Fig 1: Vitamin D suppresses the development of diabetic non-alcoholic fatty liver disease in db/db mice. Db/db mice were fed a standard chow containing various vitamin D doses (vitD def, vitD nor, and vitD Sup) for 12 weeks. (A) HE and Oil Red O staining of liver sections after treatment with VD. Scale bar = 50 μm. (B) Ultrastructural features of VD-treated mouse livers by transmission electron microscopy. Yellow arrows indicate autophagosomes (scale bar = 1 μm). Red arrows indicate fenestra (scale bar = 500 nm). (C) The protein levels of LN, PLVAP, autophagy- and pyroptosis-related markers were determined by Western blotting. (D) Representative immunohistochemical staining of autophagy- and pyroptosis-related protein in three groups (40×). (E,F) The body weight and liver weight. Results are means ± SD, * p < 0.05 vs. Vit D def group, # p < 0.05 vs. Vit D nor group. Abbreviations: VitD-def, Vitamin D deficient; VitD-nor, Vitamin D normal; VitD-sup, Vitamin D supplementation.
Fig 2: The effects of different concentrations of glucose on the expression of LN and PLVAP in HLSECs at different time points. (A,B) The expression of the hepatic sinus structural protein (PLVAP) and basement membrane protein (LN) following exposure to HG. The protein fraction was analyzed by Western blotting. * p < 0.05 compared with the control group. Abbreviations: NC, normal control; HG, high glucose; LN, laminin; PLVAP, plasmalemma vesicle-associated protein.
Fig 3: Inhibitory effect of vitamin D on HG-induced pyroptosis and hepatic sinusoidal capillarization of HLSECs. (A) Effects of MCC950 and vitamin D on pyroptosis-related markers, hepatic sinus structural protein (PLVAP), and basement membrane protein (LN) in HG-treated HLSECs. (B,C) The IL-1β and IL-18 content in HLSEC culture supernatants were determined by ELISA. (D) Lipid accumulation in HLSECs was assessed by Cell Navigator fluorescence. Scale bar = 50 μm. * p < 0.05 compared with the control group; # p < 0.05 vs. HG-treated group. Abbreviations: HG, high glucose; LN, laminin; PLVAP, plasmalemma vesicle-associated protein; NLRP3, NLR family pyrin domain containing 3; GSDMD, gasdermin D; IL, interleukin; Caspase, cysteinyl aspartate specific proteinase.
Fig 4: Vitamin D improves hepatic sinusoidal capillarization induced by high glucose. (A) Effects of vitamin D on cell viability in HLSECs. (B) The expression of LN and PLVAP proteins following exposure to HG and VD. The protein fraction was analyzed by Western blotting. (C) Effects of vitamin D on the HG-induced accumulation of lipids. (D) Ultrastructural features of HG-treated HLSECs by scanning electron microscopy. The red triangular mark represents the fenestra structures. * p < 0.05 compared with the control group; # p < 0.05 vs. HG-treated group. Abbreviations: NC, normal control; HG, high glucose; LN, laminin; PLVAP, plasmalemma vesicle-associated protein.
Fig 5: VD treatment induces autophagy and restores autophagic flux to alleviate hepatic sinusoidal capillarization. (A) HLSECs were cultured in the presence of HG with or without VD and treated with Baf-A1 for 24 h. The protein levels of autophagy-related markers, hepatic sinus structural protein (PLVAP), and basement membrane protein (LN) were determined by Western blotting. (B) Confocal microscopy observation of mRFP-GFP-LC3 adenovirus transfected HLSECs treated as indicated (magnification: ×40; scale bars = 20 μm). (C) Lipid accumulation in HLSECs was assessed by Cell Navigator fluorescence. Scale bar = 50 μm. * p < 0.05 compared with the control group; # p < 0.05 vs. HG-treated group; & p < 0.05 compared with the HG + Bafilomycin A1. Abbreviations: HG, high glucose; LN, laminin; PLVAP, plasmalemma vesicle-associated protein; LC3, microtubule-associated proteins light chain 3.
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