Study On The Methylation Status Of α-SMA In The Phenotypic-change Of Renal Residential Cells

Objective: The sequence of transcription initiation region of α-SMA that is symbolic protein of phenotypic-changed of renal residentialcells was location and ration detected the methylation status bypyrosequencing.Then to analyze the difference of illustration and amoutof methylation that belong to α-SMA gene in the front or back ofphenotypic transition HBZY-1and NRK-49F.It will help us to clarify thepossible mechanism of phenotypic-change of renal residential cells byepigenetics theory and thus provide a new and feasible direction for theprevention and treatment of kidney fibrosis. Methods:(1)Hadsynchronized HBZY-1and NRK-49F were treated with recombinanthuman TGF-β1(10ug/L,48h) and no.The morphological of cells,proteinexpressions of α-SMA in cells and typeⅠcollagen of culture medium inevery group were detected by immunocytochemistry and ennzyme-linkedimmunosorbent assay(ELISA).Then to judge the result of phenotypictransition of induced cells.(2) HBZY-1and NRK-49F were treated withthe same with front of methord,the genomic DNA were extracted.Then,the methylation status of α-SMA gene transcription initiation region wasdetected by pyrosequencing and analyzed the difference each group in thetwo cells. Results:(1)The morphologic of HBZY-1and NRK-49F in response to TGF-β1treatment were changed.The cells and nucleolus werebigger,the kytoplasm was more.HBZY-1from fusiformis or irregular starbecame to polygon or branching and outstretched protoplasmic extensionconnected to each other.NRK-49F from elongated fusiformis became toshortened fusiformis or polygon and vacuole were observed in somecells.(2)Indirect immunocytochemistry staining showed that α-SMAwas weakly positive expressed in untreated HBZY-1,but it wasupregulated and enhanced in treated ones.What was more,expression ofα-SMA could not be seen in untreated NRK-49F,but it was arised andupregulated in treated ones(P<0.05).(3)The results of ELISA showed thattypeⅠcollagen could be expressed in untreated HBZY-1and NRK-49F,while there were significant expression in treated ones(P<0.05).(4)Themethylation status of α-SMA gene transcription initiation region inuntreated HBZY-1and NRK-49F were difference.The methylationfrequency of HBZY-1was from20%to50%.It was hypomethylation.The methylation frequency of HBZY-1was more than50%. It washypermethylation.However,the treaed two cells were not altered onillustration and amount of α-SMA gene rmethylation.Conclusion:(1)TGF-β1can induce HBZY-1and NRK-49F phenotypic transition andto become myofibroblast that can be high-express α-SMA and secrtegreat amount extracellular matrix.It playes an important role in theformation of renal fibrosis.(2)The effect of α-SMA methylation status of transcription initiation region in renal residential cells phenotypictransition is not be sured in our experiment.f phenotypic-change of renalresidential cells by epigenetics theory.It that renal fibrosis is developed bychange the α-SMA methylation status needs further study.