Orexins, produced in the lateral hypothalamus, are important neuropeptides that participate in the sleep/wake cycle, and their expres- sion coincides with the projection area of the vagus nerve in the brain. Vagus ner...Orexins, produced in the lateral hypothalamus, are important neuropeptides that participate in the sleep/wake cycle, and their expres- sion coincides with the projection area of the vagus nerve in the brain. Vagus nerve stimulation has been shown to decrease the amounts of daytime sleep and rapid eye movement in epilepsy patients with traumatic brain injury. In the present study, we investigated whether vagus nerve stimulation promotes wakefulness and affects orexin expression. A rat model of traumatic brain injury was established using the free fall drop method. In the stimulated group, rats with traumatic brain injury received vagus nerve stimulation （frequency, 30 Hz, current, 1.0 mA; pulse width, 0.5 ms; total stimulation time, 15 minutes）. In the antagonist group, rats with traumatic brain injury were intracerebroventricularly injected with the orexin receptor type 1 （OXIR） antagonist SB334867 and received vagus nerve stimulation. Changes in consciousness were observed after stimulation in each group. Enzyme-linked immunosorbent assay, western blot assay and immunohistochemistry were used to assess the levels of orexin-A and OX1R expression in the prefrontal cortex. In the stimulated group, consciousness was substantially improved, orexin-A protein expression gradually increased within 24 hours after injury and OX1R expres- sion reached a peak at 12 hours, compared with rats subjected to traumatic brain injury only. In the antagonist group, the wake-promoting effect of vagus nerve stimulation was diminished, and orexin-A and OX1R expression were decreased, compared with that of the stim- ulated group. Taken together, our findings suggest that vagus nerve stimulation promotes the recovery of consciousness in comatose rats after traumatic brain injury. The upregulation of orexin-A and OXIR expression in the prefrontal cortex might be involved in the wake-promoting effects of vagus nerve stimulation.展开更多
Recent studies have shown the potential of artificially synthesized conduits in the repair of peripheral nerve injury.Natural biopolymers have received much attention because of their biocompatibility.To investigate t...Recent studies have shown the potential of artificially synthesized conduits in the repair of peripheral nerve injury.Natural biopolymers have received much attention because of their biocompatibility.To investigate the effects of novel electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits(biopolymer nanofiber conduits)on the repair of peripheral nerve injury,we bridged 10-mm-long sciatic nerve defects with electrospun absorbable biopolymer nanofiber conduits,poly(ε-caprolactone)or silicone conduits in Sprague-Dawley rats.Rat neurologica1 function was weekly evaluated using sciatic function index within 8 weeks after repair.Eight weeks after repair,sciatic nerve myelin sheaths and axon morphology were observed by osmium tetroxide staining,hematoxylin-eosin staining,and transmission electron microscopy.S-100(Schwann cell marker)and CD4(inflammatory marker)immunoreactivities in sciatic nerve were detected by immunohistochemistry.In rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits,no serious inflammatory reactions were observed in rat hind limbs,the morphology of myelin sheaths in the injured sciatic nerve was close to normal.CD4 immunoreactivity was obviously weaker in rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits than in those subjected to repair with poly(ε-caprolactone)or silicone.Rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits tended to have greater sciatic nerve function recovery than those receiving poly(ε-caprolactone)or silicone repair.These results suggest that electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits have the potential of repairing sciatic nerve defects and exhibit good biocompatibility.All experimental procedures were approved by Institutional Animal Care and Use Committee of Taichung Veteran General Hospital,Taiwan,China(La-1031218)on October 2,2014.展开更多
