A method for assessing recovery of fine motor function of the hand in a rhesus monkey model of cortical injury: an adaptation of the Fugl-Meyer Scale and Eshkol-Wachman Movement Notation.

Motor dysfunction of the upper extremity can result from stroke, cortical injury and neurological diseases and causes significant disruption of activities of daily living. While some spontaneous recovery in terms of compensatory movements does occur after injury to cortical motor areas, full recovery is rare. The distinction between complete recovery and compensatory recovery is important as the development of compensatory movements in the upper extremity may not translate into full functional use in human patients. However, current animal models of stroke do not distinguish full recovery from compensatory recovery. We have developed a Non-Human Primate Grasp Assessment Scale (GRAS) to quantify the precise recovery of composite movement, individual digit action, and finger-thumb pinch in our rhesus monkey model of cortical injury. To date, we have applied this GRAS scale to assess the recovery of fine motor function of the hand in young control and cell-therapy treated monkeys with cortical injury confined to the hand representation in the dominant primary motor cortex. We have demonstrated that with this scale we can detect and quantify significant impairments in fine motor function of the hand, the development of compensatory function during recovery and finally a return to full fine motor function of the hand in monkeys treated with a cell therapy.

Oral curcumin supplementation improves fine motor function in the middle-aged rhesus monkey.

Aged individuals experience decreased fine motor function of the hand and digits, which could result, in part, from the chronic, systemic state of inflammation that occurs with aging. Recent research for treating age-related inflammation has focused on the effects of nutraceuticals that have anti-inflammatory properties. One particular dietary polyphenol, curcumin, the principal curcuminoid of the spice turmeric, has been shown to have significant anti-inflammatory effects and there is mounting evidence that curcumin may serve to reduce systemic inflammation. Therefore, it could be useful for alleviating age-related impairments in fine motor function. To test this hypothesis we assessed the efficacy of a dietary intervention with a commercially available optimized curcumin to ameliorate or delay the effects of aging on fine motor function of the hand of rhesus monkeys. We administered oral daily doses of curcumin or a control vehicle to 11 monkeys over a 14- to 18-month period in which they completed two rounds of fine motor function testing. The monkeys receiving curcumin were significantly faster at retrieving a food reward by round 2 of testing than monkeys receiving a control vehicle. Further, the monkeys receiving curcumin demonstrated a greater degree of improvement in performance on our fine motor task by round 2 of testing than monkeys receiving a control vehicle. These findings reveal that fine motor function of the hand and digits is improved in middle-aged monkeys receiving chronic daily administration of curcumin.

Hippocampal network connections account for differences in memory performance in the middle-aged rhesus monkey.

Recent neurophysiological and functional neuroimaging studies suggest that the memory decline found with normal aging is not solely due to regional disruptions in the hippocampus, but also is brought about by alterations in the functional coupling between the hippocampus and long-distance neocortical regions. However, the anatomical basis for this functional "dyscoupling" has not been fully revealed. In this study, we applied a multimodal magnetic resonance imaging technique to noninvasively examine the large-scale anatomical and functional hippocampal network of a group of middle aged rhesus monkeys. Using diffusion spectrum imaging, we have found that monkeys with lower memory performance had weaker structural white matter connections between the hippocampus and neocortical association areas. Resting state functional imaging revealed somewhat of an opposite result. Monkeys with low memory performance displayed elevated coupling strengths in the network between the hippocampus and the neocortical areas. Taken together with recent findings, this contradictory pattern may be the result of either underlying physiological burden or abnormal neuronal decoupling due to the structural alterations, which induce a neuronal compensation mechanism for the structural loss or interference on task related neuronal activation, respectively.

Neural recovery after cortical injury: Effects of MSC derived extracellular vesicles on motor circuit remodeling in rhesus monkeys.

Reorganization of motor circuits in the cortex and corticospinal tract are thought to underlie functional recovery after cortical injury, but the mechanisms of neural plasticity that could be therapeutic targets remain unclear. Recent work from our group have shown that systemic treatment with mesenchymal stem cell derived (MSCd) extracellular vesicles (EVs) administered after cortical damage to the primary motor cortex (M1) of rhesus monkeys resulted in a robust recovery of fine motor function and reduced chronic inflammation. Here, we used immunohistochemistry for cfos, an activity-dependent intermediate early gene, to label task-related neurons in the surviving primary motor and premotor cortices, and markers of axonal and synaptic plasticity in the spinal cord. Compared to vehicle, EV treatment was associated with a greater density of cfos+ pyramidal neurons in the deep layers of M1, greater density of cfos+ inhibitory interneurons in premotor areas, and lower density of synapses on MAP2+ lower motor neurons in the cervical spinal cord. These data suggest that the anti-inflammatory effects of EVs may reduce injury-related upper motor neuron damage and hyperexcitability, as well as aberrant compensatory re-organization in the cervical spinal cord to improve motor function.

