Treatment of intestinal and liver features in cystic fibrosis mice by the osmotic laxative polyethylene glycol.

BACKGROUND: Cystic Fibrosis (CF) is a genetic disease affecting multiple organs, primarily the lungs and digestive system. Improved pulmonary management significantly improved life expectancy of CF patients. As a result, extrapulmonary manifestations, including gastrointestinal and liver-related symptoms, have become more relevant. We previously reported that the osmotic laxative polyethylene glycol (PEG), which hydrates the CF gut, decreased fecal bile acid loss in a CF knockout mouse model. In the current study we investigated the effect of PEG on intestinal fat and cholesterol absorption and on CF-related liver features in a CF mouse model with the most common CF-causing mutation. METHODS: CftrΔF508/ΔF508 (n=13) and wild-type (WT) (n=12) mice were treated with PEG for 2 weeks. The intestinal and hepatic effects of PEG were assessed by analysis of intestinal bile acid, cholesterol, and fat fluxes, transcriptome analysis as well as histology. RESULTS: PEG improved intestinal malabsorption of bile acids, fat, and cholesterol in CftrΔF508/ΔF508 mice. Transcriptome analysis showed that PEG partially restored the intestinal signaling of nuclear receptors RXR, FXR, and CAR/PXR, which are involved in bile acid and xenobiotic metabolism. PEG also reduced liver inflammation in CF mice as assessed by transcriptome and histological analyses. CONCLUSIONS: PEG, a non-absorbable osmotic laxative, improved intestinal nutrient absorption, intestinal bile acid and xenobiotic signaling, as well as CF-related liver features. These findings highlight the potential for osmotic laxation to improve gastrointestinal complications of CF in humans.

The chemotherapeutic drug doxorubicin does not exacerbate p16Ink4a-positive senescent cell accumulation and cardiometabolic disease development in young adult female LDLR-deficient mice.

Cancer survivors who received chemotherapy, such as the anthracycline doxorubicin (DOX), have an increased risk of developing complications later in life, including the development of chronic metabolic diseases. Although the etiology of this increased risk for late metabolic complications in cancer survivors is poorly understood, a causal role of therapy-induced senescent cells has been suggested. To study the role of cellular senescence in chemotherapy-induced metabolic complications, young adult female low-density lipoprotein receptor-deficient (Ldlr-/-)-p16-3MR mice, in which p16Ink4a-positive (p16Ink4a+) senescent cells can be genetically eliminated, were treated with four weekly injections of DOX (2.5 mg/kg) followed by a high-fat high-cholesterol diet for 12 weeks. While DOX treatment induced known short-term effects, such as reduction in body weight, gonadal fat mass, and adipose tissue inflammation, it was not associated with significant long-term effects on glucose homeostasis, hepatic steatosis, or atherosclerosis. We further found no evidence of DOX-induced accumulation of p16Ink4a+-senescent cells at 1 or 12 weeks after DOX treatment. Neither did we observe an effect of elimination of p16Ink4a+-senescent cells on the development of diet-induced cardiometabolic complications in DOX-treated mice. Other markers for senescence were generally also not affected except for an increase in p21 and Cxcl10 in gonadal white adipose tissue long-term after DOX treatment. Together, our study does not support a significant role for p16Ink4a+-senescent cells in the development of diet-induced cardiometabolic disease in young adult DOX-treated female Ldlr-/- mice. These findings illustrate the need of further studies to understand the link between cancer therapy and cardiometabolic disease development in cancer survivors.

Pharmacological inhibition of MEK1/2 signaling disrupts bile acid metabolism through loss of Shp and enhanced Cyp7a1 expression.

The RAS-MAPK signaling pathway is one of the most frequently dysregulated pathways in human cancer. Small molecule inhibitors directed against this pathway have clinical activity in patients with various cancer types and can improve patient outcomes. However, the use of these drugs is associated with adverse effects, which can result in dose reduction or treatment interruption. A better molecular understanding of on-target, off-tumor effects may improve toxicity management. In the present study, we aimed to identify early initiating biological changes in the liver upon pharmacological inhibition of the RAS-MAPK signaling pathway. To this end, we tested the effect of MEK inhibitor PD0325901 using mice and human hepatocyte cell lines. Male C57BL/6 mice were treated with either vehicle or PD0325901 for six days, followed by transcriptome analysis of the liver and phenotypic characterization. Pharmacological MEK inhibition altered the expression of 423 genes, of which 78 were upregulated and 345 were downregulated. We identified Shp, a transcriptional repressor, and Cyp7a1, the rate-limiting enzyme in converting cholesterol to bile acids, as the top differentially expressed genes. PD0325901 treatment also affected other genes involved in bile acid regulation, which was associated with changes in the composition of plasma bile acids and composition and total levels of fecal bile acids and elevated predictive biomarkers of early liver toxicity. In conclusion, short-term pharmacological MEK inhibition results in profound changes in bile acid metabolism, which may explain some of the clinical adverse effects of pharmacological inhibition of the RAS-MAPK pathway, including gastrointestinal complications and hepatotoxicity.

