NGF Expression Research Articles

Shuttle Transfer of mRNA Transcripts via Extracellular Vesicles From Male Reproductive Tract Cells to the Cumulus–Oocyte Complex in Rabbits (Oryctolagus cuniculus)

Semen is known to contain an ovulation-inducing factor (identified as a nerve growth factor, NGF) that shows a significant increase in ovulation after semen deposition in induced ovulatory species. However, the interplay between the male reproductive tract cells and oocyte maturation through messenger RNA (mRNA) cargo is yet to be investigated. Extracellular vesicles (EVs) from the primary culture of rabbit prostate (pEVs), epididymis (eEVs), and testis (tEVs) were isolated to examine their contents for several mRNA transcripts through relative quantitative PCR (RT-qPCR). The expressions of NGF, neurotrophin (NTF3), vascular endothelial growth factor A (VEGFA), A disintegrin and metalloprotease 17 (ADAM17), midkine (MDK), kisspeptin (KISS1), and gonadotrophin-releasing hormone (GNRH1) were examined in isolated EVs. EVs were characterized through transmission electron microscopy. EV uptake by cumulus cell culture was confirmed through microscopic detection of PKH26-stained EVs. Furthermore, the effects of pEVs, eEVs, and tEVs were compared with NGF (10, 20, and 30 ng/ml) supplementation on oocyte in vitro maturation (IVM) and transcript expression. KISS1, NTF3, MDK, ADAM17, GAPDH, and ACTB were detected in all EV types. GNRH1 was detected in tEVs. NGF was detected in pEVs, whereas VEGFA was detected in eEVs. pEVs, eEVs, and 20 ng/ml NGF showed the highest grade of cumulus expansion, followed by tEVs and 10 ng/ml NGF. Control groups and 30 ng/ml NGF showed the least grade of cumulus expansion. Similarly, first polar body (PB) extrusion was significantly increased in oocytes matured with eEVs, pEVs, tEVs, NGF20 (20 ng/ml NGF), NGF10 (10 ng/ml NGF), control, and NGF30 (30 ng/ml NGF). Additionally, the expression of NGFR showed a 1.5-fold increase in cumulus cells supplemented with eEVs compared with the control group, while the expression of PTGS2 (COX2) and NTRK showed 3-fold and 5-fold increase in NGF20-supplemented cumulus-oocyte complexes (COCs), respectively. Oocyte PMP15 expression showed a 1.8-fold increase in IVM medium supplemented with eEVs. Additionally, oocyte NGFR and NTRK expressions were drastically increased in IVM medium supplemented with pEVS (3.2- and 1.6-fold, respectively) and tEVs (4- and 1.7-fold, respectively). This is the first report to examine the presence of mRNA cargo in the EVs of male rabbit reproductive tract cells that provides a model for the stimulation of female rabbits after semen deposition.

ENT-A010, a Novel Steroid Derivative, Displays Neuroprotective Functions and Modulates Microglial Responses

Tackling neurodegeneration and neuroinflammation is particularly challenging due to the complexity of central nervous system (CNS) disorders, as well as the limited drug accessibility to the brain. The activation of tropomyosin-related kinase A (TRKA) receptor signaling by the nerve growth factor (NGF) or the neurosteroid dehydroepiandrosterone (DHEA) may combat neurodegeneration and regulate microglial function. In the present study, we synthesized a C-17-spiro-cyclopropyl DHEA derivative (ENT-A010), which was capable of activating TRKA. ENT-A010 protected PC12 cells against serum starvation-induced cell death, dorsal root ganglia (DRG) neurons against NGF deprivation-induced apoptosis and hippocampal neurons against Aβ-induced apoptosis. In addition, ENT-A010 pretreatment partially restored homeostatic features of microglia in the hippocampus of lipopolysaccharide (LPS)-treated mice, enhanced Aβ phagocytosis, and increased Ngf expression in microglia in vitro. In conclusion, the small molecule ENT-A010 elicited neuroprotective effects and modulated microglial function, thereby emerging as an interesting compound, which merits further study in the treatment of CNS disorders.

Localization of Nerve Growth Factor Expression to Structurally Damaged Cartilaginous Tissues in Human Lumbar Facet Joint Osteoarthritis.

