Promoted Angiographic Reperfusion And Neuroprotection In Acute Ischemic Stroke

Objective: Without cerebral revascularization is a major reason of infarct lesion growth in acute ischemic stroke.Fortunately,this may be reversed by recent advances that provide increased revascularization with increased availability of endovascular procedures.However,the current improved effects of endovascular therapy are not able to match to the higher rate of revascularization due to lack of angiographic reperfusion which partly aroused from inflammation-microthrombus in microvasculature.Recent advances in susceptibility weighted imaging(SWI)have dramatically improved the visibility of small veins such as the medullary veins and venules.However,SWI cannot primarily used for in vivo detection of arterioles.The main reason is that there is tiny susceptibility difference between the arteries and the surrounding tissue.Therefore,to make the arteries visible as we see veins,we introduce a susceptibility shift in the vasculature by using an ultra-small superparamagnetic iron oxide(USPIO)contrast agent to enhance susceptibility weighted imaging(USPIO-SWI).Several animal studies demonstrated the efficacy of immune modulators like fingolimod and anti-CD49 d monoclonal antibody on different ischemic stroke animal models.Based on these promising results,several trials were initiated by us and others to preliminarily test the feasibility of immune modulation of ischemic stroke patients.However,the results of these trials are mixed.So,the purpose of this study was to develop a novel MRI method for evaluation of angiographic reperfusion in embolic middle cerebral artery occlusion rat model,and to compare alteplase based angiographic reperfusion with and without fingolimod in a preclinical randomized,blinded,placebo-controlled experiment study(p RCT).Leukemia Inhibitory Factor(LIF)is a member of the interleukin-6 family of cytokines that stimulates or inhibits cells proliferation,differentiation and survival.LIF can promote self-renewal of neural stem cells(NSCs)and plays an important role in neural support cells.Leukemia inhibitory factor(LIF)is a contributing factor to the neuroprotection of neural stem cells(NSCs)after ischemic stroke.Our aim was to explore whether LIF-transfected NSCs(LIF-NSCs)can ameliorate brain injury and promote neuroprotection in a rat model of cerebral ischemia.Methods: Experiment 1: To setup a method on angiographic reperfusion evaluation with MRI,a total of 15 rats were studied.Nine normal rats were used to optimize USPIO dosage for enhanced effect on arterial of SWI.The other 6 animals were subjected to embolic stroke model surgeries to investigate the feasibility of imaging cerebral arterioles with optimize dosage in the focal ischemic lesion.Experiment 2: To examine whether fingolimod enhance the action of tissue plasminogen activator(t PA)for acute stroke,the preclinical randomized,blinded,placebo-controlled(p RCT)experiment study was performed.Animals with a visible vessel occlusion on MR angiography and without intracranial hemorrhage on SWI were included at 2h after embolic middle cerebral artery occlusion(e MCAO).The primary end point was the lesion volume growth over 24 hours and the shift in the direction of a better outcome on hemorrhage transformation at 24 hours after ischemia.The exploring end points were the shift in the direction of a better outcome on angiographic reperfusion at 24 hours.The number of the sample size is 30 per group.Experiment 3: We transfected NSCs with a lentivirus carrying the LIF gene to stably overexpress LIF.This study consisted of both in vitro and in vivo experiments.In vitro,we tested the LIF-secreting capacity,proliferation and differentiation of LIF-NSCs.Next,we introduced the oxygen glucose deprivation(OGD)model,which mimics stroke conditions in vitro,to investigated the LIF-secreting capacity of LIF-NSCs,and the activation of caspase 3.In vivo,transient cerebral ischemia was induced in rats by 2 h of middle cerebral artery occlusion(MCAO),and LIF-NSCs were intravenously injected at 6 h post-ischemia.The ADC of DWI(day 1)and T2(day 3)and FA of DTI(day 28)images served to calculate lesion volume and white matter injury.Long term functional benefit derived from LIF-NSCs treatment initiated at 14 days after MCAO in stroke rats was assessed using m NSS tests.Moreover,we investigated the level of LIF,GFAP and MBP in rat brains through immunohistochemistry techonolgy.Results: USPIO-SWI showed cerebra arteriole at the dose of 300 μmol /kg,and imaging cerebral arterials to monitor angiographic perfusion of stroke models with this dose is feasible.The results from this p RCT trial confirmed that t PA treatment with fingolimod significantly reduced infarct growth(the mean infarct lesion growth: 34 mm3 vs 18 mm3,p = 0.015)and promote angiographic perfusion(m TICI: 69% vs 17%,p = 0.001)without impact on hemorrhage transformation.In this study,LIF-NSCs could stably and succefully express LIF and promoted the proliferation of NSCs in vitro.Under the condition of OGD,the LIF-NSCs increased the level of LIF and reduced caspase 3 activation.Both the LIF-NSCs and their differentiated astrocytes could successfully secret LIF at a relatively high level in the ischemic hemisphere of MCAO rats.In addition,we found that the LIF-NSC treatment improved functional recovery and decreased the lesion volume based on MRI detection in MCAO rats.Moreover,based on the pathology assessment,the LIF-NSC treatment also promoted white matter remodeling and increased glial cell regeneration.Conclusions: This study supports the feasibility of performing p RCTs with angiographic perfusion evaluation with USPIO-SWI in revascularization treatment translational study.And,this study suggests that the benefits of lymphocyte-targeted intervention may depend on vascular-protectants and promoting angiographic perfusion.LIF-NSCs reduced the number of apoptotic cells following OGD in vitro.LIF-NSCs treatment reduced infarction volume and improved neurological recovery,it also could ameliorate white matter injury in MCAO rats.The protective effect of LIF-NSCs on cerebral infarction may be that NSCs act as carriers to increase the expression of LIF in the lesions,which could be a potential treatment for cerebral infarction.