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15 April 2020, Volume 15 Issue 4 Previous Issue    Next Issue

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Extracellular matrix and biomimetic engineering microenvironment for neuronal differentiation Deepak Jain, Sabrina Mattiassi, Eyleen L. Goh, Evelyn K.F. Yim 2020, 15 (4):  573-585.  doi: 10.4103/1673-5374.266907 Abstract ( 97 )   PDF (1749KB) ( 212 )   Save Extracellular matrix (ECM) influences cell differentiation through its structural and biochemical properties. In nervous system, neuronal behavior is influenced by these ECMs structures which are present in a meshwork, fibrous, or tubular forms encompassing specific molecular compositions. In addition to contact guidance, ECM composition and structures also exert its effect on neuronal differentiation. This short report reviewed the native ECM structure and composition in central nervous system and peripheral nervous system, and their impact on neural regeneration and neuronal differentiation. Using topographies, stem cells have been differentiated to neurons. Further, focussing on engineered biomimicking topographies, we highlighted the role of anisotropic topographies in stem cell differentiation to neurons and its recent temporal application for efficient neuronal differentiation. Related Articles | Metrics

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Extracellular vesicles in the diagnosis and treatment of central nervous system diseases Alisa A. Shaimardanova, Valeriya V. Solovyeva, Daria S. Chulpanova, Victoria James, Kristina V. Kitaeva, Albert A. Rizvanov 2020, 15 (4):  586-596.  doi: 10.4103/1673-5374.266908 Abstract ( 232 )   PDF (1400KB) ( 165 )   Save Extracellular vesicles, including exosomes and microvesicles, play a fundamental role in the activity of the nervous system, participating in signal transmission between neurons and providing the interaction of central nervous system with all body systems. In many neurodegenerative diseases, neurons pack toxic substances into vesicles and release them into the extracellular space, which leads to the spread of misfolded neurotoxic proteins. The contents of neuron-derived extracellular vesicles may indicate pathological changes in the central nervous system, and the analysis of extracellular vesicle molecular content contributes to the development of non-invasive methods for the diagnosis of many central nervous system diseases. Extracellular vesicles of neuronal origin can be isolated from various biological fluids due to their ability to cross the blood-brain barrier. Today, the diagnostic potential of almost all toxic proteins involved in nervous system disease pathogenesis, specifically α-synuclein, tau protein, superoxide dismutase 1, FUS, leucine-rich repeat kinase 2, as well as some synaptic proteins, has been well evidenced. Special attention is paid to extracellular RNAs mostly associated with extracellular vesicles, which are important in the onset and development of many neurodegenerative diseases. Depending on parental cell type, extracellular vesicles may have different therapeutic properties, including neuroprotective, regenerative, and anti-inflammatory. Due to nano size, biosafety, ability to cross the blood-brain barrier, possibility of targeted delivery and the lack of an immune response, extracellular vesicles are a promising vehicle for the delivery of therapeutic substances for the treatment of neurodegenerative diseases and drug delivery to the brain. This review describes modern approaches of diagnosis and treatment of central nervous system diseases using extracellular vesicles. Related Articles | Metrics

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Evidence and explanation for the involvement of the nucleus accumbens in pain processing Haley N. Harris, Yuan B. Peng 2020, 15 (4):  597-605.  doi: 10.4103/1673-5374.266909 Abstract ( 108 )   PDF (1135KB) ( 151 )   Save The nucleus accumbens (NAc) is a subcortical brain structure known primarily for its roles in pleasure, reward, and addiction. Despite less focus on the NAc in pain research, it also plays a large role in the mediation of pain and is effective as a source of analgesia. Evidence for this involvement lies in the NAc’s cortical connections, functions, pharmacology, and therapeutic targeting. The NAc projects to and receives information from notable pain structures, such as the prefrontal cortex, anterior cingulate cortex, periaqueductal gray, habenula, thalamus, etc. Additionally, the NAc and other pain-modulating structures share functions involving opioid regulation and motivational and emotional processing, which each work beyond simply the rewarding experience of pain offset. Pharmacologically speaking, the NAc responds heavily to painful stimuli, due to its high density of μ opioid receptors and the activation of several different neurotransmitter systems in the NAc, such as opioids, dopamine, calcitonin gene-related peptide, γ-aminobutyric acid, glutamate, and substance P, each of which have been shown to elicit analgesic effects. In both preclinical and clinical models, deep brain stimulation of the NAc has elicited successful analgesia. The multi-functional NAc is important in motivational behavior, and the motivation for avoiding pain is just as important to survival as the motivation for seeking pleasure. It is possible, then, that the NAc must be involved in both pleasure and pain in order to help determine the motivational salience of positive and negative events. Related Articles | Metrics

