Single cell RNA sequencing reveals immunomodulatory effects of stem cell factor and granulocyte colony-stimulating factor treatment in the brains of aged APP/PS1 mice.

Alzheimers disease leads to progressive neurodegeneration and dementia. Alzheimers disease primarily affects older adults with neuropathological changes including amyloid beta deposition, neuroinflammation, and neurodegeneration. We have previously demonstrated that systemic treatment with combined stem cell factor, SCF, and granulocyte colony stimulating factor, GCSF, reduces amyloid beta load, increases amyloid beta uptake by activated microglia and macrophages, reduces neuroinflammation, and restores dendrites and synapses in the brains of aged APP-PS1 mice. However, the mechanisms underlying SCF-GCSF-enhanced brain repair in aged APP-PS1 mice remain unclear. This study used a transcriptomic approach to identify potential mechanisms by which SCF-GCSF treatment modulates microglia and peripheral myeloid cells to mitigate Alzheimers disease pathology in the aged brain. After injections of SCF-GCSF for 5 consecutive days, single cell RNA sequencing was performed on CD11b positive cells isolated from the brains of 28-month-old APP-PS1 mice. The vast majority of cell clusters aligned with transcriptional profiles of microglia in various activation states. However, SCF-GCSF treatment dramatically increased a cell population showing upregulation of marker genes related to peripheral myeloid cells. Flow cytometry data also revealed an SCF-GCSF-induced increase of cerebral CD45high-CD11b positive active phagocytes. SCF-GCSF treatment robustly increased the transcription of genes implicated in immune cell activation, including gene sets that regulate inflammatory processes and cell migration. Expression of S100a8 and S100a9 were robustly enhanced following SCF-GCSF treatment in all CD11b positive cell clusters. Moreover, the topmost genes differentially expressed with SCF-GCSF treatment were largely upregulated in S100a8-S100a9 positive cells, suggesting a well-conserved transcriptional profile related to SCF-GCSF treatment in resident and peripherally derived CD11b positive immune cells. This S100a8-S100a9-associated transcriptional profile contained notable genes related to proinflammatory and antiinflammatory responses, neuroprotection, and amyloid beta plaque inhibition or clearance. Altogether, this study reveals immunomodulatory effects of SCF-GCSF treatment in the aged brain with Alzheimers disease pathology, which will guide future studies to further uncover the therapeutic mechanisms.

Activating the healing process: three-dimensional culture of stem cells in Matrigel for tissue repair.

BACKGROUND: To establish a strategy for stem cell-related tissue regeneration therapy, human gingival mesenchymal stem cells (hGMSCs) were loaded with three-dimensional (3D) bioengineered Matrigel matrix scaffolds in high-cell density microtissues to promote local tissue restoration. METHODS: The biological performance and stemness of hGMSCs under 3D culture conditions were investigated by viability and multidirectional differentiation analyses. A Sprague‒Dawley (SD) rat full-thickness buccal mucosa wound model was established, and hGMSCs/Matrigel were injected into the submucosa of the wound. Autologous stem cell proliferation and wound repair in local tissue were assessed by histomorphometry and immunohistochemical staining. RESULTS: Three-dimensional suspension culture can provide a more natural environment for extensions and contacts between hGMSCs, and the viability and adipogenic differentiation capacity of hGMSCs were significantly enhanced. An animal study showed that hGMSCs/Matrigel significantly accelerated soft tissue repair by promoting autologous stem cell proliferation and enhancing the generation of collagen fibers in local tissue. CONCLUSION: Three-dimensional cell culture with hydrogel scaffolds, such as Matrigel, can effectively improve the biological function and maintain the stemness of stem cells. The therapeutic efficacy of hGMSCs/Matrigel was confirmed, as these cells could effectively stimulate soft tissue repair to promote the healing process by activating the host microenvironment and autologous stem cells.

Hematopoietic growth factors Regulate Entry of Monocytes into the Adult Brain via Chemokine Receptor CCR5.

