RESUMO
Objective To study the effects of dendritic cells (DC) modified with transforming growth factor ?1 (TGF-?1) gene on experimental autoimmune myasthenia gravis (EAMG) in Lewis rats. Methods 30 female Lewis rats were divided randomly into 6 groups: normal group, EAMG group, DCs treatment group, pcDNA_3-TGF-?1-DCs treatment group, pcDNA_3-DCs treatment group and normal saline group. The rats were immunized with the acetylcholine receptor (AChR) protein extracted from electric organ of Narcine timilei and completed Freund’s adjuvant (CFA) in the experiment groups except normal group. 2?106 pcDNA_3-TGF-?1-DCs/rat were injected subcutaneously into the backs of the rats that had been immunized 5 days earlier with AChR+CFA. The rats in DCs treatment group, pcDNA_3-DCs treatment group and normal saline group were injected in parallel with untreated DCs, pcDNA_3-DCs and normal saline respectively. Then the clinical manifestations were observed everyday. And 7 weeks after the first immunization, repetitive nerve stimulation, detection of acetylcholine receptor antibody (AChRab) and ultrastructural study of neuromuscular junction (NMJ) were performed. Results (1) The mild symptoms were observed on 1 or 2 rats in the experiment groups except normal group after a week, which lasted for 2 to 5 days. After about 5 weeks, the rats in EAMG group, DCs treatment group, pcDNA_3-DCs treatment group and normal saline group presented some symptoms at different degree like myasthenia gravis, and only one of the rats in pcDNA_3-TGF-?1-DCs treatment group presented mildly decreased activity. (2) The significant decrement of repetitive nerve stimulation were found in EAMG group, DCs treatment group, pcDNA_3-DCs treatment group and normal saline group(16.75?6.13, 17.75?7.81, 18.25?8.22 and 16.50?7.14, respectively), but there was no attenuation in pcDNA_3-TGF-?1-DCs treatment group and normal group(3.20?3.70 and 5.60?2.70, respectively). The percentage of decrement in pcDNA_3-TGF-?1-DCs treatment group was lower than that in EAMG group(5.60?2.70 and 16.75?6.13, respectively,P0.05). (4) The combined AChRs in NMJ of the rats in pcDNA_3-TGF-?1-DCs group were higher than that in EAMG group, and the structure changes of the synapse were relieved.Conclusion It suggests that DCs, transfected with pcDNA_3-TGF-?1, when injected subcutaneously into Lewis rats with incipient EAMG, could inhibit the production of AChR-Ab, relieve the pathologic changes in NMJ and ameliorate the development of EAMG.
RESUMO
Objective To induce experimental autoimmune myasthenia gravis (EAMG) rabbits by using T?125～147 Methods Peptides T?125～147 were synthesized referring to the residue sequence of acetylcholine receptor of Torpedo California and the rabbits were inoculated with the peptides Clinical manifestation was graded in 4 levels Electrophysiological function was assessed by repetitive nerve stimulation (RNS) and single fibre electromyography (SFEMG) tests Anti peptides antibodies were evaluated by enzyme linked immunosorbent assay (ELISA) Student t test was used to analyze the difference between the EAMG and healthy rabbits Results Following the second inoculation,the rabbits appeared weakness Clinical symptoms were improved by neostigmine At 3,5 and 10 Hz,the decrement of compound muscle action potential (CMAP) and the mean jitter (MCD) of the immunized rabbits were higher than in the healthy ones The percentage of decrement of CMAP in order of the control group and the T?125～147 group were:3 Hz: 1 625?1 317,25 375?7 945; 5 Hz: 2 000?1 732,25 625?9 102; 10 Hz: 1 750?1 392,28 875?8 709.Following the above sequence,the MCD were:3 Hz: (9 875?1 126) ?s,(25 875?7 945) ?s; 5 Hz: (11 375?0 916) ?s,(27 500?3 381) ?s; 10 Hz: (12 375?1 061) ?s,(31 000?4 811) ?s Anti peptide antibodies in immunized groups were significantly higher than those in the control group Conclusion T?125～147 might be served as the immunogenic to induce EAMG in rabbits,accompanied by elevation of anti peptide antibodies and the blockage of neuromuscular transmission
RESUMO
