Overexpression Of Nerve Growth Factor Research Articles

Acceleration of bone repairation by BMSCs overexpressing NGF combined with NSA and allograft bone scaffolds

BackgroundRepairation of bone defects remains a major clinical problem. Constructing bone tissue engineering containing growth factors, stem cells, and material scaffolds to repair bone defects has recently become a hot research topic. Nerve growth factor (NGF) can promote osteogenesis of bone marrow mesenchymal stem cells (BMSCs), but the low survival rate of the BMSCs during transplantation remains an unresolved issue. In this study, we investigated the therapeutic effect of BMSCs overexpression of NGF on bone defect by inhibiting pyroptosis.MethodsThe relationship between the low survival rate and pyroptosis of BMSCs overexpressing NGF in localized inflammation of fractures was explored by detecting pyroptosis protein levels. Then, the NGF+/BMSCs-NSA-Sca bone tissue engineering was constructed by seeding BMSCs overexpressing NGF on the allograft bone scaffold and adding the pyroptosis inhibitor necrosulfonamide(NSA). The femoral condylar defect model in the Sprague–Dawley (SD) rat was studied by micro-CT, histological, WB and PCR analyses in vitro and in vivo to evaluate the regenerative effect of bone repair.ResultsThe pyroptosis that occurs in BMSCs overexpressing NGF is associated with the nerve growth factor receptor (P75NTR) during osteogenic differentiation. Furthermore, NSA can block pyroptosis in BMSCs overexpression NGF. Notably, the analyses using the critical-size femoral condylar defect model indicated that the NGF+/BMSCs-NSA-Sca group inhibited pyroptosis significantly and had higher osteogenesis in defects.ConclusionNGF+/BMSCs-NSA had strong osteogenic properties in repairing bone defects. Moreover, NGF+/BMSCs-NSA-Sca mixture developed in this study opens new horizons for developing novel tissue engineering constructs.

Targeting nerve growth factor-mediated osteosarcoma metastasis: mechanistic insights and therapeutic opportunities using larotrectinib

Osteosarcoma (OS) therapy presents numerous challenges, due largely to a low survival rate following metastasis onset. Nerve growth factor (NGF) has been implicated in the metastasis and progression of various cancers; however, the mechanism by which NGF promotes metastasis in osteosarcoma has yet to be elucidated. This study investigated the influence of NGF on the migration and metastasis of osteosarcoma patients (88 cases) as well as the underlying molecular mechanisms, based on RNA-sequencing and gene expression data from a public database (TARGET-OS). In osteosarcoma patients, the expression of NGF was significantly higher than that of other growth factors. This observation was confirmed in bone tissue arrays from 91 osteosarcoma patients, in which the expression levels of NGF and matrix metallopeptidase-2 (MMP-2) protein were significantly higher than in normal bone, and strongly correlated with tumor stage. In summary, NGF is positively correlated with MMP-2 in human osteosarcoma tissue and NGF promotes osteosarcoma cell metastasis by upregulating MMP-2 expression. In cellular experiments using human osteosarcoma cells (143B and MG63), NGF upregulated MMP-2 expression and promoted wound healing, cell migration, and cell invasion. Pre-treatment with MEK and ERK inhibitors or siRNA attenuated the effects of NGF on cell migration and invasion. Stimulation with NGF was shown to promote phosphorylation along the MEK/ERK signaling pathway and decrease the expression of microRNA-92a-1-5p (miR-92a-1-5p). In in vivo experiments involving an orthotopic mouse model, the overexpression of NGF enhanced the effects of NGF on lung metastasis. Note that larotrectinib (a tropomyosin kinase receptor) strongly inhibited the effect of NGF on lung metastasis. In conclusion, it appears that NGF promotes MMP-2-dependent cell migration by inhibiting the effects of miR-92a-1-5p via the MEK/ERK signaling cascade. Larotrectinib emerged as a potential drug for the treatment of NGF-mediated metastasis in osteosarcoma.

Association of Uterine Tissue Innervation and Peripheral Nerve Density with Adenomyosis Related Pain. A Systematic Review.

