Monomeric CXCL12-Engineered Adipose-Derived Stem Cells Transplantation for the Treatment of Ischemic Stroke.

Adipose-derived stem cells (ASCs) possess therapeutic potential for ischemic brain injury, and the chemokine CXCL12 has been shown to enhance their functional properties. However, the cumulative effects of ASCs when combined with various structures of CXCL12 on ischemic stroke and its underlying molecular mechanisms remain unclear. In this study, we genetically engineered mouse adipose-derived ASCs with CXCL12 variants and transplanted them to the infarct region in a mice transient middle cerebral artery occlusion (tMCAO) model of stroke. We subsequently compared the post-ischemic stroke efficacy of ASC-mCXCL12 with ASC-dCXCL12, ASC-wtCXCL12, and unmodified ASCs. Neurobehavior recovery was assessed using modified neurological severity scores, the hanging wire test, and the elevated body swing test. Changes at the tissue level were evaluated through cresyl violet and immunofluorescent staining, while molecular level alterations were examined via Western blot and real-time PCR. The results of the modified neurological severity score and cresyl violet staining indicated that both ASC-mCXCL12 and ASC-dCXCL12 treatment enhanced neurobehavioral recovery and mitigated brain atrophy at the third and fifth weeks post-tMCAO. Additionally, we observed that ASC-mCXCL12 and ASC-dCXCL12 promoted angiogenesis and neurogenesis, accompanied by an increased expression of bFGF and VEGF in the peri-infarct area of the brain. Notably, in the third week after tMCAO, the ASC-mCXCL12 exhibited superior outcomes compared to ASC-dCXCL12. However, when treated with the CXCR4 antagonist AMD3100, the beneficial effects of ASC-mCXCL12 were reversed. The AMD3100-treated group demonstrated worsened neurological function, aggravated edema volume, and brain atrophy. This outcome is likely attributed to the interaction of monomeric CXCL12 with CXCR4, which regulates the recruitment of bFGF and VEGF. This study introduces an innovative approach to enhance the therapeutic potential of ASCs in treating ischemic stroke by genetically engineering them with the monomeric structure of CXCL12.

Low-intensity focused ultrasound stimulation promotes stroke recovery via astrocytic HMGB1 and CAMK2N1 in mice.

BACKGROUND: Low-intensity focused ultrasound stimulation (LIFUS) has been developed to enhance neurological repair and remodelling during the late acute stage of ischaemic stroke in rodents. However, the cellular and molecular mechanisms of neurological repair and remodelling after LIFUS in ischaemic stroke are unclear. METHODS: Ultrasound stimulation was treated in adult male mice 7 days after transient middle cerebral artery occlusion. Angiogenesis was measured by laser speckle imaging and histological analyses. Electromyography and fibre photometry records were used for synaptogenesis. Brain atrophy volume and neurobehaviour were assessed 0-14 days after ischaemia. iTRAQ proteomic analysis was performed to explore the differentially expressed protein. scRNA-seq was used for subcluster analysis of astrocytes. Fluorescence in situ hybridisation and Western blot detected the expression of HMGB1 and CAMK2N1. RESULTS: Optimal ultrasound stimulation increased cerebral blood flow, and improved neurobehavioural outcomes in ischaemic mice (p<0.05). iTRAQ proteomic analysis revealed that the expression of HMGB1 increased and CAMK2N1 decreased in the ipsilateral hemisphere of the brain at 14 days after focal cerebral ischaemia with ultrasound treatment (p<0.05). scRNA-seq revealed that this expression pattern belonged to a subcluster of astrocytes after LIFUS in the ischaemic brain. LIFUS upregulated HMGB1 expression, accompanied by VEGFA elevation compared with the control group (p<0.05). Inhibition of HMGB1 expression in astrocytes decreased microvessels counts and cerebral blood flow (p<0.05). LIFUS reduced CAMK2N1 expression level, accompanied by increased extracellular calcium ions and glutamatergic synapses (p<0.05). CAMK2N1 overexpression in astrocytes decreased dendritic spines, and aggravated neurobehavioural outcomes (p<0.05). CONCLUSION: Our results demonstrated that LIFUS promoted angiogenesis and synaptogenesis after focal cerebral ischaemia by upregulating HMGB1 and downregulating CAMK2N1 in a subcluster of astrocytes, suggesting that LIFUS activated specific astrocyte subcluster could be a key target for ischaemic brain therapy.

