected tissues would indicate that they may constitute a reservoir for rabies because cadaver ingestion is a natural feeding source for many species . Furthermore they may permit the passage of rabies virus in carnivorous, animals since they are an important prey for them . The present observations indicate two situations which may increase rabies risks to man through rodent bites. Probl Tuberk, 1990, (9), 60 - 2 {Microscopic method of the determination of Mycobacterium tuberculosis sensitivity to chemotherapeutic agents}; Bil'ko IP; A microscopic method of testing M . tuberculosis sensitivity to chemotherapeutic drugs was developed and tried out . According to it, M . tuberculosis are inoculated and cultivated on agar-agar gel disks, put on a Petri dish with an egg medium and chemotherapeutic drugs . Following the incubation lasting for 5-7 days at 37 degrees C, agar-agar gel disks with inoculated material are removed from the medium and subjected to microscopic examination as to confirm or deny the growth of mycobacteria in the form of microcolonies . The developed method is simple and easy to use, and makes it possible to shorten the period of testing M . tuberculosis sensitivity to chemotherapeutic drugs up to 5-7 days since the examination is started. Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi, 1990, 8(3), 199 - 202 {Successful axenic cultivation of a local human strain of Giardia lamblia in suckling gerbil}; Lu SQ et al.; G . lamblia cysts isolated from the fresh feces of a Giardia-infected boy in Beijing rural area were inoculated into suckling gerbils (Meriones unguiculatus) . Trophozoites of G . lamblia obtained from the intestines of infected gerbils were cultivated in modified TYI-S-33 medium enriched with dehydrated bovine bile . The parasites grew luxuriantly and formed an intensive monolayer on the surface of the culture tube on day 14 after initial cultivation . The culture has been maintained for more than 12 months and more than 120 subcultures have been made . The growth curve of the organism showed that the peak growth of the trophozoites was attained at the 120th hour after seeding . The generation time was 15 +/- 2.0 hours . Periodic examinations of Giardia cultures for bacteria contamination, with Petri dishes of blood agar and beef broth, proved negative . After being cryopreserved in liquid nitrogen for 1 week or longer, the average viable rate of the organism was 65.7% and the resuscitated parasites grew luxuriantly in subcultures. Child Nephrol Urol, 1990, 10(4), 186 - 8 Significance of the monocyte crossmatch in living-related transplantation; Leichter HE et al.; It is claimed that antibodies to the vascular-endothelial (VEM) antigen system are responsible for early renal transplant rejection . To evaluate the significance of these anti-VEM antibodies, we performed a retrospective study in 16 patients aged 14.8 +/- 5 (SD) years, who received either a living-related HLA-identical (n = 5) or haplo-identical (n = 11) transplant between 4/79 and 3/84 . All patients had been transfused prior to transplantation . Lymphocyte and monocyte complement dependent cytotoxic crossmatches were performed using pre- and posttransplant recipients' sera and donor lymphocytes and monocytes . Lymphocytes were isolated with Ficoll-Hypaque; monocytes by adherence to plastic Petri dishes . Of the 16 patients studied, only 1 had a positive pre- and posttransplant monocyte crossmatch . Crossmatches using recipients' T and B lymphocytes were uniformly negative, indicating the presence of anti-VEM antibodies in the absence of anti-HLA antibodies in this 1 patient . This patient rejected the transplant in the immediate posttransplant period . Of the remaining 15 patients, 4 lost their kidneys within 16 days posttransplant, whereas 11 have good graft function . We conclude that anti-VEM antibody occurs rarely pretransplant and is an unusual cause of immediate rejection in the living-related transplant situation . However, when anti-VEM antibody is identified, transplantation should be avoided because of the likelihood of immediate early rejection. Pathobiology, 1990, 58(6), 323 - 8 Cloning of primary human tumors in capillary tube versus Petri dish: a head-to-head comparison; Peng XG et al.; Plating efficiencies (PEs) of primary human tumors cloned in a capillary assay system were compared to those derived from the conventional two-layer agar Petri dish assay system . A total of 143 consecutively received primary human tumors of 24 different pathologies were simultaneously tested in both assay