Stepwise impairment of neural stem cell proliferation and neurogenesis concomitant with disruption of blood-brain barrier in recurrent ischemic stroke
Introduction
Stroke is the fifth leading cause of death and the leading cause of disability in the US. Dementia is a common sequela in post-stroke patients. Cognitive impairment is three times more common in stroke patients than in those who have not had a stroke (Leys et al., 2005). Post-stroke dementia can be due to direct damage of the brain from injury itself or secondary damage due to inflammation, oxidative stress, and microglial activation (Hu et al., 2014; Zeng et al., 2013; Iadecola and Anrather, 2011a; Weishaupt et al., 2016; Schmidt et al., 2015). Importantly, not all cases of post-stroke dementia involve dementia-related brain structures and even remote or mild ischemia events can increase the incidence of dementia (Okada et al., 1995; Yonemori et al., 1999).
The hippocampus plays a particularly important role in cognitive function, memory formation and learning. Notably, the subgranular zone (SGZ) of the dentate gyrus in the hippocampus remains a site of neurogenesis throughout adult life (Seri et al., 2001; Palmer et al., 1997). Moreover, after middle cerebral artery occlusion (MCAO) in rats, there is significant neural stem cell (NSC) activation and enhanced neurogenesis observed in the SGZ (Lin et al., 2015). Similarly, other adult stem cell niches, including the subventricular zone (SVZ) of the lateral ventricle and novel niches along the third and fourth ventricles also become sites of much enhanced neurogenesis after stroke (Lin et al., 2015). Although the hippocampus is not thought to become ischemic in stroke (Okada et al., 1995; Yonemori et al., 1999), pathological changes to the hippocampus which impact neurogenesis could contribute to the cognitive deficits commonly seen in post-stroke patients.
Also, potentially impacting this neurogenic capability is the fact that the stem cell pool can deplete under a variety of conditions (Monje et al., 2003; Brazel et al., 2004; Sierra et al., 2015, Sierra et al., 2010; Kuhn et al., 1996; Darsalia et al., 2005; Spalding et al., 2013; Yu et al., 2010; Ekdahl et al., 2003; Khacho et al., 2016; Tepavčević et al., 2011; Walter et al., 2011). Indeed, Encinas et al. (2011) described the concept of “disposable” NSCs in the aging hippocampus, wherein quiescent NSCs in the SGZ undergo a series of approximately three rapid, asymmetric divisions after activation, and then differentiate into astrocytes. According to their results, repeated activation of quiescent NSCs eventually leads to depletion of the SGZ stem cell pool, which can result in a decline in cognitive ability.
We wondered whether multiple ischemic strokes could also impair brain stem cell niches like the SGZ. Although patients who survive a first stroke are at higher risk for recurrent stroke and post-stroke dementia compared to the general population (Leys et al., 2005; Mohan et al., 2011; Pendlebury and Rothwell, 2009), the effects of recurrent stroke on stem cell proliferation and neurogenesis have not yet been investigated. In fact, the only model of recurrent stroke utilizes a small focal photothrombotic injury and in these studies neurogenesis has not been examined (Schmidt et al., 2015). Since recurrent stroke most commonly occurs in patients with a previous stroke resulting from large-artery atherosclerosis (Lovett et al., 2004), we developed a more clinically relevant animal model of recurrent stroke using successive MCAOs. Utilizing this model, we examined whether first ever stroke-activated stem cell proliferation and neurogenesis in the SGZ and SVZ leads to the impairment of neurogenesis after a recurrent event. We further investigated the underlying mechanisms through which this might occur and whether the process is exacerbated by recurrent stroke.
Section snippets
Animals
All procedures were carried out in accordance with the Guide for the Care and Use of Laboratory Animals of the NIH and approved by the IACUC Committee of Thomas Jefferson University.
Animal treatment protocols and BrdU administration
Adult male Sprague-Dawley rats aged 8 weeks were anesthetized using ketamine hydrochloride, xylazine and acepromazine maleate (60 mg/kg, 10 mg/kg, 5 mg/kg respectively). Body temperature was monitored with a rectal temperature probe, and maintained with a heating pad and/or a small fan to within 0.5 °C.
For the first
Stroke activates stem cell proliferation and neurogenesis in SGZ
In Protocol 1, we found that 14 days after MCAO (group 2), there was a significant increase in the number of proliferating (BrdU+) NSCs in the ipsilateral SGZ and a smaller increase in the contralateral SGZ compared to controls (group 1) (Fig. 1). We also found that most BrdU+ cells in the SGZ co-expressed the neuroblast marker DCX after MCAO (Fig. 1B). These findings indicated that like the SVZ (Lin et al., 2015), neurogenesis was up-regulated 2–2.5 fold 14 days after MCAO in both SGZs (Fig. 1
Discussion
Stroke is one of the leading causes of death and disability in the world. Recurrent stroke, usually more devastating and lethal than the first, accounts for 25–30% of all preventable stroke (Hankey, 2014). The rate of recurrent ischemic stroke is estimated to be 12.6–13.2% per year in the US (Allen et al., 2010). Risk for recurrent stroke increases over time from a 3.1% risk at 30 days to 39.2% risk at 10 years after initial stroke (Mohan et al., 2011).
As one of the major causes of dependency
Acknowledgments
We acknowledge the excellent support of Dr. Lawrence Kenyon and Mrs. Bodil Tuma for EM, Jingli Cai and Eric Kostuk for neurosphere assay, Aurelie Ky for animal caring.
Sources of funding
This research was funded by the Joseph and Marie Field Foundation to RR and LI.
Disclosures
None.
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