BACKGROUND:Vascular endothelial growth factors (VEGF) can promote the division of endothelial cells and accelerate the growth of newborn vessels, whereas the expression and distribution of VEGF receptors (VEGFR) in spinal cord should be observed.OBJECTIVE: To observe the expression and distribution of VEGFR-1 and VEGFR-2 in rat spinal cord.DESIGN: A randomized controlled observation.SETTING: Department of Orthopaedic Surgery, the First Affiliated Hospital of Dalian Medical University; Department of Hand Surgery, Union Hospital affiliated to Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: Ten adult male Wistar rats of clean degree, weighing 180-200 g, were provided by the animal experimental center of Tongji Medical College, Huazhong University of Science and Technology. Immunohistochemical primary antibody was purchased from Santa Cruz Company; the second and third antibodies from Beijing Zhongshan Biotechnology Co., Ltd. Reverse transcription-polymerase chain reaction (RT-PCR) kit was the product of Promega,Trizol reagents were purchased from Invitrogen Company, and the VEGFR-1 and VEGFR-2 primers were designed by Beijing Aoke Company.METHODS: The experiment was carried out in the laboratory of Department of Hand Surgery of Wuhan Union Hospital from March to June in 2004. ① Detection of VEGFR expression in spinal cord anterior horn: The rats were anesthetized by 100 g/L chloral hydrate, spinal cord of lumbar 4-6 (L4-6) was fixed in fixation solution at 4 ℃ for overnight, then routine dehydration, hyalinization and paraffin embedding were performed, and serial sections of about 5 μm were prepared for observing the VEGFR expressions using immuniohistochemical staining. ② Detection of VEGFR mRNA expression in spinal cord: Five rats were selected, L4-6 spinal cord (50 mg) was removed and centrifugated, then the content of total RNA was determined with ultraviolet spectrophotometer. The synthetized cDNA was amplified with PCR, and the PCR products (10 μL) were treated with 20 g/L agarose gel electrophoresis, and stained with 0.1 mg/L ethidium bromide, the results were observed and recorded under ultraviolet lamp and photographed. The products were scanned and quantified with gel imaging analytical system to record the gray value of each band, the gray value of β-actin band was taken as 1,and those of the other objective fragments were compared with it to record the gray value ratio and analyze the expressions of the objective fragments.MAIN OUTCOMEMEASURES: ① Expression and distribution of VEGFR-1 and VEGFR-2 in rat spinal cord anterior horn;② Results of VEGFR mRNA expression in spinal cord.RESULTS: ① Both VEGFR-1 and VEGFR-2 were expressed in the microvessels of normal rat spinal cord tissue.Besides, VEGFR-2 mainly expressed in motor neurons, glial cells and the nerve fibers in surrounding white matter. ②The results of gel imaging analytical system showed that the VEGFR-2 content in normal spinal cord was obviously higher than VEGFR-1 (0.874±0.222, 0.486±0.181, P ＜ 0.05).CONCLUSION: VEGF promote the formation of microvessels through the combined effects of VEGFR-1 and VEGFR-2,and it plays the neurotrophic and neuroprotective role through VEGFR-2.

BACKGROUND:Recent development of bioengineering technology and tissue-engineered nerve brings a new hope for the treatment of peripheral nerve injuries, which has gradual y become a research spot. OBJECTIVE:To review the new progress in the repair of peripheral nerve injuries using seed cells, biomaterials and tissue-engineered nerve construction technology. METHODS:PubMed and CNKI were searched by the first authors for articles concerning nerve tissue engineering and repair of peripheral nerve injuries published prior to July 2013. The keywords were“tissue engineering, peripheral nerves, nerve injuries, stem cells, Schwann cells, scaffold, growth factor”in English and Chinese, respectively. The articles published recently or in the authorized journals were preferred in the same field. Final y, 63 articles were included in result analysis. RESULTS AND CONCLUSION:Up to now, there is a great advance in the tissue engineering technology for the repair of peripheral nerve injuries. However, most studies are stil in experimental step. For the clinical application of nerve tissue engineering, some problems to be solved include:(1) source and ethics of seed cells;(2) immunological rejection fol owing cellproliferation and transplantation;(3) stability and oncogenicity of transplanted cells;(4) degradation rate, optimal porosity, tube thickness and shape;(5) repair timing for in vitro tissue-engineered nerve construction;(6) local release and regulation of various neurobiological factors. With the development of science, many patients with nerve injuries can profit from the solve of these problems.

