Effect of Topical Recombinant Human Nerve Growth Factor on Corneal Epithelial Regeneration in Refractory Epithelial Keratopathy.

PURPOSE: To report the effect of topical application of recombinant human nerve growth factor (rhNGF) eye drops on corneal epithelial regeneration in patients with refractory epitheliopathy. METHODS: A retrospective chart review was conducted on patients treated with topical rhNGF for refractory epithelial keratopathy due to stage I neurotrophic keratitis (NK). Data regarding demographics and ocular/systemic past medical history was extracted from patient charts. Visual acuity and corneal staining scores were recorded at baseline and subsequent follow-up visits at 8 weeks and 3 months. Measurements from the worse eye were used to compare before and after treatment values. RESULTS: We identified 14 patients (median age 68 years, 21% male) who received rhNGF treatment for refractory epithelial keratopathy. After an 8-week treatment with topical rhNGF, the median corneal staining score in the worse eye improved from 4 to 1 (p = 0.001). All patients showed at least one-grade improvement in corneal staining at 8 weeks, with sustained effect in seven patients at 3 months. A better response was observed in eyes with post-radiation epithelial keratopathy, LASIK, and Sjogren's disease. Those with chronic use of other topical treatments and uncontrolled diabetes mellitus demonstrated incomplete responses. Eight patients reported mild-to-moderate ocular discomfort from drop application that fully resolved after completion of treatment. CONCLUSIONS: Topical rhNGF was effective and safe for refractory epithelial keratopathy in our small cohort, but sustained effects were seen only in certain etiologies for up to 3 months. Further studies are needed for optimal dosing and duration based on underlying causes.

Phase IV Multicenter, Prospective, Open-Label Clinical Trial of Cenegermin (rhNGF) for Stage 1 Neurotrophic Keratopathy (DEFENDO).

INTRODUCTION: Cenegermin is approved for treatment of neurotrophic keratopathy (NK) and has been studied in patients with stage 2 or 3 NK. This study evaluated the efficacy and safety of cenegermin in adults with stage 1 NK. METHODS: This was a phase IV, multicenter, prospective, open-label, uncontrolled trial. Adults with stage 1 NK (Mackie criteria) and decreased corneal sensitivity (≤ 4 cm) received 1 drop of cenegermin 20 mcg/ml in the affected eye(s) 6 times/day for 8 weeks with a 24-week follow-up. RESULTS: Of 37 patients, corneal epithelial healing was observed in 84.8% (95% confidence interval [CI] 68.1-94.9%; P < 0.001) at week 8; 95.2% (95% CI 76.2-99.9%; P < 0.001) of those patients remained healed at the end of the 24-week follow-up (week 32). At week 8, 91.2% (95% CI 76.3-98.1%; P < 0.001) of patients experienced improved corneal sensitivity; this improvement was observed in 82.1% (95% CI 63.1-93.9%; P < 0.001) of patients at week 32. Mean best-corrected distance visual acuity change from baseline at week 8 was - 0.10 logMAR (standard deviation [SD], 0.15; 95% CI - 0.16 to - 0.05; P < 0.001) and at week 32 was - 0.05 logMAR (SD, 0.16; 95% CI - 0.11 to 0.01; P = 0.122). At weeks 8 and 32, 15.2% (95% CI 5.1-31.9%; P < 0.001) and 10.7% (95% CI 2.3-28.2%; P < 0.001) of patients, respectively, had a 15-letter gain from baseline. At least one adverse event (AE) was reported by 73.0% and 45.7% of patients during the treatment and follow-up periods, respectively. The most common treatment-related, treatment-emergent AEs were eye pain (37.8%), blurred vision (10.8%), and eyelid pain (8.1%); these were mostly mild or moderate and were only reported during the treatment period. CONCLUSIONS: These results support the potential use of cenegermin for treating patients with stage 1 NK, and future confirmatory studies would be beneficial to elaborate on these findings. TRIAL REGISTRATION: DEFENDO; NCT04485546.

Enhanced neuroprotective activity of ophthalmic delivered nerve growth factor conjugated with cell penetrating peptide against optic nerve injury.