Adenomyosis is associated with dysmenorrhea and chronic pelvic pain; however, the triggering mechanisms of painful stimuli and the role of uterine nerve fibers in the manifestation of pain remain poorly understood. The objective of this study was to systematically review the role of uterine nerve fibers' presence and density in the occurrence of pain in patients with adenomyosis. An electronic search was performed using the Embase, PubMed/Medline, and Cochrane databases. We included all studies from inception to November 2023. A total of ten studies that compared uterine biopsies samples of women with and without adenomyosis were included. The biomarker antiprotein gene product 9.5 was decreased or absent in the endometrium of most included women with adenomyosis. None of the included studies observed a difference in neurofilament (NF) staining between the adenomyosis and non-adenomyosis groups. Studies that assessed nerve growth factor (NGF) staining were heterogeneous in design. One study reported no difference in immunohistochemistry staining in any endometrial layer between the adenomyosis and non-adenomyosis groups, while another reported increased staining in the adenomyosis functional endometrial layer, and a third study reported overexpression of NGF, synaptophysin (SYN), and microtubule-associated protein 2 mRNA in focal adenomyosis alone. Preliminary data from poor-quality studies suggest an increase in the uterine density of nerve fibers in patients with adenomyosis. Well-designed studies are essential to assess the cause-and-effect relationship between uterine nerve fibers and pain in patients with adenomyosis.

Innervation pattern of the unclosed detrusor muscle in classic bladder exstrophy: a study of patients with urothelial overexpression of nerve growth factor

PurposeAn overexpression of nerve growth factor (NGF) in the urothelium is discussed to lead to neuronal hyperinnervation of the bladder detrusor. The aim was to assess the sensory and sympathetic innervation of the detrusor in unclosed exstrophic bladders patients with known overexpression of NGF in the urothelium.MethodsFull-thickness bladder biopsies were prospectively obtained from 34 infants at delayed primary bladder closure between 01/2015 and 04/2020. The bladder biopsies were immunohistochemically stained with antibodies against S100, calcitonin gene-related peptide (anti-CGRP), Neurofilament 200 (anti-NF200), and tyrosine-hydroxylase (anti-TH). Specimens from 6 children with congenital vesicoureterorenal reflux (VUR) served as controls.ResultsThere was no statistically significant difference in nerve fiber density in any of the immunohistochemical assessments (anti-S100 [p = 0.210], anti-CGRP [p = 0.897], anti-NF200 [p = 0.897]), and anti-TH [p = 0.956]) between patients with BE and patients with VUR. However, we observed a trend toward lower nerve fiber densities in exstrophic detrusor.ConclusionOverall our results showed an unharmed innervation pattern in this cohort but a lower density of nerve fibers in the detrusor compared to controls. Further studies in patients after successful primary closure are needed to clarify the potential impact of the urothelial overexpression of NGF modulating the innervation pattern in exstrophic bladders.

Generation of Periventricular Reactive Astrocytes Overexpressing Aquaporin 4 Is Stimulated by Mesenchymal Stem Cell Therapy.

Aquaporin-4 (AQP4) plays a crucial role in brain water circulation and is considered a therapeutic target in hydrocephalus. Congenital hydrocephalus is associated with a reaction of astrocytes in the periventricular white matter both in experimental models and human cases. A previous report showed that bone marrow-derived mesenchymal stem cells (BM-MSCs) transplanted into the lateral ventricles of hyh mice exhibiting severe congenital hydrocephalus are attracted by the periventricular astrocyte reaction, and the cerebral tissue displays recovery. The present investigation aimed to test the effect of BM-MSC treatment on astrocyte reaction formation. BM-MSCs were injected into the lateral ventricles of four-day-old hyh mice, and the periventricular reaction was detected two weeks later. A protein expression analysis of the cerebral tissue differentiated the BM-MSC-treated mice from the controls and revealed effects on neural development. In in vivo and in vitro experiments, BM-MSCs stimulated the generation of periventricular reactive astrocytes overexpressing AQP4 and its regulatory protein kinase D-interacting substrate of 220 kDa (Kidins220). In the cerebral tissue, mRNA overexpression of nerve growth factor (NGF), vascular endothelial growth factor (VEGF), hypoxia-inducible factor-1 (HIF1α), and transforming growth factor beta 1 (TGFβ1) could be related to the regulation of the astrocyte reaction and AQP4 expression. In conclusion, BM-MSC treatment in hydrocephalus can stimulate a key developmental process such as the periventricular astrocyte reaction, where AQP4 overexpression could be implicated in tissue recovery.

