Indian J Med Res, 2000 Jan, 111, 11 - 3 Isolation of emetine resistant clones of Entamoeba histolytica by petri dish agar method; Prabhu R et al.; Emetine resistant clones of Entamoeba histolytica strain HM1:IMSS were isolated by using petri dish agar method after mutation with ethyl-methanesulphonate . Two emetine resistant clones were obtained and both were resistant to emetine at a concentration of 24 micrograms/ml of emetine . The 50 per cent inhibitory concentration (IC50) for both emetine sensitive and resistant clones was 5 and 14 micrograms/ml respectively . The colony forming efficiency of E . histolytica strain HM1:IMSS varied from 44 to 54 per cent . This method is useful for isolating clones from different strains of the parasite for molecular and immunological studies. J Cell Biochem, 2000 Apr, 77(4), 569 - 83 Epidermal growth factor induces cell cycle arrest and apoptosis of squamous carcinoma cells through reduction of cell adhesion; Cao L et al.; Most squamous epithelial cells are strictly anchorage-dependent cell types . We observed that epidermal growth factor (EGF) promoted the growth of A431 squamous carcinoma cells in suspension cultures but suppressed cell growth and induced apoptosis in monolayer cultures, suggesting that loss of adhesion is responsible for the effects observed in monolayer culture, before cell death . Consistent with this finding, we demonstrated that EGF reduced cell attachment, cell-cell interaction, and cell spreading . Treatment with EGF increased cell adhesion-regulated expression of p21 but suppressed expressions of cyclin A, D1, cdk2, and retinoblastoma protein (pRb), leading to cell cycle arrest and adhesion-regulated programmed cell death . To test directly whether promoting cell adhesion could reduce the effects of EGF, we grew cultures on plates coated with type II collagen . On these plates, cell adhesion was enhanced and EGF treatment had little effect on cell adhesion and apoptosis when cells were attached to the collagen . The collagen effects were dose dependent, and cell cycle and cell cycle-associated proteins were altered accordingly . Finally, when cultures were plated on bacterial Petri dishes, which completely disrupted cell attachment to substratum, the level of apoptosis was greatly higher and cell cycle was arrested as compared with monolayer cultures . Taken together, our results strongly suggest that the EGF-induced cell cycle arrest and apoptosis in monolayer cultures was the result of a decline in cell adhesion . Clin Orthop, 2000 Mar, (372), 272 - 9 Effect of microwave oven induced mild hyperthermia on bone viability and strength; Liebergall M et al.; Extracorporeal hyperthermia treatment of bone followed by reimplantation may be an option for treating bone tumors . However, intensive heat treatment, such as autoclaving, causes a decline of mechanical and biologic functions of bone tissue . In the current study, a microwave oven was used for minimal hyperthermic treatment, and it was found that complete eradication of all viable cells in rat bone could be achieved with minimal reduction in mechanical function . When the cells were evaluated histologically by special lactate dehydrogenase activity staining, complete bone cell death occurred after 60 seconds of heating in an empty Petri dish and after 30 seconds when heated in a Petri dish containing normal saline . Mechanical stiffness and strength of the bones, tested in three-point bending, showed no decrease after this heating . Microwave oven induced hyperthermia eradication of viable cells without significant damage to the mechanical properties may have clinical relevance in limb salvage tumor surgery. Theriogenology, 1999 Feb, 51(3), 637 - 46 Comparison of the longevity of motility of stallion spermatozoa incubated at 38 degrees C in different capacitating media and containers; Bedford SJ et al.; This study was designed to compare the effects of different media and containers on longevity of motility of spermatozoa during in vitro incubation at 38 degrees C in either air or 5% CO2 atmosphere . Three ejaculates were collected from each of 4 stallions . The media tested were skim milk-glucose, modified Krebs/Ringer and Hank's salts solution for incubation in an air atmosphere, and modified Krebs/Ringer and Brackett and Oliphant (BO) defined medium for incubation in a 5% CO2 atmosphere . All samples were incubated in 5-mL borosilicate glass tubes filled with 3 mL of extended spermatozoa, 5-mL borosilicate tubes filled with 6 mL (topped) of extended spermatozoa, 35-mm Petri dishes filled with 3 mL of extended spermatozoa, and 35-mm Petri dishes with 200-microL microdroplets of extended spermatozoa under sterile mineral oil . For all treatments, individual samples were removed at 2, 4, 6 and 12 h of incubation to determine the percentage of motile cells . Overall, spermatozoa incubated in Petri dishes in both 3-mL and microdroplet treatments had significantly higher motility than those incubated in glass tubes (P<0.01) . At 6 and 12 h of incubation in Petri dishes, progressive motility was significantly higher for spermatozoa extended in the Hank's salts solution than in the other media . Both the medium and container used significantly affected the longevity of motility of spermatozoa incubated at 38 degrees C. Theriogenology, 1999 Feb, 51(3), 519 - 29 Live birth of a bear cub following nonsurgical embryo collection; Boone WR et al.; In the near future, 6 of 8 bear species will face extinction mainly because of loss of their natural habitat . This loss of habitat will ultimately require some of these bears to be maintained in zoos and wildlife preserves in the hope of conserving genetic diversity . If the giant panda is representative of other bear species, reproductive performance will be inhibited in such an environment . In this study, we used the nonendangered American black bear (Ursus americanus) as the model for developing appropriate embryo transfer procedures . The donor bear mated numerous times between late May and early June . In late July we anesthetized her and used a series of telescoping sheaths to gain access to the uterus Then we passed a catheter through the largest sheath, inflated the balloon, and, using a 20-mL syringe, repeatedly infused into and then aspirated from the uterus PBS + BSA . We emptied the syringe into Petri dishes and observed 2 embryos . We rinsed the embryos, placed them in human tubal fluid + HSA + HEPES and then held them at 35 degrees C for 5 h . The recipient mated during mid-June; in late July we anesthetized her and, with the aid of laparoscopy, transferred an embryo into the cranial portion of the uterine horn ipsilateral to the ovary containing a CL . The recipient delivered 2 cubs in January . Necropsy results indicated that the neonates lived for 6 to 8 wk before succumbing to flooding in the den . The DNA from hair samples belonging to the neonates indicated that the male cub belonged to the donor, the female cub to the recipient . The delayed implantation mechanism in bears probably allowed for the successful development of the embryo in the presence of a substantial asynchrony between the donor and the recipient (13 d) . We conclude that embryo transfer is possible in the American black bear and can lead to the birth of live cubs. J Chromatogr A, 2000 Feb 18, 870(1-2), 405 - 11 Determination of catecholamines in pheochromocytoma cell (PC-12) culture medium by microdialysis-microbore liquid chromatography; Cheng FC et al.; An in vitro microdialysis system was constructed for the measurement of catecholamines in pheochromocytoma cell culture medium . The novel microdialysis device is composed of a petri dish, a dialysis membrane and two transmission tubes . The dialysis membrane is located in the space of a petri dish such that it is immersed in the culture medium . Catecholamines contained in the culture medium diffused into a designed dialysis membrane with sufficient recovery (about 60%) . Dialysates were collected by a sampling loop and introduced by an on-line injector to a microbore liquid chromatographic system for analysis of catecholamines . This assay yielded a detection limit of 0.2-0.5 pg/injection with acceptable intra- and inter-assay reproducibilities in 5 microl of dialysates . To evaluate the on-line microdialysis system, PC-12 cells were cultured in a petri dish within an incubator . The baseline concentration of dopamine in PC-12 cell culture medium was about 0.29 ng/ml which was elevated to 2.43 ng/ml after treatment with 0.5 mM potassium cyanide . In conclusion, the present microassay provides for the sensitive, direct measurement of catecholamines in culture medium while minimizing pretreatment procedures for sample preparation. Int J Radiat Biol, 2000 Feb, 76(2), 169 - 76 Dose-response relationship for radiation-induced mutations at micro- and minisatellite loci in human somatic cells in culture; Boyd M et al.; PURPOSE: The study was designed to determine the dose-response relationship for radiation induction of mutations at mini- and microsatellite loci in human somatic cells . Mutations induced by graded doses of gamma-irradiation were quantified by screening clones derived from single irradiated cells for micro- and minisatellite alterations following irradiation with 1, 2 or 3 Gy . MATERIALS AND METHODS: After irradiation, the moderately radioresistant glioma cell line UVW was seeded at low density into Petri dishes to allow formation of discrete colonies, 100 of which were examined at each dose . All the cells within a colony were presumed to have arisen from a single irradiated cell . Radiation-induced microsatellite alterations were determined at 16 different loci, by PCR amplification and visualization on polyacrylamide gels . Minisatellite alterations were identified at four different minisatellite loci by restriction enzyme digestion and Southern blotting . RESULTS: A dose-response curve for mutation frequency was obtained by analysis of 100 clones, yielding a minisatellite mutation rate of 5.5x10(-3) mutations/locus/Gy/cell and a microsatellite mutation rate of 8.75x10(-4) mutations/locus/ Gy/cell . At microsatellite loci, alterations were predominantly simple loss or gain of repeat units and loss of heterozygosity (LOH) . The mutations in minisatellite loci resulted predominantly in LOH and variation in repeat number . The background instability at each locus was determined by analysis of non-irradiated clones . Only 2% and 1% of the micro-and minisatellite loci respectively showed altered bands . CONCLUSIONS: This is the first report of a dose-response relationship for radiation-induced micro- and minisatellite mutations in human somatic cells . Described is a sensitive method for analysis of low-dose radiation mutagenesis in somatic cells that may prove to be a useful tool for radiation protection and dosimetry. J Epidemiol, 1999 Dec, 9(6 Suppl), S66 - 71 Somatic cell mutation induced by sunlight in Drosophila; Negishi T et al.; There is ample epidemiological evidence showing that sunlight can cause skin cancer in the human . In experimental studies, simulated sunlight or UV lamps are used for demonstrating carcinogenesis and other biological effects . Little studies, however, have been performed using natural sunlight itself . In this work, we have examined the mutagenicity of natural sunlight in Drosophila . The Drosophila wing spot test is useful to detect somatic cell mutations . Third instar larvae in petri dishes were exposed to sunlight (ultraviolet region with < 290 nm wavelength cut off by a plastic cover) in the yard of Okayama University campus (north latitude: 34 degrees 39', east longitude: 133 degrees 55') . The sunlight was mutagenic in Drosophila larvae and produced pyrimidine dimers in their DNA . In the observed mutagenicity, there was dependence on the exposure period and UV fluence . During the two-year monitoring, the highest induction of mutant spot observed was 1.98 total spots/wing on June 25, 1998, and the lowest was 0.64 on December 29, 1998, while non-exposure spontaneous spots were 0.29 and 0.32 on these days, respectively . Thus, solar radiation was mutagenic both in summer and in winter. Microvasc Res, 2000 Mar, 59(2), 221 - 32 Generational analysis reveals that TGF-beta1 inhibits the rate of angiogenesis in vivo by selective decrease in the number of new vessels; Parsons-Wingerter P et al.; Quantitative analysis of vascular generational branching demonstrated that transforming growth factor-beta1 (TGF-beta1), a multifunctional cytokine and angiogenic regulator, strongly inhibited angiogenesis in the arterial tree of the developing quail chorioallantoic membrane (CAM) by inhibition of the normal increase in the number of new, small vessels . The cytokine was applied uniformly in solution at embryonic day 7 (E7) to the CAMs of quail embryos cultured in petri dishes . After 24 h the rate of arterial growth was inhibited by as much as 105% as a function of increasing TGF-beta1 concentration . Inhibition of the rate of angiogenesis in the arterial tree by TGF-beta1 relative to controls was measured in digital images by three well-correlated, computerized methods . The first computerized method, direct measurement by the computer code VESGEN of vascular morphological parameters according to branching generations G(1) through G(>/=5), revealed that TGF-beta1 selectively inhibited the increase in the number density of small vessels, N(v>/=5) (382 +/- 85 cm(-2) for specimens treated with 1 microg TGF-beta1/CAM for 24 h, compared to 583 +/- 99 cm(-2) for controls), but did not significantly affect other parameters such as average vessel length or vessel diameter . The second and third methods, the fractal dimension (D(f)) and grid intersection (rho(v)), are statistical descriptors of spatial pattern and density . According to D(f) and rho(v), arterial density increased in control specimens from 1.382 +/- 0.007 and 662 +/- 52 cm(-2) at E7 (0 h) to 1.439 +/- 0.013 and 884 +/- 55 cm(-2) at E8 (24 h), compared to 1 . 