systems . The successful clonal growth rate in the capillary assay was 82.7%, while in the Petri dish it was 64.7% (p less than 0.001) . The median PE was 0.017% in the capillary assay demonstrating a 4.25-fold increase over the 0.004% PE of the Petri dish system . The data confirmed previous results showing that cancer cells of ovarian, breast, and lung origins clone with higher PE in capillary tubes . In contrast, we confirmed that stomach carcinoma cells were the only tumor type that showed a higher PE with the Petri dish method . In addition, this study shows for the first time that lymphomas and renal cell carcinoma, when they survive in vitro, clone equally well in both methods . However, the capillary cloning method resulted in a 66% success rate for lymphoma cell cloning, but Petri dish cloning resulted in only a 33% success rate . Thus, for some types of cancers (i.e., lymphoma), capillary cloning may be advantageous because it improves the probability of obtaining evaluable results . In other cases, the advantage of capillary cloning may be only the decreased amount of specimen and reagents needed for the assay. J Egypt Soc Parasitol, 1989 Dec, 19(2 Suppl), 887 - 94 Dot-ELISA in diagnosis of schistosomiasis; Madwar MA et al.; One microliter of S . mansoni egg antigen was dotted directly on the nitrocellulose paper sheet acting as the adsorbent surface (9 dots/paper) . The sera of 25 Egyptian patients and 15 healthy persons (2 microliters of each) were dotted over the antigen dots, then 2 ml of each of the blocking, washing, HRP-conjugated IgG and DAB adding procedures, were added over the nitrocellulose paper in the petri-dish at room temperature . An intact brown circle (by naked-eye) indicates a positive in Dot-ELISA . There is an insignificant dot colour intensities in different clinical stages of S . mansoni infected Egyptians whereas, a direct relation was obtained between egg count and the colour intensity of the dots . The test had 100% sensitivity and 86% specificity thus it appears to be useful for both laboratory and field studies. Cancer Res, 1989 Dec 1, 49(23), 6738 - 44 High invasiveness associated with augmentation of motility in a fos-transferred highly metastatic rat 3Y1 cell line; Taniguchi S et al.; We previously reported that v-fos transfer to a src-transformed rat 3Y1 cell line enhanced lung metastasis . To clarify the mechanism of this enhancement, we compared various biological factors related to metastatic potential between a fos-transferred highly metastatic cell line (fos-SR-3Y1-202) and the control cell line transferred with genetic marker (pSV2-neo) plus pBR322, neo-SR-3Y1-200 . Lung arrest, the effect of lung extract on cell growth, or sensitivity to natural killer cells could not explain the higher metastasis of fos-SR-3Y1-202, compared to findings with neo-SR-3Y1-200 . The invasiveness, assessed by penetration through a Matrigel-coated filter was about 5 times higher in fos-SR-3Y1-202 than in neo-SR-3Y1-200; high invasiveness in vitro was also observed in a fos-transferred mixed-population cell line (fos-SR-3Y1-200) and fos-transferred highly metastatic clones . Histopathological evidence of an in vivo tumor also showed the high invasiveness of fos-SR-3Y1-202 cells . To elucidate the causes of the increased invasiveness of fos-SR-3Y1-202, attachment of the cells to Matrigel and its components, such as laminin and type IV collagen, type IV collagenase activity, and motility were examined . Attachment of the cells to the substrate coated on Petri dishes or the activity of type IV collagenase did not differ significantly . On the other hand, cell motility, determined by a new method to directly quantitate alteration of cell shape continuously, using video image analysis and computer techniques, was higher in fos-SR-3Y1-202 than in neo-SR-3Y1-200 . Thus, the fos-transferred cell line, fos-SR-3Y1-202 has a high invasiveness, in association with augmentation of motility, hence the enhancement of metastatic potential. Int J Radiat Biol, 1989 Dec, 56(6), 989 - 98 Cell density dependence of transformation frequencies in C3H10T1/2 cells exposed to X-rays; Bettega D et al.; The effects of cell density on transformation frequencies were studied in C3H10T1/2 cells exposed to 0.5 and 7 Gy of 200 kVp X-rays . Initial cell density strongly influenced transformation frequency; this decreased by a factor of between 4 and 10 when the initial seeding density was changed from 50 to 2500 cells/10 cm diameter Petri dish . The data were fitted with two equations: (a) an allometric fu