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Pleural effusion is one of the complications of human non-small cell lung cancer (NSCLC). High mobility group box-1 protein (HMGB1) correlates highly with invasion and metastasis in multiple tumors. The aim of this study was to explore the clinical value of HMGB1 in NSCLC patients, and to investigate the role of HMGB1 in the development of pleural effusion. In addition, we also investigated the regulatory role of HMGB1 in the sensitivity of NSCLC cells to cisplatin. 46 NSCLC malignant pleural effusion (MPE) and 31 benign pleural effusion samples were quantitatively analyzed with Enzyme-Linked Immunosorbent Assay (ELISA) for cytokines, such as IL-1beta, IL-6, IL-8 and HMGB1. The HMGB1 expression in NSCLC tissues was examined with RT-qPCR and western blotting methods. Then the influence by HMGB1 on the chemosensitivity of lung cancer A549 cells was examined with MTT assay and colony forming assay for the A549 cells post the treatment with cisplatin or (and) HMGB1. The results demonstrated that HMGB1 was up-regulated in the pleural effusion of NSCLC patients, along with the up-regulated levels of proinflammatory cytokines such as IL-6 and IL-8. And the up-regulation of HMGB1 was confirmed at both the mRNA and protein levels in the NSCLC tissues. Recombinant HMGB1 reduced the sensitivity of A549 cells to cisplatin in vitro. In conclusion, HMGB1 was up-regulated in the pleural effusion and tumor tissues of NSCLC patients. HMGB1 reduced the sensitivity of NSCLC A549 cells to cisplatin in vitro