Role of regulatory B-cells in pathogenesis and progression of autoimmune myocarditis

自身免疫性心肌炎可发展为慢性扩张型心肌病，并增加心力衰竭的风险，其发病机制尚不完全清楚。这表明，功能失调的细胞免疫调节可能会导致不平衡的促炎和抗炎过程，从而支持这种复杂的疾病。虽然T细胞在自身免疫性心肌炎中的重要作用已被广泛接受，但B细胞的作用仍不完全清楚。首先，研究仅限于伴随疾病进展的自身抗体（自身抗体）。特别是调节性B细胞（布雷格）的亚群，可能能够潜在地抵消炎症过程尚未进行研究。我们在心肌肌钙蛋白I（cTnI）诱导的实验性自身免疫性心肌炎（EAM）小鼠模型中的初步工作已经指出了布雷格在控制这种疾病方面的强大影响：虽然从cTnI免疫的小鼠中过继转移T细胞导致大多数未免疫的小鼠发生EAM，正如所预期的，这些T细胞与自体B细胞一起过继共转移强烈降低了EAM的发生率和程度。这意味着可能存在不同的B细胞亚群，具有支持或抵消心肌炎的潜力。在本项目提案中，我们追求3个目标：1）表征EAM中布雷格被激活和发挥功能的条件：为此，在将cTnI特异性T细胞转移到未免疫小鼠中之前，将在不同条件下刺激B细胞。此外，在免疫接种之前或之后将布雷格转移至EAM-小鼠，以评估它们是否可以保护免受EAM或甚至下调炎症环境中的症状. 2）评估布雷格对细胞环境的影响.在此，将使用来自cTnI免疫和布雷格处理的小鼠的各种免疫细胞进行功能测定，以研究布雷格对免疫应答的调节作用。进一步，我们尝试通过分选可能代表布雷格的B细胞亚型并测试其调节能力来揭示布雷格表型。3）评估布雷格亚型的作用和自身抗体对EAM的影响。为此，将用cTnI免疫B细胞缺陷小鼠（JH-/-），并与B细胞感受态小鼠进行比较。此外，还添加了布雷格细胞（或第2部分中发现的布雷格候选细胞）。此外，将布雷格与cTnI-T细胞共转移至布雷格缺陷小鼠中。通过添加B细胞培养物上清液和EAM-血清（具有和不具有所含抗体的消耗）来补充实验，以评估自身抗体在EAM进展中的影响。通过研究布雷格在EAM中的作用，为恢复心肌炎的免疫平衡提供线索，为开发新型的免疫治疗药物奠定基础。因此，我们计划将我们在肌钙蛋白I-EAM（卡亚博士）方面的长期专业知识与布雷格在慢性炎症方面的专业知识（特雷特博士）合并。

The pathogenesis of autoimmune myocarditis that can progress into chronically dilated cardiomyopathy with an increased risk for heart failure is not completely understood. It is suggested that dysfunctional cellular immunoregulation might lead to an imbalance of pro- and anti-inflammatory processes and thereby support this complex disease. While the crucial impact of the T cells in autoimmune myocarditis is widely accepted, the role of B cells is still incompletely understood. Primarily, research is restricted to the auto-antibodies (auto-Abs) that accompany disease progression. Especially the subpopulation of regulatory B cells (Breg) that might be able to potentially counteract the inflammatory process has not been investigated, yet. Preliminary work in our mouse model of cardiac troponin I (cTnI) -induced experimental autoimmune myocarditis (EAM) already points to a strong impact of Breg in control of this disease: while adoptive transfer of T cells from cTnI immunized mice resulted in development of EAM in the majority of non-immunized mice, as expected, adoptive cotransfer of those T cells together with autologous B-cells strongly reduced incidence and extent of EAM. This implies that different subpopulations of B cells might exist, with the potential to support or to counteract myocardits. In this project proposal we pursue 3 aims: 1) Characterize the conditions under which Breg in EAM are activated and functional: To this end B cells will be stimulated under different conditions before transfer with cTnI specific T cells into non-immunized mice. Further Breg will be transferred to EAM- mice before or after their immunization to evaluate, whether they can protect from EAM or even down regulate symptoms in an inflammatory environment.2) Assess the influence of Breg on the cellular environment. Here functional assays with various immune cells from cTnI immunized and Breg treated mice will be performed to investigate the modulatory effects of Breg on immune responses. Further we attempt to reveal the Breg phenotype by sorting B-cell subtypes that might represent Breg and test their regulatory abilities.3) Evaluate the role of Breg subtypes and impact of auto-Abs on EAM. To this end B cell deficient mice (JH-/-) will be immunized with cTnI and compared to B cell competent mice. In addition Breg cells (or Breg candidates found in part 2) are added. Further, cotransfer of Breg, with cTnI-T cells into Breg deficient mice will be performed. The experiments are supplemented by addition of B cell culture supernatant and EAM-serum with and without depletion of containing antibodies to assess the impact of auto-Abs in progression of EAM. By investigating of the role of Breg in EAM we aim to find clues to restore the immunological balance in myocarditis This might help to develop novel immunotherapeutics in future. We therefore plan to merge our long-standing expertise in the cTnI-EAM (Dr.Kaya) with that of Breg in chronic inflammation (Dr.Tretter).