379 +/- 0.039 and 650 +/- 111 cm(-2) for specimens treated with 1 microg TGF-beta1/CAM for 24 h . TGF-beta1 therefore regulates vascular pattern and the rate of angiogenesis in a unique "fingerprint" manner, as do other major angiogenic regulators that include VEGF, FGF-2 (bFGF), and angiostatin . TGF-beta1 did not stimulate angiogenesis significantly at low cytokine concentrations, which suggests that this quail CAM model of angiogenesis is not associated with an inflammatory response . J Clin Periodontol, 2000 Jan, 27(1), 69 - 73 Leukocyte functions in 2 cases of Papillon-Lefèvre syndrome; Liu R et al.; AIM: To investigate the role of leukocytes in the pathogenesis of Papillon-Lefevre syndrome (PLS) . METHODS: Peripheral blood polymorphonuclear neutrophils (PMNs), monocytes (MNs) and gingival crevicular fluid (GCF) were obtained from 2 cases of PLS with typical features . The chemotaxis of PMNs and MNs were evaluated using a modified Boyden chamber . The adherence of PMNs was determined by adherence of PMNs to petri dishes . Interleukin-8 (IL-8) in GCF was detected by sandwich ELISA . Elastase activity in GCF was measured with a low molecular weight substrate (S-2484) specific for granulocyte elastase . RESULTS: PMNs from both patients showed depressed chemotactic response to FMLP and IL-8 . Total amounts of IL-8 in GCF from the 2 patients were much higher than those of the normal controls . Elastase activity was not significantly different from that of the controls . The adherence of PMN and the chemotaxis of MN in the 2 patients were normal . CONCLUSION: The depressed chemotactic response of PMN leads to decreased recruitment of PMN and/or release of lysozyme from PMN in the diseased gingival tissue, increasing the susceptibility of PLS patients to periodontal infection. Plant J, 1999 Dec, 20(5), 581 - 90 Sucrose availability on the aerial part of the plant promotes morphogenesis and flowering of Arabidopsis in the dark; Roldan M et al.; Conditions to promote dark morphogenesis and flower-ing in Arabidopsis have previously been limited to liquid cultures and to a few laboratory ecotypes . We have obtained development and flowering of Arabidopsis plants under complete darkness by growing them on vertical Petri dishes containing solid agar medium with sucrose . Under these conditions, all the ecotypes tested were able to develop, giving rise to etiolated plants that flowered after producing a certain number of leaves . Dark-grown plants showed similarities with phytochrome-deficient mutants and were different from de-etiolated or constitutive photomorphogenesis mutants such as det and cop . Late- and early-flowering ecotypes, showing large differences in flowering time and leaf number under long days, flowered with a similar number of leaves when grown in the dark . Rapid dark flowering of late-flowering ecotypes was not an effect of darkness but the result of the interaction between dark and sucrose availability at the aerial part of the plant, since sucrose also had an effect when plants were grown in the light . Gibberellin-deficient and insensitive mutants were delayed in the initiation of flowers in the dark, indicating a role for these hormones in flowering promotion in the dark . The late-flowering phenotype of mutants at different loci of the autonomous and long-day-dependent flowering induction pathways was rescued in dark growth conditions . However, the late-flowering phenotype of ft and fwa mutants was not rescued by sucrose either in the dark or in the light, suggesting a different role for these genes in flowering induction. Hum Gene Ther, 2000 Jan 1, 11(1), 43 - 51 High-efficiency retroviral transduction of mammalian cells on positively charged surfaces; Hennemann B et al.; The efficiency of retroviruses as transducing agents has been appreciated for many years, particularly for hematopoietic cell targets for which alternative strategies applicable to adherent cells are not effective . Advances in vector design, pseudotyping, and infection conditions have eliminated the need to cocultivate the target cells with virus-producing cells . Nevertheless, improvements are still needed for many applications, including those with a therapeutic or clinical cell-tracking objective . In this study we show that more positively charged surfaces, including those designed for the culture of anchorage-dependent cells, allow measurable levels of adhesion by different pseudotypes of retroviruses, which can result in increased gene transfer efficiencies to a variety of target cells including normal primary human hematopoietic cells as well as human leukemic cell lines and rat and murine fibroblasts . In the experiments with primary human cells, equal aliquots of enriched CD34+ cord blood cells were first stimulated for 2 days with cytokines (Flt3 ligand, Steel factor, IL-3, IL-6, and G-CSF) and then exposed for 4 days to a green fluorescent protein (GFP)- and Neo(r)-encoding retrovirus produced in PG13 cells . Both the final yield (approximately 300% relative to initial numbers), and the proportion (approximately 60%) of transduced CD34+ cells, colony-forming cells, and long-term culture-initiating cells were the same for cells infected either in tissue culture dishes or in fibronectin-coated petri dishes . Similar proportions (approximately 10%) and absolute yields of GFP+ human cells were also found in multilineage engrafted NOD/SCID mice assessed 6 to 8 weeks after being transplanted with these two types of transduced, but unselected, cells . These findings suggest a new and simpler approach for achieving high gene transfer efficiencies to hematopoietic cells. Free Radic Biol Med, 1999 Dec, 27(11-12), 1219 - 27 Yeast superoxide dismutase mutants reveal a pro-oxidant action of weak organic acid food preservatives; Piper PW; Saccharomyces cerevisiae could provide a simple experimental system for testing the antioxidant or pro-oxidant actions of chemicals, because it has the capacity for aerobic and anaerobic growth and can readily lose its mitochondrial electron transport chain (the major endogenous source of reactive oxygen species {ROS}) . This study showed that yeast superoxide dismutase mutants, in a simple petri dish test, readily distinguish a compound that enhances the detrimental effects of endogenous ROS production by the mitochondrial respiratory chain from another chemical that generates oxidative stress by redox cycling . Using this system, weak organic acid food preservatives are shown to exert a strong pro-oxidant action on aerobic yeast cells . In addition these acids are mutagenic toward the yeast mitochondrial genome, even at levels that are subinhibitory to growth . This raises the concern that the large-scale consumption of these preservatives in the human diet may generate oxidative stress within the epithelia of the gastrointestinal tract. J Periodontal Res, 1999 Aug, 34(6), 323 - 8 Graft of autologous fibroblasts in gingival tissue in vivo after culture in vitro . Preliminary study on rats; Simain-Sato F et al.; Several grafting techniques and guided tissue regeneration techniques (GTR) have been well-developed in periodontal surgery . However, these techniques could induce pain and side effects, such as a gingival recession during the healing period following the therapy . The graft of a small autologous connective tissue, using non-invasive surgical techniques could yield several benefits for the patients . Our preliminary study explores the feasibility of collecting healthy gingival tissues, culturing them in vitro to amplify rat gingival fibroblasts (RGF) and inoculating the obtained cells into autologous rat gingival tissues in vivo . Gingival tissues samples were cultured as explants as described by Freshney et al . and Adolphe . Confluent cells surrounding explants were detached after 7 d of culture from Petri dishes using 0.05% trypsin and designated "first transferred cells" (T1) . At the third passage (T3), cells cultured as monolayer were either examined under microscopy--phase contrast, scanning, or transmission electron--or numerated after trypan blue exclusion test . Autologous RGF labelled with fluorochrome were inoculated at the vestibular and palatine site of gingival tissue close to the superior incisors . In this preliminary study, 12 Wistar rats were used; for each, 2 biopsies were dissected and fixed for phase contrast or fluorescence microscopy . On d 1, 3 and 7 after injection in rat gingival tissues, fluorochrome-labelled cells could be detected in all these. Pancreas, 2000 Jan, 20(1), 67 - 76 Growth and function of isolated canine pancreatic ductal cells; Zhang M et al.; These studies investigated the growth characteristics and functional properties of isolated canine pancreatic ductal epithelial cells . Cells were isolated from the accessory pancreatic duct and cultured by using three conditions: on vitrogen-coated petri dishes with fibroblast conditioned medium (nonpolarized); in vitrogen-coated Transwells above a fibroblast feeder layer (polarized); or as organotypic rafts above a fibroblast-embedded collagen layer (polarized) . Growth characteristics, transepithelial resistances, and carbonic anhydrase and cyclic adenosine monophosphate (AMP) responses were evaluated . Under polarized conditions, the cells grew as monolayers with columnar epithelial characteristics . The monolayers developed high transepithelial resistance and became impervious to the passage of horseradish peroxidase . Epithelial growth factor (EGF) (2 ng/ml) stimulated ductal cell growth and accelerated the formation of a high-resistance monolayer . Forskolin (10 microM) rapidly decreased transepithelial resistance . Carbonic anhydrase activity, which was lower in nonpolarized compared with polarized conditions, was stimulated by carbachol (175 microM) . Secretin, however, did not stimulate carbonic anhydrase activity in these cells . Although secretin stimulated adenylyl cyclase activity in early-passage cells, this response was lost in later-passage cells . Both vasoactive intestinal polypeptide (VIP; 1 microM) and forskolin (10 microM) consistently increased adenylyl cyclase activity . Isolated canine pancreatic ductal epithelial cells proliferate in vitro, develop high-resistance epithelial monolayers, and respond to stimuli that activate adenylyl cyclase . These cells should provide a useful model for regulatory studies of ductal cell functions. J Med Entomol, 1999 Nov, 36(6), 903 - 5 Baseline susceptibility of a laboratory strain of Pediculus humanus humanus (Anoplura: Pediculidae) using a modified World Health Organization testing protocol; Zeichner BC; The World Health Organization (WHO) protocol for determining resistance in body lice, Pediculus humanus humanus (L.), requires holding lice for long periods, which makes successful execution of the test difficult in field settings . The purpose of this study was to modify the WHO test procedure to make the holding period of lice shorter and the handling of lice easier . Susceptible lice from a laboratory colony were placed in a petri dish containing a paper that had been treated with an insecticide solution . After 6 h, the petri dish was turned on its side and lightly tapped on the table . Lice that were unable to cling to the paper were counted as knocked down . The KD50 in mg (AI)/ml of the insecticide solution used to treat the papers was as follows: lindane 0.060, permethrin 0.115, d-phenothrin 0.554, and malathion 1.008 . If the diagnostic dose is set at 2 times the KD99, for this test procedure the diagnostic doses and WHO equivalent dose would be lindane, 0.368 mg (AI)/ml (WHO 0.132%); permethrin, 0.498 mg (AI)/ml (WHO 0.206%); d-phenothrin, 2.680 mg (AI)/ml (WHO 1.107%); and malathion, 5.212 mg (AI)/ml (WHO 2.020%). Ann Acad Med Singapore, 1999 May, 28(3), 409 - 16 Cancer gene therapy--fantasy or foresight? Kong HL. Gene therapy is an exciting new method of treatment that may have far-reaching implications on the way we manage diseases in the future . Cancer has become the principal focus of this futuristic research . The breathtaking pace of gene discovery in the last two decades, coupled with the birth of recombinant DNA technology, gave rise to the concept that genes may be manipulated and used as drugs . Genetic modification of cells can be carried out in petri dishes (ex vivo) or within the living system (in vivo) . In order for the therapeutic genes to exert their effect, they have to be transported into the cell nucleus where transcription takes place . Liposomes and genetically modified viruses have been extensively used as gene vectors . The ideal vector remains elusive . It would be one that can achieve tumour-specific, sustained, and regulatable gene expression without host toxicity . As a result of the past decade of intense gene therapy research, we have learned that it is a rational scientific concept that works remarkably well in petri dishes and in laboratory animals . However, early clinical gene therapy experimentations paled in comparison . This apparent disparity between dramatic preclinical successes and the very modest clinical results of gene therapy does not in anyway nullify the concept of gene therapy . Instead, it exposes the folly of underestimating the technical complexity of gene manipulation in human diseases . Fortunately, technical hurdles such as those confronting gene therapy today are not insurmountable; they need, however, much ingenuity, resolution and time to be overcome . It is reassuring that recent advances in gene therapy provide abundant evidence that the premature infant, born of unrealistic pressure, is indeed healthy and thriving . With proper nurturing and patience, there is no doubt that, in time, it will bear fruit. Zhonghua Yi Xue Za Zhi (Taipei), 1999 Oct, 62(10), 710 - 6 Effect of oncostatin-M on proliferation and activity in osteoblastic MC3T3-E1 cells; Shih C et al.; BACKGROUND: Osteogenic cells (osteoprogenitor cells and osteoblasts) respond to specific cytokines, growth factors and hormones . Understanding which cytokines affect osteogenic cells and the consequences of those effects are central to understanding normal and pathologic