Hypoxia induces HMGB1 secretion through exosomes in pancreatic cancer cells
WANG Ming1, SONG Lian2, ZHANG Lirong2, GONG Aihua3, ZHU Haitao2, WANG Dongqing2
（1. School of Medicine, Jiangsu University, Zhenjiang Jiangsu 212013; 2. Department of Radiology, Affiliated Hospital of Jiangsu University, Zhenjiang Jiangsu 212001, China)
Abstract：Objective: To study the secretory pathway of high mobility group box1 protein(HMGB1) in hypoxiainduced human pancreatic cancer cells.Methods: The expression of HMGB1 in pancreatic cancer was analyzed by GEO and TCGA database. Western blotting was used to detect the content of HMGB1 in the supernatant of pancreatic cancer PaTu8988 cells under different hypoxic time points(0, 6, 12, 24, 48 h); the exosomes in pancreatic cancer PaTu8988 cells culture medium under normal and hypoxic conditions were separated by exosome extraction kit. The exosomes were identified by transmission electron microscopy and Western blotting, and HMGB1 in exosomes was detected by Western blotting. Immunofluorescence staining was used to observe the distribution of HMGB1 in cells by confocal microscope under normoxia and hypoxia.Results: The results of GEO and TCGA database showed that HMGB1 was highly expressed in pancreatic cancer tissues compared with normal pancreatic tissue (P<0.01), and the survival time of highexpression patients was significantly shorter than that of lowexpression patients(P<0.05). After hypoxic treatment for 48 h, the expression of HMGB1 in the cell culture supernatant was the highest, significantly higher than that of 6,12,24 h. Transmission electron microscopy showed that the culture had microvesicle structure with a diameter of about 100 nm. Western blotting results showed the expression of exosome membrane markers CD9, CD63, CD81, HSP70 and the expression of HMGB1 in exosomes. Immunofluorescence results showed that HMGB1 and exosomes were colocalized in hypoxia. Conclusion: HMGB1 could be released through the exosomal pathway in PaTu8988 pancreatic cancer cells under hypoxia.
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2019, Vol. 29 
