Our previous study showed that miR was highly expressed in

Our previous study showed that miR-210 was highly expressed in neural precursor hiv integrase inhibitor (NPCs) under hypoxic conditions (Liu et al., 2011). To further understand the role of miR-210 in NPCs, TargetScan was used to search for miR-210 putative target genes. Based on conserved seed sequences and the observed functional effects of miR-210 on NPC growth, we confirmed that caspase-8 associated protein-2 (Casp8ap2), iron–sulfur scaffold protein (ISCU) and brain-derived neurotrophic factor (BDNF) are potential targets of miR-210 in NPCs under hypoxia (Supplementary information, Fig. S1), which is consistent with previous reports (Chen et al., 2010; Kim et al., 2009). Interestingly, we found that the Bcl-2 adenovirus E1B 19kDa-interacting protein 3 (BNIP3) is a potential target of miR-210 in the NPCs under hypoxia. BNIP3 was first identified using the adenovirus E1B-19K protein as bait in a yeast two-hybrid screen (Zhang and Ney, 2009). The promoter of BNIP3 contains a functional hypoxia response element (HRE), and its mRNA and protein expression are dramatically increased in multiple cell types in response to hypoxia (Bruick, 2000). Thus, BNIP3 is transcriptionally regulated by the hypoxia inducible factor (HIF) complex under hypoxic conditions. Increasing evidence indicates that BNIP3 functions as a pro-apoptotic BH3-only protein associated with the pathogenesis of many diseases, including cancer and cardiovascular disease (Azad et al., 2008; Vande Velde et al., 2000). Recent studies suggested that BNIP3 may have a dual function in the myocardium, where it regulates both mitochondrial turnover via autophagy and cell death (Zhang and Ney, 2009). In the present study, we report for the first time that miR-210 directly regulates BNIP3 expression to reduce the hypoxia-induced cell death of neural progenitor cells.
Materials and methods
Results
Discussion The present study is the first to identify BNIP3 as a target of miR-210. Although BNIP3 is transcriptionally regulated by HIF-1 (Bruick, 2000), posttranscriptional control by miR-210 contributes significantly to BNIP3 regulation. Moreover, we showed that hypoxia-induced upregulation of miR-210 plays an anti-apoptotic role in NPCs via suppressing BNIP3-induced translocation of AIF into nucleus. This anti-apoptotic effect may contribute to NPC survival under severe hypoxia. And, as both miR-210 and BNIP3 are key target genes of HIF-1, the discovery of miR-210-BNIP3 contributes to the regulation network of HIF-1 under hypoxia in NPC and other cell types, which deserves further investigation. MiR-210 is a target gene of HIF-1, and its promoter region, immediately upstream of miR-210 stem-loop structure, contains functional hypoxia-responsive element (HRE) (Huang et al., 2009). MiR-210 is the only miRNA consistently upregulated in all published studies, in both normal and transformed cells exposed to hypoxia (Biswas et al., 2010; Chen et al., 2010; Cheng et al., 2005; Fasanaro et al., 2008; Gee et al., 2010; Huang et al., 2009, 2010; Kim et al., 2009; Pulkkinen et al., 2008). Our previous work showed that miR-210 was highly expressed in NPCs under hypoxic conditions. NPCs are self-renewing and can differentiate into neurons, astrocytes or oligodendrocytes in vitro, exist widely in the developing and adult mammalian brain. Our previous studies showed that hypoxia promoted the proliferation of NPCs in vitro and in vivo (Zhao et al., 2008; Zhu et al., 2005). Therefore, upregulation of miR-210 may be involved in hypoxia-induced proliferation of NPCs. A number of independent studies have identified diverse functions of miR-210 involve in almost every aspect of hypoxia related biology, such as angiogenesis, apoptosis, proliferation, differentiation, cell cycle regulation, DNA damage repair, mitochondrial metabolism and tumor growth. Cheng et al. first described that blockade of miR-210 increased the apoptosis of HeLa cells (Cheng et al., 2005). Most recently, studies showed that miR-210 was induced in both brain and blood plasma of rat middle cerebral artery occlusion (MCAO) model (Jeyaseelan et al., 2008; Liu et al., 2010). Zeng et al. reported that miR-210 level in stroke patients with good outcome was significantly higher than patients with poor outcome (Zeng et al., 2011). We found that miR-210 mimics can reduce apoptosis of hypoxia treated NPCs, whereas miR-210 inhibitor plays an opposite role. These studies suggested that hypoxia-induced miR-210 may have cytoprotective effects on promoting the survival of brain cells.