The calcium channel blocker,verapamil,has been shown to reduce scar formation by inhibiting fibroblast adhesion and proliferation in vitro.It was not clear whether topical application of verapamil after surgical repai...The calcium channel blocker,verapamil,has been shown to reduce scar formation by inhibiting fibroblast adhesion and proliferation in vitro.It was not clear whether topical application of verapamil after surgical repair of the nerve in vivo could inhibit the formation of excessive scar tissue.In this study,the right sciatic nerve of adult Sprague-Dawley rats was transected and sutured with No.10-0 suture.The stoma was wrapped with gelfoam soaked with verapamil solution for 4 weeks.Compared with the control group（stoma wrapped with gelfoam soaked with physiological saline）,the verapamil application inhibited the secretion of extracellular matrix from fibroblasts in vivo,suppressed type I and III collagen secretion and increased the total number of axons and the number of myelinated axons.These findings suggest that verapamil could reduce the formation of scar tissue and promote axon growth after peripheral nerve repair.展开更多
The Wnt/Frizzled signaling pathway participates in many inflammation-linked diseases. However, the inflammatory response mediated by the Wnt/Frizzled signaling pathway in experimental subarachnoid hemorrhage has not b...The Wnt/Frizzled signaling pathway participates in many inflammation-linked diseases. However, the inflammatory response mediated by the Wnt/Frizzled signaling pathway in experimental subarachnoid hemorrhage has not been thoroughly investigated. Consequently, in this study, we examined the potential role of the Wnt/Frizzled signaling pathway in early brain injury in rat models of subarachnoid hemorrhage.Simultaneously, possible neuroprotective mechanisms were also investigated. Experimental subarachnoid hemorrhage rat models were induced by injecting autologous blood into the prechiasmatic cistern. Experiment 1 was designed to examine expression of the Wnt/Frizzled signaling pathway in early brain injury induced by subarachnoid hemorrhage. In total, 42 adult rats were divided into sham(injection of equivalent volume of saline), 6-, 12-, 24-, 48-, 72-hour, and 1-week subarachnoid hemorrhage groups. Experiment 2 was designed to examine neuroprotective mechanisms of the Wnt/Frizzled signaling pathway in early brain injury induced by subarachnoid hemorrhage. Rats were treated with recombinant human Wnt1(rhwnt1), small interfering Wnt1(siwnt1) RNA, and monoclonal antibody of Frizzled1(anti-Frizzled1) at 48 hours after subarachnoid hemorrhage. Expression levels of Wnt1, Frizzled1, β-catenin, peroxisome proliferator-activated receptor-γ, CD36, and active nuclear factor-κB were examined by western blot assay and immunofluorescence staining. Microglia type conversion and inflammatory cytokine levels in brain tissue were examined by immunofluorescence staining and enzyme-linked immunosorbent assay. Our results show that compared with the sham group, expression levels of Wnt1, Frizzled1, and β-catenin were low and reduced to a minimum at 48 hours, gradually returning to baseline at 1 week after subarachnoid hemorrhage. rhwnt1 treatment markedly increased Wnt1 expression and alleviated subarachnoid hemorrhage-induced early brain injury(within 72 hours), including cortical cell apoptosis, brain edema, and neurobehavioral deficits, accompanied by increasing protein levels of β-catenin, CD36, and peroxisome proliferator-activated receptor-γ and decreasing protein levels of nuclear factor-κB. Of note, rhwnt1 promoted M2-type microglia conversion and inhibited release of inflammatory cytokines(interleukin-1β, interleukin-6, and tumor necrosis factor-α). In contrast, siwnt1 RNA and anti-Frizzled1 treatment both resulted in an opposite effect. In conclusion, the Wnt/Frizzled1 signaling pathway may participate in subarachnoid hemorrhage-induced early brain injury via inhibiting the inflammatory response, including regulating microglia type conversion and decreasing inflammatory cytokine release. The study was approved by the Animal Ethics Committee of Anhui Medical University and First Affiliated Hospital of USTC,Division of Life Sciences and Medicine, University of Science and Technology of China(approval No. LLSC-20180202) in May 2017.展开更多
