The Study Of Correlation And Mechanism Between Kai1 And Autophagy In Pancreatic Cancer

KAI1, as a metastasis-suppressor gene, correlates with metastasis of pancreatic cancer closely. Our previous findings revealed that KAI1 could inhibited human pancreatic cancer metastasis and proliferation in vitro. Furthermore, MIA PaCa-2 cells of overexpressing KAI1/CD82 subcutaneous injection into nude mice reduced metastases significantly without affecting primary tumor growth in vivo. However, the mechanism remains unclear. One of the main difference between in vivo and in vitro is the hypoxiain conditons in the microenvironment. Recent studies have shown that hypoxic conditions of solid tumors induce high-level autophagy and KAI1 expression. The hypoxia in is known to increase the malignant potential of pancreatic cancers. Because autonomously proliferating cancer cells are often exposed to hypoxic conditions, there must be an alternative metabolic pathway, such as autophagy, that allows them to obtain energy when both oxygen and glucose are depleted. Studies have also shown that strong autophagy in the peripheral area of pancreatic cancer tissue was correlated with a poor outcome and a short disease-free period. Activated autophagy was thought to be a response to factors associated with the cancer microenvironment, such as hypoxia and a poor nutrient supply . Despite recent advances in the understanding of the molecular mechanisms and biological function of both autophagy and KAI1, the relationship between autophagy and KAI1 has not been reported. We hypothesized that KAI1 may influence the autophagy of cancer cells, which may further change the effect of KAI1 on cell survival and studied this area. The study offered an opportunity to partly explain why KAI1 has no influence on primary tumorigenicity and raise the potential of use of KAI1 for cancer therapy.【Objective】To study the relationship and mechanism between autophagy and KAI1 in human pancreatic cancer cell line MIA PaCa-2. To investigate the effect and mechanism of Ad5-KAI1-induced autophagy on proliferation and apoptosis in human pancreatic cancer cell line MIA PaCa-2.【Methods】Human pancreatic cancer cells of MIA PaCa-2 do not express the protein of KAI1. After infection of Ad5-KAI1, the cells could express the protein of KAI1. The formation of autophagosomes was observed by TEM . The expression level of Beclin 1, AKT, ERK, the phosphorylation of AKT and ERK protein and the ratio of LC3-II to LC3-I was detected by western blot. The green fluorescent protein-labeled light chain 3 association with autophagosome membranes was detected by confocal microscopy. The change of autophagy in human pancreatic cancer cell line MIA PaCa-2 infected by Ad5-KAI1 was observed to judge the relationship between KAI1 and autophagy. Pretreatment of LY294002 and PD98059 were used to inhibit the the phosphorylation of AKT and ERK. Then we observed whether the relationship between KAI1 and autophagy could be inhibited too. The proliferation of cells was observed by CCK8. The apoptosis of cells was observed by AnnexinV-FITC/PI,PARP cleavage and caspase-3 activation. Pretreatment of 3-MA were used to inhibit the the autophagy. Then, we observed whether the effect of KAI1 on survival of cells could be changed. Pretreatment of 3-MA was used to inhibit the autophagy before inhibit the activation of caspase-3 and the phosphorylation of ERK. Then, we observed whether the effect of autophagy on apoptosis of cell could be inhibited too.【Conclusions】KAI1 can induce autophagy in human pancreatic cell line MIA PaCa-2 through phosphorylation of extracellular signal-related kinases rather than that of AKT. KAI1-induced autophagy promoted MIA PaCa-2 cells proliferation and protected the MIA PaCa-2 cells from apoptosis. KAI1-induced autophagy protected the MIA PaCa-2 cells from apoptosis through the downregulation of PARP cleavage and caspase-3 activation but not through the downregulation of ERK phosphorylation.