The Regulation And Mechanism Of PDZK1and It’s Related Members In Renal Tubular Epithelial Cells By Inflammation Signal

ObjectiveThis study is aimed to investigate the inflammatory signals’influence on gene transcription levels of PDZK1and its coupling proteins and protein expression levels of PDZK1and ABCG2, PDZK1protein subcellular localization, and explore the signaling pathways in human renal tubular epithelial cells(HK-2).MethodsInflammatory signals IL-1β、IL-18、IL-37、IL-10、TGF-p and IL-1β、combined IL-18or IL-37and IL-18combined IL-37stimulate HK-2cells by real time quantitative PCR (Real-time PCR) to detect mRNA expression levels of PDZK1、 ABCG2、 SLC22A12(URAT1、SLC22A13、 SLC5A12、 SLC22A11、 SLC17A1and SLC17A3, Western Blotto detect protein expression levels of ABCG2and PDZK1and immunofluorescence to detect the expression and subcellular localization of PDZK1 protein. Join inhibitor PDTC (NF-κB) or Wortmanin (PI3K) and then adding inflammatory stimuli, Western Blot to detect PDZK1protein expression changes of PDZK1and ABCG2in HK-2cells.Results1.IL-1β inhibit mRNA expression of SLC17A1、SLC17A3、SLC22A13、 SLC22A12、PDZK1、SLC22A11(P<0.05), IL-1β stimulation decrease SLC5A12and ABCG2s mRNA expression, but the difference is not statistically significant. IL-lp and IL-18or IL-37combined stimulation still decrease SLC17A1、SLC17A3、 SLC22A13、 SLC22A12、PDZK1、 SLC22A11mRNA expression, and it is not statistically significant with L-1β stimulation alone、 SLC17A1、 SLC17A3、 PDZK1mRNA expression decrease by TGF-P stimulation (P<0.05), which also caused SLC22A11、 ABCG2mRNA downregulated, but the difference is not statistically significant. IL-18、 IL-37、 IL-10regulate part of mRNA levels of uric acid pathway gene,but the function is far away from IL-1β and TGF-β.2. IL-1β、 TGF-Pdecrease protein expression of PDZK1,IL-18、IL-10、IL-37have no effect on protein expression of PDZK1,IL-1joint IL-18or IL-37also decrease protein expression of PDZK1IL-1βis found to promote ABCG2protein expression and IL-1β with IL-37or IL-18also promotes its expression, IL-37、 TGF-β、 IL-18、IL-10s function is not obviously3. PDZK1protein expression is significantly up-regulated by PDTC (NF-κB inhibitor) combined with IL-1β、 IL-1β+IL-18、IL-1β+IL-37stimulation, whereas PDTC combined with IL-37、IL-10downregulate PDZK1protein expression, and PDTC combined with IL-18、IL-18+IL-37makes PDZK1protein expression increase slightly; TGF-β has no effect on PDZK1protein expression. Wortmannin (PI3K/Akt inhibitor) has no influence on the expression of PDZK1protein expression. PDTC joined with IL-1β+IL-18、IL-1β+IL-37、IL-18+IL-37downregulate protein expression of ABCG2, whereas IL-18stimulation upregulate ABCG2protein expression,IL-1β、IL-37、IL-10and TGF-β have no effect. Wortmannin combined with IL-1β+IL-18. IL-1β+IL-37. IL-18+IL-37upregulate protein expression of ABCG2, Wortmannin combined with IL-18makes its downregulated, for Wortmannin combined with IL-1β、IL-37、IL-10and TGF-β make no sense.ConclusionIL-1p. TGF-β plays an important regulatory role in mRNA and protein expression of various uric acid channels gene, the function of IL-1β with IL-18or IL-37is consistent with IL-1β alone, IL-18、IL-37、IL-10have a moderate effect on part of mRNA expression of uric acid channels gene. Inflammatory signals are influence on the expression of PDZK1protein levels consistent with function in mRNA levels. However, IL-1β’s role on mRNA and protein expression of ABCG2is opposite. Inflammatory factors regulate protein expression of PDZK1is associated with NF-κB signaling pathway, because IL-1β inhibit the protein expression of PDZKland inhibit of NF-κB pathway significantly increased the expression of PDZK1, but the PI3K/Akt pathway has no effect on the protein expression of PDZK1. The NF-κB and PI3K/Akt pathways associate with the ABCG2protein expression with the former promoting the expression of it, and the latter inhibiting its expression.