Aims: Nerve growth factor is a well characterised neurotrophic factor that play a critical role in the survival, growth and differentiation of neurons both in central and peripheral nervous system. However, it is difficult for the conventional exogenous nerve growth factor administration delivery to the central nervous system due to the biological barrier in human bodies.Results: We validated a series of cell penetrating peptides and found that L-PenetraMax significantly enhanced the efficiency of recombinant human nerve growth factor entry into the rat retina. In the optic nerve crush mice model, eye drop administration of recombinant human nerve growth factor alone promoted retinal ganglion cell survival and axon regeneration at high dose, while the combination of recombinant human nerve growth factor with L-PenetraMax significantly enhanced the neuroprotective efficacy at lower dose, thus potentially enhancing the availability of recombinant human nerve growth factor eye drops in patients with optic neuropathy.Conclusions: This study provides the evidence that the noncovalent coadministration of recombinant human nerve growth factor with L-PenetraMax could be a potent strategy for the non-invasive and sustained ocular delivery of therapeutic proteins for improving the optic nerve injury.

Effect of recombinant human nerve growth factor treatment on corneal nerve regeneration in patients with neurotrophic keratopathy.

Introduction: Neurotrophic Keratopathy (NK) is a neurodegenerative corneal disease that results in diminished corneal sensation. Previous studies have found that Cenegermin 0.002%, a recombinant human nerve growth factor (rhNGF), improves corneal epithelial healing in stage 2 and 3 NK patients. However, rhNGF effect on corneal sensation and nerve regeneration has not been well established. Thus, this study aims to analyze the effect of rhNGF on corneal nerve regeneration using in vivo confocal microscopy (IVCM) and on corneal sensitivity in NK patients. Methods: This is a retrospective, longitudinal, case-control study that included patients with NK, treated with rhNGF for at least 4 weeks, with pre- and post-treatment IVCM images available for analysis. Chart reviews were conducted documenting prior medical and surgical history, clinical signs and symptoms, and corneal sensation using Cochet-Bonnet esthesiometry. Corneal nerve parameters were assessed by IVCM. Sex- and age-matched reference controls were selected from a database of healthy subjects for comparison. Results: The study included 25 patients, with 22 (88%) stage 1, two (8%) stage 2, and 1 (4%) stage 3 NK patients, with a median age of 64 years (range: 30-93 years). Total, main, and branch nerve densities [median (range) in mm/mm2] were lower in the NK group pre-treatment [2.3 (0.0-21.1); 1.7 (0.0-13.0); 0.5 (0.0-10.2); respectively] vs. controls [22.3 (14.9-29.0); 10.1 (3.2-15.4); and 12.1 (6.2-18.4), (p < 0.0001 for all), respectively]. Post-treatment nerve densities increased compared to pre-treatment to 5.3 (0.0-19.4, p = 0.0083) for total, 3.5 (0.0-13.2, p = 0.0059) for main, and 2.0 (0.0-10.4, p = 0.0251) for branch nerves, but remained lower than controls (p < 0.0001 for all). Corneal sensation increased from 2.3 ± 1.1 cm pre-treatment to 4.1 ± 1.4 cm post-treatment (p = 0.001). Median best corrected visual acuity significantly increased following rhNGF treatment from 0.4 (0.0-1.6) to 0.12 (-0.1 to 1.6) (p = 0.007). Conclusion: Patients with NK treated with at least 4 weeks of rhNGF, showed a significant increase in corneal nerve densities after treatment. A significant increase in corneal sensation, as well as best corrected visual acuity, was observed following treatment.

Neuroma Treatment With the Acellular Nerve Allograft Reconstruction Technique.

Treatment of a painful neuroma is a challenging problem for both the patient and the providers. Current surgical treatment options typically include excision of the neuroma and stump relation. However, with both treatment options, patients have high rates of persistent pain and rates of neuroma recurrence. We describe two patients with neuromas treated with our acellular nerve allograft reconstruction technique. This technique involves the excision of the neuroma and bridging the proximal nerve end to the surrounding tissue with an acellular nerve allograft. Both patients had immediate resolution of their neuropathic pain that was maintained at their final follow-up. Acellular nerve allograft reconstruction is a promising treatment option for the treatment of painful neuromas.

Morphology and morphometry of the human obturator nerve in males and females.

