Thirty-five SD rats were divided into 3 groups, in which 5 rats were served as control. The other 2 groups, 15 rats in each received either NGF solution or normal saline. The injury at the level of T8 spinal segment of the rat in these two groups were made by dropping a weight of 10 g from a height 2.5 cm after a total laminectomly from T7-11 segments. A thin plastic tube was inserted into the subarachnoid space below the injured segments. NGF was introduced through the tube at intervals of 1, 2, 3, 4, 8, 12, 24, 48 hours in the NGF group, and normal saline in the normal saline group. At 4, 8, 24 hours following surgery, 5 rats in each group were sacrificed and the injured segments were resected for examination. The contents of water and calcium were measured by dry-wet method and atomic absorption spectroscopy. The result showed that total calcium and water contents in normal saline group were markly increased, however, the changes of these two parametere were not so prominent in NGF group. It was suggested that NGF play a role in protecting the spinal cord by maintaining the calcium level of the injured segment.
ObjectiveTo explore the protective effect and mechanism of Krüppel-like factor 7 (KLF7) overexpression on retinal ganglion cells (RGC) in mice after optic nerve crush (ONC). MethodsA total of sixty five 10-week-old C57BL/6J mice were randomly divided into five groups: blank control group (group A), intravitreal injection (IVT)-KLF7 group (group B), IVT-phosphate buffer saline-ONC group (group C), IVT-KLF7-ONC group (group D), and IVT-recombinant adeno-associated virus 2-enhanced green fluorescent protein-ONC group (group E), with 13 mice in each group. On the 7 days after the ONC model, the mice in each group were killed. RGC survival rate was counted by whole retina flat mount and immunofluorescence techniques. KLF7, nerve growth factor (NGF), tyrosine kinase A (TrkA), phosphorylated TrkA (pTrkA), tyrosine kinase B (TrkB) and phosphorylated TrkB (pTrk) were detected by western blot, growth associated protein 43 (GAP43), brain-derived neurotrophic factor, B lymphoblastoma-2 (Bcl-2), Bcl-2 associated X protein (BAX), Caspase-3 protein relative expression levels. One-way analysis of variance or Kruskal-Wallis test were used for comparison between groups. ResultsOn the 7 days after the ONC model, the density of RGC in the retina of groups A, B, C, D and E were (3 707.4±12.8), (3 582.4±13.3), (1 396.3±16.1), (1 658.3±22.2) and (1 323.6±16.9)/mm2, respectively. Compared with groups C and E, RGC density in group D was significantly increased, and the difference was statistically significant (P=0.028, 0.007). Compared with groups A, B, C and E, the relative expression levels of NGF, pTrkA, pTrkB, GAP43 and Bcl-2 proteins in the retina of mice in group D were increased, while the relative expression levels of BAX and Caspase-3 proteins were decreased, with statistical significance (P＜0.01). ConclusionIn mouse ONC model, overexpression of KLF7 can improve RGC survival rate, increase the relative expression levels of NGF, pTrkA, pTrkB, GAP43 and Bcl-2 proteins in retina, and decrease the relative expression levels of BAX and Caspase-3 proteins.
OBJECTIVE: To study the nerve growth factor (NGF) expression and the influence of IL-1 alpha or IL-1 beta on NGF secretion in newborn rat astrocytes. METHODS: Astrocytes obtained from the brain cortex of newborn rats were cultured and purified, and they were divided into three groups, experimental, control and blank groups. IL-1 alpha or IL-1 beta were added into the experimental group with 25, 50 and 100 U/ml, each group was cultured for 24, 48 or 72 hours, and then the NGF contents in cultured astrocytes suspension media were measured by a two-cite enzymelinked immunoserbent assay (ELISA). RESULTS: Astrocytes could secret NGF by themselves and each concentration of IL-1 alpha or IL-1 beta media at any testing time could enhance NGF secreting in newborn rat astrocytes in certain degrees. The effects of IL-1 beta were ber than IL-1 alpha, the best effect in the unit time was observed in IL-1 beta with 50 U/ml for 24 hours. CONCLUSION: Astrocytes can express NGF, and IL-1 alpha or IL-1 beta can enhance the NGF expression in newborn rat astrocytes.
