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Abstract: TH-PO766

PFKFB3 Downregulation Aggravates Angiotensin II-Induced Podocyte Detachment

Session Information

Category: Glomerular Diseases

  • 1403 Podocyte Biology

Authors

  • Chen, Zhaowei, Renmin Hospital of Wuhan University, Wuhan, China
  • Huang, Xiaoxiao, Renmin Hospital of Wuhan University, Wuhan, China
  • Ding, Guohua, Renmin Hospital of Wuhan University, Wuhan, China
Background

Podocytes are known to play a critical role in the maintenance of normal glomerular filtration. However, the exact mechanism of podocyte loss remains unclear. Fructose-2,6-biphosphatase 3 (PFKFB3) is a bifunctional enzyme and has been demonstrated to play a crucial role not only in glycolysis, cell proliferation, and cell survival but also in cell adhesion. The purpose of this study is to evaluate the role of PFKFB3 in Angiotensin II (Ang II)-induced kidney injury.

Methods

Immunohistochemical staining and immunofluorescence double staining techniques were employed to determine the localization of PFKFB3 in podocytes. Western blot analysis was performed to assess the expression levels of PFKFB3, p-talin1, and ITGB1 in glomeruli. The adhesive capacity of podocytes was assessed using an adhesion assay. Western blot was conducted to evaluate the expression levels of p-talin1 and ITGB1 in podocytes. Flow cytometry was employed to assess the impact of PFKFB3 on podocyte apoptosis. To examine the impact of PFKFB3 on podocyte adhesion under Ang II treatment, pEnCMV-PFKFB3 plasmid and PFKFB3 siRNA were transfected into podocytes.

Results

PAS and HE staining of mouse kidney sections revealed significant glomerulosclerosis in the Ang II-infused mice. Urinary protein examination demonstrated that the angiotensin II-infused mouse model exhibited higher levels of proteinuria. Immunofluorescence staining and immunofluorescence double staining revealed that Ang II inhibited the PFKFB3 expression in both the cytoplasm and nucleus of cultured human podocytes. Western blot experiments further confirmed that Ang II inhibited PFKFB3, p-talin1, and ITGB1 expression in glomeruli. In vitro study, adhesion assay demonstrated that Ang II stimulation of podocytes resulted in decreased adhesion compared to the control groups. Inhibition of PFKFB3 further enhanced the suppression of p-talin1 and ITGB1 expression in podocytes induced by Ang II. Additionally, stimulation of podocytes with Ang II suppressed PFKFB3, leading to podocyte apoptosis. All these effects were significantly exacerbated following transfection with PFKFB3 siRNA. However, the overexpression of PFKFB3 alleviated these effects.

Conclusion

These findings suggest that Ang II leads to the decrease in podocyte adhesion by suppressing PFKFB3 expression and indicates a potential therapeutic target for podocyte injury in CKD.