Peripheral nerve injury and the nerves' subsequent repair and regeneration continues to be marked clinically by poor functional recovery. The analysis of nerve morphology is an aspect which may provide an impact on successful clinical outcomes through better prediction of donor and recipient matching. In this study, we evaluated the morphological aspects of the human obturator nerve for a better understanding of its potential in nerve transplantation. Morphological characteristics of donor obturator nerves were analysed, including nerve diameter and length, fascicle count and the ratio of neural to non-neural tissue present within the cross-sectional area of the nerve's epineurium, with respect to laterality and sex. Statistical significance (p < 0.10) was determined for male obturator nerves having an average diameter of 2.67 mm compared to female obturator nerves at 1.91 mm, as well as left obturator nerves having an average of 11.21 fascicles compared to the right having an average of 10.17 fascicles. Strong positive correlations were determined between cross-sectional nerve area and limb size index, as well as between percentage of non-neural tissue and area of non-neural tissue, among males. Separately, strong correlation between percentage of non-neural tissue and area of non-neural tissue among right obturator nerves in males and females was determined . These findings indicate that there are associations and predictions that can be made about nerve morphology and that these when combined with other patient characteristics may enhance patient functional recovery following a peripheral nerve's repair.

Clinical observation of recombinant human nerve growth factor in the treatment of neurotrophic keratitis.

AIM: To characterize changes of corneal nerve morphology and tear indices in patients with neurotrophic keratitis (NK) treated with recombinant human nerve growth factor (rhNGF). METHODS: In a prospective observational study, six patients (nine eyes) were locally treated with rhNGF. Visual acuity, corneal fluorescein staining score, the heights of the tear river, lipid layer thickness (LLT), tear ferning (TF) test, conjunctival impression cytology (CIC) examination, the densities of cornea subbasal nerve fibers were determined before and after treatment. RESULTS: Compared with baseline, there was a significant difference in corneal fluorescence staining scores (P<0.01); all patient corneal epithelial defects recovered completely within 8wk, but there was no significant improvement in the height of the tear river (P=0.202). LLT was significantly increased when compared with baseline (P=0.042); however, the function of conjunctival goblet cells and mucin content did not significantly improve using the TF test and CIC examination (P=0.557, P=0.539). After 8wk of treatment, the average corneal subbasal nerve fiber density increased significantly (P<0.01), as did the number of corneal nerve fiber branches (P=0.001). CONCLUSION: RhNGF can increase the density of corneal subbasal nerve fibers, promote the healing of persistent corneal epithelial defects and corneal ulcers in patients with NK, also improving tear function partially.

Neurotrophic Keratopathy Treated with Topical Recombinant Human Nerve Growth Factor (Cenegermin): Case Series Study with Long-Term Follow-Up.

The authors report the use of topical recombinant human nerve growth factor cenegermin 0.02% in 5 patients diagnosed with neurotrophic keratopathy (NK) in a real-life setting. These 5 patients affected with stage II and III NK mainly of herpetic cause received cenegermin six times daily for 8 weeks. It was initiated upon refractoriness to prior conventional topical treatment. Visual acuity, corneal sensitivity test at four corneal quadrants, fluorescein staining, OC,T and photography were performed weekly during 9 weeks of follow-up from the completion of treatment. At the ninth week of follow-up, corneal sensitivity improvement and healing of corneal ulcers were found in all patients. No adverse events were reported, and no corneal ulcer recurrence was observed over a 4-year follow-up period. Cenegermin should be used in combination with conventional therapy for advanced NK, as it is an effective treatment for healing corneal ulcers, improving the corneal surface homeostasis and avoiding surgery.

Long-term clinical efficacy of topical treatment with recombinant human nerve growth factor in neurotrophic keratopathy: a novel cure for a rare degenerative corneal disease?

