title: Role of transient receptor potential vanilloid 2 in LPS-induced cytokine production in macrophages.
abstract: There is considerable evidence indicating that intracellular Ca2+ participates as a second messenger in TLR4-dependent signaling. However, how intracellular free Ca2+ concentrations ([Ca2+]i) is increased in response to LPS and how they affect cytokine production are poorly understood. Here we examined the role of transient receptor potential (TRP), a major Ca2+ permeation pathway in non-excitable cells, in the LPS-induced cytokine production in macrophages. Pharmacologic experiments suggested that TRPV family members, but neither TRPC nor TRPM family members, are involved in the LPS-induced TNFalpha and IL-6 production in RAW264 macrophages. RT-PCR and immunoblot analyses showed that TRPV2 is the sole member of TRPV family expressed in macrophages. ShRNA against TRPV2 inhibited the LPS-induced TNFalpha and IL-6 production as well as IkappaBalpha degradation. Experiments using BAPTA/AM and EGTA, and Ca2+ imaging suggested that the LPS-induced increase in [Ca2+]i involves both the TRPV2-mediated intracellular and extracellular Ca2+ mobilizations. BAPTA/AM abolished LPS-induced TNFalpha and IL-6 production, while EGTA only partially suppressed LPS-induced IL-6 production, but not TNFalpha production. These data indicate that TRPV2 is involved in the LPS-induced Ca2+ mobilization from intracellular Ca2+ store and extracellular Ca2+. In addition to Ca2+ mobilization through the IP3-receptor, TRPV2-mediated intracellular Ca2+ mobilization is involved in NFkappaB-dependent TNFalpha and IL-6 expression, while extracellular Ca2+ entry is involved in NFkappaB-independent IL-6 production.
authors.affiliation: Division of Diabetes and Endocrinology, Department of Internal Medicine, Jikei University School of Medicine, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Yamashiro
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Sasano
authors.forename: Katsuyoshi
authors.initials: K
authors.lastname: Tojo
authors.forename: Iyuki
authors.initials: I
authors.lastname: Namekata
authors.forename: Junko
authors.initials: J
authors.lastname: Kurokawa
authors.forename: Naoki
authors.initials: N
authors.lastname: Sawada
authors.forename: Takayoshi
authors.initials: T
authors.lastname: Suganami
authors.forename: Yasutomi
authors.initials: Y
authors.lastname: Kamei
authors.forename: Hikaru
authors.initials: H
authors.lastname: Tanaka
authors.forename: Naoko
authors.initials: N
authors.lastname: Tajima
authors.forename: Kazunori
authors.initials: K
authors.lastname: Utsunomiya
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Ogawa
authors.forename: Tetsushi
authors.initials: T
authors.lastname: Furukawa
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D016207:Cytokines / Q000096:biosynthesis*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D020672:I-kappa B Proteins / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D000072000:NF-KappaB Inhibitor alpha; D050916:TRPV Cation Channels / Q000235:genetics / Q000502:physiology*
chemical_list: D015220:Calcium Channels; D016207:Cytokines; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; C000606205:Nfkbia protein, mouse; D050916:TRPV Cation Channels; C485717:Trpv2 protein, mouse; D000072000:NF-KappaB Inhibitor alpha; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D002118:Calcium