Cholesterol Accumulation as a Driver of Hepatic Inflammation Under Translational Dietary Conditions Can Be Attenuated by a Multicomponent Medicine

Abstract

BackgroundNon-alcoholic fatty liver disease (NAFLD) is a complex multifactorial disorder that is characterised by dysfunctional lipid metabolism and cholesterol homeostasis, and a related chronic inflammatory response. NAFLD has become the most common cause of chronic liver disease in many countries, and its prevalence continues to rise in parallel with increasing rates of obesity. Here, we evaluated the putative NAFLD-attenuating effects of a multicomponent medicine consisting of 24 natural ingredients: Hepar compositum (HC-24).MethodsLdlr-/-.Leiden mice were fed a high-fat diet (HFD) with a macronutrient composition and cholesterol content comparable to human diets for 24 weeks to induce obesity-associated metabolic dysfunction, including hepatic steatosis and inflammation. HC-24 or vehicle control was administered intraperitoneally 3 times/week (1.5 ml/kg) for the last 18 weeks of the study. Histological analyses of liver and adipose tissue were combined with extensive hepatic transcriptomics analysis. Transcriptomics results were further substantiated with ELISA, immunohistochemical and liver lipid analyses.ResultsHFD feeding induced obesity and metabolic dysfunction including adipose tissue inflammation and increased gut permeability. In the liver, HFD-feeding resulted in a disturbance of cholesterol homeostasis and an associated inflammatory response. HC-24 did not affect body weight, metabolic risk factors, adipose tissue inflammation or gut permeability. While HC-24 did not alter total liver steatosis, there was a pronounced reduction in lobular inflammation in HC-24-treated animals, which was associated with modulation of genes and proteins involved in inflammation (e.g., neutrophil chemokine Cxcl1) and cholesterol homeostasis (i.e., predicted effect on ‘cholesterol’ as an upstream regulator, based on gene expression changes associated with cholesterol handling). These effects were confirmed by CXCL1 ELISA, immunohistochemical staining of neutrophils and biochemical analysis of hepatic free cholesterol content. Intrahepatic free cholesterol levels were found to correlate significantly with the number of inflammatory aggregates in the liver, thereby providing a potential rationale for the observed anti-inflammatory effects of HC-24.ConclusionsFree cholesterol accumulates in the liver of Ldlr-/-.Leiden mice under physiologically translational dietary conditions, and this is associated with the development of hepatic inflammation. The multicomponent medicine HC-24 reduces accumulation of free cholesterol and has molecular and cellular anti-inflammatory effects in the liver.

Krill Oil Treatment Increases Distinct PUFAs and Oxylipins in Adipose Tissue and Liver and Attenuates Obesity-Associated Inflammation via Direct and Indirect Mechanisms

“…Furthermore, both tissues were subject to a comprehensive fatty acid composition analysis, including oxylipins using LC-MS/MS. Ldlr−/− Leiden mice were chosen as a model because these mice develop obesity in combination with pronounced hyperinsulinemia and adipose tissue and liver inflammation when treated with energy-dense diets for 28 weeks (to allow for the histological analysis of organ inflammation) with a human-diet-like macronutrient composition, not requiring dietary supplementation with cholesterol [ 20 , 21 , 22 , 23 ]. Under the experimental conditions employed herein, Ldlr−/− Leiden mice have been shown to develop histopathological features similar to humans and they recapitulate human disease pathways, as demonstrated by recent comparative transcriptomics, metabolomics and proteomics studies [ 21 , 22 , 24 ].…”

Chronic Oral Administration of Mineral Oil Compared With Corn Oil: Effects on Gut Permeability and Plasma Inflammatory and Lipid Biomarkers

“…Future work could involve use of a more advanced disease model, for example mice on a high fat diet, containing a higher amount of saturated fat (45 kcal%) than used in the current study (36 kcal%), to mimic obesity and insulin resistance in the Western world. Such a high fat diet is known to aggravate intestinal permeability in mice (Mueller et al, 2021), which may be further worsened by mineral oil. Future work could also assess where in the body mineral oil accumulates and whether (and how) long-term mineral oil administration affects tissue composition, local cytokine levels in the intestine and liver, and the composition of the gut microbiota.…”

Butyrate Protects against Diet-Induced NASH and Liver Fibrosis and Suppresses Specific Non-Canonical TGF-β Signaling Pathways in Human Hepatic Stellate Cells

“…Adipose tissue inflammation was assessed as described previously [ 17 ]. In short, the number of crown-like structures (CLS) were scored in hematoxylin-phloxine-saffron stained epidydimal WAT (eWAT) and mesenteric WAT (mWAT) cross sections in three non-overlapping fields (100 × magnification) per mouse.…”

“…These observations have been confirmed in rodents, where dietary supplementation with BU was shown to reduce food intake and attenuate body weight gain [ 16 ]. BU has also been shown to improve liver steatosis and inflammation [ 12 , 17 , 18 ]; however, it is unclear if these effects are secondary to its anti-obesogenic effects, or result from a direct effect of BU on these processes. Furthermore, while BU has been shown to improve many of the metabolic dysregulations that underlie development of inflammation and fibrosis in the liver, it is unknown whether BU can also reduce development of fibrosis in the context of obesity-associated NASH.…”

“…To study the potential anti-fibrotic effects of butyrate we used a well-established translational mouse model for diet-induced NASH and liver fibrosis: high-fat diet (HFD)-fed Ldlr-/-.Leiden mice. Under translational dietary conditions (i.e., using a HFD with a macronutrient composition and cholesterol content comparable to human diets) [ 17 ], these mice develop NASH with fibrosis in the context of obesity, insulin resistance, adipose tissue inflammation, and gut dysfunction, as is typical for many NASH patients [ 18 ]. In addition, these mice express underlying disease pathways relevant for human NASH, as demonstrated by recent transcriptomics, metabolomics, and proteomics studies [ 18 , 19 ].…”