Defective Lipid Droplet-Lysosome Interaction Causes Fatty Liver Disease as Evidenced by Human Mutations in TMEM199 and CCDC115.

BACKGROUND & AIMS: Recently, novel inborn errors of metabolism were identified because of mutations in V-ATPase assembly factors TMEM199 and CCDC115. Patients are characterized by generalized protein glycosylation defects, hypercholesterolemia, and fatty liver disease. Here, we set out to characterize the lipid and fatty liver phenotype in human plasma, cell models, and a mouse model. METHODS AND RESULTS: Patients with TMEM199 and CCDC115 mutations displayed hyperlipidemia, characterized by increased levels of lipoproteins in the very low density lipoprotein range. HepG2 hepatoma cells, in which the expression of TMEM199 and CCDC115 was silenced, and induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells from patients with TMEM199 mutations showed markedly increased secretion of apolipoprotein B (apoB) compared with controls. A mouse model for TMEM199 deficiency with a CRISPR/Cas9-mediated knock-in of the human A7E mutation had marked hepatic steatosis on chow diet. Plasma N-glycans were hypogalactosylated, consistent with the patient phenotype, but no clear plasma lipid abnormalities were observed in the mouse model. In the siTMEM199 and siCCDC115 HepG2 hepatocyte models, increased numbers and size of lipid droplets were observed, including abnormally large lipid droplets, which colocalized with lysosomes. Excessive de novo lipogenesis, failing oxidative capacity, and elevated lipid uptake were not observed. Further investigation of lysosomal function revealed impaired acidification combined with impaired autophagic capacity. CONCLUSIONS: Our data suggest that the hypercholesterolemia in TMEM199 and CCDC115 deficiency is due to increased secretion of apoB-containing particles. This may in turn be secondary to the hepatic steatosis observed in these patients as well as in the mouse model. Mechanistically, we observed impaired lysosomal function characterized by reduced acidification, autophagy, and increased lysosomal lipid accumulation. These findings could explain the hepatic steatosis seen in patients and highlight the importance of lipophagy in fatty liver disease. Because this pathway remains understudied and its regulation is largely untargeted, further exploration of this pathway may offer novel strategies for therapeutic interventions to reduce lipotoxicity in fatty liver disease.

ß-Catenin Directs Nuclear Factor-κB p65 Output via CREB-Binding Protein/p300 in Human Airway Smooth Muscle.

ß-Catenin is a multifunctional protein that apart from its role in proliferative and differentiation events, also acts upon inflammatory processes, mainly via interaction with nuclear factor-κB (NF-κB). However, there is still controversy as to whether ß-catenin facilitates or represses NF-κB output. Insights into the molecular mechanisms underlying the interaction between ß-catenin and NF-κB have highlighted the cofactors CREB-binding protein (CBP) and p300 as important candidates. Here, we hypothesized that the interaction of ß-catenin with CBP/p300 directs NF-κB output. Using human airway smooth muscle (ASM) cells, we found that ß-catenin is essential in interleukin -1ß (IL-1ß)-mediated expression of interleukin-6 (IL-6) by promoting nuclear translocation of the p65 subunit of NF-κB. These effects were independent from WNT pathway activation or other factors that promote ß-catenin signaling. In the nucleus, inhibition of either the CBP- or p300-ß-catenin interaction could regulate NF-κB output, by enhancing (CBP inhibition) or inhibiting (p300 inhibition) IL-1ß-induced expression of IL-6, respectively. Acetylation of p65 by p300 likely underlies these events, as inhibition of the p300-ß-catenin interaction diminished levels of acetylated p65 at lysine 310, thereby reducing p65 transcriptional activity. In conclusion, ß-catenin is a critical component of NF-κB-mediated inflammation in human ASM, affecting transcriptional output by interacting with the nuclear cofactors CBP and p300. Targeting ß-catenin may be an alternative strategy to treat airway inflammation in patients with airway disease, such as asthma.