Carnosic Acid Protects INS-1 β-Cells against Streptozotocin-Induced Damage by Inhibiting Apoptosis and Improving Insulin Secretion and Glucose Uptake

Figures

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  Figure 1. Impact of CA or STZ treatment on cell viability and apoptosis in INS-1 cells. (A) CA chemical structure. (B) MTT assay in INS-1 cells preincubated with various concentrations of CA (2.5, 5, 10 µM) for 24 h compared to untreated cells. (C) Crystal violet staining for INS-1 cells. (D) MTT assay in STZ-treated cells (3 mM) or preincubated cells with 2.5 or 5 µM of CA for 24 h followed by STZ-treated versus untreated control cells. (E) Annexin V-PI analysis of apoptosis in untreated control cells, STZ-treated cells for 3 h, or cells pretreated with CA for 24 h prior to STZ exposure. Q1, Q2, Q3, and Q4 represent the necrotic, late apoptotic, early apoptotic, and viable cell populations. (F) Apoptotic index summarizes the apoptosis results. Data were obtained from three independent experiments. ** p < 0.01. Bars represent mean ± SD.

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  Figure 2. Impact of CA on insulin secretion, content, insulin expression in INS-1 cell treated with 3 mM STZ treatment. (A) Normalized stimulated insulin secretion was determined by ELISA in response to 2.8 and 16.7 mM glucose, respectively. (B) Normalized insulin content. (C) Western blot analysis of INS (Pro/Insulin), all in STZ-treated cells or cells preincubated for 24 h with CA followed by STZ treatment compared to control cells. β-actin was used as a reference control for Western blot. Data presented are obtained from three independent experiments. ** p < 0.01, and bars represent mean ± SD.

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  Figure 3. Impact of CA on (A) ROS production, (B) MDA levels, and (C) glucose uptake in STZ-treated cells. Levels of ROS and MDA were determined by fluorescence intensity, while glucose uptake assessment was measured by flow cytometry. Data presented are obtained from three independent experiments. ** p < 0.01, *** p < 0.001. Bars represent mean ± SD.

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  Figure 4. Effect of CA on protein expression. Western blot analysis of PI3K (A), AKT/p-AKT (B), PDX1 (C), BCL2 (D), BAX (E), BAD (F), and cleaved PARP1 (G) in STZ-treated cells for 3 h or cells preincubated with CA for 24 h followed by STZ treatment, compared to control cells. β-actin was used as an internal control. Data presented are obtained from three independent experiments. * p < 0.05 and ** p < 0.01. Bars represent mean ± SD.