bone remodeling . Oncostatin-M (OSM) is a glycoprotein interleukin-6 cytokine known to inhibit bone resorption in fetal long bone cultures . However, it is still unclear whether OSM affects bone formation . The aim of this study was to investigate the effects of OSM on bone formation (bone cell proliferation, differentiation and function) . METHODS: For the in vitro bone formation bioassay, MC3T3-E1 cells were plated into 35 mm Petri dishes and cultured in Dulbecco's modified eagle medium (DMEM) . Various concentrations (1, 10, 100 or 1,000 units/ml) of OSM were added daily to the experimental dishes, while only DMEM was added in the control dishes during the proliferative and differentiated stages of MC3T3-E1 cell growth . The effect of OSM on bone formation was evaluated using 3H-thymidine incorporation, alkaline phosphatase expression, type I collagen synthesis assay and phase-contrast light microscopy . RESULTS: OSM significantly decreased osteoprogenitor cell proliferation and alkaline phosphatase activity in a dose-related fashion . There was no effect on type I collagen synthesis noted at any OSM dose . Thus, OSM exerted significant inhibition on bone formation . CONCLUSIONS: OSM is a potent inhibitor of bone formation by decreasing both osteoprogenitor cell proliferation and alkaline phosphatase activity. Clin Oral Investig, 1999 Mar, 3(1), 25 - 9 Cytopathologic effects of arecoline on human gingival fibroblasts in vitro; Chang YC et al.; Arecoline, a major betel nut alkaloid, has been detected in saliva obtained during betel nut chewing in concentrations up to 140 micrograms/ml, corresponding to 0.9 mM . Arecoline in the millimolar concentration range might participate in the initiation and/or progression of periodontal disease during the long-term effects of betel nut chewing . In this study, cell growth, cell proliferation, assessment of cytoplasmic enzyme lactate dehydrogenase (LDH) and collagen synthesis were used to investigate the effects of human gingival fibroblasts exposed to arecoline levels of 0-200 micrograms/ml . Control culture exhibited a normal monolayer of long spindle-shaped fibroblast morphology . Arecoline-treated human gingival fibroblasts showed a more rounded appearance and detached at the higher concentrations . At concentrations higher than 75 micrograms/ml, many cells had detached from the surface of the petri dish and numerous floating cells could be seen under the inverted microscope . At a concentrations higher than 25 micrograms/ml, arecoline inhibited cell growth, proliferation and collagen synthesis and increased LDH leakage in a dose-dependent manner (P < 0.05) . These results indicate that arecoline is a cytotoxic agent to human gingival fibroblasts . Repeated and long-term exposure to arecoline could impair gingival fibroblast function . Betel quid chewers might be more susceptible to destruction of the periodontium and less responsive to a regeneration procedures during periodontal therapy. J Eur Acad Dermatol Venereol, 1999 Sep, 12 Suppl 1, S10 - 6; discussion S17 Itraconazole and terbinafine in perspective: from petri dish to patient; De Doncker P; OBJECTIVE: To compare the antifungal activity of itraconazole and terbinafine in vitro and to relate them to their experimental in vivo activity and to their efficacy in patients with superficial fungal infections (tinea pedis and onychomycosis) . RESULTS: Fungal infections such as onychomycosis and tinea pedis are often treated with oral antifungals . With the introduction of newer agents such as terbinafine and itraconazole, efficacy and safety have been improved . In vitro evaluation showed somewhat better results against dermatophytes for terbinafine than for itraconazole, but in vivo results were at least equivalent . Moreover, itraconazole is a broad-spectrum agent with higher cure rates for infections other than dermatophytosis (e.g . for Candida infections) than terbinafine, according to ex vivo studies . A review of all published clinical trials, comparing the efficacy and safety of terbinafine and itraconazole in a meta-analysis revealed similar and high cure rates (>70%) for both antifungal agents and similar adverse event profiles . Both treatments were safe and well tolerated . CONCLUSIONS: Antifungal research has responded to the challenges of treating superficial infections by developing effective, well-tolerated, fast-acting antifungal therapies . The reduction in treatment duration has also led to improved patient's compliance . The most noticeable difference between itraconazole and terbinafine is the 1-week pulse concept of itraconazole in contrast to the continuous treatment concept of terbinafine. Rev Argent Microbiol, 1999 Jul-Sep, 31(3), 120 - 6 {Effect of calcium and light on the development of Iodophanus carneus}; Diorio LA; Ca2+ affects the mycelial morphological characteristics in I . carneus, modulating the development and conditioning the sexual differentiation . In Ca2+ 0 mM there was an interaction with light and development, in darkness the colonies did not reach the edge of Petri dishes and spiral growth was observed . In darkness the number of hyphae/mm was greater than those grown in light, except in Ca2+ 1 mM, at 28 mm of inoculum, and Ca2+ 2 mM, at 16 and 28 mm, where the number of hyphae/mm was greater . In cords and apothecial number and location a similar behavior was observed, evidencing a direct relation between these, Ca2+ concentration and light. Postgrad Med, 1999 Jul, Spec No, 6 - 11 Itraconazole and terbinafine in perspective . From petri dish to patient; De Doncker P et al.; This paper compares the antifungal activity and efficacy of itraconazole and terbinafine in vitro with their experimental activity and efficacy in vivo in patients with superficial fungal infections (tinea pedis and onychomycosis) . Onychomycosis and tinea pedis are often treated with oral antifungals . With the introduction of newer agents, such as terbinafine and itraconazole, efficacy and safety have been improved . In vitro evaluation showed somewhat better results for terbinafine over itraconazole against dermatophytes, but in vivo results were at least equivalent . Moreover, according to ex vivo studies, itraconazole is a broad-spectrum agent with higher cure rates than terbinafine for fungal infections other than dermatophytosis (e.g., Candida infections) . A meta-analysis of data from all published clinical trials comparing the efficacy and safety of terbinafine and itraconazole revealed similar high cure rates (> 70%) for both antifungal agents and similar adverse-event profiles . Both treatments were safe and well tolerated . Antifungal research has responded to the challenges of treating superficial infections by developing effective, well-tolerated, fast-acting antifungal therapies . The reduction in treatment duration also has led to improved patient compliance . The most notable difference between itraconazole and terbinafine is the 1-week pulse regimen available with itraconazole as opposed to the continuous treatment course available for terbinafine. Gene Ther, 1999 Sep, 6(9), 1617 - 25 Endothelial cell DNA transfer and expression using petri dish electroporation and the nonreplicating vaccinia virus/T7 RNA polymerase hybrid system; Lewis E et al.; The nonreplicating vaccinia virus MVA/T7 RNA polymerase hybrid system was tested with Petri dish electroporation for ectopic gene expression in human umbilical vein endothelial cells (HUVECs) . A range of voltages (150-450 V), pulse times (10-40 ms), DNA concentrations (0-20 microg/ml) and infection levels (0-15 multiplicities of infection) were tested for effects on T7 promoter-directed chloramphenicol acetyltransferase (CAT) activity after MVA/T7RP infection . MVA/T7RP-directed expression was transient and at least 10 000-fold in excess of nonviral, cytomegalovirus enhancer-directed expression . Use of a Petri dish electrode with the MVA/T7RP system showed increased viability compared with a cuvette electrode . Overexpression of interleukin-2 alpha subunit (IL2Ralpha) pro- tein followed by anti-IL2Ralpha-directed magnetic immunoaffinity cell sorting allowed isolation of the transfected population . The high fidelity of cellular sorting was shown by segregation of CAT activity in the IL2Ralpha-sorted population after transfection of T7 promoter-directed bicistronic IL2Ralpha/CAT DNA . Expression of a panel of proteins including the fluorophore green fluorescent protein as detected by fluorescence microscopy and p21cip1, p27kip1, pp60c-src, FGF-1, pRb, p107 and pRb2/p130 proteins was also achieved . Thus, use of the nonreplicating vaccinia virus/T7 RNA polymerase expression system with Petri dish electroporation is feasible for certain applications for the manipulation of HUVECs by gene transfer. In Vitro Cell Dev Biol Anim, 1999 Feb, 35(2), 87 - 93 Bio-stretch, a computerized cell strain apparatus for three-dimensional organotypic cultures; Liu M et al.; In the present study, a unique mechanical strain apparatus for three-dimensional organotypic cultures was developed into a computerized system . It consists of a personal computer running Windows-based software, the Bio-Stretch Manager, a Bio-Stretch Controller, and three sets of magnet boards . Cells are cultured on a Gelfoam sponge that is placed in a 35 mm petri dish with one end glued to the dish, and the other end attached to a coated steel bar . The petri dish is placed in front of a magnet, and the movement of the steel bar is controlled by dynamically changing the magnetic field . Up to five stretch patterns of variable frequency, duty cycle, and magnitude can be designed for each stretch regimen . Three different stretch regimens can be tested simultaneously . The operational characteristics of sponges were examined . Attachment of cells to the sponges was observed on several cell types . These features provide wide options for using this system to study the effects of mechanical stretch on cells. Boll Chim Farm, 1999 Jun, 138(6), 243 - 8 Study of accelerated storage conditions affecting physical characteristics, in-vitro dissolution and stability of bioadhesive containing tablets Hosny EA. The effect of storage at different temperatures and relative humidities on directly compressed tablets of indometacin containing 20% polycarbophil was studied for up to 16 weeks . The prepared tablets were stored in closed containers of type III glass under different conditions of temperature (30 degrees C, 40 degrees C and 50 degrees C), in open petri dishes at room temperature, at relative humidity (RH) (31%, 79.3% and 98%), and combinations of temperature and relative humidity (30 degrees C/92.9% RH, 40 degrees C/79.5% RH and 50 degrees C/65% RH) . The tablet properties such as mean weight, hardness, disintegration, in-vitro dissolution and drug content were monitored just after preparation and after 2, 4, 8, 12 and 16 weeks of storage . The tablets mean weight was significantly (p < 0.05) increased at RH 98% and at 30 degrees C/92.9% RH and non-significantly increased at 79.3% RH, 40 degrees C/79.5% RH and 50 degrees C/65% RH . There was no effect of temperature on mean weight . The hardness of the tablets increased slightly at 31% RH and decreased at 79.3% RH and at 40 degrees C/79.5% RH and became zero at 98% RH and at 30 degrees C/92.9% RH . The increase in temperature caused slight increase in tablets hardness . Tablets stored at 98% RH and at 30 degrees C/92% RH showed an increase in disintegration time associated with a decrease in rate of dissolution and indometacin content . No effect on drug content or dissolution profile was observed as a result of storage under other conditions . These findings point out to the importance of proper protection of indometacin tablets containing polycarbophil from high temperature and humidity conditions in order to ensure stability of the drug. Microbiology, 1999 Aug, 145 ( Pt 8), 1927 - 36 Spore surface glycoproteins of Colletotrichum lindemuthianum are recognized by a monoclonal antibody which inhibits adhesion to polystyrene; Hughes HB et al.; Conidia (spores) of Colletotrichum lindemuthianum, a fungal plant pathogen causing bean anthracnose, adhere to the aerial parts of host plants to initiate the infection process . These spores possess a fibrillar 'spore coat' as well as a cell wall . In a previous study a mAb, UB20, was raised that recognized glycoproteins on the spore surface . In this study UB20 was used to localize and characterize these glycoproteins and to investigate their possible role in adhesion . Glycoproteins recognized by UB20 were concentrated on the outer surface of the spore coat and, to a lesser extent, at the plasma membrane/cell wall interface . Extraction of spores with hot water or 0.2% SDS resulted in removal of the spore coat . Western blotting with UB20 showed that a relatively small number of glycoproteins were extracted by these procedures, including a major component at 110 kDa . Biotinylation of carbohydrate moieties, together with cell fractionation, confirmed that these glycoproteins were exposed at the surface of the spores . In adhesion assays, > 90% of ungerminated conidia attached to polystyrene Petri dishes within 30 min . UB20 IgG at low concentrations inhibited attachment in an antigen-specific manner . This suggests that the glycoproteins recognized by this mAb may function in the initial rapid attachment of conidia to hydrophobic substrata . Polystyrene microspheres bound selectively to the 110 kDa glycoprotein in Western blots, providing further evidence that this component could mediate interactions with hydrophobic substrata. Cell Transplant, 1999 May-Jun, 8(3), 233 - 46 Enhanced cytochrome P450 IA1 activity of self-assembled rat hepatocyte spheroids; Wu FJ et al.; Primary rat hepatocytes can self-assemble to form multicellular spheroids when plated onto Primaria petri dishes . Spheroids have been observed to exhibit enhanced liver-specific functions and differentiated ultrastructure compared to monolayer cultures on dry collagen . With confocal scanning laser microscopy, CYP1A1 activity was evaluated