The incidence of ischemic stroke in patients with diabetes is increasing. While brachial-ankle pulse wave velocity （BaPWV） and ankle-bra- chial index （ABI） are known to be associated with ischemic cardiovascular a...The incidence of ischemic stroke in patients with diabetes is increasing. While brachial-ankle pulse wave velocity （BaPWV） and ankle-bra- chial index （ABI） are known to be associated with ischemic cardiovascular and cerebrovascular diseases, whether these measures predict the risk of ischemic cerebrovascular disease in diabetic patients remains unclear. 117 patients with type 2 diabetes were enrolled in this study. According to the results of head magnetic resonance imaging, the patients were divided into a diabetes-only group （n = 55） and a diabetes and ischemic stroke group （n = 62）. We then performed ABI and BaPWV examinations for all patients. Compared with the diabe- tes-only group, we found decreased ABI and increased BaPWV in the diabetes and ischemic stroke group. Multivariate logistic regression analyses revealed that BaPWV and ABI were risk factors for ischemic stroke in patients with type 2 diabetes. Our findings indicate that decreased ABI and increased BaPWV are objective indicators of increased risk of ischemic stroke in patients with type 2 diabetes.展开更多
We previously found that the K141N mutation in heat shock protein B8 （HSPB8） was responsible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a tr...We previously found that the K141N mutation in heat shock protein B8 （HSPB8） was responsible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a transgenic mouse model bearing the K141N mutation in the human HSPB8 gene, and to determine whether this K141NHSPB8 transgenic mouse model would manifest the clinical phenotype of Charcot-Marie-Tooth disease type 2L, and consequently be suitable for use in studies of disease pathogenesis. Transgenic mice overexpressing K141N HSPB8 were generated using K141N mutant HSPB8 cDNA cloned into a pCAGGS plasmid driven by a human cytomegalovirus expression system. PCR and western blot analysis confirmed integration of the KI41NHSPB8 gene and widespread expression in tissues of the transgenic mice. The K141N HSPB8 transgenic mice exhibited decreased muscle strength in the hind limbs and impaired motor coordination, but no obvious sensory disturbance at 6 months of age by behavioral assessment. Electrophysiological analysis showed that the compound motor action potential amplitude in the sciatic nerve was significantly decreased, but motor nerve conduction velocity remained normal at 6 months of age. Pathological analysis of the sciatic nerve showed reduced myelinated fiber density, notable axonal edema and vacuolar degeneration in K141N HSPB8 transgenic mice, suggesting axonal involvement in the peripheral nerve damage in these animals. These findings indicate that the KI4mHSPB8 transgenic mouse successfully models Charcot-Marie-Tooth disease type 2L and can be used to study the pathogenesis of the disease.展开更多
The best tissue-engineered spinal cord grafts not only match the structural characteristics of the spinal cord but also allow the seed cells to grow and function in situ.Platelet-derived growth factor（PDGF） has been...The best tissue-engineered spinal cord grafts not only match the structural characteristics of the spinal cord but also allow the seed cells to grow and function in situ.Platelet-derived growth factor（PDGF） has been shown to promote the migration of bone marrow stromal cells;however,cytokines need to be released at a steady rate to maintain a stable concentration in vivo.Therefore,new methods are needed to maintain an optimal concentration of cytokines over an extended period of time to effectively promote seed cell localization,proliferation and differentiation.In the present study,a partition-type tubular scaffold matching the anatomical features of the thoracic 8–10 spinal cord of the rat was fabricated using chitosan and then subsequently loaded with chitosan-encapsulated PDGF-BB microspheres（PDGF-MSs）.The PDGF-MS-containing scaffold was then examined in vitro for sustained-release capacity,biocompatibility,and its effect on neural progenitor cells differentiated in vitro from multilineage-differentiating stress-enduring cells（MUSE-NPCs）.We found that pre-freezing for 2 hours at-20°C significantly increased the yield of partition-type tubular scaffolds,and 30 μL of 25% glutaraldehyde ensured optimal crosslinking of PDGF-MSs.The resulting PDGF-MSs cumulatively released 52% of the PDGF-BB at 4 weeks in vitro without burst release.The PDGF-MS-containing tubular scaffold showed suitable biocompatibility towards MUSE-NPCs and could promote the directional migration and growth of these cells.These findings indicate that the combination of a partition-type tubular scaffold,PDGF-MSs and MUSENPCs may be a promising model for the fabrication of tissue-engineered spinal cord grafts.展开更多
