调节性T细胞和辅助性T细胞在重症症肌无力患者中的变化及免疫抑制剂对它们的影响
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摘要
重症肌无力（MG）是一典型的器官特异性自身免疫性疾病，是自身免疫耐受失败的结果。CD4~+CD25~+调节性T细胞（Tregs）是调控和维持机体自身免疫耐受的主要免疫细胞，其中在数量和功能上起扮演主要角色的是胸腺来源的自然Tregs（nTregs）。Tregs与效应T细胞（Teffs）在机体内处于相互依存的关系，且维持着一定的平衡状态，自身免疫性疾病的发生是与这种平衡被先天或后天的因素所破坏有关。近年来，关于Tregs在MG发病中的作用备受国内外学者关注，但研究得出的结论并非完全一致，可能涉及的因素包括选取不同的Tregs相关分子、年龄偏差等。因此，我们认为在全面系统评估MG患者体内Tregs和Teffs的变化时需考虑到这些混杂因素的干扰。此外，我们也想了解那些常规治疗MG患者的免疫抑制剂如糖皮质激素（GC）和丙种球蛋白（Ig）是如何发挥免疫调节作用的。
本研究分为三个部分：一、检测MG患者外周血中代表Tregs的相关分子，探索它们与患者病情严重程度的相关性以及GC等免疫抑制剂对它们影响。二、了解Tregs、naǐve T细胞和Th17细胞在MG患者胸腺组织中的表达情况。三、观察Th_1/Th_2/Th_(17)等Th亚群在MG患者外周血中的变化以及GC等免疫抑制剂对它们的影响。
一、MG患者外周血中Tregs相关分子的变化以及强的松等免疫抑制剂对它们的影响
CD4~+FOXP3~+T细胞、 CD4~+CD25highT细胞、 CD4~+CD25~+CD127-T细胞和CD4~+CD25~+FOXP3~+T细胞均被不同的研究者看做是人源Tregs的代表，在本次研究中我们一一考察它们在MG患者外周血中的变化，并探索它们与患者病情严重程度的相关性以及GC等免疫抑制剂对它们影响。
本实验我们采用流式细胞仪的六色荧光分析技术。为了解Tregs对患者病情的影响，将65例MG患者分为活动期患者57例和缓解期患者8例。为探讨GC、Ig等免疫抑制剂对Tregs和Th亚群的影响，我们又将活动期的57例MG患者分为未治疗组17例、GC治疗组29例和IVIg治疗组11例。健康对照组90例。患者组平均年龄是39.0±13.2岁，健康对照组平均年龄34.5±9.0岁。患者组性别比是11:8（男：女），健康对照组是3:2（男：女）。
结果发现外周血中CD4~+CD25~+CD127-T细胞的比例和绝对数无论在活动状态的患者还是处于缓解期的患者与健康对照者相比均无明显差异。我们对CD4~+CD25highT细胞的检测结果显示活动期患者CD4~+CD25highT细胞比例显著低于健康对照者，与缓解期患者相比较时却无差异。
CD4~+FOXP3~+T细胞在未治疗组和GC治疗组与健康对照组比较时均未见差异，但IVIg治疗组的百分比显著升高。CD4~+CD25~+FOXP3~+T细胞无论在百分比和绝对数上活动期患者显著地低于缓解期患者和健康对照者，后两者之间则无差异。在应用GC和Ig等免疫抑制剂后多数患者肌无力症状得到了不同程度的改善，伴随的是患者外周血中CD4~+CD25~+FOXP3~+T细胞比例的升高，它们之间呈显著的负相关。
二、Tregs、naǐve T细胞和Th17细胞在MG患者胸腺组织中的变化
为探讨胸腺在MG发病中的价值，我们进行了胸腺标本的免疫组化分析。为防止年龄等混杂因素对结果的影响，我们还同时设立了多组对照，包括健康对照组和MG（-）的胸腺瘤组。健康对照来自因胸部外伤或心血管畸形接受心脏手术的患者，他们既往均未罹患自身免疫性疾病，也未服用相关药物。MG患者组39例，平均年龄40.92±14.4岁，男女性别比是10:29；健康对照组14例，平均年龄14.43±6.28岁，男女性别比是3:11；MG(-)的胸腺瘤患者组29例，平均年龄45.24±12.70岁，男女性别比是8:21。CD25在胸腺中的表达强烈提示是Tregs，而FOXP3的表达又是决定胸腺产生的Tregs是否成熟及能否发挥抑制功能的关键，因此，我们同时检测了CD25和FOXP3在胸腺中的表达情况作为胸腺Tregs的代表
结果显示MG患者胸腺中CD25阳性细胞和FOXP3阳性细胞的数量和表达强度显著低于健康对照组，但两者存在着年龄的偏差，因此差异可能是由年龄造成的。因此，我们又将年龄与MG患者匹配的MG（-）的胸腺瘤患者与之相比较，结果发现无论在数量还是强度上两者均无明显差异。
与Tregs相对应的Teffs前体naiveT细胞是在胸腺中发育并输出的，我们的免疫组化结果同时显示MG（+）胸腺瘤患者胸腺中CD4~+细胞和CD8+细胞较MG（-）胸腺瘤患者和健康对照者显著增多。
我们用流式的方法检测了几例新鲜胸腺标本中Th17细胞的表达，结果发现无论是MG（+）的胸腺瘤患者还是MG（-）胸腺瘤患者胸腺中均未发现Th17细胞，同样在健康对照者胸腺中也未发现。
三、Th_1/Th_2/Th_(17)等Th细胞亚群在MG患者外周血中的变化及免疫抑制剂对它们的影响
Th细胞的不同亚群在MG发病中扮演什么样的角色是我们此次研究的目标之一。我们将新鲜提取的外周血单个核细胞（PBMNC）在加入适当浓度的刺激剂后放置CO2孵育箱培养4~6个小时。然后进行流式抗体标记。考虑到CD4~+T细胞在增殖过程中可能会出现包膜内陷CD4分子难以检测到，我们采用CD3和CD8抗体反相标记CD4~+T细胞。Th亚群的代表性细胞因子均为胞内表达，故在标记表面分子后须固定、破膜。Th亚群的流式检测结果显示患者外周血中的Th_1/Th_2/Th_(17)无论在比例和绝对数上均与健康对照者无明显差异，GC和Ig等免疫抑制剂对它们也无明显的影响。与此相对应的是，MG患者血浆中分别代表不同Th亚群的炎症细胞因子与健康对照者相比较也无明显不同。
总之：根据我们的研究结果可以得出这样的几个结论：
1）在免疫稳态中具有最强的免疫调节功能也是nTregs中一个最稳定的亚群CD4~+CD25~+FOXP3~+T细胞在MG患者外周循环中显著减少，可能的原因并不是胸腺发育的缺陷。另外，我们在研究中发现患者外周循环中CD4~+FOXP3~+T细胞百分比和绝对数与健康对照者比较均没有明显减少。
2）患者外周血中的三个Th亚群无论是绝对数还是百分比与健康对照者相比较时均未发现明显变化。
3）大部分胸腺瘤或胸腺增生的患者接受治疗性胸腺切除（Tx）手术后肌无力症状会改善，可能的原因在于患者胸腺增生活跃，生成和输出的naǐve T细胞增多。
4） GC和Ig仍然是目前治疗MG的最常用药物，我们的研究结果证实它们的治疗作用是与其提升Tregs比例有关。
Myasthenia gravis (MG) is a typical organ specific autoimmune disease, and it isthe result of immune tolerance failure. CD4~+CD25~+regulatory T cells (Tregs) areinstrumental in the maintenance of immunologic self-tolerance, and thymus derivednatural Tregs (nTregs) playe the key roles in the number and function. Tregs andeffector T cells (Teffs) are in the interdependent relationship in the body and maintain acertain equilibrium state. Occurrence of autoimmune disease is associated with thisdamaged balance because of some congenital or acquired factors. The view has beenwidely accepted.
Recently, the effect of Tregs in the pathogenesis of MG has been drawn attentionby domestic and international scholars, but the conclusions were not consistent.Relevant factors may include selecting Tregs associated molecule, different age bias.Therefore, we think it necessary to consider the interference of these confoundingfactors when we would like to comprehensively assess the changes of Tregs and Teffs inMG patients. Moreover, we want to understand how to the conventionalimmunosuppressants (IS) inhibitors such as GC and Ig, which are prescribed to MGpatients, work.
This study was divided into three parts:1) detected Treg-association molecules inperipheral blood of MG patients and compared to those of the heathy donors, andobserved the effect of immunomodulators such as prednisone and gamma globulin onthem.2) investigated the expressions of Tregs、naǐve T cells and Th17cells in thethymus of the MG patients, and compare with those of the MG (-) thymoma patientsand the healthy subjects.3) explored the changes of different Th cells in peripheral blood of the MG patients, and investigated the effects of immunomodulators such asprednisone and gamma globulin on them.
Changes of Treg-association Molecules in the Peripheral Blood of the MG Patientsand the Effects of Immunosuppresants such as Prednisone and Immunoglobulin onthem
CD4~+FOXP3~+T cells、 CD4~+CD25highT cells、 CD4~+CD25~+CD127-T cells andCD4~+CD25~+FOXP3~+T cells respectively were regarded as human Tregs, we detected thechanges of them in the peripheral blood of the MG patients, and explored theircorrelation with the severity of clinical sympotom and the effects of immunosuppresantssuch as pred on them.
We used the six colors fluorescent analysis technique of FACSCanto Ⅱflowcytometry by newly developed by BD, and take use of the characteristic of its simpleand quick operation and highly accurate test results.
They were divided into two groups in order to understand the impact of Tregs onthe patient condition:57MG patients in active stage and8MG patients in remission.57MG patients in active stage were also divided into three groups according to thetreatment strategy in order to investigate the effect of GC, Ig and other immuneinhibitor on Tregs and Th subsets:1)17MG patients (untreated group): who did notundergo any IS therapy before enrollment;2)29MG patients (GCs treated group): whohad previously received oral prednisone (pred) alone or pred and azathioprine treatment;3)11MG patients (IVIG group): who were suffering from progressive generalweakness and diagnosed clinically as myasthenia crisis. The healthy control group is90cases. The average age of the MG patients was39.4±13.5years old, and the averageage of the healthy donors was34.5±9.0years old. The sex ratio was11:8(male: female)in the patients, and it was3:2in the healthy donors.
Our results showed that no significant difference was observed in the absolutenumber and frequency of CD4~+CD25~+CD127-T cells among different groups.
The frequency of CD4~+CD25highT cells in the patients in active stage obviouslydecreased when compared to that of the healthy donors, but no measurable differencewhen compared to that of the patents in remission.
CD4~+FOXP3~+T cells are often taken as Tregs. Our results showed that nosignificant difference was observed in the absolute number and frequency ofCD4~+FOXP3~+T cells between different groups.
Our results indicated that the numbers and frequencies of CD4~+CD25~+FOXP3~+Tcells significantly decreased in the patients in active stage when compared to those ofthe patients in remission and the healthy donors, and no diffenrence was seen beweenthe latters. The dynamic analysis of Treg cell level in MG patients who accepted IStherapy suggested that IS such as GC and Ig had the capacity of increasing the size ofthe peripheral Tregs population. The analysis of relationship between the change rate ofQMGs and Treg cell levels suggested significant that there was significant negativerelationship between them.
The changes of Tregs and naǐve T cells in the Thymus Tissue of the MG Patients
Considering some confounding factors such as age, so we used multiple groupanalysis to investigate the expression of Tregs in the thymus of the MG patients.Control thymus tissues were obtained from the young adults suffering from bluntthoracic trauma and accepting thoracic surgery and the adolescents accepting correctivetherapy because of suffering from congenital cardiovascular malformation. They allsigned informed consent. They had not suffered from autoimmune disease and took anydrugs. The MG patient group included39patients, their mean age was40.92±14.4years, and the male to female ratio was10:29. The healthy donor group included14,their mean age was14.43±6.28years, and the male to female ratio was3:11. Thethymoma patients without myasthenia gravis were29, and their mean age was45.24±12.70years, and the male to female ratio was8:21. Different from the peripheralcirculation, CD25expression in thymus strongly is suggestive of Tregs, and FOXP3 expression in thymus also decided whether thymus produced Tregs are mature and playa key role in the inhibition function. Therefore, we examined the expression of CD25and FOXP3in the thymus.
Our results showed that the numbers and expression intensities of CD25~+thymocytes and FOXP3~+thymocytes, significantly decreased when compared to those ofthe healthy donors. However, there was age bias between them, and the discrepancymay be ascribed to age bias. Therefore, we compared the age-mathched MG (-) patientswith thymoma to the MG patients, and found no measurably difference in the numbersand expression intensities of CD25~+thymocytes and FOXP3~+thymocytes between them.