Construction and Application of a Training Program for ICU Nurses to Manage Artificial Airway Gasbags to Prevent Ventilator-Associated Pneumonia.

Purpose: To construct a training program for ICU nurses to manage artificial airway gasbags to prevent ventilator-associated pneumonia (VAP) and explore its application in ICU nurses. Methods: From January to March 2023, 17 experts were consulted using the Delphi method to construct the draft of the training program. 52 ICU nurses were selected from a third-level, first-class hospital in Henan Province. They received training using the program for 4 weeks. The training effects were evaluated by examining the theoretical knowledge, testing operation skills, and the ICU medical staff gasbag management knowledge-attitude-behavior questionnaire before and one week after the training in April 2023. Results: The coefficients of expert authority for the two round inquiries were 0.816 and 0.837, respectively. The coordination coefficient of expert opinions ranges from 0.2 to 0.3. The final training program for ICU nurses on managing artificial airway gasbags to prevent VAP was constructed and included 4 primary indicators, 25 secondary indicators, 47 tertiary indicators. After the training by the program, the scores of the ICU nurses' theoretical knowledge (before, 73.73 ± 8.54 VS after, 88.31 ± 6.29; t = 11.017, P<0.001) and technical operation (before, 75.29 ± 7.48 VS after, 86.92 ± 4.72; t = 8.986, P<0.001) were significantly increased. The total scores of the ICU nurses' knowledge-attitude-behavior on gasbag management (before, 76.67 ± 10.68 VS after, 109.04 ± 9.87; t = 19.916, P<0.001) were also significantly increased compared to the performance before training. Conclusion: The training program for ICU nurses on managing artificial airway gasbags to prevent VAP based on the Miller pyramid model is scientific and practical and can enhance the knowledge-attitude-behavior level of ICU nurses. This training program constructed in this study can be recommended and applied after further verification for the ICU nurses to manage the artificial airway gasbags in order to reducing the occurrence of the VAP in the clinical practice.

M2 Microglial Extracellular Vesicles Attenuated Blood-brain Barrier Disruption via MiR-23a-5p in Cerebral Ischemic Mice.

Protecting the integrity of the blood-brain barrier (BBB) is crucial for maintaining brain homeostasis after ischemic stroke. Previous studies showed that M2 microglial extracellular vesicles (EVs) played a neuroprotective role in cerebral ischemia. However, the role of M2 microglial EVs in maintaining BBB integrity is unclear. Therefore, we explored the mechanisms of M2 microglial EVs in regulating BBB integrity. To identify microglial EVs, we used nanoparticle tracking analysis, transmission electron microscopy, and western blot analysis. Adult male ICR mice were subjected to 90-min middle cerebral artery occlusion (MCAO), followed by the injection of PKH26-labeled M2 microglial EVs via the tail vein. After MCAO, we assessed brain infarct and edema volume, as well as modified neurological severity score. BBB integrity was measured by assessing IgG leakage. The effects of M2 microglial EVs on astrocytes and endothelial cells were also examined. To investigate the molecular mechanisms, we performed RNA sequencing, miR-23a-5p knockdown, and luciferase reporter assays. Our results showed that PKH26-labeled microglial EVs were mainly taken up by neurons and glial cells. M2 microglial EVs treatment decreased brain infarct and edema volume, modified neurological severity score, and IgG leakage, while increasing the ZO-1, occludin, and claudin-5 expression after MCAO. Knockdown of miR-23a-5p reversed these effects. RNA sequencing revealed that the TNF, MMP3 and NFκB signaling pathway involved in regulating BBB integrity. Luciferase reporter assay showed that miR-23a-5p could bind to the 3' UTR of TNF. M2 microglial EVs-derived miR-23a-5p decreased TNF, MMP3 and NFκB p65 expression in astrocytes after oxygen-glucose deprivation, thereby increasing ZO-1 and Claudin-5 expression in bEnd.3 cells. In conclusion, our findings demonstrated that M2 microglial EVs attenuated BBB disruption after cerebral ischemia by delivering miR-23a-5p, which targeted TNF and regulated MMP3 and NFκB p65 expression.

An Adhesive Bioink toward Biofabrication under Wet Conditions.