PurposeNerve Growth Factor (NGF) is a pivotal mediator of chronic pain and plays a role in bone remodelling. Through its high affinity receptor TrkA, NGF induces substance P (SP) as key downstream mediator of pain and local inflammation. Here we analysed NGF, TrkA and SP tissue distribution in facet joint osteoarthritis (FJOA), a major cause of chronic low back pain.MethodsFJOA specimens (n=19) were harvested from patients undergoing intervertebral fusion surgery. Radiologic grading of FJOA and spinal stenosis, followed by immunohistochemistry for NGF, TrkA and SP on consecutive tissue sections, was performed in ten specimens. Explant cultures (n=9) were used to assess secretion of NGF, IL-6, and SP by FJOA osteochondral tissues under basal and inflammatory conditions.ResultsNGF was predominantly expressed in damaged cartilaginous tissues (80%), occasionally in bone marrow (20%), but not in osteochondral vascular channels. NGF area fraction in cartilage was not associated with the extent of proteoglycan loss or radiologic FJOA severity. Consecutive sections showed that NGF and SP expression was localized at structurally damaged cartilage, in absence of TrkA expression. SP and TrkA were expressed in subchondral bone marrow in both presence and absence of NGF. Low level NGF, but not SP secretion, was detected in four out of eighteen FJOA explants under both basal or inflammatory conditions (n=2 each).ConclusionNGF is associated with SP expression and structural cartilage damage in osteoarthritic facet joints, but not with radiologic disease severity. NGF tissue distribution in FJOA differs from predominant subchondral bone expression reported for knee OA.

Preconditioning with secretome of neural crest-derived stem cells enhanced neurotrophic expression in mesenchymal stem cells

During the last 20 years, stem cell therapy has been considered as an effective approach for regenerative medicine. Due to poor ability of stem cells to survive following transplantation, it has been proposed that beneficial effects of stem cells mainly depend on paracrine function. Therefore, the present study was designed to reinforce mesenchymal stem cells (MSCs) to express higher levels of trophic factors especially the ones with the neurotrophic properties. Here, bone marrow (BM)-MSCs and adipose-MSCs were treated with conditioned medium (CM) of dental pulp stem cells (DPSCs) or hair follicle stem cells (HFSCs) for up to three days. The relative expression of five key trophic factors that have critical effects on the central nervous system regeneration were evaluated using qRT-PCR technique. Furthermore, to assess the impacts of conditioned mediums on the fate of MSCs, expression of seven neuronal/glial markers were evaluated 3 days after the treatments. The obtained data revealed priming of BM-MSCs with HFSC-CM or DPSC-CM increases the BDNF expression over time. Such effect was also observed in adipose-MSCs following DPSC-CM treatment. Secretome preconditioning remarkably increased NGF expression in the adipose-MSCs. In addition, although priming of adipose-MSCs with HFSC-CM increased GDNF expression one day after the treatment, DPSC-CM enhanced GDNF mRNA in BM-MSCs at a later time point. It seemed priming of BM-MSCs with HFSC-CM, promoted differentiation into the glial lineage. Our findings showed that MSCs preconditioning with secretome of neural crest-derived stem cells could be a promising approach to enhance the neurotrophic potential of these stem cells.

Gastrodin injection alleviates lung injury caused by focal cerebral ischemia in rats through NGF/TrkA pathway-mediated activation of the anti-inflammatory pathway

To investigate the therapeutic mechanism of gastrodin injection for alleviating lung injury caused by focal cerebral ischemia in rats and the role of the NGF-TrkA pathway in mediating this effect. Forty SD rats were equally randomized into normal group, sham-operated group, model group and gastrodin group, and in the latter two groups, rat models of focal cerebral ischemia were established by embolization of the right middle cerebral artery. After successful modeling, the rats were treated with intraperitoneal injection of gastrodin injection at the daily dose of 10 mg/kg for 14 days. After the treatment, the wet/dry weight ratio of the lung tissue was determined, the pathological changes in the lung tissue were observed using HE staining, and the levels of IL-10 and TNF-α in the arterial blood were detected with ELISA. The expressions of NF-κB p65 and TNF-α in the lung tissue were detected with Western blotting, and the expressions of NGF and TrkA were detected using immunohistochemical staining and Western blotting. Compared with the normal control and sham-operated groups, the rats in the model group showed obvious inflammatory lung injury, significantly increased wet/ dry weight ratio of the lungs (P < 0.01), increased TNF-α level in arterial blood (P < 0.01), and significantly up-regulated protein expressions of NF-κB p65 (P < 0.01), TNF-α (P < 0.01), NGF (P < 0.05) and TrkA(P < 0.05) in the lung tissue. Treatment with gastrodin injection obviously alleviated lung inflammation, decreased the wet/dry weight ratio of the lungs (P < 0.05), and significantly lowered TNF-α level (P < 0.01) and increased IL-10 level in the arterial blood in the rat models (P < 0.01); gastrodin injection also significantly decreased the protein expressions of NF-κB p65 and TNF-α (P < 0.05) and up-regulated the expressions of NGF and TrkA in the lung tissue of the rats (P < 0.05). The NGF/TrkA pathway may participate in cerebral ischemia-induced inflammatory lung injury, which can be obviously alleviated by gastrodin through the activation of the anti-inflammatory pathway mediated by the NGF/TrkA pathway.

