Analysis of the mechanism of receptor tyrosine kinase activation in monocytes

Involvement of JNK pathway in GnRH I and II induced MMP 26 production The mitogen activated protein kinase signaling cascade has been shown to mediate the action of GnRH in human trophoblast cells. To address the role of the MAPK cascade in regulating MMP 26 expression by GnRH I and GnRH II, we examined the phosphorylation [You must be registered and logged in to see this link.] of JNK and ERK12 in B6Tert 1 cells treated with GnRH I or GnRH II for 0, 5, 10 and 30 minutes. As shown in Figure 4A, addition of 100 nM of GnRH I or GnRH II resulted in a transient activation of JNK at 5 min or 10 min, respectively. However, the addition of both GnRH I and GnRH II had no effects on ERK12 activity in B6Tert 1 cells. To further clarify the involvement of JNK and ERK12 in GnRH induced MMP 26 expression, B6Tert 1 cells were cultured with GnRH I or GnRH II in the absence or presence of the JNK inhibitor or ERK12 inhibitor.

SP600125, but not PD98059, significantly [You must be registered and logged in to see this link.] blocked MMP 26 induction. These data indicate that stimulation of MMP 26 expression by GnRH I and II requires the activation of JNK, but not ERK12 signaling molecules. Regulatory effects of GnRH I and GnRH II on MMP 26 expression in primary cultured CTBs We further confirmed the regulatory effects of GnRH I and GnRH II on MMP 26 expression using the primary isolates of first trimester CTB cells. The cells were incu bated with GnRH I and GnRH II at concentrations of 0. 1 100 nM for 24 h. A significant increase in MMP 26 mRNA levels was only observed in trophoblast cultured in the presence of higher concentrations of these two hormones.

GnRH II at a con centration of 100 nM induced the maximal increase in MMP 26 expression. To further determine [You must be registered and logged in to see this link.] the signaling mediators involved in GnRH induced MMP 26 expression, CTB cells were cultured with 100 nM GnRH I and II in the presence of 10 uM SP600125 or PD98059. As shown in Figure 5B, SP600125 inhibited the stimulatory effects of GnRH I and GnRH II on MMP 26 mRNA levels, whereas PD98059 had no effects on MMP 26 expression. Discussion MMP 26 is a recently discovered and only partially characterized human proteinase. Unlike all other MMPs, the Mmp 26 gene does not exist in the murine genome. Besides its extensive distribution in can cer cells of epithelial origin, MMP 26 is also restricted in human placenta and uterus, but nor in other normal tissues.

Previous studies showed its intensive expression in human trophoblasts in vivo and invasive promoting effect on trophoblast cells in vitro, suggesting the unique role of MMP 26 in tropho blast invasion at the feto maternal interface. However, the regulation of MMP 26 expression in trophoblast cells remains to be determined. In this study, we utilized human trophoblast like cell line, B6Tert 1, to show that 1 both GnRH I and GnRH II could up regulate MMP 26 expression, 2 GnRH I and GnRH II had differential effects on MMP 26 expression, and 3 GnRH I and II induced MMP 26 expression was mediated by JNK, but not ERK12 signaling pathway. The B6Tert 1 cell line is an immortalized cytotropho blast like cell line developed from normal human pla cental villi during the first trimester. These cells are transfected with human telomerase catalytic subunit gene and thus maintain reconstituted telomerase activity.