Three-dimensional (3D) bioprinting is driving significant innovations in biomedicine over recent years. Under certain scenarios such as in intraoperative bioprinting, the bioinks used should exhibit not only cyto/biocompatibility but also adhesiveness in wet conditions. Herein, an adhesive bioink composed of gelatin methacryloyl, gelatin, methacrylated hyaluronic acid, and skin secretion of Andrias davidianus is designed. The bioink exhibits favorable cohesion to allow faithful extrusion bioprinting in wet conditions, while simultaneously showing good adhesion to a variety of surfaces of different chemical properties, possibly achieved through the diverse bonds presented in the bioink formulation. As such, this bioink is able to fabricate sophisticated planar and volumetric constructs using extrusion bioprinting, where the dexterity is further enhanced using ergonomic handheld bioprinters to realize in situ bioprinting. In vitro experiments reveal that cells maintain high viability; further in vivo studies demonstrate good integration and immediate injury sealing. The characteristics of the bioink indicate its potential widespread utility in extrusion bioprinting and will likely broaden the applications of bioprinting toward situations such as in situ dressing and minimally invasive tissue regeneration.

Progranulin released from microglial lysosomes reduces neuronal ferroptosis after cerebral ischemia in mice.

The cellular redox state is essential for inhibiting ferroptosis. Progranulin (PGRN) plays an important role in maintaining the cellular redox state after ischemic brain injury. However, the effect of PGRN on ferroptosis and its underlying mechanism after cerebral ischemia remains unclear. This study assesses whether PGRN affects ferroptosis and explores its mechanism of action on ferroptosis after cerebral ischemia. We found endogenous PGRN expression in microglia increased on day 3 after ischemia. In addition, PGRN agonists chloroquine and trehalose upregulated PGRN expression, reduced brain infarct volume, and improved neurobehavioral outcomes after cerebral ischemia compared to controls (p < 0.05). Moreover, PGRN upregulation attenuated ferroptosis by decreasing malondialdehyde and increasing Gpx4, Nrf2, and Slc7a11 expression and glutathione content (p < 0.05). Furthermore, chloroquine induced microglial lysosome PGRN release, which was associated with increased neuron survival. Our results indicate that PGRN derived from microglial lysosomes effectively inhibits ferroptosis during ischemic brain injury, identifying it as a promising target for ischemic stroke therapy.

Endothelial progenitor cell transplantation attenuates synaptic loss associated with enhancing complement receptor 3-dependent microglial/macrophage phagocytosis in ischemic mice.

Endothelial progenitor cell (EPC) transplantation has therapeutic effects in cerebral ischemia. However, how EPCs modulate microglial activity remains unclear. In the study, we explored whether EPCs modulated microglial/macrophage activity and facilitated injured brain repair. Adult male mice (n = 184) underwent transient middle cerebral artery occlusion, and EPCs were transplanted into the brain immediately after ischemia. Microglial/macrophage activity and complement receptor 3 (CR3) expression were evaluated in ischemic brains and cultured microglia. CR3 agonist leukadherin-1 was administrated into mice immediately after ischemia to imitate the effects of EPCs. Synaptophysin and postsynaptic density protein 95 (PSD-95) expressions were detected in EPC- and leukadherin-1 treated mice. We found that EPC transplantation increased the number of M2 microglia/macrophage-phagocytizing apoptotic cells and CR3 expression in ischemic brains at 3 days after ischemia (p < 0.05). EPC-conditional medium or cultured EPCs increased microglial migration and phagocytosis and upregulated CR3 expression in cultured microglia under oxygen-glucose deprivation condition (p < 0.05). Leukadherin-1 reduced brain atrophy volume and neurological deficits at 14 days after ischemia (p < 0.05). Both EPC transplantation and leukadherin-1 increased synaptophysin and PSD-95 expression at 14 days after ischemia (p < 0.05). EPC transplantation promoted CR3-mediated microglial/macrophage phagocytosis and subsequently attenuated synaptic loss. Our study provided a novel therapeutic mechanism for EPCs.

Blocking C3d+/GFAP+ A1 Astrocyte Conversion with Semaglutide Attenuates Blood-Brain Barrier Disruption in Mice after Ischemic Stroke.