This study is to investigate the effect of local phVEGF(165) injection on sciatic nerve regeneration in the rats and to search for a new way in the further treatment of peripheral nerve injuries. Forty-five adult male Wistar rats received a neurotomy to bilateral sciatic nerves, which were subsequently reconnected with 10/0 epineurial nylon sutures. The injured segments was locally injected with normal saline (group A), or 25 microg of phVEGF(165) (group B) or 50 microg phVEGF(165) (group C). Nerve conduction and regeneration were evaluated in terms of the histological changes, weight of gastrocnemius muscles, electrophysiology and morphometric results. Our study demonstrated that rats of group C showed the best results in terms of nerve regeneration, followed by group B and group A. Our findings suggested that local injection of phVEGF165 can facilitate nerve regeneration and promote functional recovery in a dose-dependent manner.

BACKGROUND: Secretion of various neurotrophic factors by Schwann cells plays important roles in neural regeneration. However, the secretion capability is affected by many factors. To seek a feasible method for promoting nerve growth factor secretion by Schwann cells is a key of regeneraion following neurologic defect.OBJECTIVE: To explore the effects of methylprednisolone(solu-medrol) on the secreted function of Schwann cells of cultured rats.METHODS: Schwann cells were isolated and cultured by enzyme digestion method. Cell growth was observed under an inverted phase contrast microscope. Following passage, purity of some Schwann cells was identified using S-100 protein immunity. Other Schwann cells were regulated using cell counting plate into 1×10~9/L, and incubated in a 6-well culture plate (15 wells) for further incubation. Following 4 days of culture, different concentrations of solu-medrol (10~(-3), 10~(-4), 10~(-6), 10~(-8) mol/L) were administrated to the cell, while blank control group (1 well) was given no drug. 24, 48 and 72 hours after administration, reverse trancription-polymerase chain reaction (RT-PCR) was used in the detection of the levels of nerve growth factor mRNA.RESULTS AND CONCLUSION: Number of primarily cultured cells was significantly increased at day 7, and 80% cells were confluent. Subcultured cells were spindle-shaped, with 2 thin long processes, showing positive fluorescence staining. Fibroblasts were round or flat, showing negative reaction of fluorescence staining. Reserve transcription-polymerase chain reaction demonstrated that nerve growth factor number at 72 hours affected by 10~(-8) mol/L radiosone was increased compared with the blank control group and other concentrations and other time points (P < 0.05). Number of nerve growth factor was reduced following treatment of 10~(-3) mol/L radiosone compared with the blank control group and other concentrations (P < 0.05). These results suggested that high concentration of solu-medrol prohibits secreted function of Schwann's cells, but long time and low dosage solu-medrol promotes secreted function of Schwann's cells.

ObjectiveTo evaluate the efficacy of mini-incisional double-tsuge suture method with 0-0 absorbable polydioxanone-cord (PDS-Ⅱ)in repair of acute achilles tendon rupture.MethodsA total of 34 patients were subjected to acute closed achilles tendon ruptures,including 25 males and 9 females at a mean age of 32 years ( range,20-45 years).Injury causes included sports injuries in 27 patients,falling injuries in six and heavy object impingement injury in one.The time from injury to operation was average 3 days (range,1-6 days).All patients underwent minimally invasive repair with double-tsuge suture method by using PDS-Ⅱ.The ankle joint was fixed with short leg plaster cast at 30° plantar flexion position and the cast was removed six weeks later to take functional exercise.The patients could walk with full weight-bearing 8-10 weeks later and could gradually return to activity 3-4 months later.Results There was one patient with poor incision healing and one patient with reflex sympathetic dystrophy postoperatively.The rest patients had stage Ⅰ incision healing without skin adhesions.No complications such as infection,lower extremity deep venous thrombosis or sural nerve injury occurred postoperatively.All the patients received follow-up of 12-24 months (average 15 months),which showed no complications like tendon rerupture occurred.According to clinical evaluation criterion of Termann,the average score was 92 points (range,76-96 points).The result was excellent in 28 patients,good in five and fair in one,with excellence rate of 97％.ConclusionsSmall incisional double-tsuge suture method achieves low rate of complications and good outcomes for repairing acute achilles tendon rupture and is an ideal tendon surgery approach.