Teffs precursor naǐve T cells, corresponding to Tregs, develop and output in thethymus.Our immunohistochemical results showed that the numbers of CD4~+Tlymphocytes and CD8+T lymphocytes in the thymus of the MG patients with thymomasignificantly were higher than those of the MG (-) patients with thymoma and thehealthy donors. The similar result was seen when compared to the non-neoplastic MGpatients. Moreover, we found the phenomenon that the frequencies of CD45RA+CD4~+Tcells in peripheral blood of some MG patients were higher than that of healthy donors.
We detected the express of Th17cell in the fresh thymus specimens of a few casesby the method of flow cytometry. Th17cells were not found in the thymi of the MGpatients with thymoma and the MG (-) patients, and the similar result was found in thehealthy donors.
Changes of Th_1/Th_2/Th_(17)in the Peripheral Blood of MG Patients and the Effects ofImmunosuppresants on them
Peripheral blood mononuclear cells (PBMNC) freshly extracted were added intoappropriate stimulating agents，placed in a CO2incubator and incubated for4~6hours.Then they were labeled by the flow type antibodies. Considering that CD4moleculemay be coated in CD4~+T cells and difficult to detect, we labeled CD4~+T cells by usingCD3and CD8antibodies. The cytokines representative of Th cell subsets are intracellularly expressed and fixed. Their cell membranes were ruptured. The resultsdetected by flow cytometry showed that no significant difference was seen in thefrequencies and absolute numbers of Th cell substs among the patients and the healthyhealthy donors. No obvious effect of IS such as GC and Ig on Th cell subsets wasobserved. Similarly, no significant difference was seen in the levels of the plasmainflammatory cytokines representive of different Th subsets among the MG patients andthe healthy donors.
In conclusion: according to our results, several conclusions can be drawn:
1) Our results indicated that significant reduction in the numbers and frequencies ofCD4~+CD25~+FOXP3~+T cells, the strongest regulation function in immune homeostasisand the most stable subset of nTregs, could not be attributed to Tregs deficiency inthymic development. Moreover, no significant difference in the number and frequenciesof CD4~+FOXP3~+T cells was seen between the patients and the healthy donors.
2) No measureable difference was observed in the absolute numbers and frequencies ofTh_1/Th_2/Th_(17)between the patients and the healthy donors.
3) Myasthenic symptoms can improve in the most patients with thymoma or thymichyperplasia, who accepted Tx therapy. The possible reason lies in the active patientswith thymic hyperplasia, generation and output of naǐve T cells.
4) IS such as GC and Ig has been the most commonly prescribed drug for MG. Ourresults confirmed that their therapeutic effect is related to increasing the number and thefequency of CD4~+CD25~+FOXP3~+T cells in peripheral blood of the MG patients.
本研究分为三个部分：一、检测MG患者外周血中代表Tregs的相关分子，探索它们与患者病情严重程度的相关性以及GC等免疫抑制剂对它们影响。二、了解Tregs、naǐve T细胞和Th17细胞在MG患者胸腺组织中的表达情况。三、观察Th_1/Th_2/Th_(17)等Th亚群在MG患者外周血中的变化以及GC等免疫抑制剂对它们的影响。
一、MG患者外周血中Tregs相关分子的变化以及强的松等免疫抑制剂对它们的影响
CD4~+FOXP3~+T细胞、 CD4~+CD25highT细胞、 CD4~+CD25~+CD127-T细胞和CD4~+CD25~+FOXP3~+T细胞均被不同的研究者看做是人源Tregs的代表，在本次研究中我们一一考察它们在MG患者外周血中的变化，并探索它们与患者病情严重程度的相关性以及GC等免疫抑制剂对它们影响。
本实验我们采用流式细胞仪的六色荧光分析技术。为了解Tregs对患者病情的影响，将65例MG患者分为活动期患者57例和缓解期患者8例。为探讨GC、Ig等免疫抑制剂对Tregs和Th亚群的影响，我们又将活动期的57例MG患者分为未治疗组17例、GC治疗组29例和IVIg治疗组11例。健康对照组90例。患者组平均年龄是39.0±13.2岁，健康对照组平均年龄34.5±9.0岁。患者组性别比是11:8（男：女），健康对照组是3:2（男：女）。
结果发现外周血中CD4~+CD25~+CD127-T细胞的比例和绝对数无论在活动状态的患者还是处于缓解期的患者与健康对照者相比均无明显差异。我们对CD4~+CD25highT细胞的检测结果显示活动期患者CD4~+CD25highT细胞比例显著低于健康对照者，与缓解期患者相比较时却无差异。
CD4~+FOXP3~+T细胞在未治疗组和GC治疗组与健康对照组比较时均未见差异，但IVIg治疗组的百分比显著升高。CD4~+CD25~+FOXP3~+T细胞无论在百分比和绝对数上活动期患者显著地低于缓解期患者和健康对照者，后两者之间则无差异。在应用GC和Ig等免疫抑制剂后多数患者肌无力症状得到了不同程度的改善，伴随的是患者外周血中CD4~+CD25~+FOXP3~+T细胞比例的升高，它们之间呈显著的负相关。
二、Tregs、naǐve T细胞和Th17细胞在MG患者胸腺组织中的变化
为探讨胸腺在MG发病中的价值，我们进行了胸腺标本的免疫组化分析。为防止年龄等混杂因素对结果的影响，我们还同时设立了多组对照，包括健康对照组和MG（-）的胸腺瘤组。健康对照来自因胸部外伤或心血管畸形接受心脏手术的患者，他们既往均未罹患自身免疫性疾病，也未服用相关药物。MG患者组39例，平均年龄40.92±14.4岁，男女性别比是10:29；健康对照组14例，平均年龄14.43±6.28岁，男女性别比是3:11；MG(-)的胸腺瘤患者组29例，平均年龄45.24±12.70岁，男女性别比是8:21。CD25在胸腺中的表达强烈提示是Tregs，而FOXP3的表达又是决定胸腺产生的Tregs是否成熟及能否发挥抑制功能的关键，因此，我们同时检测了CD25和FOXP3在胸腺中的表达情况作为胸腺Tregs的代表
结果显示MG患者胸腺中CD25阳性细胞和FOXP3阳性细胞的数量和表达强度显著低于健康对照组，但两者存在着年龄的偏差，因此差异可能是由年龄造成的。因此，我们又将年龄与MG患者匹配的MG（-）的胸腺瘤患者与之相比较，结果发现无论在数量还是强度上两者均无明显差异。
与Tregs相对应的Teffs前体naiveT细胞是在胸腺中发育并输出的，我们的免疫组化结果同时显示MG（+）胸腺瘤患者胸腺中CD4~+细胞和CD8+细胞较MG（-）胸腺瘤患者和健康对照者显著增多。
我们用流式的方法检测了几例新鲜胸腺标本中Th17细胞的表达，结果发现无论是MG（+）的胸腺瘤患者还是MG（-）胸腺瘤患者胸腺中均未发现Th17细胞，同样在健康对照者胸腺中也未发现。
三、Th_1/Th_2/Th_(17)等Th细胞亚群在MG患者外周血中的变化及免疫抑制剂对它们的影响
Th细胞的不同亚群在MG发病中扮演什么样的角色是我们此次研究的目标之一。我们将新鲜提取的外周血单个核细胞（PBMNC）在加入适当浓度的刺激剂后放置CO2孵育箱培养4~6个小时。然后进行流式抗体标记。考虑到CD4~+T细胞在增殖过程中可能会出现包膜内陷CD4分子难以检测到，我们采用CD3和CD8抗体反相标记CD4~+T细胞。Th亚群的代表性细胞因子均为胞内表达，故在标记表面分子后须固定、破膜。Th亚群的流式检测结果显示患者外周血中的Th_1/Th_2/Th_(17)无论在比例和绝对数上均与健康对照者无明显差异，GC和Ig等免疫抑制剂对它们也无明显的影响。与此相对应的是，MG患者血浆中分别代表不同Th亚群的炎症细胞因子与健康对照者相比较也无明显不同。
总之：根据我们的研究结果可以得出这样的几个结论：
1）在免疫稳态中具有最强的免疫调节功能也是nTregs中一个最稳定的亚群CD4~+CD25~+FOXP3~+T细胞在MG患者外周循环中显著减少，可能的原因并不是胸腺发育的缺陷。另外，我们在研究中发现患者外周循环中CD4~+FOXP3~+T细胞百分比和绝对数与健康对照者比较均没有明显减少。
2）患者外周血中的三个Th亚群无论是绝对数还是百分比与健康对照者相比较时均未发现明显变化。
3）大部分胸腺瘤或胸腺增生的患者接受治疗性胸腺切除（Tx）手术后肌无力症状会改善，可能的原因在于患者胸腺增生活跃，生成和输出的naǐve T细胞增多。
4） GC和Ig仍然是目前治疗MG的最常用药物，我们的研究结果证实它们的治疗作用是与其提升Tregs比例有关。
Myasthenia gravis (MG) is a typical organ specific autoimmune disease, and it isthe result of immune tolerance failure. CD4~+CD25~+regulatory T cells (Tregs) areinstrumental in the maintenance of immunologic self-tolerance, and thymus derivednatural Tregs (nTregs) playe the key roles in the number and function. Tregs andeffector T cells (Teffs) are in the interdependent relationship in the body and maintain acertain equilibrium state. Occurrence of autoimmune disease is associated with thisdamaged balance because of some congenital or acquired factors. The view has beenwidely accepted.
Recently, the effect of Tregs in the pathogenesis of MG has been drawn attentionby domestic and international scholars, but the conclusions were not consistent.Relevant factors may include selecting Tregs associated molecule, different age bias.Therefore, we think it necessary to consider the interference of these confoundingfactors when we would like to comprehensively assess the changes of Tregs and Teffs inMG patients. Moreover, we want to understand how to the conventionalimmunosuppressants (IS) inhibitors such as GC and Ig, which are prescribed to MGpatients, work.
This study was divided into three parts:1) detected Treg-association molecules inperipheral blood of MG patients and compared to those of the heathy donors, andobserved the effect of immunomodulators such as prednisone and gamma globulin onthem.2) investigated the expressions of Tregs、naǐve T cells and Th17cells in thethymus of the MG patients, and compare with those of the MG (-) thymoma patientsand the healthy subjects.3) explored the changes of different Th cells in peripheral blood of the MG patients, and investigated the effects of immunomodulators such asprednisone and gamma globulin on them.
Changes of Treg-association Molecules in the Peripheral Blood of the MG Patientsand the Effects of Immunosuppresants such as Prednisone and Immunoglobulin onthem
CD4~+FOXP3~+T cells、 CD4~+CD25highT cells、 CD4~+CD25~+CD127-T cells andCD4~+CD25~+FOXP3~+T cells respectively were regarded as human Tregs, we detected thechanges of them in the peripheral blood of the MG patients, and explored theircorrelation with the severity of clinical sympotom and the effects of immunosuppresantssuch as pred on them.
We used the six colors fluorescent analysis technique of FACSCanto Ⅱflowcytometry by newly developed by BD, and take use of the characteristic of its simpleand quick operation and highly accurate test results.
They were divided into two groups in order to understand the impact of Tregs onthe patient condition:57MG patients in active stage and8MG patients in remission.57MG patients in active stage were also divided into three groups according to thetreatment strategy in order to investigate the effect of GC, Ig and other immuneinhibitor on Tregs and Th subsets:1)17MG patients (untreated group): who did notundergo any IS therapy before enrollment;2)29MG patients (GCs treated group): whohad previously received oral prednisone (pred) alone or pred and azathioprine treatment;3)11MG patients (IVIG group): who were suffering from progressive generalweakness and diagnosed clinically as myasthenia crisis. The healthy control group is90cases. The average age of the MG patients was39.4±13.5years old, and the averageage of the healthy donors was34.5±9.0years old. The sex ratio was11:8(male: female)in the patients, and it was3:2in the healthy donors.
Our results showed that no significant difference was observed in the absolutenumber and frequency of CD4~+CD25~+CD127-T cells among different groups.
The frequency of CD4~+CD25highT cells in the patients in active stage obviouslydecreased when compared to that of the healthy donors, but no measurable differencewhen compared to that of the patents in remission.
CD4~+FOXP3~+T cells are often taken as Tregs. Our results showed that nosignificant difference was observed in the absolute number and frequency ofCD4~+FOXP3~+T cells between different groups.
Our results indicated that the numbers and frequencies of CD4~+CD25~+FOXP3~+Tcells significantly decreased in the patients in active stage when compared to those ofthe patients in remission and the healthy donors, and no diffenrence was seen beweenthe latters. The dynamic analysis of Treg cell level in MG patients who accepted IStherapy suggested that IS such as GC and Ig had the capacity of increasing the size ofthe peripheral Tregs population. The analysis of relationship between the change rate ofQMGs and Treg cell levels suggested significant that there was significant negativerelationship between them.