Astrocytes play an essential role in the modulation of blood-brain barrier function. Neurological diseases induce the transformation of astrocytes into a neurotoxic A1 phenotype, exacerbating brain injury. However, the effect of A1 astrocytes on the BBB dysfunction after stroke is unknown. Adult male ICR mice (n=97) were subjected to 90-minute transient middle cerebral artery occlusion (tMCAO). Immunohistochemical staining of A1 (C3d) and A2 (S100A10) was performed to characterize phenotypic changes in astrocytes over time after tMCAO. The glucagon-like peptide-1 receptor agonist semaglutide was intraperitoneally injected into mice to inhibit A1 astrocytes. Infarct volume, atrophy volume, neurobehavioral outcomes, and BBB permeability were evaluated. RNA-seq was adopted to explore the potential targets and signaling pathways of A1 astrocyte-induced BBB dysfunction. Astrocytic C3d expression was increased, while expression of S100A10 was decreased in the first two weeks after tMCAO, reflecting a shift in the astrocytic phenotype. Semaglutide treatment reduced the expression of CD16/32 in microglia and C3d in astrocytes after ischemic stroke (p<0.05). Ischemia-induced brain infarct volume, atrophy volume and neuroinflammation were reduced in the semaglutide-treated mice, and neurobehavioral outcomes were improved compared to control mice (p<0.05). We further demonstrated that semaglutide treatment reduced the gap formation of tight junction proteins ZO-1, claudin-5 and occludin, as well as IgG leakage three days following tMCAO (p<0.05). In vitro experiments revealed that A1 astrocyte-conditioned medium disrupted BBB integrity. RNA-seq showed that A1 astrocytes were enriched in inflammatory factors and chemokines and significantly modulated the TNF and chemokine signaling pathways, which are closely related to barrier damage. We concluded that astrocytes undergo a phenotypic shift over time after ischemic stroke. C3d+/GFAP+ astrocytes aggravate BBB disruption, suggesting that inhibiting C3d+/GFAP+ astrocyte formation represents a novel strategy for the treatment of ischemic stroke.

Targeted delivery of fat extract by platelet membrane-cloaked nanocarriers for the treatment of ischemic stroke.

BACKGROUND: Our previous studies suggest that human fat extract (FE) contains a variety of angiogenic factors and may provide an alternative treatment option for stroke. However, the therapeutic effect is largely limited due to its short half-life, and inaccurate targeting. RESULTS: Herein, we leverage the targeting abilities of platelets (PLTs) to the lesion area of stroke and Arg-Gly-Asp (RGD) peptides to the angiogenic blood vessels to develop a biomimetic nanocarrier that capable of delivering FE precisely to treat stroke. The biomimetic nanocarriers are comprised of FE-encapsulated PLGA (poly(lactic-co-glycolic acid)) core enclosed by RGD peptides decorated plasma membrane of PLTs, namely RGD-PLT@PLGA-FE. We found that RGD-PLT@PLGA-FE not only targeted damaged and inflamed blood vessels but also achieved rapid accumulation in the lesion area of ischemic brain. In addition, RGD-PLT@PLGA-FE kept a sustained release behavior of FE at the lesion site, effectively increased its half-life and promoted angiogenesis and neurogenesis with delivering neurotrophic factors including BDNF, GDNF and bFGF to the brain, that ultimately resulted in blood flow increase and neurobehavioral recovery. CONCLUSIONS: In conclusion, our study provides a new strategy to design a biomimetic system for FE delivery and it is a promising modality for stroke therapy.

M2 microglia-derived extracellular vesicles promote white matter repair and functional recovery via miR-23a-5p after cerebral ischemia in mice.

