BHMT在糖尿病性白内障晶状体中的表达及功能研究OA

Objective To investigate the expression of betaine-homocysteine methyltransferase(BHMT)in the lens of diabetic cataract(DC)and its effect on homocysteine(Hcy)metabolism,apoptosis and oxidative damage in lens epithelial cells exposed to high glu-cose condition.Methods Lens samples were collected from corneal transplant donors(CL group),age-related cataract patients(ARC group),and diabetic cataract patients(DC group),with 10 specimens in each group.The mRNA and protein expression of BHMT,as well as Hcy levels,were detected in in the lens cortex and nucleus of these samples.Human lens epithelial cells HLEC-B3 were divided into four groups:control group,high glucose incubation group(HG group,30 mmol/L glucose),BHMT overexpression group(BHMT-OE group),and BHMT overexpression plus high glucose incubation group(BHMT-OE+HG group).Cells in each group were cultured for 24 hours.Cell proliferation of HLEC-B3 cells was accessed by MTT assay and EdU assay,and cell apoptosis by TUNEL assay.The levels of Hcy and reactive oxygen species(ROS),as well as the activities of reduced glutathione(GSH)and oxidized gluta-thione(GSSG),were measured using commercial kits following the manufacturers' instructions.The mRNA levels of BHMT and αA-crystallin(CRYAA)were detected by RT-qPCR.Western blot was applied to measure the protein levels of BHMT,endoplasmic reticu-lum stress(ERS)-related proteins,B-cell lymphoma-2(Bcl-2)and CRYAA.Results Compared with the CL group,the mRNA and protein levels of BHMT in the lens cortex and nucleus were decreased,while Hcy levels were increased in both the ARC group and the DC group(all P<0.05).Compared with the ARC group,the mRNA and protein levels of BHMT in the lens cortex were decreased and the Hcy level was increased in the DC group(all P<0.05);in contrast,in the lens nucleus of the DC group,the mRNA and protein levels of BHMT were increased(P<0.05),while Hcy levels were decreased(P<0.05).Compared with the control group,the mRNA and protein levels of BHMT and CRYAA,the relative cell viability,EdU positive rate,Bcl-2 protein level and GSH content in the HG group were decreased(P<0.05);conversely the TUNEL positive rate,relative fluorescence intensity of ROS,contents of Hcy and GSSG,and levels of ERS-related proteins were increased(P<0.05).Compared with the HG group,the mRNA and protein levels of BHMT and CRYAA,the relative cell viability,EdU positive rate,Bcl-2 protein level and GSH content in the BHMT-OE+HG group were increased(P<0.05);however the TUNEL positive rate,relative fluorescence intensity of ROS,contents of Hcy and GSSG,and levels of ERS-related proteins were decreased(P<0.05).Conclusion BHMT is down-regulated in the lenses of DC patients.U-pregula-tion of BHMT promotes lens epithelial cells proliferation and inhibits apoptosis by modulating Hcy metabolism,alleviating oxidative and endoplasmic reticulum stress(ERS),and upregulating CRYAA expression.