The changes of Tregs and naǐve T cells in the Thymus Tissue of the MG Patients
Considering some confounding factors such as age, so we used multiple groupanalysis to investigate the expression of Tregs in the thymus of the MG patients.Control thymus tissues were obtained from the young adults suffering from bluntthoracic trauma and accepting thoracic surgery and the adolescents accepting correctivetherapy because of suffering from congenital cardiovascular malformation. They allsigned informed consent. They had not suffered from autoimmune disease and took anydrugs. The MG patient group included39patients, their mean age was40.92±14.4years, and the male to female ratio was10:29. The healthy donor group included14,their mean age was14.43±6.28years, and the male to female ratio was3:11. Thethymoma patients without myasthenia gravis were29, and their mean age was45.24±12.70years, and the male to female ratio was8:21. Different from the peripheralcirculation, CD25expression in thymus strongly is suggestive of Tregs, and FOXP3 expression in thymus also decided whether thymus produced Tregs are mature and playa key role in the inhibition function. Therefore, we examined the expression of CD25and FOXP3in the thymus.
Our results showed that the numbers and expression intensities of CD25~+thymocytes and FOXP3~+thymocytes, significantly decreased when compared to those ofthe healthy donors. However, there was age bias between them, and the discrepancymay be ascribed to age bias. Therefore, we compared the age-mathched MG (-) patientswith thymoma to the MG patients, and found no measurably difference in the numbersand expression intensities of CD25~+thymocytes and FOXP3~+thymocytes between them.
Teffs precursor naǐve T cells, corresponding to Tregs, develop and output in thethymus.Our immunohistochemical results showed that the numbers of CD4~+Tlymphocytes and CD8+T lymphocytes in the thymus of the MG patients with thymomasignificantly were higher than those of the MG (-) patients with thymoma and thehealthy donors. The similar result was seen when compared to the non-neoplastic MGpatients. Moreover, we found the phenomenon that the frequencies of CD45RA+CD4~+Tcells in peripheral blood of some MG patients were higher than that of healthy donors.
We detected the express of Th17cell in the fresh thymus specimens of a few casesby the method of flow cytometry. Th17cells were not found in the thymi of the MGpatients with thymoma and the MG (-) patients, and the similar result was found in thehealthy donors.
Changes of Th_1/Th_2/Th_(17)in the Peripheral Blood of MG Patients and the Effects ofImmunosuppresants on them
Peripheral blood mononuclear cells (PBMNC) freshly extracted were added intoappropriate stimulating agents，placed in a CO2incubator and incubated for4~6hours.Then they were labeled by the flow type antibodies. Considering that CD4moleculemay be coated in CD4~+T cells and difficult to detect, we labeled CD4~+T cells by usingCD3and CD8antibodies. The cytokines representative of Th cell subsets are intracellularly expressed and fixed. Their cell membranes were ruptured. The resultsdetected by flow cytometry showed that no significant difference was seen in thefrequencies and absolute numbers of Th cell substs among the patients and the healthyhealthy donors. No obvious effect of IS such as GC and Ig on Th cell subsets wasobserved. Similarly, no significant difference was seen in the levels of the plasmainflammatory cytokines representive of different Th subsets among the MG patients andthe healthy donors.
In conclusion: according to our results, several conclusions can be drawn:
1) Our results indicated that significant reduction in the numbers and frequencies ofCD4~+CD25~+FOXP3~+T cells, the strongest regulation function in immune homeostasisand the most stable subset of nTregs, could not be attributed to Tregs deficiency inthymic development. Moreover, no significant difference in the number and frequenciesof CD4~+FOXP3~+T cells was seen between the patients and the healthy donors.
2) No measureable difference was observed in the absolute numbers and frequencies ofTh_1/Th_2/Th_(17)between the patients and the healthy donors.
3) Myasthenic symptoms can improve in the most patients with thymoma or thymichyperplasia, who accepted Tx therapy. The possible reason lies in the active patientswith thymic hyperplasia, generation and output of naǐve T cells.
4) IS such as GC and Ig has been the most commonly prescribed drug for MG. Ourresults confirmed that their therapeutic effect is related to increasing the number and thefequency of CD4~+CD25~+FOXP3~+T cells in peripheral blood of the MG patients.
引文
1.Yi HJ, Chae CS, So JS, Tzartos SJ, Souroujon MC, Fuchs S, Im SH. Suppression ofexperimental myasthenia gravis by a B-cell epitope-free recombinant acetylcholinereceptor. Mol Immunol.2008,46(1):192-201
2.Sommer N, Willcox N, Harcourt GC, et al. Myasthenic thymus and thymoma areselectively enriched in acetylcholine receptor-reactive T cells. Ann Neurol.1990,28(3):312-9.
3.Huter EN, Punkosdy GA, Glass DD, Cheng LI, Ward JM, Shevach EM.TGF-beta-induced Foxp3+regulatory T cells rescue scurfy mice. Eur J Immunol.2008,38(7):1814-21.
4. Adel-Patient K, Wavrin S, Bernard H, Meziti N, Ah-Leung S, Wal JM. Oral toleranceand Treg cells are induced in BALB/c mice after gavage with bovineβ-lactoglobulin.Allergy.2011,66(10):1312-21
5.Ono M, Yaguchi H, Ohkura N, Kitabayashi I, Nagamura Y, Nomura T, Miyachi Y,Tsukada T, Sakaguchi S. Foxp3controls regulatory T-cell function by interacting withAML1/Runx1. Nature.2007,446(7136):685-9.
6.Zhang M, Zhou J, Zhao T, Huang G, Tan Y, Tan S, Fu X, Niu W, Meng G, Chen X,Shang X, Liu D, Ni B, Wang L, Wu Y. Dissection of a circulating and intrahepaticCD4(+)Foxp3(+) T-cell subpopulation in chronic hepatitis B virus (HBV) infection: ahighly informative strategy for distinguishing chronic HBV infection states. J InfectDis.2012,205(7):1111-20.
7.Banham AH. Cell-surface IL-7receptor expression facilitates the purification ofFOXP3(+) regulatory T cells. Trends Immunol.2006,27(12):541-4
8.Yu N, Li X, Song W, Li D, Yu D, Zeng X, Li M, Leng X, Li X. CD4(+)CD25(+)CD127(low/-) T cells: a more specific Treg population in human peripheral blood.Inflammation.2012,35(6):1773-80.
9.Tsakiri N, Papadopoulos D, Denis MC, Mitsikostas DD, Kollias G. TNFR2onnon-haematopoietic cells is required for Foxp3+Treg-cell function and diseasesuppression in EAE. Eur J Immunol.2012,42(2):403-12
10.Chen X, Subleski JJ, Hamano R, Howard OM, Wiltrout RH, Oppenheim JJ.Co-expression of TNFR2and CD25identifies more of the functionalCD4+FOXP3+regulatory T cells in human peripheral blood. Eur J Immunol.2010,40(4):1099-106.
11.Heiber JF, Geiger TL. Context and location dependence of adaptive Foxp3(+)regulatory T cell formation during immunopathological conditions. Cell Immunol.2012,279(1):60-5.
12. Qiao M, Thornton AM, Shevach EM. CD4+CD25+[corrected] regulatory T cellsrender naive CD4+CD25-T cells anergic and suppressive. Immunology.2007，120(4):447-55.
13. Haribhai D, Williams JB, Jia S, Nickerson D, Schmitt EG, Edwards B, ZiegelbauerJ, Yassai M, Li SH, Relland LM, Wise PM, Chen A, Zheng YQ, Simpson PM,Gorski J, Salzman NH, Hessner MJ, Chatila TA, Williams CB. A requisite role forinduced regulatory T cells in tolerance based on expanding antigen receptordiversity. Immunity.2011,35(1):109-22.
14. Hippen KL, Merkel SC, Schirm DK, Nelson C, Tennis NC, Riley JL, June CH,Miller JS, Wagner JE, Blazar BR. Generation and large-scale expansion of humaninducible regulatory T cells that suppress graft-versus-host disease.Am J Transplant.2011,11(6):1148-57
15. Komatsu N, Mariotti-Ferrandiz ME, Wang Y, Malissen B, Waldmann H, Hori S.Heterogeneity of natural Foxp3+T cells: a committed regulatory T-cell lineage andan uncommitted minor population retaining plasticity. Proc Natl Acad Sci U S A.2009,106(6):1903-8
16.Yang XO, Nurieva R, Martinez GJ, Kang HS, Chung Y, Pappu BP, Shah B, ChangSH, Schluns KS, Watowich SS, Feng XH, Jetten AM, Dong C. Molecular antagonismand plasticity of regulatory and inflammatory T-cell programs. Immunity.2008,29:44–56.
17. Laurence A, Amarnath S, Mariotti J, Kim YC, Foley J, Eckhaus M, O'Shea JJ,Fowler DH. STAT3transcription factor promotes instability of nTreg cells andlimits generation of iTreg cells during acute murine graft-versus-hostdisease.Immunity.2012,37(2):209-22.
18.Ryba-Stanis awowska M, Skrzypkowska M, My liwska J, My liwiec M. The serumIL-6profile and Treg/Th17peripheral cell populations in patients with type1diabetes. Mediators Inflamm.2013,2013:205284
19.Mann MK, Maresz K, Shriver LP, Tan Y, Dittel BN. B cell regulation ofCD4+CD25+T regulatory cells and IL-10via B7is essential for recovery fromexperimental autoimmune encephalomyelitis. J Immunol.2007,178(6):3447-56.
20.Marra LE, Zhang ZX, Joe B, Campbell J, Levy GA, Penninger J, Zhang L. IL-10induces regulatory T cell apoptosis by up-regulation of the membrane form ofTNF-alpha. J Immunol.2004,172(2):1028-35.
21.Barron L, Dooms H, Hoyer KK, Kuswanto W, Hofmann J, O'Gorman WE, AbbasAK. Cutting edge: mechanisms of IL-2-dependent maintenance of functionalregulatory T cells. J Immunol.2010,185(11):6426-30.
22.Chen Q, Kim YC, Laurence A, Punkosdy GA, Shevach EM. IL-2controls thestability of Foxp3expression in TGF-beta-induced Foxp3+T cells in vivo. J Immunol.2011,186(11):6329-37.
23.Grinberg-Bleyer Y, Saadoun D, Baeyens A, Billiard F, Goldstein JD, Grégoire S,Martin GH, Elhage R, Derian N, Carpentier W, Marodon G, Klatzmann D, Piaggio E,Salomon BL. Pathogenic T cells have a paradoxical protective effect in murineautoimmune diabetes by boosting Tregs. J Clin Invest.2010,120(12):4558-68.
24.Nakahara M, Nagayama Y, Ichikawa T, Yu L, Eisenbarth GS, Abiru N. The effect ofregulatory T-cell depletion on the spectrum of organ-specific autoimmune diseases innonobese diabetic mice at different ages. Autoimmunity.2011,44(6):504-10.
25.Shiono H, Roxanis I, Zhang W, Sims GP, Meager A, Jacobson LW, Liu JL, MatthewsI, Wong YL, Bonifati M, Micklem K, Stott DI, Todd JA, Beeson D, Vincent A,Willcox N. Scenarios for autoimmunization of T and B cells in myasthenia gravis.Ann N YAcad Sci.2003,998:237–56.
26.Berrih S, Morel E, Gaud C, Raimond F, Le Brigand H, Bach JF. Anti-AChRantibodies, thymic histology, and T cell subsets in myasthenia gravis. Neurology.1984,34(1):66-71.
27.Kirchner T, Tzartos S, Hoppe F, Schalke B, Wekerle H, Müller-Hermelink HK.Pathogenesis of myasthenia gravis. Acetylcholine receptor-related antigenicdeterminants in tumor-free thymuses and thymic epithelial tumors. Am J Pathol.1988,130(2):268-80.
28.Melms A, Schalke BC, Kirchner T, Müller-Hermelink HK, Albert E, Wekerle H.Thymus in myasthenia gravis. Isolation of T-lymphocyte lines specific for thenicotinic acetylcholine receptor from thymuses of myasthenic patients. J Clin Invest.1988,81(3):902-8.
29.Engler JB, Undeutsch R, Kloke L, Rosenberger S, Backhaus M, Schneider U, EgererK, Dragun D, Hofmann J, Huscher D, Burmester GR, Humrich JY, Enghard P,Riemekasten G. Unmasking of autoreactive CD4T cells by depletion of CD25regulatory T cells in systemic lupus erythematosus. Ann Rheum Dis.2011,70(12):2176-83.