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MicroRNAs in blood and cerebrospinal fluid as diagnostic biomarkers of multiple sclerosis and to monitor disease progression Bridget Martinez, Philip V. Peplow 2020, 15 (4):  606-619.  doi: 10.4103/1673-5374.266905 Abstract ( 108 )   PDF (406KB) ( 148 )   Save Multiple sclerosis is a chronic autoimmune disease of the central nervous system. It is the main cause of non-traumatic neurological disability in young adults. Multiple sclerosis mostly affects people aged 20–50 years; however, it can occur in young children and much older adults. Factors identified in the distribution of MS include age, gender, genetics, environment, and ethnic background. Multiple sclerosis is usually associated with progressive degrees of disability. The disease involves demyelination of axons of the central nervous system and causes brain and spinal cord neuronal loss and atrophy. Diagnosing multiple sclerosis is based on a patient’s medical history including symptoms, physical examination, and various tests such as magnetic resonance imaging, cerebrospinal fluid and blood tests, and electrophysiology. The disease course of multiple sclerosis is not well correlated with the biomarkers presently used in clinical practice. Blood-derived biomarkers that can detect and distinguish the different phenotypes in multiple sclerosis may be advantageous in personalized treatment with disease-modifying drugs and to predict response to treatment. The studies reviewed have shown that the expression levels of a large number of miRNAs in peripheral blood, serum, exosomes isolated from serum, and cerebrospinal fluid are altered in multiple sclerosis and can distinguish the disease phenotypes from each other. Further studies are warranted to independently validate these findings so that individual or pairs of miRNAs in serum or cerebrospinal fluid can be used as potential diagnostic markers for adult and pediatric multiple sclerosis and for monitoring disease progression and response to therapy. Related Articles | Metrics

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Urokinase-type plasminogen activator is a modulator of synaptic plasticity in the central nervous system: implications for neurorepair in the ischemic brain Manuel Yepes 2020, 15 (4):  620-624.  doi: 10.4103/1673-5374.266904 Abstract ( 113 )   PDF (393KB) ( 133 )   Save The last two decades have witnessed a rapid decrease in mortality due to acute cerebral ischemia that paradoxically has led to a rapid increase in the number of patients that survive an acute ischemic stroke with various degrees of disability. Unfortunately, the lack of an effective therapeutic strategy to promote neurological recovery among stroke survivors has led to a rapidly growing population of disabled patients. Thus, understanding the mechanisms of neurorepair in the ischemic brain is a priority with wide scientific, social and economic implications. Cerebral ischemia has a harmful effect on synaptic structure associated with the development of functional impairment. In agreement with these observations, experimental evidence indicates that synaptic repair underlies the recovery of neurological function following an ischemic stroke. Furthermore, it has become evident that synaptic plasticity is crucial not only during development and learning, but also for synaptic repair after an ischemic insult. The plasminogen activating system is assembled by a cascade of enzymes and their inhibitors initially thought to be solely involved in the generation of plasmin. However, recent work has shown that in the brain this system has an important function regulating the development of synaptic plasticity via mechanisms that not always require plasmin generation. Urokinase-type plasminogen activator (uPA) is a serine proteinase and one of the plasminogen activators, that upon binding to its receptor (uPAR) not only catalyzes the conversion of plasminogen into plasmin on the cell surface, but also activates cell signaling pathways that promote cell migration, proliferation and survival. The role of uPA is the brain is not fully understood. However, it has been reported while uPA and uPAR are abundantly found in the developing central nervous system, in the mature brain their expression is restricted to a limited group of cells. Remarkably, following an ischemic injury to the mature brain the expression of uPA and uPAR increases to levels comparable to those observed during development. More specifically, neurons release uPA during the recovery phase from an ischemic injury, and astrocytes, axonal boutons and dendritic spines recruit uPAR to their plasma membrane. Here we will review recent evidence indicating that binding of uPA to uPAR promotes the repair of synapses damaged by an ischemic injury, with the resultant recovery of neurological function. Furthermore, we will discuss data indicating that treatment with recombinant uPA is a potential therapeutic strategy to promote neurological recovery among ischemic stroke survivors. Related Articles | Metrics

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Neuroinflammation and oxidative stress act in concert to promote neurodegeneration in the diabetic retina and optic nerve: galectin-3 participation Henrique Rocha Mendonça, Raul Carpi-Santos, Karin da Costa Calaza, Ana Maria Blanco Martinez 2020, 15 (4):  625-635.  doi: 10.4103/1673-5374.266910 Abstract ( 97 )   PDF (885KB) ( 183 )   Save Diabetes is a lifelong disease characterized by glucose metabolic imbalance, in which low insulin levels or impaired insulin signaling lead to hyperglycemic state. Within 20 years of diabetes progression, 95% of patients will have diabetic retinopathy, the leading cause of visual defects in working-age people worldwide. Although diabetes is considered a microvascular disease, recent studies have shown that neurodegeneration precedes vascular changes within the diabetic visual system, albeit its mechanisms are still under investigation. Neuroinflammation and oxidative stress are intrinsically related phenomena, since macrophage/ microglia and astrocytes are the main sources of reactive oxygen species during central nervous system chronic degenerative diseases, and both pathological processes are increased in the visual system during diabetes. The present review will focus on recent findings of the contribution of oxidative stress derived from neuroinflammation in the early neurodegenerative aspects of the diabetic visual system and their relationship with galectin-3. Related Articles | Metrics

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Unfolded protein response in myelin disorders Wensheng Lin, Sarrabeth Stone 2020, 15 (4):  636-645.  doi: 10.4103/1673-5374.266903 Abstract ( 80 )   PDF (531KB) ( 257 )   Save Activation of the unfolded protein response in response to endoplasmic reticulum stress preserves cell viability and function under stressful conditions. Nevertheless, persistent, unresolvable activation of the unfolded protein response can trigger apoptosis to eliminate stressed cells. Recent studies show that the unfolded protein response plays an important role in the pathogenesis of various disorders of myelin, including multiples sclerosis, Charcot-Marie-Tooth disease, Pelizaeus-Merzbacher disease, vanishing white matter disease, spinal cord injury, tuberous sclerosis complex, and hypoxia-induced perinatal white matter injury. In this review we summarize the current literature on the unfolded protein response and the evidence for its role in the pathogenesis of myelin disorders. Related Articles | Metrics

