Kv1.3通道调节动脉粥样硬化适应性免疫的作用及分子机制的研究

Modulation of the activities of macrophages and T lymphocytes contributes to the stability of atherosclerotic plaque. In the preliminary study, we successfully invented a novel specific human Kv1.3 (hKv1.3) channel blocker-the antibody against the hKv1.3-E314 peptide. By blocking Kv1.3 channels expressed on human macrophage membranes, the antibody played an important role in the modulation of atherosclerosis (AS) innate immunity and exhibited a beneficial effect on cholesterol metabolism, which suggests that the Kv1.3 channel may be a good target for the treatment of AS. Further researches are needed to address the role of the Kv1.3 channel in AS adaptive immunity. We suppose that the Kv1.3 channel can regulate the imbalance between T lymphocyte-mediated pro-inflammatory and anti-inflammatory response in AS, which promotes the stability of AS plaque. We intend to develop the monoclonal antibody against the hKv1.3-E314 peptide as an agent to specifically block Kv1.3 channels expressed on T lymphocyte membranes in AS. In vitro and vivo experiments are going to be performed around the effect and mechanism of Kv1.3 blockade on secretion, homing, migration and differentiation of T lymphocytes in AS. The results will clarify the role of Kv1.3 channels in AS adaptive immunity. This study will provide experimental evidence for comprehensive evaluation about the Kv1.3 channel as a target for the treatment of AS.

调节动脉粥样硬化(AS)中巨噬细胞和T淋巴细胞等炎症细胞的活性，有助于促进AS斑块的稳定。我们在前期的研究中，成功研制了一种新型、特异性人Kv1.3（hKv1.3）通道阻滞剂- - 抗hKv1.3-E314肽段抗体。该抗体能阻滞人巨噬细胞膜表面的Kv1.3通道，有效地调节AS的固有免疫，改善巨噬细胞的胆固醇代谢。Kv1.3通道可能是AS良好的干预靶点。进一步研究须阐明Kv1.3通道对AS适应性免疫的调节作用及机制。我们设想，Kv1.3通道能够调节T淋巴细胞介导的AS促炎/抗炎反应平衡，促进AS斑块的稳定。我们拟研制抗hKv1.3-E314肽段单克隆抗体，阻滞人AS中T细胞膜上的Kv1.3通道，通过离体和在体实验，观察其对T细胞的分泌、归巢和迁移、分化功能的作用及机制。本研究将阐明Kv1.3通道调节AS适应性免疫的作用及分子机制, 从而为全面评价Kv1.3通道能否作为AS的干预靶点提供实验依据。

调节动脉粥样硬化(AS)中巨噬细胞和T淋巴细胞等炎症细胞的活性，有助于促进 AS 斑块的稳定。我们在前期的研究中，成功研制了一种新型、特异性人 Kv1.3（hKv1.3）通道阻滞剂—抗hKv1.3-E314 肽段抗体。该抗体能阻滞人巨噬细胞膜表面的 Kv1.3通道，有效地调节AS的固有免疫，改善巨噬细胞的胆固醇代谢，减少泡沫细胞的形成。 Kv1.3通道亦表达于T淋巴细胞。因此，Kv1.3通道可能通过调节AS固有免疫和适应性免疫， 从而成为AS 良好的干预靶点。. 本研究从离体和在体两方面，通过抗hKv1.3-E314肽段单克隆抗体（专利申请号：201110044416.X）特异性地阻滞巨噬细胞和T淋巴细胞膜上的 Kv1.3 通道，调节巨噬细胞和T淋巴细胞的活性，抑制其促炎反应，增强其抗炎效应，进而促进 AS 斑块的稳定。. 目前我们已经成功完成hKv1.3胞外环肽段E314的合成和抗hKv1.3-E314肽段单克隆抗体的制备，并通过免疫学方法及膜片钳技术验证该单克隆抗体的特异性和亲和力。通过hKv1.3-E314单克隆抗体特异性阻滞巨噬细胞和T淋巴细胞膜上的Kv1.3通道，调节巨噬细胞和T淋巴细胞介导的AS 促炎/抗炎反应平衡，促进 AS 斑块的稳定。此外，我们成功引进并繁殖Kv1.3基因敲除小鼠，建立动脉粥样硬化模型，证实Kv1.3表达缺失可以改善AS斑块内巨噬细胞的胆固醇代谢，并调节 Mo/MΦ亚群的活性及其流入/流出平衡；同时，Kv1.3表达缺失能抑制促炎性Ly6Chigh型MΦ生成和浸润、下调促炎性M1型MΦ活性、上调抑炎性M2型MΦ，能促进T细胞向抑炎型Th2/Treg的分化、抑制其向Th1/Th17的分化；最终有效减少脂质核心的大小、增加纤维帽的厚度，增强斑块稳定性。进一步阐明 Kv1.3通道调节AS固有免疫和适应性免疫的作用及分子机制，从而为AS的治疗提供新的干预靶点。