Cigarette Smoke Extract Promotes Senescence By Reducing Expression Of HDAC2 In Murineskeletal Muscle Cells

PART ?Effects of cigarette smoke extract on murine skeletal muscle cells senescenceAIM:To investigatethe cigarette smoke extract(CSE)can cause murineskeletal muscle cells senescenceMETHODS:To culture C2C12 myoblast and induce it to differentiate into skeletal muscle cells,which were stained and numbered byspecific antibody MY-32.After exposure of cigarette smoke,to observe the effects of CSE on skeletal muscle cellsactivities by MTT.The percentage of skeletal musclesenescent cells were detected by?-galactosidasestaining.RESULTS:After the cells differentiation,the rate of positive cells was 98%,the cell differentiation was successful.The activity of skeletal muscle cells after intervention of CSE,compared with control group,there was no statistical difference(P>0.05)with the concentration of CSE nofewer than 60 mL/L in 12 h,24 h,while the concentration of CSE was more than 80 mL/L,the growth of cells were restricted obviously(P<0.05).After 24hintervention of CSE,at the certain range of concentration(CSE ?60 mL/L),with the increase of concentration of CSE,the number of senescent cells was increased.While the CSE>60 mL/L,continued to increase the concentration of CSE,the cell changed on morphology and then died.The positive cell rates of ?-galactosidase staining in the skeletal muscle cells was decreased.CONCIUSION:CSE can cause skeletal muscle cells senescence.After 24h intervention of CSE,when the CSE within the certain concentration range(CSE?60 mL/L),with the increase of concentration of CSE,the positive cell rates of ?-galactosidase staining in the cells was increased.PART ?Effect of HDAC2 expression on senescence in murine skeletal muscle cells exposed to cigarette smoke extractAIM:To investigate therelationshipbetween HDAC2 and senescence in skeletal muscle cellsexposed to cigarette smoke extract.METHODS:The expression of HDAC2 at protein levels in skeletal muscle cells was observed after intervention of CSE(60mL/L)at different time points(24 h,48h,72h).Using the histone deacetylase activator(TBB)and inhibitor(valproic acid),the cells were divided into six groups:control group,CSE group,TBB group,TBB + CSE group,valproic acid group,valproic acid + CSE group.The expression of HDAC2 at mRNA and protein levels was detected by qRT-PCR and Western blotting,and the positive cellrates of ?-galactosidase staining were detected.RESULTS:After intervention of 60 mL/LCSE,compared with the control group,the expression of HDAC2 of CSE group had statistically significant differencesat each time point(P<0.05).The longer the intervention time,the less HDAC2expression.The expression of HDAC2 at mRNA and protein levels was increased by TBB,and the positive cellrates of ?-galactosidase staining were decreased.On the contrary,HDAC2 expression at mRNA and protein levels was reduced by histone deacetylase inhibitor valproic acid,the positive cell rates of ?-galactosidase staining was increased.CONCIUSION:(1)The expression ofHDAC2 was decreased in skeletal muscle cellsexposed to cigarette smoke extract,The longer the intervention time,the less HDAC2expression;(2)CSE promotes the senescence by reducing the expression of HD AC2 in skeletal muscle cells.