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Objective To constructan eukaryotic expression recombinant plasmid named pEGFP-N2-PRNP .Methods Total RNA was extracted from alzheimer (AD) disease peripheral blood ,and the PRNP gene was amplified by reverse transcription-poly-merase chain reaction(RT-PCR) .By using gene recombination technique ,human PRNP cDNA was inserted into retroviral vector pEGFP-N2 .The recombinant plasmid was identified by a pair of specified primers containing the restriction sites of Xho Ⅰ and EcoRⅠ .Results The PRNP gene could be obtained by RT-PCR ,the recombinant plasmid was identified by restriction endonucle-ase analysis ,PCR and sequence analysis ,and the expression vector pEGFP-N2-PRNP ,which could be stably expressed in SH-SY5Y cells .Conclusion The recombinant plasmid pEGFP-N2-PRNP is constructed successfully ,Which offers a basic for the further re-search on PRNP biological fuction .
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BACKGROUND: Mature brain contains neural stem cells. It is very important that how to directionally induce the differentiation of neural stem cells into a specific neuron, substitute for damaged neurons, in effective treatment of nervous system disease.OBJECTIVE: To study the role of vascular endothelial growth factor in the proliferation and differentiation of neural stem cells in vivo.Neurobiology, Xiangya School of Medicine, Central South University from June 2007 to June 2008.MATERIALS: A total of 24 healthy male Sprague Dawley rats were randomly assigned to model control, an'dbody treatment and sham operation groups.METHODS: Rat models of hippocampal fimbria and fomix transsection were established in the model control and antibody treatment groups. Rats in the sham operation only received digging skull operation. Immediately following model induction, rats in the antibody treatment group were subjected to 4 μL anti-vascular endothelial growth factor antibody. The needle was inserted at anterior fontanelle+0.6 mm, lateral side+0.6 mm and ventral side-5.5 mm at the affected side. Rats in the model control and sham operation groups were subjected to an equal volume of saline.MAIN OUTCOME MEASURES: The expression of vascular endothelial growth factor, Nestin and proliferating cell nuclear antigen was observed using immunohistochemical method. Proliferating cell nuclear antigen index was calculated.RESULTS: Compared with the sham operation group, the expression of vascular endothelial growth factor and Nastin was significantly increased at the septal area following hippocampal fimbria and fornix transsection in the model control group (P < 0.01). Compared with the model control group, Nestin expression was significantly decreased in the antibody treatment group (P< 0.01). The proliferating cell nuclear antigen diffusely expressed in neuronal cytoplasm in the sham operation group, with the presence of non-specific staining, and the proliferation index was nearly 0. A little proliferating cell nuclear antigen was found in the model control group, with a proliferating index of 1%. Following antibody treatment, the proliferating index was decreased to 0.CONCLUSION: Following septal area damage, the increased expression of vascular endothelial growth factor induces the occurrence of neural stem cells. High expression of vascular endothelial growth factor may be the promoting factor of occurrence and differentiation of neural stem cells, and the basis of self-repair following bran damage.
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Objective To study the temporo-spatial changing laws of calcitonin gene-related peptide(CGRP)in the rat model of sciatic nerve ligation.Methods Totally 48 Sprague-Dawley rats were randomly divided into sham-operation group(n=6)and experimental groups which survived for 1,3,5,7,14,21,28 d respectively after sciatic nerve ligation(n=6 at each time point).The distribution and quantity of CGRP in sciatic nerve,dorsal root ganglion(DRG)and spinal cord were detected by immunohistochemistry and image analysis methods.Results A large amount of CGRP piled up in the sciatic nerve at 1 d after the ligation,significantly higher in proximal segment than that in distal segment,gradually dropping till disappear basically at 14 d in proximal segment and at 7 d in distal segment.The expression of CGRP in DRG at 7 d began to drop after the ligation,at 21 d to the minimal value and at 28 d lower than normal.The CGRP-immunoreactivety in ipsilateral spinal dorsal horn at 14 d decreased after operation,but the immuno-positive areas of CGRP were of no changes.There are no changes in the CGRP-positive spinal ventral motoneurons of all groups.Conclusion The changes of CGRP presented a temporo-spatial pattern following peripheral nerve legation,maybe resulting from the deficiency of neurothrophins from target organs.
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Objective To evaluate the feasibility of endoscopic transoral-transpharyngeal approach to the upper cervical. Methods Anatomic characteristics were observed and measured in the anterior column of 50 dry atlas and axis specimens. Conventional and endoscopic methods to decompress the spinal cord and excise the cartilage surface of the atlantoaxial joint by transoral-transpharyngeal approach were taken respectively in two groups of cadaveric heads and necks. All the cadaveric specimens were then open dissected to evaluate endoscopic operation methods, decompression size and the "safe zone". Results The anterior arch of atlas was of a length of (19.8?2.3) mm, the height of odontoid was (15.9?1.9) mm, the width (10.5?0.6) mm, and the thickness (11.5?1.9) mm; the maximal transverse diameter of superior facet of axis was (15.1?1.6) mm, and the anteroposterior one was (17.7?1.3) mm. The anterior tubercle of the atlas could be acted as landmark leading to the endoscopic atlantoaxis surgery. The arch could be drilled either from the tubercle to the lateral side or broken from the junction to the lateral mass. Endoscopic odontoid dissection should begin at the apex of the odontoid, and proceed inferiorly. It was necessary to move or slope the working tube to explore atlantoaxial lateral joint and dissect its cartilage, but the width and depth of cartilage dissection should be limited to 12 mm and 10 mm in order to avoid damage to vertebral artery and spinal cord. Measurements after postoperative open dissection showed that endoscopic decompression size were not significantly different from that of conventional method. There was a "safe zone" in the front of atlantoaxis of transoral-transpharyngeal approach, with (45.9?3.6) mm wide and (29.4?2.5) mm high. Conclusion Endoscopic transoral-transpharyngeal approach to the upper cervical is technically feasible, which had a good exploration and could get the same decompressing size with conventional transoral-transpharyngeal approach.