Rationale: White matter repair is critical for the cognitive and neurological functional recovery after ischemic stroke. M2 microglia are well-documented to enhance remyelination and their extracellular vesicles (EVs) mediate cellular function after brain injury. However, whether M2 microglia-derived EVs could promote white matter repair after cerebral ischemia and its underlying mechanism are largely unknown. Methods: EVs were isolated from IL-4 treated microglia (M2-EVs) and untreated microglia (M0-EVs). Adult ICR mice subjected to 90-minute transient middle cerebral artery occlusion received intravenous EVs treatment for seven consecutive days. Brain atrophy volume, neurobehavioral tests were examined within 28 days following ischemia. Immunohistochemistry, myelin transmission electron microscope and compound action potential measurement were performed to assess white matter structural remodeling, functional repair and oligodendrogenesis. The effects of M2-EVs on oligodendrocyte precursor cells (OPCs) were also examined in vitro. EVs' miRNA sequencing, specific miR-23a-5p knockdown in M2-EVs and luciferase reporter assay were used to explore the underlying mechanism. Results: M2-EVs reduced brain atrophy volume, promoted functional recovery, oligodendrogenesis and white matter repair in vivo, increased OPC proliferation, survival and differentiation in vitro. miR-23a-5p was enriched in M2-EVs and could promote OPC proliferation, survival and maturation, while knocking down miR-23a-5p in M2-EVs reversed the beneficial effects of M2-EVs both in vitro and in vivo. Luciferase reporter assay showed that miR-23a-5p directly targeted Olig3. Conclusion: Our results demonstrated that M2 microglia could communicate to OPCs through M2-EVs and promote white matter repair via miR-23a-5p possibly by directly targeting Olig3 after ischemic stroke, suggesting M2-EVs is a novel and promising therapeutic strategy for white matter repair in stroke and demyelinating disease.

Monocyte-derived SDF1 supports optic nerve regeneration and alters retinal ganglion cells' response to Pten deletion.

Although mammalian retinal ganglion cells (RGCs) normally cannot regenerate axons nor survive after optic nerve injury, this failure is partially reversed by inducing sterile inflammation in the eye. Infiltrative myeloid cells express the axogenic protein oncomodulin (Ocm) but additional, as-yet-unidentified, factors are also required. We show here that infiltrative macrophages express stromal cell­derived factor 1 (SDF1, CXCL12), which plays a central role in this regard. Among many growth factors tested in culture, only SDF1 enhances Ocm activity, an effect mediated through intracellular cyclic AMP (cAMP) elevation and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) activation. SDF1 deficiency in myeloid cells (CXCL12flx/flxLysM-Cre−/+ mice) or deletion of the SDF1 receptor CXCR4 in RGCs (intraocular AAV2-Cre in CXCR4flx/flx mice) or SDF1 antagonist AMD3100 greatly suppresses inflammation-induced regeneration and decreases RGC survival to baseline levels. Conversely, SDF1 induces optic nerve regeneration and RGC survival, and, when combined with Ocm/cAMP, SDF1 increases axon regeneration to levels similar to those induced by intraocular inflammation. In contrast to deletion of phosphatase and tensin homolog (Pten), which promotes regeneration selectively from αRGCs, SDF1 promotes regeneration from non-αRGCs and enables the latter cells to respond robustly to Pten deletion; however, SDF1 surprisingly diminishes the response of αRGCs to Pten deletion. When combined with inflammation and Pten deletion, SDF1 enables many RGCs to regenerate axons the entire length of the optic nerve. Thus, SDF1 complements the effects of Ocm in mediating inflammation-induced regeneration and enables different RGC subtypes to respond to Pten deletion.

Extracellular vesicles from adipose-derived stem cells promote microglia M2 polarization and neurological recovery in a mouse model of transient middle cerebral artery occlusion.

BACKGROUND: Adipose-derived stem cells (ADSCs) and their extracellular vesicles (EVs) have therapeutic potential in ischemic brain injury, but the underlying mechanism is poorly understood. The current study aimed to explore the contribution of miRNAs in ADSC-EVs to the treatment of cerebral ischemia. METHODS: After the intravenous injection of ADSC-EVs, therapeutic efficacy was evaluated by neurobehavioral tests and brain atrophy volume. The polarization of microglia was assessed by immunostaining and qPCR. We further performed miRNA sequencing of ADSC-EVs and analyzed the relationship between the upregulated miRNAs in ADSC-EVs and microglial polarization-related proteins using Ingenuity Pathway Analysis (IPA). RESULTS: The results showed that ADSC-EVs reduced brain atrophy volume, improved neuromotor and cognitive functions after mouse ischemic stroke. The loss of oligodendrocytes was attenuated after ADSC-EVs injection. The number of blood vessels, as well as newly proliferated endothelial cells in the peri-ischemia area were higher in the ADSC-EVs treated group than that in the PBS group. In addition, ADSC-EVs regulated the polarization of microglia, resulting in increased repair-promoting M2 phenotype and decreased pro-inflammatory M1 phenotype. Finally, STAT1 and PTEN were highlighted as two downstream targets of up-regulated miRNAs in ADSC-EVs among 85 microglia/macrophage polarization related proteins by IPA. The inhibition of STAT1 and PTEN by ADSC-EVs were confirmed in cultured microglia. CONCLUSIONS: In summary, ADSC-EVs reduced ischemic brain injury, which was associated with the regulation of microglial polarization. miRNAs in ADSC-EVs partly contributed to their function in regulating microglial polarization by targeting PTEN and STAT1.

