Effects of S1P/S1PR1 pathway on high glucose induced epithelial-mesenchymal transition of rat renal tubular epithelial cells by regulating ROS/NLRP3 / 中国药理学通报

Aim To explore the effect of S1P/S1PR1 signaling pathway on high glucose(HG)-induced epithelial-mesenchymal transition of rat renal tubular epithelial cells and its possible mechanism. Methods Cells were treated with different concentrations of glucose, and intracellular S1P expression was detected by ELISA and S1PR1 protein expression was detected by Western blot. The cells were divided into normal control group, HG group and HG + siS1PR1 group. The expression of E-cadherin, Vimentin, Fibronectin and Twist mRNA were detected by RT-qPCR and E-cadherin, α-SMA, Vimentin, NLRP3, ASC and NF-κB protein expression were detected by Western blot, and the levels of reactive oxygen species(ROS) were detected by flow cytometry. The cells were divided into normal control group, S1P group and S1P + siS1PR1 group. Vimentin, Snail, α-SMA, NLRP3, ASC and NF-κB protein expressions were detected by Western blot, and ROS levels were measured by fluorescence microscopy. Results ELISA results showed that the content of S1P in cells increased significantly under high glucose stimulation. Western blot results showed that S1PR1 protein expression was significantly higher at 30 mmol · L

Intermedin inhibits lipopolysaccharide-induced polarization of macrophages by activating AMPK signaling pathway / 中国药理学通报

Aim To evaluate the mechanism by which intermdin(IMD)inhibits lipopolysaccha ride(LPS)-induced polarization in RAW264.7 cells.Methods RAW264.7 cells were divided into control groups, LPS groups, LPS+IMD groups, LPS+IMD+Compound C groups.The mRNA expressions of tumor necrosis factor-α,(TNF-α,), CD86, inducible nitric oxide synthase(iNOS), Arginase-1(Arg-1)and CD206 were detected by Realtime-PCR.The protein expressions of p-AMPK, AMPK, TNF-α, intereukin-6(IL-6)and intereukin-10(IL-10)were detected by Western blot.The proportion of CD86+ M1 type cells was detected by Flow cytometry.In addition, the expression levels of supernatant cytokines, including IL-6 and TNF-α, were detected by ELISA.Results Compared with control and LPS groups, IMD treatment could up-regulate the expression level of p-AMPK and the ratio of p-AMPK/AMPK.LPS promoted M1 polarization, since the expressions of CD86, TNF-α and iNOS increased, while the expressions of CD206 and Arg-1 decreased by LPS induction.The proportion of M1 type cells increased and the secretion of TNF-α, IL-6 in the cell supernatant increased.And IMD treatment could inhibit the polarization of M1 induced by LPS.These effects were reversed by Compound C, an inhibitor of AMPK.Conclusion IMD can inhibit the M1-type polarization of LPS-induced macrophages by activating AMPK signaling pathway.

Resveratrol inhibits progression of IgA nephropathy in rats by inhibiting proliferation of mesangial cells and down regulating expression of PDGF-B and TGF-β1 / 中国药理学通报

Aim To investigate the therapeutic effect of SirTl agonist resveratrol on IgA nephropathy rats and its mechanisms. Methods An IgA nephropathy rat model was established. The rats were divided into four groups randomly: control group, IgA nephropathy group, control treated with Res group and IgA nephropathy treated with Res group. The urine protein was detected by Ponceau S; the biochemical indexes were detected by automatic biochemical analyzer; the pathological changes of kidney were observed by PAS and Masson staining; IgA deposition was observed by immunofluorescence; the expressions of PDGF-B and TGF-fil were detected by immunohistochemistry. Results Compared with IgA nephropathy group, the volume of 24-hour urinary protein and the expression of BUN and Scr in Res group decreased significantly, and the fluorescence of IgA in glomerulus was less in resveratrol group; mesangial cells and matrix proliferated and glomerular volume increased in IgA nephropathy group at the later stage, and both of them were significantly inhibited. Resveratrol could significantly reduce the high expression of PDGF-B and TGF-β1 in IgA nephropathy group. Conclusions Res can inhibit the deposition of IgA immune complex in mesangial region of IgA nephropathy rats and reduce glomerulosclerosis by down-regulating the expression of PDGF-B and TGF-β1, in turn it suppresses cell proliferation in mesangial region. It suggests that resveratrol plays an important role in slowing down the progression of IgA nephropathy.

