Fig. 3: Overexpression of SLC25A51 in hepatocytes increases mitochondrial NAD+, promotes liver regeneration and maintains mitochondrial respiratory capacity post injury.
From: Hepatocyte mitochondrial NAD+ content is limiting for liver regeneration
Male mice were infected with AAV-expressing human SLC25A51 (OE) or eGFP under the control of the TBG promoter and studied 3–4 weeks post infection. a, The total SLC25A51 transcript in liver (primers designed in a homologous region of human and mouse coding sequence (CDS)), (n = 8 for eGFP and n = 11 for A51 OE). b,c, The body weights (b, n = 10 for eGFP and n = 12 for A51 OE) and the liver weights (c, n = 10 for eGFP and n = 11 for A51 OE). d,e, The fasting and refed blood glucose (d, n = 10 for fasting GFP and n = 13 for fasting A51 OE; n = 6 for refed for both eGFP and A51 OE) and insulin (e, n = 5 for GFP and n = 7 for A51 OE) levels. f, The enrichment analysis of the liver transcripts. g,h, The untargeted proteomics (g) and metabolomics (h) profiling from liver tissue. The changes with a nominal P value less than 0.05 are coloured in red (upregulated) or in green (downregulated). For untargeted proteomics profiling, n = 3–6 per group, LC–MS experiments for metabolomics, n = 6–8 per group. i, The subcellular fractionation with stable isotope labelling by NAD-SILEC scheme in HAP1 cells j, The adjusted NAD+ concentrations in cellular compartments by NAD-SILEC (n = 6 for eGFP and n = 7 for A51 OE). The data for the mitochondrial fraction were analysed using a one-tailed t-test. Two-tailed t-tests were performed for the other subcellular fractions. k, The SLC25A51-OE and eGFP male mice were subjected to two thirds PHx and analysed 48 h later. l, The liver-to-body-weight ratio 48 h post PHx (n = 10 for eGFP and n = 11 for A51 OE). m, The representative regenerating liver sections stained in H&E and Ki-67 at 40× showing mitotic figures (in white arrows) and micro- and macrovesicular fatty changes. n, The mitosis as determined by counting mitotic figures in hepatocytes under high power in H&E sections (n = 9 for both eGFP and A51 OE). o, The percent of Ki-67 positively stained cells (n = 6 for both eGFP and A51 OE). p, The hepatic TG content (n = 10 for eGFP pre and n = 8 for post PHx; n = 13 for A51 OE pre and n = 9 for post PHx). q, The hepatic ATP content (n = 7 for eGFP, n = 8 for A51 OE pre and n = 7 for post PHx). r,s, The total liver NAD+ (n = 10 for eGFP pre and n = 9 for post PHx; n = 13 for A51 OE pre and n = 8 for post PHx) (r) and NADP(H) (n = 4–5 for eGFP and n = 6–7 for A51 OE) (s) content. t, The mitochondrial NAD+ and NADH content (n = 21 for eGFP pre NAD(H); n = 22 and 21 for A51 OE NAD(H), respectively; n = 8 and 4 for eGFP NAD(H) post, respectively; n = 11 and 7 for A51 OE NAD(H) post, respectively). The violin plots show individual data points and the median. u–w, The state three-coupled respiration in mitochondria from resected (u) and regenerated (v) livers, using complex I (CI, pyruvate), complex II (CII, succinate) and complex IV (CIV, TMPD-ASC)-dependent substrates (n = 9–10 for CI, n = 7 for CII and CIV in eGFP; n = 11 for A51 OE). w, The mitochondrial oxygen consumption using fatty acid oxidation substrates (n = 5 for both genotypes). The data are presented as the mean, and the error bars represent s.e.m. The ANOVA results are shown as the main effects of genotype ($), time ($) and interaction (&). The individual P values are shown where significant (P ≤ 0.05). The tests for normality and equality of variances, along with the resulting statistical test selected for each panel, are provided in the statistical source data. 10HPF, ten high-power fields; O2 flux(mt), mitochondrial oxygen consumption flux. Panel i created with BioRender.com.