BACKGROUND: Neurotrophic keratopathy (NK) is a rare, degenerative ocular disease characterized by reduction or loss of corneal sensitivity and development of non-healing corneal epithelial defects and ulcers. Cenegermin, a recombinant human nerve growth factor (rhNGF) eye drop solution, is the first drug approved for the treatment of NK. The aim of our study is to evaluate the long-term efficacy of this innovative topical treatment in patients with NK. METHODS: Retrospective, consecutive, observational case series study from a single-center setting (Department of Sense Organs, University Sapienza of Rome, Rome, Italy). 18 patients with diagnosis of stage 2 or 3 NK, treated with Cenegermin 20 mcg/ml eye drops were followed for up to 48 months. Recurrence of lesion during follow-up was evaluated at 12, 24, 36, and 48 months. In addition, corneal sensitivity, Schirmer tear test, and visual acuity (VA) were recorded at baseline, end of treatment, and at 12, 24, 36, and 48 months. RESULTS: Three patients experienced recurrence of persistent epithelial defects (PEDs) within 12 months and one patient experienced recurrence of a corneal ulcer within 36 months. Corneal sensitivity was significantly improved at all timepoints (P < 0.05). Significant improvements in visual acuity and tear production were seen at the completion of treatment as well as at 12, 24, and 36 months (P < 0.05) when compared to baseline. CONCLUSIONS: A single 8-week treatment regimen of Cenegermin eye drops has clinical efficacy that can persist for up to 48 months. The long-term clinical utility of treatment with Cenegermin for NK was demonstrated through the low rate of lesion recurrence along with improvements in corneal sensitivity and tear production.

Ultrasound Appearance of In Vitro Nerve Allografts and Conduits for Peripheral Nerve Reconstruction.

Ultrasound enables the accurate assessment of traumatic disorders of small peripheral nerves of extremities. Human nerve allografts and nerve conduits are increasingly used for the surgical treatment of nerve trauma but ultrasound reports on this field are scarce in the radiological literature. We present the macroscopic and in vitro ultrasound appearance of human allografts, and synthetic and biological conduits. In addition, we describe the ultrasound findings in some patients operated upon using the same devices. The in vitro ultrasound appearance correlated well with the macroscopic appearance of the devices. Awareness of their appearance in vitro can help sonologists when examining postsurgical patients.

Long-term clinical outcome and satisfaction survey in patients with neurotrophic keratopathy after treatment with cenegermin eye drops or amniotic membrane transplantation.

PURPOSE: Neurotrophic keratopathy (NK) is a degenerative corneal disease caused by damage of trigeminal innervation. The purpose of this study is to evaluate the clinical outcomes and patient-reported satisfaction of treatment with amniotic membrane transplantation (AMT) or cenegermin eye drops in patients with NK. METHODS: Clinical charts of patients with NK treated with AMT (group A) or cenegermin eye drops (group B), with at least 12 months of follow-up, were reviewed for demographics, medical history, corneal healing, and disease recurrence. Patient satisfaction was evaluated by a newly developed questionnaire investigating patient's appreciation of treatment of NK (2 items) and satisfaction with NK treatment outcomes (5 items). RESULTS: At the end of treatment, complete corneal healing was observed in 13/15 (86%) patients in group A and in 23/24 (96%) in group B. At 12 months follow-up, 6/13 patients (46%) in group A and 3/23 patients (13%) in group B showed recurrence of NK (p = 0.037). Survival analysis showed that group B remained recurrence free for a significantly longer period of time than the group A (p = 0.028). Patients in group B showed a significantly higher satisfaction when compared with patients in group A (total score: 65.7 ± 15.7 vs 47.4 ± 12.8, p = 0.003), both in terms of patients' appreciation of treatment (78.3 ± 15.9 vs 52.2 ± 30, p = 0.020) and satisfaction with treatment outcomes (60.7 ± 21 vs 45.4 ± 13.3, p = 0.037). CONCLUSIONS: Treatment of NK with cenegermin was associated with long-term maintenance of corneal integrity and a higher degree of patient satisfaction.

Efficacy of rhNGF-loaded amniotic membrane transplantation for rabbit corneal epithelial and nerve regeneration.

AIM: To evaluate the efficacy of recombinant human nerve growth factor-loaded amniotic membrane (rhNGF-AM) on corneal epithelial and nerve regeneration in rabbit model. METHODS: Freshly prepared human amniotic membrane (AM) were immersed into PBS buffer containing 100 or 500 µg/mL rhNGF for 15, 30, and 60min at 4°C. The in vitro release kinetics of rhNGF was measured with ELISA. For in vivo evaluation, the AM were immersed with 500 µg/mL rhNGF for 30min. Fifty-seven rabbits were selected to establish corneal epithelial defect model. In addition to the 19 rabbits in control group, 38 rabbits received AM transplantation with or without rhNGF after the removal of central epithelium. Corneal epithelial defect area, sub-epithelial nerve fiber density, corneal sensitivity, rhNGF contents in resident AM and corneas were measured after the surgery. RESULTS: rhNGF was sustained release from the AM within 14d in vitro, with the positive correlation with initial immersion concentration. The immersion of AM in 500 µg/mL rhNGF for 30min achieved the most stable release within 14d. After transplantation in rabbit cornea, a high concentration of rhNGF in resident rhNGF-AM and cornea was maintained within 8d. Corneal epithelial healing, nerve fiber regeneration and the recovery of corneal sensitivity were significantly accelerated after the rhNGF-AM transplantation when compared to simple AM transplantation (all P<0.05). CONCLUSION: Simple immersion of AM achieves the sustained release of rhNGF, and promotes corneal epithelial wound healing and nerve regeneration, as well as the recovery of corneal sensitivity in rabbit.