30.Zhao Xiaoyan, R Pirskanen, V Malmstrom, A K Lefvert. Expression of OX40(CD134) on CD4+T-cells from patients with myasthenia gravis. Clin Exp Immunol.2006,143(1):110-6.
31.Ruolan Liu, Qinghua Zhou, Antonio La Cava, Denise I. Campagnolo, Luc Van Kaer,Fu-Dong Shi. Expansion of regulatory T cells via IL-2/anti-IL-2mAb complexessuppresses experimental myasthenia. Eur J Immunol.2010,40(6):1577-89.
32.Mu L, Sun B, Kong Q, Wang J, Wang G, Zhang S, Wang D, Liu Y, Liu Y, An H, Li H.Disequilibrium of T helper type1,2and17cells and regulatory T cells during thedevelopment of experimental autoimmune myasthenia gravis. Immunology.2009,128(1Suppl):e826-36.
33.Kong QF, Sun B, Bai SS, Zhai DX, Wang GY, Liu YM, Zhang SJ, Li R, Zhao W,Sun YY, Li N, Wang Q, Peng HS, Jin LH, Li HL. Administration of bone marrowstromal cells ameliorates experimental autoimmune myasthenia gravis by altering thebalance of Th1/Th2/Th17/Treg cell subsets through the secretion of TGF-beta. JNeuroimmunol.2009,207(1-2):83-91.
34.Kong QF, Sun B, Wang GY, Zhai DX, Mu LL, Wang DD, Wang JH, Li R, Li HL.BM stromal cells ameliorate experimental autoimmune myasthenia gravis by alteringthe balance of Th cells through the secretion of IDO. Eur J Immunol.2009,39(3):800-9.
35.Li XL, Liu Y, Cao LL, Li H, Yue LT, Wang S, Zhang M, Li XH, Dou YC, Duan RS.Atorvastatin-modified dendritic cells in vitro ameliorate experimental autoimmunemyasthenia gravis by up-regulated Treg cells and shifted Th1/Th17to Th2cytokines.Mol Cell Neurosci.2013,56C:85-95.
36.Jaretzki A3rd, Barohn RJ, Ernstoff RM, Kaminski HJ, Keesey JC, Penn AS,Sanders DB. Myasthenia gravis: recommendations for clinical research standards.Task Force of the Medical Scientific Advisory Board of the Myasthenia GravisFoundation of America. Neurology.2000;55:16–23.
37.Bedlack RS, Simel DL, Bosworth H, Samsa G, Tucker-Lipscomb B, Sanders DB.Quantitative myasthenia gravis score: Assessment of responsiveness and longitudinalvalidity. Neurology.2005;64:1968–70.
38.Chen X, Subleski JJ, Kopf H, Howard OM, M nnel DN, Oppenheim JJ. Cuttingedge: expression of TNFR2defines a maximally suppressive subset of mouseCD4+CD25+FoxP3+T regulatory cells: applicability to tumor-infiltrating Tregulatory cells. J Immunol.2008,180(10):6467-71.
39.Otsubo K, Kanegane H, Kamachi Y, Kobayashi I, Tsuge I, Imaizumi M, Sasahara Y,Hayakawa A, Nozu K, Iijima K, Ito S, Horikawa R, Nagai Y, Takatsu K, Mori H,Ochs HD, Miyawaki T. Identification of FOXP3-negative regulatory T-like(CD4(+)CD25(+)CD127(low)) cells in patients with immune dysregulation,polyendocrinopathy, enteropathy, X-linked syndrome. Clin Immunol.2011,141(1):111-20.
40.Zhang B, Zhang X, Tang F, Zhu L, Liu Y. Reduction of forkhead box P3levels inCD4+CD25high T cells in patients with new-onset systemic lupus erythematosus.Clin Exp Immunol.2008,153(2):182-7.
41.Xuyu Zhou, Samantha Bailey-Bucktrout, Lukas T, Jeker and Jeffrey A Blue stone.Plasticity of CD4+FoxP3+T cells. Curr Opin Immunol.2009,21(3):281–285.
42. Sugimoto N, Oida T, Hirota K, Nakamura K, Nomura T, Uchiyama T, Sakaguchi S.Foxp3-dependent and-independent molecules specific for CD25+CD4+naturalregulatory T cells revealed by DNA microarray analysis. Int Immunol.2006,18(8):1197-209.
43. Getnet D, Grosso JF, Goldberg MV, Harris TJ, Yen HR, Bruno TC, Durham NM,Hipkiss EL, Pyle KJ, Wada S, Pan F, Pardoll DM, Drake CG. A role for thetranscription factor Helios in human CD4(+)CD25(+) regulatory T cells. MolImmunol.2010,47:1595–1600.
44. Thornton AM, Korty PE, Tran DQ, Wohlfert EA, Murray PE, Belkaid Y, ShevachEM. Expression of Helios, an Ikaros transcription factor family member,differentiates thymic-derived from peripherally induced Foxp3+T regulatory cells. JImmunol.2010,184:3433–3441.
45. Kim YC, Bhairavabhotla R, Yoon J, Golding A, Thornton AM, Tran DQ, ShevachEM. Oligodeoxynucleotides stabilize Helios-expressing Foxp3+human T regulatorycells during in vitro expansion. Blood.2012,119:2810–2818.
46. Akimova T, Beier UH, Wang L, Levine MH, Hancock WW. Helios expression is amarker of T cell activation and proliferation. PLoS One.2011,6:e24226.
47. Serre K, Benezech C, Desanti G, Bobat S, Toellner KM, Bird R, Chan S, Kastner P,Cunningham AF, Maclennan IC, Mohr E. Helios is associated with CD4T cellsdifferentiating to T helper2and follicular helper T cells in vivo independently ofFoxp3expression. PLoS One.2011,6:e20731.
48. Gottschalk RA, Corse E, Allison JP. Expression of Helios in peripherally inducedFoxp3+regulatory T cells. J Immunol.2012,188:976–980.
49. Zabransky DJ, Nirschl CJ, Durham NM, Park BV, Ceccato CM, Bruno TC, Tam AJ,Getnet D, Drake CG. Phenotypic and functional properties of Helios+regulatory Tcells. PLoS One.2012;7:e34547.
50.Chen Y, Adams E, Regateiro FS, Vaux DJ, Betz AG, Andersen KG, Waldmann H,Howie D. Activation rather than Foxp3expression determines that TGF-β-inducedregulatory T cells out-compete na ve T cells in dendritic cell clustering. Eur JImmunol.2012,42(6):1436-48
51.Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A.Circulatingand thymic CD4CD25T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005,116(1):134-41
52.Stary G, Klein I, Bauer W, Koszik F, Reininger B, Kohlhofer S, Gruber K, Skvara H,Jung T, Stingl G. Glucocorticosteroids modify Langerhans cells to produce TGF-βand expand regulatory T cells. J Immunol.2011,186(1):103-12.
53. Tu E, Bourges D, Gleeson PA, Ang DK, van Driel IR. Pathogenic T cells persistafter reversal of autoimmune disease by immunosuppression with regulatory T cells.Eur J Immunol.2013,43(5):1286-96.
54.Beres A, Komorowski R, Mihara M, Drobyski WR. Instability of Foxp3expressionlimits the ability of induced regulatory T cells to mitigate graft versus hostdisease.Clin Cancer Res.2011,17(12):3969-83
55.Hirabayashi Y, Takahashi Y, Xu Y, et al. Lack of CD4(+)CD25(+)FOXP3(+)regulatory T cells is associated with resistance to intravenous immunoglobulintherapy in patients with Kawasaki disease. Eur J Pediatr.2013Jan23.
56.Deng H, Yang X, Jin T, Wu J, Hu LS, Chang M, Sun XJ, Adem A, Winblad B, ZhuJ. The role of IL-12and TNF-alpha in AIDP and AMAN. Eur J Neurol.2008,15(10):1100-5.
57.Ephrem A, Chamat S, Miquel C, Fisson S, Mouthon L, Caligiuri G, Delignat S,Elluru S, Bayry J, Lacroix-Desmazes S, Cohen JL, Salomon BL, Kazatchkine MD,Kaveri SV, Misra N. Expansion of CD4+CD25+regulatory T cells by intravenousimmunoglobulin: a critical factor in controlling experimental autoimmuneencephalomyelitis. Blood.2008,111(2):715-22.
58.Housley WJ, Adams CO, Nichols FC, Puddington L, Lingenheld EG, Zhu L, RajanTV, Clark RB. Natural but not inducible regulatory T cells require TNF-alphasignaling for in vivo function. J Immunol.2011,186(12):6779-87.
59.Tulic MK, Andrews D, Crook ML, Charles A, Tourigny MR, Moqbel R, Prescott SL.Changes in thymic regulatory T-cell maturation from birth to puberty: differences inatopic children. J Allergy Clin Immunol.2012,129(1):199-206.e1-4.
60.Bacchetta R, Passerini L, Gambineri E, Dai M, Allan SE, Perroni L,Dagna-Bricarelli F, Sartirana C, Matthes-Martin S, Lawitschka A, Azzari C, ZieglerSF, Levings MK, Roncarolo MG. Defective regulatory and effector T cell functionsin patients with FOXP3mutations. J Clin Invest.2006,116(6):1713-22.
61.Stephens LA, Mason D. CD25is a marker for CD4+thymocytes that preventautoimmune diabetes in rats, but peripheral T cells with this function are found inboth CD25+and CD25-subpopulations. J Immunol.2000,165(6):3105-10.
62.Bachmann K, Burkhardt D, Schreiter I, Kaifi J, Schurr P, Busch C, Thayssen G,Izbicki JR, Strate T. Thymectomy is more effective than conservative treatment formyasthenia gravis regarding outcome and clinical improvement. Surgery.2009;145(4):392–8.
63.Leite MI, Str bel P, Jones M, Micklem K, Moritz R, Gold R, Niks EH, Berrih-AkninS, Scaravilli F, Canelhas A, Marx A, Newsom-Davis J, Willcox N, Vincent A. Fewerthymic changes in MuSK antibody-positive than in MuSK antibody-negative MG.Ann Neurol.2005,57(3):444-8.
64.Chen G, Marx A, Chen WH, Yong J, Puppe B, Stroebel P, Mueller-Hermelink HK.New WHO histologic classification predicts prognosis of thymic epithelial tumors: aclinicopathologic study of200thymoma cases from China.Cancer.2002,95(2):420-9.
65.Balandina A, Lécart S, Dartevelle P, Saoudi A, Berrih-Aknin S.Functional defect ofregulatory CD4(+)CD25+T cells in the thymus of patients with autoimmunemyasthenia gravis. Blood.2005,105(2):735-41.
66.C. Luther, S. Poeschel, M. Varga, A. Melms, E. Tolosa. Decreased frequency ofintrathymic regulatory T cells in patients with myasthenia-associated thymoma.Journal of Neuroimmunology164(2005)124–128.
67.Matsui N, Nakane S, Saito F, Ohigashi I, Nakagawa Y, Kurobe H, Takizawa H,Mitsui T, Kondo K, Kitagawa T, Takahama Y, Kaji R. Undiminished regulatory Tcells in the thymus of patients with myasthenia gravis. Neurology.2010,74(10):816-20.
68. Toker A, Engelbert D, Garg G, Polansky JK, Floess S, Miyao T, Baron U, Düber S,Geffers R, Giehr P, Schallenberg S, Kretschmer K, Olek S, Walter J, Weiss S, Hori S,Hamann A, Huehn J. Active demethylation of the Foxp3locus leads to thegeneration of stable regulatory T cells within the thymus. J Immunol.2013,190(7):3180-8.
69.Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A. Circulatingand thymic CD4CD25T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005.116(1):134-41.
70. Mabarrack NH, Turner NL, Mayrhofer G. Recent thymic origin, differentiation, andturnover of regulatory T cells. J Leukoc Biol.2008,84(5):1287-97.
71.Vukmanovic-Stejic M, Zhang Y, Cook JE, Fletcher JM, McQuaid A, Masters JE,Rustin MH, Taams LS, Beverley PC, Macallan DC, Akbar AN.Human CD4+CD25hiFoxp3+regulatory T cells are derived by rapid turnover of memory populations invivo. J Clin Invest.2006,116(9):2423-33.
72.Aslam R, Hu Y, Gebremeskel S, Segel GB, Speck ER, Guo L, Kim M, Ni H,Freedman J, Semple JW. Thymic retention of CD4+CD25+FOXP3+T regulatorycells is associated with their peripheral deficiency and thrombocytopenia in a murinemodel of immune thrombocytopenia. Blood.2012,120(10):2127-32.
