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Objective To analyze the stability of chromosome variant ratio of three available transformed corneal cell lines. Methods Chromosome specimens of transformed cells including human corneal epithelial cells(HCE),bovine corneal endothelial cells(BCE) and rabbit corneal epithelial cells(RCE) were prepared by a direct method using regular Giemsa staining. Chromosomes of cells in metaphase were counted under the microscope. Then, the variant ratio of chromosomes and their nuclear types were analyzed. Results The chromosome numbers were 56 to 65, 27 to 34 and 74 to 88 for HCE, BCE and RCE, respectively. Chromosome numbers in the three commonly used and transformed corneal cell lines were changed in comparison to their parent tissues. Conclusion Genotyping study may provide important information for using HCE、BCE、RCE in functional studies.
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Objective:To investigate the effects of adramycin on cell line of FRH-0201.Methods:Human hilar cholangiocarcinona cell line were cultured with adramycin (2?g/ml) through lasting different times,finally this cell line can survive after 72h cultured in the culture liquid with the adramycin(1?g/ml) .Then the medium was replaced with a fresh one without adramycin,and the cells were continuously cultured for 1 month.The in vitro studies included the observation of the appearance with optical microscope,MTT cell proliferation assay,analysis of the cell cycle by Flow cytometry(FACS),assays of tumor marker using en{yme-linked immunosor bent assay(ELISA),the fluorescence density of adramycin.Results:Compared to the parent FRH-0201cell line:The morphology showed typical morphological characteristics,the doubling time prolonged about 3h,the cell cycle by flow cytometry identified in G_1,G_2 and S phase of cell cycle were 40.50%,19.74%and 39.77% in trial group,and 69.83%,7.29%and 22.88% in parent group,respectively.Tumor marker has no difference between them,the fluorescence density of epirubicin descending 5.2 times.Conclusion:The adramycin can affect the cell cycle and cause the change of metabolism.
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The bioengineering research is essential in the development of ideal combination of biomaterials and cultured cells to produce the permanent wound coverage. The experimental model of cultured keratinocytes presents all steps of the culture, since the isolation of the keratinocytes, preparation of the human acellular dermis, preparation of the composite skin graft and their elevation to the air-liquid interface. The research in cultured keratinocytes model advances in two main ways: 1. optimization of the methods in vitro to the skin cells culture and proliferation and 2. developing biomaterials that present similar skin properties.
A pesquisa em bioengenharia é primordial no desenvolvimento da combinação ideal de biomateriais e células cultivadas para produzir a cobertura definitiva das lesões. O modelo experimental da cultura de queratinócitos apresenta toda as etapas do cultivo, desde o isolamento dos queratinócitos, preparação da derme acelular humana, do enxerto composto e da sua elevação à interface ar-líquido. A pesquisa em modelo de cultura de queratinócitos desenvolve-se em duas vias principais: 1. otimização dos métodos in vitro para cultivo e proliferação de células da pele e 2. desenvolvimento de biomateriais que mimetizem as propriedades da pele.
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Aim To investigate the effects of EDT on anoxia and ischemic injury in cultured PC12 cells. Methods Cultured PC12 cells were treated with 1 mmol?L -1 Na 2S 2O 4 and 20 mmol?L -1 NaCN in combination with glucose deprivation. The protective effects of EDT on these two models were evaluated by lactate dehydrogenate (LDH) efflux assay and colormetric MTT assay.ResultsEDT, within the range of 10 -8～ 10 -6 mol?L -1, significantly antagonized LDH efflux induced by two models and increased the optical density at 570 nm tested by colorimetric MTT assay in concentration-dependent manner. 10 -6 mol?L -1 EDT might time-dependently inhibit two injuries and reach maximal level at 48 h. Conclusion EDT can protect PC12 cells from anoxia and ischemic injury.
