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The Role of TLR4 on PGC-1α-Mediated Oxidative Stress in Tubular Cell in Diabetic Kidney Disease
Shuguang Yuan1, Xuemei Liu1, Xuejing Zhu1
1Department of Nephrology, The Second Xiangya Hospital, Central South University, Changsha, Hunan, China.
Abstract:
The role and precise mechanism of TLR4 in mitochondria-related oxidative damage and apoptosis of renal tubules in diabetic kidney disease (DKD) remain unclear. We examined the expression of TLR4 in renal biopsy tissues. Db/db diabetic mice and HK-2 cells cultured under high glucose (HG) were used as in vivo and vitro models. Real-time RT-PCR, Western blot, and immunohistochemistry were performed to examine the mRNA and protein levels of TLR4, NF-κΒ, PGC-1α, cytochrome C, and cleaved caspase-3. ATP level, activity of electron transport chain complex III, and antioxidant enzymes were investigated for mitochondrial function. Electron microscopy (EM) and MitoTracker Red CMXRos were used for mitochondrial morphology alteration. DHE staining and TUNEL assay were detected for ROS accumulation and apoptosis. PGC-1α plasmids were used for the overexpression of PGC-1α in HK-2. TAK242 and parthenolide were used as TLR4 and NF-κB blockers, respectively. Results showed that TLR4 was extensively expressed in the renal tubules of DKD patients and db/db diabetic mice, which was positively related to the tubular interstitial damage score and urinary β-NAG levels. In diabetic mice, inhibition of TLR4 could reverse the decreased expression of PGC-1α, increased expression of cytochrome C and cleaved caspase-3, mitochondrial dysfunction and deformation, increased accumulation of ROS, and activation of tubular cell apoptosis. In vitro, inhibition of TLR4 or NF-κB showed consistent results. PGC-1α overexpression could reverse the mitochondrial dysfunction, increased cleaved caspase-3, and apoptosis in HK-2 cells treated with HG. Data indicated that the TLR4/NF-κB signaling pathway might be the upstream pathway of PGC-1α and promote the tubular damage of DKD by modulating the mitochondria-related oxidative damage and apoptosis.
Insights
Toll-like receptor 4 (TLR4) exacerbates diabetic kidney disease (DKD) by promoting mitochondrial dysfunction and apoptosis in renal tubules. Inhibiting TLR4 or NF-κB protects against DKD progression.
Area of Science:
- Nephrology
- Immunology
- Molecular Biology
Background:
- Diabetic kidney disease (DKD) involves renal tubule damage, but the precise role of Toll-like receptor 4 (TLR4) in mitochondria-related injury remains unclear.
- Mitochondrial dysfunction, oxidative stress, and apoptosis are key pathological features of DKD.
Purpose of the Study:
- To elucidate the mechanism of TLR4 in mitochondria-related oxidative damage and apoptosis of renal tubules in DKD.
- To investigate the relationship between TLR4 expression and renal tubule injury in DKD patients and animal models.
Main Methods:
- Examined TLR4 expression in DKD patient renal biopsies and db/db diabetic mice.
- Utilized HK-2 cells under high glucose (HG) conditions as an in vitro model.
- Assessed mRNA and protein levels of TLR4, NF-κB, PGC-1α, cytochrome C, and cleaved caspase-3 via RT-PCR, Western blot, and immunohistochemistry.
- Evaluated mitochondrial function (ATP, electron transport chain activity, antioxidant enzymes), morphology (EM), ROS accumulation (DHE), and apoptosis (TUNEL assay).
- Investigated the effects of TLR4/NF-κB inhibition (TAK242, parthenolide) and PGC-1α overexpression.
Main Results:
- TLR4 was highly expressed in renal tubules of DKD patients and diabetic mice, correlating with interstitial damage and urinary β-NAG levels.
- Inhibition of TLR4 in diabetic mice reversed PGC-1α downregulation, cytochrome C and cleaved caspase-3 upregulation, mitochondrial dysfunction, ROS accumulation, and apoptosis.
- In vitro studies showed similar protective effects with TLR4 or NF-κB inhibition, and PGC-1α overexpression mitigated HG-induced mitochondrial dysfunction and apoptosis.
Conclusions:
- The TLR4/NF-κB signaling pathway acts upstream of PGC-1α, driving renal tubule damage in DKD.
- TLR4 promotes DKD progression by modulating mitochondria-related oxidative damage and apoptosis in renal tubules.
- Targeting the TLR4/NF-κB pathway may offer a therapeutic strategy for diabetic kidney disease.
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