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Relationship between MRI perfusion and clinical severity in multiple sclerosis Maria Marcella Laganà, Laura Pelizzari, Francesca Baglio 2020, 15 (4):  646-652.  doi: 10.4103/1673-5374.266906 Abstract ( 112 )   PDF (203KB) ( 371 )   Save Perfusion alterations within several brain regions have been shown in multiple sclerosis patients using different magnetic resonance imaging (MRI) techniques. Furthermore, MRI-derived brain perfusion metrics have been investigated in association with multiple sclerosis phenotypes, physical disability, and cognitive impairment. However, a review focused on these aspects is still missing. Our aim was to review all the studies investigating the relationship between perfusion MRI and clinical severity during the last fifteen years to understand the clinical relevance of these findings. Perfusion differences among phenotypes were observed both with 1.5T and 3T scanners, with progressive multiple sclerosis presenting with lower perfusion values than relapsing-remitting multiple sclerosis patients. However, only 3T scanners showed a statistically significant distinction. Controversial results about the association between MRI-derived perfusion metrics and physical disability scores were found. However, the majority of the studies showed that lower brain perfusion and longer transit time are associated with more severe physical disability and worse cognitive performances. Related Articles | Metrics

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Therapeutic importance of hydrogen sulfide in age-associated neurodegenerative diseases Rubaiya Tabassum, Na Young Jeong, Junyang Jung 2020, 15 (4):  653-662.  doi: 10.4103/1673-5374.266911 Abstract ( 195 )   PDF (2445KB) ( 163 )   Save Hydrogen sulfide (H2S) is a gasotransmitter that acts as an antioxidant and exhibits a wide variety of cytoprotective and physiological functions in age-associated diseases. One of the major causes of age-related diseases is oxidative stress. In recent years, the importance of H2S has become clear, although its antioxidant function has not yet been fully explored. The enzymes cystathionine β-synthase, cystathionine γ-lya-se, and 3-mercaptopyruvate sulfurtransferase are involved in the enzymatic production of H2S. Previously, H2S was considered a neuromodulator, given its role in long-term hippocampal potentiation, but it is now also recognized as an antioxidant in age-related neurodegeneration. Due to aerobic metabolism, the central nervous system is vulnerable to oxidative stress in brain aging, resulting in age-associated degenerative diseases. H2S exerts its antioxidant effect by limiting free radical reactions through the activation of antioxidant enzymes, including superoxide dismutase, catalase, and glutathione peroxidase, which protect against the effects of aging by regulating apoptosis-related genes, including p53, Bax, and Bcl-2. This review explores the implications and mechanisms of H2S as an antioxidant in age-associated neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, and Down syndrome. Related Articles | Metrics

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Role of CD20+ T cells in multiple sclerosis: implications for treatment with ocrelizumab Stefan Gingele, Thomas Skripuletz, Roland Jacobs 2020, 15 (4):  663-664.  doi: 10.4103/1673-5374.266913 Abstract ( 94 )   PDF (361KB) ( 179 )   Save CD20 is a membrane-spanning hosphoprotein strongly expressed on the cell surface of B lineage cells and is widely regarded as a B cell specific marker. However, CD20 (Figure 1) is also expressed at a low level on a small subset of CD3+ T cells which therefore are sometimes referred to as CD20dimCD3+ T cells in contrast to CD20brightCD19+ B cells which represent the majority of cells expressing CD20 (Hultin et al., 1993). The amount of the CD20 antigen has been assessed to be 25 to 50 times higher on CD20+CD19+ B cells compared to CD20+CD3+ T cells (Hultin et al., 1993). At first description of this cell population the frequency of these CD20+ T cells has been described to represent an average of 2.4% of all peripheral blood lymphocytes (50 healthy controls) (Hultin et al., 1993). In the largest characterized cohort with 142 healthy individuals CD20+CD3+ T cells constituted a mean proportion of 1.6% (range from 0.1–6.8%) of all circulating CD3+ T cells and in absolute numbers accounted for approximately 28 cells/μL (Wilk et al., 2009). Related Articles | Metrics

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Nerve growth factor catches copper in neuronal inning. Diego La Mendola 2020, 15 (4):  665-666.  doi: 10.4103/1673-5374.266912 Abstract ( 74 )   PDF (230KB) ( 156 )   Save Neurotrophins (NTs) are a family of homologues proteins that play an essential role in neuronal cells growth, survival and differentiation. These proteins include the nerve growth factor (NGF), the brain derived neurotrophic factor (BDNF), NT-3 and NT-3, also known as NT-4/5. Related Articles | Metrics

