MCLR-elicited hepatic fibrosis and carcinogenic gene expression changes persist in rats with diet-induced nonalcoholic steatohepatitis through a 4-week recovery period.

Nonalcoholic steatohepatitis (NASH) causes liver extracellular matrix (ECM) remodeling and is a risk factor for fibrosis and hepatocellular carcinoma (HCC). Microcystin-LR (MCLR) is a hepatotoxin produced by fresh-water cyanobacteria that causes a NASH-like phenotype, liver fibrosis, and is also a risk factor for HCC. The focus of the current study was to investigate and compare hepatic recovery after cessation of MCLR exposure in healthy versus NASH animals. Male Sprague-Dawley rats were fed either a control or a high fat/high cholesterol (HFHC) diet for eight weeks. Animals received either vehicle or 30 µg/kg MCLR (i.p: 2 weeks, alternate days). Animals were euthanized at one of three time points: at the completion of the MCLR exposure period and after 2 and 4 weeks of recovery. Histological staining suggested that after four weeks of recovery the MCLR-exposed HFHC group had less steatosis and more fibrosis compared to the vehicle-exposed HFHC group and MCLR-exposed control group. RNA-Seq analysis revealed dysregulation of ECM genes after MCLR exposure in both control and HFHC groups that persisted only in the HFHC groups during recovery. After 4 weeks of recovery, MCLR hepatotoxicity in pre-existing NASH persistently dysregulated genes related to cellular differentiation and HCC. These data demonstrate impaired hepatic recovery and persistent carcinogenic changes after MCLR toxicity in pre-existing NASH.

Insulin-induced lipid body accumulation is accompanied by lipid remodelling in model mast cells.

Mast cell lipid bodies are key to initiation, maintenance and resolution of inflammatory responses in tissue. Mast cell lines, primary bone marrow-derived mast cells and peripheral blood basophils present a 'steatotic' phenotype in response to chronic insulin exposure, where cells become loaded with lipid bodies. Here we show this state is associated with reduced histamine release, but increased capacity to release bioactive lipids. We describe the overall lipid phenotype of mast cells in this insulin-induced steatotic state and the consequences for critical cellular lipid classes involved in stages of inflammation. We show significant insulin-induced shifts in specific lipid classes, especially arachidonic acid derivatives, MUFA and PUFA, the EPA/DHA ratio, and in cardiolipins, especially those conjugated to certain DHA and EPAs. Functionally, insulin exposure markedly alters the FcεRI-induced release of Series 4 leukotriene LTC4, Series 2 prostaglandin PGD2, Resolvin-D1, Resolvin-D2 and Resolvin-1, reflecting the expanded precursor pools and impact on both the pro-inflammation and pro-resolution bioactive lipids that are released during mast cell activation. Chronic hyperinsulinemia is a feature of obesity and progression to Type 2 Diabetes, these data suggest that mast cell release of key lipid mediators is altered in patients with metabolic syndrome.

Comparative analysis of lipotoxicity induced by endocrine, pharmacological, and innate immune stimuli in rat basophilic leukemia cells.

Cellular lipotoxicity manifests as the steatotic accumulation of lipid droplets or lipid bodies, and/or induction of phospholipidosis. Lipotoxicity can be induced by hyperinsulinemia/nutrient overload, cationic amphiphilic drugs (CAD), and innate immunological stimuli, all of which are stimuli relevant to mast cell physiology. Hyper-accumulation of mast cell lipid bodies in response to hyperinsulinemia has been documented, but lipotoxicity in response to CAD or innate immunologic stimuli has not been analysed comparatively. Moreover, gaps in our understanding of this steatosis remain, specifically as to whether hyperinsulinemia-driven steatosis in these cells attains lipotoxic levels or is accompanied by phospholipidosis. To compare endocrine, pharmacological, and innate immunological stimuli for their ability to induce steatosis and phospholipidosis in a rat basophilic leukemia mast cell model (RBL2H3), differential fluorescence microscopy staining and quantitation of phospholipidosis and steatosis in the RBL2H3 cell line was examined. The three classes of stimuli differentially induced phospholipidosis and steatosis. PPARγ up-regulation was not uniformly associated with the expansion of the lipid body population. Fluorescence imaging of lipid-enriched structures generated in response to lipotoxic cationic amphiphilic drugs, chronic insulin exposure, and TLR2/4 ligands revealed differential staining patterns when visualized using lipophilic dyes. It is concluded that lipotoxicity-inducing pathways in this model mast cell system are diverse, and include steatotic responses to an endocrine stimulus, as well as phospholipidosis responses to cationic lipophilic drugs not previously described in this cell type.