73.Haas J, Fritzsching B, Trübswetter P, Korporal M, Milkova L, Fritz B, Vobis D,Krammer PH, Suri-Payer E, Wildemann B. Prevalence of newly generated naiveregulatory T cells (Treg) is critical for Treg suppressive function and determines Tregdysfunction in multiple sclerosis. J Immunol.2007,179(2):1322-30.
74.Str bel P, Helmreich M, Menioudakis G, Lewin SR, Rüdiger T, Bauer A, Hoffacker V,Gold R, Nix W, Schalke B, Elert O, Semik M, Müller-Hermelink HK, Marx A.Paraneoplastic myasthenia gravis correlates with generation of mature naive CD4(+)T cells in thymomas. Blood.2002,100(1):159-66.
75.Hoffacker V, Schultz A, Tiesinga JJ, Gold R, Schalke B, Nix W, Kiefer R,Müller-Hermelink HK, Marx A. Thymomas alter the T-cell subset composition in theblood: a potential mechanism for thymoma-associated autoimmune disease. Blood.2000,96(12):3872-9.
76.Fattorossi A, Battaglia A, Buzzonetti A, Minicuci G, Riso R, Peri L, Scambia G,Evoli A. Thymopoiesis, regulatory T cells, and TCRVbeta expression in thymomawith and without myasthenia gravis, and modulatory effects of steroid therapy. J ClinImmunol.2008,28(2):194-206.
77.Sempowski G, Thomasch J, Gooding M, Hale L, Edwards L, Ciafaloni E, Sanders D,Massey J, Douek D, Koup R, Haynes B. Effect of thymectomy on human peripheralblood T cell pools in myasthenia gravis. J Immunol.2001,166(4):2808-17.
78.Wang L, Xiao W, Zheng Y, Xiao R, Zhu G, Wang M, Li Y, Peng S, Tan X, He Y,Tan J. High dose lipopolysaccharide triggers polarization of mouse thymic Th17cellsin vitro in the presence of mature dendritic cells. Cell Immunol.2012，274(1-2):98-108.
79.Kiros TG, van Kessel J, Babiuk LA, Gerdts V. Induction, regulation andphysiological role of IL-17secreting helper T-cells isolated from PBMC, thymus, andlung lymphocytes of young pigs. Vet Immunol Immunopathol.2011,144(3-4):448-54.
80. Aricha R, Feferman T, Scott HS, Souroujon MC, Berrih-Aknin S, Fuchs S. Thesusceptibility of Aire(-/-) mice to experimental myasthenia gravis involvesalterations in regulatory T cells. J Autoimmun.2011,36(1):16-24.
81.Cosmi L, De Palma R, Santarlasci V, Maggi L, Capone M, Frosali F, Rodolico G,Querci V, Abbate G, Angeli R, Berrino L, Fambrini M, Caproni M, Tonelli F, LazzeriE, Parronchi P, Liotta F, Maggi E, Romagnani S, Annunziato F. Human interleukin17-producing cells originate from a CD161+CD4+T cell precursor. J Exp Med.2008,205(8):1903-16.
82.Sofi MH, Liu Z, Zhu L, Yu Q, Kaplan MH, Chang CH. Regulation of IL-17expression by the developmental pathway of CD4T cells in the thymus. MolImmunol.2010,47(6):1262-8.
83.Kim JS, Smith-Garvin JE, Koretzky GA, Jordan MS. The requirements for naturalTh17cell development are distinct from those of conventional Th17cells. J Exp Med.2011,208(11):2201-7.
84.Raju R, Navaneetham D, Protti MP, Horton RM, Hoppe BL, Howard J Jr, Conti-FineBM. TCR V beta usage by acetylcholine receptor-specific CD4+T cells inmyasthenia gravis. Autoimmun.1997,10(2):203-17.
85.Nakahara M, Nagayama Y, Ichikawa T, Yu L, Eisenbarth GS, Abiru N. The effect ofregulatory T-cell depletion on the spectrum of organ-specific autoimmune diseases innonobese diabetic mice at different ages. Autoimmunity.2011,44(6):504-10
86.Montane J, Bischoff L, Soukhatcheva G, Dai DL, Hardenberg G, Levings MK,Orban PC, Kieffer TJ, Tan R, Verchere CB. Prevention of murine autoimmunediabetes by CCL22-mediated Treg recruitment to the pancreatic islets. J Clin Invest.2011,121(8):3024-8.
87.Viret C, Leung-Theung-Long S, Serre L, Lamare C, Vignali DA, Malissen B, CarrierA, Guerder S. Thymus-specific serine protease controls autoreactive CD4T celldevelopment and autoimmune diabetes in mice. J Clin Invest.2011,121(5):1810-21.
88.Str bel P, Chuang WY, Chuvpilo S, Zettl A, Katzenberger T, Kalbacher H,Rieckmann P, Nix W, Schalke B, Gold R, Müller-Hermelink HK, Peterson P, MarxA. Common cellular and diverse genetic basis of thymoma-associated myastheniagravis: role of MHC class II and AIRE genes and genetic polymorphisms. Ann N YAcad Sci.2008,1132:143-56.
89.Close PM, Kirchner T, Uys CJ, Müller-Hermelink HK. Reproducibility of ahistogenetic classification of thymic epithelial tumours. Histopathology.1995,26(4):339-4
90.Marx A, Schultz A, Wilisch A, Helmreich M, Nenninger R, Müller-Hermelink HK.Paraneoplastic autoimmunity in thymus tumors. Dev Immunol.1998,6(1-2):129-40.
91.Poussin MA, Tüzün E, Goluszko E, Scott BG, Yang H, Franco JU, Christadoss P.B7-1costimulatory molecule is critical for the development of experimentalautoimmune myasthenia gravis. J Immunol.2003,170(8):4389-96.
92.Wang W, Milani M, Ostlie N, et al. C57BL/6mice genetically deficient inIL-12/IL-23and IFN-gamma are susceptible to experimental autoimmune myastheniagravis, suggesting a pathogenic role of non-Th1cells. J Immunol.2007,178(11):7072-80.
93.Ostlie N, Milani M, Wang W, et al. Absence of IL-4facilitates the development ofchronic autoimmune myasthenia gravis in C57BL/6mice. J Immunol.2003,170(1):604-12.
94.Yang H, Zhang Y, Wu M, et al. Suppression of ongoing experimental autoimmunemyasthenia gravis by transfer of RelB-silenced bone marrow dentritic cells isassociated with a change from a T helper Th17/Th1to a Th2and FoxP3+regulatoryT-cell profile. inflamm Res.2010,59(3):197-205.
95.Patakas A, Platt AM, Butcher JP, Maffia P, McInnes IB, Brewer JM, Garside P,Benson RA. Putative existence of reciprocal dialogue between Tfh and B cells and itsimpact on infectious and autoimmune disease. Immunol Lett.2011,138(1):38-46.
96.Maehara T, Moriyama M, Hayashida JN, Tanaka A, Shinozaki S, Kubo Y,Matsumura K, Nakamura S. Selective localization of T helper subsets in labialsalivary glands from primary Sj gren's syndrome patients. Clin Exp Immunol.2012,169(2):89-99.
97.Simpson N, Gatenby PA, Wilson A, Malik S, Fulcher DA, Tangye SG, Manku H,Vyse TJ, Roncador G, Huttley GA, Goodnow CC, Vinuesa CG, Cook MC.Expansion of circulating T cells resembling follicular helper T cells is a fixedphenotype that identifies a subset of severe systemic lupus erythematosus. ArthritisRheum.2010,62(1):234-44.
98.Ma J, Zhu C, Ma B, Tian J, Baidoo SE, Mao C, Wu W, Chen J, Tong J, Yang M, JiaoZ, Xu H, Lu L, Wang S. Increased frequency of circulating follicular helper T cells inpatients with rheumatoid arthritis. Clin Dev Immunol.2012,2012:827480.
99.Luo C, Li Y, Liu W, Feng H, Wang H, Huang X, Qiu L, Ouyang J. Expansion ofcirculating counterparts of follicular helper T cells in patients with myasthenia gravis.Neuroimmunol.2013,256(1-2):55-61.
100.Stummvoll GH, DiPaolo RJ, Huter EN, Davidson TS, Glass D, Ward JM, ShevachEM. Th1, Th2, and Th17effector T cell-induced autoimmune gastritis differs inpathological pattern and in susceptibility to suppression by regulatory T cells. JImmunol.2008,181(3):1908-16.
101.Kinnunen T, Chamberlain N, Morbach H, et al. Accumulation of peripheralautoreactive B cells in the absence of functional human regulatory T cells. Blood.2013,121(9):1595-603.
102.Leonardo SM, De Santis JL, Gehrand A, et al. Expansion of follicular helper T cellsin the absence of Treg cells: implications for loss of B-cell anergy. Eur J Immunol.2012,42(10):2597-607.
1. Wang W, Ostlie NS, Conti-Fine BM, Milani M. The susceptibility to experimentalmyasthenia gravis of STAT6-/-and STAT4-/-BALB/c mice suggests a pathogenicrole of Th1cells. J Immunol.2004,172(1):97-103.
2. Baggi F, Annoni A, Ubiali F, Milani M, Longhi R, Scaioli W, Cornelio F,Mantegazza R, Antozzi C. Breakdown of tolerance to a self-peptide of acetylcholinereceptor alpha-subunit induces experimental myasthenia gravis in rats. J Immunol.2004,172(4):2697-703.
3. Li N, Mu L, Wang J, Zhang J, Xie X, Kong Q, Tang W, Yao X, Liu Y, Wang L,Wang G, Wang D, Jin L, Sun B, Li H. Activation of the adenosine A2A receptorattenuates experimental autoimmune myasthenia gravis severity.Eur J Immunol.2012,42(5):1140-51
4. Mu L, Sun B, Kong Q, Wang J, Wang G, Zhang S, Wang D, Liu Y, Liu Y, An H, Li H.Disequilibrium of T helper type1,2and17cells and regulatory T cells during thedevelopment of experimental autoimmune myasthenia gravis. Immunology.2009,128(1Suppl):826-36.
5. Masuda M, Matsumoto M, Tanaka S, Nakajima K, Yamada N, Ido N, Ohtsuka T,Nishida M, Hirano T, Utsumi H. Clinical implication of peripheral CD4+CD25+regulatory T cells and Th17cells in myasthenia gravis patients. J Neuroimmunol.2010,225(1-2):123-31.
6. Li X, Xiao BG, Xi JY, Lu CZ, Lu JH. Decrease of CD4+CD25highFoxp3+regulatory Tcells and elevation of CD19+BAFF-R+B cells and soluble ICAM-1in myastheniagravis. Clinical Immunology.2008,126:180–188.
7. Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A. Circulatingand thymic CD4+CD25+T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005,116(1):134-41.
8. Xu WH, Zhang AM, Ren MS, Zhang XD, Wang F, Xu XC, Li Q, Wang J, Din BS,Wu YB, Chen GH. Changes of Treg-associated molecules on CD4+CD25+Treg cellsin myasthenia gravis and effects of immunosuppressants. J Clin Immunol.2012,32(5):975-83.
9. Le Panse R, Cizeron-Clairac G, Cuvelier M, Truffault F, Bismuth J, Nancy P, DeRosbo NK, Berrih-Aknin S. Regulatory and pathogenic mechanisms in humanautoimmune myasthenia gravis. Ann N Y Acad Sci.2008,1132:135-42
10. Luther C, Adamopoulou E, Stoeckle C, Brucklacher-Waldert V, Rosenkranz D,Stoltze L, Lauer S, Poeschel S, Melms A, Tolosa E. Prednisolone treatment inducestolerogenic dendritic cells and a regulatory milieu in myasthenia gravis patients.Immunol.2009,183(2):841-8.
11. Sheng JR, Li L, Ganesh BB, Vasu C, Prabhakar BS, Meriggioli MN. Suppression ofexperimental autoimmune myasthenia gravis by granulocyte-macrophagecolony-stimulating factor is associated with an expansion of FoxP3+regulatory Tcells. J Immunol.2006,177(8):5296-306.
12. Sheng JR, Li LC, Ganesh BB, Prabhakar BS, Meriggioli MN. Regulatory T cellsinduced by GM-CSF suppress ongoing experimental myasthenia gravis. ClinImmunol.2008,128(2):172-80.
13. Yang H, Zhang Y, Wu M, Li J, Zhou W, Li G, Li X, Xiao B, Christadoss P.Suppression of ongoing experimental autoimmune myasthenia gravis by transfer ofRelB-silenced bone marrow dentritic cells is associated with a change from a Thelper Th17/Th1to a Th2and FoxP3+regulatory T-cell profile. Inflamm Res.2010,59(3):197-205.
14. Chang HW, Chow YH, Chong P, Sia C. The Cross-Regulatory Relationship betweenHuman Dendritic and Regulatory T Cells and its Role in Type1Diabetes Mellitus.Rev Diabet Stud.2007,4(2):68–76.
