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Márquez MG, del Carmen Fernández-Tome M, Favale NO, Pescio LG, Sterin-Speziale NB. Bradykinin modulates focal adhesions and induces stress fiber remodeling in renal papillary collecting duct cells. Am J Physiol Renal Physiol 294: F603-F613, 2008. First published December 26, 2007 doi:10.1152/ajprenal.00234.2007 are specialized regions of cell attachment to the extracellular matrix. Previous works have suggested that bradykinin (BK) can modulate cell-matrix interaction. In the present study, we used a physiological cellular model to evaluate the potential role of BK in modulating FAs and stress fibers. We performed a quantitative morphometric analysis of FAs in primary cultured rat renal papillary collecting duct cells, which included size, axial ratio (shape), and average length. After 1, 5, or 10 min of incubation with BK, cultured cells were immunostained and analyzed by confocal microscopy. Although the shape of FAs was not altered, BK induced a decrease in the number of vinculin-stained FAs per cell, and a decrease in both their size and their average length, but not in talin-containing FAs, thus suggesting that BK could be inducing a restructuring of FAs. BK also induced a remodeling of the actin filament assemblies rather than their dissipation. Since we have previously demonstrated that BK stimulates activation of PLC␤ in rat renal papillae, we attempted to determine whether BK can modulate FA restructuring by this mechanism, by pretreating cultured cells with the PLC␤ inhibitor U73122. The present study, performed under physiological conditions with cells that were not genetically manipulated, provides new experimental evidence supporting the notion that the intrarenal hormone BK modulates FAs and actin cytoskeleton organization through a mechanism that involves the activation of PLC␤. We propose this finding as a novel mechanism for BK modulation of tubular collecting duct function. stress fibers; cell-matrix adhesion; vinculin; talin; actin filaments FOCAL ADHESIONS (FAs) are specialized regions of the cell involved in the attachment to the extracellular matrix, where actin stress fibers and the extracellular matrix are connected (4, 35). FA assembly occurs through the binding of the integrinextracellular domain to extracellular matrix proteins, followed by the interaction of the ␤-integrin cytoplasmic domain with talin. Talin can then recruit vinculin, which, in turn, binds other FA proteins (15,21,33). In typical adherent cultured cells, a large number of distinct FAs, ranging in size from less than a square micrometer to several square micrometers, can be detected. The number of FAs, their size, and distribution can vary from one cell to another or even within a single cell, and their morphological diversity may be affected by multiple factors (36). In migrating cells, integrin-mediated adhesions are molecularly heterogeneous and appear in different forms such as "classic" FAs, fibrillar adhesions, and focal complexes. Each of these has a typical morphology and molecular composition (34). Besides h...

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Background and aims Non‐alcoholic fatty liver (NAFLD) and its more serious form non‐alcoholic steatohepatitis increase risk of hepatocellular carcinoma (HCC). Lipid metabolic alterations and its role in HCC development remain unclear. SPARC (Secreted Protein, Acidic and Rich in Cysteine) is involved in lipid metabolism, NAFLD and diabetes, but the effects on hepatic lipid metabolism and HCC development is unknown. The aim of this study was to evaluate the role of SPARC in HCC development in the context of NAFLD. Methods Primary hepatocyte cultures from knockout (SPARC−/−) or wild‐type (SPARC+/+) mice, and HepG2 cells were used to assess the effects of free fatty acids on lipid accumulation, expression of lipogenic genes and de novo triglyceride (TG) synthesis. A NAFLD‐HCC model was stabilized on SPARC−/− or SPARC+/+ mice. Correlations among SPARC, lipid metabolism‐related gene expression patterns and clinical prognosis were studied using HCC gene expression dataset. Results SPARC−/− mice increases hepatic lipid deposits over time. Hepatocytes from SPARC−/− mice or inhibition of SPARC by an antisense adenovirus in HepG2 cells resulted in increased TG deposit, expression of lipid‐related genes and nuclear translocation of SREBP1c. Human HCC database analysis revealed that SPARC negatively correlated with genes involved in lipid metabolism, and with poor survival. In NAFLD‐HCC murine model, the absence of SPARC accelerates HCC development. RNA‐seq study revealed that pathways related to lipid metabolism, cellular detoxification and proliferation were upregulated in SPARC−/− tumour‐bearing mice. Conclusions The absence of SPARC is associated with an altered hepatic lipid metabolism, and an accelerated NAFLD‐related HCC development.