title: Airway smooth muscle CXCR3 ligand production: regulation by JAK-STAT1 and intracellular Ca²⁺.
abstract: In asthma, airway smooth muscle (ASM) chemokine (C-X-C motif) receptor 3 (CXCR3) ligand production may attract mast cells or T lymphocytes to the ASM, where they can modulate ASM functions. In ASM cells (ASMCs) from people with or without asthma, we aimed to investigate JAK-STAT1, JNK, and Ca²⁺ involvement in chemokine (C-X-C motif) ligand (CXCL)10 and CXCL11 production stimulated by interferon-γ, IL-1β, and TNF-α combined (cytomix). Confluent, growth-arrested ASMC were treated with inhibitors for pan-JAK (pyridone-6), JAK2 (AG-490), JNK (SP-600125), or the sarco(endo)plasmic reticulum Ca²⁺ATPase (SERCA) pump (thapsigargin), Ca²⁺ chelator (BAPTA-AM), or vehicle before and during cytomix stimulation for up to 24 h. Signaling protein activation as well as CXCL10/CXCL11 mRNA and protein production were examined using immunoblot analysis, real-time PCR, and ELISA, respectively. Cytomix-induced STAT1 activation was lower and CXCR3 ligand mRNA production was more sensitive to pyridone-6 and AG-490 in asthmatic than nonasthmatic ASMCs, but CXCL10/CXCL11 release was inhibited by the same proportion. Neither agent caused additional inhibition of release when used in combination with the JNK inhibitor SP-600125. Conversely, p65 NF-κB activation was higher in asthmatic than nonasthmatic ASMCs. BAPTA-AM abolished early CXCL10/CXCL11 mRNA production, whereas thapsigargin reduced it in asthmatic cells and inhibited CXCL10/CXCL11 release by both ASMC types. Despite these inhibitory effects, neither Ca²⁺ agent affected early activation of STAT1, JNK, or p65 NF-κB. In conclusion, intracellular Ca²⁺ regulated CXCL10/CXCL11 production but not early activation of the signaling molecules involved. In asthma, reduced ASM STAT1-JNK activation, increased NF-κB activation, and altered Ca²⁺ handling may contribute to rapid CXCR3 ligand production and enhanced inflammatory cell recruitment.
authors.affiliation: Faculty of Pharmacy, The University of Sydney, Camperdown, New South Wales, Australia. xiahui.tan@umassmed.edu
authors.forename: X
authors.initials: X
authors.lastname: Tan
authors.forename: Y A
authors.initials: YA
authors.lastname: Alrashdan
authors.forename: H
authors.initials: H
authors.lastname: Alkhouri
authors.forename: B G G
authors.initials: BG
authors.lastname: Oliver
authors.forename: C L
authors.initials: CL
authors.lastname: Armour
authors.forename: J M
authors.initials: JM
authors.lastname: Hughes
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001249:Asthma / Q000378:metabolism; D002118:Calcium / Q000502:physiology*; D054357:Chemokine CXCL10 / Q000096:biosynthesis*; D054371:Chemokine CXCL11 / Q000096:biosynthesis*; D019743:Chemokines, CXC / Q000378:metabolism; D005260:Female; D006801:Humans; D007371:Interferon-gamma; D053583:Interleukin-1beta / Q000494:pharmacology; D008168:Lung / Q000378:metabolism*; D008171:Lung Diseases / Q000378:metabolism; D008175:Lung Neoplasms / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D054367:Receptors, CXCR3 / Q000378:metabolism; D050794:STAT1 Transcription Factor; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: C114753:CXCL10 protein, human; C515202:CXCL11 protein, human; C515187:CXCR3 protein, human; D054357:Chemokine CXCL10; D054371:Chemokine CXCL11; D019743:Chemokines, CXC; D053583:Interleukin-1beta; D016328:NF-kappa B; D054367:Receptors, CXCR3; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D002118:Calcium
keywords: asthma; calcium signaling; chemokine (C-X-C motif) ligand 10; chemokine (C-X-C motif) ligand 11; signal transducer and activator of transcription 1