in situ by detecting resorufin . This highly fluorescent molecule is the P450-mediated product of 7-ethoxyresorufin O-dealkylation (EROD) . Significantly higher P450 activity was observed in spheroids compared to monolayers on collagen upon induction with 50 microM beta-naphthoflavone (BNF), a CYP1A inducer . This was confirmed by measuring microsomal EROD activity . The distribution of CYP1A1 activity within spheroids was heterogeneous, with higher activity localized to the hepatocytes in the interior . During the process of spheroid formation, cells were initially seen to attach and spread out as a monolayer . This stage was associated with relatively low CYP1A1 activity . As cells formed multicellular structures and aggregated into spheroids, the level of CYP1A1 activity increased over time . At least a fivefold higher fluorescence intensity was observed in spheroids compared to that of monolayers maintained on collagen . The higher P450 activity within spheroids may be associated with their ability to maintain a greater degree of differentiation compared to monolayers . These studies demonstrate the potential of hepatocyte spheroids as a model system for investigating drug metabolism, tissue engineering, and tissue self-assembly. Genes Chromosomes Cancer, 1999 Sep, 26(1), 84 - 91 De novo retrotransposition of unbiased sequences in a human breast cancer cell clone; Werle-Schneider G et al.; It has been demonstrated recently that certain repetitive sequences and even expressed single-copy genes are capable of retrotransposition, but little is known about the endogenous or exogenous modifiers of this process in human cells . Retrotransposition may contribute to gene inactivation and genetic instability in cancer development . We have used the human cell line MCF-7 to generate a method for investigating de novo retrotransposition in breast cancer cells . The strategy employs a reporter construct transfected into MCF-7 cells that encodes neomycin phosphotransferase gene (neoR) sequences interrupted by an intron derived from the gamma-globin gene and sandwiched between two promoters in opposite orientation; the phosphotransferase is not produced in transfected cells expressing the plasmid until transposition via a spliced antisense neoR RNA intermediate has occurred, conferring a functional gene product and thereby resistance to G418 . A stable transfectant line that showed presence of reporter plasmid DNA and expression of reporter antisense neoR was obtained and used to demonstrate spontaneous retrotransposition of neoR sequences: tester cells were subjected to selection in G418 medium, and neomycin-resistant clones were isolated at a frequency of 10(-7) . A simple PCR-based prescreening of colonies fixed and stained in Petri dishes can be used to verify intronless neoR DNA . Expanded populations of G418-resistant colonies were determined to be derived from reporter sequences that had transposed via an RNA intermediate by Southern blot genotyping . This experimental assay may be used for exploring endogenous and environmental factors that influence host cell-mediated retrotransposition of unbiased cellular sequences in breast tumor cells . Genes Chromosomes Cancer 26:84-91, 1999 . Acta Chir Plast, 1999, 41(2), 54 - 8 Experience with banked skin in the Prague Burn Center; Broz L et al.; Despite progress in materials science, the use of human allografts and xenografts of pig origin is in the Prague Burn Center among the preferred means of temporary burn wound cover since 1973 . True closure is achieved only with living autografts or isografts (identical twins) . The method for preparing fresh porcine grafts was introduced in Prague 25 years ago: dermoepidermal sheets are retrieved in strips, are treated with a lavage of chemotherapeutics and antibiotics, are spread onto sterile wet gauze and stored in Petri dishes at 4 degrees centigrade in a refrigerator . Cellular viability is maintained for 10-14 days when transferred to patients . The Prague Skin Bank commenced its activity in 1986 . The Protocol for the cryopreservation of skin was established: the pretreated skin is kept in aluminium vessels in containers with vapours of liquid nitrogen . Cryoprotective Medium is used with 15% glycerol . The skin viability has been verified by investigation of glucose metabolism . The production of fresh and long-term stored viable skin grafts has been increasing continuously and at present, the production represents 2 million square centimeters per year . About 15% of the harvest is distributed to other surgical and trauma departments . Any burn wound dressing may fail due to a failure to use them properly-lack of attention to the details in burn wound care can lead to disappointment. Tissue Eng, 1999 Jun, 5(3), 207 - 21 Receding cytochrome P450 activity in disassembling hepatocyte spheroids; Hsiao CC et al.; Primary rat hepatocytes can self-assemble to form multicellular spheroids when plated onto Primaria petri dishes or suspended in stirred vessels . These spheroids exhibit prolonged viability, enhanced liver-specific functions and differentiated ultrastructure compared to monolayer cultures . Upon transfer to collagen coated surface, or upon the addition of fetal bovine serum (FBS) to the culture, these spheroids began to disassemble and spread on the surface . The dynamics of cytochrome P450 CYP1A1/2 activity in the course of spheroid disassembly was examined in situ by detection of the fluorescent product, resorufin, of ethoxyresorufin O-dealkylation . Optical sectioning of the disassembling spheroids by confocal microscopy demonstrated that hepatocytes that reverted to monolayer exhibited markedly lower CYP1A1/2 activity than those that remained in a multilayered structure . This occurred whether the disassembly was caused by incubation with FBS-containing medium or by cultivation on a collagen-coated surface . When spheroids were cultured on the surface of agar, the disassembly process was retarded even in the presence of FBS . However, even in those intact spheroids, the exposure to FBS markedly decreased CYP1A1/2 activity . The decreased CYP1A1/2 activity was correlated to a diminished smooth endoplasmic reticulum as seen in the transmission electron micrograph . The results clearly demonstrate that the disassembly of hepatocyte spheroids led to decreased CYP1A1/2 activity . Furthermore, FBS contained a factor that caused CYP1A1/2 to decrease even in intact spheroids. Parasitol Res, 1999 Aug, 85(8-9), 765 - 9 Paramphistomum daubneyi and Fasciola hepatica: influence of temperature changes on the shedding of cercariae from dually infected Lymnaea truncatula; Abrous M et al.; Dual infections of Lymnaea truncatula with Paramphistomum daubneyi and Fasciola hepatica were performed to determine whether temperature changes in snails (daily water change with spring water at 6 degrees-8 degrees C, which subsequently increased to room temperature at 20 degrees C) would influence snail infection and the production of cercariae by both trematodes . At day 30 post-exposure the surviving snails were individually placed in petri dishes to constitute two groups . Snails from the first group were maintained at a temperature of 20 degrees C, and the water in the petri dishes was changed daily . The protocol was identical for the second group of snails except that the water temperature was 6 degrees-8 degrees C when changed . The interval between exposure and the first shedding of cercariae in snails immersed in cold water for a short period was longer (67-69 days instead of 48-50 days in the 20 degrees C group) . In both groups, snails infected only with F . hepatica or P . daubneyi or with both trematodes were detected . In snails infected only with F . hepatica the frequency of cercaria-shedding snails and the total number of metacercariae were significantly greater in the 20 degrees C group . Inversely, in snails infected only with P . daubneyi the frequency of cercaria-shedding snails and the number of metacercariae were significantly greater in the 6 degrees-8 degrees C group . In snails harboring both trematode larval forms, no significant difference in the frequencies of cercaria-shedding snails between the two groups was noted . Metacercariae of both trematodes were obtained from these snails . In the 20 degrees C group, F . hepatica metacercariae were more numerous, whereas in the 6 degrees-8 degrees C group the number of P . daubneyi metacercariae was greater . From these results it appears that greater activity of P . daubneyi cercariae occurs in snails subjected to daily temperature changes (from 6 degrees to 20 degrees C). Biol Reprod, 1999 Aug, 61(2), 548 - 52 Regulation of monocyte chemotactic protein-1 expression in human endometrial stromal cells by integrin-dependent cell adhesion; Garcia-Velasco JA et al.; Shed menstrual endometrium is viable and has the ability to implant and grow in women, who eventually develop endometriosis . Many of the cell-to-cell or cell-to-extracellular matrix (ECM) connections are mediated by integrins . Monocyte chemotactic protein (MCP)-1, a potent chemotactic factor produced in many cell types, is elevated in the peritoneal fluid of women with endometriosis . In this study, we investigated whether endometrial stromal cell (ESC) adhesion itself induces the expression of MCP-1 and whether this process is integrin mediated . ESC were plated on Petri dishes and 24-well plates coated with fibronectin, laminin, collagen IV, poly-L-lysine, or mouse anti-human integrin beta(1) and beta(2) monoclonal antibodies . Adherence of ESC to various ECM substrates, except for poly-L-lysine, a non-integrin-dependent adhesion matrix, induced the expression of MCP-1 at both mRNA and protein levels . Engagement of beta(1)-containing integrins was associated with ESC adhesion and resulted in up-regulation of MCP-1 gene expression and protein secretion . Disruption of the actin cytoskeleton by treating ESC with cytochalasin D completely blocked the increase of MCP-1 induced in response to integrin activation . These findings indicate a novel mechanism of MCP-1 regulation . Cell adhesion to ECM is an important event that leads to stimulation of MCP-1 expression, and this process is mediated by integrins. Biol Reprod, 1999 Aug, 61(2), 406 - 10 Analysis of in vitro migration patterns of human spermatozoa by a petri dish-based horizontal column; Hossain AM et al.; Spermatozoa are required to travel a considerable distance in vivo to meet the oocyte at the fertilization site . However, none of the existing in vitro tests critically evaluates migration of sperm to assess their potential of reaching the oocyte . On the other hand, an in vivo model is not suitable for this type of study because of ethical and technical constraints . In the present study we utilized a horizontal column technique to analyze sperm migration . Migratory characteristics of fresh, unwashed semen sperm and sperm undergoing various treatments were examined in vitro using a Petri dish-based horizontal fluid column . The procedure involved loading a sperm sample into the column and determining sperm concentration, motility, and viability at different column segments for different migration durations (6, 12, 24, 48, and 72 h) . All sperm samples produced an exponential migration pattern in all durations of migration . Propagation along the column edge, tendency to exit from the column, and hiding in the blind pouches were some of the important characteristic features exhibited by the migratory sperm . Variations in migration patterns were documented among semen donors, between fresh and frozen semen, and between washed and unwashed sperm . Prolonged postejaculation time diminished migratory potential . The recovery of sperm in the column end was independent of seminal variables with the exception of oligozoospermia . These observations suggest that the Petri dish-based horizontal column is effective for analyzing sperm migration characteristics for prolonged periods . The potential of this migration assay in predicting the in vivo potential of spermatozoa to reach the fertilization site will be worth exploring. Biotechnol Bioeng, 1999 Jul 5, 64(1), 92 - 100 Medium optimization for spore production of coniothyrium minitans using statistically-based experimental designs Ooijkaas LP, Wilkinson EC, Tramper J, Buitelaar RM. Statistically-based experimental designs were used to optimize a chemically defined solid medium for the spore production of Coniothyrium minitans . In the first optimization step the influence of starch, urea, phosphate, magnesium, calcium, thiamin and trace elements on spore production was evaluated using a fractional factorial design . Starch and trace elements influenced spore production positively while urea affected spore production negatively . The other components had no significant influence on spore production . In the second and third steps the concentrations of starch, urea and trace elements were further optimized using central composite designs and response surface analysis . This optimization strategy allowed the spore production to be increased by a factor 7 from 4 x 10(9) to almost 3 x 10(10) spores per Petri dish of 9 cm diameter . J Assist Reprod Genet, 1999 Jul, 16(6), 306 - 9 Randomized autocontrolled comparison of the embryo culture performance of Nunc and Falcon petri dishes; Van den Bergh M et al.; PURPOSE: Our purpose was to compare the embryo culture performance of two types of petri dishes (Nunc and Falcon) . METHODS: Mouse zygotes were cultured up to the expanded blastocyst stage in both types of dishes . The oocytes from 50 in vitro fertilization cycles were randomly divided between the two types of dishes . Fertilization, cleavage, and embryo quality were compared . Oocytes from another 50 cycles were all cultured at random in either type of dish . Pregnancy and implantation rates were compared between the two types . RESULTS: Of 91 mouse zygotes, 81 cleaved to two-cell-stage embryos, and 64 became expanded blastocysts in Falcon dishes; of 99 zygotes, 81 cleaved to two-cell-stage embryos