Objective To investigate the effects of epidermal growth factor (EGF)，fibroblast growth factor(FGF), and bovine serum on proliferation and apoptosis of the cultured fetal human retinal cells.Methods EGF and FGF were added or not to the medium of fetal human retinal cells cultured by bovine serum in vitro. The number of cells, bromodeoxyuridine(BrdU) incorporation and Tdt-mediated dUTP nick end labelling(TUNEL) were detected to determine the proliferation and apoptosis. Immunohistochemical staining of neuron specific enolase(NSE), Thy1.1, glial fibrillary acidic protein(GFAP) and scan electromicroscopy were performed to identify cell components. The expression of transcription factor c-fos, c-jun and apoptosis regulation factor bcl-2 and Bax were examined by immunohistochemical staining to explore the underlying mechanism.Results The increased number of NSE and Thy1.1 positive cells and BrdU incorporation, and decreased apoptotic cells were found in the groups treated with EGF and FGF. Meanwhile, the up-regulation of c-fos, c-jun and bcl-2 were also found. Conclusion EGF and FGF can promote the survival and proliferation of cultured retinal cells by up-regulating the expression of c-fos, c-jun and bcl-2. (Chin J Ocul Fundus Dis，2003，19：113-116)
Objective Using nerve growth factor ( NGF) and anti-NGF microspheres injected directly into the asthmatic rat adrenal gland, to explore the possible role of anti-NGF microsphere treatment in asthma.Methods 32 male SD rats were randomly divided into a normal control group, an asthma group, a NGF microspheres group, and an anti-NGF microspheres group. The behavior of rats, lung function testing, light microscopy of lung biopsy, electron microscopy of adrenal medulla cell ultrastructure changes, NGF and phenylethanolamine N-methyltransferase ( PNMT) expressions in the adrenal gland were assayed by immunohistochemistry method, and serum NGF, cortisol, corticosterone, epinephrine and norepinephrine concentrations were detected by ELISA. Results Behavior in the asthma rats showed varying degrees of sneezing, runny nose, wheezing, scratching the head and face, irritability holes, incontinence, increased aggression and other acts, while in the anti-NGF rats showed relatively slighter symptoms. The rats in the asthma, anti-NGF and NGF groups showed significant airway hyperresponsiveness, while RL value reduced and Cdyn value increased in the anti-NGF group compared with the asthma group. HE staining of lung tissue revealed obvious bronchoconstriction, inflammatory cell infiltration around small vessels and alveolar spaces and in interstitum, bronchial epithelial cells desquamation in the asthma group. In anti-NGF group, tracheal epithelium was relatively complete, inflammatory exudation, bronchoconstriction and inflammatory cell infiltration were milder compared to the asthma group. Electron microscopy showed vacuolated changes of adrenal medulla cells, uneven distribution of chromaffin granules in the asthma group and the NGFgroup, and the quantity and concentration of chromaffin granules were significantly lower than normal. There were villous clubbing processes on the adrenal medulla cell membrane in the NGF group. While the anti-NGF group had no significant vacuolar changes in chromaffin granules and the concentration was close to normal. Image analysis showed that mean gray values of PNMT and NGF in the anti-NGF group were significantly different fromthe asthma group. The ELISA results showed that: ( 1) The average concentrations of epinephrine in each group were as follows, ie. the control group gt; anti-NGF group gt; asthma group gt; NGF group. ( 2) The average concentrations of norepinephrine in each group were as follows, ie. the NGF group gt; asthma group gt; anti-NGF group gt; control group. ( 3) There was no significant difference among the groups in the average concentration of cortisol. ( 4) The average concentrations of norepinephrine in each group were as follows, ie. , the control group gt; anti-NGF group gt; asthma group gt; NGF group. Conclusions Local embedding of anti-NGF microspheres can alleviate inflammatory infiltration in lung tissue and improve lung function of rat model with asthma. The mechanismmay be the anti-NGF antagonists the NGF receptor and reverse adrenal medulla cell transdifferentiation process primined by NGF.
ObjectiveTo investigate the effect of nerve growth factor (NGF) on recuperate of optic nerve after contusion by clamping in adult rabbits.