Mechanisms of rapamycin on regulating rat mesangial cell proliferation and cell cycle through mTORC1/p70S6K pathways / 药学学报

To explore the affect and mechanisms of rapamycin on mesangial cell proliferation and cell cycle, rat mesangial cells (HBZY-1) were cultured and divided into the six groups: normal; normal with platelet derived growth factor (PDGF) 20 ng·mL-1; PDGF + rapamycin 1, 10, 100, 1 000 nmol·L-1. The cell proliferation was measured by MTT in 24 and 48 h; flow cytometry was used to detect the cell cycle phase. Western blot was performed to determine cyclin D1,cyclin E, cyclin-dependent kinase 2 (CDK2), cyclin-dependent kinase 4 (CDK4), p27, p70S6K/p-p70S6K protein expression. The p27 mRNA was detect by Real-time PCR. The results showed that rapamycin significantly suppressed PDGF induced glomerular mesangial cells (MCs) proliferation in a dose and time-dependent manner, but with the dose increased (1 to 1 000 nmol·L-1), the time dependence gradually weakened. Rapamycin inhibited mesangial cell proliferation and arrested the cell cycle in the G0/G1 phase. PDGF at 20 ng·mL-1 significantly increased the expression of cyclin D1, cyclin E and CDK2, CDK4 (P < 0.05), but rapamycin did not affect the expression of cyclin D1, cyclin E and CDK2, CDK4. Rapamycin can significantly inhibited p70S6K phosphorylation, up-regulated the expression of p27 protein and mRNA. Collectively, rapamycin has the effect of inhibiting the glomerular mesangial cells proliferation of mesangial cells by regulating the transcription of p27 mRNA, increasing its protein expression through the mTORC1/p70S6K pathway, resulting in decreased activity of cyclin-CDK, and blocking cell cycle in G0/G1 phase.

Inhibition of proliferation of breast cancer cells by down-regulation of cyclin d1 and survivin protein expression by erk inhibitor u0126 / 中国药理学通报

AimTo investigate the inhibitory effect of ERK signaling pathway inhibitor UO126 on breast cancer cell proliferation and explore its specific regulatory mechanism. Methods Cultured human breast cancer cell MCF-7, MDA-MB231, the cell were divided into different groups and intervened. MTT was used to measure the cell proliferation; Flow cytometry was employed to test cell cycle and cell apoptosis; Western blot was applied to test p-ERK/ERK, cyclin D1, survivin and cleaved caspase-3 protein expression. Results The results of MTT showed that the inhibitory rate of MCF-7 and MDA-MB231 cells increased significantly after U0126 intervention for 24 h and 48 h(P < 0.01). Cell cycle and apoptosis were detected by MCF-7 and MDA-MB231 cells treated with U0126 for 24 h. The proportion of cells in G0/G1 phase was significantly higher than that in control group, and the proportion of cells in S and G2 phase decreased(P < 0.05). The apoptotic rate in intervention group was significantly higher than that in non-intervention group, and the difference was significant(P < 0.01); U0126 treatment of human breast cancer cells could block ERK phosphorylation, increase cyclin D1 protein expression, inhibit survivin and up-regulate cleaved caspase3 expression, which were significantly different from control group(P < 0.05). Conclusions U0126 blocks ERK signaling pathway in MCF-7, MDA-MB231, down-regulates the cyclin D1, and blocks cell cycle in G0/G1 phase, then it inhibits the expression of survivin and increases cleaved caspase-3, promotes cell apoptosis, and inhibits the proliferation of breast cancer cell MCF-7, MDA-MB231.