Study On The Induction Of Osteoblasts By Gastrin Through PI3K/AKT Signaling Pathway

Objective To investigate the effect of(ghrelin)on osteoblast induction and the role of PI3K/AKT signaling pathway in osteoblast induction.Methods Mouse osteoblast precursor cells(MC 3T3-E1)were selected as the object of study,and the morphology and changes of the cells were observed under microscope.Different concentrations of ghrelin(0 ? mol / L,0.05 ? mol / L,0.10 ? mol / L)were added to the cell culture medium,and the proliferation activity of the cells was detected by CCK-8 kit.Detection of Alkaline Phosphatase activity by Alkaline Phosphatase Assay Kit;Western blot method to detect the expression of PI3 K,p-PI3 K,AKT,p-AKT in PI3K/AKT signaling Pathway Change.According to the experimental data obtained by the above methods,the statistical analysis software SPSS 22.0 was used to analyze the data,and the mapping software Graphpad Prism 5 was used to map the data.One-way ANOVA was used for comparison between groups.P < 0.05 was considered statistically significant.Results(1)Under the microscope,we can see that the cell adheres to the wall,showing irregular morphology such as spindles,triangles,polygons and so on,and the cells can connect with each other through dendrites of different length,which accords with the morphological characteristics of the growth of osteoblasts.Compared with the control group,the growth of the cells in the experimental group was faster than that in the control group.In a certain concentration range,the higher the concentration of ghrelin,the faster the growth of the cells.(2)CCK-8 was used to detect the proliferation activity of the cells.It was found that the proliferation of the cells was promoted by the addition of different concentrations of ghrelin compared with the control group(P<0.05),and within a certain range of concentration,the proliferation of the cells was increased with the addition of different concentrations of ghrelin.The effect of ghrelin on cell proliferation was also increased with the increase of the concentration gradient of ghrelin(P<0.05).In this experiment,0.10?mol/L of ghrelin was more effective in promoting cell proliferation.(3)The alkaline phosphatase activity of the cells was measured by alkaline phosphatase assay kit.It was found that the alkaline phosphatase activity of the cells in each group was higher than that of the control group(P<0.05),after adding different concentrations of hunger hormone,the alkaline phosphatase activity of the cells was higher than that of the control group.In a certain concentration range,the alkaline phosphatase activity increased with the increase of hungry concentration gradient(P<0.05).In this experiment,0.10?mol/L of ghrelin increased the alkaline phosphatase activity more obviously than that of the control group(P<0.05),and the alkaline phosphatase activity of the cells increased with the increase of the concentration gradient of ghrelin(P<0.05).(4)Western blot method was used to detect the expression of PI3 K,p-PI3 K,AKT,p-AKT in PI3K/AKT signaling pathway.It was found that after adding different concentrations of hunger hormone,compared with the control group,the expression of PI3 K,p-PI3 K,AKT,in each group was significantly higher than that in the control group.The expression of PI3K/AKT signaling pathway-related proteins,such as p-AKT,increased in a certain concentration range,and increased with the increase of the concentration gradient of ghrelin.Conclusion(1)Under the microscope,we can see that the cell adheres to the wall,showing irregular morphology such as spindles,triangles,polygons and so on,and the cells can connect with each other through dendrites of different length,which accords with the morphological characteristics of the growth of osteoblasts.(2)Ghrelin can promote the proliferation and differentiation of 3T3-E1 in osteoblasts,and within a certain concentration range,the proliferation and differentiation ability of osteoblasts increased with the increase of the concentration of ghrelin.(3)Hungry can affect the proliferation and differentiation of osteoblast 3T3-E1 through PI3K/AKT signaling pathway.At the protein level,the expression of PI3K/AKT signaling pathway-related protein PI3 K,p-PI3 K,AKT,p-AKT is related to the level of ghrelin concentration.In a certain concentration range,the protein expression increased with the increase of the concentration gradient of ghrelin.