Extracellular Vesicle-Derived microRNA-410 From Mesenchymal Stem Cells Protects Against Neonatal Hypoxia-Ischemia Brain Damage Through an HDAC1-Dependent EGR2/Bcl2 Axis

Abstract

Hypoxia-ischemia brain damage (HIBD) is a neurological disorder occring in neonates, which is exacerbated by neuronal apoptosis. Mesenchymal stem cells (MSCs)-derived extracellular vesicles (EVs) have been proposed as a promising strategy for treating or preventing ischemia-related diseases. However, their mechanisms in HIBD remain unclear. Thus, we aimed to address the role of EV-derived microRNA (miR)-410 in HIBD. Neonatal HIBD mouse model was constructed using HI insult, from which neurons were isolated, followed by exposure to oxygen glucose deprivation (OGD). EVs were isolated from human umbilical cord (hUC)-derived MSCs. In silico analyses, dual-luciferase reporter gene and chromatin immunoprecipitation assays were adopted to determine relationships among miR-410, histone deacetylase 1 (HDAC1), early growth response protein 2 (EGR2), and B cell lymphoma/leukemia 2 (Bcl2). The functional roles of EV-derived miR-410 were determined using loss- and gain-of functions experiments, and by evaluating neuronal viability, cell-cycle distribution and neuronal apoptosis in vitro as well as modified neurological severity score (mNSS), edema formation, and cerebral infarction volume in vivo. hUC-MSCs-derived EVs protected against HIBD in vivo and inhibited the OGD-induced neuronal apoptosis in vitro. miR-410 was successfully delivered to neurons by hUC-MSCs-EVs and negatively targeted HDAC1, which inversely mediated the expression of EGR2/Bcl2. Upregulation of EV-derived miR-410 promoted the viability but inhibited apoptosis of neurons, which was reversed by HDAC1 overexpression. EV-derived miR-410 elevation reduced mNSS, edema formation, and cerebral infarction volume by increasing EGR2/Bcl2 expression through downregulating HDAC1 expression in vivo. In summary, EV-derived miR-410 impeded neuronal apoptosis by elevating the expression of EGR2/Bcl2 via HDAC1 downregulation, thereby providing a potential strategy for treating or preventing HIBD.

The Role of Extracellular Vesicles in the Developing Brain: Current Perspective and Promising Source of Biomarkers and Therapy for Perinatal Brain Injury

“…Similarly, neuropathological assessment in cresyl violet stained sections of the neonatal mouse brains collected 7 days after HI and following repeated (1, 3, and 5 days after HI) bolus IP injections of human bone marrow MSC-EVs demonstrated a significant decrease in tissue atrophy within the striatum region compared to both platelet-derived EV (control EVs) and untreated HI controls (Kaminski et al, 2020). Additionally, studies of repeated IP injections of umbilical cord MSC-EVs to neonatal mice both 14 h before HI and immediately before and 3 h following HI, have revealed a significant reduction in edema and cerebral infarction volume compared to untreated HI mice (Han et al, 2020). Further, histological examination of neonatal mice brains 72 h following HI revealed systemic administration of mouse bone marrow MSC-EVs 24 h following HI, attenuated both edema and infarction volume of the ipsilateral hemisphere, as well as cortical tissue loss compared to untreated animals (Xin et al, 2020).…”

“…Thirteen studies that utilize EVs as a therapeutic intervention for the treatment of perinatal brain injury using an in-vivo model were identified in this unbiased review of the literature (Figure 2 and Table 1; Ophelders et al, 2016;Drommelschmidt et al, 2017;Shu et al, 2018;Gussenhoven et al, 2019;Sisa et al, 2019;Thomi et al, 2019a,b;Chu et al, 2020;Han et al, 2020;Kaminski et al, 2020;Xin et al, 2020;Ahn et al, 2021;Du et al, 2021). Eight of these studies used models of HI alone (Ophelders et al, 2016;Gussenhoven et al, 2019;Sisa et al, 2019;Chu et al, 2020;Han et al, 2020;Kaminski et al, 2020;Xin et al, 2020;Du et al, 2021), three studies used models of simulated infection in the setting of HI (Drommelschmidt et al, 2017;Thomi et al, 2019a,b), one study utilized a model of IVH (Ahn et al, 2021) and one study did not provide sufficient details within the publication to determine the model of brain injury (Shu et al, 2018; Table 1). 12/13 studies utilized MSC-derived EVs.…”

“…Of the eight perinatal brain injury studies which utilized bone marrow-derived MSC-EVs, only one study administered rat MSC-EVs in a rat model (Shu et al, 2018), two studies administered mouse MSC-EVs in a mouse model (Chu et al, 2020;Xin et al, 2020), whereas all other studies utilized human bone marrow-derived MSC-EVs on either a rat (Drommelschmidt et al, 2017), mouse (Sisa et al, 2019;Kaminski et al, 2020) or fetal sheep (Ophelders et al, 2016;Gussenhoven et al, 2019) model (Table 1 and Figure 3). In addition, 2/12 studies utilized human umbilical cord Wharton's jelly derived MSCs-EVs in a rat model (Thomi et al, 2019a,b), 1/12 studies utilized human umbilical cord blood MSC-EVs in a rat model (Ahn et al, 2021) and 1/12 studies utilized umbilical cord-derived MSC-EVs in a mouse model (Han et al, 2020; Table 1 and Figure 3). Interestingly, one study, which met the eligibility criteria used hydrogen sulfide to pretreat MSCs in order confer a neuroprotective phenotype in MSC-EVs.…”

“…However, only 3/13 of the perinatal studies identified in this systemic review have utilized intranasal administration of EVs in their animal models (Figure 3; Sisa et al, 2019;Thomi et al, 2019a,b). Despite the intranasal route offering several advantages, the authors of the other studies identified in this review adopted either intraperitoneal (IP) (Drommelschmidt et al, 2017;Han et al, 2020;Kaminski et al, 2020;Du et al, 2021), IV (Ophelders et al, 2016;Gussenhoven et al, 2019), intracardial (IC) (Chu et al, 2020;Xin et al, 2020), intracerebroventricular (ICV) (Shu et al, 2018;Ahn et al, 2021), or ipsilateral hemisphere (Han et al, 2020) routes of administration of EVs (Figure 3).…”

“…Furthermore, some studies administered EV therapy as a single dose post-injury (Sisa et al, 2019;Thomi et al, 2019b;Chu et al, 2020;Xin et al, 2020;Ahn et al, 2021), whereas others as a repeated dose either prior to and/or following injury (Ophelders et al, 2016;Drommelschmidt et al, 2017;Gussenhoven et al, 2019;Han et al, 2020;Kaminski et al, 2020;Du et al, 2021). There were also differences in the normalization methods of dose; it was either based on a mg/kg body weight (Thomi et al, 2019b), cell equivalents/kg body weight (Ophelders et al, 2016;Drommelschmidt et al, 2017;Gussenhoven et al, 2019;Han et al, 2020;Kaminski et al, 2020), particles/dose (Sisa et al, 2019) or µg of protein (Chu et al, 2020;Xin et al, 2020;Ahn et al, 2021;Du et al, 2021) approach (Table 1). In addition, one study did not provide clear information of the timing and dosage regimen of EVs (Shu et al, 2018).…”