Oligodendrocyte precursor cell transplantation promotes angiogenesis and remyelination via Wnt/ß-catenin pathway in a mouse model of middle cerebral artery occlusion.

White matter injury is a critical pathological characteristic during ischemic stroke. Oligodendrocyte precursor cells participate in white matter repairing and remodeling during ischemic brain injury. Since oligodendrocyte precursor cells could promote Wnt-dependent angiogenesis and migrate along vasculature for the myelination during the development in the central nervous system, we explore whether exogenous oligodendrocyte precursor cell transplantation promotes angiogenesis and remyelination after middle cerebral artery occlusion in mice. Here, oligodendrocyte precursor cell transplantation improved motor and cognitive function, and alleviated brain atrophy. Furthermore, oligodendrocyte precursor cell transplantation promoted functional angiogenesis, and increased myelin basic protein expression after ischemic stroke. The further study suggested that white matter repairing after oligodendrocyte precursor cell transplantation depended on angiogenesis induced by Wnt/ß-catenin signal pathway. Our results demonstrated a novel pathway that Wnt7a from oligodendrocyte precursor cells acting on endothelial ß-catenin promoted angiogenesis and improved neurobehavioral outcomes, which facilitated white matter repair and remodeling during ischemic stroke.

Oligodendrocyte Precursor Cells Transplantation Improves Stroke Recovery via Oligodendrogenesis, Neurite Growth and Synaptogenesis.

Ischemic-induced white matter injury is strongly correlated with the poor neurological outcomes in stroke patients. The transplantation of oligodendrocyte precursor cells (OPCs) is an effective candidate for enhancing re-myelination in congenitally dysmyelinated brain and spinal cord. Nevertheless, mechanisms governing the recovery of white matter and axon after OPCs transplantation are incompletely understood in ischemic stroke. In this study, OPCs were transplanted into the ischemic brain at 7 days after transient middle cerebral artery occlusion (tMCAO). We observed improved behavior recovery and reduced brain atrophy volume at 28 days after OPCs transplantation. Moreover, our results identified that myelin sheath integrity and endogenous OPCs proliferation and migration were promoted after OPCs transplantation. By contrast, AMD3100, an antagonist of C-X-C chemokine receptor type 4, eliminated the beneficial effects of OPCs transplantation on white matter integrity and endogenous oligodendrogenesis. In addition, the improvement of neurite growth and synaptogenesis after OPCs transplantation in ischemic brain or OPC co-cultured neurons, potentially through the upregulation of Netrin-1, was indicated by increased protein levels of synaptophysin and postsynaptic density protein 95. Knockdown of Deleted in Colorectal Carcinoma, a receptor of Netrin-1, prevented increased neurite growth and synaptogenesis in neurons co-cultured with OPCs. In conclusion, our studies suggested that engrafted OPCs promoted the recovery after ischemic stroke by enhancing endogenous oligodendrogenesis, neurite growth, and synaptogenesis; the last two being mediated by the Netrin-1/DCC axis.

Stroke subtype-dependent synapse elimination by reactive gliosis in mice.

The pathological role of reactive gliosis in CNS repair remains controversial. In this study, using murine ischemic and hemorrhagic stroke models, we demonstrated that microglia/macrophages and astrocytes are differentially involved in engulfing synapses in the reactive gliosis region. By specifically deleting MEGF10 and MERTK phagocytic receptors, we determined that inhibiting phagocytosis of microglia/macrophages or astrocytes in ischemic stroke improved neurobehavioral outcomes and attenuated brain damage. In hemorrhagic stroke, inhibiting phagocytosis of microglia/macrophages but not astrocytes improved neurobehavioral outcomes. Single-cell RNA sequencing revealed that phagocytosis related biological processes and pathways were downregulated in astrocytes of the hemorrhagic brain compared to the ischemic brain. Together, these findings suggest that reactive microgliosis and astrogliosis play individual roles in mediating synapse engulfment in pathologically distinct murine stroke models and preventing this process could rescue synapse loss.