Sex differences in recovery of motor function in a rhesus monkey model of cortical injury.

BACKGROUND: Stroke disproportionately affects men and women, with women over 65 years experiencing increased severity of impairment and higher mortality rates than men. Human studies have explored risk factors that contribute to these differences, but additional research is needed to investigate how sex differences affect functional recovery and hence the severity of impairment. In the present study, we used our rhesus monkey model of cortical injury and fine motor impairment to compare sex differences in the rate and degree of motor recovery following this injury. METHODS: Aged male and female rhesus monkeys were trained on a task of fine motor function of the hand before undergoing surgery to produce a cortical lesion limited to the hand area representation of the primary motor cortex. Post-operative testing began two weeks after the surgery and continued for 12 weeks. All trials were video recorded and latency to retrieve a reward was quantitatively measured to assess the trajectory of post-operative response latency and grasp pattern compared to pre-operative levels. RESULTS: Postmortem analysis showed no differences in lesion volume between male and female monkeys. However, female monkeys returned to their pre-operative latency and grasp patterns significantly faster than males. CONCLUSIONS: These findings demonstrate the need for additional studies to further investigate the role of estrogens and other sex hormones that may differentially affect recovery outcomes in the primate brain.

Extracellular vesicles derived from bone marrow mesenchymal stem cells enhance myelin maintenance after cortical injury in aged rhesus monkeys.

Cortical injury, such as stroke, causes neurotoxic cascades that lead to rapid death and/or damage to neurons and glia. Axonal and myelin damage in particular, are critical factors that lead to neuronal dysfunction and impair recovery of function after injury. These factors can be exacerbated in the aged brain where white matter damage is prevalent. Therapies that can ameliorate myelin damage and promote repair by targeting oligodendroglia, the cells that produce and maintain myelin, may facilitate recovery after injury, especially in the aged brain where these processes are already compromised. We previously reported that a novel therapeutic, Mesenchymal Stem Cell derived extracellular vesicles (MSC-EVs), administered intravenously at both 24 h and 14 days after cortical injury, reduced microgliosis (Go et al. 2019), reduced neuronal pathology (Medalla et al. 2020), and improved motor recovery (Moore et al. 2019) in aged female rhesus monkeys. Here, we evaluated the effect of MSC-EV treatment on changes in oligodendrocyte maturation and associated myelin markers in the sublesional white matter using immunohistochemistry, confocal microscopy, stereology, qRT-PCR, and ELISA. Compared to vehicle control monkeys, EV-treated monkeys showed a reduction in the density of damaged oligodendrocytes. Further, EV-treatment was associated with enhanced myelin maintenance, evidenced by upregulation of myelin-related genes and increases in actively myelinating oligodendrocytes in sublesional white matter. These changes in myelination correlate with the rate of motor recovery, suggesting that improved myelin maintenance facilitates this recovery. Overall, our results suggest that EVs act on oligodendrocytes to support myelination and improves functional recovery after injury in the aged brain. SIGNIFICANCE: We previously reported that EVs facilitate recovery of function after cortical injury in the aged monkey brain, while also reducing neuronal pathology (Medalla et al. 2020) and microgliosis (Go et al. 2019). However, the effect of injury and EVs on oligodendrocytes and myelination has not been characterized in the primate brain (Dewar et al. 1999; Sozmen et al. 2012; Zhang et al. 2013). In the present study, we assessed changes in myelination after cortical injury in aged monkeys. Our results show, for the first time, that MSC-EVs support recovery of function after cortical injury by enhancing myelin maintenance in the aged primate brain.

Extracellular vesicles from mesenchymal stem cells reduce microglial-mediated neuroinflammation after cortical injury in aged Rhesus monkeys.

