Activation and functional changes of astrocytes cultured in vitro under inflammatory stimulation
Objective To observe the activation and functional changes of astrocytes cultured in vitro under inflammatory stimulation.Methods Astrocytes from the cortex of C57BL/6 mice were isolated and cultured in vitro,the changes of astrocyte morphology were observed by immunocytochemistry and the expression levels of proinflammatory factors(iNOS,IL-6,TNF-α)and anti-inflammatory factors(ARG1,IL-10,TGF-β1)were determined by Western blot protein analysis.Results After inflammatory stimulation,immunofluorescence cell staining showed that astrocytes in M1(LPS+IFN-γ)group were abnormally activated,cell body swelling,enlargement,distortion and other morphological changes were also obvious,and the number of GFAP positive astrocytes in M1 group were evidently increased(P＜0.001 vs.control).CCK-8 test showed that compared with the control group,the absorbance values of LPS group,IFN-γ group and M1 group were increased,especially the absorbance values of M1 group was significantly increased(P＜0.001 vs.control).Western blot results showed that the expression levels of iNOS,IL-6 and TNF-α in M1 group were markedly increased after LPS and IFN-γ combined stimulation(P＜0.001 vs.control).The expression levels of anti-inflammatory factors(ARG1,IL-10 and TGF-β1)in each group were also significantly upregulated(P＜0.01 or 0.001 vs.control).Conclusions Under inflammatory stimulation,the cultured-astrocytes in vitro can be abnormally activated,its morphological and functional changes are distinct,and the expression level of Al-like inflammatory factors in astrocytes are increased.
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