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Objective To investigate the effects of the serum contained Shuluo sterile powder for injection (SLI) on injury induced by anoxia and hypoglycemia in cultured PC12 cells. Methods Cultured PC12 cells were treated with Na2S2O4 and Earle′ s liquid with glucose free to induce models of anoxia and hypoglycemia. The protective effects of the serum contained SLI on the two kinds of models were evaluated by the activity of lactate dehydrogenate (LDH) and staining with MTT assay. Results The serum contained SLI counteracted the injury induced by anoxia and hypoglycemia to various degrees and prevent the efflux of LDH. Conclusion The serum contained SLI can protect PC12 cells from injury induced by anoxia and hypoglycemia.
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Nephrotoxicity is the most common dose-limiting factor of cyclosporine A (CSA) in clinical usage. But the mechanism of CSA-induced nephrotoxicity still remains unresolved. Many authors insisted that CSA induced renal proximal tubular cell injury is due to the secondary effects following hemodynamic changes or endothelial cell damage, instead of direct toxicity by CSA. To find out that CSA has a direct toxicity to the proximal tubular cells, the author used primary cultures of human proximal tubular cells to eliminate the hemodynamic or endothelial influences that could be produced in in vivo model. In the present study, the viability against CSA was tested by the neutral red assay method with modulation of Ca2+ amount in incubating media and observed electron microscopically. The viability test showed direct toxic effect of CSA on human proximal tubular cells and this was enhanced by Ca2+ depletion in incubating media. Morphologically noted are accumulation of lipid droplets and polyribosomal dispersion, which may be association with inhibition of cellular synthetic activity. These results suggest the toxixity is a direct effect of cyclosporine and that toxic mechanism may be due to inhibition of cellular synthetic activity. And this experiment also showed that primary cultures of human renal proximal tubular cells can be a good in in vivo model for investigating CSA nephrotoxicity.
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HumanosRESUMEN
Cyclosporine A (CSA), a lipophilic cyclic undecapeptide, is not accumulated evently in all tissues and has a high affinity to several tissues such as lymphoid organs, liver, and kidneys. From this point of view, it is reasonable to assume that the amount of CSA uptake would be correlated with the extent of cell injury. On the other hand, verapamil, a Ca2+ channel blocker, bas been shown to ameliorate CSA nephrotoxicity. Since proximal tubule is the major site of drug transport and CSA uptake and its interaction with verapamil in isolated human renal proximal tubular cells. The CSA uptake rapidly increased over the first 5 min and then achieved almost steady-state after 10 min at all concentrations (0.5-10 uM). Kinetic analysis yielded that the Km and Vmax values of CSA were 5.6 uM and 86.2 p mol/mg cell protein/min, respectively. And Ca2+ depletion in media enhanced CSA uptake significantly but verapamil reduced it. These results suggest that the Ca2+ channels and CSA transporting sites on cell membrane are closely associated and that Ca2+ and CSA might be taken up competitively by proximal tubular cells.
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HumanosRESUMEN
Objective To study effects of GDNF and HSV GDNF on apoptosis of spinal cord motoneurons after scratch injury in vitro. Methods In the period of culture cell,motor neurons were periodically observed and counted.Scratch injury was executed on culturing 12th day,in the same time,cultured neurons were divided into 4 groups,and each group was given corresponding medium(medium serum free control group,serum group,HSV GDNF group,GDNF group).On the 4th and 7th day after scratch injury,TUNEL staining was respectively performed,and the number and the mean densities of apoptotic motoneurons were observed. Results The number of living motoneurons was in inverse proportion to time of scratch injury in each group.The number of apoptotic motoneurons from control group,HSV GDNF group to GDNF group was successively decreased as well as the mean densities of apoptotic motoneurons on the 4th and 7th day after scratch injury.Furthermore,the effects of groups with serum were no better than those of medium serum free groups,in the same time,difference was not obviously in HSV GDNF group and GDNF group. Conclusion GDNF and HSV GDNF can decrease apoptosis of injured motoneurons in vitro .It suggests that GDNF and HSV GDNF might play an important role in the growth and development of motor neurons.