Nonhuman Primate Model Of Stroke Research Articles

Sleep dysfunction and gut dysbiosis related amino acids metabolism disorders in cynomolgus monkeys after middle cerebral artery occlusion

IntroductionThis study aimed to explore the characteristics of post-stroke sleep dysfunction and verify their association with gut dysbiosis and the related amino acid metabolism disorders. This was achieved by using fecal microbiota transplantation (FMT) in a non-human primate stroke model. MethodsTwenty adult male cynomolgus monkeys were divided into the sham (n = 4), middle cerebral artery occlusion (MCAO, n = 5), MCAO + FMT (n = 3), and donor (n = 8) groups. The MCAO+FMT group received FMT at post-MCAO week 4. Sleep parameters, gut microbiota, gamma-aminobutyric acid (GABA), and glutamine (Gln) in the cerebrospinal fluid (CSF) were measured at baseline and postoperative weeks 4, 8, and 12. ResultsAt postoperative weeks 4, 8, and 12, the MCAO group showed decreased sleep efficiency, measured as the percentage of sleep during the whole night (82.3 ± 3.2 % vs 91.3 ± 2.5 %, 79.0 ± 3.75 % vs 90.8 ± 3.2 %, and 69.5 ± 4.8 % vs 90.5 ± 2.7 %; all P < 0.05), lower relative abundance of Lactobacillus (all P < 0.05), and reduced GABA concentrations in the CSF (317.3 ± 30.6 nmol/L vs 437.7 ± 25.6 nmol/L, 303.1 ± 48.9 nmol/L vs 4 40.9 ± 37.8 nmol/L, and 337.9 ± 49.4 nmol/L vs 457.4 ± 39.2 nmol/L; all P < 0.05) compared with the sham group. Sleep efficiency at post-FMT weeks 4 and 8 (84.7 ± 1.1 % vs 79.0 ± 3.75 %, and 84.1 ± 2.0 % vs 69.5 ± 4.8 %; both P < 0.05) and GABA concentration in the CSF at post-FMT week 4 (403.1 ± 25.4 nmol/L vs 303.1 ± 48.9 nmol/L, P < 0.05) was higher in the MCAO+FMT group than in the MCAO group. ConclusionsPost-stroke sleep dysfunction in monkeys is characterized by impaired sleep coherence, associated with decreased levels of probiotics such as Lactobacillus, GABA, and Gln in the CSF and can be ameliorated using FMT.

Hypothermic neuroprotection by targeted cold autologous blood transfusion in a non-human primate stroke model

Over decades, nearly all attempts to translate the benefits of therapeutic hypothermia in stroke models of lower-order species to stroke patients have failed. Potentially overlooked reasons may be biological gaps between different species and the mismatched initiation of therapeutic hypothermia in translational studies. Here, we introduce a novel strategy of selective therapeutic hypothermia in a non-human primate ischemia-reperfusion model, in which autologous blood was cooled ex vivo and the cool blood transfusion was administered at the middle cerebral artery just after the onset of reperfusion. Cold autologous blood cooled the targeted brain rapidly to below 34 °C while the rectal temperature remained around 36 °C with the assistance of a heat blanket during a 2-h hypothermic process. Therapeutic hypothermia or extracorporeal-circulation related complications were not observed. Cold autologous blood treatment reduced infarct sizes, preserved white matter integrity, and improved functional outcomes. Together, our results suggest that therapeutic hypothermia, induced by cold autologous blood transfusion, was achieved in a feasible, swift, and safe way in a non-human primate model of stroke. More importantly, this novel hypothermic approach conferred neuroprotection in a clinically relevant model of ischemic stroke due to reduced brain damage and improved neurofunction. This study reveals an underappreciated potential for this novel hypothermic modality for acute ischemic stroke in the era of effective reperfusion.

CDC42 Might Be a Molecular Signature of DWI-FLAIR Mismatch in a Nonhuman Primate Stroke Model.