Monocytes are circulating macrophage precursors and are generated from bone marrow hematopoietic stem cells. In the adults, monocytes continuously replenish cerebral border-associated macrophages under a physiological condition. Monocytes also rapidly infiltrate into the brain in the settings of pathological conditions. The mechanisms of recruiting monocyte-derived macrophages into the brain under pathological conditions have been extensively studied. However, it remains unclear how monocytes enter the brain for renewal of border-associated macrophages under the physiological condition. Using both in vitro and in vivo approaches, this study reveals that the combination of two hematopoietic growth factors, stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF), complementarily and synergistically enhances adhesion of monocytes to cerebral endothelial cells in a dose dependent manner. Cysteine-cysteine chemokine receptor 5 (CCR5) in brain endothelial cells, but not cell adhesion molecules mediating neuroinflammation-related infiltration of monocyte-derived macrophages, modulates the SCF+G-CSF-enhanced monocyte-endothelial cell adhesion. Blocking CCR5 or genetically deleting CCR5 reduces monocyte-endothelial cell adhesion induced by SCF+G-CSF. SCF+G-CSF-enhanced recruitment of bone marrow-derived monocytes/macrophages in cerebral perivascular space is also reduced in adult CCR5 knockout mice. This study demonstrates the contribution of SCF and G-CSF in regulating the entry of monocytes into the adult brain to replenish perivascular macrophages.

Maxillofacial growth changes after maxillary protraction therapy in children with class III malocclusion: a dual control group retrospective study.

PURPOSE: To investigate the balance between post-treatment effect and continued nature growth after maxillary protraction treatment in patients with skeletal class III malocclusion. METHODS: 31 patients aged 8.79 ± 1.65 years with skeletal Class III malocclusion had been treated with maxillary protraction and the treatment lasted an average of 1.16 years. The average observation duration after treatment in the maxillary protraction group was 2.05 ± 0.39 years. In the control groups, a sample of 22 patients (9.64 ± 2.53 years) with untreated skeletal class III malocclusion and 24 patients (9.28 ± 0.96 years) with skeletal class I malocclusion were matched to the treatment group according to age, sex and observation period. The mean observation interval of the control groups was 2.39 ± 1.29 years in the class III group and 1.97 ± 0.49 years in the class I group. RESULTS: The active orthopedic treatment effect showed a opposite trend to the natural craniomaxillofacial growth effect after treatment in many aspects. In the observation duration of treatment group, decrease in ANB, Wits appraisal and BAr-AAr were statistically significant compared to class I control group (p < 0.001), and there was a significant increase in NA-FH (P < 0.001) which was contrary to class III control group. Treatment group presented a significant increase in Gn-Co (P < 0.01) and Co-Go (P < 0.001), except for changes in the extent of the mandibular base (Pog-Go, P = 0.149) compared to class I control group. The vertical maxillomandibular skeletal variables (Gonial; MP-SN; MP-FH; Y-axis) in treatment group decreased significantly compared to those in class III control group (P < 0.01). U1-SN and L1-MP showed a significant increase, which was similar to the class I group (P > 0.05), and overjet decreased significantly relative to both of the two control groups (P < 0.05). CONCLUSION: Maxillary protraction therapy led to stable outcomes in approximately 77.42% of children with Class III malocclusion approximately 2 years after treatment. Unfavorable skeletal changes were mainly due to the greater protrusion of the mandible but maxillary protraction did have a certain degree of postimpact on the mandibular base. Protraction therapy does not fundamentally change the mode of maxillary growth in Class III subjects except for the advancement of the maxilla. Craniomaxillofacial region tend to restabilize after treatment and lead to skeletal growth rotation and more dentoalveolar compensation.

Rab27a-mediated extracellular vesicle secretion contributes to osteogenesis in periodontal ligament-bone niche communication.