and 66 became expanded blastocysts in Nunc dishes . Of 248 oocyte-cumulus complexes (OCC), 145 fertilized in Falcon dishes, and of 269 OCC, 175 fertilized in Nunc dishes . The high quality embryo ratio was 51 out of 118 in Falcon dishes, not different from that in Nunc dishes, 58 out of 139 . In Falcon dishes 72 out of 118 embryos were at least at the four-cell stage after 45 hr, versus 70 out of 139 in Nunc dishes . Twenty-three clinical pregnancies were obtained in the first 50 cycles with sibling oocytes . In the second group, with randomization of the cycles between Nunc and Falcon, 8 pregnancies were obtained in the Nunc and 10 in the Falcon dishes . The implantation rate in this second group of 50 cycles was 9 out of 61 in Falcon and 11 out of 57 in Nunc dishes. Mutagenesis, 1999 May, 14(3), 335 - 8 Effects of oleic acid, docosahexaenoic acid and eicosapentaenoic acid on background and genotoxin-induced frequencies of SCEs in Indian muntjac fibroblasts; Higgins S et al.; Muntjac cells were cultured at 5 X 10(5) cells/10 cm Petri dish for 24 h prior to addition of fatty acids (50 microM) which were delivered to the cells complexed with 2% bovine serum albumin (fatty acid-free) and incubated for a further 24 h . Parallel dishes were processed for lipid extraction and GC analysis . This analysis showed highly significant (P < 0.01) uptake by the cells of each fatty acid . Genotoxins (75 microM hydrogen peroxide, 20 microM t-butylhydroperoxide and 2.4 microM mitomycin C) were added to the cells for 1 h prior to the end of the 24 h fatty acid incubation period . Control (no genotoxin or fatty acid) treatments were included . No difference was observed in background frequencies of SCEs between controls and fatty acid treatments, thus indicating that these fatty acids per se do not cause DNA damage . The cells incubated with the genotoxins showed increased (P < 0.05) frequencies of SCEs when compared with control frequencies . Cells incubated with genotoxins in the presence of fatty acids also showed significantly higher (P < 0.05) levels of SCEs when compared with control frequencies . When cells supplemented with genotoxins in the presence of fatty acids were compared with cells treated with genotoxins alone, higher levels of SCEs were observed in the former, suggesting that the fatty acids exacerbate DNA damage caused by these genotoxins. Exp Physiol, 1999 May, 84(3), 489 - 99 Activation of ionic channels by deoxycholate in frog and human cell lines; Mauricio AC et al.; Humans, after extensive ileal resection, frequently suffer from diarrhoea, which may be due to an increased delivery of deoxycholate (DOC) to the large intestine . In the frog skin the addition of DOC (0.5 mM) to the apical side induced the activation of amiloride-sensitive Na+ channels and an increase in the unidirectional Cl- fluxes . Here we used two established cell lines (A6 and Caco2) to study the effect of DOC on ion channels at cell and membrane level using the patch-clamp technique . In A6 cells subcultured directly on Petri dishes and studied in the whole-cell configuration, DOC induced an increase in cell conductance of 110.3 +/- 4 pS pF-1 (N = 8) which was reduced to 89 +/- 14 pS pF-1 (N = 8) by the addition of DIDS (0.5 mM), The absolute values of these two effects were not statistically different (P < 0.2) . In Caco2 cells, the addition of DOC (0.5 mM) induced, after 1 min, an increase in cell conductance of 583 +/- 16 pS pF-1 (N = 8) which was reduced to 560.4 +/- 16 pS pF-1 (N = 8) by DIDS (0.5 mM) and N-phenylanthranilic acid (DPC; 0.5 mM) . The two values were not statistically different (P < 0.4) . In Caco2 cells subcultured under the same conditions, DOC induced an increase in cell conductance of 1710 +/- 64 pS pF-1 (N = 6) . Subsequent addition of amiloride (0.1 mM) reduced the cell conductance to 1558 +/- 33 pS pF-1 (N = 6) . These two mean values were statistically different allowing for an error of the second kind < 0 . 05 . In cells in which DOC produced a conductance increase of 1010 +/- 10 pS pF-1, gadolinium (0.5 mM) induced a fall in cell conductance of 1800 +/- 10 pS pF-1 . In Caco2 cells, addition of DOC (0.5 mM) to the bath reversibly induced the appearance of or an increase in channel activity in patches studied in cell-attached and excised inside-out configuration . In inside-out experiments (N = 13) DOC (0 . 5 mM) induced the appearance of channel activity with conductances and reversal potentials (Er) of 27.7 +/- 1.9 pS and 0.8 +/- 5.7 mV, respectively . In cell-attached patches (N = 13) these values were 24.9 +/- 4.4 pS and -18.1 +/- 6.4 mV . In excised inside-out patches from Caco2 cells, subjected to electrochemical gradients for Na+, K+ and Cl-, (+85, -85 and 0 mV, respectively), addition of DOC also induced an increase in the baseline conductance and a shift in the reversal potential from values around +25 mV to values around 0 mV . Bile salts activated both anionic and cationic channels and did not require the presence of intracellular factors for these effects . We suggest that they act at the membrane level. Trends Biotechnol, 1999 Jun, 17(6), 226 - 30 Towards electronic Petri dishes and picolitre-scale single-cell technologies; Cooper JM; Microsensors have traditionally been made as one-off, hand-crafted probes . Recently, however, there has been a concerted drive to exploit the microfabrication methods developed within the semiconductor industry in order to mass produce cheap, planar microsensing arrays . Such devices might be 'electronic Petri dishes' for the direct stimulation of, and measurement from, a variety of single cells, including neurones . In addition, micromachining has been used to construct picolitre-scale analytical sensors to extend the range of single-cell analyses. Braz J Med Biol Res, 1999 Jan, 32(1), 79 - 83 Differential in vitro pathogenicity of predatory fungi of the genus Monacrosporium for phytonematodes, free-living nematodes and parasitic nematodes of cattle; Gomes AP et al.; In vitro tests were carried out on the pathogenicity of nine isolates of the predatory fungi of the genus Monacrosporium (5 M . sinense isolates, 3 M . appendiculatum and 1 M . thaumasium isolate) for a phytonematode (second stage juveniles from Meloidogyne incognita, race 3), a free-living nematode (Panagrellus spp), and two gastrointestinal parasitic nematodes of cattle (infective larvae of Cooperia punctata and Haemonchus placei) . A suspension containing 2,000 nematodes from each species was added to Petri dishes containing fungi and grown on 2% water-agar medium at 25 degrees C in the dark for up to 7 days . The dishes were examined every other day for 7 days and predation-free nematodes were counted . The results showed that the free-living nematodes, Panagrellus spp . were the most susceptible (P < 0.05), followed by the phytonematode M . incognita, while the controls were > or = 98.5% viable . However, a variable susceptibility of the nematodes to different fungi was observed . This indicates that the use of predatory fungi for the environmental control of nematodes will be limited by the multiplicity of nematodes in the environment and their differential susceptibility to fungal isolates of the same genus. Rev Argent Microbiol, 1999 Jan-Mar, 31(1), 13 - 8 {Evaluation of Trichoderma spp . as antagonist of Rhizoctonia solani in vitro and as biocontrol of greenhouse tomato plants}; Durman S et al.; Five Trichoderma isolates were compared in their ability for controlling Rhizoctonia solani attack to tomato plants in greenhouse and as antagonists of this pathogen in three independent laboratory assays . Four out of five isolates showed biocontrol ability and decreased pathogen growth and survival of its sclerotia in soil . Results suggest that dual cultures in Petri dishes and mycoparasitism assays against R . solani sclerotia may be useful for detecting isolates effective as biological control agents against this pathogen in tomato plants. Biomaterials, 1999 May, 20(9), 879 - 84 Porous calcium phosphate coating over phosphorylated chitosan film by a biomimetic method; Varma HK et al.; A porous calcium phosphate coating deposited on chitosan films was studied using scanning electron microscopy, energy-dispersive X-ray analysis, micro-Fourier transform infrared spectroscopy (micro-FTIR) and thin-film X-ray diffractometry (XRD) . Chitosan films were first prepared by dissolving chitosan powder in dilute acetic acid and drying in a flat petri dish . The films were phosphorylated using urea and H3PO4 with the P content being 0.1-0.2 wt% . Phosphorylated films soaked in saturated Ca(OH)2 solution for 8 days led to the formation of a calcium phosphate precursor phase over the entire surface . This precursor phase stimulated the growth of a porous coating of calcium-deficient hydroxy apatite when immersed in 1.5 x SBF for more than 20 days . Phosphorylated films not treated with Ca(OH)2 did not show any calcium phosphate growth upon immersion in SBF solution . The precursor phase is thought to be octacalcium phosphate, which nucleates a HAP phase during SBF treatment . Initially, this treatment in SBF results in the formation of a single-layer calcium phosphate particles over the film surface . As immersion time in SBF increases, further nucleation and growth produce a porous HAP coating . The Ca/P ratio of the HAP coating is a function of SBF immersion time. J Biomech Eng, 1999 Feb, 121(1), 89 - 98 Surfactant-spreading and surface-compression disturbance on a thin viscous film; Bull JL et al.; Spreading of a new surfactant in the presence of a pre-existing surfactant distribution is investigated both experimentally and theoretically for a thin viscous substrate . The experiments are designed to provide a better understanding of the fundamental interfacial and fluid dynamics for spreading of surfactants instilled into the lung . Quantitative measurements of spreading rates were conducted using a fluorescent new surfactant that was excited by argon laser light as it spread on an air-glycerin interface in a petri dish . It is found that pre-existing surfactant impedes surfactant spreading . However, fluorescent microspheres used as surface markers show that pre-existing surfactant facilitates the propagation of a surface-compression disturbance, which travels faster than the leading edge of the new surfactant . The experimental results compare well with the theory developed using lubrication approximations . An effective diffusivity of the thin film system is found to be Deff = (E*gamma)/(mu/H), which indicates that the surface-compression disturbance propagates faster for larger background surfactant concentration, gamma, larger constant slope of the sigma*-gamma* relation, -E*, and smaller viscous resistance, mu/H . Note that sigma* and gamma* are the dimensional surface tension and concentration, respectively, mu is fluid viscosity, and H is the unperturbed film thickness. Phys Med Biol, 1999 Feb, 44(2), 525 - 35 Experimental validation of arthroscopic cartilage stiffness measurement using enzymatically degraded cartilage samples; Lyyra T et al.; In order to evaluate the ability of the arthroscopic indentation instrument, originally developed for the measurement of cartilage stiffness during arthroscopy, to detect cartilage degeneration, we compared changes in the stiffness with the structural and constitutional alterations induced by enzymes on the tissue in vitro . The culturing of osteochondral plugs on Petri dishes was initiated in Minimum Essential Medium with Earle's salts and the baseline stiffness was measured . Then, the experimental specimens were digested using 50 microg ml(-1) trypsin for 24 h, 0.1 U ml(-1) chondroitinase ABC or 30 U ml(-1) purified collagenase (type VII) for 24 h or 48 h (n = 8-15 per group) . The control specimens were incubated in the medium . After the enzyme digestion, the end-point stiffness was measured and the specimens for the microscopic analyses were processed . The proteoglycan (PG) distribution was analysed using quantitative microspectrophotometry and the quantitative evaluation of the collagen network was made using a computer-based polarized light microscopy analysis . Decrease (p < 0.05) of cartilage stiffness was found after 24 h trypsin (36%) and 48 h chondroitinase ABC (24%) digestion corresponding to a decrease (p < 0.01) of up to 80% and up to 30% in the PG content respectively . Decrease of the superficial zone collagen content or arrangement (78%, p < 0.001) after 48 h collagenase digestion also induced a decrease (30%, p < 0.001) in cartilage stiffness . We conclude that our instrument is capable of detecting early structural and compositional changes related to cartilage degeneration. Clin Exp Allergy, 1999 Feb, 29(2), 193 - 200 Do mite avoidance measures affect mite and cat airborne allergens? Carswell F, Oliver J, Weeks J. BACKGROUND: Effective mite allergen avoidance measures are presumed to reduce airborne allergens yet the quantity in the air is rarely measured . OBJECTIVE: To monitor airborne allergen during a placebo-controlled mite allergen avoidance study . METHODS: Bedrooms of 56 atopic asthmatic children were randomly allocated to hot washing and encasing covers + acaricide (active regime) or placebo treatment . Dust was collected from the mattress, bedding and carpets; airborne allergen was measured using Casella samplers and dust settling in open Petri dishes . Der p 1, Der p 2 and Fel d 1 were measured . RESULTS: After 24 weeks of mite allergen avoidance the Casella air-samplers collected Der p 1 less frequently in active than placebo-treated bedrooms (0 vs . 29%, P<0.05) and Petri dishes in the active group collected less than baseline (0.2 vs . 