No definitive blood markers of DWI-FLAIR mismatch, a pivotal indicator of salvageable ischemic penumbra brain tissue, are known. We previously reported that CDC42 and RHOA are associated with the ischemic penumbra. Here, we investigated whether plasma CDC42 and RHOA are surrogate markers of DWI-FLAIR mismatch. Sixteen cynomolgus macaques (3 as controls and 13 for the stroke model) were included. Guided by digital subtraction angiography (DSA), a middle cerebral artery occlusion (MCAO) model was established by occluding the middle cerebral artery (MCA) with a balloon. MRI and neurological deficit scoring were performed to evaluate postinfarction changes. Plasma CDC42 and RHOA levels were measured by enzyme-linked immunosorbent assay (ELISA). The stroke model was successfully established in eight monkeys. Based on postinfarction MRI images, experimental animals were divided into a FLAIR (-) group (N = 4) and a FLAIR (+) group (N = 4). Plasma CDC42 in the FLAIR (-) group showed a significant decrease compared with that in the FLAIR (+) group (p < 0.05). No statistically significant difference was observed for plasma RHOA. The FLAIR (-) group showed a milder neurological function deficit and a smaller infarct volume than the FLAIR (+) group (p < 0.05). Therefore, plasma CDC42 might be a new surrogate marker for DWI-FLAIR mismatch.

A Reliable Nonhuman Primate Model of Ischemic Stroke with Reproducible Infarct Size and Long-term Sensorimotor Deficits.

A nonhuman primate model of ischemic stroke is considered as an ideal preclinical model to replicate various aspects of human stroke because of their similarity to humans in genetics, neuroanatomy, physiology, and immunology. However, it remains challenging to produce a reliable and reproducible stroke model in nonhuman primates due to high mortality and variable outcomes. Here, we developed a focal cerebral ischemic model induced by topical application of 50% ferric chloride (FeCl3) onto the MCA-M1 segment through a cranial window in the cynomolgus monkeys. We found that FeCl3 rapidly produced a stable intraarterial thrombus that caused complete occlusion of the MCA, leading to the quick decrease of the regional cerebral blood flow in 10 min. A typical cortical infarct was detected 24 hours by magnetic resonance imaging (MRI) and was stable at least for 1 month after surgery. The sensorimotor deficit assessed by nonhuman primate stroke scale was observed at 1 day and up to 3 months after ischemic stroke. No spontaneous revascularization or autolysis of thrombus was observed, and vital signs were not affected. All operated cynomolgus monkeys survived. Our data suggested that FeCl3-induced stroke in nonhuman primates was a replicable and reliable model that is necessary for the correct prediction of the relevance of experimental therapeutic approaches in human beings.

Adenosine A1R/A3R agonist AST-004 reduces brain infarction in mouse and rat models of acute ischemic stroke.

Acute ischemic stroke (AIS) is the second leading cause of death globally. No Food and Drug Administration (FDA) approved therapies exist that target cerebroprotection following stroke. Our group recently reported significant cerebroprotection with the adenosine A1/A3 receptor agonist, AST-004, in a transient stroke model in non-human primates (NHP) and in a preclinical mouse model of traumatic brain injury (TBI). However, the specific receptor pathway activated was only inferred based on in vitro binding studies. The current study investigated the underlying mechanism of AST-004 cerebroprotection in two independent models of AIS: permanent photothrombotic stroke in mice and transient middle cerebral artery occlusion (MCAO) in rats. AST-004 treatments across a range of doses were cerebroprotective and efficacy could be blocked by A3R antagonism, indicating a mechanism of action that does not require A1R agonism. The high affinity A3R agonist MRS5698 was also cerebroprotective following stroke, but not the A3R agonist Cl-IB-MECA under our experimental conditions. AST-004 efficacy was blocked by the astrocyte specific mitochondrial toxin fluoroacetate, confirming an underlying mechanism of cerebroprotection that was dependent on astrocyte mitochondrial metabolism. An increase in A3R mRNA levels following stroke suggested an intrinsic cerebroprotective response that was mediated by A3R signaling. Together, these studies confirm that certain A3R agonists, such as AST-004, may be exciting new therapeutic avenues to develop for AIS.

