Objective: To investigate the effects of signal pathway inhibitors PD98059 and LY294002 on cell proliferation, apoptosis, expressions of phosphorylated extracellular signal-regulared kinase (p-ERK) and phosphorylated protein kinase B (p-Akt) in endometrial carcinoma xenografts.
Methods: Human endometrial carcinoma Ishikawa cells were cultured in vitro. The effects of PD98059 and LY294002 on proliferation, apoptosis, and cell cycle distribution of endometrial cancer cells were detected by monotetrazolium (MTT) assay and fluorescence-activated cell sorting technique. The models of xenografted tumor were established by the subcutaneous inoculation in 24 nude mice, and then they were randomly divided into 4 groups (n = 6), normal saline group, PD98059 group (PD group), LY294002 group (LY group) or PD98059 + LY294002 group (PD + LY group) by intraperitoneal injections, respectively. The anti-tumor efficacy was evaluated by measuring tumor volume and tumor growth status. The histopathological change of tumor specimens was observed using HE staining and terminal deoxynucleotidyl transferase-mediated dUTP-digoxigen in nick and labeling method (TUNEL) testing and the expression levels of p-ERK and p-Akt were detected by immunohistochemistry method.
Results: (1) The proliferation of Ishikawa cells were suppressed after treated by PD98059 and(or) Y294002, in which A(570) values of cells decreased showing both time-dependent and concentration-dependent manner (LY294002: F(group) = 9.801, P = 0.002; F(time) = 10.398, P = 0.001. PD98059: F(group) = 8.213, P = 0.015; F(time) = 6.839, P = 0.036). Cell cycle distribution analysis revealed that percentage of Ishikawa cells at G(0)/G(1) phase (F(time) = 35.049, P = 0.004; F(group) = 32.024, P < 0.01) increased and percentage of S phase cells (F(time) = 7.789, P = 0.049; F(group) = 30.132, P < 0.01) decreased significantly. The percentage of apoptotic cells increased significantly among PD group, LY group and PD + LY group, in which there were significant difference [(63.3 ± 0.5)% vs (30.7 ± 20.1)% vs (40.8 ± 1.3)%; F = 621.059, P < 0.01]. (2) Compared with the control group, the increasing of transplanting tumor volume in the treated groups were obviously (F = 23.545, P < 0.01), and the inhibited rate of the tumor was higher in PD + LY group than that in PD group or LY group [(68 ± 9)% vs (32 ± 16)% or (38 ± 17)%; F = 10.283, P < 0.05]. (3) HE staining shown that there were different degrees of necrosis for endometrial carcinoma cell in different groups. The apoptosis of tumor cells were significantly increased in treated groups by TUNEL testing [(13.7 ± 1.5)%, (14.1 ± 1.2)%, (29.0 ± 1.8)%; F = 320.344, P < 0.01]. Immunohistochemistry results demonstrated that the expressions of p-ERK and p-Akt in treated groups were lower than that in control group, of which LY + PD group was the lowest one.
Conclusion: The signal pathway inhibitors PD98059 and LY294002 could inhibit the growth of human endometrial carcinoma in vivo and in vitro, in which may induce cell apoptosis.