0.6 ng/day P<0.05) . Homes without cats had less cat allergen than cat-owning homes and when actively treated for 24 weeks showed a greater reduction (P = 0.03) in mattress cat allergen than the placebo group . CONCLUSION: Encasing covers and hot washing of bed linen reduced mite aeroallergen (and mattress cat allergen in the absence of cats) . This could mean dual benefits to a patient sensitive to both mite and cat. Clin Exp Allergy, 1998 Dec, 28(12), 1487 - 92 House dust mite allergen exposure in infancy; Mahmic A et al.; BACKGROUND: Infancy may be a critical time for exposure to house dust mite allergens, when exposure to high levels can increase the risk of allergic sensitization and the development of asthma in later life . OBJECTIVE: To measure house dust mite allergen (Der p 1) concentration in the infants' environment and examine lifestyle factors which may influence mite allergen exposure . METHODS: Infants aged between 4 and 12 months (n = 134) from the western region of Sydney, Australia . participated . Reservoir dust samples were collected from four sites within each home: infant's bed, second bed (adult or second child's bed), lounge floor and sheepskins (where available) . Settling aeroallergen was collected for 10-14 d in Petri dishes in the infant's room . Der p 1 was measured by ELISA . A questionnaire on types of bedding, sleeping and playing patterns of the infant was completed by the parents at the time of dust collection . RESULTS: All infants were exposed to at least one site with Der p 1 concentrations greater than 10 microg/g fine dust . The mean settling aeroallergen level in the infants' room was 24 ng De p l/m2 day and this was weakly related to bed allergen levels (r=0.21, P<0.05) . Mattress type had a weak effect on Der p 1 levels as measured in the whole bed (P = 0.07), while bed cover and bed type had no effect (P>0.6) . The mean product of time spent at a site and its allergen concentration was highest for beds in 69% of infants . CONCLUSION: The high level of allergen exposure in the environment of this group of infants places them at an increased risk of early sensitization and development of asthma . Any strategy to reduce asthma prevalence should address these high and avoidable levels. Reprod Nutr Dev, 1998 Nov-Dec, 38(6), 605 - 13 Recent developments in embryo sexing and its field application; Bredbacka P; This review focuses on polymerase chain reaction (PCR) sexing of bovine embryos in commercial situations with emphasis on new developments . Simplifications of the biopsy technique is one of the major simplifications over the last few years . The stabilization of the embryo by means of protein-free medium or scratches produced on the bottom of the Petri dish makes it possible to perform a biopsy with a single microinstrument . The traditional PCR sexing approach utilizes electrophoresis, which involves the risk of deoxyribonucleic acid (DNA) contamination of subsequent assays . Such contamination, resulting in females misdiagnosed as males, is avoided efficiently by using a non-electrophoretic method in which the sex is determined based on fluorescence of unopened tubes . However, female samples cannot be distinguished from blank samples in the non-electrophoretic assay, which thus relies on accurate transfer of biopsy into tubes . Nevertheless, an accuracy of about 95% can be reached with both approaches . High pregnancy rates (50-70%) can be reached with biopsied Grade 1 embryos, but there is evidence that pregnancy rates with Grade 2 embryos is 15-20% lower . Recent data indicate that pregnancy rates of 50% can be achieved with frozen-thawed biopsied Grade 1 embryos . In conclusion, recent developments in biopsy techniques, detection systems and freezing should increase interest in PCR sexing. Can J Vet Res, 1999 Jan, 63(1), 1 - 4 Isolation and short term culture of canine adrenal microvascular endothelial cells; Trochta OA et al.; A rapid method for the isolation of endothelial cells from canine adrenal glands is described . The technique is based on collagenase digestion of adrenal glands following mechanical tissue disruption . The endothelial cells attach and spread onto plastic petri dishes coated with a commercial attachment factor approximately one hour after seeding . Duration to first passage was approximately one week . Doubling times were 2-3 days . Cells were positively identified with immunoperoxidase staining for von Willebrand factor antigen. Exp Cell Res, 1999 Jan 10, 246(1), 221 - 32 Enhanced oxygen delivery reverses anaerobic metabolic states in prolonged sandwich rat hepatocyte culture; Bader A et al.; It must be assumed that current petri dish primary hepatocyte culture models do not supply sufficient amounts of oxygen and thus cause anaerobic metabolism of the cells . This is contrary to the physiologic state of the cells . In vivo the liver is a highly vascularized organ with a rather high blood flow rate of a mixture of arterial and venous blood . The aim of the present study was to show the oxygen dependence of primary rat hepatocytes in long-term culture and to define appropriate conditions that could allow hepatocytes to maintain tissue specific functions in an aerobic environment . To this purpose matrix overlaid hepatocytes were either cultured on gas-permeable (fluorinated hydrocarbon films) or gas-impermeable (polystyrene) supports at 10% and 20% ambient oxygen concentration (v/v), respectively . Tissue-specific functions were assessed by studying albumin and urea secretion as well as xenobiotic metabolism . The mRNA expression and catalytic activities of the cytoprotective antioxidant enzymes mitochondrial manganese superoxide dismutase (MnSOD), cytosolic copper and zinc superoxide dismutase, peroxisomal catalase, and cytosolic glutathione peroxidase were investigated to assess intracellular responses to the defined variations in oxygen supply . Hepatocytes could successfully be maintained at aerobic conditions in long-term culture on gas-permeable PTFE films . At 50% (10%, v/v) of currently used oxygen levels lactate accumulation was prevented, a plateau-like albumin secretion reestablished, urea secretion improved, and xenobiotic metabolism proceeded at physiological rates . mRNA expression of cytoprotective enzymes responded to the pericellular availability of oxygen and was most pronounced in the case of MnSOD . However, the biggest stress factor for the hepatocytes still appeared to be the isolation procedure, as mRNA expression and catalytic activities were most elevated shortly thereafter . In conclusion, this study clearly shows the oxygen dependence of primary rat hepatocytes in long-term culture and indicates means to establish appropriate conditions for the aerobic culture of primary rat sandwich hepatocytes with full maintenance of function . The long-term culture of hepatocytes on oxygenating supports at in vivo-like oxygen tensions therefore appears to be more physiologic and beneficial for the cells . J Investig Med, 1998 Dec, 46(9), 453 - 9 Matrix-mesangial cell interaction modulates migration of macrophages; Singhal PC et al.; BACKGROUND: Macrophages (Mos) have been demonstrated to play an important role in immune-mediated renal injury . Accumulation of macrophages in the mesangium has been reported to be a key event in the development of focal glomerulosclerosis . We hypothesized that mesangial cells (MCs) and matrix interaction may be a determinant for the migration of Mos into the mesangium . Therefore, we studied the effect of the interaction between matrix and MCs on the migration of Mos . METHODS: Mouse MCs were plated on Petri dishes coated either with buffer, collagen type I, III, IV, or Matrigel in media containing 1% fetal calf serum for 48 hours . Subsequently, supernatants were collected and stored . The effect of these supernatants (conditioned media) was evaluated on the migration of Mos across a filter in a modified Boyden chamber . RESULTS: Conditioned media from MCs grown on Matrigel (MC-Matrigel interaction products, MC-MGP) enhanced the migration of macrophages across a filter in a modified Boyden chamber when compared with conditioned media from MCs grown on plastic, collagen type I, type III, or type IV (MC-PP, MC-CI, MC-CIII, and MC-CIV) . MC-MGP enhanced the migration of Mos in a dose dependent manner . Anti-MCP-1 antibodies attenuated (P < 0.05) the MC-MGP-induced Mo migration (MC-MGP, 16.8 +/- 2.5 vs MC-MGP + anti-MCP-1 antibody, 6.5 +/- 1.2 migrated macrophages/field, n = 12) . Anti-TGF-beta antibodies did not attenuate MC-MGP-induced Mo migration . MCs grown on Matrigel showed a 5-fold increase of MCP-1 mRNA when compared with cells grown on plastic or collagen type IV . CONCLUSIONS: The present study suggests that matrix components may modulate the migration of Mos . This effect of MC-matrix interaction on macrophage migration may be mediated through the generation of MCP-1. Anim Behav, 1998 Nov, 56(5), 1129 - 1136 Olfactory traces and spatial learning in rats; Lavenex P et al.; We conducted an experiment to assess the use of olfactory traces for spatial orientation in an open environment in rats, Rattus norvegicus . We trained rats to locate a food source at a fixed location from different starting points, in the presence or absence of visual information . A single food source was hidden in an array of 19 petri dishes regularly arranged in an open-field arena . Rats were trained to locate the food source either in white light (with full access to distant visuospatial information) or in darkness (without any visual information) . In both cases, the goal was in a fixed location relative to the spatial frame of reference . The results of this experiment revealed that the presence of noncontrolled olfactory traces coherent with the spatial frame of reference enables rats to locate a unique position as accurately in darkness as with full access to visuospatial information . We hypothesize that the olfactory traces complement the use of other orientation mechanisms, such as path integration or the reliance on visuospatial information . This experiment demonstrates that rats can rely on olfactory traces for accurate orientation, and raises questions about the establishment of such traces in the absence of any other orientation mechanism . Anim Behav, 1998 Jul, 56(1), 35 - 9 Effects of pollen quality and genotype on the dance of foraging honey bees; Waddington KD et al.; Animals assess the quality and quantity of food and choose among different foods based on these assessments . We explored whether there was genetic variation for assessment of pollen quality by foraging honey bees, Apis mellifera . Honey bees derived from two genotypic strains foraged for pollen of varying quality from a petri dish placed inside an outdoor flight cage . The strains were the result of a colony-level, two-way selection on amount of stored pollen . We used the forager's round dance to quantify the assessments of pollen quality by individually marked worker bees . The dance rate (number of 180 degrees turns per minute) and the probability of dancing were each greater when bees foraged for pure pollen compared with a lower-quality mixture of pollen and alpha-cellulose (1:1 by volume) . Bees from the high-pollen genotypic strain had a higher dance rate than those from the low-pollen strain, suggesting different assessments . Bees from the low-pollen strain, however, had a higher probability of dancing than did bees from the high-pollen strain . Dance duration was not affected by a bee's strain or by the quality of pollen . We conclude that the dance rate may be used to quantify a forager's subjective evaluation of pollen quality and that this evaluation has a genetic component . Our results also suggest that the dance may function at the colony level to recruit bees to more profitable pollen sources . Vis Neurosci, 1998 Nov-Dec, 15(6), 1189 - 93 Perfusion system components release agents that distort functional properties of rod cyclic nucleotide-gated ion channels; Crary JI et al.; In switching from studying native cyclic nucleotide-gated (CNG) ion channels in rod cells to studying the corresponding cloned channels expressed in Xenopus oocytes, we changed our perfusion system to a more efficient one . This change involved replacing culture flasks and a small plexiglass/glass chamber with plastic syringes, metal needles, and plastic petri dishes . We now report that these new perfusion system components release agents that distort or obscure measured functional properties of rod CNG channels . The magnitude and time course of appearance of the artifacts vary widely among individual components (e.g . from syringe to syringe) . The effects most resemble voltage-dependent block of the channels, giving a decrease in current at positive potentials, and producing distortions of the kinetics and voltage dependence of channel activation. FEBS Lett, 1998 Nov 6, 438(3), 236 - 40 Factors responsible for inhibiting the motility of zoospores of the phytopathogenic fungus Aphanomyces cochlioides isolated from the non-host plant Portulaca oleracea; Mizutani M et al.; In a survey of plant secondary metabolites regulating the behaviour of Aphanomyces cochlioides zoospores, we found that root extracts of Portulaca oleracea inhibited zoospore motility . Bioassay-directed fractionation of Portulaca constituents revealed that the inhibitory activity was dependent on the interaction of two chemically different factors . These were identified as a phenolic compound, N-trans-feruloyltyramine, which by itself was active as a zoospore stimulant, and an acidic compound, 1-linoleoyl-2-lysophosphatidic acid monomethyl ester, which had zoospore-repellent activity . When Chromosorb W AW particles coated with a mixture of these pure compounds were bioassayed in Petri dishes, the inhibitory effect on zoospore motility was identical with that caused by root tip or root extracts of P . oleracea . Inhibited zoospores rapidly settled to the bottom of the Petri dishes where they initially encysted, and then germinated within 1-2 h . This is the first report of factors which inhibit zoospore motility without killing or bursting the zoospores. Int J Radiat Biol, 1998 Oct, 74(4), 501 - 9 RBE-LET relationships for cell inactivation and mutation induced by low energy protons in V79 cells: further results at the LNL facility; Belli M et al.; PURPOSE: RBE-LET relationships for cell inactivation and hprt mutation in V79 cells have been studied