PMID- 26321737 OWN - NLM STAT- MEDLINE DCOM- 20151228 LR - 20220311 IS - 1618-095X (Electronic) IS - 0944-7113 (Linking) VI - 22 IP - 10 DP - 2015 Sep 15 TI - Apocynin inhibits the upregulation of TGF-beta1 expression and ROS production induced by TGF-beta in skeletal muscle cells. PG - 885-93 LID - S0944-7113(15)00210-X [pii] LID - 10.1016/j.phymed.2015.06.011 [doi] AB - BACKGROUND: Pure apocynin, which can be traditionally isolated and purified from several plant species such as Picrorhiza kurroa Royle ex Benth (Scrophulariaceae), acts as an inhibitor of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity inhibiting its production of reactive oxygen species (ROS). Transforming growth factor type beta 1 (TGF-beta1) is a growth factor that produces inhibition of myogenesis, diminution of regeneration and induction of atrophy in skeletal muscle. The typical signalling that is activated by TGF-beta involves the Smad pathway. PURPOSE: To evaluate the effect of TGF-beta and the effect of apocynin on TGF-beta1 expression in skeletal muscle cells. STUDY DESIGN: Controlled laboratory study. In vitro assays were performed with C2C12 cells incubated with TGF-beta1 in presence or absence of apocynin (NOX inhibitor), SB525334 (TGF-beta-receptor I inhibitor), or chelerythrine (PKC inhibitor). METHODS: TGF-beta1 and atrogin-1 expression was evaluated by RT-qPCR and/or ELISA; Smad3 phosphorylation by western blot; Smad4 nuclear translocation by indirect immunofluorescence; and ROS levels by DCF probe fluorescent measurements. RESULTS: We show that myoblasts respond to TGF-beta1 by increasing its own gene expression in a time- and dose-dependent fashion which was abolished by SB525334 and siRNA for Smad2/3. TGF-beta1 also induced ROS. Remarkably, apocynin inhibited the TGF-beta1 induced ROS as well as the autoinduction of TGF-beta1 gene expression. We also show that TGF-beta-induced ROS production and TGF-beta1 expression require PKC activity as indicated by the inhibition using chelerythrine. CONCLUSION: These results strongly suggest that TGF-beta induces its own expression through a TGF-beta-receptor/Smad-dependent mechanism and apocynin is able to inhibit this process, suggesting that requires NOX-induced ROS in skeletal muscle cells. CI - Copyright (c) 2015 Elsevier GmbH. All rights reserved. FAU - Abrigo, Johanna AU - Abrigo J AD - Laboratorio de Biologia y Fisiopatologia Molecular, Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas and Facultad de Medicina, Universidad Andres Bello, Santiago, Chile; Millennium Institute on Immunology and Immunotherapy, Santiago, Chile. FAU - Morales, Maria Gabriela AU - Morales MG AD - Laboratorio de Biologia y Fisiopatologia Molecular, Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas and Facultad de Medicina, Universidad Andres Bello, Santiago, Chile; Millennium Institute on Immunology and Immunotherapy, Santiago, Chile. FAU - Simon, Felipe AU - Simon F AD - Laboratorio de Fisiopatologia Integrativa, Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas and Facultad de Medicina, Universidad Andres Bello, Santiago, Chile; Millennium Institute on Immunology and Immunotherapy, Santiago, Chile. FAU - Cabrera, Daniel AU - Cabrera D AD - Departamento de Ciencias Quimicas y Biologicas, Universidad Bernardo O Higgins, Santiago, Chile. FAU - Di Capua, Gabriella AU - Di Capua G AD - Laboratorio de Biologia y Fisiopatologia Molecular, Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas and Facultad de Medicina, Universidad Andres Bello, Santiago, Chile; Millennium Institute on Immunology and Immunotherapy, Santiago, Chile. FAU - Cabello-Verrugio, Claudio AU - Cabello-Verrugio C AD - Laboratorio de Biologia y Fisiopatologia Molecular, Departamento de Ciencias Biologicas, Facultad de Ciencias Biologicas and Facultad de Medicina, Universidad Andres Bello, Santiago, Chile; Millennium Institute on Immunology and Immunotherapy, Santiago, Chile. Electronic address: claudio.cabello.verrugio@gmail.com. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20150708 PL - Germany TA - Phytomedicine JT - Phytomedicine : international journal of phytotherapy and phytopharmacology JID - 9438794 RN - 0 (6-(2-tert-butyl-5-(6-methylpyridin-2-yl)-1H-imidazol-4-yl)quinoxaline) RN - 0 (Acetophenones) RN - 0 (Benzophenanthridines) RN - 0 (Imidazoles) RN - 0 (Quinoxalines) RN - 0 (Reactive Oxygen Species) RN - 0 (Smad2 Protein) RN - 0 (Smad2 protein, mouse) RN - 0 (Smad3 Protein) RN - 0 (Smad3 protein, mouse) RN - 0 (Tgfb1 protein, mouse) RN - 0 (Transforming Growth Factor beta) RN - 0 (Transforming Growth Factor beta1) RN - B6J7B9UDTR (acetovanillone) RN - E3B045W6X0 (chelerythrine) RN - EC 2.7.11.13 (Protein Kinase C) SB - IM MH - Acetophenones/*pharmacology MH - Animals MH - Benzophenanthridines/pharmacology MH - Cell Line MH - Imidazoles/pharmacology MH - Mice MH - Muscle Fibers, Skeletal/*drug effects/metabolism MH - Protein Kinase C/metabolism MH - Quinoxalines/pharmacology MH - Reactive Oxygen Species/*metabolism MH - Smad2 Protein/metabolism MH - Smad3 Protein/metabolism MH - Transforming Growth Factor beta/*pharmacology MH - Transforming Growth Factor beta1/*metabolism MH - Up-Regulation OTO - NOTNLM OT - Atrophy OT - NOX OT - ROS OT - Skeletal muscle OT - Smad OT - TGF-beta1 EDAT- 2015/09/01 06:00 MHDA- 2015/12/29 06:00 CRDT- 2015/09/01 06:00 PHST- 2015/03/02 00:00 [received] PHST- 2015/06/18 00:00 [revised] PHST- 2015/06/21 00:00 [accepted] PHST- 2015/09/01 06:00 [entrez] PHST- 2015/09/01 06:00 [pubmed] PHST- 2015/12/29 06:00 [medline] AID - S0944-7113(15)00210-X [pii] AID - 10.1016/j.phymed.2015.06.011 [doi] PST - ppublish SO - Phytomedicine. 2015 Sep 15;22(10):885-93. doi: 10.1016/j.phymed.2015.06.011. Epub 2015 Jul 8.