sincalide and Brain-Ischemia

To explore whether local transplantation of mesenchymal stem cells (MSCs) in temporal muscle can promote collateral angiogenesis and to analyze its main mechanisms of promoting angiogenesis.. Bilateral carotid artery stenosis (BCAS) treated mice were administrated with encephalo-myo-synangiosis (EMS), and bone marrow mesenchymal stem cells (BMSCs) were transplanted into the temporal muscle near the cerebral cortex. On the 30th day after EMS, the Morris water maze, immunofluorescence, laser speckle imaging, and light sheet microscopy were performed to evaluate angiogenesis; In addition, rats with bilateral common carotid artery occlusion were also followed by EMS surgery, and BMSCs from GFP reporter rats were transplanted into the temporal muscle to observe the survival time of BMSCs. Then, the concentrated BMSC-derived conditioned medium (BMSC-CM) was used to stimulate HUVECs and BMECs for ki-67 immunocytochemistry, CCK-8, transwell and chick chorioallantoic membrane assays. Finally, the cortical tissue near the temporal muscle was extracted after EMS, and proteome profiler (angiogenesis array) as well as RT-qPCR of mRNA or miRNA was performed.. The results of the Morris water maze 30 days after BMSC transplantation in BCAS mice during the EMS operation, showed that the cognitive impairment in the BCAS + EMS + BMSC group was alleviated (P < 0.05). The results of immunofluorescence, laser speckle imaging, and light sheet microscopy showed that the number of blood vessels, blood flow and astrocytes increased in the BCAS + EMS + BMSC group (P < 0.05). The BMSCs of GFP reporter rats were applied to EMS and showed that the transplanted BMSCs could survive for up to 14 days. Then, the results of ki-67 immunocytochemistry, CCK-8 and transwell assays showed that the concentrated BMSC-CM could promote the proliferation and migration of HUVECs and BMECs (P < 0.05). Finally, the results of proteome profiler (angiogenesis array) in the cerebral cortex showed that the several pro-angiogenesis factors (such as MMP-3, MMP-9, IGFBP-2 or IGFBP-3) were notably highly expressed in MSC transplantation group compared to others.. Local MSCs transplantation together with EMS surgery can promote angiogenesis and cognitive behavior in chronic brain ischemia mice. Our study illustrated that MSC local transplantation can be the potential therapeutical option for improving EMS treatment efficiency which might be translated into clinical application.

Topics: Animals; Brain Ischemia; Ki-67 Antigen; Mesenchymal Stem Cells; Mice; Neovascularization, Pathologic; Proteome; Rats; Sincalide

Cerebral stroke is a leading cause of death and permanent disability. The current therapeutic outcome of ischemic stroke (>85% of all strokes) is very poor, thus novel therapeutic drug is urgently needed. In vitro cell model of ischemia was established by oxygen-glucose deprivation (OGD) and in vivo animal model of ischemia was established by middle cerebral artery occlusion (MCAO). The effects of Spatholobus suberctus Dunn extract (SSCE) on OGD-induced cell injury, MCAO-induced neural injury and miR-494 level were all evaluated. The possible target genes were virtually screened utilizing bioinformatics and verified by luciferase assay. Subsequently, the effects of abnormally expressed miR-494 on OGD-induced cell injury and target gene expression were determined. Additionally, whether SSCE affected target gene expression through modulation of miR-494 was studied. Finally, the effects of aberrantly expressed Sox8 on OGD-induced injury and signaling pathways were estimated. SSCE reduced OGD-induced cell injury and ameliorated MCAO-induced neuronal injury, along with down-regulation of miR-494. Then, OGD-induced cell injury was increased by miR-494 overexpression but decreased by miR-494 silence. Sox8 was a target gene of miR-494, and SSCE could up-regulate Sox8 expression via down-regulating miR-494. Afterwards, OGD-induced cell injury was proved to be increased by Sox8 inhibition but reduced by Sox8 overexpression. Finally, OGD-induced inhibition of PI3K/AKT/mTOR and MAPK pathways was further inhibited by Sox8 silence but activated by Sox8 overexpression. SSCE ameliorates ischemia-induced injury both in vitro and in vivo by miR-494-mediated modulation of Sox8, involving activations of PI3K/AKT/mTOR and MAPK pathways.

Topics: Animals; Apoptosis; Blotting, Western; Brain Ischemia; Cell Survival; Fabaceae; Male; MicroRNAs; PC12 Cells; Plant Exudates; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; Sincalide

Ischemic stroke causes extensive cellular loss that impairs brain functions, resulting in severe disabilities. No effective treatments are currently available for brain tissue regeneration. The need to develop effective therapeutic approaches for treating stroke is compelling. A tissue engineering approach employing a hydrogel carrying both cells and neurotrophic cytokines to damaged regions is an encouraging alternative for neuronal repair. However, this approach is often challenged by low in vivo cell survival rate, and low encapsulation efficiency and loss of cytokines. To address these limitations, we propose to develop a biomaterial that can form a matrix capable of improving in vivo survival of transplanted cells and reducing in vivo loss of cytokines. Here, we report that using sericin, a natural protein from silk, we have fabricated a genipin-cross-linked sericin hydrogel (GSH) with porous structure and mild swelling ratio. The GSH supports the effective attachment and growth of neurons in vitro. Strikingly, our data reveal that sericin protein is intrinsically neurotrophic and neuroprotective, promoting axon extension and branching as well as preventing primary neurons from hypoxia-induced cell death. Notably, these functions are inherited by the GSH's degradation products, which might spare a need of incorporating costly cytokines. We further demonstrate that this neurotrophic effect is dependent on the Lkb1-Nuak1 pathway, while the neuroprotective effect is realized through regulating the Bcl-2/Bax protein ratio. Importantly, when transplanted in vivo, the GSH gives a high cell survival rate and allows the cells to continuously proliferate. Together, this work unmasks the neurotrophic and neuroprotective functions for sericin and provides strong evidence justifying the GSH's suitability as a potential neuronal cell delivery vehicle for ischemic stroke repair.

Topics: Animals; Biocompatible Materials; Bombyx; Brain Ischemia; Cell Hypoxia; Cell Line; Cell Proliferation; Cell Survival; Cross-Linking Reagents; Cytokines; Glutathione Transferase; Green Fluorescent Proteins; Humans; Hydrogels; Iridoids; Mice; Mice, Inbred C57BL; Mice, Nude; Neurons; Neuroprotective Agents; Porosity; Regeneration; Sericins; Silk; Sincalide; Spectroscopy, Fourier Transform Infrared; Stroke; Tissue Engineering

We examined the effect of cholecystokinin octapeptide sulfated type (CCK-8S) on dysfunction of CA1 pyramidal neurons induced by in vitro ischemic insult in hippocampal slices of stroke-prone spontaneously hypertensive rats (SHRSP). CCK-8S shortened the time required for partial recovery from block of a population spike produced by ischemia. Furthermore, CCK-8S reduced ischemic insult-induced accumulation of K+ in extracellular space. Suppression of the K+ conductance by the CCKB receptor activation is suggested to contribute to neuroprotection by CCK-8S.

Topics: Animals; Benzodiazepinones; Brain Ischemia; Cerebrovascular Disorders; Devazepide; Electrophysiology; Hippocampus; In Vitro Techniques; Male; Neurons; Potassium; Rats; Rats, Inbred SHR; Receptors, Cholecystokinin; Sincalide