Quantitative behavioral evaluation of a non-human primate stroke model using a new monitoring system.

BackgroundRecently, the common marmoset (Callithrix jacchus) has attracted significant interest as a non-human primate stroke model. Functional impairment in non-human primate stroke models should be evaluated quantitatively and successively after stroke, but conventional observational assessments of behavior cannot fully fit this purpose. In this paper, we report a behavioral analysis using MarmoDetector, a three-dimensional motion analysis, in an ischemic stroke model using photosensitive dye, along with an observational behavioral assessment and imaging examination.MethodsIschemic stroke was induced in the left hemisphere of three marmosets. Cerebral infarction was induced by intravenous injection of rose bengal and irradiation with green light. The following day, the success of the procedure was confirmed by magnetic resonance imaging (MRI). The distance traveled, speed, activity time, and jumps/climbs were observed for 28 days after stroke using MarmoDetector. We also assessed the marmosets’ specific movements and postural abnormalities using conventional neurological scores.ResultsMagnetic resonance imaging diffusion-weighted and T2-weighted images showed hyperintense signals, indicating cerebral infarction in all three marmosets. MarmoDetector data showed that the both indices immediately after stroke onset and gradually improved over weeks. Neurological scores were the worst immediately after stroke and did not recover to pre-infarction levels during the observation period (28 days). A significant correlation was observed between MarmoDetector data and conventional neurological scores.ConclusionIn this study, we showed that MarmoDetector can quantitatively evaluate behavioral changes in the acute to subacute phases stroke models. This technique can be practical for research on the pathophysiology of ischemic stroke and for the development of new therapeutic methods.

A clinically relevant model of focal embolic cerebral ischemia by thrombus and thrombolysis in rhesus monkeys.

Over decades of research into the treatment of stroke, nearly all attempts to translate experimental treatments from discovery in cells and rodents to use in humans have failed. The prevailing belief is that it might be necessary to pretest pharmacological neuroprotection in higher-order brains, especially those of nonhuman primates (NHPs). Over the past few years, chemical thrombolysis and mechanical thrombectomy have been established as the standard of care for ischemic stroke in patients. The spotlight is now shifting towards emphasizing both focal ischemia and subsequent reperfusion in developing a clinically relevant stroke model in NHPs. This protocol describes an embolic model of middle cerebral artery occlusion in adult rhesus monkeys. An autologous clot is combined with a microcatheter or microwire through endovascular procedures, and reperfusion is achieved through local intra-artery thrombolysis with tissue plasminogen activator. These NHP models formed relatively stable infarct sizes, delivered predictable reperfusion and survival outcomes, and recapitulated key characteristics of patients with ischemic stroke as observed on MRI images and behavioral assays. Importantly, treated animals could survive 30 d after the surgery for post-stroke neurologic deficit analyses. Thus far, this model has been used in several translational studies. Here we describe in detail the teamwork necessary for developing stroke models of NHPs, including the preoperation preparations, endovascular surgery, postoperation management and histopathological analysis. The model can be established by the following procedures over a 45-d period, including preparation steps (14 d), endovascular operation (1 d) and evaluation steps (30 d).

Adenosine A1R/A3R (Adenosine A1 and A3 Receptor) Agonist AST-004 Reduces Brain Infarction in a Nonhuman Primate Model of Stroke.