Demonstration of the protective effect of propofol in rat model of cisplatin-induced neuropathy.

Cisplatin is commonly used in the treatment of lung, genitourinary, and gastrointestinal cancers. Peripheral neuropathy is the most important side effect, leading to a decrease in the dose of cisplatin or its complete cessation in the early period. 16 rats were given cisplatin at a dose of 2.5 mg/ kg/day twice a week for 4 weeks to induce neuropathy model. The rats taking Cisplatin were divided into 2 groups. Group 1 rats (n = 8) were given 1 ml/kg/day 0.9 % NaCl intraperitoneally, and Group 2 rats were given 10 mg/kg/day Propofol intraperitoneally daily for 4 weeks. The remaining 8 rats served as the control group. At the end of the study, all animals were tested for motor functions. Blood samples were collected for the measurement of plasma lipid peroxidation (malondialdehyde; MDA), tumor necrosis factor (TNF-α), glutathione (GSH), IL-6 and HSP-70 levels. Electromyography findings revealed that compound muscle action potential (CMAP) amplitude was significantly higher in the cisplatin-Propofol group than in the cisplatin-saline group. Also, cisplatin-Propofol treated group showed significantly lower TNF-α, MDA and IL-6 levels and higher GSH and HSP-70 levels than cispalatin-Saline group (p < 0.01, p < 0.001). In addition, while the CMAP latency was decreased in the propofol group, the CMAP amplitude was increased, and a significant improvement was observed in the Inclined test scores. Besides, histological examinations showed an increase in axon diameter and NGF expression with Propofol treatment. This study demonstrated that Propofol exerts protective activity against cisplatin-induced neurotoxicity by increasing endogenous antioxidants and reducing lipid peroxidation and inflammation (Tab. 3, Fig. 4, Ref. 30). Text in PDF www.elis.sk Keywords: cisplatin, neuropathy, propofol, oxidative damage, ınflammation.

The effects of Anisodamine-Tirofiban Combined Therapy in acute myocardial infarction treated with Percutaneous Coronary Intervention (PCI).

Objectives:To study the effects of anisodamine-tirofiban combined therapy on cardiac function and serological expression of serum NGF and ESM-1 in patients with acute myocardial infarction treated with percutaneous coronary intervention (PCI).Methods:Eighty patients with myocardial infarction treated in Cangzhou Medical College, Hebei, China from February 2015 to April 2017 were selected and divided into the control group and the research group according to the principle of random draw, 40 patients per group. The patients in the control group received symptomatic routine treatment, while the patients in the research group received anisodamine-tirofiban combined therapy on the top of symptomatic routine treatment. Differences between the two groups in TIMI flow grades, cardiac function, levels of NGF and ESM-1 and adverse response were observed.Results:The recovery of cardiac function in the research group was statistically significant with P value (p<0.05) and better than the control group in TIMI flow grades, myocardial perfusion capacity and cardiac function. The serological indicators in the research group had a higher level of NGF and a lower level of ESM-1 than the control group, and the differences were statistically significant (p<0.05). In terms of safety, neither group showed significant hepatorenal disorders.Conclusion:The combined treatment of anisodamine-tirofiban in patients with acute myocardial infarction after percutaneous coronary intervention (PCI) can recover NGF and ESM-1 related proteins, improve postoperative myocardial perfusion, and accelerate the recovery of cardiac function. It is worth promoting in clinic.

