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91.
Isaline Rowe Marco Chiaravalli Klaus B. Piontek Gregory G. Germino Alessandra Boletta 《Biochemical and biophysical research communications》2014
The PKD1 gene is essential for a number of biological functions, and its loss-of-function causes autosomal dominant polycystic kidney disease (ADPKD). The gene is developmentally regulated and believed to play an essential role in renal development. Previous studies have shown that manipulating murine renal organ cultures with dominant-negative forms of the Pkd1 gene impaired ureteric bud (UB) branching. In the current study, we analyzed different stages of renal development in two distinct mouse models carrying either a null mutation or inactivation of the last two exons of Pkd1. Surprisingly, metanephric explants from Pkd1-deleted kidneys harvested at day E11.5 did not show defects of UB branching and elongation, estimated by cytokeratin staining on fixed tissues or by Hoxb7-GFP time-lapse imaging. However, renal explants from Pkd1-mutants isolated at day E14.5 showed impaired nephrogenesis. Notably, we observed cell migratory defects in the developing endothelial compartment. Previous studies had implicated the Pkd1 gene in controlling cell migration and collagen deposition through PI3 kinases. In line with these studies, our results show that wild-type explants treated with PI3-kinase inhibitors recapitulate the endothelial defects observed in Pkd1 mutants, whereas treatment with VEGF only partially rescued the defects. Our data are consistent with a role for the Pkd1 gene in the endothelium that may be required for proper nephrogenesis. 相似文献
92.
Ming-Jen Hsu Joen-Rong Sheu Chien-Huang Lin Ming-Yi Shen Chung Y. Hsu 《Biochimica et Biophysica Acta (BBA)/General Subjects》2010
Prevailing evidence suggests that amyloid beta peptide (Aβ), a key mediator in age-dependent neuronal and cerebrovascular degeneration, activates death signaling processes leading to neuronal as well as non-neuronal cell death in the central nervous system. A major cellular event in Aβ-induced death of non-neuronal cells, including cerebral endothelial cells, astrocytes and oligodendrocytes, is mitochondrial dysfunction. The death signaling cascade upstream of mitochondria entails Aβ activation of neutral sphingomyelinase, resulting in the release of ceramide from membrane sphingomyelin. Ceramide then activates protein phosphatase 2A (PP2A), a member in the ceramide-activated protein phosphatase (CAPP) family. PP2A dephosphorylation of Akt and FKHRL1 plays a pivotal role in Aβ-induced Bad translocation to mitochondria and transactivation of Bim. Bad and Bim are pro-apoptotic proteins that cause mitochondrial dysfunction characterized by excessive ROS formation, mitochnondrial DNA (mtDNA) damage, and release of mitochondrial apoptotic proteins including cytochrome c, apoptosis inducing factor (AIF), endonuclease G and Smac. The cellular events activated by Aβ to induce death of non-neuronal cells are complex. Understanding these death signaling processes will aid in the development of more effective strategies to slow down age-dependent cerebrovascular degeneration caused by progressive cerebrovascular Aβ deposition. 相似文献
93.
