Following liver regeneration after partial hepatectomy, liver grows back precisely to its original mass and does not exceed it. The mechanism regulating this "hepatostat" is not clear and no exceptions have been found to date. Although pathways initiating liver regeneration have been well studied, mechanisms involved in the termination of liver regeneration are unclear. Here, we report that integrin-linked kinase (

Human breast tumors harbor supernumerary centrosomes in almost 80% of tumor cells. Although amplified centrosomes compromise cell viability via multipolar spindles resulting in death-inducing aneuploidy, cancer cells tend to cluster extra centrosomes during mitosis. As a result cancer cells display bipolar spindle phenotypes to maintain a tolerable level of aneuploidy, an edge to their survival. HSET/KifC1, a kinesin-like minus-end directed microtubule motor has recently found fame as a crucial centrosome clustering molecule. Here we show that HSET promotes tumor progression via mechanisms independent of centrosome clustering. We found that HSET is overexpressed in breast carcinomas wherein nuclear HSET accumulation correlated with histological grade and predicted poor progression-free and overall survival. In addition, deregulated HSET protein expression was associated with gene amplification and/or translocation. Our data provide compelling evidence that HSET overexpression is pro-proliferative, promotes clonogenic-survival and enhances cell-cycle kinetics through G2 and M-phases. Importantly, HSET co-immunoprecipitates with survivin, and its overexpression protects survivin from proteasome-mediated degradation, resulting in its increased steady-state levels. We provide the first evidence of centrosome clustering-independent activities of HSET that fuel tumor progression and firmly establish that HSET can serve both as a potential prognostic biomarker and as a valuable cancer-selective therapeutic target.

Centrosome amplification (CA), a cell-biological trait, characterizes pre-neoplastic and pre-invasive lesions and is associated with tumor aggressiveness. Recent studies suggest that CA leads to malignant transformation and promotes invasion in mammary epithelial cells. Triple negative breast cancer (TNBC), a histologically-aggressive subtype shows high recurrence, metastases, and mortality rates. Since TNBC and non-TNBC follow variable kinetics of metastatic progression, they constitute a novel test bed to explore if severity and nature of CA can distinguish them apart. We quantitatively assessed structural and numerical centrosomal aberrations for each patient sample in a large-cohort of grade-matched TNBC (n = 30) and non-TNBC (n = 98) cases employing multi-color confocal imaging. Our data establish differences in incidence and severity of CA between TNBC and non-TNBC cell lines and clinical specimens. We found strong correlation between CA and aggressiveness markers associated with metastasis in 20 pairs of grade-matched TNBC and non-TNBC specimens (p < 0.02). Time-lapse imaging of MDA-MB-231 cells harboring amplified centrosomes demonstrated enhanced migratory ability. Our study bridges a vital knowledge gap by pinpointing that CA underlies breast cancer aggressiveness. This previously unrecognized organellar inequality at the centrosome level may allow early-risk prediction and explain higher tumor aggressiveness and mortality rates in TNBC patients.

BackgroundAmplified centrosomes in cancers are recently garnering a lot of attention as an emerging hub of diagnostic, prognostic and therapeutic targets. Ovarian adenocarcinomas commonly harbor supernumerary centrosomes that drive chromosomal instability. A centrosome clustering molecule, KIFC1, is indispensable for the viability of extra centrosome-bearing cancer cells, and may underlie progression of ovarian cancers.MethodsCentrosome amplification in low- and high- grade serous ovarian adenocarcinomas was quantitated employing confocal imaging. KIFC1 expression was analyzed in ovarian tumors using publically-available databases. Associated grade, stage and clinical information from these databases were plotted for KIFC1 gene expression values. Furthermore, interactions and functional annotation of KIFC1 and its highly correlated genes were studied using DAVID and STRING 9.1.ResultsClinical specimens of ovarian cancers display robust centrosome amplification and deploy centrosome clustering to execute an error-prone mitosis to enable karyotypic heterogeneity that fosters tumor progression and aggressiveness. Our in silico analyses showed KIFC1 overexpression in human ovarian tumors (n = 1090) and its upregulation associated with tumor aggressiveness utilizing publically-available gene expression databases. KIFC1 expression correlated with advanced tumor grade and stage. Dichotomization of KIFC1 levels revealed a significantly lower overall survival time for patients in high KIFC1 group. Intriguingly, in a matched-cohort of primary (n = 7) and metastatic (n = 7) ovarian samples, no significant differences in KIFC1 expression were detectable, suggesting that high KIFC1 expression may serve as a marker of metastases onset. Nonetheless, KIFC1 levels in both primary and matched metastatic sites were significantly higher compared to normal tissue . Ingenuity based network prediction algorithms combined with pre-established protein interaction networks uncovered several novel cell-cycle related partner genes on the basis of interconnectivity, illuminating the centrosome clustering independent agenda of KIFC1 in ovarian tumor progression.ConclusionsOvarian cancers display amplified centrosomes, a feature of aggressive tumors. To cope up with the abnormal centrosomal load, ovarian cancer cells upregulate genes like KIFC1 that are known to induce centrosome clustering. Our data underscore KIFC1 as a putative biomarker that predicts worse prognosis, poor overall survival and may serve as a potential marker of onset of metastatic dissemination in ovarian cancer patients.

