Role of p38 MAPKs in hypericin photodynamic therapy-induced apoptosis of nasopharyngeal carcinoma cells.

The present study aims to determine the role of mitogen-activated protein kinases (MAPKs) in hypericin-mediated photodynamic therapy (HY-PDT)-induced apoptosis of the HK-1 nasopharyngeal carcinoma (NPC) cells. HY-PDT was found to induce proteolytic cleavage of procaspase-9 and -3 in HK-1 cells. Apoptotic nuclei were observed at 6 h after PDT whereas B-cell leukemia/lymphoma-2-associated-X-protein (Bax) translocation and formation of Bax channel is responsible for the cell death. Increase in phosphorylation of p38 MAPKs and c-Jun N-terminal kinase 1/2 (JNK1/2) was detected at 15-30 min after HY-PDT. The appearance of phosphorylated form of p38 MAPKs and JNK1/2 was inhibited by the singlet oxygen scavenger l-histidine. HY-PDT-induced cell death was enhanced by the chemical inhibitors for p38 MAPKs (SB202190 and SB203580), but not by the JNKs inhibitor SP600125. Knockdown of the p38alpha and p38beta MAPK isoforms by small interfering RNA (siRNA) are more effective than the p38delta in enhancing PDT-induced cell death. Augmentation of apoptosis by p38alpha or p38beta knockdown is also correlated with the increased proteolytic cleavage of procaspase-9 after HY-PDT treatment. Our results suggested that HY-PDT activated p38 MAPKs through the production of singlet oxygen. Inhibition of p38 MAPKs with chemical inhibitors or siRNA enhances HY-PDT-induced apoptosis of the HK-1 NPC cells.

Hypoxia-inducible factor-1-mediated activation of stanniocalcin-1 in human cancer cells.

Stanniocalcin-1 (STC1) is an endocrine hormone originally discovered in the corpuscles of Stannius, endocrine glands on kidneys of bony fishes, and also has been identified in mammals. The mammalian STC1 gene is widely expressed in various tissues and appears to be involved in diverse biological processes. There is growing evidence to suggest that altered patterns of gene expression have a role in human cancer development. Recently STC1 has been identified as a stimulator of mitochondrial respiration and has been hypothesized to be functionally related to the Warburg effect, of which hypoxia-inducible factor (HIF)-1 plays a key role in reprogramming tumor metabolism. This prompted us to examine the involvement of HIF-1 in the regulation of STC1 expression in tumor hypoxia. Our data reveal that hypoxia can stimulate STC1 gene expression in various human cancer cell lines, including those derived from colon carcinomas, nasopharyngeal cancer (CNE-2, HONE-1, HK-1), and ovarian cancer (CaOV3, OVCAR3, SKOV3). By far, the greatest response was observed in CNE-2 cells. In further studies on CNE-2 cells, desferrioxamine, cobalt chloride, and O(2) depletion all increased HIF-1alpha protein and STC1 mRNA levels. Desferrioxamine treatment, when coupled with Fe replenishment, abolished these effects. RNA interference studies further confirmed that endogenous HIF-1alpha was a key factor in hypoxia-induced STC1 expression. The ability of vascular endothelial growth factor to stimulate STC1 expression in CNE-2 cells was comparatively low. Collectively, the present findings provide the first evidence of HIF-1 regulation of STC1 expression in human cancer cells. The studies have implications as to the role of STC1 in hypoxia induced adaptive responses in tumor cells.

Identification of signal transduction pathways that modulate dibutyryl cyclic adenosine monophosphate activation of stanniocalcin gene expression in neuroblastoma cells.

Stanniocalcin (STC) is a new mammalian polypeptide hormone and appears to be a regulator of neuronal function. We have already shown that the induction of STC mRNA and protein expression by cAMP is integral to neuroblastoma cell differentiation, particularly neurite outgrowth. In this study, we examined the cAMP pathway in greater detail. Some common neuritogenic agents, euxanthone (PW1) and trans-retinoic acid (RA), were studied for possible interactions with the dibutyryl cAMP (dbcAMP)-mediated response. Our results showed that STC mRNA induction by dbcAMP was mediated by protein kinase A-cAMP response element binding protein (CREB) pathway, accompanied with phosphorylation of CREB and a reduction of p50, p65, and phosphorylated inhibitor kappaBalpha levels. Using a synthetic peptide nuclear factor-kappaB SN50, stimulation of dbcAMP-mediated STC expression was observed; indicating the nuclear translocation of nuclear factor kappaB might possibly repress STC expression. dbcAMP-induced STC mRNA expression was enhanced by PW1. In contrast, RA had highly suppressive effects. Cotreatment of cell with PW1 and cAMP provoked an increase in phosphorylated CREB (pCREB). Conversely, cotreatment with RA suppressed pCREB. The results highlighted the importance of phosphorylation of CREB in mediating STC gene expression. Taking a step further to dissect the possible regulatory pathways involved, with the aid of phorbol 12-myristate 13-acetate or ionomycin, additive effects on STC gene expression were observed. The induction was aided by further elevation of pCREB, which was completely abolished by Gö 6976, a Ca2+-dependent protein kinase C (PKC) alpha and PKCbeta1 inhibitor. Our results indicated that cross-talk with PKC and/or Ca2+ signaling pathways might sensitize cAMP-mediated effects, on CREB phosphorylation and STC gene expression.

Anti-inflammatory and antiviral effects of indirubin derivatives in influenza A (H5N1) virus infected primary human peripheral blood-derived macrophages and alveolar epithelial cells.

Human disease caused by highly pathogenic avian influenza A (HPAI) (H5N1) is associated with fulminant viral pneumonia and mortality rates in excess of 60%. Acute respiratory syndrome (ARDS) has been found to be the most severe form of acute lung injury caused by H5N1 virus infection while cytokine dysregulation and viral replication are thought to contribute to its pathogenesis. In this study, the antiviral and anti-inflammatory effects of two indirubin derivatives: indirubin-3'-oxime (IM) and E804 on primary human peripherial blood-derived macrophages and type-I like pneumocytes (human alveolar epithelial cells) during influenza A (H5N1) virus infection were investigated. We found that both of the indirubin derivatives strongly suppress the pro-inflammatory cytokines including IP-10 (CXCL10), one of the key factors which contribute to the lung inflammation during H5N1 virus infection. In addition, we also demonstrated that the indirubin derivative delays the virus replication in the primary cell culture models. Our results showed that indirubin derivatives have a potential to be used as an adjunct to antiviral therapy for the treatment of severe human H5N1 disease.

Bisindigotin, a TCDD antagonist from the Chinese medicinal herb Isatis indigotica.

A new indigoid derivative, bisindigotin (1), with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-antagonistic activity was isolated from the ethanol extract of the Chinese medicinal herb Isatis indigotica. Its structure was determined by spectroscopic methods. In the human HepG2 hepatoma cell model, 1 (50 nM to 2 microM) was found to dose-dependently inhibit TCDD-induced ethoxyresorufin O-deethylase (EROD) activity.