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Inhibiting endogenous tissue plasminogen activator enhanced neuronal apoptosis and axonal injury after traumatic brain injury Jun-Jie Zhao, Zun-Wei Liu, Bo Wang, Ting-Qin Huang, Dan Guo, Yong-Lin Zhao, Jin-Ning Song 2020, 15 (4):  667-675.  doi: 10.4103/1673-5374.266914 Abstract ( 100 )   PDF (5690KB) ( 185 )   Save Tissue plasminogen activator is usually used for the treatment of acute ischemic stroke, but the role of endogenous tissue plasminogen activator in traumatic brain injury has been rarely reported. A rat model of traumatic brain injury was established by weight-drop method. The tissue plasminogen activator inhibitor neuroserpin (5 μL, 0.25 mg/mL) was injected into the lateral ventricle. Neurological function was assessed by neurological severity score. Neuronal and axonal injuries were assessed by hematoxylin-eosin staining and Bielschowsky silver staining. Protein level of endogenous tissue plasminogen activator was analyzed by western blot assay. Apoptotic marker cleaved caspase-3, neuronal marker neurofilament light chain, astrocyte marker glial fibrillary acidic protein and microglial marker Iba-1 were analyzed by immunohistochemical staining. Apoptotic cell types were detected by immunofluorescence double labeling. Apoptotic cells in the damaged cortex were detected by terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP-biotin nick-end labeling staining. Degenerating neurons in the damaged cortex were detected by Fluoro-Jade B staining. Expression of tissue plasminogen activator was increased at 6 hours, and peaked at 3 days after traumatic brain injury. Neuronal apoptosis and axonal injury were detected after traumatic brain injury. Moreover, neuroserpin enhanced neuronal apoptosis, neuronal injury and axonal injury, and activated microglia and astrocytes. Neuroserpin further deteriorated neurobehavioral function in rats with traumatic brain injury. Our findings confirm that inhibition of endogenous tissue plasminogen activator aggravates neuronal apoptosis and axonal injury after traumatic brain injury, and activates microglia and astrocytes. This study was approved by the Biomedical Ethics Committee of Animal Experiments of Shaanxi Province in China in June 2015 (approval No. SYXK [Shaan] 2015-002). Related Articles | Metrics

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Inflammation-related gene expression profiles of salivary extracellular vesicles in patients with head trauma Yan Cheng, Mandy Pereira, Neha P. Raukar, John L. Reagan, Mathew Quesenberry, Laura Goldberg, Theodor Borgovan, W Curt LaFrance Jr, Mark Dooner, Maria Deregibus, Giovanni Camussi, Bharat Ramratnam, Peter Quesenberry 2020, 15 (4):  676-681.  doi: 10.4103/1673-5374.266924 Abstract ( 102 )   PDF (597KB) ( 133 )   Save At present, there is no reliable biomarker for the diagnosis of traumatic brain injury (TBI). Studies have shown that extracellular vesicles released by damaged cells into biological fluids can be used as potential biomarkers for diagnosis of TBI and evaluation of TBI severity. We hypothesize that the genetic profile of salivary extracellular vesicles in patients with head trauma differs from that in uninjured subjects. Findings from this hypothesis would help investigate the severity of TBI. This study included 19 subjects, consisting of seven healthy controls who denied history of head trauma, six patients diagnosed with concussion injury from an outpatient concussion clinic, and six patients with TBI who received treatment in the emergency department within 24 hours after injury. Real-time PCR analysis of salivary extracellular vesicles in participants was performed using TaqMan Human Inflammation array. Gene expression analysis revealed nine upregulated genes in emergency department patients (LOX5, ANXA3, CASP1, IL2RG, ITGAM, ITGB2, LTA4H, MAPK14, and TNFRSF1A) and 13 upregulated genes in concussion clinic patients compared with healthy participants (ADRB1, ADRB2, BDKRB1, HRH1, HRH2, LTB4R2, LTB4R, PTAFR, CYSLTR1, CES1, KLK1, MC2R, and PTGER3). Each patient group had a unique profile. Comparison between groups showed that 15 inflammation-related genes had significant expression change. Our results indicate that inflammation biomarkers can be used for diagnosis of TBI and evaluation of disease severity. This study was approved by the Institutional Review Board on December 18, 2015 (approval No. 0078-12) and on June 9, 2016 (approval No. 4093-16). Related Articles | Metrics

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Protective effects of organic extracts of Alpinia oxyphylla against hydrogen peroxide-induced cytotoxicity in PC12 cells Li-Hong Duan, Meng Li, Chun-Bao Wang, Qing-Mei Wang, Quan-Quan Liu, Wan-Feng Shang, Ya-Jin Shen, Zhuo-Hua Lin, Tong-Yang Sun, Zheng-Zhi Wu, Ying-Hong Li, Yu-Long Wang, Xun Luo 2020, 15 (4):  682-689.  doi: 10.4103/1673-5374.266918 Abstract ( 129 )   PDF (6030KB) ( 252 )   Save Alpinia oxyphylla, a traditional herb, is widely used for its neuroprotective, antioxidant and memory-improving effects. However, the neuroprotective mechanisms of action of its active ingredients are unclear. In this study, we investigated the neuroprotective effects of various organic extracts of Alpinia oxyphylla on PC12 cells exposed to hydrogen peroxide-induced oxidative injury in vitro. Alpinia oxyphylla was extracted three times with 95% ethanol (representing extracts 1–3). The third 95% ethanol extract was dried and resuspended in water, and then extracted successively with petroleum ether, ethyl acetate and n-butanol (representing extracts 4–6). The cell counting kit-8 assay and microscopy were used to evaluate cell viability and observe the morphology of PC12 cells. The protective effect of the three ethanol extracts (at tested concentrations of 50, 100 and 200 μg/mL) against cytotoxicity to PC12 cells increased in a concentration-dependent manner. The ethyl acetate, petroleum ether and n-butanol extracts (each tested at 100, 150 and 200 μg/mL) had neuroprotective effects as well. The optimum effective concentration ranged from 50–200 μg/mL, and the protective effect of the ethyl acetate extract was comparatively robust. These results demonstrate that organic extracts of Alpinia oxyphylla protect PC12 cells against apoptosis induced by hydrogen peroxide. Our findings should help identify the bioactive neuroprotective components in Alpinia oxyphylla. Related Articles | Metrics