Periodontitis, an infectious and common disease worldwide, leads to the destruction of the periodontal ligament-alveolar bone complex. Within the bone metabolic niche, communication between periodontal ligament stem cells (PDLSCs) and bone marrow mesenchymal stem cells (BMMSCs) has been considered a major contributor to osteogenesis. PDLSC-derived extracellular vesicles (P-EVs) have shown great potential for bone regeneration. However, the secretion and uptake mechanisms of P-EVs remain elusive. Herein, the biogenesis of extracellular vesicles (EVs) from PDLSCs was observed using scanning and transmission electron microscopy. PDLSCs were transduced with Ras-associated protein 27a (Rab27a) siRNA (PDLSCsiRab27a) to inhibit EV secretion. The effect of P-EVs on BMMSCs was evaluated using a non-contact transwell co-culture system. We observed that Rab27a knockdown decreased EV secretion, and PDLSCsiRab27a remarkably attenuated co-culture-enhanced osteogenesis of BMMSCs. Isolated PDLSC-derived EVs enhanced osteogenic differentiation of BMMSCs in vitro and induced bone regeneration in a calvarial defect model in vivo. PDLSC-derived EVs were rapidly endocytosed by BMMSCs via the lipid raft/cholesterol endocytosis pathway and triggered the phosphorylation of extracellular signal-regulated kinase 1/2. In conclusion, PDLSCs contribute to the osteogenesis of BMMSCs through Rab27a-mediated EV secretion, thereby providing a potential cell-free approach for bone regeneration.

Stem Cell Factor and Granulocyte Colony-Stimulating Factor Promote Remyelination in the Chronic Phase of Severe Traumatic Brain Injury.

Severe traumatic brain injury (TBI) causes long-term disability and death in young adults. White matter is vulnerable to TBI damage. Demyelination is a major pathological change of white matter injury after TBI. Demyelination, which is characterized by myelin sheath disruption and oligodendrocyte cell death, leads to long-term neurological function deficits. Stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) treatments have shown neuroprotective and neurorestorative effects in the subacute and chronic phases of experimental TBI. Our previous study has revealed that combined SCF and G-CSF treatment (SCF + G-CSF) enhances myelin repair in the chronic phase of TBI. However, the long-term effect and mechanism of SCF + G-CSF-enhanced myelin repair remain unclear. In this study, we uncovered persistent and progressive myelin loss in the chronic phase of severe TBI. SCF + G-CSF treatment in the chronic phase of severe TBI enhanced remyelination in the ipsilateral external capsule and striatum. The SCF + G-CSF-enhanced myelin repair is positively correlated with the proliferation of oligodendrocyte progenitor cells in the subventricular zone. These findings reveal the therapeutic potential of SCF + G-CSF in myelin repair in the chronic phase of severe TBI and shed light on the mechanism underlying SCF + G-CSF-enhanced remyelination in chronic TBI.

Stem cell factor and granulocyte colony-stimulating factor promote remyelination in the chronic phase of severe traumatic brain injury.

Severe traumatic brain injury (TBI) causes long-term disability and death in young adults. White matter is vulnerable to TBI damage. Demyelination is a major pathological change of white matter injury after TBI. Demyelination which is characterized by myelin sheath disruption and oligodendrocyte cell death leads to long-term neurological function deficits. Stem cell factor (SCF) and granulocyte colonyâ€"stimulating factor (G-CSF) treatments have shown neuroprotective and neurorestorative effects in the subacute and chronic phases of experimental TBI. Our previous study has revealed that combined SCF and G-CSF treatment (SCF+G-CSF) enhances myelin repair in the chronic phase of TBI. However, the long-term effect and mechanism of SCF+G-CSF-enhanced myelin repair remain unclear. In this study, we uncovered persistent and progressive myelin loss in the chronic phase of severe TBI. SCF+G-CSF treatment in the chronic phase of severe TBI enhanced remyelination in the ipsilateral external capsule and striatum. The SCF+G-CSF-enhanced myelin repair is positively correlated with the proliferation of oligodendrocyte progenitor cells in the subventricular zone. These findings reveal the therapeutic potential of SCF+G-CSF in myelin repair in the chronic phase of severe TBI and shed light on the mechanism underlying SCF+G-CSF-enhanced remyelination in chronic TBI.

