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Expression and Significance of Pleiotrophin and Vascular Endothelial Growth Factor in Intervertebral Disc

Author: ZhangShengFei
Tutor: JiaChangQing
School: China Medical University
Course: Surgery
Keywords: Multi-effect growth factor Vascular endothelial growth factor Intervertebral disc Degeneration Rats
CLC: R681.53
Type: Master's thesis
Year: 2009
Downloads: 41
Quote: 2
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Abstract


The purpose of disc degeneration (intervertebral disc degeneration, IVDD) is the intervertebral discs lose their normal structure and associated with progressive fibrosis, the gel-like nucleus pulposus disappear, the original nucleus pulposus and annulus fibrosus clear boundaries gradually disappear, the annulus fibrosus board layer rough and fibrosis, the last occurrence of cracks and age pigment deposition. Disc degeneration and disc tissue studies confirm the change of enzyme activity and the release of certain cytokines. Multi-the efficiency growth factor (pleiotrophin, PTN) and vascular endothelial growth factor (vascular endothelial growthfactor, VEGF) is an important stimulus for angiogenesis factor in disc degeneration is not yet very clear, this study different age Wistar rats The morphological changes of lumbar intervertebral disc which PTN and VEGF content changes to clarify the physiological state of rat lumbar intervertebral disc PTN and VEGF expression trend further explore the relationship of PTN and VEGF lumbar disc degeneration. Materials and methods, the grouping of the experimental animals and specimens collected from 50 healthy Wistar rats, male, weighing not limited months of age, divided into experimental group 1, 3, 6, 12 and 18,5 per Group 10. 25g / L sodium pentobarbital intraperitoneal overdose of anesthesia, the rats were killed, each rat to take the fourth and fifth lumbar disc tissue specimens. 5 lumbar disc specimens after removal in 4% paraformaldehyde solution, fixed, 10% ethylene diamine tetra-acetic acid (EDTA) solution decalcified, paraffin-embedded, 4μm serial sections; 4th lumbar disc specimens placed in -70 ℃ refrigerator . 2, the experimental methods (1) → the HE staining xylene dewaxing → gradient alcohol hydration hematoxylin - eosin staining → gradient alcohol dehydration → xylene → gum Fengpian; (2) SABC immunohistochemical staining dewaxing water of → 3% hydrogen peroxide (H 2 O 2 ) inhibition of endogenous peroxidase activity → heat antigen Repair → → BSA closed primary antibody, secondary antibody incubation → dropping SABC reagent → DAB chromogenic → nuclear stained transparent → 1% hydrochloric acid alcohol differentiation → dehydration → the gum Fengpian; choice of phosphate-buffered saline (PBS) instead of primary antibody as a negative control. (3) Western-blot western blot stained tissue crushed → ultrasonic homogenizer, centrifuged, and the extracted proteins, protein quantitation, the the sample → SDS-PAGE gel electrophoresis → transfer membrane → closed, after washing in a primary antibody solution in 4 ° C → secondary antibody solution at room temperature overnight to soak for 2 hours → washing liquid washing the membrane staining to strip to show termination staining. Selection of β-actin as an internal reference, PBS instead of primary antibody as a negative control. (4) The results of determining the PTN and VEGF are mainly located in the cytoplasm to disc cytoplasmic brown staining particles as positive, according to the percentage of positive cells were analyzed. Western-blot test results striped gray value analysis by gel image analysis system. (5) data analysis data expressed as mean ± standard deviation ((?) ± s), and SPSS13.0 statistical analysis software for the analysis of variance, P <0.05 was considered statistically significant. 1, HE staining results as rats-month-old increase in the degree of structural disorder of the disc tissue gradually increased, the nucleus pulposus matrix degradation, the middle appear cavity, the proliferation of collagen fibers, coarse, disorganized, and visible fiber breakage or missing . Expression of PTN and VEGF increase (1-12 months) As the month-old rats, PTN and VEGF expression in the intervertebral disc tissue gradually reduced to 18 months of age, PTN and VEGF expression have increased; PTN and VEGF expression in intervertebral disc cells 6 and 12-month-old group was significantly lower than the 1-month-old group (P <0.01), and 18-month-old group PTN and VEGF expression was significantly higher than the 12-month-old group ( P <0.01). Intervertebral disc cells in the same month group (P <0.05) higher than the expression in the nucleus pulposus and annulus of PTN and VEGF expression in the endplate PTN and VEGF expression in the nucleus pulposus and annulus no significant difference (P> 0.05). PTN and VEGF expression is also visible in some specimens extracellular matrix. Conclusion 1, rat intervertebral disc tissue structure gradually increase as the month increased occurrence of degenerative changes; 2, PTN and vEGF, involved in the rat intervertebral disc degeneration, and may promote the disc tissue angiogenesis, delaying repair intervertebral disc degeneration.

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CLC: > Medicine, health > Surgery > Orthopaedic Surgery ( movement system diseases,orthopedic surgery ) > Bone diseases > Spine and back disorders > Spinal joint disease
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