Cholera toxin (CTX) is produced by virulent stain of Vibrio cholerae. CTX binding to the alpha-subunit of G-proteins causes its ADP-ribosylation and thus constitutive activation of the Gs/adenylyl cyclase pathway leading to increased intracellular cyclic AMP levels. cAMP can modulate the expression of various genes including interleukin-1 (IL-1). This study was undertaken to examine the CTX-mediated cross regulation between the regulation of G-proteins and IL-1. Isolated pieces of intestine (40 mg) from neonatal mice were kept in RPMI medium supplemented with 10% fetal calf serum and treated with CTX (1 mu g/ml) for various time periods until 4 h The G protein and IL-1 alpha levels were measured by western blots and ELISA respectively. The results show that CTX caused marked reduction in the Gs alpha protein with out any effect on Gi or Go proteins. CTX treatment also caused 2-3 fold increase in the production of IL-1 alpha levels compared to control untreated tissues and the effects were mimicked by treatment with forskolin (a direct activator of adenylyl cyclase) and dibutyryl-cAMP (a cAMP analogue) indicating that CTX-mediated effects on IL-1 alpha were cAMP-dependent. Pretreatment with the nitric oxide (NO) synthase inhibitor, L-NAME (10 mu M), decreased the levels of IL-1 alpha (60%) compared to control. Similarly pretreatment of tissues with cyclooxygenase inhibitor (indomethacin; 10 mu M) blocked an increase in IL-1 alpha levels. It seems that the CTX-mediated regulation of IL-1 alpha is cAMP-dependent and involves multiple signalling pathways, i.e., the activation of the NO and COX pathways in mouse intestines.
ZAFAR NAWAZ ,BUKHTIAR H SHAH ,
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