Non-alcoholic fatty liver disease (NAFLD) is a common condition that can progress to the more severe conditions like non-alcoholic steatohepatitis (NASH) for which limited effective therapeutic... Show moreNon-alcoholic fatty liver disease (NAFLD) is a common condition that can progress to the more severe conditions like non-alcoholic steatohepatitis (NASH) for which limited effective therapeutic options are available. In this study, we set out to evaluate the novel glucocorticoid receptor modulator CORT125385, an analogue of the previously studied miricorilant but without mineralocorticoid receptor binding activity. Male and female mice that received high-fat diet and fructose water were treated with either vehicle, CORT125385 or mifepristone. We found that CORT125385 significantly lowered hepatic triglyceride levels in male mice, and hepatic triglyceride and cholesterol levels in female mice. Mifepristone treatment had no effect in male mice, but significantly lowered hepatic triglyceride and cholesterol levels in female mice. In reporter assays in vitro, CORT125385 showed weak partial agonism on the progesterone receptor (PR) at high doses, as well as PR antagonism at a potency 1000-fold lower than mifepristone. In vivo, CORT125385 treatment did not influence PR-responsive gene expression in the oviduct, while mifepristone treatment strongly influenced these genes in the oviduct, thus excluding in vivo PR cross-reactivity of CORT125385 at a therapeutically active dose. We conclude that CORT125385 is a promising glucocorticoid receptor modulator that effectively reduces liver steatosis in male and female mice without affecting other steroid receptors at doses that lower hepatic lipid content. Show less
Bone morphogenetic protein (BMP)-9, a member of the TGF beta-family of cytokines, is believed to be mainly produced in the liver. The serum levels of BMP-9 were reported to be reduced in newly... Show moreBone morphogenetic protein (BMP)-9, a member of the TGF beta-family of cytokines, is believed to be mainly produced in the liver. The serum levels of BMP-9 were reported to be reduced in newly diagnosed diabetic patients and BMP-9 overexpression ameliorated steatosis in the high fat diet-induced obesity mouse model. Furthermore, injection of BMP-9 in mice enhanced expression of fibroblast growth factor (FGF)21. However, whether BMP-9 also regulates the expression of the related FGF19 is not clear. Because both FGF21 and 19 were described to protect the liver from steatosis, we have further investigated the role of BMP-9 in this context.We first analyzed BMP-9 levels in the serum of streptozotocin (STZ)-induced diabetic rats (a model of type I diabetes) and confirmed that BMP-9 serum levels decrease during diabetes. Microarray analyses of RNA samples from hepatic and intestinal tissue from BMP-9 KO-and wild-type mice (C57/Bl6 background) pointed to basal expression of BMP-9 in both organs and revealed a down-regulation of hepatic Fgf21 and intestinal Fgf19 in the KO mice. Next, we analyzed BMP-9 levels in a cohort of obese patients with or without diabetes. Serum BMP-9 levels did not correlate with diabetes, but hepatic BMP-9 mRNA expression negatively correlated with steatosis in those patients that did not yet develop diabetes. Likewise, hepatic BMP-9 expression also negatively correlated with serum LPS levels. In situ hybridization analyses confirmed intestinal BMP-9 expression. Intestinal (but not hepatic) BMP-9 mRNA levels were decreased with diabetes and positively correlated with intestinal E-Cadherin expression. In vitro studies using organoids demonstrated that BMP-9 directly induces FGF19 in gut but not hepatocyte organoids, whereas no evidence of a direct induction of hepatic FGF21 by BMP-9 was found. Consistent with the in vitro data, a correlation between intestinal BMP-9 and FGF19 mRNA expression was seen in the patients' samples.In summary, our data confirm that BMP-9 is involved in diabetes development in humans and in the control of the FGF-axis. More importantly, our data imply that not only hepatic but also intestinal BMP-9 associates with diabetes and steatosis development and controls FGF19 expression. The data support the conclusion that increased levels of BMP-9 would most likely be beneficial under pre-steatotic conditions, making supplemen-tation of BMP-9 an interesting new approach for future therapies aiming at prevention of the development of a metabolic syndrome and liver steatosis. Show less
Lingen, E. van; Tushuizen, M.E.; Steenhuis, M.E.J.; Deynen, T. van; Martens, J.; Morales, D.D.I.; ... ; Maljaars, P.W.J. 2021