Treatment with A1R/A3R (adenosine A1 and A3 receptor) agonists in rodent models of acute ischemic stroke results in significantly reduced lesion volume, indicating activation of adenosine A1R or A3R is cerebroprotective. However, dosing and timing required for cerebroprotection has yet to be established, and whether adenosine A1R/A3R activation will lead to cerebroprotection in a gyrencephalic species has yet to be determined. The current study used clinical study intervention timelines in a nonhuman primate model of transient, 4-hour middle cerebral artery occlusion to investigate a potential cerebroprotective effect of the dual adenosine A1R/A3R agonist AST-004. Bolus and then 22 hours intravenous infusion of AST-004 was initiated 2 hours after transient middle cerebral artery occlusion. Primary outcome measures included lesion volume, lesion growth kinetics, penumbra volume as well as initial pharmacokinetic-pharmacodynamic relationships measured up to 5 days after transient middle cerebral artery occlusion. Secondary outcome measures included physiological parameters and neurological function. Administration of AST-004 resulted in rapid and statistically significant decreases in lesion growth rate and total lesion volume. In addition, penumbra volume decline over time was significantly less under AST-004 treatment compared with vehicle treatment. These changes correlated with unbound AST-004 concentrations in the plasma and cerebrospinal fluid as well as estimated brain A1R and A3R occupancy. No relevant changes in physiological parameters were observed during AST-004 treatment. These findings suggest that administration of AST-004 and combined A1R/A3R agonism in the brain are efficacious pharmacological interventions in acute ischemic stroke and warrant further clinical evaluation.

CCL2 (C-C Motif Chemokine Ligand 2) Biomarker Responses in Central Versus Peripheral Compartments After Focal Cerebral Ischemia.

Inflammatory mediators in blood have been proposed as potential biomarkers in stroke. However, a direct relationship between these circulating factors and brain-specific ischemic injury remains to be fully defined. An unbiased screen in a nonhuman primate model of stroke was used to find out the most responsive circulating biomarker flowing ischemic stroke. Then this phenomenon was checked in human beings and mice. Finally, we observed the temporospatial responsive characteristics of this biomarker after ischemic brain injury in mice to evaluate the direct relationship between this circulating factor and central nervous system–specific ischemic injury. In a nonhuman primate model, an unbiased screen revealed CCL2 (C-C motif chemokine ligand 2) as a major response factor in plasma after stroke. In mouse models of focal cerebral ischemia, plasma levels of CCL2 showed a transient response, that is, rapidly elevated by 2 to 3 hours postischemia but then renormalized back to baseline levels by 24 hours. However, a different CCL2 temporal profile was observed in whole brain homogenate, cerebrospinal fluid, and isolated brain microvessels, with a progressive increase over 24 hours, demonstrating a mismatch between brain versus plasma responses. In contrast to the lack of correlation with central nervous system responses, 2 peripheral compartments showed transient profiles that matched circulating plasma signatures. CCL2 protein in lymph nodes and adipose tissue was significantly increased at 2 hours and renormalized by 24 hours. These findings may provide a cautionary tale for biomarker pursuits in plasma. Besides a direct central nervous system response, peripheral organs may also contribute to blood signatures in complex and indirect ways.

Robotic Assessment of Upper Limb Function in a Nonhuman Primate Model of Chronic Stroke.

Stroke is a leading cause of death and disability worldwide and survivors are frequently left with long-term disabilities that diminish their autonomy and result in the need for chronic care. There is an urgent need for the development of therapies that improve stroke recovery, as well as accurate and quantitative tools to measure function. Nonhuman primates closely resemble humans in neuroanatomy and upper limb function and may be crucial in randomized pre-clinical trials for testing the efficacy of stroke therapies. To test the feasibility of robotic assessment of motor function in a NHP model of stroke, two cynomolgus macaques were trained to perform a visually guided reaching task and were also assessed in a passive stretch task using the Kinarm robot. Strokes were then induced in these animals by transiently occluding the middle cerebral artery, and their motor performance on the same tasks was assessed after recovery. Relative to pre-stroke performance, post-stroke hand movements of the affected limb became slower and less accurate. Regression analyses revealed both recovered and compensatory movements to complete movements in different spatial directions. Lastly, we noted decreased range of motion in the elbow joint of the affected limb post-stroke associated with spasticity during passive stretch. Taken together, these studies highlight that sensorimotor deficits in reaching movements following stroke in cynomolgus macaques resemble those in human patients and validate the use of robotic assessment tools in a nonhuman primate model of stroke for identifying and characterizing such deficits.