Abstract 10766: Sovateltide Induces Neuroprotection in a Neonatal Rat Model of Hypoxic Ischemic Encephalopathy

Introduction: Hypoxic-ischemic encephalopathy(HIE) is a major cause of morbidity, mortality, and severe neurodevelopmental disability in term neonates. The development of a treatment strategy that could provide neuroprotection is still required. We have shown that agonism of endothelin B(ETB) receptors mediated through sovateltide provides neuroprotection and enhances neurovascular remodeling in several neuropathological conditions. Hypothesis: We hypothesize that administering sovateltide alone or as an adjuvant therapy with hypothermia could provide neuroprotection by reducing oxidative stress and promoting cell survival post-HIE brain injury. Methods: Sprague-Dawley male and female rat pups were grouped separately and were divided into 5 different subgroups (1) Control; (2) HIE+Vehicle; (3) HIE+Hypothermia; (4) HIE+sovateltide; and (5) HIE+sovateltide+hypothermia. HIE was induced by ligating the right carotid artery on a postnatal day (PND) 7, followed by hypoxia for 120 min and hypothermia (32-34°C) for 5 h. On PND 7, sovateltide (5 μg/kg, ICV) was injected after hypoxic-ischemic injury. Pups were euthanized on PND 10. Brains were isolated, and ETB receptors, VEGF, NGF, cell death and oxidative stress makers - malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD) were assessed. Results: Animals receiving sovateltide showed a significant (p&lt;0.0001) upregulation of ETB receptor (135%), VEGF (110%), and NGF (183%) expression in the brain compared to vehicle. Vehicle-treated animals had high oxidative stress levels, as indicated by increased lipid peroxidation, MDA, and decreased antioxidants, SOD, and GSH compared to control. These effects were reversed (p&lt;0.001) in sovateltide alone or in combination with the hypothermia, indicating reduced oxidative stress in these animals. Additionally, sovateltide alone or in combination with hypothermia reduced (p&lt;0.001) cell death compared to vehicle or hypothermia alone. Similar findings were obtained in both male and female rats. Conclusion: Sovateltide alone or as an adjunct to hypothermia exerts neuroprotective effects in HIE-induced brain injury in rat pups, suggesting sovateltide could be a potential therapeutic agent for neonatal HIE.

What's New in Musculoskeletal Basic Science.

What's New in Musculoskeletal Basic Science.

Bone Marrow Mesenchymal Stem Cells (BMSCs) Promote Neuronal Cell Repair in Spinal Cord Injury by Regulating Toll-Like Receptor 4/Nuclear Factor-Kappa B Signaling Pathway

SCI (SCI) poses a challenge to nerve cell repair strategies. SCI injury can lead to the development of inflammation, which in turn can exacerbate nerve cell damage. The TLR4/NF-kappa B signaling pathway is a common inflammatory signaling pathway. Since BMSCs are involved in injury repair, whether they can promote the repair of SCI neuronal cells have not been reported. Spinal cord nerve cells were cultured in vitro and divided into mechanical injury group and BMSCs group followed by analysis of cell proliferation activity and detection of altered apoptotic activity. Changes in the concentrations of IL-6 and IL-1β were measured by ELISA and cellular mitochondrial alterations was assessed by JG-B staining along with analysis of NF-kappa B, TLR4, related neurodevelopmental factor BDNF, and NGF expression by western blot. Mechanical damage to neuronal cells resulted in decreased cell proliferation, increased apoptotic activity, decreased cellular mitochondrial activity, increased TLR4 and NF-kappa B expression, decreased BDNF and NGF expression, as well as increased secertions of IL-6 and IL-1β (P &lt; 0.05). In contrast, co-culture with BMSCs resulted in increased proliferation and decreased apoptosis of mechanically injured neuronal cells, increased cellular mitochondrial activity, with observation of the inverse changes in other factors (P &lt; 0.05). In conclusion, BMSCs can suppress inflammation and promote repair of injured neuronal cells by inhibiting TLR4/NF-kappa B signaling.

Cartilage tissue miR-214-3p regulates the TrkB/ShcB pathway paracrine VEGF to promote endothelial cell migration and angiogenesis.