Periodontal Ligament Stem Cell-Derived Small Extracellular Vesicles Embedded in Matrigel Enhance Bone Repair Through the Adenosine Receptor Signaling Pathway.

PURPOSE: Small extracellular vesicles (sEVs) are natural biocarriers for biomolecule transfer between cells and promising therapeutic strategies for bone defect repair. In this study, human periodontal ligament stem cell (PDLSC)-derived sEVs (P-EVs) were immobilized in Matrigel to establish a topical cell-free transplantation strategy for bone repair. METHODS: PDLSCs were cultured and P-EVs were isolated from the culture supernatant. In a rat bilateral calvarial defect model, P-EV/Matrigel was plugged into one defect and PBS/Matrigel was applied to the other. Bone repair in vivo was assessed by micro-computed tomography, histomorphometry, and immunohistochemical staining. In vitro, we investigated the effects of P-EVs on the proliferation and migration capabilities of bone marrow mesenchymal stem cells (BMMSCs) and explored the potential mechanism of action. RESULTS: The in vivo study showed that P-EV/Matrigel accelerated bone tissue repair by increasing cell infiltration when compared with the control. In vitro, P-EVs enhanced proliferation and migration of BMMSCs via increased phosphorylation of AKT and extracellular signal-regulated kinase 1/2 (ERK1/2). The role of P-EV-induced adenosine receptor signaling in AKT and ERK1/2 phosphorylation was a key mediator during enhanced BMMSC migration. CONCLUSION: These results are the first to demonstrate that P-EVs accelerated the repair of bone defects, partially through promoting cell proliferation and migration. P-EV/Matrigel, which combines topical EV-implantation and extracellular matrix scaffolds, provides a new cell-free strategy for bone tissue repair.

SCF + G-CSF treatment in the chronic phase of severe TBI enhances axonal sprouting in the spinal cord and synaptic pruning in the hippocampus.

Traumatic brain injury (TBI) is a major cause of long-term disability in young adults. An evidence-based treatment for TBI recovery, especially in the chronic phase, is not yet available. Using a severe TBI mouse model, we demonstrate that the neurorestorative efficacy of repeated treatments with stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF + G-CSF) in the chronic phase is superior to SCF + G-CSF single treatment. SCF + G-CSF treatment initiated at 3 months post-TBI enhances contralesional corticospinal tract sprouting into the denervated side of the cervical spinal cord and re-balances the TBI-induced overgrown synapses in the hippocampus by enhancing microglial function of synaptic pruning. These neurorestorative changes are associated with SCF + G-CSF-improved somatosensory-motor function and spatial learning. In the chronic phase of TBI, severe TBI-caused microglial degeneration in the cortex and hippocampus is ameliorated by SCF + G-CSF treatment. These findings reveal the therapeutic potential and possible mechanism of SCF + G-CSF treatment in brain repair during the chronic phase of severe TBI.

Brain-derived CCR5 Contributes to Neuroprotection and Brain Repair after Experimental Stroke.