BACKGROUND:The principle of lyophilization is to sublimate the solvent of frozen materials in vacuum and retain the solute, thus making a pore structure. OBJECTIVE: To produce a chitosan tubular scaffold by lyophilization, and to test its physicochemical properties. METHODS: The chitosan tubular material was prepared by lyophilization method, folowed by gross observation and electron microscopic observation. The chitosan tubular material samples were placed into PBS solution and pure water for 50 days, respectively, and then immersed in trypsin liquid for 1 day folowed by embedded into the muscle and dorsal skin of neonatal Sprague-Dawley rats for 30 days. The degradation rate and porosity of the material were observed and calculated. The breaking strength and compressive strength of the material were determined both under drying and soaking conditions using tensile instrument and pressure meter, respectively. RESULTS AND CONCLUSION:The external form of the chitosan tubular material was normaly tubular. Under the electron microscope, it was composited by different size pores, and the pore size was 50-200μm. The degradation rates of the material were (5.33±0.12)% in PBS, (11.26±0.15) in water, 0.012% in the trypsin liquid and (35.2±3.7)in vivo. The porosity rate was (97.5±1.5)%. The breaking strength and compressive strength of the material was higher under the drying state than under the soaking state (P < 0.05). These findings indicate that the lyophilization method can produce the chitosan tubular material with good porosity rate and degradation rate as wel as good tensile ability and compressive capability.

This study is to investigate the effect of local phVEGF165 injection on sciatic nerve regeneration in the rats and to search for a new way in the further treatment of peripheral nerve injuries.Forty-five adult male Wistar rats received a neurotomy to bilateral sciatic nerves, which were subsequently reconnected with 10/0 epineurial nylon sutures. The injured segments was locally injected with normal saline (group A), or 25 μg of phVEGF165 (group B) or 50 μg phVEGF165 (group C).Nerve conduction and regeneration were evaluated in terms of the histological changes, weight of gastrocnemius muscles, electrophysiology and morphometric results. Our study demonstrated that rats of group C showed the best results in terms of nerve regeneration, followed by group B and group A. Our findings suggested that local injection of phVEGF165 can facilitate nerve regeneration and promote functional recovery in a dose-dependent manner.

Objective: To investigate the effects of Kaixin Powder on learning and memory, ATP/AMP ratio, GABA andiNOS levels in rats with multiple infarct dementia. Methods: The rat model of multi-infarct dementia was established bymicro-thromboembolism. Morris water maze and opening experiment were used to evaluate learning and memoryfunction. The pathological changes of hippocampal CA1 area were evaluated by HE staining. The contents of ATP andAMP in brain tissue were determined by HPLC. The content of γ-aminobutyric acid and iNOS in peripheral blood weredetected by ELISA kit. Results: Compared with the rats of model group, rats of Kaixin Powder group can significantlyshorten the escape latency, increase the number of crossing platforms, Results: Compared with the model group, increasethe standing times in the opening experiment, prolong the exercise time, shorten the resting time, improve the nerve celldamage in the hippocampal CA1 area, and significantly increase the ratio of ATP/AMP of brain tissue, decreased brainGABA content and serum iNOS content (P ＜ 0.05) . Conclusion: Kaixin Powder has the effect of improving learning andmemory function and abnormal motor behavior in rats with multiple infarct dementia. The mechanism is related toreducing GABA and i NOS content in brain tissue, increasing ATP/AMP ratio in brain tissue and improving energysupply in brain tissue.