This study was designed to explore the mechanisms through which chondrocytes regulated endothelial cell migration and angiogenesis in osteoarthritis (OA). The expressions of related genes of OA were detected by Western blot and real-time quantitative PCR. Chondrocytes were co-cultured with endothelial cells, and migration as well as angiogenesis rates, and vascular endothelial growth factor (VEGF) secretion of the cells were detected. The relationship between miRNA and TrkB were analyzed by bioinformatics analysis, RNA immunoprecipitation and dual-luciferase assays. The effects of miRNA on the histopathology of the OA mice were determined. The expressions of NGF, TrkA, TrkB, and ShcB were increased significantly in OA patients. IL-1β promoted the expressions of TrkA, TrkB, and ShcB in chondrocytes and inhibited the expressions of chondrogenic differentiation markers, but shTrkB partially reversed IL-1β-mediated chondrogenic differentiation. Overexpression of TrkB promoted cell migration, angiogenesis, and VEGF levels, while silencing ShcB reversed the regulation of TrkB. Moreover, chondrocytes miR-214-3p regulated endothelial cell migration and angiogenesis by targeting TrkB paracrine VEGF to activate PI3K/Akt pathway proteins. In addition, overexpressed miR-214-3p improved collagenase-induced cartilage and synovial damage in OA mice. The activation of TrkB/ShcB signaling pathway paracrine VEGF is mediated by miR-214-3p in chondrocytes and it regulates endothelial cell migration and angiogenesis in the development of OA.

Comprehensive Nursing can Relieve the Negative Emotions of Patients undergoing Breast Cancer Resection and Reduce Postoperative

To explore the effect of comprehensive nursing on adverse emotions and postoperative complications of breast cancer patients undergoing mastectomy. Altogether 180 patients who received treatment in our hospital from May 2017 to May 2019 were selected as the research participants and divided into group A and group B. Among them, 100 cases in group A received comprehensive nursing, 80 cases in group B received routine nursing. The surgical indications, upper limb function, serum NGF, TK1 and CA15-3 expression level, VAS score, SAS, SDS score, quality of life SF-36 score were detected, and the incidence rate of postoperative complications and nursing quality score were compared. Compared with group B, group A had less postoperative bed time, intraoperative blood loss, length of hospital stay, better recovery of upper limb function, lower expression levels of serum NGF, TK1 and CA15-3, lower VAS score, SAS and SDS score, higher quality of life SF-36 score, lower incidence of postoperative complications and higher nursing quality score. Comprehensive nursing can relieve the negative emotions of patients undergoing breast cancer resection and reduce the incidence of postoperative complications.

NGF Expression and Elevation in Hip Osteoarthritis Patients with Pain and Central Sensitization.

Osteoarthritis (OA) is a chronic degenerative musculoskeletal disease that causes articular cartilage degeneration and chronic pain. Research into OA animal models suggests that elevated NGF levels in the synovium contribute to pain and central sensitization (CS). However, it is unclear whether synovial NGF contributes to CS in patients with OA. We investigated the association between synovial NGF expression and clinical assessments of pain and CS in hip OA (hOA) patients. We also aimed to identify which cells in the synovium of hOA patients express NGF. Sixty-six patients who received total hip replacement and a diagnosis of hOA were enrolled. We measured NGF mRNA expression in synovial samples obtained from 50 patients using qPCR and analyzed the correlation of NGF expression with the CS inventory (CSI) score and Japanese Orthopaedic Association (JOA) score, a clinical scoring system for OA. To identify the synovial cells expressing NGF, we analyzed NGF mRNA expression in CD14+ and CD14- cells, which represent macrophage-rich and fibroblast-rich fractions, respectively, extracted from 8 patients. To further identify which macrophage subtypes express NGF, we examined NGF mRNA expression in CD14high and CD14low cells sorted from 8 patients. Synovial NGF mRNA expression was negatively correlated with JOA score but positively correlated with CSI score (JOA pain, r = −0.337, P = 0.017; CSI score, r = 0.358, P = 0.011). Significantly greater levels of NGF were observed in CD14- cells compared to CD14+ cells (P = 0.036) and in CD14high cells compared to CD14low cells (P = 0.008). In conclusion, synovial NGF expression is correlated with the degree of pain and CS in hOA patients. NGF is predominantly expressed in synovial fibroblasts. Further, CD14high synovial macrophages expressed higher levels of NGF. Our results may provide a novel NGF-targeted therapeutic strategy for hOA pain.