[Effects of gender and age on protein secretion in saliva of transgenic mice expressing human nerve growth factor].

Nerve growth factor (NGF) can promote the development, differentiation and regeneration of neurons. Recently, in order to efficiently produce human NGF (hNGF) drugs with better efficacy, we created transgenic mice expressing hNGF specifically in their salivary glands, and purified highly active hNGF protein from their saliva. Some studies reported that the NGF secretion in mouse saliva is affected by gender and age. Here, in order to select hNGF transgenic mice with high NGF secretion for saliva collection and hNGF purification, we divided transgenic mice into 4 groups, including 28-day-old young males and females, 63-day-old adult males and females. We compared their saliva volume, total salivary protein amount, salivary mNGF protein amount and salivary hNGF protein amount. The results showed that the saliva volume as well as amounts of total salivary protein, salivary mNGF protein and salivary hNGF protein secreted by 63-day-old transgenic mice were significantly higher than those secreted by sex-match 28-day-old transgenic mice, and the salivary hNGF protein amount secreted by male transgenic mice at the age of 63 days was significantly higher than that of female transgenic mice at the same age; Among 4 groups of mice, 63-day-old male transgenic mice secreted the highest salivary hNGF content, which was about 46 times higher than that secreted by the 28-day-old female transgenic mice. Therefore, 63-day-old male transgenic mice should be selected for saliva collection and hNGF purification.

Outcome of human peripheral nerve repair interventions using conduits: a systematic review.

BACKGROUND & OBJECTIVE: Peripheral nerve injury is very common, but repair is a challenging medical problem. Advances in medical sciences and technologies have however made tremendous breakthroughs in understanding repair mechanisms in nerve injury making this a fascinating area in neurotherapeutics. However, a systematic analysis of existing data is lacking, the present study was attempted to review existing literature in nerve repair studies in human beings and analyse outcome systematically. METHODS: A detailed search was made from various databases published in the last 10 years. The studies were included based on availability of data on the age of the patients, type of injuries, type of intervention and also on the minimal follow up period. Studies satisfying these criteria were subjected to a homogeneity test. On 263 patients from 3 homogeneous studies outcome parameters such as the functional improvement, sensory and motor recovery parameters were analysed. RESULTS: Results showed that conduits were safe and significantly more effective compared to the conventional sutures in effecting repair of sensory nerve injuries (Odds ratio 3.78; P < .00001). CONCLUSION: In conclusion, repair of human sensory peripheral nerve using conduits is safe and more effective than direct nerve suture.

Nerve-targeted probes for fluorescence-guided intraoperative imaging.

A fundamental goal of many surgeries is nerve preservation, as inadvertent injury can lead to patient morbidity including numbness, pain, localized paralysis and incontinence. Nerve identification during surgery relies on multiple parameters including anatomy, texture, color and relationship to surrounding structures using white light illumination. We propose that fluorescent labeling of nerves can enhance the contrast between nerves and adjacent tissue during surgery which may lead to improved outcomes. Methods: Nerve binding peptide sequences including HNP401 were identified by phage display using selective binding to dissected nerve tissue. Peptide dye conjugates including FAM-HNP401 and structural variants were synthesized and screened for nerve binding after topical application on fresh rodent and human tissue and in-vivo after systemic IV administration into both mice and rats. Nerve to muscle contrast was quantified by measuring fluorescent intensity after topical or systemic administration of peptide dye conjugate. Results: Peptide dye conjugate FAM-HNP401 showed selective binding to human sural nerve with 10.9x fluorescence signal intensity (1374.44 ± 425.96) compared to a previously identified peptide FAM-NP41 (126.17 ± 61.03). FAM-HNP401 showed nerve-to-muscle contrast of 3.03 ± 0.57. FAM-HNP401 binds and highlight multiple human peripheral nerves including lower leg sural, upper arm medial antebrachial as well as autonomic nerves isolated from human prostate. Conclusion: Phage display has identified a novel peptide that selectively binds to ex-vivo human nerves and in-vivo using rodent models. FAM-HNP401 or an optimized variant could be translated for use in a clinical setting for intraoperative identification of human nerves to improve visualization and potentially decrease the incidence of intra-surgical nerve injury.

