[Mechanism of tetramethylpyrazine combined with neural stem cells transplantation on cerebral ischemia-reperfusion rats].

This study aimed to investigate the intervention effect of tetramethylpyrazine(TMP) combined with transplantation of neural stem cells(NSCs) on middle cerebral artery occlusion(MCAO) rat model and to explore the mechanism of TMP combined with NSCs transplantation on ischemic stroke based on the regulation of stem cell biological behavior. MCAO rats were randomly divided into a model group, a TMP group, an NSCs transplantation group, and a TMP combined with NSCs transplantation group according to neurological function scores. A sham group was set up at the same time. The neurological function score was used to evaluate the improvement of neurological function in MCAO rats after TMP combined with NSCs transplantation. The proliferation, migration, and differentiation of NSCs were evaluated by BrdU, BrdU/DCX, BrdU/NeuN, and BrdU/GFAP immunofluorescence labeling. The protein expression of stromal cell-derived factor 1(SDF-1), C-X-C motif chemokine receptor 4(CXCR4), as well as oxidative stress pathway proteins nuclear factor erythroid 2-related factor 2(Nrf2), Kelch-like ECH-associated protein 1(KEAP1), heme oxygenase 1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1) was detected by Western blot to study the migration mechanism of TMP combined with NSCs. The results showed that TMP combined with NSCs transplantation significantly improved the neurological function score in MCAO rats. Immunofluorescence staining showed a significant increase in the number of BrdU~+, BrdU~+/DCX~+, BrdU~+/NeuN~+, and BrdU~+/GFAP~+ cells in the TMP, NSCs transplantation, and combined treatment groups, with the combined treatment group showing the most significant increase. Further Western blot analysis revealed significantly elevated expression of CXCR4 protein in the TMP, NSCs transplantation, and combined treatment groups, along with up-regulated protein expression of Nrf2, HO-1, and NQO1, and decreased KEAP1 protein expression. This study showed that both TMP and NSCs transplantation can promote the recovery of neurological function by promoting the proliferation, migration, and differentiation of NSCs, and the effect of TMP combined with NSCs transplantation is superior. The mechanism of action may be related to the activation of the Nrf2/HO-1/CXCR4 pathway.

Fucosyltransferase 9 promotes neuronal differentiation and functional recovery after spinal cord injury by suppressing the activation of Notch signaling.

Individuals with spinal cord injury (SCI) suffer from permanent disabilities such as severe motor, sensory and autonomic dysfunction. Neural stem cell transplantation has proven to be a potential strategy to promote regeneration of the spinal cord, since NSCs can produce neurotrophic growth factors and differentiate into mature neurons to reconstruct the injured site. However, it is necessary to optimize the differentiation of NSCs before transplantation to achieve a better regenerative outcome. Inhibition of Notch signaling leads to a transition from NSCs to neurons, while the underlying mechanism remains inadequately understood. Our results demonstrate that overexpression of fucosyltransferase 9 (Fut9), which is upregulated by Wnt4, promotes neuronal differentiation by suppressing the activation of Notch signaling through disruption of furin-like enzyme activity during S1 cleavage. In an in vivo study, Fut9-modified NSCs efficiently differentiates into neurons to promote functional and histological recovery after SCI. Our research provides insight into the mechanisms of Notch signaling and a potential treatment strategy for SCI.

Restorative effects and mechanisms of neural stem cell transplantation on ischemic brain injury based on the Wnt signaling pathway.

OBJECTIVE: To explore the restorative effects and mechanisms of neural stem cell (NSC) transplantation on ischemic brain injury based on the Wnt signaling pathway. METHODS: Out of 102 male KM mice, 15 were randomly selected as the control group without any intervention, while the remaining 87 underwent middle cerebral artery occlusion (MCAO) using the Zea-Longa suture method. Seven mice that did not successfully model MCAO were excluded, leaving 80 mice that successfully underwent MCAO, randomized into two groups: the Ischemic Brain Injury group (n = 40) receiving 10 µL of sterile PBS solution injected into the lateral ventricle, and the Ischemic Brain Injury + NSCs Transplantation group (n = 40) receiving 10 µL of NSCs suspension injected into the lateral ventricle. RESULTS: Compared to the ischemic brain injury group, mice in the Ischemic Brain Injury + NSCs Transplantation group exhibited significantly alleviated edema in the middle cerebral artery supply area, with neurons displaying more normal morphological characteristics and fewer signs of degeneration and necrosis. The mice with NSC transplantation had significantly smaller infarct volume than those in the ischemic brain injury group (p < 0.05). The mice with NSC transplantation showed significantly lower Zea-Longa scores and a lower proportion of TUNEL-positive cells compared to those in the ischemic brain injury group (p < 0.05). CONCLUSION: NSC transplantation can significantly inhibit neuronal apoptosis in the ischemic region of mice with ischemic brain injury, alleviate brain tissue edema, reduce infarct volume, and improve neurological function. The mechanism may be related to Wnt signaling pathway activation.

