Effect of hypoxia on HIF-1α and NOS3 expressions in CD34+ cells of JAK2V617F-positive myeloproliferative neoplasms.

PURPOSE: Myeloproliferative neoplasms (MPN) are a heterogeneous group of hematopoietic stem-cell diseases with excessive proliferation of one or more blood cell lines. In this study, we evaluated the effect of different oxygen concentrations on HIF-1α and NOS3 gene expression to determine the effect of the bone marrow microenvironment on JAK2V617F positive Philadelphia chromosome negative (Ph-) MPNs. PATIENTS AND METHODS: Peripheral blood mononuclear cells (MNC) of 12 patients with Ph- MPN were collected. The presence of JAK2V617F allele status was determined with allele-specific nested PCR analysis. MPN CD34+ and CD34depleted populations were isolated from MNC by magnetic beads. Separate cell cultures of CD34+/depleted populations were managed at different oxygen concentrations including anoxia (∼0%), hypoxia (∼3%), and normoxia (∼20%) conditions for 24 â€‹h. HIF-1α and NOS3 gene expression changes were examined in each population related to JAK2V617F status with real time RT-PCR. RESULT: It was revealed that relative HIF-1α and NOS3 expressions were significantly increased in response to decreased oxygen concentration in all samples. Relative HIF-1α and NOS3 expressions were found to be higher especially in CD34+ and CD34depleted populations carrying JAK2V617F mutations compared to MPN patients carrying wild-type JAK2. CONCLUSION: JAK2V617F might have specific role in HIF-1α and NOS3 regulations with respect to low oxygen concentrations in Ph- MPN. Further evaluations might reveal the effect of JAK2V617F on Ph- MPN pathogenesis in bone marrow microenvironment.

A new enrichment approach for candidate gene detection in unexplained recurrent pregnancy loss and implantation failure.

Recurrent pregnancy loss (RPL) and implantation failure (RIF) are obstacles to livebirth and multifactorial conditions in which nearly half of the cases remain unexplained, and we aimed to identify maternal candidate gene variants and pathways for RPL and RIF by analyzing whole-exome sequencing (WES) data via a new detailed bioinformatics approach. A retrospective cohort study was applied to 35 women with normal chromosomal configuration diagnosed with unexplained RPL and/or RIF. WES and comprehensive bioinformatics analyses were performed. Published gene expression datasets (n = 46) were investigated for candidate genes. Variant effects on protein structure were analyzed for 12 proteins, and BUB1B was visualized in silico. WES and bioinformatics analyses are effective and applicable for studying URPL and RIF to detect mutations, as we suggest new candidates to explain the etiology. Forty-three variants in 39 genes were detected in 29 women, 7 of them contributing to oligogenic inheritance. These genes were related to implantation, placentation, coagulation, metabolism, immune system, embryological development, cell cycle-associated processes, and ovarian functions. WES, genomic variant analyses, expression data, and protein configuration studies offer new and promising ways to investigate the etiology of URPL and RIF. Discovering etiology-identifying genetic factors can help manage couples' needs and develop personalized therapies and new pharmaceutical products in the future. The classical approach with chromosomal analysis and targeted gene panel testing is insufficient in these cases; the exome data provide a promising way to detect and understand the possible clinical effects of the variant and its alteration on protein structure.