Effects of dexamethasone and HA1077 on actin cytoskeleton and ß-catenin in cultured human trabecular meshwork cells.

AIM: To investigate the effects of dexamethasone (DEX) and 1-(5-isoquinolinesulfonyl)-homopiperazine (HA1077) on actin cytoskeleton and ß-catenin in cultured human trabecular meshwork (HTM) cells. METHODS: The HTM cells were separated from human eyeball and cultured in vitro. They were divided into control group, DEX (1×10-6 mol/L) group, HA1077 (3×10-5 mol/L) group, and DEX (1×10-6 mol/L) and HA1077 (3×10-5 mol/L) group. Actin cytoskeleton and ß-catenin in HTM cells of the four groups were examined by immunofluorescence and Western blot analyses. RESULTS: In DEX group, there were reorganization of actin cytoskeleton and formation of cross linked actin networks (CLANs), which were partially reversed in DEX and HA1077 group. DEX treatment also induced an increased expression of ß-catenin, which was obviously reduced in DEX and HA1077 group. Meanwhile, the cultured HTM cells in HA1077 group had lower expression of ß-catenin than that in the control group. CONCLUSION: Our results show that HA1077 can reverse the changes of actin organization and expression of ß-catenin induced by DEX in cultured HTM cells, suggesting that HA1077 may play an important role in increasing outflow and reducing intraocular pressure.

Tumor necrosis factor-α increases angiopoietin-like protein 2 gene expression by activating Foxo1 in 3T3-L1 adipocytes.

Angiopoietin-like protein 2 (Angptl2) is a key adipocyte-derived inflammatory mediator linking obesity to systemic insulin resistance, which is overexpressed in obesity and related metabolic diseases. However, its regulatory mechanism remains unclear. In this study, we showed that tumor necrosis factor (TNF)-α treatment increased the expression of Angptl2 gene in 3T3-L1 adipocytes. The cloning and sequence analysis of the Angptl2 gene promoter revealed the presence of several putative-binding sites for transcriptional factors, including two IREs. Insulin suppressed Angptl2 mRNA expression in dose-dependent manners, which could be attenuated by a phosphoinositide 3-kinase (PI3K) inhibitor LY294002. The interactions between IRE sites within Angptl2 promoter and forkhead transcription factor Foxo1 were identified by EMSA and ChIP assay. Furthermore, lentivirus-mediated knockdown of Foxo1 expression inhibited the transcriptional activity of Angptl2 promoter and decreased Angptl2 mRNA expression. Finally, TNF-α inhibited Foxo1 phosphorylation and enhanced its transcriptional activity, through which TNF-α increased the expression of Angptl2 in adipocytes. These results suggest that TNF-α up-regulates Angptl2 mRNA expression via PI3K/Foxo1 pathway in 3T3-L1 adipocytes, which may be involved in obesity-induced inflammation and insulin resistance.