Nanomedicine Directs Neuronal Differentiation of Neural Stem Cells via Silencing Long Noncoding RNA for Stroke Therapy.

Transplantation of neural stem cells (NSCs) is a promising treatment paradigm to replace lost neurons and reconstruct the damaged neural circuit after ischemic stroke. However, most transplanted NSCs often differentiate into astrocytes rather than functional neurons, and the poor neuronal differentiation adversely affects the therapeutic outcome of NSCs and limits their clinical translation for stroke therapy. Herein, a theranostic nanomedicine is developed to codeliver superparamagnetic iron oxide nanoparticles (SPIO) and small interfering RNA/antisense oligonucleotides (siRNA/ASO) against Pnky long noncoding RNA (lncRNA) into NSCs. This nanomedicine not only directs neuronal differentiation of NSCs through silencing the Pnky lncRNA but also allows an in vivo tracking of NSCs with magnetic resonance imaging. The enhanced neuronal differentiation of NSCs significantly improved the structural and functional recovery of the damaged brain after a stroke. The results demonstrate the great potential of the multifunctional nanomedicine targeting lncRNA to enhance stem cell-based therapies for a stroke.

Wnt4-modified NSC transplantation promotes functional recovery after spinal cord injury.

Spinal cord injury (SCI) can lead to severe motor and sensory dysfunction, yet there are no effective therapies currently due to the failure of reconstructing the interruption of the neuroanatomical circuit. While neural stem cell (NSC) transplantation has been considered a potential strategy to repair the neural circuit after SCI, the efficacy of this strategy remains unproven. The main reason is that most of the transplanted NSC differentiates into astrocyte rather than neuron in the microenvironment of SCI. Our results demonstrated that Wnt4 significantly promotes the differentiation of NSC into neuron by activating both ß-catenin and MAPK/JNK pathways and suppressing the activation of Notch signaling, which is acknowledged as prevention of NSC differentiation into neuron, through downregulating NICD expression, translocating and preventing the combination of NICD and RbpJ in nucleus. In addition, Wnt4 rescues the negative effect of Jagged, the ligand of Notch signaling, to promote neuronal differentiation. Moreover, in vivo study, transplantation of Wnt4-modified NSC efficaciously repairs the injured spinal cord and recovers the motor function of hind limbs after SCI. This study sheds new light into mechanisms that Wnt4-modified NSC transplantation is sufficient to repair the injured spinal cord and recover the motor dysfunction after SCI.

MiR-7 inhibited peripheral nerve injury repair by affecting neural stem cells migration and proliferation through cdc42.

Objective Neural stem cells play an important role in the recovery and regeneration of peripheral nerve injury, and the microRNA-7 (miR-7) regulates differentiation of neural stem cells. This study aimed to explore the role of miR-7 in neural stem cells homing and proliferation and its influence on peripheral nerve injury repair. Methods The mice model of peripheral nerve injury was created by segmental sciatic nerve defect (sciatic nerve injury), and neural stem cells treatment was performed with a gelatin hydrogel conduit containing neural stem cells inserted into the sciatic nerve injury mice. The Sciatic Function Index was used to quantify sciatic nerve functional recovery in the mice. The messenger RNA and protein expression were detected by reverse transcription polymerase chain reaction and Western blot, respectively. Luciferase reporter assay was used to confirm the binding between miR-7 and the 3'UTR of cell division cycle protein 42 (cdc42). The neural stem cells migration and proliferation were analyzed by transwell assay and a Cell-LightTM EdU DNA Cell Proliferation kit, respectively. Results Neural stem cells treatment significantly promoted nerve repair in sciatic nerve injury mice. MiR-7 expression was decreased in sciatic nerve injury mice with neural stem cells treatment, and miR-7 mimic transfected into neural stem cells suppressed migration and proliferation, while miR-7 inhibitor promoted migration and proliferation. The expression level and effect of cdc42 on neural stem cells migration and proliferation were opposite to miR-7, and the luciferase reporter assay proved that cdc42 was a target of miR-7. Using co-transfection into neural stem cells, we found pcDNA3.1-cdc42 and si-cdc42 could reverse respectively the role of miR-7 mimic and miR-7 inhibitor on neural stem cells migration and proliferation. In addition, miR-7 mimic-transfected neural stem cells could abolish the protective role of neural stem cells on peripheral nerve injury. Conclusion MiR-7 inhibited peripheral nerve injury repair by affecting neural stem cells migration and proliferation through cdc42.