Maria Luisa Balestrieri Alfonso Giovane Lara Milone Francesca Felice Carmela Fiorito Valeria Crudele Annaclaudia Esposito Raffaele Rossiello Pellegrino Biagio Minucci Bartolomeo Farzati Luigi Servillo Claudio Napoli 《Journal of biochemical and molecular toxicology》2010,24(6):351-360
Exposure of human endothelial progenitor cells (EPCs) to tumor necrosis factor‐α (TNF‐α) reduced their number and biological activity. Yet, signal transduction events linked to TNF‐α action are still poorly understood. To address this issue, we examined the possible effect of fasudil and Y27632, two inhibitors of Rho kinase pathway, which is involved in endothelial dysfunction, atherosclerosis, and in‐ flammation. Results demonstrated that incubation with fasudil starting from 50 μM but not Y27632 determined a dose‐dependent improvement of EPC number during exposure to TNF‐α (P < 0.05 vs. TNF‐α alone). Analysis of the signal transduction pathway activated by TNF‐α revealed that the increased expression of p‐p38 was not significantly altered by fasudil. Instead, fasudil blocked the TNF‐α induced phosphorylation of Erk1/2 (P < 0.05 vs. TNF‐α) as well as the inhibitor of Erk1/2‐specific phosphorylated form, i.e., PD98059 (P < 0.05 vs. TNF‐α). These results were confirmed by analysis of these kinases by confocal microscopy. Finally, 2D‐DIGE and MALDI‐TOF/TOF analysis of EPCs treated with fasudil revealed increased expression levels of an actin‐related protein and an adenylyl cyclase associated protein and decreased expression levels of proteins related to radical scavenger and nucleotide metabolism. These findings suggest that fasudil positively affects EPC number and that other major signals might take part to this complex pathway. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:351–360, 2010; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.20345 相似文献
94.
Hanna Mannell Ariane Hammitzsch Ramona Mettler Ulrich Pohl Florian Krtz 《Cellular signalling》2010,22(1):88-96
Angiogenesis initiation is crucially dependent on endothelial proliferation and can be stimulated by the fibroblast growth factor 2 (FGF-2). The DNA dependent protein kinase (DNA-PK), long known for its importance in repairing DNA double strand breaks, belongs to the phosphatidylinositol-3 kinase (PI3-K) super family and has recently been identified as one of the enzymes phosphorylating and activating Akt. Due to its similarity with PI3-K, we hypothesized that DNA-PK may have similar effects on endothelial angiogenic processes and signalling. We used primary endothelial cells (HUVEC and PAEC) and human microvascular endothelial cells (HMEC) to study the role of DNA-PK in endothelial proliferation and signalling. DNA-PKcs suppression with the compound NU7026 or with siRNA induced basal endothelial cell proliferation as well as enhanced FGF-2 dependent proliferation. This was associated with an increase in phosphorylated Akt. Tube formation was not affected by DNA-PKcs clearly showing that the role of DNA-PK in endothelial processes differs from that of PI3-K. Our findings indicate DNA-PK as an important enzyme maintaining the quiescent endothelial phenotype by actively inhibiting Akt thus restraining endothelial cell proliferation preventing excessive growth. 相似文献
95.
Su Li Hui Qu Jianwei Hao Jinfu Sun Huancheng Guo Changming Guo Boxing Sun Changchun Tu 《Biochimica et Biophysica Acta - Proteins and Proteomics》2010,1804(9):1882-1888
Endothelial cells are the main target of classical swine fever virus during infection, and extensive hemorrhage is the most typical clinical sign of classical swine fever. To investigate the molecular mechanism of hemorrhagic pathogenesis, two-dimensional difference gel electrophoresis with fluorescent dyes (2D-DIGE) was used to analyze the proteomic profile of primary porcine umbilical vein endothelial cells (PUVECs) following CSFV infection. Of 15 protein spots with differential expression, 8 were characterized by MALDI-TOF-MS/MS in infected PUVECs at 48 h p.i.: moesin, peroxiredoxin 6, stathmin-1, a protein similar to nascent polypeptide-associated complex alpha subunit isoform 2, phosphoglycerate kinase 1, glucosidase II, transketolase and α-tubulin. These could be sorted into 5 functional groups: glycometabolism, cell proliferation, anti-oxidative stress, inflammatory response and cytoskeleton. Western blot and real-time RT-PCR analysis confirmed the down-regulation of phosphoglycerate kinase 1 (PGK1) and up-regulation of moesin identified by 2D-DIGE. Pathway analysis of these 15 differentially expressed proteins showed that CSFV infection altered the metabolism, cytoskeleton and cell proliferation of PUVECs, and that consequently an inflammatory response was induced. 相似文献
96.