Assessment of Hand Motor Function in a Non-human Primate Model of Ischemic Stroke.

Ischemic stroke results from arterial occlusion and can cause irreversible brain injury. A non-human primate (NHP) model of ischemic stroke was previously developed to investigate its pathophysiology and for efficacy testing of therapeutic candidates; however, fine motor impairment remains to be well-characterized. We evaluated hand motor function in a cynomolgus monkey model of ischemic stroke. Endovascular transient middle cerebral artery occlusion (MCAO) with an angiographic microcatheter induced cerebral infarction. In vivo magnetic resonance imaging mapped and measured the ischemia-induced infarct lesion. In vivo diffusion tensor imaging (DTI) of the stroke lesion to assess the neuroplastic changes and fiber tractography demonstrated three-dimensional patterns in the corticospinal tract 12 weeks after MCAO. The hand dexterity task (HDT) was used to evaluate fine motor movement of upper extremity digits. The HDT was modified for a home cage-based training system, instead of conventional chair restraint training. The lesion was localized in the middle cerebral artery territory, including the sensorimotor cortex. Maximum infarct volume was exhibited over the first week after MCAO, which progressively inhibited ischemic core expansion, manifested by enhanced functional recovery of the affected hand over 12 weeks after MCAO. The total performance time decreased with increasing success rate for both hands on the HDT. Compensatory strategies and retrieval failure improved in the chronic phase after stroke. Our findings demonstrate the recovery of fine motor skill after stroke, and outline the behavioral characteristics and features of functional disorder of NHP stroke model, providing a basis for assessing hand motor function after stroke.

Reperfusion plus Selective Intra-arterial Cooling (SI-AC) Improve Recovery in a Nonhuman Primate Model of Stroke.

Early reperfusion is increasingly prioritized in ischemic stroke care, but outcomes remain suboptimal. Therefore, there is an urgent need to find neuroprotective approaches that can be combined with reperfusion to maximize efficacy. Here, the neuroprotective mechanisms behind therapeutic hypothermia were evaluated in a monkey model of ischemic stroke. Focal ischemia was induced in adult rhesus monkeys by placing autologous clots in the middle cerebral artery. Monkeys were treated with tissue plasminogen activator (t-PA) alone or t-PA plus selective intra-arterial cooling (SI-AC). Serial MRI scans and functional deficit were evaluated after ischemia. Histopathology and immunohistochemistry analysis were performed after the final MRI scan. t-PA plus SI-AC treatment led to a higher rate of MRI tissue rescue, and significantly improved neurologic deficits and daily activity scores compared with t-PA alone. In peri-infarct areas, higher fractional anisotropy values and greater fiber numbers were observed in models receiving t-PA plus SI-AC. Histological findings indicated that myelin damage, spheroids, and spongiosis were significantly ameliorated in models receiving SI-AC treatment. White matter integrity was also improved by SI-AC based on immunochemical staining. Our study demonstrates that SI-AC can be effectively combined with t-PA to improve both structural and functional recovery in a monkey model of focal ischemia. These findings provide proof-of-concept that it may be feasible to add neuroprotective agents as adjunctive treatments to reperfusion therapy for stroke.

