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Calyculin A retraction of mature megakaryocytes proplatelets from embryonic stem cells |
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| Authors: | Tamaru Satoshi Kitajima Kenji Nakano Tohru Eto Koji Yazaki Akira Kobayashi Toshihiko Matsumoto Takeshi Wada Hideo Katayama Naoyuki Nishikawa Masakatsu |
| Affiliation: | a Institute of Human Research Promotion and Drug Development, Mie University, Faculty of Medicine, Mie 514-8507, Japan b Development of Pathology, Medical School and Graduate School of Frontier Biosciences, Osaka University, Osaka 565-0871, Japan c Laboratory of Stem Cell Therapy, Center for Experimental Medicine, The Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan d Department of Hematology and Oncology, Mie University Graduate School of Medicine, Mie 514-8507, Japan e Department of Molecular and Laboratory Medicine, Mie University Graduate School of Medicine, Mie 514-8507, Japan |
| Abstract: | Platelets are produced by megakaryocytes (MKs) through proplatelet formation (PPF), or cytoplasmic extensions, in vitro. Through the use of video-enhanced light microscopy, as well as localization of cytoskeletal proteins by confocal microscopy, the reaction of fully mature MK proplatelets, derived from murine embryonic stem cells, to various agents was studied. Calyculin A (protein phosphatase 1/2A inhibitor) treatment induced proplatelet retraction. In MKs with PPF, the expression of actin, myosin IIA, monophosphorylated myosin light chain (MLC-P1), and diphosphorylated myosin light chain (MLC-P2) was diffusely located. Following calyculin A treatment, actin was diffusely localized in retracted MKs and was expressed particularly in the periphery. MLC-P1 was also localized primarily in the periphery; however, MLC-P2 was expressed mostly in the inner area of proplatelets. Protein phosphatase inhibitors may result in increased hyperphosphorylation of localized MLC, which could alter the balance of actomyosin force in a cell, and therefore induce proplatelets retraction. |
| Keywords: | Actin Calyculin A Embryonic stem cell Megakaryocyte Myosin light chain phosphorylation OP9 Proplatelet formation |
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