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Volume 124 (2004)
- Issue 6 Pages 375-
- Issue 5 Pages 301-
- Issue 4 Pages 201-
- Issue 3 Pages 135-
- Issue 2 Pages 69-
- Issue 1 Pages 3-
Volume 124, Issue 5
Displaying 1-6 of 6 articles from this issue
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Reviews: Novel Functions of Calmodulin-Dependent Signal Transduction and Drug Development
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Masahiro SOKABE, Keiji NARUSE, Tang Qiong-Yao2004 Volume 124 Issue 5 Pages 301-310
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSCells can respond to a variety of mechanical stimuli such as tension, pressure, and shear stress. However, the mechanisms of mechanotransduction are largely unknown. The major reason for this lies in the ambiguity of the molecular entity of cell mechanosensors. Currently only MS (mechanosensitive) channels conform to an established class of mechanosensors due to the firm and detailed analyses by electrophysiolgy. Although molecular structures of MS channels are known for limited members, higher order structures of bacterial MS channels have been resolved and their detailed structure-function studies are in progress. In contrast, molecular and biophysical analyses of eukaryote MS channels, which may attract much attention, are yet not well-studied. Although many candidate molecules have been proposed as the cell mechanosensor, currently only 2-pore-domain K channels (TREK/TRAAK) and SAKCA, a new class of MS channel introduced here, may be the subjects eligible for rigorous electrophysiological analyses. On the other hand, lack of specific blockers to MS channels is another reason why the progress in this field is slow. Gadolinium (Gd3+) has been extensively used as a potent blocker of MS channels, but its nonspecific actions have limited its usefulness. Very recently, a promising 35 mer peptide, which can be more specific for MS channels, named GsMTx-4 has been isolated from spider venom. This peptide is interesting because it inhibits stretch-induced atrial fibrillation, which may involve MS channel activation and thus can be used as a basis for developing a new class of drugs to cure heart failure. This short review deals with recent progresses in MS channel studies and the structure-function of SAKCA, a recently cloned MS channel from heart, as well as its interaction with the new MS channel blocker GsMTx-4.
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Michitaka MASUDA, Naoko KOGATA, Naoki MOCHIZUKI2004 Volume 124 Issue 5 Pages 311-318
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSFluid shear stress (FSS) induces many forms of responses, including phosphorylation of ERK in endothelial cells (ECs). We have earlier reported that FSS and hyper-osmotic shock rapidly induce tyrosine phosphorylation of PECAM-1 (CD31). The phosphorylated PECAM-1 acts as a plasma membrane anchoring site for SHP2, a protein tyrosine phosphatase involved in the signal transmission from receptor tyrosine kinases to ERK. Osmotic shock also induces transient ERK activation in ECs. The osmotic-shock-induced ERK activation but not p38 MAP kinase activation was dependent on the PECAM-1 engagement and was blocked by its downregulation. When magnetic beads coated with antibodies against the extracellular domain of PECAM-1 were attached to ECs and tugged by magnetic force, PECAM-1 associated with the beads was tyrosine phosphorylated. ERK was also phosphorylated in these cells. Binding of the beads by itself or pulling on the cell surface using poly-L-lysine coated beads did not induce phosphorylation of PECAM-1 and ERK. These results suggest that PECAM-1 is a mechanotransduction molecule.
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Kimiko YAMAMOTO, Joji ANDO2004 Volume 124 Issue 5 Pages 319-328
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSThe mechanisms by which shear stress elevates intracellular Ca2+ in endothelial cells (EC) are not fully understood. Here we report that endogenously released ATP contributes to shear stress-induced Ca2+ responses. Application of a flow of Hank's balanced solution to human pulmonary artery EC (HPAEC) elicited shear stress-dependent increases in Ca2+ concentration. Chelation of extracellular Ca2+ with EGTA completely abolished the Ca2+ responses, whereas the phospholipase C inhibitor U-73122 and the Ca2+-ATPase inhibitor thapsigargin had no effect, indicating that the response was due to the influx of extracellular Ca2+. The Ca2+ influx was significantly suppressed by apyrase, which degrades ATP, and by antisense oligonucleotide targeted to P2X4 receptors. A luciferase luminometric assay showed that shear stress induced dose-dependent release of ATP. When the ATP release was inhibited by the ATP synthase inhibitors angiostatin or oligomycin, the Ca2+ influx was markedly suppressed but was restored by removal of these inhibitors or addition of extracellular ATP. These results suggest that shear stress stimulates HPAEC to release ATP, which activates Ca2+ influx via P2X4 receptors.
