ABSTRACT
@#BACKGROUND: To determine the protective role of mesencephalic astrocyte-derived neurotrophic factor (MANF) in regulating sepsis-associated acute kidney injury (S-AKI). METHODS: A total of 96 mice were randomly divided into the control group, control+MANF group, S-AKI group, and S-AKI+MANF group. The S-AKI model was established by injecting lipopolysaccharide (LPS) at 10 mg/kg intraperitoneally. MANF (200 μg/kg) was administered to the control+MANF and S-AKI+MANF groups. An equal dose of normal saline was administered daily intraperitoneally in the control and S-AKI groups. Serum and kidney tissue samples were obtained for biochemical analysis. Western blotting was used to detect the protein expression of MANF in the kidney, and enzyme-linked immunosorbent assay (ELISA) was used to determine expression of MANF in the serum, pro-inflammatory cytokines (tumor necrosis factor-α [TNF-α] and interleukin-6 [IL-6]). Serum creatinine (SCr), and blood urea nitrogen (BUN) were examined using an automatic biochemical analyzer. In addition, the kidney tissue was observed for pathological changes by hematoxylin-eosin staining. The comparison between two groups was performed by unpaired Student’s t-test, and statistics among multiple groups were carried out using Tukey’s post hoc test following one-way analysis of variance (ANOVA). A P-value <0.05 was considered statistically significant. RESULTS: At the early stage of S-AKI, MANF in the kidney tissue was up-regulated, but with the development of the disease, it was down-regulated. Renal function was worsened in the S-AKI group, and TNF-α and IL-6 were elevated. The administration of MANF significantly alleviated the elevated levels of SCr and BUN and inhibited the expression of TNF-α and IL-6 in the kidney. The pathological changes were more extensive in the S-AKI group than in the S-AKI+MANF group. CONCLUSION: MANF treatment may significantly alleviate renal injury, reduce the inflammatory response, and alleviate or reverse kidney tissue damage. MANF may have a protective effect on S-AKI, suggesting a potential treatment for S-AKI.
ABSTRACT
Objective @#To investigate the effect of mesencephalic astrocyte-derived neurotrophic factor (MANF) on the adaptive expression of bile acid transporter in human hepatoellular carcinomas (HepG2) induced by rifampicin (RFP) ．@*Methods@#The control group cell line (Y07) and the knockdown group cell line (Y25) were constructed by lentiviral stable transfection technology．The Y07 and Y25 cells were treated with RFP of 200 μmol / L for 48 h， and qRT-PCR and Western blot were used to detect the protein and gene expression levels of MANF，bile salt export pump ( BSEP) ，multidrug resistance-related proteins 2 /3 /4 ( MRP2 ，MRP3 ，MRP4) ，multidrug resistance protein 1 (MDR1) ，organic solute transporter a / β ( OSTα/ β) ，organic anion transporter ( OATP2B1) ．The protein and gene expression levels of proliferating cell nuclear antigen ( PCNA) ，proliferating cell marker Ki67 were used to evaluate the proliferation of cells in each group changes in levels．Changes in the protein and gene expression levels of C / EBP homologous protein( CHOP) and cysteinyl aspartate specific proteinase-3 ( Caspase-3) were used to evaluate the apoptosis of cells in each group．The relative contents of alanine aminotransferase(ALT) ，aspartate aminotransferase(AST) ，alkaline phosphatase ( ALP) ，total bilirubin ( TBIL) ，indirect bilirubin ( IBIL) and total bile acid(TBA) in the supernatant of cell culture medium of each group were detected by kits. @*Results@#RFP could induce the protein and gene expression of MANF，BSEP ，MRP2 ，MRP3 ，MRP4 ，MDR1 ，OSTα , OSTβ , OATP2B1 in HepG2 cells (P ＜0. 05 ) ，while the protein and gene expression levels of BSEP ，MRP2 ，MRP3， MRP4，MDR1，OSTα、OSTβ、OATP2B1 decreased after MANF knockdown(P＜0. 05) ．Moreover，under the action of RFP，the protein expression of PCNA and Ki67 in the knockdown group was still higher．The protein and gene levels of CHOP and Caspase-3 significantly increased after MANF knockdown(P＜0. 05) ．The levels of the hepatic cell injury markers in the cell supernatant increased significantly(P＜0. 05) ．@*Conclusion @#RFP can induce the expression of bile acid transporter such as BSEP，MRP2，MRP3，MRP4，MDR1，OSTα , OSTβ and OATP2B1 to increase in HepG2 cells(P＜0. 05) ，but the expression of bile acid transporter of HepG2 after MANF knockdown will significantly decrease under the induction of rifampicin(P＜0. 05) ，and cell indury is aggravated，indicating that MANF plays a protective role in RFP-induced adaptive responses by regulating the bile acid transporter.
