The middle cerebral artery occlusion (MCAO) model in rats closely imitates ischemic stroke and is widely used. Existing instrumental methods provide a certain level of MCAO guidance, but monitoring of the MCA-occluding intraluminal filament position and possible complications can be improved. The goal of this study was to develop a MRI-based method of simultaneous control of the filament position, blood flow in the intracranial vessels, and hemorrhagic complications. Rats were subjected to either MRI-guided MCAO (group 1, n = 51) or MCAO without MRI control (group 2, n = 38). After operation, group 1 rats were transferred into a MRI scanner for the control of the filament position and possible complications. Ninety minutes after the onset of MCAO, the filament was removed in rats of both groups and MRI control of the infarct volume and hemorrhagic complications performed. High-resolution T1- and T2-weighted imaging performed immediately after filament insertion provided visualization of the filament position, blood flow in brain arteries, and complications related to inappropriate filament insertion. It permitted replacement of wrongly positioned filaments and exclusion of animals with complications from the experiment. MRI-based MCAO guiding provided real-time intra-operational monitoring of crucial parameters determining MCAO suitability for stroke modeling, including better assessment of the operation outcomes in individual animals and significant enhancement of the model success rate. The possibility of simultaneous visualization of the filament, blood flow in the arteries, brain tissue, and hemorrhagic complications is the principal advantage of the proposed method over other instrumental methods of MCAO quality control. Graphical AbstractMRI-guided middle cerebral artery occlusion technique permits intra-operational monitoring via direct non-invasive simultaneous visualization of the filament, blood flow in the arteries, brain tissue, and hemorrhagic complications. It provides better assessment of MCAO outcomes in individual animals and significant enhancement of MCAO success rate.

Cell therapy is an emerging approach to stroke treatment with a potential to limit brain damage and enhance its restoration after the acute phase of the disease. In this study we tested directly reprogrammed neural precursor cells (drNPC) derived from adult human bone marrow cells in the rat middle cerebral artery occlusion (MCAO) model of acute ischemic stroke using human placenta mesenchymal stem cells (pMSC) as a positive control with previously confirmed efficacy. Cells were infused into the ipsilateral (right) internal carotid artery of male Wistar rats 24 h after MCAO. The main goal of this work was to evaluate real-time distribution and subsequent homing of transplanted cells in the brain. This was achieved by performing intra-arterial infusion directly inside the MRI scanner and allowed transplanted cells tracing starting from their first pass through the brain vessels. Immediately after transplantation, cells were observed in the periphery of the infarct zone and in the brain stem, 15 min later small numbers of cells could be discovered deep in the infarct core and in the contralateral hemisphere, where drNPC were seen earlier and in greater numbers than pMSC. Transplanted cells in both groups could no longer be detected in the rat brain 48–72 h after infusion. Histological and histochemical analysis demonstrated that both the drNPC and pMSC were localized inside blood vessels in close contact with the vascular wall. No passage of labeled cells through the blood brain barrier was observed. Additionally, the therapeutic effects of drNPC and pMSC were compared. Both drNPC and pMSC induced substantial attenuation of neurological deficits evaluated at the 7th and 14th day after transplantation using the modified neurological severity score (mNSS). Some of the effects of drNPC and pMSC, such as the influence on the infarct volume and the survival rate of animals, differed. The results suggest a paracrine mechanism of the positive therapeutic effects of IA drNPC and pMSC infusion, potentially enhanced by the cell-cell interactions. Our data also indicate that the long-term homing of transplanted cells in the brain is not necessary for the brain’s functional recovery.

Transplantation of various types of stem cells as a possible therapy for stroke has been tested for years, and the results are promising. Recent investigations have shown that the administration of the conditioned media obtained after stem cell cultivation can also be effective in the therapy of the central nervous system pathology (hypothesis of their paracrine action). The aim of this study was to evaluate the therapeutic effects of the conditioned medium of hiPSC-derived glial and neuronal progenitor cells in the rat middle cerebral artery occlusion model of the ischemic stroke. Secretory activity of the cultured neuronal and glial progenitor cells was evaluated by proteomic and immunosorbent-based approaches. Therapeutic effects were assessed by overall survival, neurologic deficit and infarct volume dynamics, as well as by the end-point values of the apoptosis- and inflammation-related gene expression levels, the extent of microglia/macrophage infiltration and the numbers of formed blood vessels in the affected area of the brain. As a result, 31% of the protein species discovered in glial progenitor cells-conditioned medium and 45% in neuronal progenitor cells-conditioned medium were cell type specific. The glial progenitor cell-conditioned media showed a higher content of neurotrophins (BDNF, GDNF, CNTF and NGF). We showed that intra-arterial administration of glial progenitor cells-conditioned medium promoted a faster decrease in neurological deficit compared to the control group, reduced microglia/macrophage infiltration, reduced expression of pro-apoptotic gene Bax and pro-inflammatory cytokine gene Tnf, increased expression of anti-inflammatory cytokine genes (Il4, Il10, Il13) and promoted the formation of blood vessels within the damaged area. None of these effects were exerted by the neuronal progenitor cell-conditioned media. The results indicate pronounced cytoprotective, anti-inflammatory and angiogenic properties of soluble factors secreted by glial progenitor cells.