Plasma from healthy donors protects blood-brain barrier integrity via FGF21 and improves the recovery in a mouse model of cerebral ischaemia.

BACKGROUND: Healthy plasma therapy reverses cognitive deficits and promotes neuroplasticity in ageing brain disease. However, whether healthy plasma therapy improve blood-brain barrier integrity after stroke remains unknown. METHODS: Here, we intravenously injected healthy female mouse plasma into adult female ischaemic stroke C57BL/6 mouse induced by 90 min transient middle cerebral artery occlusion for eight consecutive days. Infarct volume, brain atrophy and neurobehavioural tests were examined to assess the outcomes of plasma treatment. Cell apoptosis, blood-brain barrier integrity and fibroblast growth factor 21 knockout mice were used to explore the underlying mechanism. RESULTS: Plasma injection improved neurobehavioural recovery and decreased infarct volume, brain oedema and atrophy after stroke. Immunostaining showed that the number of transferase dUTP nick end labelling+/NeuN+ cells decreased in the plasma-injected group. Meanwhile, plasma injection reduced ZO-1, occluding and claudin-5 tight junction gap formation and IgG extravasation at 3 days after ischaemic stroke. Western blot results showed that the FGF21 expression increased in the plasma-injected mice. However, using FGF21 knockout mouse plasma injecting to the ischaemic wild-type mice diminished the neuroprotective effects. CONCLUSIONS: Our study demonstrated that healthy adult plasma treatment protected the structural and functional integrity of blood-brain barrier, reduced neuronal apoptosis and improved functional recovery via FGF21, opening a new avenue for ischaemic stroke therapy.

M2 microglial small extracellular vesicles reduce glial scar formation via the miR-124/STAT3 pathway after ischemic stroke in mice.

Rationale: Glial scars present a major obstacle for neuronal regeneration after stroke. Thus, approaches to promote their degradation and inhibit their formation are beneficial for stroke recovery. The interaction of microglia and astrocytes is known to be involved in glial scar formation after stroke; however, how microglia affect glial scar formation remains unclear. Methods: Mice were treated daily with M2 microglial small extracellular vesicles through tail intravenous injections from day 1 to day 7 after middle cerebral artery occlusion. Glial scar, infarct volume, neurological score were detected after ischemia. microRNA and related protein were examined in peri-infarct areas of the brain following ischemia. Results: M2 microglial small extracellular vesicles reduced glial scar formation and promoted recovery after stroke and were enriched in miR-124. Furthermore, M2 microglial small extracellular vesicle treatment decreased the expression of the astrocyte proliferation gene signal transducer and activator of transcription 3, one of the targets of miR-124, and glial fibrillary acidic protein and inhibited astrocyte proliferation both in vitro and in vivo. It also decreased Notch 1 expression and increased Sox2 expression in astrocytes, which suggested that astrocytes had transformed into neuronal progenitor cells. Finally, miR-124 knockdown in M2 microglial small extracellular vesicles blocked their effects on glial scars and stroke recovery. Conclusions: Our results showed, for the first time, that microglia regulate glial scar formation via small extracellular vesicles, indicating that M2 microglial small extracellular vesicles could represent a new therapeutic approach for stroke.

Cortical Hemodynamic Response to Multi-afferent Stimulation: an optical imaging study.

Multisensory stimulation plays an important role in the recovery of ischemic stroke. However, little is known about the interactions between neuronal activities with multi-afferent stimulations and their effects on hemodynamic responses. Optogenetics has been a useful tool in neuroscience research to unravel the mechanisms of neurovascular coupling at cell-specific level. In this study, we applied laser speckle contrast imaging (LSCI) to map the cortical hemodynamic response with high spatiotemporal resolution. The results showed that optogenetic inhibition of pyramidal neurons in sensorimotor cortex induced both local and distant increases of cerebral blood flow (CBF) with dual peaks, and the full width at half maximum (FWHM) was significantly larger than that of the CBF response to optogenetic excitation. Furthermore, optogenetic excitation of pyramidal neurons could significantly increase the local CBF response to sensory stimulation, whereas optogenetic inhibition of pyramidal neurons decreased the local CBF response at the early stage after sensory stimulation and increased the distant CBF response during the recovery period. Our work provided useful insights into the mechanisms of brain stimulation, which might help in clinical neurological applications.