ABSTRACT
The mesencephalic astrocyte-derived neurotrophic factor (MANF) has been recently identified as a neurotrophic factor, but its role in hepatic fibrosis is unknown. Here, we found that MANF was upregulated in the fibrotic liver tissues of the patients with chronic liver diseases and of mice treated with CCl4. MANF deficiency in either hepatocytes or hepatic mono-macrophages, particularly in hepatic mono-macrophages, clearly exacerbated hepatic fibrosis. Myeloid-specific MANF knockout increased the population of hepatic Ly6Chigh macrophages and promoted HSCs activation. Furthermore, MANF-sufficient macrophages (from WT mice) transfusion ameliorated CCl4-induced hepatic fibrosis in myeloid cells-specific MANF knockout (MKO) mice. Mechanistically, MANF interacted with S100A8 to competitively block S100A8/A9 heterodimer formation and inhibited S100A8/A9-mediated TLR4-NF-κB signal activation. Pharmacologically, systemic administration of recombinant human MANF significantly alleviated CCl4-induced hepatic fibrosis in both WT and hepatocytes-specific MANF knockout (HKO) mice. This study reveals a mechanism by which MANF targets S100A8/A9-TLR4 as a "brake" on the upstream of NF-κB pathway, which exerts an impact on macrophage differentiation and shed light on hepatic fibrosis treatment.
ABSTRACT
Objective To investigate protective effects of dexmedetomidine on oxygen-glucose deprivation/reperfusion(OGD/R)-induced neuronal apoptosis.Methods SH-SY5Y cells were differentiated to neurons with ATRA and followed by TPA.According to the results of preliminary experiment, OGD/R modle was constructed by oxygen-glucose deprivation(OGD) for 12 h and reperfusion(R) for another 12 h.During the start of the OGD, neurons were immediately divided into six groups: group D0(0 μmol/L dexmedetomidine), group D1(0.1 μmol/L dexmedetomidine), group D2 (1 μmol/L dexmedetomidine), group D3 (10 μmol/L dexmedetomidine), group D4(100 μmol/L dexmedetomidine), group D5 (1 000 μmol/L dexmedetomidine).After reperfusion 12 h, the cell viability was evaluated by the method of MTT.The cellular apoptosis was observed by flow cytometry method.The protective effects of different concentration dexmedetomidine on OGD/R-induced neuronal apoptosis were investigated.Then in chosen the exact group having protective effects, endoplasmic reticulum stress specific protein mesencephalic astrocyte-derived neurotrophic factor (MANF) and pro-apoptotic protein Caspase-3 and CHOP were detected by Westernblot method.Results Compared with group D0, there was no difference on the cell viability and cellular apoptosis induced by OGD/R in groups D1 and D2, but a significant decrease and increase in groups D4 and D5 (P<0.01 or P<0.05).And only group D3 had a neuroprotective effect, significantly increased the cell viability and inhibited the apoptosis (P<0.01).Further studys found that group D3 significantly up-regulated ER stress specific protein MANF (P<0.01) and inhibited up-regulation of Caspase-3 and CHOP (P<0.01).Conclusion These data suggest that 10 μmol/L dexmedetomidine had neuroprotective effect on OGD/R-induced neuronal apoptosis and significantly increased cell viability.Our results also indicate that up-regulation of ER stress specific protein MANF and inhibition of CHOP and Caspase-3 by MANF are involved in the neuroprotective effects of Dexmedetomidine.