|Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1993: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1992: ¥1,000,000 (Direct Cost: ¥1,000,000)
In this study, we tried to elucidate signal transduction pathway in primitive hematopoietic progenitors of mice stimulated with interleukin-11(IL-11). First, effects of calphostin C, a specific inhibitor for protein Kinase C(PKC), and genistein and herbimycin A, specific tyrosine kinase (TK) inhibitors on colony formation of 5-FU resistant, dormant, primitive hematopoietic progenitors supported by IL-6 which receptor has the same signal transducer gp 130 of IL-11 receptor and 12-o-tetradecanoyl phorbol-13-acetate(TPA). Although IL-6 and TPA alone did not support colonies, IL-6 in combination with TPA gave rise to significant number of colonies which suppressed by calphostin C or herbimycin A, indicating the signal of IL-6 + TPA was transduced through an avtivation of PKC and TK.The concentration of calphostin C at which inhibited the effect of TPA was 100-200nM, which was 2-3 times higher concentration of 50% inhibition constant(IC50) for PKC.On the other hand, the concentration of herbimycin A which suppress the effect of IL-l was 100-200ng/ml. Chromic exposure of bone marrow cells to TPA ( down regulation of PKC ) resulted a suppression of colony formation with IL-6 + TPA.
Next, we studied the effect of IL-11 on IL-3-dependent colony formation and its inhibition by these protein kinase inhibitors. IL-11 enhanced IL-3-dependent colony formation by 2-3 times. Calphostin C at concentration of 200nM suppressed colony formation supported IL-3 alone or in combination with IL-11. Preincubation of bone marrow cells with IL-11 in combination with calphostin C or herbimycin A canceled the augmentation of IL-3-dependent colony formation by IL-11. These findings suggested that signal of IL-11 in colony formation of primitive hematopoietic progenitors was transduced through an activation of PKC and TK.Confirmation of these findings by immunocytochemical examination on activation of PKC and TK by IL-11 is on going.