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Hisayuki OHATA, Takeharu NIIOKA, Myung-sook KIM, Sanae ANDO, Masayuki ...2004 Volume 124 Issue 5 Pages 329-335
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSThe mechanotransduction mechanisms play an important role in regulation of specific cellular response or maintenance of cellular homeostasis in a wide variety of cell types. Increase in intracellular free Ca2+ concentration ([Ca2+]i) is an important signal in the first step of mechanotransduction. Mechanosensitive (MS) cation channels are thought to be a putative pathway of Ca2+ entry; however, the molecular mechanisms remain unclear. We have previously demonstrated that lysophosphatidic acid (LPA), a bioactive phospholipid present in human plasma, sensitizes the response of [Ca2+]i to mechanical stress in cultured smooth muscle cells, cultured lung epithelial cells, and cultured lens epithelial cells. Using real-time confocal microscopy, local increases in [Ca2+]i in several regions within the cell subjected to mechanical stress were clearly visualized in cultured bovine lens epithelial cells and cultured vascular endothelial cells in the presence of LPA. We called the phenomenon “Ca2+ spots”. Pharmacological studies revealed that the Ca2+ spot is an elementary Ca2+-influx event through MS channels. In this review, possible physiological and pathophysiological roles of LPA as a mechanosensitizer are discussed.
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Yoshiyuki TANABE, Koichi NAKAYAMA2004 Volume 124 Issue 5 Pages 337-344
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSObesity frequently promotes a variety of cardiovascular diseases including atherosclerosis, hypertension, and type 2 diabetes. In a view of both the preventive and therapeutic aspects of the abovementioned diseases, most intensive clinical interventions have been primarily directed at decreasing excessive amounts of fat tissue by a change in the balance between intake and expenditure of energy; such changes are typically effected via daily exercise and diet control. Mechanical stimuli such as stretching and rubbing of fat tissues using gymnastic exercises or massage are believed to decrease obesity; however, there is no report concerning the direct effect of the mechanical stimulation on adipocytes. Here, we demonstrated that cyclic stretch inhibited adipocyte differentiation of mouse 3T3-L1 cells, which was attributable to a reduced expression of adipogenic transcription factor peroxisome proliferator-activated receptor (PPAR)γ2 via the activation of an extracellular signal-regulated protein kinase (ERK) pathway. The inhibitory effect of the cyclic stretching on the differentiation of 3T3-L1 cells could be restored by troglitazone, a synthetic ligand for PPARγ. Our results provide a molecular basis for the physiological significance of the local application of mechanical stimuli to fat tissues, which is totally independent of a mechanism for systemic energy consumption.
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Technical Note
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Masafumi KAKEI, Katsuya DEZAKI, Toshihiko YADA2004 Volume 124 Issue 5 Pages 345-352
Published: 2004
Released on J-STAGE: October 22, 2004
JOURNAL FREE ACCESSRegulation of blood glucose is a fundamental homeostasis in the body. Insulin is released from pancreatic β-cells in response to changes in blood glucose, the defect of which leads to impaired insulin secretion and diabetes mellitus. Pancreatic β-cells that release insulin occupy approximately 70% of the islet cells, while α, δ, PP-cells are also present in islets. Therefore, analysis of β-cells is a direct approach for studying mechanisms of insulin secretion. Insulin secretion is regulated by cytosolic Ca2+ in β-cells, and its concentration and localization can be measured in real time by fluorescence imaging using indicators. Glucose metabolism is assessed by measurements of NAD(P)H by its autofluorescence. Furthermore, β-cells are equipped with ion channels, which transduce glucose-evoked metabolic signals to electric signals. Electrophysiological analysis by patch clamp techniques detects the activity of various ion channels and membrane potential. Several β-cell lines including HIT, MIN6, INS1, RIN, and βTC are used; however, they do not necessarily retain normal responsiveness to glucose. Therefore, analysis of physiologic functions of β-cells requires the use of acutely isolated or primary cultured β-cells from normal animals. The methods for preparation of islets and single β-cells using collagenase can be applied to a variety of animals of small to large sizes, which can produce islets and β-cells with physiological responsiveness to glucose. These primary cultured β-cells can be used for elucidating signal transduction mechanisms, and evaluating effects of drugs, providing excellent tools for physiological, pharmacological, and disease-oriented studies.
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