| IntroductionRheumatoid arthritis (RA) is a chronic inflammatory joint disease led by aberrant immune function. The major pathological changes in RA lesions include irregular proliferation of synovial cells, neoangiogenesis, bone and cartilage destruction in the affected joints. Recently, as discussing the abnormity of immunology in RA, researchers think highly of synovial pathological changes in RA increasingly. It was found that the synovium in RA is a kind of tissue with strong encroachment, its growth and pathological behavior resemble tumor in many ways: synovium growing with hyperplasy and rodent, synovium getting thickening , and pannus being formed. There are lots of cytokines and growth factors involved in the development of pathological changes of synovium, and one of them - vascular endothelial growth factor (VEGF) - may play a considerable role in the development of synovial inflammatory and the forming of synovial pannus.Prolactin(PRL) originally identified as a pituitary hormone with activity of modulating galactogogue action has been proved to have immunoregulatory potential. The relationship between PRL and autoimmune disease has attracted many researchers. Clinical observations and experimental animal studies have already implicated that PRL plays an important role in the pathogenesis of autoimmune disease such as systemical lipus erythematosus, autoimmune thyroid disease and autoimmune Addison's disease etc. Numerous reports described the abnormal plasma level of PRL in RA. The plasma PRL levels elevate and show a significant correlation with RA.The functional mechanism of PRL in RA has multiplicity including (1) the regulation of immune system, (2) the interaction with the inflammatory medi-urns and (3) mediation of the cytokines, etc. However, it needs a further study to make this mechanism precise. To determine the exact role of PRL in the path-ogenesis of RA and supply experimental basis for clinical treatment of RA, we examine the expression of VEGF of adjuvant arthritis at both hypoprolactinemic and hyperprolactinemic levels.Materials and methodsExperimental animals: forty male wistar rats, 6 -8weeks old, weighing 160 ±20 grams.Experimental reagent: Freud's complete adjuvant, diethylstilbestrol, bro-mocriptine (SIGMA) , rabbit anti rat VEGF antibody, immunoglobulin marked with HRP(Boshide Company, Wuhan) ,ECL kit, immunohistochemistry SP kit, NC (Zhongshan Company, Beijing) .Experimental instruments: stir apparatus, low temperature centrifugal machine, electrophoresis apparatus, transmark apparatus, freeze microtome, Luze - F image analysis system.Methods;The forty male wistar rats were devided into the following groups with random: group I : normal rats;group II : adjuvant arthritis rats;group HI : Bromcryptine - induced hypoprolactinemic adjuvant arthritis rats;group IV : Diethylstilbestrol — induced hyperprolactinemic adjuvant arthritis rats. The animals were killed the 21 day after the adjuvant injections. Collected the serum to detect the level of PRL by radioimmunoassay. The synovium was removed and then frozen immediately for Western blot to analyze the expression levels of VEGF. The synovium of some rats was done and kept for immunohistochemistry to analyze the expression levels of VEGF.Data from above methods were expressed as mean ± standard deviation (SD) . The differences between groups were determined by unpaired student t -test. P<0.05 was statistically significant.Result1. The result of radioimmunoassay: The level of PRL in the serum of Group II is more than Group I (625. 34 ±30. 6 vs. 495. 59 ±21. 2, p <0. 05) , and GroupIV is more than Group II (695. 67 ± 30. 8 vs. 625. 34 ± 30. 6, p < 0. 05) , and Group I is less than Group II (562.08 ±28.2 vs. 625.34 ±30.6, p<0.05).2. The result of Western blot method: Western blot analysis revealed that the molecular weight of VEGF was 45kDa. Comparing the IDV ratio of VEGF to P - actin, Group II is more than Group I ( 1. 2941 ±0. 0295 vs. 0. 8761 ±0. 0198, p <0. 05 ) , and GroupIV is more than Group II ( 1. 5387 ± 0. 0376 vs. 1. 2941 ± 0. 0295, p < 0. 05) , and Group I is less than Group II (1. 0082 ± 0. 0264 vs. 1.2941 ± 0.0295, p < 0.05).3. The result joi immunohistochemiatry: Immunohistochemical analysis clearly showed positive staining in synovial cell. Comparing the OD of VEGF, Group II is more than Group I (0. 425 ± 0. 0144 vs. 0. 309 ± 0. 0255 , p < 0. 05) , and Group IV is more than Group H (0. 501 ± 0. 0182 vs. 0. 425 ± 0. 0144, p <0. 05) , and GroupIH is less than Group II (0. 374 ±0. 0188 vs. 0. 425 ±0.0144, p<0.05).DiscussionThere were numerous reports about the relationship between PRL and RA. Clinical observations and experimental animal studies have already implicated the dysfunction of the secretion of PRL in RA, and the plasma PRL levels showed a significant correlation with the severity of RA. After removing the pituitary , the rats not only had low immune function, but also could not be induced to experimental arthritis;while after embedding pituitary or given exogenous PRL, the rats could be induced to experimental arthritis again.VEGF affects specifically vascular endothelial cells. Combining with the receptor, VEGF leads to the endotheliocyte division and hyperplasy. Moreover, VEGF can induce endotheliocyte to secrete lots of catheptic enzyme which degrade extracellular matrix and cause endotheliocytes to migrate and infiltrate and cause neovascularization.There are abundant VEGF in the synovial fluid and synovium of RA which activates monocytes and lymphocytes and promotes the secretion of inflammatory mediators such as b - FGF to aggravate the destruction of joint and the connected tissue. The over expression of VEGF in RA can promote the release of fibrinoly-sin which activates metalloprotease to destruct arthrodial cartilage.This study took the AA rats induced by CFA injection as the animal model of RA, the result of it showed that the VEGF of synovium of knee joint increased in hyperprolactinemic AA and decreased in hypoprolectinemic AA comparing to the A A control.PRL could affect multiple processes of RA including: the regulation of the immune system, the interaction with the inflammatory medium and the regulation of the cytokins, etc.The result of this study indicates that PRL may affect the development of RA by changing the secretion of VEGF. It has been reported that with PRL and BRC treating the RA patients the inflammatory factor and proteolytic enzyme were stimulated and inhibited respectively, which conforms to the result of this study.This study first time revealed the effect of hypoprolectinemia and hyperpro-lectinemia to the expression of VEGF. The changes of VEGF in this study may be led directly by PRL, or maybe led by cytokines affected by PRL indirectly, or may be led by both of them. To make the effect clear still need a further study.Conclusion1. After induction of adjuvant arthritis, the expression of VEGF in synoviumwas increased.2. The level of PRL in rats with adjuvant arthritis could regulate the expression of VEGF in synovium. The hyperprolacineniia caused the high expression of VEGF;however, the hypoprolactinemia lead to the low expression of VEGF.
|
PDF Full Text Request