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Protective effect of rhodioloside and bone marrow mesenchymal stem cells infected with HIF-1- expressing adenovirus on acute spinal cord injury Xiao-Qin Ha, Bo Yang, Huai-Jing Hou, Xiao-Ling Cai, Wan-Yuan Xiong, Xu-Pan Wei 2020, 15 (4):  690-696.  doi: 10.4103/1673-5374.266920 Abstract ( 120 )   PDF (4632KB) ( 193 )   Save Rhodioloside has been shown to protect cells from hypoxia injury, and bone marrow mesenchymal stem cells have a good effect on tissue repair. To study the effects of rhodioloside and bone marrow mesenchymal stem cells on spinal cord injury, a rat model of spinal cord injury was established using the Infinite Horizons method. After establishing the model, the rats were randomly divided into five groups. Rats in the control group were intragastrically injected with phosphate buffered saline (PBS) (5 μL). PBS was injected at 6 equidistant points around 5 mm from the injury site and at a depth of 5 mm. Rats in the rhodioloside group were intragastrically injected with rhodioloside (5 g/kg) and intramuscularly injected with PBS. Rats in the mesenchymal stem cell (MSC) group were intramuscularly injected with PBS and intramuscularly with MSCs (8 × 106/mL in a 50-μL cell suspension). Rats in the Ad-HIF-MSC group were intragastrically injected with PBS and intramuscularly injected with HIF-1 adenovirus-infected MSCs. Rats in the rhodioloside + Ad-HIF-MSC group were intramuscularly injected with MSCs infected with the HIF-1 adenovirus and intragastrically injected with rhodioloside. One week after treatment, exercise recovery was evaluated with a modified combined behavioral score scale. Hematoxylin-eosin staining and Pischingert’s methylene blue staining were used to detect any histological or pathological changes in spinal cord tissue. Levels of adenovirus IX and Sry mRNA were detected by real-time quantitative polymerase chain reaction and used to determine the number of adenovirus and mesenchymal stem cells that were transfected into the spinal cord. Immunohistochemical staining was applied to detect HIF-1 protein levels in the spinal cord. The results showed that: (1) compared with the other groups, the rhodioloside + Ad-HIF-MSC group exhibited the highest combined behavioral score (P < 0.05), the most recovered tissue, and the greatest number of neurons, as indicated by Pischingert’s methylene blue staining. (2) Compared with the PBS group, HIF-1 protein expression was greater in the rhodioloside group (P < 0.05). (3) Compared with the Ad-HIF-MSC group, Sry mRNA levels were higher in the rhodioloside + Ad-HIF-MSC group (P < 0.05). These results confirm that rhodioloside combined with bone marrow mesenchymal stem cells can promote the recovery of spinal cord injury and activate the HIF-1 pathway to promote the survival of bone marrow mesenchymal stem cells and repair damaged neurons within spinal cord tissue. This experiment was approved by the Animal Ethics Committee of Gansu University of Traditional Chinese Medicine, China (approval No. 2015KYLL029) in June 2015. Related Articles | Metrics

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Changes in neurological and pathological outcomes in a modified rat spinal cord injury model with closed canal Xin Sun, Xing-Zhen Liu, Jia Wang, Hai-Rong Tao, Tong Zhu, Wen-Jie Jin, Kang-Ping Shen 2020, 15 (4):  697-704.  doi: 10.4103/1673-5374.266919 Abstract ( 103 )   PDF (2595KB) ( 191 )   Save Most animal spinal cord injury models involve a laminectomy, such as the weight drop model or the transection model. However, in clinical practice, many patients undergo spinal cord injury while maintaining a relatively complete spinal canal. Thus, open spinal cord injury models often do not simulate real injuries, and few previous studies have investigated whether having a closed spinal canal after a primary spinal cord injury may influence secondary processes. Therefore, we aimed to assess the differences in neurological dysfunction and pathological changes between rat spinal cord injury models with closed and open spinal canals. Sprague-Dawley rats were randomly divided into three groups. In the sham group, the tunnel was expanded only, without inserting a screw into the spinal canal. In the spinal cord injury with open canal group, a screw was inserted into the spinal canal to cause spinal cord injury for 5 minutes, and then the screw was pulled out, leaving a hole in the vertebral plate. In the spinal cord injury with closed canal group, after inserting a screw into the spinal canal for 5 minutes, the screw was pulled out by approximately 1.5 mm and the flat end of the screw remained in the hole in the vertebral plate so that the spinal canal remained closed; this group was the modified model, which used a screw both to compress the spinal cord and to seal the spinal canal. At 7 days post-operation, the Basso-Beattie-Bresnahan scale was used to measure changes in neurological outcomes. Hematoxylin- eosin staining was used to assess histopathology. To evaluate the degree of local secondary hypoxia, immunohistochemical staining and western blot assays were applied to detect the expression of hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF). Compared with the spinal cord injury with open canal group, in the closed canal group the Basso-Beattie-Bresnahan scores were lower, cell morphology was more irregular, the percentage of morphologically normal neurons was lower, the percentages of HIF-1α- and VEGF-immunoreactive cells were higher, and HIF-1α and VEGF protein expression was also higher. In conclusion, we successfully established a rat spinal cord injury model with closed canal. This model could result in more serious neurological dysfunction and histopathological changes than in open canal models. All experimental procedures were approved by the Institutional Animal Care Committee of Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, China (approval No. HKDL201810) on January 30, 2018. Related Articles | Metrics

