Alcohol promotes liver fibrosis in high fat diet induced diabetic rats.

OBJECTIVES: Type 2 diabetes (T2DM) and alcoholism are considered to be lifestyle-associated independent risk factors in fatty liver diseases (FLD) mediated cirrhosis and hepatocellular carcinoma (HCC). A combined effect of both these conditions may exacerbate the pathological changes and a pre-clinical exploration of this is expected to provide a mechanical detail of the pathophysiology. The present study aims to understand the effect of alcohol on pre- diabetic and type 2 diabetic female Wistar rats. METHODS: In this experimental study, 12 Wistar rats (180-220 g) were randomly assigned into three groups: Normal (fed normal rat chow), alcohol (20 %) fed diabetic (HFD + STZ), and pre-diabetic rats (HFD alone). After, two months of the experimental period, blood and liver tissues were collected lipid metabolic alteration, liver injury, and fibrosis were determined following biochemical and histological methods. Data were analyzed using one-way ANOVA and Dunnett's Post Hoc test. RESULTS: Significant dyslipidemia was observed in the liver tissues of diabetic and pre-diabetic rats following alcohol ingestion. A significant (p<0.05) increase in lipid peroxidation status, and hepatic marker enzyme activities (p<0.0001) were observed in diabetic animals. In corroborating with these observations, hematoxylin and eosin staining of hepatic tissue revealed the presence of sinusoidal dilation along with heavily damaged hepatocytes and inflammatory cell infiltration. Further, significantly (p<0.001) increased hepatic hydroxyproline content and extended picrosirius red stained areas of collagen in liver tissue indicated initiation of fibrosis in alcohol-fed diabetic rats. CONCLUSIONS: Overall, the results indicate that alcohol consumption in T2DM conditions is more deleterious than pre diabetic conditions in progressing to hepatic fibrosis.

Thermally Oxidized Coconut Oil as Fat Source in High-Fat Diet Induces Hepatic Fibrosis in Diabetic Rat Model.

In the present study, HFD/STZ-mediated type 2 diabetic rodent model was used to comparatively evaluate coconut oil (CO) and thermally oxidized CO (TCO) as fat sources for the development of NAFLD. Female Wistar rats (six in each group; average bwt 200 g) fed HFD containing either CO or TCO for 2 months along with an intraperitoneal injection of streptozotocin (30 mg/kg bwt) at the end of 1-month feeding were found to develop fatty liver and subsequent inflammatory changes when compared to the normal laboratory diet-fed animals over 2-month period. Dyslipidemia as well as enhanced activities of serum hepatic marker enzymes (e.g., AST, ALT, and ALP) were prominent in TCO-fed animals. Further, HFD-fed animals showed alterations in their hepatic redox equilibrium. Hepatic GSH and antioxidant enzyme activities that form the part of a protective mechanism against oxidative/carbonyl stress were found to be increased in HFD-fed rats. Supporting this, CO- and TCO-containing-HFD-fed animals had enhanced lipid peroxidation (increased TBARs). Thus, fatty liver with heightened antioxidant defense, lipid peroxidation, and inflammation indicate hepatosteatosis. Histological details of the hepatic tissues corroborated sufficiently with these observations and showed an increased incidence of macrovesicles, inflammation, and hepatocyte ballooning in the TCO-fed rats than in CO-fed animals. Further, in support of this proposition, hydroxyproline, an index of collagen formation, was found to be significantly increased in TCO-fed rats than in the CO-fed group. Overall, the study shows that the formulation of HFD incorporated with TCO as a fat source, combined with STZ injection, is an efficient dietary model for developing hepatosteatosis with fibrotic stage in rats within 2 months. Administration of this modified diet for a more extended period may be a good model for cirrhotic and hepatocellular carcinoma studies, which need to be further assessed.

Quantitative proteome profiling stratifies fibroepithelial lesions of the breast.

Breast fibroepithelial lesions (FELs) include heterogeneous pathological tumors, involving indolent fibroadenoma (FAD) to potentially aggressive phyllodes tumors (PTs). The current grading system remains unreliable in differentiating these tumors due to histological heterogeneity and lack of appropriate markers to monitor the sudden and unpredictable malignant transformation of PTs. Thus, there exists an imminent need for a marker-based diagnostic approach to augment the conventional histological platform that could lead to accurate diagnosis and distinction of FELs. The high- throughput quantitative proteomic analysis suggested that FAD and PTs form distinct clusters away from borderline and malignant though there exist marked differences between them. Interestingly, over-expression of extracellular matrices (ECM) related proteins and epithelial-mesenchymal transition (EMT) markers in borderline PTs led us to hypothesize a model of deposition and degradation leading to ECM remodeling and EMT acquisition triggering its malignant transformation. We also identified three candidate biomarkers such as MUCL1, HTRA1, and VEGDF uniquely expressed in FAD, borderline, and malignant PTs, respectively, which were further validated using immunohistochemistry. The present work shed light on a brief mechanistic framework of PTs aggressive nature and present potential biomarkers to differentiate overlapping FELs that would be of practical utility in augmenting existing diagnosis and disease management for this rare tumor.

Inhibition of Twist1-mediated invasion by Chk2 promotes premature senescence in p53-defective cancer cells.

Twist1, a basic helix-loop-helix transcription factor is implicated as a key mediator of epithelial-mesenchymal transition (EMT) and metastatic dissemination in p53-deficient cancer cells. On the other hand, checkpoint kinase 2 (Chk2), a major cell cycle regulatory protein provides a barrier to tumorigenesis due to DNA damage response by preserving genomic stability of the cells. Here we demonstrate that Chk2 induction proficiently abrogates invasion, cell scattering and invadopodia formation ability of p53-mutated invasive cells by suppressing Twist1, indicating Chk2 confers vital role in metastasis prevention. In addition, ectopic Chk2, as well as its (Chk2) induction by natural podophyllotoxin analog, 4'-demethyl-deoxypodophyllotoxin glucoside (4DPG), strongly restrain Twist1 activity along with other mesenchymal markers, for example, ZEB-1, vimentin and Snail1, whereas the epithelial markers such as E-cadherin and TIMP-1 expression augmented robustly. However, downregulation of endogenous Chk2 by siRNA as well as Chk2 selective inhibitor PV1019 implies that 4DPG-mediated inhibition of Twist1 is Chk2-dependent. Further, mechanistic studies unveil that Chk2 negatively regulates Twist1 promoter activity and it (Chk2) interacts steadily with Snail1 protein to curb EMT. Strikingly, Chk2 overexpression triggers premature senescence in these cells with distinctive increase in senescence-associated ß-galactosidase (SA-ß-gal) activity, G2/M cell cycle arrest and induction of senescence-specific marker p21waf1/Cip1. Importantly, stable knockdown of Twist1 by shRNA markedly augments p21 expression, its nuclear accumulation, senescence-associated heterochromatin foci (SAHF) and amplifies the number of SA-ß-gal-positive cells. Moreover, our in vivo studies also validate that 4DPG treatment significantly abrogates tumor growth as well as metastatic lung nodules formation by elevating the level of phospho-Chk2, Chk2 and suppressing Twist1 activity in mouse mammary carcinoma model. In a nutshell, this report conceives a novel strategy of Twist1 suppression through Chk2 induction, which prevents metastatic dissemination and promotes premature senescence in p53-defective invasive cancer cells.