利用斑馬魚模式動物探討經缺氧回復後參與脊髓神經修復的特殊細胞群

Abstract

Following hypoxic stress, neuron stem/progenitor cells (NSPCs) and other uncharacterized cells of zebrafish central nervous system (CNS) thrive during recovery. To characterize the remaining cell population, we employed a zebrafish transgenic line, huORFZ, which harbors an inhibitory upstream open reading frame of human chop (huORFchop) fused with GFP reporter and driven by cytomegalovirus promoter. When huORFZ embryos were treated with hypoxic stress, followed by oxygen recovery, the appearance of GFP indicated that some CNS cells survived and successfully repressed the translational inhibition caused by huORFchop. These GFP-(+) cells, termed hypoxia-responsive recovering cells, or HrRCs, were primarily some NSPCs and reactive radial glia cells (RGs), along with some oligodendrocyte progenitor cells (OLPs) and oligodendrocytes (OLs). By in vitro assay, we demonstrated that these cultured HrRCs were able to differentiate into mature unipolar neurons. By in vivo examination, we found that (1) GFP-(+) HrRCs did not undergo apoptosis, while GFP-(-) neurons did. (2) HrRCs were able to migrate; (3) among HrRCs, only GFP-(+) NSPCs and GFP-(+) RGs proliferated and differentiated into mature functional neurons after oxygen recovery; (4) prolonged recovery time after hypoxic stress correlated with higher proportions of GFP-(+) NSPCs and GFP-(+) RGs that had differentiated into neurons, in contrast to lower proportions of proliferating/differentiating GFP-(-) NSPCs and GFP(-) RGs; (5) the number of NSPCs and RGs differentiating into neurons was low in unstressed embryos, suggesting that embryonic development is not associated with the differentiation of HrRCs into neurons; and (6) specific ablation of 15 HrRCs in the spinal cord of each stress-treated huORFZ embryo severely impaired its swimming performance. Therefore, we demonstrated that type-specific cell populations which respond sensitively to hypoxic stress play an important role during the process of neuronal regeneration of zebrafish spinal cord.