May the force be with you: Transfer of healthy mitochondria from stem cells to stroke cells

Accumulating evidence has implicated dysfunctional oxygen consumption and metabolic activity in stroke.^1,2 We now recognize a close interaction between neuronal impairment of mitochondria, key organelles for energy production, and the onset and progression of secondary cell death in stroke. Many stroke symptoms may be the result of reduced oxygen availability and consequent energy production deficits. Accordingly, mitochondria-based regenerative medicine may represent as an appealing therapeutic approach for stroke.^3,4

Mitochondrial dysfunction in stroke leads to deficits in bioenergetics, pointing to mitochondria as a primary exacerbating factor in stroke secondary cell death. Thus, restoration of mitochondrial function may have a critical role in preventing stroke progression. Our overarching hypothesis (Figure 1(a)) entails that following the primary ischemic insult, a cascade of secondary cell death events ensues, with dysfunctional mitochondria as major exacerbating factor, resulting in impaired cellular bioenergetics of the stroke brain. Multiple downstream pathways involve cell death processes that specifically target inflammation, ⁵ which in turn may compromise the neurovascular unit (neuronal, astrocytic, pericyte, and endothelial cell death). As impaired mitochondria play a critical role in pro-inflammatory signaling, the repair of mitochondrial function by stem cell therapy may sequester inflammation and facilitate central nervous system (CNS) homeostasis. The putative therapeutic action of stem cell in stroke may involve functional repair via healthy mitochondria transfer into the ischemic brain, fostering a therapeutic vasculome ⁶ capable of dampening the deleterious inflammation mediators leading to secondary cell death. OPEN IN VIEWER

Recently, the pioneering discovery of mitochondrial transfer from healthy astrocytic cells to ischemic neuronal cells ⁷ provided a solid rationale to pursue therapeutic applications of mitochondria-based regenerative medicine in stroke. The successful extrapolation of mitochondrial transfer to a stem cell setting was recognized by transfer of healthy mitochondria from endothelial progenitor cells (EPCs) into ischemic brain endothelial cells, ⁸ as revealed by increased ATP levels and oxygen consumption coupled with enhanced angiogenesis and decreased transcellular permeability of brain endothelial cells. Transfer of EPC-derived extracellular mitochondria was further tested in oxygen-glucose deprivation (OGD) stroke model and showed increased transcellular endothelial permeability, accompanied by increased mitochondrial biogenesis, mitochondrial DNA copy number, and intracellular ATP levels, and reinstatement of endothelial tightness. ⁸ However, evidence of in vivo demonstration of mitochondrial transfer from stem cells into the ischemic brain is limited and warranted for translational guidance of this stem cell-mediated mitochondria therapy.

Here, we show evidence that stem cell therapy attenuates experimental stroke deficits in cultured neuronal cells by restoring mitochondrial function (Figure 1(b) to (f)). Neuronal mitochondrial morphology and distribution were altered following OGD exposure, with reduced number of tubular mitochondria and higher density of mitochondria in the soma relative to projections accompanying decreased cell viability (Figure 1(b) to (d)). OGD also reduced neurite outgrowth processes which were rescued by EPC co-culture (Figure 1(e)). Assessment of mitochondrial function showed decreased respiration in OGD-exposed neurons compared to controls, which was partially restored with EPC co-culture (Figure 1(f)). Next, in line with our long-standing goal of translating experimental therapeutics from the laboratory to the clinic, we discuss the safety and efficacy of stem cell therapy in restoring mitochondrial structure and function in ischemic neurons, resulting in behavioral recovery and reduction of histopathological deficits in stroke animals. We showed here that oxidative phosphorylation (OXPHOS) capacity was rescued in mitochondria isolated from the brains of stroke animals transplanted with EPCs compared to those stroke animals that received vehicle infusion (Figure 1(g)). Confocal microscopy confirmed transfer of mitochondria from EPCs to ischemic neurons in transplanted stroke animals (Figure 1(h)).

The use of Rho0 cells, devoid of functional mitochondria (Figure 1(i)), and specific pharmacologic mitochondrial inhibitors, combined with our stem cell technology for generating EPCs, as well as our established stroke paradigms of in vitro oxygen-glucose deprivation and in vivo thromboembolic animal model, allow us to directly probe the contribution of mitochondria in stroke pathology and its treatment. The concept that mitochondrial function reflects stroke disease states, as well as therapeutic readouts of neurobehavioral recovery, should impact on our understanding of cell death mechanisms and regenerative medicine. Our immediate goal is to evaluate the status of the mitochondria in stroke before and after stem cell treatments. Our long-term goal is the therapeutic application of stem cells, and the use of mitochondrial readouts as a robust biomarker in stroke patients. The demonstration that stem cell transplantation exerts therapeutic effects against stroke may similarly benefit other brain diseases characterized by mitochondrial deficits. Abnormalities in mitochondrial function have been implicated in the progression of hallmark pathological symptoms of Alzheimer’s disease and Parkinson’s disease.^9,10 Acute ischemic stroke shares some pathological features typical of mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes or MELAS; in particular, these patients display mitochondrial dysfunction that triggers a cascade of cell death events, such as oxidative stress, apoptosis, inflammation, blood–brain barrier breakdown altogether reminiscent of stroke, further supporting the notion that mitochondrial disruption plays a key role in stroke pathology. Elucidating the role of mitochondria in stroke-associated cell death processes will likely provide not only a better understanding of the disease pathology but equally offer innovative insights into mitochondria-based cerebroprotective therapies, thereby guiding both the diagnosis and the treatment of stroke. Paramount to these investigations is our pursuit to reveal how extracellular mitochondria can penetrate into the cell, which should aid in the development of novel treatment strategies designed to transfer healthy mitochondria from stem cells into ischemic cells.