A non-human primate model of stroke reproducing endovascular thrombectomy and allowing long-term imaging and neurological read-outs

Stroke is a devastating disease. Endovascular mechanical thrombectomy is dramatically changing the management of acute ischemic stroke, raising new challenges regarding brain outcome and opening up new avenues for brain protection. In this context, relevant experiment models are required for testing new therapies and addressing important questions about infarct progression despite successful recanalization, reversibility of ischemic lesions, blood–brain barrier disruption and reperfusion damage. Here, we developed a minimally invasive non-human primate model of cerebral ischemia (Macaca fascicularis) based on an endovascular transient occlusion and recanalization of the middle cerebral artery (MCA). We evaluated per-occlusion and post-recanalization impairment on PET-MRI, in addition to acute and chronic neuro-functional assessment. Voxel-based analyses between per-occlusion PET-MRI and day-7 MRI showed two different patterns of lesion evolution: “symptomatic salvaged tissue” (SST) and “asymptomatic infarcted tissue” (AIT). Extended SST was present in all cases. AIT, remote from the area at risk, represented 45% of the final lesion. This model also expresses both worsening of fine motor skills and dysexecutive behavior over the chronic post-stroke period, a result in agreement with cortical-subcortical lesions. We thus fully characterized an original translational model of ischemia–reperfusion damage after stroke, with consistent ischemia time, and thrombus retrieval for effective recanalization.

Abstract 179: Significant Efficacy of Adenosine A3 Receptor Agonist AST-004 in Rat and Non-human Primate Models of Transient Middle Cerebral Artery Occlusion

Acute Ischemic Stroke (AIS) is the second-leading cause of global mortality with an estimated 6.7 million victims dying each year. Our research has focused on a non-neuronal cell type, the astrocyte, and has demonstrated that GPCR receptor-mediated maintenance of astrocyte function enhances critical homeostatic mechanisms in the brain including protection against AIS-associated edema, glutamate excitotoxicity and oxidative stress. AST-004 is a small-molecule agonist of the GPCR Adenosine A3 receptor (ADORA3), demonstrating excellent pharmacokinetics and blood-brain barrier permeability in preclinical species. The efficacy and dose-response of AST-004 was assessed in rat and non-human primate stroke models of transient occlusion of the middle cerebral artery (tMCAO). In rats, a 1.5-hour tMCAO was conducted with a 24-hour steady-state intravenous infusion of vehicle or AST-004 initiated at time of reperfusion (vehicle or three AST-004 dose levels, n=12/group); neurological deficit and stroke lesion volume (TTC staining/image analysis) were assessed 24 hours post-reperfusion. Compared to vehicle, AST-004 reduced brain lesion volume by 50% (p=0.04) and improved neurological function by 36% (p=0.03). In non-human primates (cynomolgus macaque), a 4-hour tMCAO was conducted with a 22-hour steady-state intravenous infusion initiated 2 hours prior to reperfusion (vehicle or three AST-004 dose levels, n=4/group). Stroke lesion, perfusion deficits and penumbral endpoints were assessed by FLAIR, T2, MRA, DWI and ASL at multiple timepoints out to 5 days post-occlusion. Neurological deficit was assessed at 5-days post-occlusion. AST-004 treatment resulted in statistically significant improvements in outcome relative to vehicle, up to a 44% decrease in lesion volume (p=0.02) and a 72% decrease in percentage of lesion growth (p=0.0006) over the 5-day study period. AST-004 treatment during occlusion also reduced the slope of lesion evolution prior to reperfusion by 72% (p=0.05) relative to vehicle. Together, these studies indicate the potential of a non-neuronal approach to reducing stroke lesion damage via the ADORA3 receptor. AST-004 is a first-in-class candidate that warrants further evaluation in human clinical stroke trials.

Poly-Arginine Peptide-18 (R18) Reduces Brain Injury and Improves Functional Outcomes in a Nonhuman Primate Stroke Model.

