Triptolide inhibits NLRP3 inflammasome activation and ameliorates podocyte epithelial-mesenchymal transition induced by high glucose / 中国中药杂志

The aim of this paper was to explore the effects of triptolide( TP),the effective component of Tripterygium wilfordii on improving podocyte epithelial-mesenchymal transition( EMT) induced by high glucose( HG),based on the regulative mechanisms of Nod-like receptor protein 3( NLRP 3) inflammasome in the kidney of diabetic kidney disease( DKD). The immortalized podocytes of mice in vitro were divided into the normal( N) group,the HG( HG) group,the low dose of TP( L-TP) group,the high dose of TP( HTP) group and the mannitol( MNT) group,and treated by the different measures,respectively. More specifically,the podocytes in each group were separately treated by D-glucose( DG,5 mmol·L~(-1)) or HG( 30 mmol·L~(-1)) or HG( 30 mmol·L~(-1)) + TP( 5 μg·L~(-1))or HG( 30 mmol·L~(-1)) + TP( 10 μg·L~(-1)) or DG( 5 mmol·L~(-1)) + MNT( 24. 5 mmol·L~(-1)). After the treatment of HG or TP at 24,48 and 72 h,firstly,the activation of podocyte proliferation was investigated. Secondly,the protein expression levels of the epithelial markers in podocytes such as nephrin and ZO-1,the mesenchymal markers such as collagen Ⅰ and fibronectin( FN) were detected,respectively. Finally,the protein expression levels of NLRP3 and apoptosis-associated speck-like protein( ASC) as the key signaling molecules of NLRP3 inflammasome activation,as well as the downstream effector proteins including caspase-1,interleutin( IL)-1β and IL-18 were examined,severally. The results indicated that,for the cultured podocytes in vitro,HG could cause the low protein expression levels of nephrin and ZO-1,induce the high protein expression levels of collagen Ⅰ and FN and trigger podocyte EMT. Also HG could cause the high protein expression levels of NLRP3,ASC,caspase-1,IL-1β and IL-18 and induce NLRP3 inflammasome activation. On the other hand,the co-treatment of TP( L-TP or H-TP) and HG for podocytes could recover the protein expression levels of nephrin and ZO-1,inhibit the protein expression levels of collagen Ⅰ and FN and ameliorate podocyte EMT. Also the co-treatment of TP( L-TP or H-TP) and HG could down-regulate the protein expression levels of NLRP3 and ASC,inhibit NLRP3 inflammasome activation and reduce the protein expression levels of the downstream effector molecules including caspase-1,IL-1β and IL-18. On the whole,HG could activate NLRP3 inflammasome and induce podocyte EMT in vitro. TP at the appropriate dose range could inhibit NLRP3 inflammasome activation and ameliorate podocyte EMT,which may be one of the critical molecular mechanisms of TP protecting againstpodocyte inflammatory injury in DKD.

Exercise Training in Hypoxia Prevent Hypoxia Induced NLRP3 Inflammasome Activation in Skeletal Muscles / 中国运动医学杂志

Objective To observe the effect of exercise training in hypoxia on the activation of NL-RP3 inflammasomes in skeletal muscles,and to explore the role of nuclear factor(erythroid-derived 2)-like 2 (Nrf2).Methods Thirty-two male Sprague-Dawley rats were randomly divided into a normoxia control (NC) group,a normoxia training (NT) group,a hypoxia control (HC) group and a hypoxia training (HT) group,each of 8.The hypoxia animals were housed in normobaric hypoxic tent with oxygen content of 11.3％ for 4 weeks consecutively.The animals exercised on a motor-driven rodent treadmill of a 5％ slope at a speed of 15 m/min for 60 min/day,5 days/week.Following the last hypoxia exposure,all rats were sacrificed by decapitation and their quadriceps were removed.Their muscle mitochondria were extracted using the differential velocity centrifugation.The generation of mitochondrial reactive oxygen species(ROS)was detected using the dichlorofluorescein method.The 8-oxodG in the mitochondral DNA was measured using the high-performance liquid chromatography.The interleukin-1β(IL-13)content and Caspase-1 relative activity of muscles was determined using the enzyme-linked immune sorbent assay and colorimetric assay.Western blotting was employed to detect the expression of NLRP3,ASC,NF-E2-related factor 2(Nrf2)and quinone oxidase reductase 1(NQO1)protein.Results The ROS generation and the level of 8-oxodG in mtDNA,Caspase-1 activity and IL-1β content,as well as NLRP3 and ASC protein expression in skeletal muscles in group HC were significantly higher than group NC(P＜0.01 for all),while the Nrf2 and NQO1 protein expression of the former were significantly lower than the latter(P＜0.05,P＜0.01).The ROS generation and the level of 8-oxodG in mtDNA,Caspase-1 activity and IL-13 content,as well as NLRP3 and ASC protein expression in skeletal muscle in group HT were significantly lower than group HC(P＜0.05,P＜0.01),while the Nrf2 and NQO1 protein expression of the former were significantly higher than the latter(P＜0.01 for all).Conclusion Exercise training in hypoxia can prevent hypoxia induced NLRP3 inflammasome activation in skeletal muscle via Nrf2-dependent pathways to suppress ROS generation and elevate the expression of antioxidant enzymes.