Abstract:

Objective To discuss the effects of kallikrein 5 （KLK5） overexpression on the proliferation， invasion and cisplatin （DDP） sensitivity of cervical cancer cells， and to clarify its mechanism. Methods Western blotting method was used to verify the stable transfection and overexpression of KLK5 in the cervical cancer cell （ME180-OE-KLK5）. The cervical cancer ME180-NC-KLK5 and ME180-OE-KLK5 cells in logarithmic growth phase were subcutaneously inoculated into the nude mice to establish the subcutaneous xenograft models. After successful modeling， the mice were randomly divided into normal saline control group （NC-KLK5+0.9% NaCl group）， DDP treatment group （NC-KLK5+DDP group）， KLK5 overexpression group（OE-KLK5+0.9% NaCl group） and KLK5 overexpression combined with DDP group（OE-KLK5+DDP group）， with 5 mice in each group. The nude mice in NC-KLK5+DDP group and OE-KLK5+DDP group were given intraperitoneal injection of DDP at a dose of 5 mg·kg?1； the nude mice in NC-KLK5+0.9% NaCl group and OE-KLK5+0.9% NaCl group were given intraperitoneal injection of normal saline at a dose of 0.01 mL·g?1. The body weights of nude mice were measured every 2 d， and the long diameter and short diameter of the tumors were recorded to calculate the tumor volume and plot the tumor growth curve. At 24 h after the last administration on day 14， the nude mice were sacrificed， and the tumors were dissected and weighed. HE staining method was used to observe the pathomorphology of tumor tissue in the nude mice in various groups； immunohistochemistry staining method was used to observe the expression levels of KLK5， Ki67 and matrix metalloproteinase-9 （MMP-9） proteins in the tumor tissues of the nude mice in various groups. Results Compared with ME180-NC-KLK5 cells， the expression level of KLK5 protein in ME180-OE-KLK5 cells was increased （P<0.05）. In the first week after subcutaneous xenograft inoculation， the nude mice in various groups showed good feeding and activity status， and their body weights gradually increased. The drug administration phase started from the second week. During the drug treatment period， the feeding and activity status as well as body weight of the nude mice in NC-KLK5+0.9%NaCl group showed no significant changes compared with the first week； compared with NC-KLK5+0.9%NaCl group， the nude mice in NC-KLK5+DDP group began to show loss of appetite， no increase in body weight， and decreased activity. During the drug treatment period in the third week， the feeding and activity status of the nude mice in NC-KLK5+0.9%NaCl group showed no significant changes compared with the second week， while they began to show no increase in body weight； compared with NC-KLK5+0.9%NaCl group， the feeding and activity status of the nude mice in NC-KLK5+DDP group were significantly weakened， and their body weights decreased. Compared with NC-KLK5+0.9%NaCl group， the volume of xenograft tumor in NC-KLK5+DDP group was decreased （P<0.01）； compared with NC-KLK5+DDP group， the volume of xenograft tumor OE-KLK5+DDP group was significantly increased （P<0.001）； compared with NC-KLK5+0.9%NaCl group， the volume of xenograft tumor of the nude mice in OE-KLK5+0.9%NaCl group was increased （P<0.001）； compared with OE-KLK5+0.9%NaCl group， the volume of xenograft tumors in the nude mice in OE-KLK5+DDP group showed no statistically significant difference （P>0.05）. Compared with NC-KLK5+0.9%NaCl group， the weight of xenograft tumor of the nude mice in NC-KLK5+DDP group was decreased （P<0.05）； compared with NC-KLK5+DDP group， the weight of xenograft tumors of the nude mice in OE-KLK5+DDP group was significantly increased （P<0.001）； compared with NC-KLK5+0.9%NaCl group， the weight of xenograft tumor of the nude mice in OE-KLK5+0.9%NaCl group was increased （P<0.001）； compared with OE-KLK5+0.9%NaCl group， the weight of xenograft tumors of the nude mice in OE-KLK5+DDP group showed no statistically significant difference （P>0.05）. Compared with NC-KLK5+0.9%NaCl group， the xenograft tumor cells of the nude mice in OE-KLK5+0.9%NaCl group showed greater nuclear heterogeneity； the xenograft tumor cells of the nude mice in OE-KLK5+DDP group and NC-KLK5+DDP group showed cytomorphological changes， manifested as nuclear pyknosis and fragmentation， reduced cell volume， and the appearance of necrosis and apoptosis. Compared with NC-KLK5+DDP group， the degree of necrosis in xenograft tumor of the nude mice in OE-KLK5+DDP group was more pronounced. Compared with NC-KLK5+0.9%NaCl group， the expression levels of KLK5， Ki67 and MMP-9 proteins in xenograft tumor tissue of the nude mice in NC-KLK5+DDP group were decreased （P<0.05）； compared with NC-KLK5+DDP group， the expression levels of KLK5， Ki67， and MMP-9 proteins in xenograft tumor tissue of the nude mice in OE-KLK5+DDP group were increased （P<0.001）； compared with NC-KLK5+0.9%NaCl group， the expression levels of KLK5， Ki67 and MMP-9 proteins in xenograft tumor tissue of the nude mice in OE-KLK5+0.9%NaCl group were increased （P<0.001）； compared with OE-KLK5+0.9%NaCl group， the expression levels of KLK5， Ki67 and MMP-9 in xenograft tumor tissue of the nude mice in OE-KLK5+DDP group showed no statistically significant differences （P>0.05）. Conclusion KLK5 overexpression can promote the growth of subcutaneous xenograft tumors of cervical cancer ME180 cells treated with DDP， up-regulate the expressions of Ki67 and MMP-9 in the xenograft tumor tissue， and reduce the sensitivity of the xenograft tumor to DDP.

Key words: Kallikrein 5, Uterine cervical neoplasm, Cisplatin, Matrix metalloproteinase-9, ME180 cells, Xenograft tumor