MethodsSixteen adult rabbits were randomly divided into NGF and the control group with 8 rabbits in each group. After the optic nerve of the right eyes was clamped，tissue engineering nerve containing 0.06 ml NGF（concentration: 5×10-4 g/L, NGF group） and 0.06 ml of PBS (control group) was immediately transplanted into the injured eyes respectively, and 0.02 ml NGF（concentration: 5×10-4 g/L, NGF group）and 0.02 ml of PBS (control group) were injected into the vitreous of right eyes respectively. Flash visual evoked potential (FVEP) test was performed on the eyes 1 day, 2 weeks and 8 weeks after the injury. The number of retinal ganglion cells (RGCs) and changes of optic nerves were observed by light microscopy and electron microscopy at the 8th week after contusion,and a computer-image-analysis system was used to count the optic nerve axons.ResultsThe ratio of amplitude of FVEP of the injured and healthy eyes was 0.765±0.150 in NGF group and 0.494±0.108 in the control at the 2th week after injury with a significant difference between the two groups (Plt;0.05); and was 0.581±0.138 and 0.409±0.119 respectively at the 8th week after contusion with statistical difference between the two groups (Plt;0.05). The results of light microscopy and electron microscopy showed that degeneration of RGCs and optic nerves in the NGF group was lighter than that in the control group 8 weeks after injury, while the amount of optic nerve axons was (10 955±608.7) axons/ mm2 in the NGF group and (7 898±608.8) axons/mm2 in the control with statistical difference between the two groups (Plt;0.05).
ConclusionNGF may redound to the survival of RGCs and regeneration of the axons in some degree, which can promote the recuperation of optic nerve and visual function. (Chin J Ocul Fundus Dis, 2005,21:253-257)
OBJECTIVE: To investigate the effect of nerve growth factor(NGF) on the burn wound healing and to study the mechanism of burn wound healing. METHODS: Six domestic pigs weighting around 20 kg were used as experimental animals. Twenty-four burn wound, each 2.5 cm in diameter, were induced on every pigs by scalding. Three different concentrations of NGF, 1 microgram/ml, 2.5 micrograms/ml, 5 micrograms/ml were topically applied after thermal injury, and saline solution used as control group. Biopsy specimens were taken at 3, 5 and 9 days following treatment and immunohistochemistry method was used to detect the epidermal growth factor(EGF), EGF receptor (EGF-R), NGF, NGF receptor (NGF-R), NGF, NGF-R, CD68 and CD3. RESULTS: The expression of EGF, EGF-R, NGF, NGF-R CD68 and CD3 were observed in the experimental group, especially at 5 and 9 days, no expression of those six items in the control group. CONCLUSION: NGF can not only act directly on burn wound, but also modulate other growth factors on the burn wound to accelerate the healing of burn wound.
OBJECTIVE: To investigate the effect of electroacupuncture on mRNA expression of NGF and IGF-1 in injured nerve. METHODS: Sciatic nerve injury model was established by transection of right side sciatic nerve in 90 male SD rats, which were randomly divided into two groups. The experimental group was treated with electroacupuncture, no treatment in the control group. The distal part of the injured nerve was harvested after 1, 2, 4, 6 and 10 weeks of operation and stored in the liquid nitrogen. The total RNA was extracted by the TRIzol reagent. Reverse transcriptase-polymerase chain reaction(RT-PCR) was used to detected the mRNA expression of NGF and IGF-1. RESULTS: The mRNA expression of NGF in the experimental group was increased quickly from the second week, and reached to highest level in the fourth week. It was much higher than that of the control group (P lt; 0.05). Then it began to decline in following time and approximately reached to the level of the first week after 10 weeks of operation. The mRNA expression of IGF-1 in the experimental group was remarkably increased in the second and fourth week, and which was much higher than that of the control group respectively(P lt; 0.05). Although the mRNA expression of IGF-1 after 10 weeks of operation in the experimental group was higher than that of the control group, but there was no significant difference between the two groups(P gt; 0.05). There was linear correlation in the fourth week between mRNA expression of NGF and IGF-1 in the experimental group. CONCLUSION: The mRNA expression of NGF and IGF-1 can be elevated in injured nerve at early stage interfered with electroacupuncture.
ObjectiveTo evaluate the effects of nerve growth factor (NGF) on angiogenesis and skeletal muscle fiber remodeling in ischemic hindlimbs, and study the relationship of NGF and vascular endothelial growth factor (VEGF) to angiogenesis.
