Dynamin 2 along with microRNA-199a reciprocally regulate hypoxia-inducible factors and ovarian cancer metastasis.

Hypoxia-driven changes in the tumor microenvironment facilitate cancer metastasis. In the present study, we investigated the regulatory cross talk between endocytic pathway, hypoxia, and tumor metastasis. Dynamin 2 (DNM2), a GTPase, is a critical mediator of endocytosis. Hypoxia decreased the levels of DNM2. DNM2 promoter has multiple hypoxia-inducible factor (HIF)-binding sites and genetic deletion of them relieved hypoxia-induced transcriptional suppression. Interestingly, DNM2 reciprocally regulated HIF. Inhibition of DNM2 GTPase activity and dominant-negative mutant of DNM2 showed a functional role for DNM2 in regulating HIF. Furthermore, the opposite strand of DNM2 gene encodes miR-199a, which is similarly reduced in cancer cells under hypoxia. miR-199a targets the 3'-UTR of HIF-1α and HIF-2α. Decreased miR-199a expression in hypoxia increased HIF levels. Exogenous expression of miR-199a decreased HIF, cell migration, and metastasis of ovarian cancer cells. miR-199a-mediated changes in HIF levels affected expression of the matrix-remodeling enzyme, lysyloxidase (LOX). LOX levels negatively correlated with progression-free survival in ovarian cancer patients. These results demonstrate a regulatory relationship between DNM2, miR-199a, and HIF, with implications in cancer metastasis.

Economical spectrophotometric method for estimation of zaltoprofen in pharmaceutical formulations.

AIM: A simple, rapid, precise, and economical spectrophotometric method has been developed for quantitative analysis of zaltoprofen (ZLT) in pharmaceutical formulations. MATERIALS AND METHODS: A mixture of methanol and water was used as a solvent. Initial stock solution of ZLT was prepared in methanol and subsequent dilution was done in water. The standard solution of ZLT in water showed two absorption maxima, one at 243.5 nm and another at 338.0 nm. RESULTS: The drug obeyed Beer-Lambert's law in the concentration range of 1-40 µg/mL with regression 0.9999 at 243.5 nm and 5-100 µg/mL with regression 0.9999 at 338.0 nm. The overall % recovery was found to be 99.53% and 99.77% at 243.5 nm and 338.0 nm, respectively, which reflect that the method is free from interference of the impurities and other additives used in tablet formulation. Relative standard deviations of absorbance from six measurements were always less than 2%. CONCLUSIONS: The results of analysis have been validated as per ICH guidelines. Both the wavelengths can be adopted in routine analysis of ZLT in tablet dosage form.

Hypoxia-induced microRNA-424 expression in human endothelial cells regulates HIF-α isoforms and promotes angiogenesis.

Adaptive changes to oxygen availability are critical for cell survival and tissue homeostasis. Prolonged oxygen deprivation due to reduced blood flow to cardiac or peripheral tissues can lead to myocardial infarction and peripheral vascular disease, respectively. Mammalian cells respond to hypoxia by modulating oxygen-sensing transducers that stabilize the transcription factor hypoxia-inducible factor 1α (HIF-1α), which transactivates genes governing angiogenesis and metabolic pathways. Oxygen-dependent changes in HIF-1α levels are regulated by proline hydroxylation and proteasomal degradation. Here we provide evidence for what we believe is a novel mechanism regulating HIF-1α levels in isolated human ECs during hypoxia. Hypoxia differentially increased microRNA-424 (miR-424) levels in ECs. miR-424 targeted cullin 2 (CUL2), a scaffolding protein critical to the assembly of the ubiquitin ligase system, thereby stabilizing HIF-α isoforms. Hypoxia-induced miR-424 was regulated by PU.1-dependent transactivation. PU.1 levels were increased in hypoxic endothelium by RUNX-1 and C/EBPα. Furthermore, miR-424 promoted angiogenesis in vitro and in mice, which was blocked by a specific morpholino. The rodent homolog of human miR-424, mu-miR-322, was significantly upregulated in parallel with HIF-1α in experimental models of ischemia. These results suggest that miR-322/424 plays an important physiological role in post-ischemic vascular remodeling and angiogenesis.