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 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 study the effects of growth factor on the proliferation of the cultured huamn retinal glial cells. Methods:EGF(0.5～100.0ng/ml) and NGF (0.5～10.0ng/ml) were added to cultures of human retinal glial cells and the proliferation rates of the cells were measured by MTT method. Results:EGF at a dosage ranging from 0.5ng/ml to 100.0ng/ml and NGF (0.05～10.0ng/ml) stimulated the cellular proliferation effectively with their EC 50 of 17ng/ml and 0.7 ng/ml respectively. Conclusion:Both EGF and NGF NGF had an effective stimulation on human retinal glial cell proliferation.They may play a role in the formation of PVR. (Chin J Ocul Fundus Dis,1998,14:33-34)
OBJECTIVE: To observe the effect of nerve growth factor (NGF) and nimodipine (NP) on fetal spinal cord graft in repair of injury of spinal cord. METHODS: A total of 144 adult Wistar rats were included in this study. All were made as the hemi-section cavity injury model at the lumbar enlargement and divided into three groups: fetal spinal cord graft (group Tr), fetal spinal cord graft with NGF (group TN), and fetal spinal cord graft with NGF and NP (group TNN). The intracellular concentration of free ionic calcium was measured at the 4th, 8th, and 24th hour, and superoxidase (SOD) and malondialdehyde (MDA) at 3rd, 6th, 12th, 24th and 72nd hour after operation. RESULTS: After spinal cord was injured, the concentration of MDA and intracellular concentration of free ionic calcium increased and reached to the peak at the 6th and 8th hour respectively, but SOD decreased and at 24th hour to its vale. The MDA was significantly lower in group TN than in group Tr, while the SOD was higher (P lt; 0.05). There was no significant difference on intracellular free ionic calcium concentration between group Tr and TN. The concentration of SOD of group TNN was the highest and the intracellular concentration of free ionic calcium was the lowest in the three groups (P lt; 0.05). The weekly mortality was 33%, 31%, 17% respectively in group Tr, TN and TNN. The mortality of group TNN was significantly lower than the other two groups (P lt; 0.01). CONCLUSION: Although the fetal spinal cord graft is an effective method to repair laboratory spinal cord injury, NGF and ND can interrupt secondary injury and increase survival rate of the host.
Objective To investigate the influence of nerve growth factor (NGF) on neuroal regeneration of somatovisceral heterogenic reinnervation using a rat phrenic-to-vagus anastomosis model. Methods Forty male SD rats, aging 3 months and weighing 200 g, were selected and randomly divided into 3 groups. In group A (n=10, control group), phrenic and vagusnerves were exposed and no neurorraphy was performed. In group B (n=15) and group C (n=15), both nerves were transected and proximal stump of phrenic nevers were microsurgically anastomosed to the distal stump of vagus nerves. Postoperatively, group C was intraperitoneally injected with NGF (20 μg/kg·d), while groups A and B were given matching sal ine solution. Twelve weeks later, cardiac function was examined under electrical stimulation of the regenerated nerve. Light and electron microscopies were used to examine the heterogenic regenerated nerve, and the passing rate of axon and thickness of myel in sheath were calculated. Results Under electrical never stimulation in groups A, B, and C, the decreases of blood pressure were (20.12 ± 2.57), (10.63 ± 2.44), and (14.18 ± 2.93) mmHg (1 mmHg=0.133 kPa), respectively; and the decreases of heart rate were (66.77 ± 9.96), (33.44 ± 11.82), and (43.27 ± 11.02)/minutes, respectively. In group B, the decrease ampl itudes of blood pressure and heart rate were 52.83% and50.08% of group A, respectively. Blood pressure and heart rate in group C also decreased dramatically; the decrease ampl itudes of blood pressure and heart rate in group C were 70.48% and 64.80% of group A. There were significant differences in the decrease ampl itudes of blood pressure and heart rate (P lt; 0.05) between group B and group C. Morphological observation showed that heterogenic nerve fibers had the structure of matured myel in sheath and their axons could regenerate into the vagus nerve. In group B and group C, the passing rates of axon were 66.83% ± 4.46% and 81.63% ± 3.56%, respectively; and the thicknesses of myel in sheath were (0.25 ± 0.10) μm and (0.46 ± 0.08) μm, respectively; showing significant differences (P lt; 0.05) between group B and group C. Conclusion Heterogenic nerve is primarily a somatic motor nerve; NGF can promote the axons of heterogenic nerve to regenerate into the parasympathetic nerve.