Chemokine (C-C motif) receptor 5 (CCR5) is expressed not only in the immune cells but also in cerebral cells such as neurons, glia, and vascular cells. Stroke triggers high expression of CCR5 in the brain. However, the role of CCR5 in stroke remains unclear. In this study, using bone marrow chimeras we have determined the involvement of brain-derived or bone marrow-derived CCR5 in neuroprotection and brain repair after experimental stroke. CCR5-/- mice that received either wild-type (WT) or CCR5-/- bone marrow transplantation showed larger infarction sizes than the WT mice that received either WT or CCR5-/- bone marrow transplantation in both the acute (48h) and subacute (2 months) phases after cerebral cortical ischemia, suggesting that the lack of CCR5 in the brain leads to severe brain damage after stroke. However, the lack of CCR5 in the bone marrow-derived cells did not affect infarction size. The impairments of somatosensory-motor function and motor coordination were exacerbated in the mice lacking CCR5 in the brain. At 2 months post-stroke, increased degenerative neurons, decreased dendrites and synapses, decreased Iba1+ microglia/ macrophages, reduced myelination and CNPase+ oligodendrocytes in the peri-infarct cortex were observed in the mice lacking CCR5 in the brain. These pathological changes are significantly correlated with the increased infarction size and exacerbated neurological deficits. These data suggest that brain-derived CCR5 plays a key role in neuroprotection and brain repair in the subacute phase of stroke. This study reveals a novel role of CCR5 in stroke, which sheds new light on post-stroke pathomechanism.

Reparative Effects of Stem Cell Factor and Granulocyte Colony-Stimulating Factor in Aged APP/PS1 Mice.

Alzheimer's disease (AD), characterized by the accumulation of ß-amyloid (Aß) plaques and tau neurofibrillary tangles in the brain, neuroinflammation and neurodegeneration, is the most common form of neurodegenerative disease among the elderly. No effective treatment is available now in restricting the pathological progression of AD. The aim of this study is to determine the therapeutic efficacy of stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF+G-CSF) in aged APPswe/PS1dE9 (APP/PS1) mice. SCF+G-CSF was subcutaneously injected for 12 days to 25-month-old male APP/PS1 mice. We observed that SCF+G-CSF treatment reduced the Aß plaques in both the cortex and hippocampus. SCF+G-CSF treatment increased the association of TREM2+/Iba1+ cells with Aß plaques and enhanced Aß uptake by Iba1+ and CD68+cells in the brains of aged APP/PS1 mice. Importantly, cerebral expression area of P2RY12+and TMEM119+ homeostatic microglia and the branches of P2RY12+ homeostatic microglia were increased in the SCF+G-CSF-treated aged APP/PS1 mice. SCF+G-CSF treatment also decreased NOS-2 and increased IL-4 in the brains of aged APP/PS1 mice. Moreover, the loss of MAP2+dendrites and PSD-95+post-synapses and the accumulation of aggregated tau in the brains of aged APP/PS1 mice were ameliorated by SCF+G-CSF treatment. Furthermore, the density of P2RY12+ microglia was negatively correlated with Aß deposits, but positively correlated with the densities of MAP2+ dendrites and PSD-95+ puncta in the brains of aged APP/PS1 mice. These findings reveal the therapeutic potential of SCF+G-CSF treatment in ameliorating AD pathology at the late stage.

A simvastatin-releasing scaffold with periodontal ligament stem cell sheets for periodontal regeneration.

Simvastatin (SIM) has been documented to induce the osteogenic differentiation of periodontal ligament stem cells (PDLSCs). To establish an efficient release system for periodontal regeneration, a polycaprolactone (PCL) membrane scaffold containing SIM was electrospun and evaluated. The obtained PCL-SIM membrane scaffold showed sustained release up to 28 days, without deleterious effect on proliferation of PDLSCs on the scaffolds. PDLSCs were seeded onto scaffolds and their osteogenic differentiation was evaluated. After 21 days, expressions of collagen type I, alkaline phosphatase and bone sialoprotein genes were significantly upregulated and mineralized matrix formation was increased on the PCL-SIM scaffolds compared with the PCL scaffolds. In a heterotopic periodontal regeneration model, a cell sheet-scaffold construct was assembled by placement of multilayers of PDLSC sheets on PCL or PCL-SIM scaffolds, and these were then placed between dentin and ceramic bovine bone for subcutaneous implantation in athymic mice. After 8 weeks, the PCL-SIM membrane showed formation of significantly more ectopic cementum-like mineral on the dentin surface. These findings demonstrated that the PCL-SIM membrane scaffold promotes cementum-like tissue formation by sustained drug release, suggesting the feasibility of its therapeutic use with PDLSC sheets to improve periodontal regeneration.