The Effect of Valproic Acid on the Transcriptional Activity of Ngf and Bdnf Genes of in Vitro Cultured Neurons Under Oxidative Stress Conditions

Brain-derived neurotrophic factor (BDNF) is a secretory molecule that promotes peripheral neurons synaptic transmission and plasticity by TrkB receptor activation. This is shown in cultured central nervous system (CNS) neurons, including hippocampal and cortical cholinergic, dopaminergic and serotonergic neurons. Hypotheses suggesting that BDNF may play a potential role in the pathophysiology of schizophrenia are based on the key role of BDNF in the synaptic plasticity and, consequently, regulation of cognitive functions. In the schizophrenia treatment valproic acid is used in complex combined therapy regimens. Treatment of schizophrenia patients with valproate increases the BDNF level. Since it is not yet clear whether the BDNF protein levels measured in serum samples and in the brain correlate, we investigated valproate effects on the cultured neurons Bdnf transcription level. The primary neuron-glia culture was obtained from the cerebellum of 8-9-day-old Wistar rats. Valproic acid was added to the neurons (at a concentration of 50 µg/ml), oxidative stress was stimulated by 40 µMof H2O2, and injury was caused by mechanical damage to the neuron culture. It was shown that valproic acid in 3-24 hours increases the transcriptional activity of the Bdnf and Ngf (nerve growth factor) genes 2–2.5-fold (p&lt;0.01) and approximately 1.5-fold (p&lt;0.01), respectively. Mechanical trauma, unlike oxidative stress, activates the transcriptional activity of the Ngf and Bdnf genes (p&lt;0.01). However, under oxidative stress and mechanical damage to neurons, the effect of valproic acid on the Ngf and Bdnf genes expression was insignificant. Fluorescence microscopy analysis using specific antibodies to neurons (anti-Map-2) showed that in the presence of valproic acid, the number of neuronal processes and contacts between them significantly increased. Evidently, valproate addition to antipsychotics can be effective for the overall clinical response. Relatively little research has been done on the signaling pathways in neurons that are activated by the valproic acid. However, we have obtained evidence of activation of the Ngf and Bdnf genes transcription in cultured neurons in vitro. We also found that in the presence of valproic acid, the number of neuronal processes and contacts between them significantly increased. However, we have also found that the oxidative stress accompanying the schizophrenia can significantly reduce the valproic acid effect on the Ngf and Bdnf genes expression. The results of the study may be potentially useful for new schizophrenia therapy strategies development.

The neuro-restorative effect of adipose-derived mesenchymal stem cell transplantation on a mouse model of diabetic neuropathy

ABSTRACT Diabetic neuropathy (DN) is the most common degenerative complication associated with Diabetes Mellitus. Despite widespread awareness about DN, the only effective treatments are blood glucose control and pain management. The aim of the current study was to determine the effect of intramuscular adipose-derived mesenchymal stem cell (AMSC) transplantation on sciatic nerves in DN using EMG and histological analyses. A total of 27 mice were randomly divided into three groups: control group, DN group and AMSC group. In EMG, CMAP amplitude in the sciatic nerves was lower, but distal latency was higher in the DN group compared with the control group. CMAP amplitude in the sciatic nerves was higher in the AMSC group compared with the DN group. Distal latency in the sciatic nerve was lower in the AMSC group compared with the DN group. Histologic examination of the tissues in the animals treated with AMSC showed a remarkable improvement in microscopic morphology. Fluorescence microscopy analyses demonstrated that intramuscularly transplanted AMSC was selectively localized in the sciatic nerves. Transplantation of AMSC increased protein expression of S100, cdk2, NGF and DHH, all of which, interfered with DN onset in sciatic nerves. The findings of the present study suggest that AMSC transplantation improved DN through a signal-regulatory effect on Schwann cells, neurotrophic actions and restoration of myelination.

Comparisons of Neurotrophic Effects of Mesenchymal Stem Cells Derived from Different Tissues on Chronic Spinal Cord Injury Rats.

