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INFLAMMATION, IMMUNOPHARMACOLOGY, AND ASTHMA

Basiliolides, a Class of Tetracyclic C19 Dilactones from Thapsia garganica, Release Ca²⁺ from the Endoplasmic Reticulum and Regulate the Activity of the Transcription Factors Nuclear Factor of Activated T Cells, Nuclear Factor-B, and Activator Protein 1 in T Lymphocytes

Departamento de Biología Celular, Fisiología e Inmunología, Universidad de Córdoba, Córdoba, Spain (C.N., R.S., F.J.C., E.M.); Dipartimento di Scienze Chimiche, Alimentari, Farmaceutiche e Farmacologiche, Università del Piemonte Orientale, Novara, Italy (F.P., G.A.); Vivacell Biotechnology GmbH, Denzlingen, Germany (B.L.F.); and Department of Organic Chemistry, Lund University, Lund, Sweden (O.S.)

Calcium concentration within the endoplasmic reticulum (ER) plays an essential role in cell physiology. We have investigated the effects of basiliolides, a novel class of C19 dilactones isolated from Thapsia garganica, on Ca²⁺ mobilization in T cells. Basiliolide A1 induced a rapid mobilization of intracellular Ca²⁺ in the leukemia T-cell line Jurkat. First, a rapid calcium peak was observed and inhibited by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester. This initial calcium mobilization was followed by a sustained elevation, mediated by the entry of extracellular calcium through store-operated calcium release-activated Ca²⁺ (CRAC) channels and sensitive to inhibition by EGTA, and by the CRAC channel inhibitor N-{4-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]phenyl}-4-methyl-1,2,3-thiadiazole-5-carboxamide (BTP-2). Basiliolide A1 mobilized Ca²⁺ from ER stores, but in contrast to thapsigargin, it did not induce apoptosis. Basiliolide A1 induced nuclear factor of activated T cells 1 dephosphorylation and activation that was inhibited by BTP-2 and cyclosporine A. In addition, we found that basiliolide A1 alone did not mediate IB degradation or RelA phosphorylation (ser536), but it synergized with phorbol 12-myristate 13-acetate to induce a complete degradation of the nuclear factor-B inhibitory protein and to activate the c-Jun NH₂-terminal kinase. Moreover, basiliolide A1 regulated both interleukin-2 and tumor necrosis factor- gene expression at the transcriptional level. In basiliolide B, oxidation of one of the two geminal methyls to a carboxymethyl group retained most of the activity of basiliolide A1. In contrast, basiliolide C, where the 15-carbon is oxidized to an acetoxymethine, was much less active. These findings qualify these compounds as new probes to investigate intracellular calcium homeostasis.

Address correspondence to: Dr. Eduardo Muñoz, Departamento de Biología Celular, Fisiología e Inmunología, Facultad de Medicina, Avda. de Menéndez Pidal s/n, Universidad de Córdoba, 14004 Córdoba, Spain. E-mail: fi1muble{at}uco.es