Transcriptomic survey of key reproductive and metabolic tissues in mouse models of polycystic ovary syndrome

Excessive androgen production and obesity are key to polycystic ovary syndrome (PCOS) pathogenesis. Prenatal androgenized (PNA), peripubertal androgenized, and overexpression of nerve growth factor in theca cells (17NF) are commonly used PCOS-like mouse models and diet-induced maternal obesity model is often included for comparsion. To reveal the molecular features of these models, we have performed transcriptome survey of the hypothalamus, adipose tissue, ovary and metaphase II (MII) oocytes. The largest number of differentially expressed genes (DEGs) is found in the ovaries of 17NF and in the adipose tissues of peripubertal androgenized models. In contrast, hypothalamus is most affected in PNA and maternal obesity models suggesting fetal programming effects. The Ms4a6e gene, membrane-spanning 4-domains subfamily A member 6E, a DEG identified in the adipose tissue in all mouse models is also differently expressed in adipose tissue of women with PCOS, highlighting a conserved disease function. Our comprehensive transcriptomic profiling of key target tissues involved in PCOS pathology highlights the effects of developmental windows for androgen exposure and maternal obesity, and provides unique resource to investigate molecular mechanisms underlying PCOS pathogenesis.

NGF/TrkA promotes the vitality, migration and adhesion of bone marrow stromal cells in hypoxia by regulating the Nrf2 pathway.

Bone marrow stromal cells (BMSCs) transplantation is a treatment strategy for ischemic stroke (IS) with great potential. However, the vitality, migration and adhesion of BMSCs are greatly impaired due to the harsh environment of the ischemic area, which affects the therapeutic effects. Herein, we aimed to investigate the roles of nerve growth factor (NGF) in regulating cell behaviors of BMSCs in IS. The mRNA and protein expressions were assessed using qRT-PCR and western blot, respectively. To simulate ischemic-like conditions in vitro, Brain microvascular (bEnd.3) cells were exposed to oxygen and glucose deprivation (OGD). Cell viability and cell proliferation were evaluated by MTT assay and BrdU assay, respectively. Transwell migration and cell adhesion assays were carried out to determine cell migration and adhesion of BMSCs, respectively, coupled with flow cytometry to evaluate cell apoptosis of bEnd.3 cells. Finally, angiogenesis assay was performed to assess the angiogenesis ability of bEnd.3 cells. NGF overexpression resulted in increased cell vitality, adhesion and migration of BMSCs, while NGF knockdown presented the opposite effects. We subsequently discovered that TrkA was a receptor for NGF, and TrkA knockdown significantly inhibited the cell viability, migration and adhesion of BMSCs. Besides, Nrf2 was confirmed as the downstream target of NGF/TrkA to promote the viability, adhesion and migration of BMSC cells. Finally, NGF-silenced BMSCs could not effectively restore the OGD-induced brain microvascular cell damage. NGF/TrkA promoted the viability, migration and adhesion of BMSCs in IS via activating Nrf2 pathway.