with mono-energetic low-energy proton beams at the radiobiological facility of the INFN-Laboratori Nazionali di Legnaro (LNL), Padova, Italy . MATERIALS AND METHODS: V79 cells were irradiated in mono-layer on mylar coated stainless steel petri dishes, in air . Inactivation data were obtained at 7.7, 34.6 and 37.8 keV/microm and hprt mutation was studied at 7 7 and 37.8 keV/microm . Additional data were also collected for both the end points with the proton LET already considered in our previous publications, namely 11.0, 20.0 and 30.5 keV/microm . RESULTS: A maximum in the RBE-LET relationship for cell inactivation was found at around 31 keV/microm, while the RBE for mutation induction increased continuously with LET . CONCLUSIONS: The proton RBE-LET relationship for cell inactivation is shifted to lower LET values compared with that for heavier ions . For mutation induction, protons of LET equal to 7.7keV/microm gave an RBE value comparable with that obtained by helium ions of about 20 keV/microm . Mutagenicity and lethality caused by protons at low doses in the LET range 7.7-31 keV/microm were proportional, while the data at 37.8 keV/microm suggest that this may not hold at higher LET values. Appl Environ Microbiol, 1998 Nov, 64(11), 4260 - 3 Method To immobilize the aphid-pathogenic fungus erynia neoaphidis in an alginate matrix for biocontrol Shah PA, Aebi M, Tuor U. Erynia neoaphidis is an important fungal pathogen of aphid pests worldwide . There have been few reported attempts to formulate this natural agent for use in biocontrol . In the current study, factors involved in the immobilization of E . neoaphidis hyphae in an alginate matrix were investigated . Hyphae of two isolates cultured in liquid medium were 220 to 620 &mgr;m in length and 7 to 19 &mgr;m in diameter with a 74 to 83% cytoplasmic content . The optimal concentration of low-viscosity sodium alginate for production of conidia from entrapped hyphae was 1.5% (wt/vol), and 0.1 and 0.25 M calcium chloride were equally suitable for use as the gelling solution . Alginate beads were rinsed with 10% sucrose after gelling . However, beads should not be left for longer than 40 min in 0.1 M calcium chloride or 10% sucrose to prevent a 10% loss in conidial production . A 40% (vol/vol) concentration of fungal biomass produced significantly more conidia than either 20% or the standard concentration of 10% . This effect persisted even after beads were dried overnight in a laminar flow hood and stored at 4 degreesC for 4 days . Conidia from freshly produced alginate beads caused 27 to 32% infection in Pea aphids as determined by standardized laboratory bioassays . This finding was not significantly different from infections in aphids inoculated with fresh mycelial mats or plugs from Petri dish cultures . In conclusion, algination appears to be a promising technique for utilizing E . neoaphidis in the biocontrol of aphid pests. J Invertebr Pathol, 1998 Nov, 72(3), 281 - 7 Routes of penetration of the entomopathogenic nematode steinernema feltiae attacking larval and adult houseflies (Musca domestica) Renn N. The way in which Steinernema feltiae (Nematoda: Rhabditida: Steinernematidae) infects the housefly, Musca domestica (Diptera: Muscidae), was investigated . Adult flies were confined in petri dishes containing 1 million S . feltiae on capillary matting and individuals were dissected at hourly intervals . First, second, and third instar larvae were placed on filter papers with 100,000 infective juveniles, and then 2 larvae were examined at 30-min intervals . Infective juveniles were aggregated on the proboscis and anal aperture of male and female houseflies after 1 h . The nematodes penetrated female flies after 2 h by moving through the cloaca, then along the oviduct, and through the ovaries . Male houseflies were penetrated via the cloaca, and then S . feltiae entered the hemcoel by penetrating the wall of the ejaculatory sac . All larval stages were penetrated via the anal aperture . Nematodes then moved through the hind gut and penetrated the wall of the ileum, immediately posterior to the pylorus . Female nematodes were observed to penetrate housefly larvae before male nematodes . Male nematodes penetrated after 10 females had successfully parasitized a larva . Vet Radiol Ultrasound, 1998 Sep-Oct, 39(5), 396 - 411 In vitro evaluation of contrast medium concentration and depth effects on the radiographic appearance of specific canine urolith mineral types; Weichselbaum RC et al.; Nine pure mineral types of canine uroliths (bladder or urethral origin only) identified in a chronologic sample from the Minnesota Urolith Center were compared to sequential dilutions of iodinated radiographic contrast medium in vitro . The uroliths studied were those composed of 100% magnesium ammonium phosphate, calcium oxalate monohydrate, calcium oxalate dihydrate, calcium phosphate appatite, calcium hydrogen phosphate dihydrate (brushite), ammonium acid urate, sodium acid urate, cystine, and silica . The radiopacity of the uroliths was classified as radiolucent, isopaque, or radiopaque, as compared to the radiopacity of the contrast medium solutions in which they were placed, using 2.0 mm and 5.0 mm depths in petri dishes radiographed using a table-top technique . A statistically significant relationship was found between the effective atomic number of the uroliths and the effective atomic number of the contrast medium solutions to which they were compared for the endpoints of isopacity, first lucency (in increasing iodine concentration sequence), and optimal visualization of internal architecture . In general, uroliths isopaque or radiolucent in contrast medium solutions weaker than 23.5 mgI2/ml are most likely ammonium acid urate or sodium acid urate . Uroliths isopaque or radiolucent in contrast medium solutions between 23.5 mgI2/ml and 44.4 mgI2/ml are probably magnesium ammonium phosphate, cystine, or silica . Uroliths that remained radiopaque in solutions stronger than 44.4 mgI2/ml, and particularly those radiopaque in contrast medium solutions stronger than 80 mgI2/ml, almost always contained calcium . This relative opacity assessment is proposed for use in double contrast cystography as an aid in differentiating urolith mineral types clinically to facilitate appropriate use of medical protocols to dissolve uroliths or to prevent their growth or recurrence. Appl Environ Microbiol, 1998 Oct, 64(10), 3846 - 53 Interlaboratory comparison of methods To quantify microsclerotia of verticillium dahliae in soil Termorshuizen AJ, Davis JR, Gort G, Harris DC, Huisman OC, Lazarovits G, Locke T, Melero Vara JM, Mol L, Paplomatas EJ, Platt HW, Powelson M, Rouse DI, Rowe RC, Tsror L. In a comparison of different methods for estimating Verticillium dahliae in soil, 14 soil samples were analyzed in a blinded fashion by 13 research groups in seven countries, using their preferred methods . One group analyzed only four samples . Twelve soil samples were naturally infested, and two had known numbers of microsclerotia of V . dahliae added to them . In addition, a control was included to determine whether transport had an effect on the results . Results differed considerably among the research groups . There was a 118-fold difference between the groups with the lowest and highest mean estimates . Results of the other groups were evenly distributed between these extremes . In general, methods based on plating dry soil samples gave higher numbers of V . dahliae than did plating of an aqueous soil suspension . Recovery of V . dahliae from samples with added microsclerotia varied from 0 to 59% . Most of the variability within each analysis was at the petri dish level . The results indicate the necessity to check the performance of detection assays regularly by comparing recoveries with other laboratories, using a common set of soil samples . We conclude that wet plating assays are less accurate than dry plating assays. J Endod, 1998 Aug, 24(8), 543 - 7 Cellular response to Mineral Trioxide Aggregate; Koh ET et al.; This investigation studied the cytomorphology of osteoblasts in the presence of Mineral Trioxide Aggregate (MTA) and examined cytokine production . MTA and Intermediate Restorative Material (IRM) were prepared and placed in separate Petri dishes . Osteoblasts (cell-line MG-63), grown to confluence in Hams F12/Dulbecco's modified Eagle's medium, were seeded into the dishes, which were incubated for 1 to 7 days . The specimens were viewed by scanning electron microscopy . For cytokine evaluation, cells were grown either alone or in other dishes containing the test materials for 1 to 144 h . Media were removed for ELISA analysis of interleukin (IL)-1 alpha, IL-1 beta, IL-6, and macrophage colony-stimulating factor . Scanning electron microscopy revealed healthy cells in contact with MTA at 1 and 3 days; in contrast, cells in the presence of IRM appeared rounded . The ELISA assays revealed raised levels of all ILs at all periods when cells were grown in the presence of MTA; in contrast, cells grown alone or with IRM produced undetectable amounts . The macrophage colony-stimulating factor was produced by cells irrespective of the group . It seems that MTA offers a biologically active substrate for bone cells and stimulates IL production. Roum Arch Microbiol Immunol, 1998 Jan-Mar, 57(1), 67 - 75 The formation of frequent additional colonies and of sectors (areas) at fungi; Cercel M; By using single-spore culture method in three points standard system, additional colonies and sectors (areas) are formed due to the dislocation of growing colonies and dissemination of conidia, respectively . Droplets (resulting from condensation of the exudate on Petri dish lids) falling on or touching the conidial masses are responsible for sectors (areas) and additional colonies formation. J Lipid Res, 1998 Sep, 39(9), 1816 - 24 Astrocytes are mainly responsible for the polyunsaturated fatty acid enrichment in blood-brain barrier endothelial cells in vitro; Bernoud N et al.; To determine the respective roles of endothelial cells from brain capillaries and astrocytes in the conversion of circulating 18:2n-6 and 18:3n-3 into 20:4n-6 and 22:6n-3, respectively, a coculture of the two cell types mimicking the in vivo blood-brain barrier was used . During the culture period, endothelial cells cultured on an insert were set above the medium of a Petri dish containing or not a stabilized culture of astrocytes . Five days after confluence, labeled 18:2n-6 and 18:3n-3 (10 microM each) were added to the endothelial cells and incubated for 48 h . Analogous experiments were also performed by using each cell type cultured alone in the culture device . The distribution of radioactivity in lipids and fatty acids was studied in all the compartments of the culture device . Endothelial cells cultured alone weakly converted the precursor fatty acids into 20:4n-6 and 22:6n-3 . When endothelial cells were cocultured with astrocytes, their content of polyunsaturated fatty acids increased dramatically . This effect was associated with the uptake of polyunsaturated fatty acids from the lower medium (astrocyte medium) . These fatty acids were released by astrocytes after they were synthesized from the precursor fatty acids that passed through the endothelial cell monolayer into the lower medium . Polyunsaturated fatty acids were released by astrocytes as unesterified fatty acids and as phospholipids (mainly phosphatidylcholine and lysophosphatidylcholine) even when the medium was devoid of serum . These results suggest that astrocytes could play a major role in the delivery of essential polyunsaturated fatty acids to the barrier itself and to the brain. Surgery, 1998 Sep, 124(3), 551 - 60 Immunoaffinity localization of the enzyme xanthine oxidase on the outside surface of the endothelial cell plasma membrane; Vickers S et al.; BACKGROUND: Reactive oxygen metabolites generated from endothelial xanthine oxidase (XO) trigger reperfusion injury in many organs . We evaluated the possibility that endothelial XO was localized on the endothelial cell surface, as well as within the cytoplasm . METHODS: Primary cultures of bovine (BAECs) and porcine (PAECs) aortic endothelial cells were grown in media documented to be free of XO . Polyclonal and monoclonal antibodies were developed against XO . These antibodies were used to evaluate BAEC and PAEC for cell surface XO through immunofluorescence staining, hybridoma cell surface labeling, and endothelial cell surface binding . RESULTS: These antibodies bound specifically to the surface of these cells when the membrane was shown to be intact and impermeable (and the cytoplasm inaccessible) to immunoglobulins Moreover, hybridoma cells expressing monoclonal antibody to XO bound specifically to the endothelial cell surface . Finally, intact endothelial cells bound specifically to the anti-XO polyclonal antibodies immobilized to the surface of a Petri dish . The integrity of these endothelial cell plasma membranes was demonstrated by the subsequent growth and replication of these cells in culture . CONCLUSIONS: These findings indicate that XO is present on the outside surface of the endothelial cell plasma membrane . This would not only explain the known in vivo efficacy of intravascularly administered large molecular weight antioxidants (such as superoxide dismutase) but could have important implications for inflammatory signaling. Allergy, 1998 Aug, 53(8), 794 - 7 Ultrasonic nebulization of hypertonic solution: a new method for obtaining specimens from nasal mucosa for morphologic and biochemical analysis in allergic rhinitis; Melillo G et al.; Various techniques are used to collect specimens from the nasal mucosa for morphologic and biochemical analysis . The purpose of this study was to devise a method that overcomes some of the disadvantages (e.g., invasive procedure, samples not suitable for cytologic and biochemical analysis, lack of standardization, and poor reproducibility) of these techniques . The new method requires subjects, with neck extended, to inhale an ultrasonic nebulization