Objective To study the prophylactic effects of nasal tolerance with a dual analogue (Lys262-Ala207) on experimental autoimmune myasthenia gravis (EAMG) and observe the underlying mechanisms, the clinical and immunological changes in Lewis rats treated with dual analogue nasally. Methods The effects of the predetermined dosage of a dual analogue Lys262-Ala207 were compared at different time points, and the dual analogues or control peptides were given nasally before (Group A or CA) or on the day (Group B or CB) of immunization with acetylcholine receptor (AChR) in complete Freund's adjuvant for 10 consecutive days. The clinical scores were evaluated for 50 days after immunization. The levels of anti-AChR IgG in serum were tested by RIA. Proliferative responses of lymphocytes to no antigen, Lys262-Ala207, AChR, AChR-?100-116, MBP peptide, or Con A were tested. The numbers of mononuclear cells expressing CD4 and/or CD25 from lymph nodes were enumerated using flow cytometry. Results As compared with the corresponding control groups, Lewis rats in group A or B developed EAMG with reduced severity and loss of AChR within the neuromuscular junction. The levels of anti-AChR IgG (21.96?3.37 and 29.41?4.59) were also decreased. Proliferative responses were suppressed in response to antigen-specific stimulations in rats receiving dual analogue, whereas the numbers of CD4+CD25+ T cells were higher in group A (11.34%?1.62%) and B (8.68%?1.83%) than in their corresponding control groups. Conclusions Nasal administration with a dual analogue Lys262-Ala207, at two different time points before and on the day of immunization ameliorated muscular weakness in EAMG rats associated with decreased levels of anti-AChR IgG in serum, suppressed antigen-specific T cell proliferation and increased numbers of CD4+CD25+ T cells from lymph nodes as compared to rats receiving control peptides. The results of our study suggest that the mucosal tolerance with dual analogue should be served as an alternative maneuver in human MG.
RESUMO
Objective To study the effect of nasal tolerance with a dual analogue (Lys262 Ala207) on experimental autoimmune myasthenia gravis (EAMG) and the underlying mechanisms, the clinical and immunological changes were observed in Lewis rats treated with dual analogue Methods Different doses of dual analogues were given to Lewis rats immunized with acetylcholine receptor (AChR) in complete Freud's adjuvant (CFA) and the medium dosage was chosen in the following studies As comparing the effects of treatment of the predetermined dosage of dual analogue at different time points, the body weight and clinical symptoms of Lewis rats immunized with AChR in CFA were evaluated The levels of anti AChR IgG in serum were tested by ELISA Proliferative responses of lymphocytes to no antigen, AChR, dual analogue, control MBP peptide, MBP, or Con A were tested Results Lewis rats receiving dual analogue nasally for 10 consecutive days at the time of immunization (Group A) or the first day after complete remission from the acute phase of the disease (Group B) sparsely developed EAMG with reduced severity than the corresponding control groups The subsiding disease was associated with decreased amount of anti AChR IgG in serum expressed as optic density ( A ) at 405 nm On day 35 post immunization, the A values in group B vs control group were 0 95?0 26 and 1 19?0 12, respectively Proliferative responses expressed as stimulation index (SI) were suppressed in response to antigen specific stimulations in rats receiving dual analogue as compared with rats receiving control peptide For instance, the values of SI in response to AChR in group A and control group were 1 71?0 78 and 3 24?1 31, respectively Those values were suppressed to a less extent in group B vs control group (1 97?0 56 vs 3 19? 1 50) Conclusions Nasal administration of a dual analogue Lys262 Ala207, at two different time points post immunization should ameliorate muscular weakness in EAMG rats involved in decreased levels of anti AChR antibodies and antigen specific T cell proliferation