ABSTRACT
Objective·To investigate the mechanism of fructose-induced monocyte chemoattratant protein-1(MCP-1) production in HK-2 cells.Methods·The HK-2 cells were divided into fructose incubated (1,5 and 10 mmol/L) group,fructose and ketohexo-kinase inhibitor (KHK-IN) coincubation (fructose 5 mmol/L,KHK-IN was 12,100 and 1 000 nmol/L,respectively) group,uric acid incubation (5,15 and 50 mg/dL) group,fructose and allopurinol co-incubation (fructose 5 mmol/L,allopurinol were 0.01,0.1 and 0.5 mmol/L) group,uric acid and allopurinol co-incubation (uric acid 50 mg/dL,allopttrinol respectively 0.01,0.1and 0.5 mmol/L) group,H2O2 incubation (0.1 and 0.3 mmol/L) group,fructose and N-acetylcysteine (NAC) coincubation (fructose 5 mmol/L,NAC respectively 5,10 and 50 mmol/L) group,and uric acid and NAC co-incubation (uric acid 50 mg/dL,NAC was 5,10and 50 mmol/L,respectively) group.The quantitative PCR method and Western blotting method were used to observe the expression ofMCP-1 mRNA and protein.The effects of fructose and uric acid on the production of ROS in HK-2 cells were observed by using a fluorescent probe.Results·Fructose doseand time-dependently induced MCP-1 gene transcription and protein production in HK-2 cells,which could be blocked by the ketohexo-kinase blockers.Exogenous uric acid induced MCP-1 production in HK-2 cells.Allopurinol inhibited fructose,but not exogenous uric acid-induced MCP-1 expression.Both fructose and uric acid induced ROS generation.Incubation with H2O2promoted MCP-1 production in HK-2 cells.NAC completely inhibited MCP-1production induced by fructose and H2O2.Conclusion·Catalyzed by the ketohexo-kinase,fructose resultes the production of MCP-1 through uric acid and reactive oxygen species.
ABSTRACT
Objective·To explore the changes of pyruvate dehydrogenase (PDH) activity and pyruvate dehydrogenase kinase 4 (PDK4) expression in the end-stage renal disease (ESRD) patients' skeletal muscles. Methods·Skeletal muscle samples were collected from non-chronic kidney disease (non-CKD) patients and ESRD patients. PDH activity was detected by ELISA assay. Real-time qPCR was performed to examine gene transcription levels of PDK1-PDK4 and PDH subunits.Western blotting analysis was used to detect protein expression levels of PDK1 and PDK4. Results·There were no demographic differences between two groups of patients. Plasma creatinine and urea nitrogen were significantly elevated in ESRD group (both P<0.05), while estimated glomerular filtration rate, hemoglobin and plasma albumin in ESRD group were significantly lower than those in non-CKD group (all P<0.05).Skeletal muscle PDH activity in ESRD group was markedly lower than that in non-CKD group(P=0.014).There were no differences in PDK1-PDK4 and PDH subunits mRNA transcription levels between ESRD and non-CKD group.PDK4 protein expression was significantly higher than that in non-CKD group (P=0.000). Conclusion·The decreased PDH activity in ESRD patients' skeletal muscle may be related to up-regulation of PDK4.
ABSTRACT
Objective: To explore the changes of pyruvate dehydrogenase (PDH) activity and pyruvate dehydrogenase kinase 4 (PDK4) expression in the end-stage renal disease (ESRD) patients' skeletal muscles. Methods: Skeletal muscle samples were collected from non-chronic kidney disease (non-CKD) patients and ESRD patients. PDH activity was detected by ELISA assay. Real-time qPCR was performed to examine gene transcription levels of PDK1-PDK4 and PDH subunits. Western blotting analysis was used to detect protein expression levels of PDK1 and PDK4. Results: There were no demographic differences between two groups of patients. Plasma creatinine and urea nitrogen were significantly elevated in ESRD group (both P<0.05), while estimated glomerular filtration rate, hemoglobin and plasma albumin in ESRD group were significantly lower than those in non-CKD group (all P<0.05). Skeletal muscle PDH activity in ESRD group was markedly lower than that in non-CKD group (P=0.014). There were no differences in PDK1-PDK4 and PDH subunits mRNA transcription levels between ESRD and non-CKD group. PDK4 protein expression was significantly higher than that in non-CKD group (P=0.000). Conclusion: The decreased PDH activity in ESRD patients' skeletal muscle may be related to up-regulation of PDK4.