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Objectives: Meloxicam, a selective cyclooxygenase-2 (COX-2) inhibitor, has been demonstrated to exert anti-tumour effects against various malignancies. However, up to now, mechanisms involved in meloxicam anti-hepatocellular carcinoma effects have remained unclear. Materials and methods: Cell viability and apoptosis were assessed by CCK-8 and flow cytometry. Endoplasmic reticulum (ER) stress and autophagyassociated molecules were analysed by western blotting and immunofluorescence assay. GRP78 and Atg5 knock-down by siRNA or chemical inhibition was used to investigate cytotoxic effects of meloxicam treatment on HCC cells. Results: We found that meloxicam led to apoptosis and autophagy in HepG2 and Bel-7402 cells via a mechanism that involved ER stress. Up-regulation of GRP78 signalling pathway from meloxicaminduced ER stress was critical for activation of autophagy. Furthermore, autophagy activation attenuated ER stress-related cell death. Blocking autophagy by 3-methyladenine (3-MA) or Atg5 siRNA knock-down enhanced meloxicam lethality for HCC by activation of ER stress-related apoptosis. In addition, GRP78 seemed to lead to autophagic activation via the AMPK-mTOR signalling pathway. Blocking AMPK with a chemical inhibitor inhibited autophagy suggesting that meloxicamregulated autophagy requires activation of AMPK. Conclusions: Our results revealed that both ER stress and autophagy were involved in cell death evoked by meloxicam in HCC cells. This inhibition of autophagy to enhance meloxicam lethality, suggests a novel therapeutic strategy against HCC. [ABSTRACT FROM AUTHOR]

Secretory clusterin (s CLU) is expressed in numerous cancers and is associated with the resistance to chemotherapy. However, the role of s CLU in the resistance of hepatocellular carcinoma ( HCC) to oxaliplatin ( OXA), a recently used third-generation platinum agent, remains unclear. The stable transfectants that are depleted of or overexpress s CLU and OXA-resistant cells were generated using human HCC cells. Overexpression of s CLU abrogated OXA-induced inhibition of cell growth and cell apoptosis, but depletion of s CLU synergized with OXA to inhibit cell growth and enhance cell apoptosis, by regulating proteins involved in mitochondrial apoptosis pathways, such as Bcl-2, Bax, Bcl-x L and caspase-9, and affecting phosphorylation of Akt and GSK-3β. Overexpression of s CLU in either OXA-resistant cells or stable transfectants that overexpress s CLU significantly increased phosphorylated Akt. However, specific inhibition of Akt enhanced sensitivity of s CLU-overexpressing cells to OXA, but had no effect on s CLU expression, suggesting that the regulatory effects between s CLU and p Akt may be in a one-way manner in HCC cells. The expression levels of s CLU affected the therapeutic efficacy of OXA to treat HCC tumors established in immunodeficiency mice. The results have demonstrated that s CLU contributes to OXA resistance by activating Akt pathway, indicating that s CLU may be a novel molecular target for overcoming OXA resistance in HCC. [ABSTRACT FROM AUTHOR]

Background and Aims: Cyclooxygenase-2 (COX-2) is associated with carcinogenesis. The aim of this study was to investigate the expression of COX-2 in four hepatocellular carcinoma (HCC) cell lines, and evaluate the effect of a selective COX-2 inhibitor, meloxicam, in HepG2, a high COX-2 expressing cell line. Methods: Expression of COX-2 was detected using RT-PCR, Western blotting and immunohistochemical analysis. Cell proliferation was measured using MTT assay. Cell cycle distribution was determined by flow cytometry. Apoptosis was detected with TUNEL method. Expression of proliferating cell nuclear antigen (PCNA), cell cycle regulatory proteins including cyclins A, B1, D1 and E, and apoptosis-related proteins including Fas, Fas ligand and Bcl-2 were examined using Western blotting. Results: Cyclooxygenase-2 was intensely expressed in HepG2, HLE and BEL7402 cells, but weakly expressed in SMMC-7402 cells. Meloxicam suppressed proliferation of HepG2 cells in a dose- and time-dependent manner, resulting in cell cycle arrest in S phase and cell accumulation in G0/G1 phase. Expression of PCNA, cyclin A but not cyclin B1, cyclin D1 or cyclin E was down-regulated by meloxicam. Meloxicam also induced apoptosis of HepG2 cells, with increased expression of Fas ligand, but the expression of Fas and Bcl-2 was not affected by meloxicam treatment. Conclusions: The present study demonstrates that the specific COX-2 inhibitor meloxicam suppresses proliferation and induces apoptosis in HCC cells that express COX-2, suggesting that COX-2 inhibition may offer a novel chemopreventive and therapeutic approach for HCC. [ABSTRACT FROM AUTHOR]