AdipoRon ameliorates vascular endothelial injury in type 2 diabetes mellitus and is associated with suppression of the NLRP3-Caspase-1-GSDMD pyroptosis axis

Figures

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  Fig. 1. Effect of AdipoRon on HG-induced HUVEC injury and NLRP3 inflammasome activation. After treatment of HUVECs with HG containing various concentrations of AdipoRon (0, 0.5, 1, 10, and 100 µg/mL), A cell viability was measured by CCK-8 assay; B ROS production was assessed via flow cytometry analysis; C the protein levels of IL-1β, NLRP3, ASC, pro-caspase-1 and cleaved-caspase-1 were measured by western blotting. Horizontal lines denote statistical comparisons between the two connected groups. Asterisks indicate the level of significance: *P < 0.05, **P < 0.01, ***P < 0.001

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  Fig. 2. AdipoRon inhibits NLRP3-Caspase-1-GSDMD axis-mediated pyroptosis in HG-treated HUVECs. After treatment of with HG containing AdipoRon (10 µg/mL) or Ac-YVAD-CMK (50 µM), A cell viability was measured by CCK-8 assay; B ROS production was assessed via flow cytometry analysis; C the protein levels of IL-1β, NLRP3, ASC, Nrf2, pro-caspase-1, cleaved-caspase-1, GSDMD and p-NF-κB p65 were measured by western blotting; D caspase-1 activity, E LDH concentration, F ET-1 and NO concentrations were assessed using biochemistry analysis; G immunofluorescent test was performed to calculate GSDMD expression; H tube length of HUVECs was measured through tube formation assay; I the apoptosis of HUVECs was assessed by TUNEL assay. Scale bar = 100 μm. Horizontal lines denote statistical comparisons between the two connected groups. Asterisks indicate the level of significance: *P < 0.05, **P < 0.01, ***P < 0.001

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  Fig. 3. AdipoRon inhibits NLRP3-Caspase-1-GSDMD axis-mediated pyroptosis in T2DM mice. After administration with AdipoRon-L (5 mg/kg), AdipoRon-H (20 mg/kg) or MCC950 (10 mg/kg) in T2DM mice, A fasting blood glucose levels were measured; B homeostasis model assessment of insulin resistance (HOMA-IR) was calculated using the following formula: HOMA-IR = (fasting blood glucose × fasting serum insulin)/22.5; C HE staining was performed to the pathological changes in artery blood vessels; D the protein levels of IL-1β, NLRP3, ASC, Nrf2, pro-caspase-1, cleaved-caspase-1, GSDMD and p-NF-κB p65 were measured by western blotting; E-F immunofluorescent test was performed to calculate GSDMD expression; G-I the levels of MDA, SOD and GSH-Px were assessed using biochemistry analysis; J TUNEL staining assay was performed to assess arterial endothelial cell apoptosis. Scale bar = 100 μm. Horizontal lines denote statistical comparisons between the two connected groups. Asterisks indicate the level of significance: *P < 0.05, **P < 0.01, ***P < 0.001