Study On The Inhibitory Effect Of PAEP Secreted By Melanoma Cells On T Lymphocytes In Vitro

Malignant melanoma is one of the most malignant tumors in the skin melanocyte system. Despite of the rapid progress in biological sciences, there is no fundamental development with melanoma clinical treatment, resulting to up to80%of skin cancer deaths. Therefore, in-depth analysis of the pathogenesis of melanoma is very important to explore effective treatment options. Our previous studies indicated that the oncogenesis and development of melanoma are closely related to the expression level of progesterone endometrial protein’s expression (progestagen-associated endometrial protein, PAEP). However, the mechanism of PAEP is unclear. This study was to analyze and explore the immune mechanism of PAEP in promoting tumor development.Progestagen-associated endometrial protein (PAEP), also known as glycodelin, is a secreted glycoprotein which was first isolated from human placenta, amniotic fluid, pregnancy deciduas and seminal plasma. Many studies have shown that it can suppress the immune activity of T cells and play an important regulatory role in the establishment of pregnancy maternal immune tolerance and HLA haploidentical embryo implantation.Our previous studies found that PAEP gene highly expressed in melanoma cell lines and promoted tumor cell migration and invasion.Considering the similarity of tumor invasion and embryo implantation, we hypothesize that PAEP gene inhibits T cells’ functions in the tumor microenvironment.In order to prove this hypothesis, we studied from the following three aspects:(1) Detect the expression of PAEP gene in melanoma cell lines by RT-PCR and Western blot;(2) Transiently Down-regurate the expression of PAEP gene in melanoma cells by chemically synthesized PAEP siRNA and establish stable melanoma cells with PAEP gene knockdown by PAEP shRNA Ientivirus transfection;(3) Isolate human peripheral blood-mononuclear cells and purify CD8+T cells from peripheral blood, mix with PAEP protein or tumor cells with silencing PAEP gene, and observe the changes such as lymphocyte proliferation, apoptosis and anti-tumor functions comparing with the control group. Our results showed that:(1) We confirmed high expression of PAEP gene in the mRNA and protein levels in melanoma cell lines;(2) Compared with non-targeting siRNA, PAEP siRNA significantly reduced PAEP mRNA expression; we obtained stable cells with PAEP gene silencing and non-targeting shRNA control cells;(3) After cell supernatant which contains PAEP was added into activated lymphocytes suspensions, the proliferation of lymphocytes was significantly inhibited and the apoptosis of lymphocytes increased;(4) PAEP protein inhibited the activity of cytotoxic T lymphocyte. In a word, our study in vitro suggested that PAEP protein inhibited lymphocyte’s proliferation and cytotoxicity. The inhibition of proliferation might be caused by the pro-apoptotic action of PAEP protein.