Muscle-derived Stem Cells Combined With Nerve Growth Factor Transplantation in the Treatment of Stress Urinary Incontinence

To investigate the efficacy of muscle-derived stem cells (MDSCs) combined with nerve growth factor (NGF) in the treatment of stress urinary incontinence (SUI) METHODS: MDSCs were isolated and extracted from 90 SD rats, and the stem cell characteristics of the cells were identified using flow cytometry. NGF overexpression (oe-NGF) plasmid was coated with adenovirus and qRT-PCR was applied to verify adenovirus transfection efficiency. The rat models of SUI were constructed and randomly divided into 5 groups: control group, phosphate buffer (PBS) group, MDSCs+oe-NGF group, MDSCs+vector group, and MDSCs group. After 8 weeks of feeding, the leakage point pressure (LPP) rats, and Masson staining of rat urethral sections were detected. The expression of NGF and vascular endothelial growth factor (VEGF) was detected by western blot and IHC staining. Compared with the control group, the LPP and the ratio of muscle fibers/collagen fibers were significantly increased in the MDSCs treated groups, with the highest increase in the MDSCs+oe-NGF group. Western blot and IHC results showed that the expression of NGF and VEGF in the urethral tissues in the MDSCs treated groups were significantly up-regulated comparing with the control group, with the highest increase in the MDSCs+oe-NGF group. MDSCs alone can relieve SUI, while MDSCs combined with NGF is more effective, which may be related to the up-regulating of VEGF.

Study of the influence of NGF-β gene overexpression in human mesenchymal stem cells on the expression level of SOX1 and neural pathway genes.

Nerve growth factor (NGF) is a protein exhibiting an influence on the neural development and also, its' impact on the stem cells remains a great potential treatment strategy. The influence of its overexpression on the neural pathway differentiation on Wharton's Jelly derived MSC (WJ-MSC) has not been studied so far, but considering the fact that these cells are relatively easy to obtain, using them may indicate an innovative change in stem cell therapies. The aim of this study was to evaluate the effect of NGF overexpression in human mesenchymal stem cells (MSC) on SOX1 and genes related to the neural pathway. The lentiviral transduction was performed in order to obtain the NGF overexpression, as well as RT-PCR to evaluate the expression level SOX1, SOX2, NES, NGF under influence of overexpressed NGF protein in WJ-MSC. During the study we have observed a decrease in SOX1 expression as the marker of neural stem cells. Other than that an increase of SOX2, NES and NGF was noticed, as they all are markers of early-neural as well as already differentiated neural cells. The results show a great potential of using those examined genes' expression as a form of a new stem cell therapy. The achieved overexpression of NGF in this study, led the modified MSC onto the neural pathway as well as caused a decrease of SOX1 expression and an increase of expression of genes related to neural differentiated cells.

Nerve growth factor promotes lysyl oxidase-dependent chondrosarcoma cell metastasis by suppressing miR-149-5p synthesis

Chondrosarcoma is a malignancy of soft tissue and bone that has a high propensity to metastasize to distant organs. Nerve growth factor (NGF) is critical for neuronal cell growth, apoptosis, and differentiation, and also appears to promote the progression and metastasis of several different types of tumors, although the effects of NGF upon chondrosarcoma mechanisms are not very clear. We report that NGF facilitates lysyl oxidase (LOX)-dependent cellular migration and invasion in human chondrosarcoma cells, and that NGF overexpression enhances lung metastasis in a mouse model of chondrosarcoma. NGF-induced stimulation of LOX production and cell motility occurs through the inhibition of miR-149-5p expression, which was reversed by PI3K, Akt, and mTOR inhibitors and their respective short interfering RNAs. Notably, levels of NGF and LOX expression correlated with tumor stage in human chondrosarcoma samples. Thus, NGF appears to be a worthwhile therapeutic target for metastatic chondrosarcoma.

Intrahippocampal Adeno-Associated Virus-Mediated Overexpression of Nerve Growth Factor Reverses 192IgG-Saporin-Induced Impairments of Hippocampal Plasticity and Behavior.