Introduction of neurosupportive cells into processed acellular nerve allografts results in greater number and more even distribution when injected compared to soaking techniques.

OBJECTIVES: Processing necessary to remove immunogenic components of nerve allograft renders it acellular. Seeding with supportive cells may improve axon regeneration. We aim to identify the method associated with implantation of the greatest volume and most even distribution of cells. METHODS: Hypodermic needle injection was compared to soaking in solution under both normal and pressurized conditions after micropuncture of the allograft. Distribution within the allograft was measured using an in vitro model of fluorescent beads, as well as cultured Schwann cells. RESULTS: Injection treatment resulted in larger volumes and a more uniform cross-sectional distribution of implanted cells. Beads and cells behaved similarly relative to the measured outcomes. CONCLUSIONS: Injection instills more cells in a more uniform distribution. In vivo testing may evaluate whether these techniques vary relative to cell survival, cell migration, and clinical outcomes. Size- and concentration-matched fluorescent beads may represent a viable model for analyzing cell implantation.

Cenegermin for the treatment of neurotrophic keratitis.

The trigeminal nerve provides corneal sensitivity and trophic supply to corneal tissues. The impairment of corneal innervation leads to development of neurotrophic keratitis (NK). NK is a rare, degenerative corneal disease characterized by corneal hypo/anesthesia and development of nonhealing corneal epithelial defects and ulcers. NK is a challenging condition with high medical need due to the lack of approved treatments that can restore corneal integrity. Current treatment of NK aims at stimulating corneal healing and preventing disease progression. Cenegermin is a recombinant human nerve growth factor that was safe and well tolerated in preclinical and clinical studies. Cenegermin eye drops were safe and effective in restoring corneal integrity in two phase II clinical trials in patients with NK. The European Commission granted a full marketing authorization to cenegermin eye drops for the treatment of moderate to severe NK in July 2017.

The Evaluation of Nerve Growth Factor Over Expression on Neural Lineage Specific Genes in Human Mesenchymal Stem Cells.

OBJECTIVE: Treatment and repair of neurodegenerative diseases such as brain tumors, spinal cord injuries, and functional disorders, including Alzheimer's disease, are challenging problems. A common treatment approach for such disorders involves the use of mesenchymal stem cells (MSCs) as an alternative cell source to replace injured cells. However, use of these cells in hosts may potentially cause adverse outcomes such as tumorigenesis and uncontrolled differentiation. In attempt to generate mesenchymal derived neural cells, we have infected MSCs with recombinant lentiviruses that expressed nerve growth factor (NGF) and assessed their neural lineage genes. MATERIALS AND METHODS: In this experimental study, we cloned the NGF gene sequence into a helper dependent lentiviral vector that contained the green fluorescent protein (GFP) gene. The recombinant vector was amplified in DH5 bacterial cells. Recombinant viruses were generated in the human embryonic kidney 293 (HEK-293) packaging cell line with the helper vectors and analyzed under fluorescent microscopy. Bone marrow mesenchymal cells were infected by recombinant viruses for three days followed by assessment of neural differentiation. We evaluated expression of NGF through measurement of the NGF protein in culture medium by ELISA; neural specific genes were quantified by real-time polymerase chain reaction (PCR). RESULTS: We observed neural morphological changes after three days. Quantitative PCR showed that expressions of NESTIN, glial derived neurotrophic factor (GDNF), glial fibrillary acidic protein (GFAP) and Microtubule-associated protein 2 (MAP2) genes increased following induction of NGF overexpression, whereas expressions of endogenous NGF and brain derived neural growth factor (BDNF) genes reduced. CONCLUSION: Ectopic expression of NGF can induce neurogenesis in MSCs. Direct injection of MSCs may cause tumorigenesis and an undesirable outcome. Therefore an alternative choice to overcome this obstacle may be the utilization of differentiated neural stem cells.