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Published on: September 12, 2019
TRPC6 Inhibition Attenuates Renal Tubulointerstitial Fibrosis via the Reactive Oxygen Species/TXNIP/NLRP3 Signaling
Linting Wei1, Chenkai Cui1, Yan Li1
1Department of Nephrology, The Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, China.
Introduction:
Chronic kidney disease is a worldwide public health issue primarily characterized by glomerulosclerosis and the renal tubulointerstitial fibrosis. Recent studies have shown that TRPC6 is essential in renal interstitial fibrosis, although the precise mechanisms involved are not yet fully understood.
Methods:
UUO model was established using C57BL/6 male mice in which HK-2 cells were stimulated with TGF-β1. H&E and Masson staining were used to observe pathological changes. IHC staining was also conducted to measure the α-SMA, fibronectin (Fn), TRPC6, reactive oxygen species (ROS), and NLRP3 expressions. Scanning electron microscopy was used to observe morphological changes in the tubular cell membrane, and flow cytometry was utilized to measure ROS levels. In addition, Western blotting was performed to detect Fn, α-SMA, TRPC6, TXNIP, NLRP3, and the downstream pyroptosis-related molecule levels.
Results:
TRPC6 protein levels were enhanced in UUO mice and HK-2 cells upon TGF-β1 stimulation, which coincided with noticeable morphological changes associated with pyroptosis. Treatment with the TRPC6 inhibitor SAR7334 effectively reduced renal fibrosis markers and diminished levels of ROS, TXNIP, and proteins related to NLRP3-mediated pyroptosis (including NLRP3, cGSDMD, and IL-1β). Furthermore, application of the NLRP3 inhibitor MCC950 in HK-2 cells reinforced our findings, as it attenuated renal fibrosis-related proteins and counteracted the elevated levels of Fn, α-SMA, and NLRP3-mediated pyroptosis proteins observed in TGF-β1-stimulated HK-2 cells. Additionally, inhibiting TRPC6 appeared to dampen the activity of the ROS/TXNIP/NLRP3 pathway.
Conclusion:
TRPC6 may represent a promising target for mitigating renal interstitial fibrosis, potentially through its effects on the ROS/TXNIP regulatory pathway involving NLRP3-mediated pyroptosis.
Insights
Targeting TRPC6 shows promise in reducing kidney fibrosis by inhibiting the ROS/TXNIP/NLRP3 pathway and NLRP3-mediated pyroptosis. This offers a potential therapeutic strategy for chronic kidney disease.
Area of Science:
- Nephrology
- Molecular Biology
- Pathology
Background:
- Chronic kidney disease (CKD) is a global health concern, marked by glomerulosclerosis and renal tubulointerstitial fibrosis.
- Transient Receptor Potential Canonical 6 (TRPC6) plays a role in renal interstitial fibrosis, but its exact mechanisms remain unclear.
Purpose of the Study:
- To investigate the role of TRPC6 in renal interstitial fibrosis.
- To elucidate the mechanisms linking TRPC6 to fibrosis, focusing on oxidative stress and pyroptosis.
Main Methods:
- Utilized a unilateral ureteral obstruction (UUO) mouse model and TGF-β1-stimulated HK-2 cells.
- Employed histological staining (HE, Masson), immunohistochemistry (IHC), scanning electron microscopy (SEM), flow cytometry, and western blotting.
- Assessed expressions of α-SMA, Fibronectin (Fn), TRPC6, ROS, TXNIP, NLRP3, and pyroptosis markers.
Main Results:
- Elevated TRPC6 levels were observed in UUO mice and TGF-β1-treated HK-2 cells, correlating with pyroptosis.
- TRPC6 inhibition reduced fibrosis markers, reactive oxygen species (ROS), TXNIP, and NLRP3-mediated pyroptosis proteins.
- NLRP3 inhibition also attenuated fibrosis and pyroptosis, confirming the pathway's involvement.
Conclusions:
- TRPC6 inhibition mitigates renal interstitial fibrosis.
- The ROS/TXNIP/NLRP3 pathway and NLRP3-mediated pyroptosis are key mechanisms in TRPC6-induced fibrosis.
- TRPC6 is a potential therapeutic target for treating renal interstitial fibrosis.