15. Harry RA, Anderson AE, Isaacs JD, Hilkens CM. Generation and characterisation oftherapeutic tolerogenic dendritic cells for rheumatoid arthritis. Ann Rheum Dis.2010Jun15.
16. Pedersen AE, Gad M, Walter MR, Claesson MH. Induction of regulatory dendriticcells by dexamethasone and1alpha,25-Dihydroxyvitamin D (3). Immunol Lett.2004,91(1):63-9.
17. Xing N, L Maldonado ML, Bachman LA, McKean DJ, Kumar R, Griffin MD.Distinctive dendritic cell modulation by vitamin D(3) and glucocorticoid pathways.Biochem Biophys Res Commun.2002,297(3):645-52.
18. Smolders J, Thewissen M, Peelen E, Menheere P, Tervaert JW, Damoiseaux J,Hupperts R. Vitamin D status is positively correlated with regulatory T cell functionin patients with multiple sclerosis. PLoS One.2009,4(8): e6635.
19. van der Mei IA, Ponsonby AL, Dwyer T, Blizzard L, Taylor BV, Kilpatrick T,Butzkueven H, McMichael AJ.Vitamin D levels in people with multiple sclerosis andcommunity controls in Tasmania, Australia. J Neurol.2007,254(5):581-90.
20. Simpson S Jr, Taylor B, Blizzard L, Ponsonby AL, Pittas F, Tremlett H, Dwyer T,Gies P, van der Mei I. Higher25-hydroxyvitamin D is associated with lower relapserisk in multiple sclerosis. Ann Neurol.2010,68(2):193-203.
21. Prietl B, Pilz S, Wolf M, Tomaschitz A, Obermayer-Pietsch B, Graninger W, PieberTR. Vitamin D supplementation and regulatory T cells in apparently healthy subjects:vitamin D treatment for autoimmune diseases? Isr Med Assoc J.2010,12(3):136-9.
22. Burton JM, Kimball S, Vieth R, Bar-Or A, Dosch HM, Cheung R, Gagne D,D'Souza C, Ursell M, O'Connor P. A phase I/II dose-escalation trial of vitamin D3and calcium in multiple sclerosis. Neurology.2010,74(23):1852-9.
2.Sommer N, Willcox N, Harcourt GC, et al. Myasthenic thymus and thymoma areselectively enriched in acetylcholine receptor-reactive T cells. Ann Neurol.1990,28(3):312-9.
3.Huter EN, Punkosdy GA, Glass DD, Cheng LI, Ward JM, Shevach EM.TGF-beta-induced Foxp3+regulatory T cells rescue scurfy mice. Eur J Immunol.2008,38(7):1814-21.
4. Adel-Patient K, Wavrin S, Bernard H, Meziti N, Ah-Leung S, Wal JM. Oral toleranceand Treg cells are induced in BALB/c mice after gavage with bovineβ-lactoglobulin.Allergy.2011,66(10):1312-21
5.Ono M, Yaguchi H, Ohkura N, Kitabayashi I, Nagamura Y, Nomura T, Miyachi Y,Tsukada T, Sakaguchi S. Foxp3controls regulatory T-cell function by interacting withAML1/Runx1. Nature.2007,446(7136):685-9.
6.Zhang M, Zhou J, Zhao T, Huang G, Tan Y, Tan S, Fu X, Niu W, Meng G, Chen X,Shang X, Liu D, Ni B, Wang L, Wu Y. Dissection of a circulating and intrahepaticCD4(+)Foxp3(+) T-cell subpopulation in chronic hepatitis B virus (HBV) infection: ahighly informative strategy for distinguishing chronic HBV infection states. J InfectDis.2012,205(7):1111-20.
7.Banham AH. Cell-surface IL-7receptor expression facilitates the purification ofFOXP3(+) regulatory T cells. Trends Immunol.2006,27(12):541-4
8.Yu N, Li X, Song W, Li D, Yu D, Zeng X, Li M, Leng X, Li X. CD4(+)CD25(+)CD127(low/-) T cells: a more specific Treg population in human peripheral blood.Inflammation.2012,35(6):1773-80.
9.Tsakiri N, Papadopoulos D, Denis MC, Mitsikostas DD, Kollias G. TNFR2onnon-haematopoietic cells is required for Foxp3+Treg-cell function and diseasesuppression in EAE. Eur J Immunol.2012,42(2):403-12
10.Chen X, Subleski JJ, Hamano R, Howard OM, Wiltrout RH, Oppenheim JJ.Co-expression of TNFR2and CD25identifies more of the functionalCD4+FOXP3+regulatory T cells in human peripheral blood. Eur J Immunol.2010,40(4):1099-106.
11.Heiber JF, Geiger TL. Context and location dependence of adaptive Foxp3(+)regulatory T cell formation during immunopathological conditions. Cell Immunol.2012,279(1):60-5.
12. Qiao M, Thornton AM, Shevach EM. CD4+CD25+[corrected] regulatory T cellsrender naive CD4+CD25-T cells anergic and suppressive. Immunology.2007，120(4):447-55.
13. Haribhai D, Williams JB, Jia S, Nickerson D, Schmitt EG, Edwards B, ZiegelbauerJ, Yassai M, Li SH, Relland LM, Wise PM, Chen A, Zheng YQ, Simpson PM,Gorski J, Salzman NH, Hessner MJ, Chatila TA, Williams CB. A requisite role forinduced regulatory T cells in tolerance based on expanding antigen receptordiversity. Immunity.2011,35(1):109-22.
14. Hippen KL, Merkel SC, Schirm DK, Nelson C, Tennis NC, Riley JL, June CH,Miller JS, Wagner JE, Blazar BR. Generation and large-scale expansion of humaninducible regulatory T cells that suppress graft-versus-host disease.Am J Transplant.2011,11(6):1148-57
15. Komatsu N, Mariotti-Ferrandiz ME, Wang Y, Malissen B, Waldmann H, Hori S.Heterogeneity of natural Foxp3+T cells: a committed regulatory T-cell lineage andan uncommitted minor population retaining plasticity. Proc Natl Acad Sci U S A.2009,106(6):1903-8
16.Yang XO, Nurieva R, Martinez GJ, Kang HS, Chung Y, Pappu BP, Shah B, ChangSH, Schluns KS, Watowich SS, Feng XH, Jetten AM, Dong C. Molecular antagonismand plasticity of regulatory and inflammatory T-cell programs. Immunity.2008,29:44–56.
17. Laurence A, Amarnath S, Mariotti J, Kim YC, Foley J, Eckhaus M, O'Shea JJ,Fowler DH. STAT3transcription factor promotes instability of nTreg cells andlimits generation of iTreg cells during acute murine graft-versus-hostdisease.Immunity.2012,37(2):209-22.
18.Ryba-Stanis awowska M, Skrzypkowska M, My liwska J, My liwiec M. The serumIL-6profile and Treg/Th17peripheral cell populations in patients with type1diabetes. Mediators Inflamm.2013,2013:205284
19.Mann MK, Maresz K, Shriver LP, Tan Y, Dittel BN. B cell regulation ofCD4+CD25+T regulatory cells and IL-10via B7is essential for recovery fromexperimental autoimmune encephalomyelitis. J Immunol.2007,178(6):3447-56.
20.Marra LE, Zhang ZX, Joe B, Campbell J, Levy GA, Penninger J, Zhang L. IL-10induces regulatory T cell apoptosis by up-regulation of the membrane form ofTNF-alpha. J Immunol.2004,172(2):1028-35.
21.Barron L, Dooms H, Hoyer KK, Kuswanto W, Hofmann J, O'Gorman WE, AbbasAK. Cutting edge: mechanisms of IL-2-dependent maintenance of functionalregulatory T cells. J Immunol.2010,185(11):6426-30.
22.Chen Q, Kim YC, Laurence A, Punkosdy GA, Shevach EM. IL-2controls thestability of Foxp3expression in TGF-beta-induced Foxp3+T cells in vivo. J Immunol.2011,186(11):6329-37.
23.Grinberg-Bleyer Y, Saadoun D, Baeyens A, Billiard F, Goldstein JD, Grégoire S,Martin GH, Elhage R, Derian N, Carpentier W, Marodon G, Klatzmann D, Piaggio E,Salomon BL. Pathogenic T cells have a paradoxical protective effect in murineautoimmune diabetes by boosting Tregs. J Clin Invest.2010,120(12):4558-68.
24.Nakahara M, Nagayama Y, Ichikawa T, Yu L, Eisenbarth GS, Abiru N. The effect ofregulatory T-cell depletion on the spectrum of organ-specific autoimmune diseases innonobese diabetic mice at different ages. Autoimmunity.2011,44(6):504-10.
25.Shiono H, Roxanis I, Zhang W, Sims GP, Meager A, Jacobson LW, Liu JL, MatthewsI, Wong YL, Bonifati M, Micklem K, Stott DI, Todd JA, Beeson D, Vincent A,Willcox N. Scenarios for autoimmunization of T and B cells in myasthenia gravis.Ann N YAcad Sci.2003,998:237–56.
26.Berrih S, Morel E, Gaud C, Raimond F, Le Brigand H, Bach JF. Anti-AChRantibodies, thymic histology, and T cell subsets in myasthenia gravis. Neurology.1984,34(1):66-71.
27.Kirchner T, Tzartos S, Hoppe F, Schalke B, Wekerle H, Müller-Hermelink HK.Pathogenesis of myasthenia gravis. Acetylcholine receptor-related antigenicdeterminants in tumor-free thymuses and thymic epithelial tumors. Am J Pathol.1988,130(2):268-80.
28.Melms A, Schalke BC, Kirchner T, Müller-Hermelink HK, Albert E, Wekerle H.Thymus in myasthenia gravis. Isolation of T-lymphocyte lines specific for thenicotinic acetylcholine receptor from thymuses of myasthenic patients. J Clin Invest.1988,81(3):902-8.
29.Engler JB, Undeutsch R, Kloke L, Rosenberger S, Backhaus M, Schneider U, EgererK, Dragun D, Hofmann J, Huscher D, Burmester GR, Humrich JY, Enghard P,Riemekasten G. Unmasking of autoreactive CD4T cells by depletion of CD25regulatory T cells in systemic lupus erythematosus. Ann Rheum Dis.2011,70(12):2176-83.
30.Zhao Xiaoyan, R Pirskanen, V Malmstrom, A K Lefvert. Expression of OX40(CD134) on CD4+T-cells from patients with myasthenia gravis. Clin Exp Immunol.2006,143(1):110-6.
31.Ruolan Liu, Qinghua Zhou, Antonio La Cava, Denise I. Campagnolo, Luc Van Kaer,Fu-Dong Shi. Expansion of regulatory T cells via IL-2/anti-IL-2mAb complexessuppresses experimental myasthenia. Eur J Immunol.2010,40(6):1577-89.
32.Mu L, Sun B, Kong Q, Wang J, Wang G, Zhang S, Wang D, Liu Y, Liu Y, An H, Li H.Disequilibrium of T helper type1,2and17cells and regulatory T cells during thedevelopment of experimental autoimmune myasthenia gravis. Immunology.2009,128(1Suppl):e826-36.
33.Kong QF, Sun B, Bai SS, Zhai DX, Wang GY, Liu YM, Zhang SJ, Li R, Zhao W,Sun YY, Li N, Wang Q, Peng HS, Jin LH, Li HL. Administration of bone marrowstromal cells ameliorates experimental autoimmune myasthenia gravis by altering thebalance of Th1/Th2/Th17/Treg cell subsets through the secretion of TGF-beta. JNeuroimmunol.2009,207(1-2):83-91.
34.Kong QF, Sun B, Wang GY, Zhai DX, Mu LL, Wang DD, Wang JH, Li R, Li HL.BM stromal cells ameliorate experimental autoimmune myasthenia gravis by alteringthe balance of Th cells through the secretion of IDO. Eur J Immunol.2009,39(3):800-9.
35.Li XL, Liu Y, Cao LL, Li H, Yue LT, Wang S, Zhang M, Li XH, Dou YC, Duan RS.Atorvastatin-modified dendritic cells in vitro ameliorate experimental autoimmunemyasthenia gravis by up-regulated Treg cells and shifted Th1/Th17to Th2cytokines.Mol Cell Neurosci.2013,56C:85-95.
36.Jaretzki A3rd, Barohn RJ, Ernstoff RM, Kaminski HJ, Keesey JC, Penn AS,Sanders DB. Myasthenia gravis: recommendations for clinical research standards.Task Force of the Medical Scientific Advisory Board of the Myasthenia GravisFoundation of America. Neurology.2000;55:16–23.
