Research The Effects Of Tacromulis On Rats With Experimental Autoimmune Encephalomyelitis

Objective: to explore the efficacy and therapeutic mechanism of tacromulis(FK506) in treating rats with experimental autoimmune encephalomyelitis (EAE),which is an animal model of multiple sclerosis, by observing clinical the features, counting heir peripheral blood lymphocytes CD3 ,CD4 ,CD8 and NK subsets,pathology changes of brain and spinal cord, and to show the evidence of nerve regeneration by GAP-43 immunohistochemistry stain.Methods: (1) the guinea pig spinal cord homogenate in complete freund's adjuvant and bordetella pertussis vaccine were both used to induce the acute EAE in SD rasts . (2) The experimental rats were divided into normal control group, EAE group, FK506A group and FK506B group,prednisolone group, randomly, respectively, to treat the FK506A group and FK506B group rats with FK506 at the dosage of 1mg/kg, 0.5mg/kg was intraperitoneal injection daily given ten days after the immunization. Prednisolone group was intraperitoneal injection daily given ten days after the immunization at the dosage of 10mg/kg. The rats in EAE model group were treated in stead of equal normal saline. (3)After immunization, change of body weight and clinical score of the experimental animal were recorded everyday. 20th day After immunization counting peripheral blood lymphocytes CD3 ,CD4 ,CD8 and NK subsets. The pathological changes of central nervous system was observed by HE staining , Luxol fast blue–HE staining , Bielschowsky staining . to show the evidence of nerve regeneration by Gap-43 immunohistochemistry staining.Result: (1) The rats immunized in our study showed typical clinical manifestation and pathological changes of acute EAE. (2) in EAE model group ,maximum clinical score was (3.02±0.98 ), maximum loss of body weight was (39.62±13.64)g , latency was (10.21±1.08) d disease duration was (16.52±0.83 )d, in FK506A group maximum clinical score was(1..21±0.31), maximum loss of body weight was ( 12.52±5.26)g , latency was (14.61±1.74)d, disease duration was (10.34±0.61)d, in FK506Bgroup maximum clinical score was(1.52±0.36 ), maximum loss of body weight was (11.34±5.19), latency was (14.08±1.39 ), disease duration was (10.91±1.21 )d. in Prednisolone group maximum clinical score was(1.67±0.22 ), maximum loss of body weight was (-26.12±7.34), latency was (10.41±0.81 ), disease duration was (16.08±1.39 )d. observed in HE staining, the inflammatory infiltration of Bain, Cerebellum and Spinal cord in EAE group were(2.61±0.47),(3.17±0.29),(3.86±0.44) respectively, which wer(e1.31±0.54), (1.94±0.27), (2.08±0.23)in FK506A group , (1.27±0.69), (1.87±0.35),(2.12±0.76)in FK506B group, (1.12±0.41),(2.31±0.14),(2.29±0.58)in Prednisolone group. Show by Luxol Fast Blue staining, the demyelination of Brain, Cerebellum and Spinal cord in EAE group were (1.59±0.83),(2.07±0.74),(2.21±0.51)respectively, which were (0.86±0.31),(1.29±0.64),(1.44±0.52)in FK506A group , (0.87±0.17),(1.34±0.48),(1.51±0.72)in FK506B group, (0.81±0.33),(1.27±0.58),(1.17±0.28), in Prednisolone group. show by Bielschowsky staining, axonal damage of spinal cord In EAE group were (1.91±0.64),which were (1.17±0.26)in FK506A group, (1.31±0.37)in FK506B group, (1.06±0.14)in Prednisolone group. In EAE model group CD3+T cell ratio is (67.22±1.21), CD4+T cell is (51.27±2.13), CD8+T cell is (28.16±1.53)NK cell is (8.52±1.16),CD4+/CD8+ratio is (1.82±0.26), in FK506A group CD3+T cell ratio is (58.17±0.96), CD4+T cell is (41.34±1.41), CD8+T cell is (14.43±0.74), CD4+/CD8+ratio is (2.61±0.21) NK cell is (1.07±0.41), FK506B group CD3+T cell ratio is (60.28±2.13), CD4+T cell is (44.12±2.69), CD8+T cell is (13.2±1.17) , CD4+/CD8+ratio is (2.84±0.51), NK cell is (1.89±0.21), Prednisolone group CD3+T cell ratio is (71.06±3.83), CD4+T cell is (55.21±4.78), CD8+T cell is (26.34±1.72), CD4+/CD8+ratio is (2.09±0.38) NK cell is (7.62±0.49).Conclusion: (1) The method establishing acute EAE model was successful, stable and reliable in our study. (2) Tacromulis is an effective drug in treating EAE as it can relieve the clinical signs and ameliorate pathological lesion in central nervous system of acute EAE rats. (3) Compare with Prednisolone group, Tacromulis withdrawal did not initiate relapse with return of clinical features and inflammatory infiltrate within the CNS, Tacromulis group has delayed the onset time of EAE, shorten the course of disease, (4) The therapeutic mechanism of tacromulis on EAE is related to suppressing immune response of T lymphocyte cell, remotes axons regeneration of central nervous system.