Guiqi Dingnian Prescription Delays Senescence of Mouse Mesangial Cells by Regulating JAK2/STAT Pathway Activity / 中国实验方剂学杂志

Objective:To investigate the effects of Guiqi Dingnian prescription （GDP） on the expression of related molecules in Janus tyrosine kinase 2 （JAK2）/signal transducer and activator of transcription （JAK2/STAT） signaling pathway of <italic>D</italic>-galactose （<italic>D</italic>-gal）-induced senescent mesangial cells. Method:The senescent mouse mesangial cells induced by 10 g·L^-1 <italic>D</italic>-gal were continuously treated with 40 mg·L^-1GDP for three days. The senescence of the treated cells was determined by senescence-associated （SA）-<italic>β</italic>-gal staining. The cell cycle was detected by flow cytometry. The cell viability was analyzed using the cell counting kit-8 （CCK-8）. The mRNA expression levels of tumor necrosis factor-<italic>α</italic> （TNF-<italic>α</italic>）， interleukin-6 （IL-6）， nuclear transcription factor-<italic>κ</italic>B （NF-<italic>κ</italic>B）， and IL-1<italic>α</italic> were detected by real-time polymerase chain reaction （Real-time PCR）. The protein expression levels of STAT1， phosphorylated STAT1 （p-STAT1）， STAT3， and p-STAT3 in the JAK2/STAT signaling pathway were determined by Western blot. Result:CCK-8 results showed that the optimal concentration of GDP was 40 mg·L^-1. Compared with the blank group， the positive rate of SA-<italic>β</italic>-gal in the model group was significantly higher（<italic>P</italic><0.01）， the percentage of cells in G₀/G₁ phase was significantly increased（<italic>P</italic><0.05）， the percentage of cells in G₂/M and S phase was significantly decreased（<italic>P</italic><0.01）. The mRNA expression levels of TNF-<italic>α</italic>，IL-6，NF-<italic>κ</italic>B and IL-1<italic>α </italic>were significantly increased（<italic>P</italic><0.01）. Compared with the model group， the model + GDP group exhibited significantly decreased SA-<italic>β</italic>-gal-positive cells （<italic>P</italic><0.05）， reduced cells in the G₀/G₁ phase （<italic>P</italic><0.05）， increased cells in the G₂/M and S phases （<italic>P</italic><0.01）， and down-regulated TNF-<italic>α</italic>， IL-6， NF-<italic>κ</italic>B， and IL-1<italic>α </italic>mRNA expression （<italic>P</italic><0.05） and STAT1， p-STAT1， STAT3， and p-STAT3 protein expression （<italic>P</italic><0.05）. Conclusion:GDP delays the senescence of mouse mesangial cells possibly by down-regulating the expression of related molecules in the JAK2/STAT pathway.

Role of periostin in PDGF-induced proliferation and extracellular matrix accumulation of mouse mesangial cells / 中国药理学通报

;Aim To investigate the role of periostin in PDGF-induced proliferation and extracellular matrix accumulation of mouse mesangial cells. Methods The effects of PDGF on periostin protein, cell proliferation and extracellular matrix accumulation were detected. The cells were collected at 0, 2,4, 6, 12 h after stimulation with PDGF(10 (ig • L " 1 ) to detect the expression of periostin, PCNA, FN and TGF-ßl by Western blot. The silencing effect of sh-periostin vector on periostin protein in mouse mesangial cells was identified by Western blot. Cells were randomly divided into control group, PDGF group, PDGF + sh-nc group and PDGF + sh-periostin group to detect the role of periostin in PDGF-induced proliferation and extracellular matrix accumulation of mouse mesangial cells. Results PDGF could elevate periostin protein expression. Western blot result showed that periostin protein expression in PDGF-stimulated groups was significantly higher than that in Oh group, which was consistent with the result of immunofluorescence. Positive expression of periostin was located in cytoplasm. Western blot result showed that PCNA, FN and TGF-ßl protein in PDGF-stimulated groups increased as compared with Oh group. shRNA vector aimed at periostin (sh-periostin vector) could partially reverse PDGF-induced mesangial cell proliferation and extracellular matrix expression. PCNA, Fn and TGF-ßl expressions were attenuated significantly. Conclusions PDGF can enhance periostin protein expression and increase mouse mesangial cell proliferation and extracellular matrix accumulation. Periostin shRNA vector can partially reverse PDGF-induced mesangial cell proliferation and extracellular matrix generation.

Influence of SRT1720 on apoptosis ofhigh glucose-induced mouse mesangial cells / 中国药理学通报

Aim To investigate the effect of Sirt1 activator SRT1720 on high glucose(HG)-induced apoptosis in mouse mesangial cells(MMCs).Methods Cultured mouse MMCs were divided into normal glucose group(NG),NG plus mannitol group(M),high glucose group(HG),HG plus SRT1720 group(HG+SRT).Apoptosis of MMCs was analyzed by DeadEndTM Fluorometric TUNEL System and flow cytometry.Reactive oxygen species(ROS)production was observed by flow cytometry.The expression levels of caspase-3,cleaved caspase-3,Bax,Bcl-2,p38 MAPK,p-p38 MAPK,p53,acetylated p53 and cytochrome C protein were observed by Western blot.The mRNA levels of Bax and Bcl-2 were detected by real-time PCR.Results Compared with normal glucose group,the production of ROS,the number of cell apoptosis,the expression of cleaved caspase-3,p-p38 MAPK and acetylated p53 and ratio of Bax/Bcl-2 were significantly increased,the expression of Sirt1 was decreased,meanwhile,the release of cytochrome C from mitochondria to cytoplasm was significantly increased in MMCs in high glucose group.Treatment with SRT1720 inhibited HG-induced increase of ROS production,cell apoptosis,expression of cleaved caspase-3,acetylated p53 and p-p38 MAPK,ratio of Bax/Bcl-2 and release of cytochrome C,and reversed HG-induced Sirt1 expression.Conclusion SRT1720 could prevent HG-induced apoptosis maybe by decreasing ROS production,preserving mitochondrial function and inhibiting p53 acetylation and activation of p38 MAPK in MMCs.