| Objective To investigate the influence of L-glutamine on the expression of heat shock protein70in Muller cells in vitro.Methods A stable method to culture Muller cells in vitro was found by dissecting the retinae of rats postnatal5-7days into cell suspension after digested by trypsin. Immunocytochemical technique was used to indentify the Muller cells, such as anti-rat glutamine synthetase (GS) antibody. Cultured Muller cells in vitro were randomly divided into normal control group, zinc sulfate group in which cells were cultured with medium containing200umol/L zinc sulfate, and glutamine groups in which cells were cultured with medium containing different concentrations of glutamine (8n12>15>20mmol/L). After cultured for six hours, the expression level of HSP70protein of Muller cells was detected by immunocytochemistry and the expression of HSP70mRNA was detected by real-time quantitative polymerase chain reaction.Results Retinal Muller cells were cultured successfully, more than95%of which were positively stained by glutamine synthetase. The results of immunocytochemical demonstrate that the expression of HSP70was faint in cells from normal control group, but increased obviously and concentration-dependently in cells from experimental group treated with different concentrations of glutamine (P<0.05), the maximum expression of HSP70was in cells from experimental group treated with15mmol/L glutamine. The results of real-time quantitative RT-PCR demonstrate that the expression level of HSP70mRNA was faint in cells from normal control group, but increased obviously and concentration-dependently in cells from experimental group treated with different concentrations of glutamine (P<0.05), the maximum expression level of HSP70mRNA was also in cells from experimental group treated with15mmol/L glutamine.Conclusion There is mild expression of HSP70in normal retinal Muller cells in vitro. Hypereffective expression of HSP70may be induced by L-glutamine and has concentration dependency. |
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