Abstract-Gelsolin, a calcium-regulated actin severing and capping protein, is highly expressed in murine and human hearts after myocardial infarction and is associated with progression of heart failure in humans. The biological role of gelsolin in cardiac remodeling and heart failure progression after injury is not defined. To elucidate the contribution of gelsolin in these processes, we randomly allocated gelsolin knockout mice (GSN Ϫ/Ϫ ) and wild-type littermates (GSN ϩ/ϩ ) to left anterior descending coronary artery ligation or sham surgery. We found that GSN Ϫ/Ϫ mice have a surprisingly lower mortality, markedly reduced hypertrophy, smaller late infarct size, less interstitial fibrosis, and improved cardiac function when compared with GSN ϩ/ϩ mice. Gene expression and protein analysis identified significantly lower levels of deoxyribonuclease (DNase) I and reduced nuclear translocation and biological activity of DNase I in GSN Ϫ/Ϫ mice. Absence of gelsolin markedly reduced DNase I-induced apoptosis. The association of hypoxia-inducible factor (HIF)-1␣ with gelsolin and actin filaments cleaved by gelsolin may contribute to the higher activation of DNase. The expression pattern of HIF-1␣ was similar to that of gelsolin, and HIF-1␣ was detected in the gelsolin complex by coprecipitation and HIF-1␣ bound to the promoter of DNase I in both gel-shift and promoter activity assays. Furthermore, the phosphorylation of Akt at Ser473 and expression of Bcl-2 were significantly increased in GSN Ϫ/Ϫ mice, suggesting that gelsolin downregulates prosurvival factors. Our investigation concludes that gelsolin is an important contributor to heart failure progression through novel mechanisms of HIF-1␣ and DNase I activation and downregulation of antiapoptotic survival factors. Gelsolin inhibition may form a novel target for heart failure therapy. Key Words: gelsolin Ⅲ myocardial infarction Ⅲ cardiac remodeling Ⅲ apoptosis Ⅲ deoxyribonuclease I G elsolin is a widely distributed actin-binding protein consisting of six domains (G1 to -6) with a salt bridge between G2 and G6 (latch helix) when it is inactive. Gelsolin mediates multiple cellular functions including cell motility, morphogenesis, and actin cytoskeletal remodeling. 1,2 The most extensively examined roles of gelsolin are its actin filament severing, capping, uncapping, and nucleating activities. The severing activity of gelsolin is regulated by Ca 2ϩ and pH, whereas polyphosphoinositides (particularly PIP2) regulate uncapping. 2,3 In addition to its remodeling of actin filaments, gelsolin can also regulate signal transduction through the integrin and small GTPase (Ras-Rac)-mediated pathways. 4,5 There are conflicting data on the pro-and antiapoptotic functions of gelsolin. 6 -10 On the one hand, full-length gelsolin, its C-terminal half, and its phosphatidylinositol 4,5-bisphosphate complexes are mostly antiapoptotic. 8,11 In contrast, the N-terminal half of gelsolin is potentially proapoptotic because gelsolin-deficient cells show retarded onset of apoptosis and tra...
The aims of the present study were to investigate the presence and distribution of NPY and the Y1 receptor in endocardial endothelial cells (EECs), to verify if EECs can release NPY, and to determine if the effect of NPY on intracellular calcium is mediated via the Y1 receptor. Immunofluorescence, 3-D confocal microscopy and radioimmunoassay techniques were used on 20-week-old human fetal EECs. Our results showed that NPY and the Y1 receptor are present in human EECs (hEECs) and that their distributions are similar, the fluorescence labelling being higher in the nucleus and more particularly at the level of the nuclear envelope when compared with the cytosol. Using radioimmunoassay, we demonstrated that EECs are a source of NPY and can secrete this peptide upon a sustained increase of intracellular calcium ([Ca]i). Using fluo-3 and 3-D confocal microscopy technique, superfusion of hEECs as well as EECs isolated from rat adult hearts with increasing concentrations of NPY induced a dose-dependent, sustained increase in free cytosolic and nuclear Ca2+ levels. This effect of NPY on EEC [Ca]i was completely reversible upon washout of NPY and was partially blocked by BIBP3226, a selective Y1 receptor antagonist. The results suggest that NPY and Y1 receptors are present in the EECs of 20-week-old human fetal heart and they share the same distribution and localization inside the cell. In addition, EECs are able to secrete NPY in response to an increase in [Ca]i, and the Y1 receptor as well as other NPY receptors seem to participate in mediating the effects of NPY on [Ca]i in these cells. Thus, NPY released by EECs may modulate excitation-secretion coupling of these cells.
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