Poly-arginine peptide-18 (R18) is neuroprotective in different rodent middle cerebral artery occlusion (MCAO) stroke models. In this study, we examined whether R18 treatment could reduce ischemic brain injury and improve functional outcome in a nonhuman primate (NHP) stroke model. A stroke was induced in male cynomolgus macaques by MCAO distal to the orbitofrontal branch of the MCA through a right pterional craniotomy, using a 5-mm titanium aneurysm clip for 90min. R18 (1000nmol/kg) or saline vehicle was administered intravenously 60min after the onset of MCAO. Magnetic resonance imaging (MRI; perfusion-weighted imaging, diffusion-weighted imaging, or T2-weighted imaging) of the brain was performed 15min, 24h, and 28days post-MCAO, and neurological outcome was assessed using the NHP stroke scale (NHPSS). Experimental endpoint was 28days post-MCAO, treatments were randomized, and all procedures were performed blinded to treatment status. R18 treatment reduced infarct lesion volume by up to 65.2% and 69.7% at 24h and 28days poststroke, respectively. Based on NHPSS scores, R18-treated animals displayed reduced functional deficits. This study confirms the effectiveness of R18 in reducing the severity of ischemic brain injury and improving functional outcomes after stroke in a NHP model, and provides further support for its clinical development as a stroke neuroprotective therapeutic.

Abstract WP156: Poly-arginine Peptides R18 and R18D Improve Functional Outcomes After Endothelin-1-induced Stroke in the Sprague Dawley Rat

Aims: We have previously demonstrated that the poly-arginine peptides R18 and R18D are neuroprotective at 24 hours following intraluminal middle cerebral artery occlusion (MCAO) in the rat. Therefore, using an independent laboratory we examined the ability of R18 and R18D to improve functional outcomes after endothelin-1 (ET-1) induced MCAO in rats over 56 days post-stroke. Methods: Peptides were administered intravenously at 100, 300 or 1000 nmol/kg, 60 minutes after ET-1 induced MCAO. Multiple forelimb placing tests and horizontal ladder tests were performed post-stroke, and the neuroprotective peptide NA-1 (TAT-NR2Bpc), which is being assessed in Phase 3 clinical stroke trials, was used as a benchmark. Results: R18 (300 and 1000nmol/kg, respectively) was the most effective peptide to improve functional outcomes, followed by R18D (300 and 1000 nmol/kg, respectively), and NA-1 (300 and 100 nmol/kg, respectively). Furthermore, R18 at 300 and 1000 nmol/kg was the most effective treatment to restore pre-stroke animal body weight (post-stroke day 4 versus day 7 for saline, and days 5-7 for all other treatments), while R18 and R18D at 300 and 1000 nmol/kg, but not NA-1 also significantly reduced the number of animals requiring special care (hand feeding) 48 hours after stroke. Conclusion: In conclusion, this study confirms that R18 and R18D can improve long-term functional outcomes after stroke, and given the superior effectiveness of R18 over NA-1 in the present, as well as in our previous studies, justifies further evaluation in a non-human primate stroke model and in a clinical stroke trial.

Stroke Longitudinal Volumetric Measures Correlate with the Behavioral Score in Non-Human Primates

Stroke is the second leading cause of death worldwide. Brain imaging data from experimental rodent stroke models suggest that size and location of the ischemic lesion relate to behavioral outcome. However, such a relationship between these two variables has not been established in Non-Human Primate (NHP) models. Thus, we aimed to evaluate whether size, location, and severity of stroke following controlled Middle Cerebral Artery Occlusion (MCAO) in NHP model correlated to neurological outcome. Forty cynomolgus macaques underwent MCAO, after four mortalities, thirty-six subjects were followed up during the longitudinal study. Structural T2 scans were obtained by magnetic resonance imaging (MRI) prior to, 48 h, and 30 days post-MCAO. Neurological function was assessed with the Non-human Primate Stroke Scale (NHPSS). T2 whole lesion volume was calculated per subject. At chronic stages, remaining brain volume was computed, and the affected hemisphere parceled into 50 regions of interest (ROIs). Whole and parceled volumetric measures were analyzed in relation to the NHPSS score. The longitudinal lesion volume evaluation showed a positive correlation with the NHPSS score, whereas the remaining brain volume negatively correlated with the NHPSS. Following ROI parcellation, NHPSS outcome correlated with frontal, temporal, occipital, and middle white matter, as well as the internal capsule, and the superior temporal and middle temporal gyri, and the caudate nucleus. These results represent an important step in stroke translational research by demonstrating close similarities between the NHP stroke model and the clinical characteristics following a human stroke and illustrating significant areas that could represent targets for novel neuroprotective strategies.