One of the aspects of Alzheimer disease is loss of cholinergic neurons in the basal forebrain, which leads to development of cognitive impairment. Here, we used a model of cholinergic deficit caused by immunotoxin 192IgG-saporin to study possible beneficial effects of adeno-associated virus (AAV)–mediated overexpression of nerve growth factor (NGF) in the hippocampus of rats with cholinergic deficit. Suspension of recombinant AAV carrying control cassette or cassette with NGF was injected into both hippocampi of control rats or rats with cholinergic deficit induced by intraseptal injection of 192IgG-saporin. Analysis of choline acetyltransferase (ChAT) immunostaining showed that NGF overexpression in the hippocampus did not prevent strong loss of ChAT-positive neurons in the septal area caused by the immunotoxin. Induction of cholinergic deficit in the hippocampus led to impairments in Y-maze and beam-walking test but did not affect behavioral indices in the T-maze, open field test, and inhibitory avoidance training. NGF overexpression in the rats with cholinergic deficit restored normal animal behavior in Y-maze and beam-walking test. Recording of field excitatory postsynaptic potentials in vivo in the hippocampal CA1 area showed that induction of cholinergic deficit decreased magnitude of long-term potentiation (LTP) and prevented a decrease in paired-pulse ratio after LTP induction, and NGF overexpression reversed these negative changes in hippocampal synaptic characteristics. The beneficial effect of NGF was not associated with compensatory changes in the number of cells that express NGF receptors TrkA and NGFR in the hippocampus and medial septal area. NGF overexpression also did not prevent a 192IgG-saporin–induced decrease in the activity of acetylcholine esterase in the hippocampus. We conclude that NGF overexpression in the hippocampus under conditions of cholinergic deficit induces beneficial effects which are not related to maintenance of cholinergic function.

The mir-423-5p/MMP-2 Axis Regulates the Nerve Growth Factor-Induced Promotion of Chondrosarcoma Metastasis

Simple SummaryA chondrosarcoma is a common tumor of the bone that has a high propensity to metastasize to distant organs. The effects of NGF in a chondrosarcoma are not confirmed although NGF is capable of promoting the progression and metastasis of several different types of tumors. Here, we found that NGF promotes the chondrosarcoma migration and metastasis in vitro and in vivo. The levels of NGF and MMP-2 in human chondrosarcoma tumor tissues correlated strongly with the tumor stage. We identified that NGF induces the MMP-2 synthesis and chondrosarcoma cell motility by inhibiting miR-423-5p expression through the FAK and c-Src pathways. We suggest that NGF is a worthwhile therapeutic target in the treatment of a metastatic chondrosarcoma.A chondrosarcoma is a common tumor of the soft tissue and bone that has a high propensity to metastasize to distant organs. Nerve growth factor (NGF) is capable of promoting the progression and metastasis of several different types of tumors although the effects of NGF in a chondrosarcoma are not confirmed. Here, we found that the levels of NGF and matrix metalloproteinase-2 (MMP-2) correlated with the tumor stage in patients with a chondrosarcoma. NGF facilitated the MMP-2-dependent cellular migration in human chondrosarcoma JJ012 cells while the overexpression of NGF enhanced the lung metastasis in a mouse model of a chondrosarcoma. NGF promoted the MMP-2 synthesis and cell migration by inhibiting miR-423-5p expression through the FAK and c-Src signaling cascades. NGF appears to be a worthwhile therapeutic target in the treatment of a metastatic chondrosarcoma.

Overexpression of Nerve Growth Factor in the Hippocampus Induces Behavioral Changes in Rats with 192IgG-Saporin-Induced Cholinergic Deficit

Overexpression of Nerve Growth Factor in the Hippocampus Induces Behavioral Changes in Rats with 192IgG-Saporin-Induced Cholinergic Deficit