The contribution of stem cell factor and granulocyte colony-stimulating factor in reducing neurodegeneration and promoting neurostructure network reorganization after traumatic brain injury.

Traumatic brain injury (TBI) is a major cause of death and disability in young adults worldwide. TBI-induced long-term cognitive deficits represent a growing clinical problem. Stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) are involved in neuroprotection and neuronal plasticity. However, the knowledge concerning reparative efficacy of SCF + G-CSF treatment in post-acute TBI recovery remains incomplete. This study aims to determine the efficacy of SCF + G-CSF on post-acute TBI recovery in young adult mice. The controlled cortical impact model of TBI was used for inducing a severe damage in the motor cortex of the right hemisphere in 8-week-old male C57BL mice. SCF + G-CSF treatment was initiated 3 weeks after induction of TBI. Severe TBI led to persistent motor functional deficits (Rota-Rod test) and impaired spatial learning function (water maze test). SCF + G-CSF treatment significantly improved the severe TBI-impaired spatial learning function 6 weeks after treatment. TBI also caused significant increases of Fluoro-Jade C positive degenerating neurons in bilateral frontal cortex, striatum and hippocampus, and significant reductions in MAP2+ apical dendrites and overgrowth of SMI312+ axons in peri-TBI cavity frontal cortex and in the ipsilateral hippocampal CA1 at 24 weeks post-TBI. SCF + G-CSF treatment significantly reduced TBI-induced neurodegeneration in the contralateral frontal cortex and hippocampal CA1, increased MAP2+ apical dendrites in the peri-TBI cavity frontal cortex, and prevented TBI-induced axonal overgrowth in both the peri-TBI cavity frontal cortex and ipsilateral hippocampal CA1.These findings reveal a novel pathology of axonal overgrowth after severe TBI and demonstrate a therapeutic potential of SCF + G-CSF in ameliorating severe TBI-induced long-term neuronal pathology, neurostructural network malformation, and impairments in spatial learning.

Long-term beneficial effects of hematopoietic growth factors on brain repair in the chronic phase of severe traumatic brain injury.

Severe traumatic brain injury (TBI) is the major cause of long-term, even life-long disability and cognitive impairments in young adults. The lack of therapeutic approaches to improve recovery in the chronic phase of severe TBI is a big challenge to the medical research field. Using a single severe TBI model in young adult mice, this study examined the restorative efficacy of two hematopoietic growth factors, stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF), on brain repair in the chronic phase of TBI. SCF and G-CSF alone or combination (SCF + G-CSF) treatment was administered at 3 months post-TBI. Functional recovery was evaluated by neurobehavioral tests during the period of 21 weeks after treatment. Neuropathology was examined 22 weeks after treatment. We observed that severe TBI caused persistent impairments in spatial learning/memory and somatosensory-motor function, long-term and widespread neuropathology, including dendritic reduction, decrease and overgrowth of axons, over-generated excitatory synapses, and demyelination in the cortex, hippocampus and striatum. SCF, G-CSF, and SCF + G-CSF treatments ameliorated severe TBI-induced widespread neuropathology. SCF + G-CSF treatment showed superior efficacy in improving long-term functional outcome, enhancing neural plasticity, rebalancing neural structure networks disturbed by severe TBI, and promoting remyelination. These novel findings demonstrate the therapeutic potential of SCF and G-CSF in enhancing recovery in the chronic phase of severe TBI .