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Spatiotemporal expression of leukemia inhibitory factor receptor protein during neural tube development in embryos with neural tube defects Dong An, Xiao-Wei Wei, He-Nan Zhang, Dan Liu, Wei Ma, Zheng-Wei Yuan 2020, 15 (4):  705-711.  doi: 10.4103/1673-5374.266921 Abstract ( 111 )   PDF (2610KB) ( 126 )   Save Leukemia inhibitory factor receptor (LIFR), as a neuroregulatory cytokine receptor, generally shows a neuroprotective effect in central nervous system injuries. In this study, to understand the effect of LIFR on pathogenesis of neural tube defects, we explored spatiotemporal expression of LIFR at different stages of fetal development in normal and neural tube defect embryos. Spina bifida aperta was induced with all-trans retinoic acid on embryonic day 10 in rats, and the spatiotemporal expression of LIFR was investigated in spina bifida aperta rats and healthy rats from embryonic day 11 to 17. Real time-polymerase chain reaction and western blot assay were used to examine mRNA and protein expression of LIFR in healthy control and neural tube defect embryos. Results of the animal experiment demonstrated that expression of LIFR protein and mRNA in the spinal cords of normal rat embryos increased with embryonic development. LIFR was significantly downregulated in the spinal cords of spina bifida aperta rats compared with healthy rats from embryonic days 11 to 17. Immunohistochemical staining showed that the expression of LIFR in placenta and spinal cord in spina bifida aperta rat embryos was decreased compared with that in control embryos at embryonic day 15. Results from human embryo specimens showed that LIFR mRNA expression was significantly down-regulated in spinal cords of human fetuses with neural tube defects compared with normal controls at a gestational age of 24 to 33 weeks. The results were consistent with the down-regulation of LIFR in the animal experiments. Our study revealed spatiotemporal changes in expression of LIFR during embryonic neurulation. Thus, LIFR might play a specific role in neural tube development. All animal and human experimental procedures were approved by the Medical Ethics Committee of Shengjing Hospital of China Medical University, China (approval No. 2016PS106K) on February 25, 2016. Related Articles | Metrics

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Long-term adenosine A1 receptor activation-induced sortilin expression promotes α-synuclein upregulation in dopaminergic neurons Yun-Cheng Lv, An-Bo Gao, Jing Yang, Li-Yuan Zhong, Bo Jia, Shu-Hui Ouyang, Le Gui, Tian-Hong Peng, Sha Sun, Francisco S. Cayabyab 2020, 15 (4):  712-723.  doi: 10.4103/1673-5374.266916 Abstract ( 205 )   PDF (6798KB) ( 192 )   Save Prolonged activation of adenosine A1 receptor likely leads to damage of dopaminergic neurons and subsequent development of neurodegenerative diseases. However, the pathogenesis underlying long-term adenosine A1 receptor activation-induced neurodegeneration remains unclear. In this study, rats were intraperitoneally injected with 5 mg/kg of the adenosine A1 receptor agonist N6-cyclopentyladenosine (CPA) for five weeks. The mobility of rats was evaluated by forced swimming test, while their cognitive capabilities were evaluated by Y-maze test. Expression of sortilin, α-synuclein, p-JUN, and c-JUN proteins in the substantia nigra were detected by western blot analysis. In addition, immunofluorescence staining of sortilin and α-synuclein was performed to detect expression in the substantia nigra. The results showed that, compared with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (5 mg/kg) + CPA co-treated rats, motor and memory abilities were reduced, surface expression of sortin and α-synuclein in dopaminergic neurons was reduced, and total sortilin and total α-synuclein were increased in CPA-treated rats. MN9D cells were incubated with 500 nM CPA alone or in combination with 10 μM SP600125 (JNK inhibitor) for 48 hours. Quantitative real-time polymerase chain reaction analysis of sortilin and α-synuclein mRNA levels in MN9D cells revealed upregulated sortilin expression in MN9D cells cultured with CPA alone, but the combination of CPA and SP600125 could inhibit this expression. Predictions made using Jasper, PROMO, and Alibaba online databases identified a highly conserved sequence in the sortilin promoter that was predicted to bind JUN in both humans and rodents. A luciferase reporter assay of sortilin promoter plasmid-transfected HEK293T cells confirmed this prediction. After sortilin expression was inhibited by sh-SORT1, expression of p-JUN and c-JUN was detected by western blot analysis. Long-term adenosine A1 receptor activation levels upregulated α-synuclein expression at the post-transcriptional level by affecting sortilin expression. The online tool Raptor-X-Binding and Discovery Studio 4.5 prediction software predicted that sortilin can bind to α-synuclein. Co-immunoprecipitation revealed an interaction between sortilin and α-synuclein in MN9D cells. Our findings indicate that suppression of prolonged adenosine A1 receptor activation potently inhibited sortilin expression and α-synuclein accumulation, and dramatically improved host cognition and kineticism. This study was approved by the University Committee of Animal Care and Supply at the University of Saskatchewan (approval No. AUP#20070090) in March 2007, and the Animals Ethics Committee of University of South China (approval No. LL0387-USC) in June 2017. Related Articles | Metrics