Magnetic Nanoparticles and Methylprednisolone Based Physico-Chemical Bifunctional Neural Stem Cells Delivery System for Spinal Cord Injury Repair.

Neural stem cells (NSCs) transplantation is an attractive and promising treatment strategy for spinal cord injury (SCI). Various pathological processes including the severe inflammatory cascade and difficulty in stable proliferation and differentiation of NSCs limit its application and translation. Here, a novel physico-chemical bifunctional neural stem cells delivery system containing magnetic nanoparticles (MNPs and methylprednisolone (MP) is designed to repair SCI, the former regulates NSCs differentiation through magnetic mechanical stimulation in the chronic phase, while the latter alleviates inflammatory response in the acute phase. The delivery system releases MP to promote microglial M2 polarization, inhibit M1 polarization, and reduce neuronal apoptosis. Meanwhile, NSCs tend to differentiate into functional neurons with magnetic mechanical stimulation generated by MNPs in the static magnetic field, which is related to the activation of the PI3K/AKT/mTOR pathway. SCI mice achieve better functional recovery after receiving NSCs transplantation via physico-chemical bifunctional delivery system, which has milder inflammation, higher number of M2 microglia, more functional neurons, and axonal regeneration. Together, this bifunctional NSCs delivery system combined physical mechanical stimulation and chemical drug therapy is demonstrated to be effective, which provides new treatment insights into clinical transformation of SCI repair.

Reducing host aldose reductase activity promotes neuronal differentiation of transplanted neural stem cells at spinal cord injury sites and facilitates locomotion recovery.

Neural stem cell (NSC) transplantation is a promising strategy for replacing lost neurons following spinal cord injury. However, the survival and differentiation of transplanted NSCs is limited, possibly owing to the neurotoxic inflammatory microenvironment. Because of the important role of glucose metabolism in M1/M2 polarization of microglia/macrophages, we hypothesized that altering the phenotype of microglia/macrophages by regulating the activity of aldose reductase (AR), a key enzyme in the polyol pathway of glucose metabolism, would provide a more beneficial microenvironment for NSC survival and differentiation. Here, we reveal that inhibition of host AR promoted the polarization of microglia/macrophages toward the M2 phenotype in lesioned spinal cord injuries. M2 macrophages promoted the differentiation of NSCs into neurons in vitro. Transplantation of NSCs into injured spinal cords either deficient in AR or treated with the AR inhibitor sorbinil promoted the survival and neuronal differentiation of NSCs at the injured spinal cord site and contributed to locomotor functional recovery. Our findings suggest that inhibition of host AR activity is beneficial in enhancing the survival and neuronal differentiation of transplanted NSCs and shows potential as a treatment of spinal cord injury.

Effects of neural stem cell transplantation on the motor function of rats with contusion spinal cord injuries: a meta-analysis.

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.

Combined acupuncture and HuangDiSan treatment affects behavior and synaptophysin levels in the hippocampus of senescence-accelerated mouse prone 8 after neural stem cell transplantation.

Sanjiao acupuncture and HuangDiSan can promote the proliferation, migration and differentiation of exogenous neural stem cells in senescence-accelerated mouse prone 8 (SAMP8) mice and can improve learning and memory impairment and behavioral function in dementia-model mice. Thus, we sought to determine whether Sanjiao acupuncture and HuangDiSan can elevate the effect of neural stem cell transplantation in Alzheimer's disease model mice. Sanjiao acupuncture was used to stimulate Danzhong (CV17), Zhongwan (CV12), Qihai (CV6), bilateral Xuehai (SP10) and bilateral Zusanli (ST36) 15 days before and after implantation of neural stem cells (5 × 105) into the hippocampal dentate gyrus of SAMP8 mice. Simultaneously, 0.2 mL HuangDiSan, containing Rehmannia Root and Chinese Angelica, was intragastrically administered. Our results demonstrated that compared with mice undergoing neural stem cell transplantation alone, learning ability was significantly improved and synaptophysin mRNA and protein levels were greatly increased in the hippocampus of mice undergoing both Sanjiao acupuncture and intragastric administration of HuangDiSan. We conclude that the combination of Sanjiao acupuncture and HuangDiSan can effectively improve dementia symptoms in mice, and the mechanism of this action might be related to the regulation of synaptophysin expression.

