Hydroxyurea ameliorates mouse hypoxia through inducing embryonic hemoglobin

Objective To investigate the ameliorative effect and mechanism of hydroxyurea (HU) on hypoxia mice in a closed environment. Methods Sixty male C57BL/6J mice (6~8 weeks old, weighing 18~22 g) were randomly divided into normoxia control group (NC, n=10), normoxia HU group (NHU, n=10), hypoxia control group (HC, n=20), and hypoxia HU group (HHU, n=20).The HU mice were injected intraperitoneally with 100 mg/kg HU daily for 4 weeks, and the control mice were injected with same volume saline for 4 weeks.Then bone marrow was extracted for transcriptome sequence in the NC and NHU groups.The mice in the HC and HHU groups were settled in a sealed bottle to observe their survival, and the brains and kidneys were isolated for immunohistochemistry when these mice were in hypoxia for 15 min, and those from the NHU and NC groups served as control.RT-qPCR and Western blotting were adopted to verify the mRNA and protein expression of bone marrow and peripheral blood molecules, respectively. Results The survival time was significantly longer in the HHU group (P < 0.05), and the hypoxia of the brains and kidneys were ameliorated (P < 0.05).The oxygen affinity of peripheral blood was obviously raised, and partial pressure of oxygen 50(P50) was negatively correlated with survival time (Pearson=-0.738, P < 0.01).A total of 65 389 genes were found in transcription sequencing, including 50 up-regulated and 272 down-regulated genes in the mice from the NHU group (log2 FC≥1, Q < 0.05).Gene ontology (GO) enrichment analysis indicated that hemoglobin subunits (GO cellular component), hemoglobin complex (GO cellular function) and erythrocyte development (GO biological process) were significant enriched.Among the differentially expressed genes, Erfe, Hbb-y, Hbb-bh1 and Hba-x were up to 1.5 fold, and the down-regulated genes were erythropoiesis-related genes, Klf1, Sox6, Gata1 etc.Our results indicated that the mRNA levels of Hbb-y, Hbb-bh1 and Hba-x were notably enhanced in the bone marrow of the NHU group, and the protein level of HbE (ε-globin) protein was remarkably increased in the peripheral blood (P < 0.05). Conclusion HU exerts protective effect on the mice in a closed hypoxic condition, which may be related with its inducing embryonic hemoglobin with high oxygen affinity, and thus promoting oxygen carrying capacity.