MethodsEighteen mice were randomly allocated to normal control group (n=6), blank control group (n=6), and NGF gene transfection group (n=6). The left hindlimb ischemia model was established by ligating the femoral artery. NGF plasmid (125μg) was injected into the mouse ischemic gastrocnemius in the NGF gene transfection group. The same volume of normal saline (200μL) was injected into the mouse ischemic gastrocnemius in the blank control group. The gastrocnemius of left hindlimb was harvested under the condition of peritoneal cavity anesthesia on the 21th day after operation, and then the mice were sacrificed. The gastrocnemius of three groups were tested by hematoxylin-eosin staining, proliferating cell nuclear antigen (PCNA) and CD34 were determined by immunohistochemistry staining. Skeletal muscle fiber type was tested by myosin ATPase staining. NGF and VEGF protein expression were detected by enzyme linked immunosorbent assay.
ResultsOn the 21th day after surgery, compared with the blank control group, the skeletal muscle atrophy degree was weaker, the functional assessment score was significantly lower (P < 0.05), the endothelial cell proliferation index, capillary density, the typeⅠskeletal muscle fiber proportion, NGF and VEGF expression were significantly higher (P < 0.05) in the NGF gene transfection group.
ConclusionsNGF gene transfection could promote NGF and VEGF expression and angiogenesis in ischemic hindlimbs, and induce typeⅠskeletal muscle fibers formation in ischemic hindlimbs. The molecular regulation mechanism still needs to be further studied.
ObjectiveTo observe the efficacy of pars plana vitrectomy (PPV) combined with internal limiting membrane (ILM) peeling and vitreous injection of mouse never growth factor (mNGF) in the high myopia macular hole (HMMH). MethodsA prospective study. Thirty-one patients (33 eyes) with HMMH diagnosed in Affiliated Eye Hospital of Nanchang University from August 2020 and February 2021 were selected. Before surgery, all included patients were subjected to a complete ophthalmologic evaluation including best corrected visual acuity (BCVA), optical coherence tomography (OCT), macular microperimetry and axial length measurement. The BCVA examination was carried out using the international standard visual acuity chart, which was converted into logarithm of minimum resolution angle (logMAR) visual acuity during statistics. The included subjects were accepted the treatment of PPV combined with ILM peeling and vitreous injection of mNGF (combined group) or PPV united with ILM peeling (simple group), 15 cases with 16 eyes, 16 cases with 17 eyes, respectively. There were no significant differences in logMAR BCVA (t=0.836), macular hole (MH) diameter (t=0.657), visual acuity (VA) (t=0.176), the missing length of external limting membrane (ELM) and ellipsoid zone (EZ) (t=1.255, 0.966) between two groups (P＞0.05). The follow-up time was at least 6 months. The BCVA, closure rate of MH, integrity of ELM and EZ and recovery of VA in macular area were compared and observed between the two groups after surgery. The logMAR BCVA, VA, the deficient lengths of ELM and EZ at different time points were compared by independent-samples t-test between two groups and analysis of variance was used to compare the repeated measurement data of each group. Fisher test was performed for comparison of count data. ResultsSix months after surgery, MH closure rates in the simple group and the combined group were 88.24% (15/17) and 93.75% (15/16), respectively, with no significant difference (P=0.523). At 3 and 6 months after surgery, the integrity recovery of ELM in the combined group was better than that in the simple group, and the difference was statistically significant (t=2.282, 3.101; P＜0.05). At 1, 3 and 6 months after surgery, EZ deletion length in the combined group was lower than that in the simple group, and the difference was statistically significant (t=1.815, 2.302, 2.784; P＜0.05). Compared with 1 week after surgery, VA in macular area of the combined group increased at 1, 3 and 6 months, and the difference was statistically significant (P=0.007, ＜0.001, ＜0.001). At 3 and 6 months after surgery, VA in macular area of affected eyes in the combined group was higher than that in the simple group, and the difference was statistically significant (t=1.897, 2.250; P＜0.05). There was an interaction effect between the surgical method and the follow-up time. The postoperative time was prolonged, and the VA in macular area was decreased in the simple group and increased in the combined group, with statistical significance (F=12.963, P＜0.001). The BCVA and BCVA changes in the two groups increased with the extension of postoperative time. The improvement of BCVA and the difference of BCVA changes in the combined group were significantly higher than those in the simple group at different time points after surgery, with statistically significant differences (F=12.374, 21.807, 5.695, 4.095; P＜0.05). ConclusionPPV combined with ILM peeling and vitreous injection of mNGF is more effective than PPV with ILM peeling for HMMH, improving both anatomical and functional outcomes.
