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Yiqi Huoxue recipe ameliorates diabetic nephropathy by mediating VAPB-PTPIP51 complex to activate autophagy and
Linghao Dai1, Mengyi Wang2, Bo Wang3
1Jinhua Academy, Zhejiang Chinese Medical University, Jinhua, China.
Background:
Diabetic nephropathy (DN), a major complication of diabetes mellitus (DM), poses a high mortality and a global health burden. Mitochondria-associated endoplasmic reticulum membranes (MAMs) and mediated autophagy are regarded as the crucial factors in the development of DN. The Yiqi Huoxue recipe (YHR), a traditional Chinese medicine formula, has been reported to treat DN and regulate autophagy, while its underlying mechanism remains unclear.
Methods:
Firstly, UPLC-MS/MS analysis was performed to identify the chemical components of YHR. Then, C57BL/6 J mice were injected with streptozotocin and fed with high-fat diet to induce DN. YHR (7.8, 15.6 g/kg/d) was administered via intragastric gavage for 8 weeks. Biochemical parameters and oxidative stress indicators were measured; H&E, PAS, and immunohistochemistry staining of nephrin were performed. Mitochondrial Ca2+ levels were assessed by flow cytometry, while autophagosomes and MAMs were examined using transmission electron microscopy (TEM). The expression levels of VAPB, PTPIP51, LC3 II/I, P62 were detected by Western blot. Podocytes overexpressing PTPIP51 or VAPB were analyzed for cell activity using the CCK-8 assay, autophagy flux by TEM, and the expression of LC3 II/I and P62 by Western blot. In si-PTPIP51-transfected and high-glucose (HG)-stimulated podocytes, CCK-8 assay, PCR, TEM, and immunofluorescence staining were performed to detect the YHR-containing serum on cell activity, mtDNA, MAMs, autophagosomes and LC3 expression.
Results:
The chemical fingerprint of YHR was constructed and composed chemicals were identified. In DN mice, YHR treatment reduced the elevated fasting blood glucose (FBG), total cholesterol (TC), triglycerides (TG), blood urea nitrogen (BUN), serum creatinine (Scr), urinary albuminuria (ALB), and microscale albuminuria (MAU) levels. It also alleviated kidney and glomerulus damage, mitochondrial Ca2+, oxidative stress, MAM abnormal contact, and activated autophagy. The enhanced expression of MAM complex, VAPB-PTPIP51, were also inhibited in YHR-treated groups. The cell activity and autophagosome formation were significantly inhibited in podocytes overexpressing PTPIP51 (oe-PTPIP51) and VAPB (oe-VAPB). In contrast, in HG-podocytes, si-PTPIP51 promoted the cell activity, mtDNA copy number, MAM contact, autophagosomes formation and LC3 expression. More importantly, the addition of YHR-containing serum enhanced this effect.
Conclusion:
YHR may protect kidneys in DN by regulating the MAM complex VAPB-PTPIP51 to trigger autophagy, providing insights into TCM's clinical application and DN drug development.
Insights
Yiqi Huoxue recipe (YHR) protects against diabetic nephropathy (DN) by regulating mitochondria-associated endoplasmic reticulum membranes (MAMs) and autophagy. This traditional Chinese medicine may offer new therapeutic strategies for DN.
Area of Science:
- Nephrology
- Endocrinology
- Pharmacology
- Cell Biology
Background:
- Diabetic nephropathy (DN) is a severe complication of diabetes mellitus (DM), contributing to high mortality and global health challenges.
- Mitochondria-associated endoplasmic reticulum membranes (MAMs) and autophagy are critical in DN pathogenesis.
- The traditional Chinese medicine formula Yiqi Huoxue recipe (YHR) shows potential in treating DN and modulating autophagy, but its mechanism is not fully understood.
Purpose of the Study:
- To elucidate the underlying mechanisms of YHR in treating diabetic nephropathy.
- To investigate the role of mitochondria-associated endoplasmic reticulum membranes (MAMs) and autophagy in YHR's therapeutic effects on DN.
- To analyze the chemical composition of YHR and its impact on specific molecular targets.
Main Methods:
- Chemical fingerprinting of YHR using UPLC-MS/MS.
- Induction of DN in C57BL/6J mice using streptozotocin and high-fat diet, followed by YHR treatment.
- Assessment of biochemical parameters, oxidative stress, kidney damage, mitochondrial calcium levels, MAMs, and autophagy markers (LC3 II/I, P62).
- In vitro studies using podocytes to analyze the effects of YHR-containing serum, PTPIP51, and VAPB manipulation on cell activity, autophagy, and MAMs.
Main Results:
- YHR treatment significantly improved biochemical markers (FBG, TC, TG, BUN, Scr, ALB, MAU) and alleviated kidney damage in DN mice.
- YHR reduced mitochondrial Ca2+ overload, oxidative stress, and normalized MAMs, while activating autophagy.
- YHR inhibited the overexpression of the VAPB-PTPIP51 complex in podocytes and promoted cell activity and autophagy in high-glucose-stimulated podocytes with si-PTPIP51 transfection.
Conclusions:
- YHR protects kidneys in diabetic nephropathy by modulating the VAPB-PTPIP51 MAM complex, thereby triggering autophagy.
- These findings provide valuable insights into the clinical applications of traditional Chinese medicine (TCM) for DN.
- The study supports YHR as a potential therapeutic agent for DN drug development.
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