Influence Of Liraglutide On Endoplasmic Reticulum Stress During The Multiplication And Differentiation Of MC3T3-E1 Cells

Objective To identify the role of liraglutide on the multiplication and differentiation of the MC3T3-E1 cells,and to survey the correlation between liraglutide and endoplasmic reticulum stress.Methods 1.MC3T3-E1 osteoblasts were cultured in complete medium in vitro,and different concentrations of liraglutide were added(0,10-6,10-7,10-8,10-9mol/L),after 24 h Cell proliferation was detected with cell counting kit-8(CCK8).2.MC3T3-E1 osteoblasts were cultured in complete medium in vitro,and different concentrations of liraglutide were added(0,10-6,10-7,10-8,10-9mol/L),after 24 h Cell proliferation was detected with Ed U.3.MC3T3-E1 pre osteoblasts were cultured in osteogenic medium in vitro,and different concentrations of liraglutide were added(0,10-6,10-7,10-8,10-9mol/L),ALP activity was determined using the ALP kit after 48 h.4.MC3T3-E1 pre osteoblasts were cultured in osteogenic medium for 5 days.On the sixth day,MC3T3-E1 cells were treated with the optimal concentration of liraglutide(10-7mol/L),and the expression of BIP and XBP1 gene were measured by the RT-PCR.5.MC3T3-E1 pre osteoblasts were cultured in osteogenic medium for 5 days.On the sixth day,MC3T3-E1 cells were treated with the optimal concentration of liraglutide(10-7mol/L),and the expression of BIP and XBP1 protein was detected by Western blot.Results 1.Compared with the control group,the OD values of 10-6,10-7,10-8,10-9mol/L liraglutide groups increased respectively by 0.042,0.073,0.038,0.032(P<0.05).The OD values of 10-7mol/L liraglutide group increased most significantly.2.Ed U results showed that the nuclei of newly proliferated cells were stained by Ed U and showed red under fluorescence microscope.With the increase of liraglutide concentration,the number of red stained nuclei increased.The density of nucleus in 10-7 mol/L liraglutide group was the highest and the effect of promoting cell proliferation was the most significant.3.Compared with the control group,the ALP activity of 10-6,10-7,10-8,10-9mol/L liraglutide groups increased respectively by 0.400,0.717,0.321,0.303(P<0.05).Liraglutide(10-7mol/L)had the strongest effect.4.After Liraglutide(10-7mol/L)treatment,compared to unstimulated cells(time 0),the expression of BIP m RNA increased with time.BIP m RNA levels were not altered at 1h and 3h(P>0.05),increased significantly at 5h(1.983±0.230,P<0.05)and 10h(2.469±0.256,P<0.05).The expression of XBP1 m RNA was not altered at 1h(P>0.05),and increased significantly at 3h(3.449±0.246,P<0.05),decreasing at 5h(1.597±0.259,P<0.05),increased at 10 h again(2.412±0.274,P<0.05).5.After Liraglutide(10-7mol/L)treatment,compared to unstimulated cells(time 0),the expression of BIP protein increased with time.BIP protein levels were not altered at 1h and 3h(P>0.05),increased significantly at 5h(1.2512±0.2592,P<0.05)and 10h(1.4053±0.3151,P<0.05).The expression of XBP1 protein was not altered at 1h(P>0.05),and increased significantly at 3h(1.4347±0.1821,P<0.05),decreasing at 5h(1.0648±0.1744,P<0.05),increased at 10 h again(1.2183±0.1446,P<0.05).Conclusion 1.Liraglutide promote the proliferation of MC3T3-E1 cells.2.Liraglutide promote the differentiation of MC3T3-E1 cells.3.Liraglutide up-regulating the m RNA expression of BIP and XBP1.4.Liraglutide up-regulating the protein expression of BIP and XBP1.