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

Abstract

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.

Keywords: cynomolgus macaque; ferric chloride; ischemic stroke; middle cerebral artery; nonhuman primate.

Figure 1.

Schematic diagram of the timeline of the ischemic stroke and the MCAO surgical procedure. (A) Experimental design. MRI, NHPSS, vital signs test and blood samples test were performed as indicated. (B) Procedure of cynomolgus monkey exposure of MCA and its branches under the microscope (left panel) and illustration of blood vessels (right panel). Step 1: Excise the skin and subcutaneous tissue to expose the skull. Step 2: Expose the dura mater after removing the bone window. Step 3: After cutting open the meninges, reveal the brain tissue and lateral fissure vessels. Step 4: Expose the deep blood vessels after separating the brain tissue. Step 5: Temporarily clip upstream of the MCA and wrap filter paper around the MCA M1 segment for 10 minutes. Step 6: Remove the filter paper to reveal degeneration of blood vessels and intravascular thrombosis. The white arrow indicates the artery. (C) MRA in a macaque before and 24 hours after surgery. The red arrow indicates the embolism. ICA: internal carotid artery. MCA: Middle Cerebral Artery. MCA-M1: M1 segment of the middle cerebral artery. MCA-M2: M2 segment of the middle cerebral artery. ACA: Anterior cerebral artery. A1: A1 segment of the anterior cerebral artery.

Figure 2.

The infarct volume assessed by means of MRI in MCAO cynomolgus monkeys. (A) The brain lesions (highlighted area) were detected by MRI at different time points after M1 MCAO. (B) Quantification of the infarct volume. Cortical infarct volume is calculated based on FLAIR after surgery by image J. One-way ANOVA (**P<0.01, *P<0.05, n=3). (C) A representative image showing FLAIR sequence at 7 days after FeCl₃-induced MCAO in cynomolgus monkeys. The highlighted area in white is the high signal area under the FLAIR sequence.

Figure 3.

Stable embolization and long-term neurological deficits in NHPs subjected to MCAO. (A) A representative MRA images in cynomolgus monkeys before and 1, 7 and 28 days and 3 months after MCAO. The red arrow indicates the location of induced embolization in the MCA. (B) NHPSS scores during the 3-month observation after FeCl₃-induced MCA thrombosis in cynomolgus monkeys. One-way ANOVA. (**P<0.01, *P<0.05, n=3).