PERINEURAL INVASION IN ORAL SQUAMOUS CELL CARCINOMA: INCIDENCE, CLINICAL IMPACT AND MOLECULAR INSIGHT

<h3>Background</h3> Perineural invasion (PNI) has been associated with poor prognosis in oral squamous cell carcinoma (OSCC). Nerve growth factor (NGF) and its receptor tyrosine kinase A (TrkA) may play a role in PNI in OSCC. <h3>Objective</h3> To characterize the incidence and prognostic correlation of PNI in OSCC and to determine whether NGF and TrkA expressions could be used as biological markers for PNI in OSCC. <h3>Methods</h3> A retrospective review of pathology reports of 430 patients with OSCC treated from 1992 to 2014 in Tayside, Scotland, and followed for 5 years or until death was undertaken. The expression of NGF and TrkA was assessed with immunohistochemistry in 132 tissue sections of OSCC. <h3>Results</h3> PNI was identified in 17.4% of OSCC. High expression of NGF and TrkA was seen in 84% and 92% of OSCC, respectively. PNI-positive OSCC expressed NGF and TrkA with greater frequency than PNI-negative OSCC (<i>P</i> < .05). PNI and high expression of NGF were associated significantly with pain in OSCC (<i>P</i> < .05). PNI-positive OSCC and high NGF/TrkA expression exhibited a greater potential for lymphovascular invasion and depth of invasion >4 mm (<i>P</i> < .05). PNI was significantly associated with advanced size, nodal metastasis, poor differentiation, and stage IV tumors (<i>P</i> < .05). PNI correlated significantly with disease-specific survival in OSCC (<i>P</i> < .05). <h3>Conclusions</h3> Higher level of expression of NGF and TrkA many predict PNI and therefore may be considered as biological markers for PNI in OSCC. Pain could be a predictive for PNI and NGF overexpression. PNI and NGF expression can predict the aggressiveness and prognosis of OSCC.

RSV infection potentiates TRPV1-mediated calcium transport in bronchial epithelium of asthmatic children.

The transient receptor potential vanilloid 1 (TRPV1) channel is expressed in human bronchial epithelium (HBE), where it transduces Ca2+ in response to airborne irritants. TRPV1 activation results in bronchoconstriction, cough, and mucus production, and may therefore contribute to the pathophysiology of obstructive airway disease. Since children with asthma face the greatest risk of developing virus-induced airway obstruction, we hypothesized that changes in TRPV1 expression, localization, and function in the airway epithelium may play a role in bronchiolitis and asthma in childhood. We sought to measure TRPV1 protein expression, localization, and function in HBE cells from children with versus without asthma, both at baseline and after RSV infection. We determined changes in TRPV1 protein expression, subcellular localization, and function both at baseline and after RSV infection in primary HBE cells from normal children and children with asthma. Basal TRPV1 protein expression was higher in HBE from children with versus without asthma and primarily localized to plasma membranes (PMs). During RSV infection, TRPV1 protein increased more in the PM of asthmatic HBE as compared with nonasthmatic cells. TRPV1-mediated increase in intracellular Ca2+ was greater in RSV-infected asthmatic cells, but this increase was attenuated when extracellular Ca2+ was removed. Nerve growth factor (NGF) recapitulated the effect of RSV on TRPV1 activation in HBE cells. Our data suggest that children with asthma have intrinsically hyperreactive airways due in part to higher TRPV1-mediated Ca2+ influx across epithelial membranes, and this abnormality is further exacerbated by NGF overexpression during RSV infection driving additional Ca2+ from intracellular stores.

CS nerve growth factor regulates sevoflurane anesthesiainduced nerve injury in nerve cells

Purpose: Sevoflurane is the most commonly used anesthetic agent for surgery. However, it is associated with deficiency in learning and memory abilities. The study was aimed at investigating the role of nerve growth factor (NGF) in sevoflurane anesthesia-induced nerve injury.Methods: RT-qPCR assay was applied to measure expressions of NGF, miR-98-5p and other factors related to apoptosis. CCK-8 assay was used for detecting cell viability while luciferase reporter assay was employed to measure binding condition between miR-98-5p and NGF. Expressions of proteins in PI3K/AKT/mTOR signaling pathway was measured with western blot.Results: Sevoflurane reduced cell viability of RGC-5 cells, promoted apoptosis and reduced the expression of NGF. In sevoflurane-induced RGC-5 cells, over-expression of NGF promoted cell viability with reduced apoptosis. Also, there was reduction in the protein expression of PI3K/AKT/mTOR signaling pathway by sevoflurane, while up-regulation of NGF promoted the expressions of these proteins. In the presence of PI3K inhibitor, reduction cell viability was reduced but apoptosis increased. Luciferase reporter assay detected MiR-98-5p as the target gene of NGF and its overexpression restored high cell viability in the over-expressed NGF. The rate of apoptosis and expressions of proteins was also restored with up-regulation of miR-98-5p.Conclusion: Sevoflurane caused damage to nerve cells, while over-expression of NGF reduced the injury through PI3K/AKT/mTOR signaling pathway and suppression of miR-98-5p.&#x0D; Keywords: Nerve growth factor, Sevoflurane, Nerve injury, Anesthesia, miR-98-5p

Nerve growth factor regulates liver cancer cell polarity and motility.