Stem Cell Factor in Combination With Granulocyte Colony-Stimulating Factor Protects the Brain From Capillary Thrombosis-Induced Ischemic Neuron Loss in a Mouse Model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarct and leukoencephalopathy (CADASIL) is a Notch3 mutation-induced cerebral small vessel disease, leading to recurrent ischemic stroke and vascular dementia. There is currently no treatment that can stop or delay CADASIL progression. We have demonstrated the efficacy of treatment with combined stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) (SCF+G-CSF) in reducing cerebral small vessel thrombosis in a TgNotch3R90C mouse model of CADASIL. However, it remains unknown whether SCF+G-CSF treatment protects neurons from microvascular thrombosis-induced ischemic damage. Using bone marrow transplantation to track thrombosis, we observed that capillary thrombosis was widely distributed in the cortex, striatum and hippocampus of 22-month-old TgNotch3R90C mice. However, the capillary thrombosis mainly occurred in the cortex. Neuron loss was seen in the area next to the thrombotic capillaries, and severe neuron loss was found in the areas adjacent to the thrombotic capillaries with bifurcations. SCF+G-CSF repeated treatment significantly attenuated neuron loss in the areas next to the thrombotic capillaries in the cortex of the 22-month-old TgNotch3R90C mice. Neuron loss caused by capillary thrombosis in the cerebral cortex may play a crucial role in the pathogenesis of CADASIL. SCF+G-CSF treatment ameliorates the capillary thrombosis-induced ischemic neuron loss in TgNotch3R90C mice. This study provides new insight into the understanding of CADASIL progression and therapeutic potential of SCF+G-CSF in neuroprotection under microvascular ischemia in CADASIL.

S100 Calcium-Binding Protein A9 Knockout Contributes to Neuroprotection and Functional Improvement after Traumatic Brain Injury.

S100 calcium-binding protein A9 (S100a9), a proinflammatory protein, has been shown to be involved in the development of neuroinflammatory disorders and neurodegenerative diseases. Upregulation of S100a9 in the brain during acute brain injury has been proposed to be associated with acute neuroinflammation. However, it remains unclear whether eliminating S100a9 expression will show beneficial outcomes after traumatic brain injury (TBI). Using S100a9 knockout mice, this study has demonstrated that S100a9 deletion ameliorates post-TBI anxiety, improves TBI-impaired motor and cognitive function, reduces lesion size, prevents perilesional neuron loss and neurodegeneration, diminishes neuroinflammation and TBI-induced neurogenesis, and enhances perilesional expression of neuroplasticity protein. These findings suggest that S100a9 plays a detrimental role in TBI. Genetic deletion of S100a9 enhances neuroprotection and improves functional outcome after TBI. This study sheds light on the pathological involvement of S100a9 in TBI, which would provide a new therapeutic target to minimize TBI-induced brain damage.

[Isolation and biological characteristics of exosomes derived from periodontal ligament stem cells].

PURPOSE: To isolate and identify exosomes derived from periodontal ligament stem cells (PDLSCs) collected by ultracentrifugation. METHODS: Using the limiting dilution technique, human PDLSCs were isolated and expanded. The cell culture supernatant of PDLSCs was collected. Exosomes were collected and purified with a ultracentrifugation method. Biological characteristics of exosomes derived from PDLSCs were measured by transmission electron microscopy (TEM), Western blot and nanosight tracing analysis (NTA). RESULTS: Exosomes could be successfully isolated from the supernatant of PDLSCs by a ultracentrifugation method. Under TEM, the PDLSC-derived exosomes exhibited elliptic or saucer-like shape and the central area had lower electron density than the circum area. The PDLSC-derived exosomes could express the common surface adhesion molecules CD9, CD63, CD81 and TSG101. NTA results showed that the collected exosomes had a size around (119±12.1) nm and an approximate concentration of (3.80±0.39)×108 particles/mL. CONCLUSIONS: Exosomes derived from PDLSCs can be collected by a ultracentrifugation method, which expresses common membrane proteins and morphological characteristics of exosomes.