37.Bedlack RS, Simel DL, Bosworth H, Samsa G, Tucker-Lipscomb B, Sanders DB.Quantitative myasthenia gravis score: Assessment of responsiveness and longitudinalvalidity. Neurology.2005;64:1968–70.
38.Chen X, Subleski JJ, Kopf H, Howard OM, M nnel DN, Oppenheim JJ. Cuttingedge: expression of TNFR2defines a maximally suppressive subset of mouseCD4+CD25+FoxP3+T regulatory cells: applicability to tumor-infiltrating Tregulatory cells. J Immunol.2008,180(10):6467-71.
39.Otsubo K, Kanegane H, Kamachi Y, Kobayashi I, Tsuge I, Imaizumi M, Sasahara Y,Hayakawa A, Nozu K, Iijima K, Ito S, Horikawa R, Nagai Y, Takatsu K, Mori H,Ochs HD, Miyawaki T. Identification of FOXP3-negative regulatory T-like(CD4(+)CD25(+)CD127(low)) cells in patients with immune dysregulation,polyendocrinopathy, enteropathy, X-linked syndrome. Clin Immunol.2011,141(1):111-20.
40.Zhang B, Zhang X, Tang F, Zhu L, Liu Y. Reduction of forkhead box P3levels inCD4+CD25high T cells in patients with new-onset systemic lupus erythematosus.Clin Exp Immunol.2008,153(2):182-7.
41.Xuyu Zhou, Samantha Bailey-Bucktrout, Lukas T, Jeker and Jeffrey A Blue stone.Plasticity of CD4+FoxP3+T cells. Curr Opin Immunol.2009,21(3):281–285.
42. Sugimoto N, Oida T, Hirota K, Nakamura K, Nomura T, Uchiyama T, Sakaguchi S.Foxp3-dependent and-independent molecules specific for CD25+CD4+naturalregulatory T cells revealed by DNA microarray analysis. Int Immunol.2006,18(8):1197-209.
43. Getnet D, Grosso JF, Goldberg MV, Harris TJ, Yen HR, Bruno TC, Durham NM,Hipkiss EL, Pyle KJ, Wada S, Pan F, Pardoll DM, Drake CG. A role for thetranscription factor Helios in human CD4(+)CD25(+) regulatory T cells. MolImmunol.2010,47:1595–1600.
44. Thornton AM, Korty PE, Tran DQ, Wohlfert EA, Murray PE, Belkaid Y, ShevachEM. Expression of Helios, an Ikaros transcription factor family member,differentiates thymic-derived from peripherally induced Foxp3+T regulatory cells. JImmunol.2010,184:3433–3441.
45. Kim YC, Bhairavabhotla R, Yoon J, Golding A, Thornton AM, Tran DQ, ShevachEM. Oligodeoxynucleotides stabilize Helios-expressing Foxp3+human T regulatorycells during in vitro expansion. Blood.2012,119:2810–2818.
46. Akimova T, Beier UH, Wang L, Levine MH, Hancock WW. Helios expression is amarker of T cell activation and proliferation. PLoS One.2011,6:e24226.
47. Serre K, Benezech C, Desanti G, Bobat S, Toellner KM, Bird R, Chan S, Kastner P,Cunningham AF, Maclennan IC, Mohr E. Helios is associated with CD4T cellsdifferentiating to T helper2and follicular helper T cells in vivo independently ofFoxp3expression. PLoS One.2011,6:e20731.
48. Gottschalk RA, Corse E, Allison JP. Expression of Helios in peripherally inducedFoxp3+regulatory T cells. J Immunol.2012,188:976–980.
49. Zabransky DJ, Nirschl CJ, Durham NM, Park BV, Ceccato CM, Bruno TC, Tam AJ,Getnet D, Drake CG. Phenotypic and functional properties of Helios+regulatory Tcells. PLoS One.2012;7:e34547.
50.Chen Y, Adams E, Regateiro FS, Vaux DJ, Betz AG, Andersen KG, Waldmann H,Howie D. Activation rather than Foxp3expression determines that TGF-β-inducedregulatory T cells out-compete na ve T cells in dendritic cell clustering. Eur JImmunol.2012,42(6):1436-48
51.Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A.Circulatingand thymic CD4CD25T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005,116(1):134-41
52.Stary G, Klein I, Bauer W, Koszik F, Reininger B, Kohlhofer S, Gruber K, Skvara H,Jung T, Stingl G. Glucocorticosteroids modify Langerhans cells to produce TGF-βand expand regulatory T cells. J Immunol.2011,186(1):103-12.
53. Tu E, Bourges D, Gleeson PA, Ang DK, van Driel IR. Pathogenic T cells persistafter reversal of autoimmune disease by immunosuppression with regulatory T cells.Eur J Immunol.2013,43(5):1286-96.
54.Beres A, Komorowski R, Mihara M, Drobyski WR. Instability of Foxp3expressionlimits the ability of induced regulatory T cells to mitigate graft versus hostdisease.Clin Cancer Res.2011,17(12):3969-83
55.Hirabayashi Y, Takahashi Y, Xu Y, et al. Lack of CD4(+)CD25(+)FOXP3(+)regulatory T cells is associated with resistance to intravenous immunoglobulintherapy in patients with Kawasaki disease. Eur J Pediatr.2013Jan23.
56.Deng H, Yang X, Jin T, Wu J, Hu LS, Chang M, Sun XJ, Adem A, Winblad B, ZhuJ. The role of IL-12and TNF-alpha in AIDP and AMAN. Eur J Neurol.2008,15(10):1100-5.
57.Ephrem A, Chamat S, Miquel C, Fisson S, Mouthon L, Caligiuri G, Delignat S,Elluru S, Bayry J, Lacroix-Desmazes S, Cohen JL, Salomon BL, Kazatchkine MD,Kaveri SV, Misra N. Expansion of CD4+CD25+regulatory T cells by intravenousimmunoglobulin: a critical factor in controlling experimental autoimmuneencephalomyelitis. Blood.2008,111(2):715-22.
58.Housley WJ, Adams CO, Nichols FC, Puddington L, Lingenheld EG, Zhu L, RajanTV, Clark RB. Natural but not inducible regulatory T cells require TNF-alphasignaling for in vivo function. J Immunol.2011,186(12):6779-87.
59.Tulic MK, Andrews D, Crook ML, Charles A, Tourigny MR, Moqbel R, Prescott SL.Changes in thymic regulatory T-cell maturation from birth to puberty: differences inatopic children. J Allergy Clin Immunol.2012,129(1):199-206.e1-4.
60.Bacchetta R, Passerini L, Gambineri E, Dai M, Allan SE, Perroni L,Dagna-Bricarelli F, Sartirana C, Matthes-Martin S, Lawitschka A, Azzari C, ZieglerSF, Levings MK, Roncarolo MG. Defective regulatory and effector T cell functionsin patients with FOXP3mutations. J Clin Invest.2006,116(6):1713-22.
61.Stephens LA, Mason D. CD25is a marker for CD4+thymocytes that preventautoimmune diabetes in rats, but peripheral T cells with this function are found inboth CD25+and CD25-subpopulations. J Immunol.2000,165(6):3105-10.
62.Bachmann K, Burkhardt D, Schreiter I, Kaifi J, Schurr P, Busch C, Thayssen G,Izbicki JR, Strate T. Thymectomy is more effective than conservative treatment formyasthenia gravis regarding outcome and clinical improvement. Surgery.2009;145(4):392–8.
63.Leite MI, Str bel P, Jones M, Micklem K, Moritz R, Gold R, Niks EH, Berrih-AkninS, Scaravilli F, Canelhas A, Marx A, Newsom-Davis J, Willcox N, Vincent A. Fewerthymic changes in MuSK antibody-positive than in MuSK antibody-negative MG.Ann Neurol.2005,57(3):444-8.
64.Chen G, Marx A, Chen WH, Yong J, Puppe B, Stroebel P, Mueller-Hermelink HK.New WHO histologic classification predicts prognosis of thymic epithelial tumors: aclinicopathologic study of200thymoma cases from China.Cancer.2002,95(2):420-9.
65.Balandina A, Lécart S, Dartevelle P, Saoudi A, Berrih-Aknin S.Functional defect ofregulatory CD4(+)CD25+T cells in the thymus of patients with autoimmunemyasthenia gravis. Blood.2005,105(2):735-41.
66.C. Luther, S. Poeschel, M. Varga, A. Melms, E. Tolosa. Decreased frequency ofintrathymic regulatory T cells in patients with myasthenia-associated thymoma.Journal of Neuroimmunology164(2005)124–128.
67.Matsui N, Nakane S, Saito F, Ohigashi I, Nakagawa Y, Kurobe H, Takizawa H,Mitsui T, Kondo K, Kitagawa T, Takahama Y, Kaji R. Undiminished regulatory Tcells in the thymus of patients with myasthenia gravis. Neurology.2010,74(10):816-20.
68. Toker A, Engelbert D, Garg G, Polansky JK, Floess S, Miyao T, Baron U, Düber S,Geffers R, Giehr P, Schallenberg S, Kretschmer K, Olek S, Walter J, Weiss S, Hori S,Hamann A, Huehn J. Active demethylation of the Foxp3locus leads to thegeneration of stable regulatory T cells within the thymus. J Immunol.2013,190(7):3180-8.
69.Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A. Circulatingand thymic CD4CD25T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005.116(1):134-41.
70. Mabarrack NH, Turner NL, Mayrhofer G. Recent thymic origin, differentiation, andturnover of regulatory T cells. J Leukoc Biol.2008,84(5):1287-97.
71.Vukmanovic-Stejic M, Zhang Y, Cook JE, Fletcher JM, McQuaid A, Masters JE,Rustin MH, Taams LS, Beverley PC, Macallan DC, Akbar AN.Human CD4+CD25hiFoxp3+regulatory T cells are derived by rapid turnover of memory populations invivo. J Clin Invest.2006,116(9):2423-33.
72.Aslam R, Hu Y, Gebremeskel S, Segel GB, Speck ER, Guo L, Kim M, Ni H,Freedman J, Semple JW. Thymic retention of CD4+CD25+FOXP3+T regulatorycells is associated with their peripheral deficiency and thrombocytopenia in a murinemodel of immune thrombocytopenia. Blood.2012,120(10):2127-32.
73.Haas J, Fritzsching B, Trübswetter P, Korporal M, Milkova L, Fritz B, Vobis D,Krammer PH, Suri-Payer E, Wildemann B. Prevalence of newly generated naiveregulatory T cells (Treg) is critical for Treg suppressive function and determines Tregdysfunction in multiple sclerosis. J Immunol.2007,179(2):1322-30.
74.Str bel P, Helmreich M, Menioudakis G, Lewin SR, Rüdiger T, Bauer A, Hoffacker V,Gold R, Nix W, Schalke B, Elert O, Semik M, Müller-Hermelink HK, Marx A.Paraneoplastic myasthenia gravis correlates with generation of mature naive CD4(+)T cells in thymomas. Blood.2002,100(1):159-66.
75.Hoffacker V, Schultz A, Tiesinga JJ, Gold R, Schalke B, Nix W, Kiefer R,Müller-Hermelink HK, Marx A. Thymomas alter the T-cell subset composition in theblood: a potential mechanism for thymoma-associated autoimmune disease. Blood.2000,96(12):3872-9.
76.Fattorossi A, Battaglia A, Buzzonetti A, Minicuci G, Riso R, Peri L, Scambia G,Evoli A. Thymopoiesis, regulatory T cells, and TCRVbeta expression in thymomawith and without myasthenia gravis, and modulatory effects of steroid therapy. J ClinImmunol.2008,28(2):194-206.
77.Sempowski G, Thomasch J, Gooding M, Hale L, Edwards L, Ciafaloni E, Sanders D,Massey J, Douek D, Koup R, Haynes B. Effect of thymectomy on human peripheralblood T cell pools in myasthenia gravis. J Immunol.2001,166(4):2808-17.
78.Wang L, Xiao W, Zheng Y, Xiao R, Zhu G, Wang M, Li Y, Peng S, Tan X, He Y,Tan J. High dose lipopolysaccharide triggers polarization of mouse thymic Th17cellsin vitro in the presence of mature dendritic cells. Cell Immunol.2012，274(1-2):98-108.
79.Kiros TG, van Kessel J, Babiuk LA, Gerdts V. Induction, regulation andphysiological role of IL-17secreting helper T-cells isolated from PBMC, thymus, andlung lymphocytes of young pigs. Vet Immunol Immunopathol.2011,144(3-4):448-54.
80. Aricha R, Feferman T, Scott HS, Souroujon MC, Berrih-Aknin S, Fuchs S. Thesusceptibility of Aire(-/-) mice to experimental myasthenia gravis involvesalterations in regulatory T cells. J Autoimmun.2011,36(1):16-24.