Nerve growth factor (NGF), a prototypical neurotrophic factor essential for neuronal cell proliferation and survival, has been implicated as a marker of tumor progression, as well as a potential target for novel therapeutic approaches in cancer. To investigate the functional potential of NGF in liver cancer in the present study, a stable NGF-overexpressing HepG2 cell line was generated. The scratch-wound assay was used to investigate cell motility and polarity. Western blotting was performed to evaluate the expression levels of epithelial-mesenchymal transition (EMT)-related proteins, including E-cadherin, N-cadherin and vimentin. Moreover, immunofluorescence was performed to investigate the arrangement of the actin cytoskeleton. Cell anoikis resistance was examined using a suspension culture model and cell apoptosis was examined via flow cytometry. The present results indicated that NGF overexpression in HepG2 cells disrupted HepG2 cell polarity and promoted cell motility. Furthermore, NGF overexpression induced EMT and actin cytoskeleton rearrangement in HepG2 cells, as well as enhanced anoikis resistance and prevented cellular apoptosis. Notably, a tropomyosin receptor kinase A receptor inhibitor blocked NGF-induced cell motility and apoptosis. Therefore, it was suggested that NGF serves a critical role in the invasion and metastasis of liver cancer. The use of NGF as a biomarker or potential new target could lead to the development of novel factors for diagnosis or for improving therapeutic strategies in liver cancer.

Evaluation of PTEN Inhibitor Following Spinal Cord Injury on Recovery of Voiding Efficiency and Motor Function Observed by Regeneration in Spinal Cord.

PurposeNeurogenic bladder (NB) associated with spinal cord injury (SCI) is a serious health problem. However, no effective treatment has been developed for SCI patients with NB. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) inhibitors have been proposed as a promising option for inducing neural regeneration. Therefore, we investigated the effects of a tissue gene nerve (TGN), PTEN inhibitor, on voiding function, motor function, and the expression of growth factors after SCI.MethodsIn this experiment, female rats were randomly divided into 3 groups (n=10 in each group): the sham-operation group, the SCI-induced group, and the SCI-induced and TGN-treated group. Cystometry; the Basso, Beattie, and Bresnahan (BBB) scale test; the ladder walking test; hematoxylin and eosin staining; and Western blotting for brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF), and nerve growth factor (NGF) were performed to evaluate functional and molecular changes.ResultsAfter SCI, the rats exhibited decreased walking ability according to the BBB scale test and impaired coordinative function according to the ladder walking test. The PTEN inhibitor promoted enhanced walking ability and coordinative function. Cystometry showed voiding impairment after SCI and improved voiding function was observed after PTEN treatment. Overexpression of VEGF, BDNF, and NGF were observed after SCI. Administration of PTEN inhibitors significantly attenuated the overexpression of growth factors due to SCI.ConclusionPTEN inhibitor treatment diminished the overexpression of growth factors and promoted the repair of damaged tissue. PTEN inhibitor-treated rats also showed improved motor function and improved voiding function. Therefore, we suggest TGN as a new therapeutic agent that can be applied after SCI.

Lentivirus-mediated silencing of P75 neurotrophin receptor combined with nerve growth factor overexpression and transfection of bone marrow mesenchymal stem cells combined with demineralized bone matrix for heterotopic osteogenesis

