Background: One in eleven people is affected by chronic kidney disease, a condition characterized by kidney fibrosis and progressive loss of kidney function. Epidemiological studies indicate that adverse intrauterine and postnatal environments have a long-lasting role in chronic kidney disease development. Epigenetic information represents a plausible carrier for mediating this programming effect. Here we demonstrate that genome-wide cytosine methylation patterns of healthy and chronic kidney disease tubule samples obtained from patients show significant differences. Results: We identify differentially methylated regions and validate these in a large replication dataset. The differentially methylated regions are rarely observed on promoters, but mostly overlap with putative enhancer regions, and they are enriched in consensus binding sequences for important renal transcription factors. This indicates their importance in gene expression regulation. A core set of genes that are known to be related to kidney fibrosis, including genes encoding collagens, show cytosine methylation changes correlating with downstream transcript levels. Conclusions: Our report raises the possibility that epigenetic dysregulation plays a role in chronic kidney disease development via influencing core pro-fibrotic pathways and can aid the development of novel biomarkers and future therapeutics.

Epigenetic changes might provide the biological explanation for the long-lasting impact of metabolic alterations of diabetic kidney disease development. Here we examined cytosine methylation of human kidney tubules using Illumina Infinium 450 K arrays from 91 subjects with and without diabetes and varying degrees of kidney disease using a cross-sectional design. We identify cytosine methylation changes associated with kidney structural damage and build a model for kidney function decline. We find that the methylation levels of 65 probes are associated with the degree of kidney fibrosis at genome wide significance. In total 471 probes improve the model for kidney function decline. Methylation probes associated with kidney damage and functional decline enrich on kidney regulatory regions and associate with gene expression changes, including epidermal growth factor (EGF). Altogether, our work shows that kidney methylation differences can be detected in patients with diabetic kidney disease and improve kidney function decline models indicating that they are potentially functionally important.

Genetic variants at the SORT1 locus in humans, which cause increased SORT1 expression in the liver, are significantly associated with reduced plasma levels of LDL cholesterol and apolipoprotein B (apoB). However, the role of hepatic sortilin remains controversial, as genetic deletion of sortilin in mice has resulted in variable and conflicting effects on apoB secretion. Here, we found that Sort1-KO mice on a chow diet and several Sort1-deficient hepatocyte lines displayed no difference in apoB secretion. When these models were challenged with high-fat diet or ER stress, the loss of Sort1 expression resulted in a significant increase in apoB-100 secretion. Sort1-overexpression studies yielded reciprocal results. Importantly, carriers of SORT1 variant with diabetes had larger decreases in plasma apoB, TG, and VLDL and LDL particle number as compared with people without diabetes with the same variants. We conclude that, under basal nonstressed conditions, loss of sortilin has little effect on hepatocyte apoB secretion, whereas, in the setting of lipid loading or ER stress, sortilin deficiency leads to increased apoB secretion. These results are consistent with the directionality of effect in human genetics studies and suggest that, under stress conditions, hepatic sortilin directs apoB toward lysosomal degradation rather than secretion, potentially serving as a quality control step in the apoB secretion pathway in hepatocytes.

Background: Common genetic variation in close proximity to the ILRUN gene are significantly associated with coronary artery disease as well as with plasma lipid traits. We recently demonstrated that hepatic inflammation and lipid regulator with ubiquitin-associated domain-like and NBR1-like domains (ILRUN) regulates lipoprotein metabolism in vivo in mice. However, whether ILRUN, which is expressed in vascular cells, directly impacts atherogenesis remains unclear. We sought to determine the role of ILRUN in atherosclerosis development in mice. Methods: For our study, we generated global Ilrun -deficient ( Ilrun KO) male and female mice on 2 hyperlipidemic backgrounds: low density lipoprotein receptor knockout ( Ldlr KO) and apolipoprotein E knockout ( Apoe KO; double knockout [DKO]). Results: Compared with littermate control mice (single Ldlr KO or Apoe KO), deletion of Ilrun in DKO mice resulted in significantly attenuated both early and advanced atherosclerotic lesion development, as well as reduced necrotic area. DKO mice also had significantly decreased plasma cholesterol levels, primarily attributable to non-HDL (high-density lipoprotein) cholesterol. Hepatic-specific reconstitution of ILRUN in DKO mice on the Apoe KO background normalized plasma lipids, but atherosclerotic lesion area and necrotic area remained reduced in DKO mice. Further analysis showed that loss of Ilrun increased efferocytosis receptor MerTK expression in macrophages, enhanced in vitro efferocytosis, and significantly improved in situ efferocytosis in advanced lesions. Conclusions: Our results support ILRUN as an important novel regulator of atherogenesis that promotes lesion progression and necrosis. It influences atherosclerosis through both plasma lipid-dependent and lipid-independent mechanisms. These findings support ILRUN as the likely causal gene responsible for genetic association of variants with coronary artery disease at this locus and suggest that suppression of ILRUN activity might be expected to reduce atherosclerosis.