81.Cosmi L, De Palma R, Santarlasci V, Maggi L, Capone M, Frosali F, Rodolico G,Querci V, Abbate G, Angeli R, Berrino L, Fambrini M, Caproni M, Tonelli F, LazzeriE, Parronchi P, Liotta F, Maggi E, Romagnani S, Annunziato F. Human interleukin17-producing cells originate from a CD161+CD4+T cell precursor. J Exp Med.2008,205(8):1903-16.
82.Sofi MH, Liu Z, Zhu L, Yu Q, Kaplan MH, Chang CH. Regulation of IL-17expression by the developmental pathway of CD4T cells in the thymus. MolImmunol.2010,47(6):1262-8.
83.Kim JS, Smith-Garvin JE, Koretzky GA, Jordan MS. The requirements for naturalTh17cell development are distinct from those of conventional Th17cells. J Exp Med.2011,208(11):2201-7.
84.Raju R, Navaneetham D, Protti MP, Horton RM, Hoppe BL, Howard J Jr, Conti-FineBM. TCR V beta usage by acetylcholine receptor-specific CD4+T cells inmyasthenia gravis. Autoimmun.1997,10(2):203-17.
85.Nakahara M, Nagayama Y, Ichikawa T, Yu L, Eisenbarth GS, Abiru N. The effect ofregulatory T-cell depletion on the spectrum of organ-specific autoimmune diseases innonobese diabetic mice at different ages. Autoimmunity.2011,44(6):504-10
86.Montane J, Bischoff L, Soukhatcheva G, Dai DL, Hardenberg G, Levings MK,Orban PC, Kieffer TJ, Tan R, Verchere CB. Prevention of murine autoimmunediabetes by CCL22-mediated Treg recruitment to the pancreatic islets. J Clin Invest.2011,121(8):3024-8.
87.Viret C, Leung-Theung-Long S, Serre L, Lamare C, Vignali DA, Malissen B, CarrierA, Guerder S. Thymus-specific serine protease controls autoreactive CD4T celldevelopment and autoimmune diabetes in mice. J Clin Invest.2011,121(5):1810-21.
88.Str bel P, Chuang WY, Chuvpilo S, Zettl A, Katzenberger T, Kalbacher H,Rieckmann P, Nix W, Schalke B, Gold R, Müller-Hermelink HK, Peterson P, MarxA. Common cellular and diverse genetic basis of thymoma-associated myastheniagravis: role of MHC class II and AIRE genes and genetic polymorphisms. Ann N YAcad Sci.2008,1132:143-56.
89.Close PM, Kirchner T, Uys CJ, Müller-Hermelink HK. Reproducibility of ahistogenetic classification of thymic epithelial tumours. Histopathology.1995,26(4):339-4
90.Marx A, Schultz A, Wilisch A, Helmreich M, Nenninger R, Müller-Hermelink HK.Paraneoplastic autoimmunity in thymus tumors. Dev Immunol.1998,6(1-2):129-40.
91.Poussin MA, Tüzün E, Goluszko E, Scott BG, Yang H, Franco JU, Christadoss P.B7-1costimulatory molecule is critical for the development of experimentalautoimmune myasthenia gravis. J Immunol.2003,170(8):4389-96.
92.Wang W, Milani M, Ostlie N, et al. C57BL/6mice genetically deficient inIL-12/IL-23and IFN-gamma are susceptible to experimental autoimmune myastheniagravis, suggesting a pathogenic role of non-Th1cells. J Immunol.2007,178(11):7072-80.
93.Ostlie N, Milani M, Wang W, et al. Absence of IL-4facilitates the development ofchronic autoimmune myasthenia gravis in C57BL/6mice. J Immunol.2003,170(1):604-12.
94.Yang H, Zhang Y, Wu M, et al. Suppression of ongoing experimental autoimmunemyasthenia gravis by transfer of RelB-silenced bone marrow dentritic cells isassociated with a change from a T helper Th17/Th1to a Th2and FoxP3+regulatoryT-cell profile. inflamm Res.2010,59(3):197-205.
95.Patakas A, Platt AM, Butcher JP, Maffia P, McInnes IB, Brewer JM, Garside P,Benson RA. Putative existence of reciprocal dialogue between Tfh and B cells and itsimpact on infectious and autoimmune disease. Immunol Lett.2011,138(1):38-46.
96.Maehara T, Moriyama M, Hayashida JN, Tanaka A, Shinozaki S, Kubo Y,Matsumura K, Nakamura S. Selective localization of T helper subsets in labialsalivary glands from primary Sj gren's syndrome patients. Clin Exp Immunol.2012,169(2):89-99.
97.Simpson N, Gatenby PA, Wilson A, Malik S, Fulcher DA, Tangye SG, Manku H,Vyse TJ, Roncador G, Huttley GA, Goodnow CC, Vinuesa CG, Cook MC.Expansion of circulating T cells resembling follicular helper T cells is a fixedphenotype that identifies a subset of severe systemic lupus erythematosus. ArthritisRheum.2010,62(1):234-44.
98.Ma J, Zhu C, Ma B, Tian J, Baidoo SE, Mao C, Wu W, Chen J, Tong J, Yang M, JiaoZ, Xu H, Lu L, Wang S. Increased frequency of circulating follicular helper T cells inpatients with rheumatoid arthritis. Clin Dev Immunol.2012,2012:827480.
99.Luo C, Li Y, Liu W, Feng H, Wang H, Huang X, Qiu L, Ouyang J. Expansion ofcirculating counterparts of follicular helper T cells in patients with myasthenia gravis.Neuroimmunol.2013,256(1-2):55-61.
100.Stummvoll GH, DiPaolo RJ, Huter EN, Davidson TS, Glass D, Ward JM, ShevachEM. Th1, Th2, and Th17effector T cell-induced autoimmune gastritis differs inpathological pattern and in susceptibility to suppression by regulatory T cells. JImmunol.2008,181(3):1908-16.
101.Kinnunen T, Chamberlain N, Morbach H, et al. Accumulation of peripheralautoreactive B cells in the absence of functional human regulatory T cells. Blood.2013,121(9):1595-603.
102.Leonardo SM, De Santis JL, Gehrand A, et al. Expansion of follicular helper T cellsin the absence of Treg cells: implications for loss of B-cell anergy. Eur J Immunol.2012,42(10):2597-607.
1. Wang W, Ostlie NS, Conti-Fine BM, Milani M. The susceptibility to experimentalmyasthenia gravis of STAT6-/-and STAT4-/-BALB/c mice suggests a pathogenicrole of Th1cells. J Immunol.2004,172(1):97-103.
2. Baggi F, Annoni A, Ubiali F, Milani M, Longhi R, Scaioli W, Cornelio F,Mantegazza R, Antozzi C. Breakdown of tolerance to a self-peptide of acetylcholinereceptor alpha-subunit induces experimental myasthenia gravis in rats. J Immunol.2004,172(4):2697-703.
3. Li N, Mu L, Wang J, Zhang J, Xie X, Kong Q, Tang W, Yao X, Liu Y, Wang L,Wang G, Wang D, Jin L, Sun B, Li H. Activation of the adenosine A2A receptorattenuates experimental autoimmune myasthenia gravis severity.Eur J Immunol.2012,42(5):1140-51
4. Mu L, Sun B, Kong Q, Wang J, Wang G, Zhang S, Wang D, Liu Y, Liu Y, An H, Li H.Disequilibrium of T helper type1,2and17cells and regulatory T cells during thedevelopment of experimental autoimmune myasthenia gravis. Immunology.2009,128(1Suppl):826-36.
5. Masuda M, Matsumoto M, Tanaka S, Nakajima K, Yamada N, Ido N, Ohtsuka T,Nishida M, Hirano T, Utsumi H. Clinical implication of peripheral CD4+CD25+regulatory T cells and Th17cells in myasthenia gravis patients. J Neuroimmunol.2010,225(1-2):123-31.
6. Li X, Xiao BG, Xi JY, Lu CZ, Lu JH. Decrease of CD4+CD25highFoxp3+regulatory Tcells and elevation of CD19+BAFF-R+B cells and soluble ICAM-1in myastheniagravis. Clinical Immunology.2008,126:180–188.
7. Fattorossi A, Battaglia A, Buzzonetti A, Ciaraffa F, Scambia G, Evoli A. Circulatingand thymic CD4+CD25+T regulatory cells in myasthenia gravis: effect ofimmunosuppressive treatment. Immunology.2005,116(1):134-41.
8. Xu WH, Zhang AM, Ren MS, Zhang XD, Wang F, Xu XC, Li Q, Wang J, Din BS,Wu YB, Chen GH. Changes of Treg-associated molecules on CD4+CD25+Treg cellsin myasthenia gravis and effects of immunosuppressants. J Clin Immunol.2012,32(5):975-83.
9. Le Panse R, Cizeron-Clairac G, Cuvelier M, Truffault F, Bismuth J, Nancy P, DeRosbo NK, Berrih-Aknin S. Regulatory and pathogenic mechanisms in humanautoimmune myasthenia gravis. Ann N Y Acad Sci.2008,1132:135-42
10. Luther C, Adamopoulou E, Stoeckle C, Brucklacher-Waldert V, Rosenkranz D,Stoltze L, Lauer S, Poeschel S, Melms A, Tolosa E. Prednisolone treatment inducestolerogenic dendritic cells and a regulatory milieu in myasthenia gravis patients.Immunol.2009,183(2):841-8.
11. Sheng JR, Li L, Ganesh BB, Vasu C, Prabhakar BS, Meriggioli MN. Suppression ofexperimental autoimmune myasthenia gravis by granulocyte-macrophagecolony-stimulating factor is associated with an expansion of FoxP3+regulatory Tcells. J Immunol.2006,177(8):5296-306.
12. Sheng JR, Li LC, Ganesh BB, Prabhakar BS, Meriggioli MN. Regulatory T cellsinduced by GM-CSF suppress ongoing experimental myasthenia gravis. ClinImmunol.2008,128(2):172-80.
13. Yang H, Zhang Y, Wu M, Li J, Zhou W, Li G, Li X, Xiao B, Christadoss P.Suppression of ongoing experimental autoimmune myasthenia gravis by transfer ofRelB-silenced bone marrow dentritic cells is associated with a change from a Thelper Th17/Th1to a Th2and FoxP3+regulatory T-cell profile. Inflamm Res.2010,59(3):197-205.
14. Chang HW, Chow YH, Chong P, Sia C. The Cross-Regulatory Relationship betweenHuman Dendritic and Regulatory T Cells and its Role in Type1Diabetes Mellitus.Rev Diabet Stud.2007,4(2):68–76.
15. Harry RA, Anderson AE, Isaacs JD, Hilkens CM. Generation and characterisation oftherapeutic tolerogenic dendritic cells for rheumatoid arthritis. Ann Rheum Dis.2010Jun15.
16. Pedersen AE, Gad M, Walter MR, Claesson MH. Induction of regulatory dendriticcells by dexamethasone and1alpha,25-Dihydroxyvitamin D (3). Immunol Lett.2004,91(1):63-9.
17. Xing N, L Maldonado ML, Bachman LA, McKean DJ, Kumar R, Griffin MD.Distinctive dendritic cell modulation by vitamin D(3) and glucocorticoid pathways.Biochem Biophys Res Commun.2002,297(3):645-52.
18. Smolders J, Thewissen M, Peelen E, Menheere P, Tervaert JW, Damoiseaux J,Hupperts R. Vitamin D status is positively correlated with regulatory T cell functionin patients with multiple sclerosis. PLoS One.2009,4(8): e6635.
19. van der Mei IA, Ponsonby AL, Dwyer T, Blizzard L, Taylor BV, Kilpatrick T,Butzkueven H, McMichael AJ.Vitamin D levels in people with multiple sclerosis andcommunity controls in Tasmania, Australia. J Neurol.2007,254(5):581-90.
20. Simpson S Jr, Taylor B, Blizzard L, Ponsonby AL, Pittas F, Tremlett H, Dwyer T,Gies P, van der Mei I. Higher25-hydroxyvitamin D is associated with lower relapserisk in multiple sclerosis. Ann Neurol.2010,68(2):193-203.
21. Prietl B, Pilz S, Wolf M, Tomaschitz A, Obermayer-Pietsch B, Graninger W, PieberTR. Vitamin D supplementation and regulatory T cells in apparently healthy subjects:vitamin D treatment for autoimmune diseases? Isr Med Assoc J.2010,12(3):136-9.
22. Burton JM, Kimball S, Vieth R, Bar-Or A, Dosch HM, Cheung R, Gagne D,D'Souza C, Ursell M, O'Connor P. A phase I/II dose-escalation trial of vitamin D3and calcium in multiple sclerosis. Neurology.2010,74(23):1852-9.