Neural Stem Cell Transplantation Promotes Functional Recovery from Traumatic Brain Injury via Brain Derived Neurotrophic Factor-Mediated Neuroplasticity.

Traumatic brain injury (TBI) induces cognitive impairments, motor and behavioral deficits. Previous evidences have suggested that neural stem cell (NSC) transplantation could facilitate functional recovery from brain insults, but their underlying mechanisms remains to be elucidated. Here, we established TBI model by an electromagnetic-controlled cortical impact device in the rats. Then, 5 µl NSCs (5.0 × 105/µl), derived from green fluorescent protein (GFP) transgenic mouse, was transplanted into the traumatic brain regions of rats at 24 h after injury. After differentiation of the NSCs was determined using immunohistochemistry, neurological severity scores (NSS) and rotarod test were conducted to detect the neurological behavior. Western blot and RT-PCR as well as ELASA were used to evaluate the expression of synaptophysin and brain-derived neurotrophic factor (BDNF). In order to elucidate the role of BDNF on the neural recovery after NSC transplantation, BDNF knockdown in NSC was performed and transplanted into the rats with TBI, and potential mechanism for BDNF knockdown in the NSC was analyzed using microassay analysis. Meanwhile, BDNF antibody blockade was conducted to further confirm the effect of BDNF on neural activity. As a result, an increasing neurological function improvement was seen in NSC transplanted rats, which was associated with the upregulation of synaptophysin and BDNF expression. Moreover, transplantation of BDNF knockdown NSCs and BDNF antibody block reduced not only the level of synaptophysin but also exacerbated neurological function deficits. Microassay analysis showed that 14 genes such as Wnt and Gsk3-ß were downregulated after BDNF knockdown. The present data therefore showed that BDNF-mediated neuroplasticity underlie the mechanism of NSC transplantation for the treatment of TBI in adult rats.

Enhanced Functional Recovery from Spinal Cord Injury in Aged Mice after Stem Cell Transplantation through HGF Induction.

The number of elderly patients with spinal cord injury (SCI) is increasing worldwide, representing a serious burden for both the affected patients and the community. Previous studies have demonstrated that neural stem cell (NSC) transplantation is an effective treatment for SCI in young animals. Here we show that NSC transplantation is as effective in aged mice as it is in young mice, even though aged mice exhibit more severe neurological deficits after SCI. NSCs grafted into aged mice exhibited better survival than those grafted into young mice. Furthermore, we show that the neurotrophic factor HGF plays a key role in the enhanced functional recovery after NSC transplantation observed in aged mice with SCI. The unexpected results of the present study suggest that NSC transplantation is a potential therapeutic modality for SCI, even in elderly patients.

Transplantation of neural progenitor cells in chronic spinal cord injury.

Previous studies demonstrated that neural progenitor cells (NPCs) transplanted into a subacute contusion injury improve motor, sensory, and bladder function. In this study we tested whether transplanted NPCs can also improve functional recovery after chronic spinal cord injury (SCI) alone or in combination with the reduction of glial scar and neurotrophic support. Adult rats received a T10 moderate contusion. Thirteen weeks after the injury they were divided into four groups and received either: 1. Medium (control), 2. NPC transplants, 3. NPC+lentivirus vector expressing chondroitinase, or 4. NPC+lentivirus vectors expressing chondroitinase and neurotrophic factors. During the 8 weeks post-transplantation the animals were tested for functional recovery and eventually analyzed by anatomical and immunohistochemical assays. The behavioral tests for motor and sensory function were performed before and after injury, and weekly after transplantation, with some animals also tested for bladder function at the end of the experiment. Transplant survival in the chronic injury model was variable and showed NPCs at the injury site in 60% of the animals in all transplantation groups. The NPC transplants comprised less than 40% of the injury site, without significant anatomical or histological differences among the groups. All groups also showed similar patterns of functional deficits and recovery in the 12 weeks after injury and in the 8 weeks after transplantation using the Basso, Beattie, and Bresnahan rating score, the grid test, and the Von Frey test for mechanical allodynia. A notable exception was group 4 (NPC together with chondroitinase and neurotrophins), which showed a significant improvement in bladder function. This study underscores the therapeutic challenges facing transplantation strategies in a chronic SCI in which even the inclusion of treatments designed to reduce scarring and increase neurotrophic support produce only modest functional improvements. Further studies will have to identify the combination of acute and chronic interventions that will augment the survival and efficacy of neural cell transplants.