Purpose Liver steatosis is a frequently reported condition in patients with inflammatory bowel disease (IBD). Different factors, both metabolic and IBD-associated, are believed to contribute to the... Show morePurpose Liver steatosis is a frequently reported condition in patients with inflammatory bowel disease (IBD). Different factors, both metabolic and IBD-associated, are believed to contribute to the pathogenesis. The aim of our study was to calculate the prevalence of liver steatosis and fibrosis in IBD patients and to evaluate which factors influence changes in steatosis and fibrosis during follow-up. Methods From June 2017 to February 2018, demographic and biochemical data was collected at baseline and after 6-12 months. Measured by transient elastography (FibroScan), liver steatosis was defined as Controlled Attenuation Parameter (CAP) >= 248 and fibrosis as liver stiffness value (Emed) >= 7.3 kPa. IBD disease activity was defined as C-reactive protein (CRP) >= 10 mg/l and/or fecal calprotectin (FCP) >= 150 mu g/g. Univariate and multivariate regression analysis was performed; a p-value of <= 0.05 was considered significant. Results Eighty-two out of 112 patients were seen for follow-up; 56% were male. The mean age was 43 +/- 16.0 years, and mean BMI was 25.1 +/- 4.7 kg/m(2). The prevalence of liver steatosis was 40% and of fibrosis was 20%. At baseline, 26 patients (32%) had an active episode of IBD. Using a multivariate analysis, disease activity at baseline was associated with an increase in liver steatosis (B = 37, 95% CI 4.31-69.35, p = 0.027) and liver fibrosis (B = 1.2, 95% CI 0.27-2.14, p = 0.016) during follow-up. Conclusions This study confirms the relatively high prevalence of liver steatosis and fibrosis in IBD patients. We demonstrate that active IBD at baseline is associated with both an increase in liver steatosis and fibrosis during follow-up. Show less
This read-across case study characterises thirteen, structurally similar carboxylic acids demonstrating the application of in vitro and in silico human-based new approach methods, to determine... Show moreThis read-across case study characterises thirteen, structurally similar carboxylic acids demonstrating the application of in vitro and in silico human-based new approach methods, to determine biological similarity. Based on data from in vivo animal studies, the read-across hypothesis is that all analogues are steatotic and so should be considered hazardous. Transcriptomic analysis to determine differentially expressed genes (DEGs) in hepatocytes served as first tier testing to confirm a common mode-of-action and identify differences in the potency of the analogues. An adverse outcome pathway (AOP) network for hepatic steatosis, informed the design of an in vitro testing battery, targeting AOP relevant MIEs and KEs, and Dempster-Shafer decision theory was used to systematically quantify uncertainty and to define the minimal testing scope. The case study shows that the read-across hypothesis is the critical core to designing a robust, NAM-based testing strategy. By summarising the current mechanistic understanding, an AOP enables the selection of NAMs covering MIEs, early KEs, and late KEs. Experimental coverage of the AOP in this way is vital since MIEs and early KEs alone are not confirmatory of progression to the AO. This strategy exemplifies the workflow previously published by the EUTOXRISK project driving a paradigm shift towards NAM-based NGRA. Show less
Nahon, J.E.; Hoekstra, M.; Harmelen, V. van; Rensen, P.C.N.; Dijk, K.W. van; Kooijman, S.; Eck, M. van 2019
ObjectiveProteoglycan 4 (Prg4) has emerged from human association studies as a possible factor contributing to weight gain, dyslipidemia and insulin resistance. In the current study, we... Show moreObjectiveProteoglycan 4 (Prg4) has emerged from human association studies as a possible factor contributing to weight gain, dyslipidemia and insulin resistance. In the current study, we investigated the causal role of Prg4 in controlling lipid and glucose metabolism in mice.MethodsPrg4 knockout (KO) mice and wild-type (WT) littermates were challenged with an obesogenic high-fat diet (45% of total calories as fat) for 16 weeks. To further stimulate the development of metabolic alterations, 10% fructose water was provided starting from week 13.ResultsPrg4 deficiency only tended to reduce diet-induced body weight gain, but significantly improved glucose handling (AUC: −29%; p < 0.05), which was also reflected by a tendency towards a reduced HOMA-IR score (−49%; p = 0.06 as compared to WT mice). This coincided with lower hepatic expression of glycolysis (Gck: −30%; p < 0.05) and lipogenesis (Acc: −21%; p < 0.05 and Scd1: −38%; p < 0.001) genes, which translated in significantly lower hepatic triglyceride levels (−56%; p < 0.001) in Prg4 KO mice as compared to WT mice. Prg4 KO mice likely had lower glucose utilization by skeletal muscle as compared to WT mice, judged by a significant reduction in the genes Glut4 (−29%; p < 0.01), Pfkm (−21%; p < 0.05) and Hk2 (−39%; p < 0.001). Moreover, Prg4 KO mice showed a favorable white adipose tissue phenotype with lower uptake of triglyceride-derived fatty acids (−46%; p < 0.05) and lower gene expression of inflammatory markers Cd68, Mcp1 and Tnfα (−65%, −81% and −63%, respectively; p < 0.01) than WT mice.ConclusionPrg4 KO mice are protected from high-fat diet-induced glucose intolerance and fatty liver disease. Show less