Abnormally reduced primary motor cortex output is related to impaired hand function in chronic stroke.

Stroke often involves primary motor cortex (M1) and its corticospinal projections (CST). As hand function is critically dependent on these structures, its recovery is often incomplete. The neuronal substrate supporting affected hand function is not well understood but likely involves reorganized M1 and CST of the lesioned hemisphere (M1IL and CSTIL). We hypothesized that affected hand function in chronic stroke is related to structural and functional reorganization of M1IL and CSTIL. We tested 18 patients with chronic ischemic stroke involving M1 or CST. Their hand function was compared with 18 age-matched healthy subjects. M1IL thickness and CSTIL fractional anisotropy (FA) were determined with MRI and compared with measures of the other hemisphere. Transcranial magnetic stimulation (TMS) was applied to M1IL to determine its input-output function [stimulus response curve (SRC)]. The plateau of the SRC (MEPmax), inflection point, and slope parameters of the curve were extracted. Results were compared with measures in 12 age-matched healthy controls. MEPmax of M1IL was significantly smaller ( P = 0.02) in the patients, indicating reduced CSTIL motor output, and was correlated with impaired hand function ( P = 0.02). M1IL thickness ( P < 0.01) and CSTIL-FA ( P < 0.01) were reduced but did not correlate with hand function. The results indicate that employed M1IL or CSTIL structural measures do not explain the extent of impairment in hand function once M1 and CST are sufficiently functional for TMS to evoke a motor potential. Instead, impairment of hand function is best explained by the abnormally low output from M1IL. NEW & NOTEWORTHY Hand function often remains impaired after stroke. While the critical role of the primary motor cortex (M1) and its corticospinal output (CST) for hand function has been described in the nonhuman primate stroke model, their structure and function have not been systematically evaluated for patients after stroke. We report that in chronic stroke patients with injury to M1 and/or CST an abnormally reduced M1 output is related to impaired hand function.

Increased functional connectivity after stroke correlates with behavioral scores in non-human primate model

Here we characterized the functional connectivity (FC) changes occurring after a controlled MCA stroke in a primate model. We hypothesize that if FC can inform about the neural changes after a stroke in the non-human primate (NHP) stroke model, then significant FC changes after the stroke would have to correlate with the remaining behavioral capacities. Eleven cynomolgus monkeys underwent an experimental middle cerebral artery occlusion while five monkeys remained as the control group. One month later the neurological function was assessed with a set of fine motor tasks and the Nonhuman Primate Stroke Scale (NHPSS). Structural and functional connectivity analyses were done to compare both groups. Three FC changes showed significant behavioral correlations: right sensorimotor-right lateral intraparietal FC with the six-well task; left posterior intraparietal-left dorsal premotor FC with the hill task; and right visual-left primary motor FC with the NHPSS. In the three instances, stronger FC correlated with better behavioral outcome. The results show that the functional changes correlating with behavioral outcomes involved sensorimotor cortices that were not restricted to the affected hemisphere. These results show that the FC analysis in NHP stroke model is a relevant methodology suitable to inform the neural changes occurring after a stroke.

O3-2-7 - Study of a novel small molecule, LT3001, in a thromboembolic stroke model in non-human primates

LT3001 is a novel small molecule designed to have thrombolytic and free radical scavenging activities. Its potential safety and efficacy in treating acute ischemic stroke was evaluated using a thromboembolic stroke model in adult Cynomolgus macaque.