The prognosis of HIBD in the developing stage mainly depends on neural injury and repair. Neural injury is difficult to reverse; thus, neural repair, including neurogenesis after injury, plays an important role in the recovery of neurological function. In this study, we reported that overexpression of TERT attenuated the apoptosis in NSCs, induced proliferation, migration, and differentiation of NSCs, and promoted myelination in the brain of neonatal rats after HIBD. Our results showed that TERT could improve the learning and memory ability and neurological function of neonatal rats after HIBD. Further research indicated that the mechanism of TERT function in neurogenesis after HIBD may be related to the SHH /Gli1 signaling pathway.
Neurogenesis of the central nervous system is a complex process that involves the proliferation of NSCs, their gradual migration to functional regions, continuous neoplastic changes, differentiation into nerve cells, and establishment of contact with other nerve cells to preserve neurological function. When neurogenesis is activated after brain injury, NSCs play a significant role in neural repair and can promote post-injury repair in the damaged brain (Yin et al. 2013). Previous studies have already reported that neurogenesis after hypoxic ischemic injury in the neonatal brain was closely related to brain repair and recovery after injury (Yang and Levison 2007; Miles and Kernie 2008).
TERT is widely expressed in embryonic and early phases of the postnatal brain and decreases with neuronal differentiation (Klapper et al. 2001). TERT expression is observed only in highly proliferative regions of the adult brain, such as the hippocampus and olfactory bulb, which can provide NSCs for neurogenesis (Lee et al. 2010). These lines of evidence indicate that TERT may play a role in neuronal survival and differentiation and support neurogenesis in the brain (Klapper et al. 2001; Lee et al. 2010). In a study on adult mice, the proliferation and self-renewal ability of neural stem cells in TERT-deficient mice were significantly reduced, together with their ability to differentiate into various functional nerve cells, leading to the damage of neurological function. In contrast, reactivation of TERT improved the proliferation and differentiation ability of NSCs, promoted remyelination, and improved neurological function (Jaskelioff et al. 2011). In another study on adult mice, TERT was reported to be involved in depression-related behavioral modification by promoting hippocampal neurogenesis (Zhou et al. 2011). Consistently, overexpression of TERT promoted the proliferation of NSCs in mouse brains both in vivo and in vitro (Liu et al. 2012). TERT is believed to serve as an effective springboard for promoting differentiation and immortalization of stem cells in stem cell therapies (Yalvaç et al. 2011). However, whether TERT is involved in the process of neurogenesis in the neonatal brain after HIBD remains unclear. In this study on neonatal HIBD, we found that TERT attenuated the apoptosis in NSCs and promoted the proliferation, migration, and differentiation of NSCs in the neonatal brain after HIBD.
Myelination is an important part of the neurodevelopment process because it allows axons to connect to neurons, accelerate nerve conduction, and strengthen circuity throughout the nervous system. During neonatal HI brain injury, myelination is disrupted, thereby impairing functional recovery from brain injury and resulting in static motor deficits (Koch et al. 2008; Kaminski et al. 2020). In the present study, myelination was evaluated by immunofluorescence of MBP, a primary protein component of the myelin sheath. Our results suggest that TERT improves the neurodevelopmental process after HIBD by promoting myelination. Research on the role of TERT in regulating myelination is rare, and further studies are required to clarify the mechanism by which TERT promotes myelination.
Neurogenesis and myelination, which are important parts of post-injury repair, are closely related to the recovery of neurological function. In this study, neurological function after injury was estimated using the MWM test and mNSS. The MWM was widely performed to evaluate learning and memory ability during the research of neonatal hypoxic ischemic brain injury (Gao et al. 2020). The MWM test showed that the escape latency was improved by overexpression of TERT, indicating that TERT can improve the dysfunction of learning and memory impairment caused by HIBD. The mNSS is a behavioral functional experiment used to evaluate neurological function, which is also applied in the research of HI brain injury (Ge et al. 2018). The mNSS score in neonatal rats after HIBD increased at 14th day, while TERT overexpression reduced the mNSS scores. Together with the MWM test, our research indicated that TERT could improve the neurological function in neonatal rats after HIBD, and therefore promote the recovery of neurological function after HIBD.
Taken together, our results indicated that TERT could improve the learning and memory ability and neurological function in neonatal rats after HIBD by improving neurogenesis and myelination. Additionally, we investigated the underlying mechanism of TERT in neurogenesis. Gli1 is an important downstream transcriptional factor of the SHH signaling pathway and can accurately reflect the activity of this pathway. Previous studies have reported that the SHH/Gli signaling pathway was involved in neurological development and central nervous system diseases (Ruiz i Altaba et al. 2002). Activation of the SHH/Gli pathway could promote the proliferation and differentiation of neural precursor cells during brain growth (Ruiz i Altaba et al. 2002). In adult mice, Gli1 has been reported to participate in regulating the proliferation of NSCs and neurogenesis in the hippocampus (Sun et al. 2018). Tayyab M et al. have reported that the SHH/Gli1 signaling pathway was an effective regulator of neurogenesis during embryonic development and in the adult hippocampus (Tayyab et al. 2018). In addition, both in vivo and in vitro studies have confirmed the neuroprotective role of the SHH/Gli1 pathway in promoting neurogenesis and thus improving neurological function after ischemic brain injury (Yu et al. 2017). However, little is known about the role of the SHH/Gli1 pathway in neonatal HIBD. Here, we found that Gli1 expression was increased after neonatal HIBD, and overexpression of TERT further increased the expression level of Gli1, indicating that the SHH/Gli1 signaling pathway may be involved in the neuroprotective effect of TERT during neonatal HIBD. A previous study in cancer cells proposed TERT as a target of the Hedgehog (HH)/Gli signaling pathway. Furthermore, Gli1 suppression reduced TERT expression in cancer cells (Mazumdar et al. 2013). However, studies on the relationship between TERT and Gli1 are quite rare, and further research is needed to elucidate the interaction between TERT and Gli1, especially in neonatal HIBD.
In summary, the present study demonstrated that TERT attenuated the apoptosis in NSCs, promoted the proliferation, migration, and differentiation of NSCs, and induced myelination in neonatal rats after HIBD. TERT could improve the learning and memory ability and neurological function in neonatal rats after HIBD. Moreover, the SHH/Gli1 signaling pathway might be involved in the neuroprotective effect of TERT in neonatal HIBD. Since neurogenesis is closely related to neural repair and prognosis after brain injury, and the repair effect of endogenous neurogenesis is limited, we believe that therapies such as those enhancing endogenous stem cell activity will become a hot topic in the treatment of HIE. To date, the role of TERT in neurogenesis after HIBD is a new field. In this study, the neural repair effect and related molecular mechanism of TERT were discussed, and a new neuroprotective pathway regulated by TERT was found, which could provide novel insights into the therapeutic strategies for neonatal HIE. In addition, it suggests new possibilities for the neuronal repair and reconstruction in other diseases affecting nervous system using TERT as the target.