Cell-based therapies with mesenchymal stem cells (MSCs) are considered as promising strategies for spinal cord injury (SCI). MSCs have unique characteristics due to differences in the derived tissues. However, relatively few studies have focused on differences in the therapeutic effects of MSCs derived from different tissues. In this study, the therapeutic effects of adipose tissue-derived MSCs, bone marrow-derived MSCs, and cranial bone-derived MSCs (cMSCs) on chronic SCI model rats were compared. MSCs were established from the collected adipose tissue, bone marrow, and cranial bone. Neurotrophic factor expression of each MSC type was analyzed by real-time PCR. SCI rats were established using the weight-drop method and transplanted intravenously with MSCs at 4 weeks after SCI. Hindlimb motor function was evaluated from before injury to 4 weeks after transplantation. Endogenous neurotrophic factor and neural repair factor expression in spinal cord (SC) tissue were examined by real-time PCR and western blot analyses. Although there were no differences in the expression levels of cell surface markers and multipotency, expression of Bdnf, Ngf, and Sort1 (Nt-3) was relatively higher in cMSCs. Transplantation of cMSCs improved motor function of chronic SCI model rats. Although there was no difference in the degree of engraftment of transplanted cells in the injured SC tissue, transplantation of cMSCs enhanced Bdnf, TrkB, and Gap-43 messenger RNA expression and synaptophysin protein expression in injured SC tissue. As compared with MSCs derived other tissues, cMSCs highly express many neurotrophic factors, which improved motor function in chronic SCI model rats by promoting endogenous neurotrophic and neural plasticity factors. These results demonstrate the efficacy of cMSCs in cell-based therapy for chronic SCI.

Bone Marrow Mesenchymal Stem Cell Condition Medium Loaded on PCL Nanofibrous Scaffold Promoted Nerve Regeneration After Sciatic Nerve Transection in Male Rats.

Nowadays, researchers pay a vast deal of attention to neural tissue regeneration due to its tremendous effect on the patient's life. There are many strategies, from using conventional autologous nerve grafts to the newly developed methods for reconstructing damaged nerves. Among the various therapeutic methods, incorporating highly potent biomolecules and growth factors, the damaged nerve site would promote nerve regeneration. The aim was to examine the efficiency of a mesenchymal stem cell condition medium (MSC-CM) loaded on a 3D-polycaprolactone (PCL) scaffold as a nerve conduit in an axotomy rat model. Twenty-four mature male rats were classified into four groups: controls (the animals of this group were intact), axotomy (10mm piece of the nerve was removed), axotomy (10-mm piece of the nerve was removed) + scaffold, and axotomy (10-mm piece of the nerve was removed) + MSC-CM-loaded scaffold. We followed up nerve motor function using a sciatic function index and electromyography activity of the gastrocnemius muscle. At 12weeks post axotomy, sciatic nerve and dorsal root ganglion specimens and L4 and L5 spinal cord segments were separated from the rats and were analyzed by stereological, immunohistochemistry, and RT-PCR procedures. The rats of the axotomy group presented the expected gross locomotor deficit. Stereological parameters, immunohistochemistry of GFAP, and gene expression of S100, NGF, and BDNF were significantly enhanced in the CM-loaded scaffold group compared with the axotomy group. The most observed similarity was noted between the results of the control group and the CM-loaded scaffold group. Our results support the potential applicability of MSC-CM-loaded PCL nanofibrous scaffold to treat peripheral nerve injury (PNI).

TGF-\u03b2 regulates nerve growth factor expression in a mouse intervertebral disc injury model

BackgroundIntervertebral disc (IVD) degeneration is a major cause of low back pain (LBP). Following disc injury, nerve growth factor (NGF) concentrations rise in IVDs, and anti-NGF therapy has been shown to attenuate LBP in humans. Increased levels of tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β) in degenerative IVDs and in in vitro studies suggest that these factors promote NGF production. However, whether these factors regulate NGF in vivo remains unclear. Thus, we studied NGF regulation in a mouse model of IVD injury.MethodsAfter inducing IVD injury, we examined mRNA levels of Tnfa, Tgfb, and Ngf in IVDs from control and IVD-injured mice across 7 days. To do this, we used magnetic cell separation to isolate CD11b ( +) (macrophage-rich) and CD11b (-) (IVD cell-rich) cell fractions from injured IVDs. To study the effect of TNF-α on Ngf expression, we examined Ngf expression in injured IVDs from C57BL/6 J and Tnfa-knockout (KO) mice (C57BL/6 J background). To study the effect of TGF-β on Ngf expression, C57/BL6J mice were given an intraperitoneal injection of either the TGF-β inhibitor SB431542 or DMSO solution (vehicle) one and two days before harvesting IVDs.ResultsmRNA expression of Tnfa, Tgfb, and Ngf was significantly increased in injured IVDs. Tnfa was predominantly expressed in the CD11b ( +) fraction, and Tgfb in the CD11b (-) fraction. Ngf expression was comparable between CD11b ( +) and CD11b (-) fractions, and between wild-type and Tnfa-KO mice at post-injury day (PID) 1, 3, and 7. SB431542 suppressed TGF-β-mediated Ngf expression and NGF production in vitro. Further, administration of SB431542 significantly reduced Ngf expression in IVDs such that levels were below those observed in vehicle-treated animals at PID3 and PID7.ConclusionA TGF-β inhibitor reduced Ngf expression in a mouse model of IVD injury, suggesting that TGF-β may regulate NGF expression in vivo.