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Characteristic response of striatal astrocytes to dopamine depletion Yao-Feng Zhu, Wei-Ping Wang, Xue-Feng Zheng, Zhi Chen, Tao Chen, Zi-Yun Huang, Lin-Ju Jia, Wan-Long Lei 2020, 15 (4):  724-730.  doi: 10.4103/1673-5374.266917 Abstract ( 149 )   PDF (6973KB) ( 39 )   Save Astrocytes and astrocyte-related proteins play important roles in maintaining normal brain function, and also regulate pathological processes in brain diseases and injury. However, the role of astrocytes in the dopamine-depleted striatum remains unclear. A rat model of Parkinson’s disease was therefore established by injecting 10 μL 6-hydroxydopamine (2.5 μg/μL) into the right medial forebrain bundle. Immunohistochemical staining was used to detect the immunoreactivity of glial fibrillary acidic protein (GFAP), calcium-binding protein B (S100B), and signal transducer and activator of transcription 3 (STAT3) in the striatum, and to investigate the co-expression of GFAP with S100B and STAT3. Western blot assay was used to measure the protein expression of GFAP, S100B, and STAT3 in the striatum. Results demonstrated that striatal GFAP-immunoreactive cells had an astrocytic appearance under normal conditions, but that dopamine depletion induced a reactive phenotype with obvious morphological changes. The normal striatum also contained S100B and STAT3 expression. S100B-immunoreactive cells were uniform in the striatum, with round bodies and sparse, thin processes. STAT3-immunoreactive cells presented round cell bodies with sparse processes, or were darkly stained with a large cell body. Dopamine deprivation induced by 6-hydroxydopamine significantly enhanced the immunohistochemical positive reaction of S100B and STAT3. Normal striatal astrocytes expressed both S100B and STAT3. Striatal dopamine deprivation increased the number of GFAP/S100B and GFAP/STAT3 double-labeled cells, and increased the protein levels of GFAP, S100B, and STAT3. The present results suggest that morphological changes in astrocytes and changes in expression levels of astrocyte-related proteins are involved in the pathological process of striatal dopamine depletion. The study was approved by Animal Care and Use Committee of Sun Yat-sen University, China (Zhongshan Medical Ethics 2014 No. 23) on September 22, 2014. Related Articles | Metrics

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Remnant neuromuscular junctions in denervated muscles contribute to functional recovery in delayed peripheral nerve repair Leyang Li, Hiroyuki Yokoyama, Hidetoshi Kaburagi, Takashi Hirai, Kunikazu Tsuji, Mitsuhiro Enomoto, Yoshiaki Wakabayashi, Atsushi Okawa 2020, 15 (4):  731-738.  doi: 10.4103/1673-5374.266925 Abstract ( 201 )   PDF (1786KB) ( 189 )   Save Schwann cell proliferation in peripheral nerve injury (PNI) enhances axonal regeneration compared to central nerve injury. However, even in PNI, long-term nerve damage without repair induces degeneration of neuromuscular junctions (NMJs), and muscle atrophy results in irreversible dysfunction. The peripheral regeneration of motor axons depends on the duration of skeletal muscle denervation. To overcome this difficulty in nerve regeneration, detailed mechanisms should be determined for not only Schwann cells but also NMJ degeneration after PNI and regeneration after nerve repair. Here, we examined motor axon denervation in the tibialis anterior muscle after peroneal nerve transection in thy1-YFP mice and regeneration with nerve reconstruction using allografts. The number of NMJs in the tibialis anterior muscle was maintained up to 4 weeks and then decreased at 6 weeks after injury. In contrast, the number of Schwann cells showed a stepwise decline and then reached a plateau at 6 weeks after injury. For regeneration, we reconstructed the degenerated nerve with an allograft at 4 and 6 weeks after injury, and evaluated functional and histological outcomes for 10 to 12 weeks after grafting. A higher number of pretzel-shaped NMJs in the tibialis anterior muscle and better functional recovery were observed in mice with a 4-week delay in surgery than in those with a 6-week delay. Nerve repair within 4 weeks after PNI is necessary for successful recovery in mice. Prevention of synaptic acetylcholine receptor degeneration may play a key role in peripheral nerve regeneration. All animal experiments were approved by the Institutional Animal Care and Use Committee of Tokyo Medical and Dental University on 5 July 2017, 30 March 2018, and 15 May 2019 (A2017-311C, A2018-297A, and A2019-248A), respectively. Related Articles | Metrics

