Bletilla striata polysaccharides alleviate metabolic dysfunction-associated steatotic liver disease through enhancing hepatocyte RelA/ HNF1α signaling

Figure 11 RELA knockout partially abolishes the therapeutic effect of Bletilla striata polysaccharides on the sodium oleate/sodium palmitate-induced lipid degeneration in hepatocytes. A: RELA-wild-type (WT) and RELA-knockout (KO) HepG2 cells were randomized into three groups, including the sodium oleate/sodium palmitate (SO/SP) group (healthy rat serum + SO/SP incubated RELA-WT HepG2 cells); RELA-KO + SO/SP group (healthy rat serum + SO/SP incubated RELA-KO HepG2 cells); RELA-KO + Bletilla striata polysaccharides (BSP) + SO/SP group (conditional rat BSP serum + SO/SP incubated RELA-KO HepG2 cells). RELA-WT and RELA-KO HepG2 cells were stimulated with SO/SP in the medium containing 10% control rat serum or the conditional rat BSP serum for 48 hours; B: Oil red O staining of lipids in HepG2 cells; C: Intracellular triglycerides content; D: Cell counting kit-8 analysis of the viability of HepG2 cells; E: Malondialdehyde levels; F: Reduced glutathione levels; G: Western blot analysis of the relative levels of nuclear factor kappa B p65 and hepatocyte nuclear factor-1 alpha expression; H: Western blot analysis of the relative levels of endoplasmic reticulum stress-related protein expression; I: Western blot analysis of the relative levels of lipid metabolism-related protein expression; J: Western blot analysis of the relative levels of cleaved caspase-3, and mixed lineage kinase domain-like protein phosphorylation; K: Flow cytometry analysis of the frequency of apoptotic cells. Data are representative images or expressed as the mean ± SD of each group from three separate experiments. ^bP < 0.01. ¹P < 0.01 vs the sodium oleate/sodium palmitate group; ²P < 0.01 vs the RELA-knockout + sodium oleate/sodium palmitate group. KO: Knockout; WT: Wild type; BSP: Bletilla striata polysaccharides; TG: Triglycerides; MDA: Malondialdehyde; GSH: Glutathione; HNF1α: Hepatocyte nuclear factor-1 alpha; GRP78: 78-kDa glucose-regulated protein; CHOP: CCAAT-enhancer-binding protein homologous protein; HRD1: 3-hydroxy-3-methyl glutaryl coenzyme A reductase degradation 1 homolog; SREBP1c: Cleaved sterol regulatory element-binding protein 1; MTP: Microsomal triglyceride transfer protein; MCAD: Medium-chain acyl-CoA dehydrogenase; GPX4: Glutathione peroxidase 4; MLKL: Mixed lineage kinase domain-like protein; APC: Allophycocyanin conjugate; OD: Optical density; ATF4: Activating transcription factor 4; RelA: Nuclear factor kappa B p65.