Cortical injury, such as injuries after stroke or age-related ischemic events, triggers a cascade of degeneration accompanied by inflammatory responses that mediate neurological deficits. Therapeutics that modulate such neuroinflammatory responses in the aging brain have the potential to reduce neurological dysfunction and promote recovery. Extracellular vesicles (EVs) from mesenchymal stem cells (MSCs) are lipid-bound, nanoscale vesicles that can modulate inflammation and enhance recovery in rodent stroke models. We recently assessed the efficacy of intravenous infusions of MSC-EVs (24-h and 14-days post-injury) as a treatment in aged rhesus monkeys (Macaca mulatta) with cortical injury that induced impairment of fine motor function of the hand. Aged monkeys treated with EVs after injury recovered motor function more rapidly and more fully than aged monkeys given a vehicle control. Here, we describe EV-mediated inflammatory changes using histological assays to quantify differences in markers of neuroinflammation in brain tissue between EV and vehicle-treated aged monkeys. The activation status of microglia, the innate macrophages of the brain, is critical to cell fate after injury. Our findings demonstrate that EV treatment after injury is associated with greater densities of ramified, homeostatic microglia, along with reduced pro-inflammatory microglial markers. These findings are consistent with a phenotypic switch of inflammatory hypertrophic microglia towards anti-inflammatory, homeostatic functions, which was correlated with enhanced functional recovery. Overall, our data suggest that EVs reduce neuroinflammation and shift microglia towards restorative functions. These findings demonstrate the therapeutic potential of MSC-derived EVs for reducing neuroinflammation after cortical injury in the aged brain.

Long-term effects of curcumin in the non-human primate brain.

Curcumin has recently been shown to be a potential treatment for slowing or ameloriating cognitive decline during aging in our nonhuman primate model of normal aging. In these same monkeys, we studied for the first time the neurological impacts of long-term curcumin treatments using longitudinal magnetic resonance imaging (MRI). Sixteen rhesus monkeys received curcumin or a vehicle control for 14-18 months. We applied a combination of structural and diffusion MRI to determine whether the curcumin resulted in structural or functional changes in focal regions of the brain. The longitudinal imaging revealed decreased microscale diffusivity (mD) measurements mainly in the hippocampus and basal forebrain structures of curcumin treated animals. Changes in generalized fractional anisotropy (GFA) and grey matter density (GMd) measurements indicated an increased grey matter density in cortical ROIs with improved white matter integrity in limbic, cerebellar, and brain stem regions. These findings suggest that noticeable changes in the neuronal environment could be induced from long-term curcumin treatments. Results may provide a neurological basis on the recent findings demonstrating improved spatial working memory and motor function in nonhuman primates.

Chronic curcumin treatment improves spatial working memory but not recognition memory in middle-aged rhesus monkeys.

Studies of both humans and non-human primates have demonstrated that aging is typically characterized by a decline in cognition that can occur as early as the fifth decade of life. Age-related changes in working memory are particularly evident and mediated, in part, by the prefrontal cortex, an area known to evidence age-related changes in myelin that is attributed to inflammation. In recent years, several nutraceuticals, including curcumin, by virtue of their anti-inflammatory and antioxidant effects, have received considerable attention as potential treatments for age-related cognitive decline and inflammation. Accordingly, we assessed for the first time in a non-human primate model of normal aging the efficacy of dietary intervention using the natural phenol curcumin to ameliorate the effects of aging on spatial working and recognition memory. Results revealed that monkeys receiving daily administration of curcumin over 14-18 months demonstrated a greater improvement in performance on repeated administration of a task of spatial working memory compared to monkeys that received a control substance.

Inosine enhances recovery of grasp following cortical injury to the primary motor cortex of the rhesus monkey.

BACKGROUND: Inosine, a naturally occurring purine nucleoside, has been shown to stimulate axonal growth in cell culture and promote corticospinal tract axons to sprout collateral branches after stroke, spinal cord injury and TBI in rodent models. OBJECTIVE: To explore the effects of inosine on the recovery of motor function following cortical injury in the rhesus monkey. METHODS: After being trained on a test of fine motor function of the hand, monkeys received a lesion limited to the area of the hand representation in primary motor cortex. Beginning 24 hours after this injury and continuing daily thereafter, monkeys received orally administered inosine (500 mg) or placebo. Retesting of motor function began on the 14th day after injury and continued for 12 weeks. RESULTS: During the first 14 days after surgery, there was evidence of significant recovery within the inosine-treated group on measures of fine motor function of the hand, measures of hand strength and digit flexion. While there was no effect of treatment on the time to retrieve a reward, the treated monkeys returned to asymptotic levels of grasp performance significantly faster than the untreated monkeys. Additionally, the treated monkeys evidenced a greater degree of recovery in terms of maturity of grasp pattern. CONCLUSION: These findings demonstrate that inosine can enhance recovery of function following cortical injury in monkeys.