Metabolic dysfunction-associated Fatty Liver Disease (MAFLD) has emerged as one of the leading cardiometabolic diseases. Friend of GATA2 (FOG2) is a transcriptional co-regulator that has been shown to regulate hepatic lipid metabolism and accumulation. Using meta-analysis from several different biobank datasets, we identified a coding variant of FOG2 (rs28374544, A1969G, S657G) predominantly found in individuals of African ancestry (minor allele frequency~20%), which is associated with liver failure/cirrhosis phenotype and liver injury. To gain insight into potential pathways associated with this variant, we interrogated a previously published genomics dataset of 38 human induced pluripotent stem cell (iPSCs) lines differentiated into hepatocytes (iHeps). Using Differential Gene Expression Analysis and Gene Set Enrichment Analysis, we identified the mTORC1 pathway as differentially regulated between iHeps from individuals with and without the variant. Transient lipid-based transfections were performed on the human hepatoma cell line (Huh7) using wild-type FOG2 and FOG2^S657G and demonstrated that FOG2S^657G increased mTORC1 signaling, de novo lipogenesis, and cellular triglyceride synthesis and mass. In addition, we observed a significant downregulation of oxidative phosphorylation in FOG2^S657G cells in fatty acid-loaded cells but not untreated cells, suggesting that FOG2^S657G may also reduce fatty acid to promote lipid accumulation. Taken together, our multi-pronged approach suggests a model whereby the FOG2^S657G may promote MAFLD through mTORC1 activation, increased de novo lipogenesis, and lipid accumulation. Our results provide insights into the molecular mechanisms by which FOG2^S657G may affect the complex molecular landscape underlying MAFLD.

BACKGROUND: The coronary microvasculature is crucial for proper cardiac function, and coronary microvascular disease (CMVD) has emerged as an underdiagnosed and undertreated cause of ischemic heart disease. FOG2 (friend of GATA 2) is a transcriptional co-regulator crucial for coronary development and the maintenance of the coronary microvasculature in adult mice. Little is known about the role of FOG2 in humans or its role in CMVD. Here, we report a genotype-first approach to determine the role of FOG2 in human CMVD. METHODS: We performed phenome-wide association studies and deep cardiac phenotyping through the Electronic Health Record in individuals with FOG2 coding variants. We interrogated MagNET heart tissue data to identify genes and pathways associated with rs28374544. We then overexpressed FOG2S657G in a cardiomyocyte cell line and assessed the effects on cardiac metabolism and paracrine angiogenic signaling. RESULTS: We identified an association between rs28374544 (A1969G, p.S657G) and CMVD. Using phenome-wide association studies and deep cardiac phenotyping through the Electronic Health Record in individuals with FOG2 coding variants, we identified an association between rs28374544 (A1969G, p.S657G) and CMVD. Individuals carrying the S657G variant, almost all of African ancestry, had increased chest pain, a smaller burden of obstructive coronary artery disease, and altered coronary blood flow. Differential gene and pathway analysis using several genomic data sets showed that carriers of S657G have increased expression of genes involved in angiogenesis, glycolysis, and the HIF (hypoxia-inducible factor) pathway. In vitro functional studies show that compared with the FOG2 wild-type protein, the FOG2 ^S657G variant protein promotes angiogenic gene expression and angiogenesis while decreasing oxygen consumption rate. CONCLUSIONS: A common functional coding variant in FOG2, S657G, is associated with CMVD in humans. Altered angiogenic gene expression, regulated in part by FOG2, may contribute to CMVD.