脂肪来源的间充质干细胞具有较强的体外增殖能力,因具有生物学特性稳定、来源充足、体外培养条件低等优势已引起各国学者的关注。脂肪间充质干细胞凭借其多向分化潜能,是人体干细胞库潜在的重要来源之一。目前,研究人员已成功地在体外将其诱导为内皮细胞,成骨细胞、成软骨细胞、脂肪前体细胞、平滑肌细胞,心肌细胞、神经样细胞等。就脂肪来源的间充质干细胞体外定向诱导分化血管细胞的研究进展作一综述。 相似文献
97.
Functional and morphological responses of endothelial cells (ECs) to fluid shear stress are thought to be mediated by several mechanosensitive molecules. However, how the force due to fluid shear stress applied to the apical surface of ECs is transmitted to the mechanosensors is poorly understood. In the present paper, we performed an analysis of an intracellular mechanical field by observation of the deformation behaviors of living ECs exposed to shear stress with a novel experimental method. Lateral images of human umbilical vein ECs before and after the onset of flow were obtained by confocal microscopy, and image correlation and finite element analysis were performed for quantitative analyses of subcellular strain due to shear stress. The shear strain of the cells changed from 1.06 ± 1.09% (mean ± SD) to 4.67 ± 1.79% as the magnitude of the shear stress increased from 2 to 10 Pa. The nuclei of ECs also exhibited shear deformation, which was similar to that observed in cytoplasm, suggesting that nuclei transmit forces from apical to intracellular components, as well as cytoskeletons. The obtained strain-stress relation resulted in a mean shear modulus of 213 Pa for adherent ECs. These results provide a mechanical perspective on the investigation of flow-sensing mechanisms of ECs. 相似文献
98.
Yosuke Ueki Yuhei Uda Masaaki Sato 《Biochemical and biophysical research communications》2010,395(3):441-446
Fluid shear stress (FSS) acting on the apical surface of endothelial cells (ECs) can be sensed by mechano-sensors in adhesive protein complexes found in focal adhesions and intercellular junctions. This sensing occurs via force transmission through cytoskeletal networks. This study quantitatively evaluated the force transmitted through cytoskeletons to the mechano-sensors by measuring the FSS-induced strain on SFs using live-cell imaging for actin stress fibers (SFs). FSS-induced bending of SFs caused the SFs to align perpendicular to the direction of the flow. In addition, the displacement vectors of the SFs were detected using image correlation and the FSS-induced axial strain of the SFs was calculated. The results indicated that FSS-induced strain on SFs spanned the range 0.01-0.1% at FSSs ranging from 2 to 10 Pa. Together with the tensile property of SFs reported in a previous study, the force exerted on SFs was estimated to range from several to several tens of pN. 相似文献
99.
100.
Bruna Vinci Ellen Murphy Elisabetta Iori Maria Cristina Marescotti Angelo Avogaro Arti Ahluwalia Professor 《Biotechnology journal》2010,5(6):618-626
Static cell culture has serious limitations in its ability to represent cellular behaviour within a live organism. In vivo, cells are constantly exposed to the flow of bodily fluids and contact with other cell types. Bioreactors provide the opportunity to study cells in an environment that more closely resembles the in vivo setting because cell cultures can be exposed to dynamic flow in contact with or in proximity to other cell types. In this study we compared the metabolic profile of a dynamic cell culture system to that of a static cell culture in three different cellular phenotypes: adipocytes, endothelial cells and hepatocytes. Albumin, glucose, free fatty acids, glycerol, and lactate were measured over 48 h. We show that all three cell types have increased glucose uptake in the presence of flow; lactate release was also significantly affected. We provide robust evidence that the presence of flow significantly modifies cellular metabolism. While flow provides a more uniform nutrient distribution and increases metabolite turnover, our results indicate that different cell types have specific metabolic responses to flow, suggesting cell-specific flow-regulated activation of metabolite signalling pathways. 相似文献