Stem cell factor and granulocyte colony-stimulating factor promote brain repair and improve cognitive function through VEGF-A in a mouse model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarct and leukoencephalopathy (CADASIL) is a cerebral small vascular disease caused by NOTCH3 gene mutation in vascular smooth muscle cells (VSMCs), leading to ischemic stroke and vascular dementia. To date, the pathogenesis of CADASIL remains poorly understood, and there is no treatment that can slow the progression of CADASIL. Using a transgenic mouse model of CADASIL (TgNotch3R90C), this study reveals novel findings for understanding CADASIL pathogenesis that decreased cerebral vascular endothelial growth factor (VEGF/VEGF-A) is linked to reduced cerebral blood vessel density. Reduced endothelial cell (EC) proliferation and angiogenesis are seen in TgNotch3R90C mouse brain-isolated ECs. Decreased dendrites, axons, and synapses in the somatosensory and motor cortex layer 2/3 and in the hippocampal CA1, and reduced neurogenesis in both the subventricular zone and subgranular zone occur in 15-month-old TgNotch3R90C mice. These reductions in neuron structures, synapses, and neurogenesis are significantly correlated to decreased cerebral vasculature in the corresponding areas. Impaired spatial learning and memory in TgNotch3R90C mice are significantly correlated with the reduced cerebral vasculature, neuron structures, and synapses. Repeated treatment of stem cell factor and granulocyte colony-stimulating factor (SCF+G-CSF) at 9 and 10 months of age improves cognitive function, increases cerebral VEGF/VEGF-A, restores cerebral vasculature, and enhances regeneration of neuronal structures, synaptogenesis and neurogenesis in TgNotch3R90C mice. Pretreatment with Avastin, an angiogenesis inhibitor by neutralizing VEGF-A, completely eliminates the SCF+G-CSF-enhanced cognitive function, vascular and neuronal structure regeneration, synaptogenesis and neurogenesis in TgNotch3R90C mice. SCF+G-CSF-enhanced EC proliferation and angiogenesis in TgNotch3R90C mouse brain-isolated ECs are also blocked by Avastin pretreatment. These data suggest that SCF+G-CSF treatment may repair Notch3R90C mutation-damaged brain through the VEGF-A-mediated angiogenesis. This study provides novel insight into the involvement of VEGF/VEGF-A in the pathogenesis of CADASIL and sheds light on the mechanism underlying the SCF+G-CSF-enhanced brain repair in CADASIL.

Stem Cell Factor in Combination with Granulocyte Colony-Stimulating Factor reduces Cerebral Capillary Thrombosis in a Mouse Model of CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy (CADASIL) is a cerebral small vascular disease caused by NOTCH3 mutation-induced vascular smooth muscle cell (VSMC) degeneration, leading to ischemic stroke and vascular dementia. Our previous study has demonstrated that repeated treatment with a combination of stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) reduces VSMC degeneration and cerebral endothelial cell (EC) damage and improves cognitive function in a mouse model of CADASIL (TgNotch3R90C). This study aimed to determine whether cerebral thrombosis occurs in TgNotch3R90C mice and whether repeated SCF+G-CSF treatment reduces cerebral thrombosis in TgNotch3R90C mice. Using the approaches of bone marrow transplantation to track bone marrow-derived cells and confocal imaging, we observed bone marrow-derived blood cell occlusion in cerebral small vessels and capillaries (thrombosis). Most thrombosis occurred in the cerebral capillaries (93% of total occluded vessels), and the thrombosis showed an increased frequency in the regions of capillary bifurcation. Degenerated capillary ECs were seen inside and surrounding the thrombosis, and the bone marrow-derived ECs were also found next to the thrombosis. IgG extravasation was seen in and next to the areas of thrombosis. SCF+G-CSF treatment significantly reduced cerebral capillary thrombosis and IgG extravasation. These data suggest that the EC damage is associated with thrombosis and blood-brain barrier leakage in the cerebral capillaries under the CADASIL-like condition, whereas SCF+G-CSF treatment diminishes these pathological alterations. This study provides new insight into the involvement of cerebral capillary thrombosis in the development of CADASIL and potential approaches to reduce the thrombosis, which may restrict the pathological progression of CADASIL.