基金supported by the Natural Science Foundation of China,No.81260295the Graduate Student Innovation Fund of Jiangxi Province of China,No.YC2015-S090
文摘Orexins, produced in the lateral hypothalamus, are important neuropeptides that participate in the sleep/wake cycle, and their expres- sion coincides with the projection area of the vagus nerve in the brain. Vagus nerve stimulation has been shown to decrease the amounts of daytime sleep and rapid eye movement in epilepsy patients with traumatic brain injury. In the present study, we investigated whether vagus nerve stimulation promotes wakefulness and affects orexin expression. A rat model of traumatic brain injury was established using the free fall drop method. In the stimulated group, rats with traumatic brain injury received vagus nerve stimulation （frequency, 30 Hz, current, 1.0 mA; pulse width, 0.5 ms; total stimulation time, 15 minutes）. In the antagonist group, rats with traumatic brain injury were intracerebroventricularly injected with the orexin receptor type 1 （OXIR） antagonist SB334867 and received vagus nerve stimulation. Changes in consciousness were observed after stimulation in each group. Enzyme-linked immunosorbent assay, western blot assay and immunohistochemistry were used to assess the levels of orexin-A and OX1R expression in the prefrontal cortex. In the stimulated group, consciousness was substantially improved, orexin-A protein expression gradually increased within 24 hours after injury and OX1R expres- sion reached a peak at 12 hours, compared with rats subjected to traumatic brain injury only. In the antagonist group, the wake-promoting effect of vagus nerve stimulation was diminished, and orexin-A and OX1R expression were decreased, compared with that of the stim- ulated group. Taken together, our findings suggest that vagus nerve stimulation promotes the recovery of consciousness in comatose rats after traumatic brain injury. The upregulation of orexin-A and OXIR expression in the prefrontal cortex might be involved in the wake-promoting effects of vagus nerve stimulation.
基金supported by grants from the Taichung Veterans General Hospital and Central Taiwan University of Science and Technology,No.TCVGH-CTUST1047701(to CCS and BSL)Taichung Veterans General Hospital,No.TCVGH-1034907C(to CCS),Taiwan,China
文摘Recent studies have shown the potential of artificially synthesized conduits in the repair of peripheral nerve injury.Natural biopolymers have received much attention because of their biocompatibility.To investigate the effects of novel electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits(biopolymer nanofiber conduits)on the repair of peripheral nerve injury,we bridged 10-mm-long sciatic nerve defects with electrospun absorbable biopolymer nanofiber conduits,poly(ε-caprolactone)or silicone conduits in Sprague-Dawley rats.Rat neurologica1 function was weekly evaluated using sciatic function index within 8 weeks after repair.Eight weeks after repair,sciatic nerve myelin sheaths and axon morphology were observed by osmium tetroxide staining,hematoxylin-eosin staining,and transmission electron microscopy.S-100(Schwann cell marker)and CD4(inflammatory marker)immunoreactivities in sciatic nerve were detected by immunohistochemistry.In rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits,no serious inflammatory reactions were observed in rat hind limbs,the morphology of myelin sheaths in the injured sciatic nerve was close to normal.CD4 immunoreactivity was obviously weaker in rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits than in those subjected to repair with poly(ε-caprolactone)or silicone.Rats subjected to repair with electrospun absorbable biopolymer nanofiber conduits tended to have greater sciatic nerve function recovery than those receiving poly(ε-caprolactone)or silicone repair.These results suggest that electrospun absorbable poly(ε-caprolactone)/type I collagen nanofiber conduits have the potential of repairing sciatic nerve defects and exhibit good biocompatibility.All experimental procedures were approved by Institutional Animal Care and Use Committee of Taichung Veteran General Hospital,Taiwan,China(La-1031218)on October 2,2014.
