Early Reperfusion After Brain Ischemia Has Beneficial Effects Beyond Rescuing Neurons

Abstract

Background and Purpose-Recent studies show that successful endovascular thrombectomy 6 to 12 hours after stroke onset enhances functional outcomes 3 months later. In this study, we investigated the effects of reperfusion after ischemia on repair processes in the ischemic areas, as well as on functional recovery, using mouse stroke models. Methods-We examined time-dependent histological changes and functional recovery after transient middle cerebral artery occlusion of different durations, including permanent middle cerebral artery occlusion, using the CB-17 (CB-17/lcr-+/+Jcl) mouse strain, which has poor pial collateral blood flow. Results-Large microtubule-associated protein 2-negative areas of neuronal death were produced in mice subjected to ≥60 minutes of ischemia followed by reperfusion on day 1, while restricted microtubule-associated protein 2-negative regions were observed in mice subjected to a 45-minute period of ischemia. A substantial reduction in microtubule-associated protein 2-negative areas was observed on day 7 in mice given early reperfusion and was associated with better functional recovery. Klüver-Barrera staining demonstrated that white matter injury on day 1 was significantly lesser in mice with reperfusion. Immunohistochemistry and electron microscopy revealed that a greater number of endothelial cells were present in the infarct areas in mice with earlier reperfusion and were associated with a more rapid recruitment of plateletderived growth factor receptor β-positive pericytes and subsequent intrainfarct fibrosis. Early reperfusion also resulted in a greater accumulation of glial fibrillary acidic protein-positive astrocytes in peri-infarct areas. Peri-infarct astrogliosis was attenuated in platelet-derived growth factor receptor β heterozygous knockout mice. Conclusions-Early reperfusion after ischemia enhances the survival of endothelial cells and pericytes within ischemic areas even after the infarct is established, resulting in efficient intrainfarct fibrosis and peri-infarct astrogliosis. These effects might be associated with efficient peri-infarct reorganization and functional recovery. in peri-infarct areas in experimental researches, although this remains controversial. 21,22 Furthermore, it is experimentally suggested that efficient fibrotic formation within areas of infarct may promote peri-infarct astrogliosis. 23 Thus, if these repair processes are associated with functional recovery, apparent infarct volumes at early phases may not be suitable for predicting functional outcome.To date, few papers have focused on the effects of reperfusion on poststroke repair processes in both animals and human. In the present study, we hypothesized that early reperfusion might elicit efficient tissue repair, such as intrainfarct fibrosis and peri-infarct astrogliosis, and lead to functional recovery even if it does not prevent neuronal death within the ischemic areas. To examine the precise effects of ischemia-reperfusion in the MCA, we performed tMCAO of various durations, as well as perm...

Pericyte-Mediated Tissue Repair through PDGFRβ Promotes Peri-Infarct Astrogliosis, Oligodendrogenesis, and Functional Recovery after Acute Ischemic Stroke

“…All data are presented as the mean 6 SEM; p , 0.05 was considered significant. The sample sizes were similar to those reported in previous publications (Tachibana et al, 2017;Komori et al, 2019) and are indicated in the relevant figure legends. No statistical method was used to predetermine sample size.…”

“…Epidemiological studies also demonstrate that various factors, including age, sex, and status of life-style diseases, can affect post-stroke functional recovery. These findings encourage the exploration of therapeutic strategies that promote recovery of function, i.e., neurorestoration, even during the subacute phase after ischemic stroke (Chen et al, 2014;Tachibana et al, 2017;Xing et al, 2017).…”

“…Whole brains were fixed with 4% PFA in PBS for approximately 3 d. PFA-fixed 2-mm-thick coronal slices were embedded in paraffin and sectioned at 4 mm. The sections were then deparaffinized, rehydrated through a graded series of ethanol solutions, and washed in PBS (Tachibana et al, 2017). After blocking with a solution of 5% skimmed milk or PBS containing 10% normal goat serum for 30 min at room temperature, the sections were incubated with primary antibodies: anti-microtubule-associated protein 2 (MAP2; 1:1000; Sigma-Aldrich #M4403), anti-PDGFRb [1:100; Cell Signaling Technology (CST) #3169], anti-CD13 (1:200; R&D Systems #AF2335), anti-CD31 (1:200; BD Biosciences #550274), anti-glial fibrillary acidic protein (GFAP; 1:200; CST #3670), anti-OLIG2 (1:200; R&D Systems #AF2418), anti-APC (1:100; Calbiochem #OP80), anti-ionized calcium-binding adapter molecule 1 (IBA1; 1:500; Abcam #ab5076), anti-myelin basic protein (MBP; 1:500; Millipore #MAB386), anti-SMI312 (1:500; BioLegend #837904), anti-doublecortin (DCX; 1:1000; Millipore #AB2253), anti-Fibrin (1:1000; DAKO #A0080), anti-NG2 (1:200; Millipore #AB5320), anti-interleukin-6 (IL6; 1:200; R&D Systems #AF206), or anti-pSTAT3 (1:1000; CST #9145) at 4°C overnight.…”

“…In the scores for severity of injury, 1 point is awarded for the inability to perform the test or for the lack of a tested reflex; thus, the higher the score, the more severe the injury. The cylinder test was performed as described previously (Tachibana et al, 2017). A mouse was placed in a clear acrylic cylinder 9 cm in diameter and 20 cm in height and videotaped during the test.…”

“…Although a difficulty of pericyte research in the CNS is the lack of pericyte-specific markers, some reports have demonstrated that PDGFRb can be used as a pericyte marker, particularly after ischemic stroke: the expression of PDGFRb is induced remarkably and exclusively in pericytes within infarct areas Arimura et al, 2012). In these situations, it is still controversial whether pericyte-mediated fibrotic repair is beneficial or detrimental to peri-infarct neural reorganization and functional recovery (Su et al, 2008;Göritz et al, 2011;Makihara et al, 2015;Tachibana et al, 2017;Gautam and Yao, 2018). Thus, this is an important issue to be solved even from the standpoint of the development of novel therapeutic strategies that promote post-stroke functional recovery.…”