The ERK/CREB/PTN/syndecan-3 pathway involves in heparin-mediated neuro-protection and neuro-regeneration against cerebral ischemia-reperfusion injury following cardiac arrest

BackgroundHeparin, a commonly used anticoagulant, has been found to improve cerebral ischemia-reperfusion injury (CIR-CA) following cardiopulmonary resuscitation (CPR). Here, we aimed to explore the role of pleiotrophin (PTN)/syndecan-3 pathway in heparin therapy for CIR-CA. Materials and methodsThe CA-CPR model was constructed in Sprague-Dawley (SD) rats, which were treated with low molecular weight heparin, and the neurological changes and brain histopathological changes were evaluated. For in-vitro experiments, the ischemic injury model of primary neurons was established by oxygen and glucose deprivation (OGD), and the neuron regeneration was detected via the Cell counting Kit-8 (CCK8) method, flow cytometry and microscopy. CREB antagonist (KG-501), ERK antagonist (PD98059) and si-PTN were used respectively to inhibit the expression of CREB, ERK and PTN in cells, so as to explore the role of heparin in regulating neuronal regeneration. ResultsCompared with the sham rats, the neurological deficits and cerebral edema of CA-CPR rats were significantly improved after heparin treatment. Heparin also attenuated OGD-mediated neuronal apoptosis and promoted neurite outgrowth in vitro. Moreover, heparin attenuated CA-CPR-mediated neuronal apoptosis and microglial neuroinflammation. In terms of the mechanism, heparin upregulated the expression of ERK, CREB, NF200, BDNF, NGF, PTN and syndecan-3 in the rat brains. Inhibition of ERK, CREB and interference with PTN expression notably weakened the heparin-mediated neuroprotective effects and restrained the expression of ERK/CREB and PTN/syndecan-3 pathway. ConclusionHeparin attenuates the secondary brain injury induced by CA-CPR through regulating the ERK/CREB-mediated PTN/syndecan-3 pathway.

Hyperoside Attenuate Inflammation in HT22 Cells via Upregulating SIRT1 to Activities Wnt/β-Catenin and Sonic Hedgehog Pathways.

Neuroinflammation plays important roles in the pathogenesis and progression of altered neurodevelopment, sensorineural hearing loss, and certain neurodegenerative diseases. Hyperoside (quercetin-3-O-β-D-galactoside) is an active compound isolated from Hypericum plants. In this study, we investigate the protective effect of hyperoside on neuroinflammation and its possible molecular mechanism. Lipopolysaccharide (LPS) and hyperoside were used to treat HT22 cells. The cell viability was measured by MTT assay. The cell apoptosis rate was measured by flow cytometry assay. The mRNA expression levels of interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-8 (IL-8), and tumor necrosis factor-α (TNF-α) were determined by quantitative reverse transcription polymerase chain reaction. The levels of oxidative stress indices superoxide dismutase (SOD), reactive oxygen species (ROS), catalase (CAT), glutathione (GSH), and malondialdehyde (MDA) were measured by the kits. The expression of neurotrophic factor and the relationship among hyperoside, silent mating type information regulation 2 homolog-1 (SIRT1) and Wnt/β-catenin, and sonic hedgehog was examined by western blotting. In the LPS-induced HT22 cells, hyperoside promotes cell survival; alleviates the level of IL-1β, IL-6, IL-8, TNF-α, ROS, MDA, Bax, and caspase-3; and increases the expression of CAT, SOD, GSH, Bcl-2, BDNF, TrkB, and NGF. In addition, hyperoside upregulated the expression of SIRT1. Further mechanistic investigation showed that hyperoside alleviated LPS-induced inflammation, oxidative stress, and apoptosis by upregulating SIRT1 to activate Wnt/β-catenin and sonic hedgehog pathways. Taken together, our data suggested that hyperoside acts as a protector in neuroinflammation.