基金supported by the Natural Science Foundation of Hubei Province of China,No.2011CDB392
文摘The calcium channel blocker,verapamil,has been shown to reduce scar formation by inhibiting fibroblast adhesion and proliferation in vitro.It was not clear whether topical application of verapamil after surgical repair of the nerve in vivo could inhibit the formation of excessive scar tissue.In this study,the right sciatic nerve of adult Sprague-Dawley rats was transected and sutured with No.10-0 suture.The stoma was wrapped with gelfoam soaked with verapamil solution for 4 weeks.Compared with the control group（stoma wrapped with gelfoam soaked with physiological saline）,the verapamil application inhibited the secretion of extracellular matrix from fibroblasts in vivo,suppressed type I and III collagen secretion and increased the total number of axons and the number of myelinated axons.These findings suggest that verapamil could reduce the formation of scar tissue and promote axon growth after peripheral nerve repair.
基金supported by the Natural Science Foundation of Anhui Province of China,No.1508085QH184(to YW)
文摘The Wnt/Frizzled signaling pathway participates in many inflammation-linked diseases. However, the inflammatory response mediated by the Wnt/Frizzled signaling pathway in experimental subarachnoid hemorrhage has not been thoroughly investigated. Consequently, in this study, we examined the potential role of the Wnt/Frizzled signaling pathway in early brain injury in rat models of subarachnoid hemorrhage.Simultaneously, possible neuroprotective mechanisms were also investigated. Experimental subarachnoid hemorrhage rat models were induced by injecting autologous blood into the prechiasmatic cistern. Experiment 1 was designed to examine expression of the Wnt/Frizzled signaling pathway in early brain injury induced by subarachnoid hemorrhage. In total, 42 adult rats were divided into sham(injection of equivalent volume of saline), 6-, 12-, 24-, 48-, 72-hour, and 1-week subarachnoid hemorrhage groups. Experiment 2 was designed to examine neuroprotective mechanisms of the Wnt/Frizzled signaling pathway in early brain injury induced by subarachnoid hemorrhage. Rats were treated with recombinant human Wnt1(rhwnt1), small interfering Wnt1(siwnt1) RNA, and monoclonal antibody of Frizzled1(anti-Frizzled1) at 48 hours after subarachnoid hemorrhage. Expression levels of Wnt1, Frizzled1, β-catenin, peroxisome proliferator-activated receptor-γ, CD36, and active nuclear factor-κB were examined by western blot assay and immunofluorescence staining. Microglia type conversion and inflammatory cytokine levels in brain tissue were examined by immunofluorescence staining and enzyme-linked immunosorbent assay. Our results show that compared with the sham group, expression levels of Wnt1, Frizzled1, and β-catenin were low and reduced to a minimum at 48 hours, gradually returning to baseline at 1 week after subarachnoid hemorrhage. rhwnt1 treatment markedly increased Wnt1 expression and alleviated subarachnoid hemorrhage-induced early brain injury(within 72 hours), including cortical cell apoptosis, brain edema, and neurobehavioral deficits, accompanied by increasing protein levels of β-catenin, CD36, and peroxisome proliferator-activated receptor-γ and decreasing protein levels of nuclear factor-κB. Of note, rhwnt1 promoted M2-type microglia conversion and inhibited release of inflammatory cytokines(interleukin-1β, interleukin-6, and tumor necrosis factor-α). In contrast, siwnt1 RNA and anti-Frizzled1 treatment both resulted in an opposite effect. In conclusion, the Wnt/Frizzled1 signaling pathway may participate in subarachnoid hemorrhage-induced early brain injury via inhibiting the inflammatory response, including regulating microglia type conversion and decreasing inflammatory cytokine release. The study was approved by the Animal Ethics Committee of Anhui Medical University and First Affiliated Hospital of USTC,Division of Life Sciences and Medicine, University of Science and Technology of China(approval No. LLSC-20180202) in May 2017.
