Abstract: Background and purpose: Pancreatic cancer is a malignant tumor which occurs primarily in the digestive system. The incidence rate is higher in men than in women, and its prognosis is poor. Recombinant human pyruvate kinase isozymes R/L (PKLR) belong to the pyruvate kinase family. There are four kinds of mammalian pyruvate kinase isozymes: L, R, M1 and M2, which are closely related to the occurrence and development of tumor. Studies have shown that PKLR promotes breast cancer metastasis. This study aimed to investigate the clinicopathological significance of PKLR protein in pancreatic cancer, and to analyze the effect of PKLR on the biological behavior of pancreatic cancer and its molecular mechanism. Methods: Immunofluorescence staining was used to detect the localization of PKLR protein in pancreatic cancer cells. The negative expression rate, positive expression rate and strong positive expression rate of PKLR in pancreatic cancer tissues were detected by immunohistochemical staining EnVision method, and the correlation between PKLR and clinicopathological features of pancreatic cancer was analyzed. Small interfering RNA transfection technology was used to knockdown PKLR expression and observe its protein expression in pancreatic cancer cells. Methylthiazolyldiphenyl-tetrazolium bromide assay was used to detect the changes of cell proliferation activity after knockdown of PKLR. Plate cloning assay was used to detect the changes of colony formation of BXPC-3 and Mia PaCa-2 cells after PKLR knockdown. The scratch healing test was used to detect the effect of PKLR on the lateral migration of pancreatic cancer cells. Transwell assay was used to detect the effect of PKLR on the vertical migration of pancreatic cancer cells. The effect of PKLR on the expression of epithelial-mesenchymal transformation (EMT) related marker proteins was detected by Western blot. Results: Immunofluorescence results showed that PKLR fluorescence was localized in the cytoplasm of BxPC-3 and MIA PaCa-2 pancreatic cancer cells. Immunohistochemical staining showed that the positive expression rate of PKLR in pancreatic cancer tissues (75.2%) and strong positive expression rate (48.6%) were significantly higher than those in normal pancreatic tissues (14.3% and 7.1%) (P<0.01). The expression was closely related to histological grading and lymph node metastasis of pancreatic cancer (P<0.05). After knockdown of PKLR expression by small interfering RNA transfection, the protein expression level of PKLR in pancreatic cancer cells was significantly downregulated. MTT results showed that cell proliferation was significantly inhibited in a time-dependent manner (P<0.05) after knockdown of PKLR. Plate cloning experiment showed that knockdown of PKLR could significantly reduce the number of colony formation of BXPC-3 and Mia PaCa-2 cells; Scratch healing test and transwell test showed that the migration ability of pancreatic cancer cells was significantly inhibited after knockdown of PKLR. The effect of PKLR on EMT markers was detected. Downregulation of PKLR expression could significantly inhibit the proliferation and migration of pancreatic cancer cells, upregulate the expression of E-cadherin, and downregulate the expression of vimentin and Snail (P<0.05). Conclusion: PKLR plays an important role in the development and progression of pancreatic cancer. We will do further research.

Key words: Pancreatic cancer, PKLR, Proliferation, Migration, Epithelial-mesenchymal transformation