This study tested whether hepatic steatosis sensitizes liver to toxicant-induced injury and investigated the potential mechanisms of hepatotoxic sensitivity. Male Sprague-Dawley rats were fed a methionine-and choline-deficient diet for 31 days to induce steatosis. On the 32nd day, administration of a nonlethal dose of CCl 4 (2 mL/kg, intraperitoneally) yielded 70% mortality in steatotic rats 12-72 hours after CCl 4 administration, whereas all nonsteatotic rats survived. Neither CYP2E1 levels nor covalent binding of [ 14 C]CCl 4 -derived radiolabel differed between the groups, suggesting that increased bioactivation is not the mechanism for this amplified toxicity. Cell division and tissue repair, assessed by [ 3 H]thymidine incorporation and proliferative cell nuclear antigen assay, were inhibited in the steatotic livers after CCl 4 administration and led to progressive expansion of liver injury culminating in mortality. The hypothesis that fatty hepatocytes undergo cell cycle arrest due to (1) an inability to replenish ATP due to overexpressed uncoupling protein-2 (UCP-2) or (2) induction of growth inhibitor p21 leading to G 1 /S phase arrest was tested. Steatotic livers showed 10-fold lower ATP levels due to upregulated UCP-2 throughout the time course after N onalcoholic fatty liver disease (NAFLD) is defined as deposition of fat in the liver (hepatic steatosis) that is not caused by chronic ingestion of alcohol. The terms NAFLD and nonalcoholic steatohepatitis (NASH) are often used interchangeably; however, NASH specifically refers to that subset of patients with NAFLD who have evidence of inflammation on liver biopsy. NAFLD and NASH are highly prevalent among obese and type II diabetic patients suggesting that obesity and diabetes are predisposing factors for NAFLD and NASH. [1][2][3] The prevalence of obesity in Western populations is currently 10% to 25% and is expected to rise in the future; this increase will result in a parallel rise in NAFLD and NASH.Ob/Ob mice and Zucker fa/fa rats are genetic models for NAFLD. 4 Liver regeneration after two thirds partial hepatectomy is inhibited in obese diabetic Zucker fa/fa rats and ob/ob mice with fatty liver, indicating that ability of the liver to regenerate is decreased in hepatic steatosis. [4][5][6] These observations show that inhibition of cell division does not require inflammation associated with NASH and can occur in a nonalcoholic fatty liver (hepatic steatosis) devoid of the onset of inflammation. However, the response of nonalcoholic fatty liver to drug/chemicalinduced hepatotoxicity is still unknown.The purpose of this study was to investigate whether steatosis alone renders the liver susceptible to amplifica-

Particularly interesting new cysteine-histidine-rich protein (PINCH) protein is part of the ternary complex known as the IPP (integrin linked kinase (ILK)-PINCH-Parvin-α) complex. PINCH itself binds to ILK and to another protein known as Rsu-1 (Ras suppressor 1). We generated PINCH 1 and PINCH 2 Double knockout mice (referred as PINCH DKO mice). PINCH2 elimination was systemic whereas PINCH1 elimination was targeted to hepatocytes. The genetically modified mice were born normal. The mice were sacrificed at different ages after birth. Soon after birth, they developed abnormal hepatic histology characterized by disorderly hepatic plates, increased proliferation of hepatocytes and biliary cells and increased deposition of extracellular matrix. After a sustained and prolonged proliferation of all epithelial components, proliferation subsided and final liver weight by the end of 30 weeks in livers with PINCH DKO deficient hepatocytes was 40% larger than the control mice. The livers of the PINCH DKO mice were also very stiff due to increased ECM deposition throughout the liver, with no observed nodularity. Mice developed liver cancer by one year. These mice regenerated normally when subjected to 70% partial hepatectomy and did not show any termination defect. Ras suppressor 1 (Rsu-1) protein, the binding partner of PINCH is frequently deleted in human liver cancers. Rsu-1 expression is dramatically decreased in PINCH DKO mouse livers. Increased expression of Rsu-1 suppressed cell proliferation and migration in HCC cell lines. These changes were brought about not by affecting activation of Ras (as its name suggests) but by suppression of Ras downstream signaling via RhoGTPase proteins. In conclusion, our studies suggest that removal of PINCH results in enlargement of liver and tumorigenesis. Decreased levels of Rsu-1, a partner for PINCH and a protein often deleted in human liver cancer, may play an important role in the development of the observed phenotype.

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Reprogramming factors have been used to induce pluripotent stem cells as an alternative to somatic cell nuclear transfer technology in studies targeting disease models and regenerative medicine. The neuronal repressor REST maintains self renewal and pluripotency in mouse embryonic stem cells by maintaining the expression Oct3/4, Nanog, and cMyc. We report that primary hepatocytes express REST and most of the reprogramming factors in culture. Their expression is upregulated by hepatocyte growth factor (HGF) and epidermal growth factor (EGF). REST inhibition results in downregulation of reprogramming factor expression, increased apoptosis, decreased proliferation, and cell death. The reprogramming factors are also upregulated after 70% partial hepatectomy in vivo. These findings show that genes inducing iPS phenotype, even though expressed at lower levels than embryonic stem cells, they nonetheless are associated with control of apoptosis and cell proliferation in hepatocytes in culture and may play a role for such processes during liver regeneration.