To investigate the effects of silencing P75 neurotrophin receptor (P75NTR) and nerve growth factor (NGF) overexpression on the proliferative activity and ectopic osteogenesis ability of bone marrow mesenchymal stem cells (BMSCs) combined with demineralized bone matrix for heterotopic osteogenesis. BMSCs of Sprague Dawley (SD) rats were cultured and passaged by adherent isolation method. The third generation BMSCs were transfected with lentivirus mediated P75NTR gene silencing (group B), NGF overexpression gene (group C), P75NTR silencing and NGF overexpression double genes (group D), respectively, and untransfected cells as control (group A). After 7 days of transfection, the expression of fluorescent protein of the target gene was observed by fluorescence microscope; cell counting kit 8 method was used to detect the cells activity for 8 days after transfection; the expressions of P75NTR and NGF proteins in each group were detected by Western blot. The adhesion of BMSCs to demineralized bone matrix (DBM) was observed by inverted phase contrast microscope and scanning electron microscope after transfection of p75NTR silencing and NGF overexpression double genes. After transfection, BMSCs and DBM were co-cultured to prepare 4 groups of tissue engineered bone, which were respectively placed in the dorsal subcutaneous tissue of 8-week-old SD rats to construct subcutaneous ectopic osteogenesis model ( n=6). HE staining was performed at 4 and 8 weeks after operation. ALP staining was used to observe the formation of calcium nodules at 8 weeks after operation. The expressions of Runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), and osteocalcin (OCN) were detected by real-time fluorescent quantitative PCR. At 7 days after transfection, there was no fluorescence expression in group A, red fluorescence expression was seen in group B, green fluorescence expression in group C, and red-green compound fluorescence expression in group D. The fluorescence expression rate of target gene was about 70%. Western blot detection showed that the relative expression of P75NTR protein in groups A and C was significantly higher than that in groups B and D, and the relative expression of NGF protein in groups C and D was significantly higher than that in groups A and B ( P<0.05). With the passage of time, the cell proliferation activity increased in all groups, especially in group D, which was significantly higher than that in group A at 3-8 days ( P<0.05). The results of inverted phase contrast microscope and scanning electron microscope showed that BMSCs could adhere well to DBM. In the subcutaneous ectopic osteogenesis experiment, HE staining showed that at 4 and 8 weeks after operation, the more bone tissue was formed in group D than in the other 3 groups. ALP staining showed that group D had the highest ALP activity and better osteogenic expression. Compared with group A, the relative expressions of Runx2, ALP, and OCN mRNAs in group D were significantly higher than those in group A ( P<0.05). Silencing P75NTR and NGF overexpression double genes co-transfected BMSCs with DBM to construct tissue engineered bone has good ectopic osteogenic ability. By increasing NGF level and closing P75NTR apoptosis channel, it can not only improve cell activity, but also promote bone tissue regeneration.

Nerve growth factor protects salivary glands from irradiation-induced damage

AimsRadiotherapy has become a basic treatment modality for head and neck cancer. However, radiotherapy results in inevitable side effects, particularly radiation sialadenitis, that significantly impairs quality of life. A previous study indicated that nerve growth factor (NGF) has a radio-protective effect, but the mechanism was not determined in salivary glands. In this study, we explored the functional role and mechanism regarding how NGF protects salivary glands against IR-induced damage. Main methodsHuman salivary gland (HSG) cells and C57BL/6 mice were selected to establish an IR-induced salivary gland damage model in vitro and in vivo. Recombinant NGF protein and NGF siRNA and over-expression plasmids were applied to manipulate NGF expression in vitro. AAV-NGF was retrogradely perfused into the submandibular gland (SMG) through the SMG duct to manipulate NGF expression in vitro. Small-molecule inhibitors and siRNAs were applied to inhibit AKT and JNK. Western blotting, quantitative PCR, flow cytometry and histology assays were performed to analyse the functional role and mechanism of NGF. Key findingsOur study demonstrated that NGF expression was upregulated following radiotherapy both in human HSG cells and mouse SMG tissues. NGF could reduce IR-induced HSG cell apoptosis, and AAV-mediated gene therapy could restore the salivary flow rate and protect the salivary gland against IR-induced apoptosis in vivo. Mechanistically, NGF protects salivary glands from IR-induced apoptosis by de-phosphorylating JNK kinase rather than promoting AKT phosphorylation. SignificanceThe current study findings indicated that the modulation of the NGF pathway might prevent IR-induced salivary hypo-function.