5-Hydroxymethylfurfural and Isoverbascoside Alleviate Oxidative Damage INS-1 and MIN6 β-Cells by Activating Autophagy and Inhibiting Apoptosis

<b>Background/Objectives</b>: In type 2 diabetes (T2DM), dysregulated glucose and lipid metabolism impair cellular energy sensing and inhibit autophagy, leading to the accumulation of dysfunctional cellular components, increased inflammation and oxidative stress, and activation of the intrinsic apoptotic pathway. Prepared Rehmannia glutinosa is an anti-diabetic traditional Chinese medicine whose active monomers, including 5-Hydroxymethylfurfural (5-HMF) and isoverbascoside, exhibit potential antioxidant and anti-apoptotic effects. However, their role in β-cell protection remains unexplored. This study aims to investigate the protective mechanisms of 5-HMF and isoverbascoside against H₂O₂-induced oxidative damage in pancreatic β-cells. <b>Methods</b>: INS-1 and MIN6 β-cells were treated with 5-HMF and isoverbascoside (20 μM, 40μM) for 24 h under H₂O₂-induced oxidative stress. Multiple techniques were employed, including transcriptomics, proteomics, machine learning, Western blot analysis, and molecular docking. Flow cytometry and Hoechst 33342 staining were used to assess apoptosis, while autophagy was evaluated via LC3 fluorescence intensity and Beclin-1 expression. Chloroquine (CQ), an autophagy inhibitor, was applied to further examine autophagy's role. <b>Conclusions</b>: 5-HMF and isoverbascoside enhance autophagic activity in pancreatic β-cells, attenuate oxidative stress-induced apoptosis, and improve cell survival and proliferation. These findings underscore their potential as protective agents in T2DM by modulating the autophagy-apoptosis balance.