of a hypertonic (3% NaCl) solution (UNHS) for 5 min . They then blow their nose into a Petri dish, one nostril at a time with the other one blocked . The secretions are dispersed with 0.1% dithiothreitol in phosphate buffer solution for 20 min . Total cell count (TCC) is evaluated, and the cellular suspension is divided into two aliquots: one is centrifuged and the supernatants are collected for eosinophil cationic protein (ECP) measurements; the other is cytocentrifuged and the slides, stained with Diff-Quik, are used for differential cell count . The results obtained with the UNHS and nasal lavage (NL) methods were compared . Eleven nonatopic healthy subjects and 19 allergic rhinitic patients were studied . Total cell count (x10(5)) was significantly higher with UNHS than with NL (13.0+/-12.3 vs 1.9+/-1.6; P<0.01) The differential cell count was similar with the two procedures . ECP levels (microg/l) were higher with UNHS than with NL (39.1+/-38.2 vs 16.7+/-41.2; P<0.01) . For evaluation of reproducibility, four healthy and six rhinitic subjects underwent UNHS on two occasions within 5 days, and the results of two samples (sample 1 vs sample 2) were analyzed . Reproducibility was good as to TCC, differential cell count, and ECP. Biochem Biophys Res Commun, 1998 Jul 30, 248(3), 812 - 6 Secondary excitotoxicity contributes to dopamine-induced apoptosis of dopaminergic neuronal cultures; Zhang J et al.; Dopamine (DA) and related catechols may contribute to selective degeneration of dopaminergic neurons in the substantia nigra in Parkinson's disease . To investigate whether DA induces apoptosis of dopaminergic neurons, we characterized the effects of various concentrations of exogenous DA on a substantia nigra/neuroblastoma hybrid cell line (MES 23.5 or MES) . The hybrid MES cells were maintained in the presence of 50 microM glutamate in logarithmic growth on poly-D-lysine-precoated T-75 flasks and plated either onto petri dishes with glass coverslips for morphological studies or onto 6-well plates for quantification of apoptosis by flow cytometry . The results showed that DA exposure (0.5-20 microM) induced time- and dose-dependent apoptotic cell death of MES cells . To further analyze the mechanism responsible for DA-mediated apoptosis, we repeated the experiments at 20 microM DA in the presence or absence of 40 microM nomifensine, a DA re-uptake inhibitor, and 25 microM 2-amino-5-phosphonopentanoic acid (AP5), an N-methyl-D-aspartate (NMDA) receptor antagonist . The data indicate that both compounds significantly prevented DA-induced apoptosis of MES cells and that combination of AP5 and nomifensine provided greater protection against DA toxicity than AP5 alone . These results suggest for the first time that DA-induced apoptosis in dopaminergic neurons is partially attributable to increased vulnerability of these cells to non-toxic levels of excitatory amino acids, i.e., secondary excitotoxicity. Int J Radiat Biol, 1998 Jul, 74(1), 139 - 44 Analysis of radiation- and 5-FU-induced inhibition of cell proliferation by an automatic colony analyser; Piasecka A et al.; Proliferation of Chinese hamster cells B-14 was inhibited by irradiation, by incubation with 5-fluorouracil (5-FU) and by a combination of both treatments . The reduction in proliferation was assayed by the colony formation test, which was evaluated by an automatic colony analyser according to the number and volume of the colonies . It was demonstrated that the number of colonies multiplied by the volume was equivalent to the number of cells in a Petri dish and is called total colony volume . Since this quantity reflects the entire proliferation of cells, it is a more sensitive parameter for measuring cell viability than the clonogenicity of cells . The drug-radiation interaction showed a supra-additive effect, if total colony volume is taken into account, while the traditional scoring of colonies yielded only an additive effect. J Control Release, 1998 Jan 2, 50(1-3), 103 - 9 Effect of fractal dimension on drug permeation through porous ethylcellulose films; Yamane S et al.; Fractal geometry was applied to quantify the complexity of an internal structure of a porous film prepared with ethylcellulose (EC) and diethylphthalate (DEP) as a plasticizer . EC was dissolved together with DEP in a water-ethanol mixture solution, and then evaporated on Teflon petri dishes in order to make porous EC films . Boundary lines of the porous structures in the EC film cross section were taken by a confocal laser microscope as image data, and these images were fed into a computer to estimate the fractal dimension . The porous structure in EC film was observed to be a typical fractal and its complexity was quantified as a non-integral fractal dimension . No clear correlation was observed between the fractal dimension and the porosity of EC films, suggesting that they were mutually independent parameters representing the porous structure in the EC films . The permeation of theophylline through the EC films was determined by using two-chamber diffusion cells . A fairly good relationship between the permeability coefficient of theophylline and the fractal dimensions was observed, suggesting the usefulness of the fractal dimension as a novel parameter for evaluating drug permeation through porous films. Ann N Y Acad Sci, 1998 Jun 29, 849, 355 - 64 Entomopathogenic nematodes as a potential biological control method for ticks; Kocan KM et al.; Entomopathogenic nematodes have been used for biological control of certain insect pests . In these studies the nematodes were tested as a possible biological control agent for engorged female ticks . Five species of infective juveniles (IJs) were tested initially for their ability to penetrate and kill ticks, including Steinernema glaseri (SG), S . riobravus (SR), S . carpocapsae (DT), S . feltiae (SF) and Heterorhabiditis bacteriophora (HP88) . Infective juveniles (IJs) of SRs and SFs appeared to be the most effective in killing ticks and invaded and killed 30 to 100% of replete females . These two nematode species were tested on several tick species including Amblyomma americanum, A . cajennense, A . maculatum, Dermacentor variabilis and Rhipicephalus sanguineus . Although the killing rate of each tick species varied, the nematodes did not appear to be host specific and were able to kill ticks of all species tested . Egg mass weights of exposed ticks of each species were significantly lower than those of the controls . Ticks were examined with microscopy to determine whether nematodes entered and multiplied inside ticks . Partially fed female Amblyomma americanum and Dermacentor variabilis exposed to 5000 IJs in petri dishes were collected at 8, 24, 48 and 96 hrs (Trial 1) and 1, 2, 3, 4, 7 and 9 days (Trial 2) post-exposure, and fixed, processed and embedded in resin for microscopy studies . Only a few nematodes were seen in the hemocoel and tissues and they were surrounded by a clear space . Bacteria, released from the nematodes, were present in the exposed ticks and appeared to increase daily causing a generalized infection . Degeneration of tick tissues and death of the ticks appeared to result from bacterial proliferation . Nematodes did not multiply within ticks as they do in insect larvae . In these controlled laboratory studies, exposure of ticks to nematodes resulted in tick mortality and reduced egg production . Entomopathogenic nematodes appear to have potential as a biological control agent of ticks, but future studies will be required to determine whether nematode/tick interactions will occur in the field. Br J Dermatol, 1998 May, 138(5), 849 - 51 A novel culturing and grafting system for the treatment of leg ulcers; Villeneuve P et al.; The purpose of this study was to develop and test an efficient culturing and grafting system for the treatment of leg ulcers . The culturing system consisted of a Petriperm culture vessel (20 cm2) aseptically placed in a larger standard Petri dish (60 cm2) . Skin cultures were established and cultivated in the Petriperm dish . The cells grew on the bottom of the Petriperm dish, which was made of a gas-permeable 25-micron thick transparent Teflon film . Grafts were produced simply by cutting the film from the bottom of the Petriperm dish with a scalpel . The system was used to produce subconfluent epidermal autografts which were used to heal a 32 cm2 chronic rheumatoid arthritis leg ulcer . The cultured autografts were transferred cell side down on to the cleaned wound bed without an enzymatic digestion . The grafts consisted of autologous keratinocytes, melanocytes and fibroblasts . Caution was taken not to disturb the wound bed for 7-9 days at which time the Teflon film was removed . The wound closed 2 weeks after the last grafting and has remained closed for more than a year post-treatment . The culturing and grafting system presented here will make it possible to develop cellular-based therapies that were previously not possible. Artif Cells Blood Substit Immobil Biotechnol, 1998 Jul, 26(4), 437 - 9 In vitro experimental research of rabbit condrocytes biostimulation with diode laser Ga-Al-As: a preliminary study; Morrone G et al.; The scope of our study was to verify the effects of a new diode laser device with active material composed of Gallium, Aluminum and Arsenic (Ga-Al-As) configured as MOCVD (780 nm., 3000 mW) for the biostimulation of the cartilage cells in vitro . The condrocytes cells, withdrawn from the cartilage of the medial condyle of the femur of the rabbit, were cultivated, incubated and subject to biostimulation treatment with the laser . The condrocytes cells were placed in 24 Petri dishes at the concentration of 0.25 x 10(5)/ml and divided into 4 groups: 3 group (I, II, III) were treated with the laser and the fourth group (IV) was used as the control group . At the end of the treatment, all four groups, were evaluated with a MTT test and a cell count of the condrocytes cells . Group III (300 J, 1 Watt, 300 Hz, 10' of exposure time with a pulsating emission) provided the best results in terms of cell viability (MTT test) and for the number of cells found in the dishes when compared to the other treated groups and the control group . The results obtained with the use of this new diode laser Ga-Al-As device in the biostimulation of the cartilage tissue, permits us to consider the use of this device clinically. Microvasc Res, 1998 May, 55(3), 201 - 14 A novel assay of angiogenesis in the quail chorioallantoic membrane: stimulation by bFGF and inhibition by angiostatin according to fractal dimension and grid intersection; Parsons-Wingerter P et al.; In a novel assay of angiogenesis in the quail chorioallantoic membrane (CAM), we measured vascular pattern and angiogenic rate after homogeneous exposure of the entire vascular tree to recognized modulators of vessel growth . In comparison to phosphate-buffered saline (PBS)-treated controls, the vascular stimulator, basic fibroblast growth factor (bFGF or FGF-2), increased the rate of angiogenesis by a maximum of 72%, whereas a recently discovered angiogenic inhibitor, angiostatin, decreased the rate of vascular growth by a maximum of 68% . The perturbants were applied in PBS to the CAM of 7-day-old embryos (E7) cultured in petri dishes, and the embryos were cultured further until fixation at E8 or E9 . For morphometry of the quasi-two-dimensional CAM vasculature, digital images of arterial endpoints from the middle region of the CAM were acquired in grayscale at a magnification of 10x, binarized to black/white, and skeletonized . The pattern of vessel branching was assessed by measurement of the fractal dimension (Df), and vessel density (rhov), with the method of grid intersection . Correlations between these two statistical techniques were linear (r2 ranged from 0.967 to 0.985) . For skeletonized images at E9, Df and rhov of bFGF-treated samples were 1.55 +/- 0.01 and 782 +/- 26/cm2, respectively (relative to 1.49 +/- 0.02 and 583 +/- 60/cm2 for controls), and of angiostatin-treated samples, 1.43 +/- 0.02 and 424 +/- 74/cm2 (relative to 1.50 +/- 0.02 and 616 +/- 59/cm2 for controls) . To establish normalization values for rates of angiogenesis, we analyzed untreated CAMs of E6 to E12 . From E7 to E10 in skeletonized images, Df increased linearly from 1.37 +/- 0.01 to 1.54 +/- 0.01 and rhov from 311 +/- 67 to 746 +/- 124/cm2 (in both cases, r2 = 1.000) . Thus, the rates of normal angiogenic growth as measured by Df and rhov were 0.06/day and 138/cm2-day, respectively . From E10 to E12, Df and rhov declined slightly . Differences between the vasculature of untreated and PBS-treated CAMs were statistically insignificant . In conclusion, vascular branching pattern and density in the quail CAM were stimulated by bFGF and inhibited by angiostatin . We quantified these changes with statistical significance by Df and rhov, which are expressed relative to the rates of normal developmental angiogenesis measured for the two parameters in untreated quail embryos . Plast Reconstr Surg, 1998 Jul, 102(1), 73 - 7 Efficient in vitro model for immunotoxicologic assessment of mammary silicone implants; Rhie JW et al.; In clinical and experimental studies, silicone gel has been assumed to cause immune alterations that may be related to macrophage activation of silicone implants . However, it has not been proven that the immunotoxicities are caused by the direct contact of macrophages and silicone gel because there has not been an adequate experimental model . In the present study, silicone gel was loaded directly onto Petri dishes and was distributed uniformly to the bottom by centrifugation . Peritoneal macrophages and splenic lymphocytes were cultured either on the silicone-coated plates or on the conventional, normal plates, and their functions were compared with each other . The experiments were repeated three times . The cytotoxic activities of peritoneal macrophages on cancer cells were markedly augmented by cultivation on silicone gel, and the primary T-dependent immunoglobulin M response in which macrophages participated as anti