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Effects of miR-219/miR-338 on microglia and astrocyte behaviors and astrocyte-oligodendrocyte precursor cell interactions Lan Huong Nguyen, William Ong, Kai Wang, Mingfeng Wang, Dean Nizetic, Sing Yian Chew 2020, 15 (4):  739-747.  doi: 10.4103/1673-5374.266922 Abstract ( 122 )   PDF (3423KB) ( 144 )   Save MiR-219 and miR-338 (miR-219/miR-338) are oligodendrocyte-specific microRNAs. The overexpression of these miRs in oligodendrocyte precursor cells promotes their differentiation and maturation into oligodendrocytes, which may enhance axonal remyelination after nerve injuries in the central nervous system (CNS). As such, the delivery of miR-219/miR-338 to the CNS to promote oligodendrocyte precursor cell differentiation, maturation and myelination could be a promising approach for nerve repair. However, nerve injuries in the CNS also involve other cell types, such as microglia and astrocytes. Herein, we investigated the effects of miR-219/miR-338 treatment on microglia and astrocytes in vitro and in vivo. We found that miR-219/miR-338 diminished microglial expression of pro-inflammatory cytokines and suppressed astrocyte activation. In addition, we showed that miR-219/miR-338 enhanced oligodendrocyte precursor cell differentiation and maturation in a scratch assay paradigm that re-created a nerve injury condition in vitro. Collectively, our results suggest miR-219/miR-338 as a promising treatment for axonal remyelination in the CNS following nerve injuries. All experimental procedures were approved by the Institutional Animal Care and Use Committee (IACUC), Nanyang Technological University (approval No. A0309 and A0333) on April 27, 2016 and October 8, 2016. Related Articles | Metrics

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Effects of neural stem cell transplantation on the motor function of rats with contusion spinal cord injuries: a meta-analysis Kai Qian, Tuo-Ye Xu, Xi Wang, Tao Ma, Kai-Xin Zhang, Kun Yang, Teng-Da Qian, Jing Shi, Li-Xin Li, Zheng Wang 2020, 15 (4):  748-758.  doi: 10.4103/1673-5374.266915 Abstract ( 97 )   PDF (486KB) ( 187 )   Save Objective: To judge the efficacies of neural stem cell (NSC) transplantation on functional recovery following contusion spinal cord injuries (SCIs). Data sources: Studies in which NSCs were transplanted into a clinically relevant, standardized rat model of contusion SCI were identified by searching the PubMed, Embase and Cochrane databases, and the extracted data were analyzed by Stata 14.0. Data selection: Inclusion criteria were that NSCs were used in in vivo animal studies to treat contusion SCIs and that behavioral assessment of locomotor functional recovery was performed using the Basso, Beattie, and Bresnahan lo-comotor rating scale. Exclusion criteria included a follow-up of less than 4 weeks and the lack of control groups. Outcome measures: The restoration of motor function was assessed by the Basso, Beattie, and Bresnahan locomotor rating scale. Results: We identified 1756 non-duplicated papers by searching the aforementioned electronic databases, and 30 full-text articles met the inclusion criteria. A total of 37 studies reported in the 30 articles were included in the meta-analysis. The meta-analysis results showed that transplanted NSCs could improve the motor function recovery of rats following contusion SCIs, to a moderate extent (pooled standardized mean difference (SMD) = 0.73; 95% confidence interval (CI): 0.47–1.00; P < 0.001). NSCs obtained from different donor species (rat: SMD = 0.74; 95% CI: 0.36–1.13; human: SMD = 0.78; 95% CI: 0.31–1.25), at different donor ages (fetal: SMD = 0.67; 95% CI: 0.43–0.92; adult: SMD = 0.86; 95% CI: 0.50–1.22) and from different origins (brain-derived: SMD = 0.59; 95% CI: 0.27–0.91; spinal cord-derived: SMD = 0.51; 95% CI: 0.22–0.79) had similar efficacies on improved functional recovery; however, adult induced pluripotent stem cell-derived NSCs showed no significant efficacies. Furthermore, the use of higher doses of transplanted NSCs or the administration of immunosuppressive agents did not promote better locomotor function recovery (SMD = 0.45; 95% CI: 0.21–0.70). However, shorter periods between the contusion induction and the NSC transplantation showed slightly higher efficacies (acute: SMD = 1.22; 95% CI: 0.81–1.63; subacute: SMD = 0.75; 95% CI: 0.42–1.09). For chronic injuries, NSC implantation did not significantly improve functional recovery (SMD = 0.25; 95% CI: –0.16 to 0.65). Conclusion: NSC transplantation alone appears to be a positive yet limited method for the treatment of contusion SCIs. Related Articles | Metrics

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Small-world networks of neuroblastoma cells cultured in three-dimensional polymeric scaffolds featuring multi-scale roughness Valentina Onesto, Angelo Accardo, Christophe Vieu, Francesco Gentile 2020, 15 (4):  759-768.  doi: 10.4103/1673-5374.266923 Abstract ( 114 )   PDF (2721KB) ( 186 )   Save Understanding the mechanisms underlying cell-surface interaction is of fundamental importance for the rational design of scaffolds aiming at tissue engineering, tissue repair and neural regeneration applications. Here, we examined patterns of neuroblastoma cells cultured in three-dimensional polymeric scaffolds obtained by two-photon lithography. Because of the intrinsic resolution of the technique, the micrometric cylinders composing the scaffold have a lateral step size of ~200 nm, a surface roughness of around 20 nm, and large values of fractal dimension approaching 2.7. We found that cells in the scaffold assemble into separate groups with many elements per group. After cell wiring, we found that resulting networks exhibit high clustering, small path lengths, and small-world characteristics. These values of the topological characteristics of the network can potentially enhance the quality, quantity and density of information transported in the network compared to equivalent random graphs of the same size. This is one of the first direct observations of cells developing into 3D small-world networks in an artificial matrix. Related Articles | Metrics