文摘The incidence of ischemic stroke in patients with diabetes is increasing. While brachial-ankle pulse wave velocity （BaPWV） and ankle-bra- chial index （ABI） are known to be associated with ischemic cardiovascular and cerebrovascular diseases, whether these measures predict the risk of ischemic cerebrovascular disease in diabetic patients remains unclear. 117 patients with type 2 diabetes were enrolled in this study. According to the results of head magnetic resonance imaging, the patients were divided into a diabetes-only group （n = 55） and a diabetes and ischemic stroke group （n = 62）. We then performed ABI and BaPWV examinations for all patients. Compared with the diabe- tes-only group, we found decreased ABI and increased BaPWV in the diabetes and ischemic stroke group. Multivariate logistic regression analyses revealed that BaPWV and ABI were risk factors for ischemic stroke in patients with type 2 diabetes. Our findings indicate that decreased ABI and increased BaPWV are objective indicators of increased risk of ischemic stroke in patients with type 2 diabetes.
基金funded by the National Natural Science Foundation of China,No.81071001,30900805
文摘We previously found that the K141N mutation in heat shock protein B8 （HSPB8） was responsible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a transgenic mouse model bearing the K141N mutation in the human HSPB8 gene, and to determine whether this K141NHSPB8 transgenic mouse model would manifest the clinical phenotype of Charcot-Marie-Tooth disease type 2L, and consequently be suitable for use in studies of disease pathogenesis. Transgenic mice overexpressing K141N HSPB8 were generated using K141N mutant HSPB8 cDNA cloned into a pCAGGS plasmid driven by a human cytomegalovirus expression system. PCR and western blot analysis confirmed integration of the KI41NHSPB8 gene and widespread expression in tissues of the transgenic mice. The K141N HSPB8 transgenic mice exhibited decreased muscle strength in the hind limbs and impaired motor coordination, but no obvious sensory disturbance at 6 months of age by behavioral assessment. Electrophysiological analysis showed that the compound motor action potential amplitude in the sciatic nerve was significantly decreased, but motor nerve conduction velocity remained normal at 6 months of age. Pathological analysis of the sciatic nerve showed reduced myelinated fiber density, notable axonal edema and vacuolar degeneration in K141N HSPB8 transgenic mice, suggesting axonal involvement in the peripheral nerve damage in these animals. These findings indicate that the KI4mHSPB8 transgenic mouse successfully models Charcot-Marie-Tooth disease type 2L and can be used to study the pathogenesis of the disease.
基金supported by the Natural Science Foundation of China,No.81501610,81350030the Priority Academic Program Development of Jiangsu Higher Education Institutes of China
文摘The best tissue-engineered spinal cord grafts not only match the structural characteristics of the spinal cord but also allow the seed cells to grow and function in situ.Platelet-derived growth factor（PDGF） has been shown to promote the migration of bone marrow stromal cells;however,cytokines need to be released at a steady rate to maintain a stable concentration in vivo.Therefore,new methods are needed to maintain an optimal concentration of cytokines over an extended period of time to effectively promote seed cell localization,proliferation and differentiation.In the present study,a partition-type tubular scaffold matching the anatomical features of the thoracic 8–10 spinal cord of the rat was fabricated using chitosan and then subsequently loaded with chitosan-encapsulated PDGF-BB microspheres（PDGF-MSs）.The PDGF-MS-containing scaffold was then examined in vitro for sustained-release capacity,biocompatibility,and its effect on neural progenitor cells differentiated in vitro from multilineage-differentiating stress-enduring cells（MUSE-NPCs）.We found that pre-freezing for 2 hours at-20°C significantly increased the yield of partition-type tubular scaffolds,and 30 μL of 25% glutaraldehyde ensured optimal crosslinking of PDGF-MSs.The resulting PDGF-MSs cumulatively released 52% of the PDGF-BB at 4 weeks in vitro without burst release.The PDGF-MS-containing tubular scaffold showed suitable biocompatibility towards MUSE-NPCs and could promote the directional migration and growth of these cells.These findings indicate that the combination of a partition-type tubular scaffold,PDGF-MSs and MUSENPCs may be a promising model for the fabrication of tissue-engineered spinal cord grafts.