It has been proved that the bovine seminal plasma contains rich source of NGF around 0.1mg of pure NGF can be isolated from 10ml bovine seminal plasma. Modifying Gregory s method, we first successfully obtained the low molecular weight form of bovine NGF in China.We chose the dorsal root sensory ganglia (DRG) of embryonic chicken as a cultured nerve tissue, the NGF purified from seminal plasma is added to cultural plate with 96 holes. The cultural process was without plus serum and showed the high biological activity. It was found that this method has the advantages of simple technique, satisfactory result. It is an ibeal method for assaying the biological effect of NGF.
Objective To investigate the possibility of constructing eukaryotic expression vector for human glial derived neurotrophic factor （hGDNF）, transfecting it to spinal cord tissue of rats so as to treat acute spinal cord injury. Methods The eukaryotic expression vector pcDNA3-hGDNF was constructed by recombinant DNA technique, transfected into glial cell and neuron of spinal cord by liposome DOTAP as experimental group. In control group, mixture of empty vector and liposome was injected. The mRNA and protein expressions of hGNDF were detected by RT-PCR and Western blot. Results After the recombinant eukaryotic expression vector for hGDNF was digested with Hind III and XbaⅠ, electrophoresis revealed 400 bp fragment for hGDNF gene and 5 400 bp fragment for pcDNA3 vector. In the transfected spinal cord tissue, the mRNA and protein expressions of hGDNF gene were detected with RT-PCR and Western blot. Conclusion The constructed eukaryotic expression vector pcDNA3hGDNF could be expressed in the transfected spinal cord tissue of rat, so it provide basis for gene therapy of acute spinal cord injury.
Objective To construct a bioengineered dermis containing microencapsulated nerve growth factor (NGF) expressing -NIH3T3 cells and to study the effect of the microencapsule on the bioengineered dermis and acute wound healing. Methods A recombinant NGF (PcDNA3.1+/NGF) was constructed and transfected intoNIH-3T3 cells using FuFENETM6 transfection reagent. Positive cell strain was cultured and enclosed in alginate-polylysine-alginate(APA) microcapsules in vitro. Bioengineered dermis was incorporated with NGF-expressing micorencapsules and human fibroblast cells as seed cells using tissue engineering method. The characteristics of the dermis were described by the content of Hydroxyproline(Hyp), HE staining. The content of NGF in the dermis culturing supernatant was measured by ELISA method. These bioengineered dermis were transplanted onto the acute circular full thickness excisional wounds on the dorsum of each swine to observe the rate of reepithelization and wound healing: NGFNIH3T3 microencapsulations(group A), NIH3T3 microencapsulations( group B), empty microencapsulations (group C), NGF incorporated with collagenⅠ( group D) and blank (group E as control group). Results NGF can be tested stably about 124.32 pg/ml in the dermis culturing supernatant after 6 weeks, and the content of Hyp in group A was 69.68±6.20(mg/g wet weight) and increased about 2 times when compared with control groups after 1 week. The tissue engineering skin grafts which can secrete NGF were used to ure the acute wounds and the rate of reepithelization was promoted. The periods of wound healing were 25±2 days in group A, 34±3 days in group B, 34±2 days in group C, 33±2 days in group D and 40±3 days in group E.The period of wound healing was decreased about 10 days at least. Conclusion NGF-expressing NIH3T3 microencapsulates can promote the quality of bioengineered dermis and alsopromote acute wound healing.
Abstract To observe the effect of exogenous high molecular weight nerve growth factor (HMW-NGF) mixed with bletilia striata gelatin (BSG) in the promotion of healing, the experiment was performed as follow: (1) In serumfree medium, the normal saline, BSG, HMWNGF, and BSG+HMW-NGF were added separately, and then, the chick embryo root ganglions (DRGs) were cultivated in the above prepared media and the axonal growth was observed. (2) 40 SD rats were divided into 4 groups. A wound of 2cm×2cm was made on the back of every rat. No treatment was given in group one. In other groups, BSG, HMW-NGF, andBSG+HMW-NGF were given separately to the wounds once daily. After 3 and 10 days, the wound area of every rat was measured, cells in the wounds were observed under light microscope and were calculated, and the time of healing was recorded. The results showed that BSG, HMW-NGF, especially BSG+HMW-NGF could promote wound healing.