Department of Rheumatology and Clinical Immunology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
State Key Laboratory of Complex Severe and Rare Diseases, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
Front Immunol. 2021 Mar 9;12:637659. doi: 10.3389/fimmu.2021.637659. eCollection 2021.
To identify the importance of the Toll-like receptor (TLR) pathway using B cell high-throughput sequencing and to explore the participation of the TLR7 signaling pathway in primary Sjogren's syndrome (pSS)-associated thrombocytopenia in patient and mouse models. High-throughput gene sequencing and bioinformatic analyses were performed for 9 patients: 3 patients with pSS and normal platelet counts, 3 patients with pSS-associated thrombocytopenia, and 3 healthy controls. Twenty-four patients with pSS were recruited for validation. Twenty-four non-obese diabetic (NOD) mice were divided into the TLR7 pathway inhibition (CA-4948), activation (Resiquimod), and control groups. Serum, peripheral blood, bone marrow, and submandibular glands were collected for thrombocytopenia and TLR7 pathway analysis. Seven hub genes enriched in the TLR pathway were identified. Compared to that in control patients, the expression of interleukin (IL)-8 and TLR7 pathway molecules in B-cells was higher in patients with pSS-associated thrombocytopenia. Platelet counts exhibited a negative correlation with serum IL-1β and IL-8 levels. In NOD mice, CA-4948/Resiquimod treatment induced the downregulation/upregulation of the TLR7 pathway, leading to consistent elevation/reduction of platelet counts. Megakaryocyte counts in the bone marrow showed an increasing trend in the Resiquimod group, with more naked nuclei. The levels of IL-1β and IL-8 in the serum and submandibular gland tissue increased in the Resiquimod group compared with that in CA-4948 and control groups. pSS-associated thrombocytopenia may be a subset of the systemic inflammatory state as the TLR7 signaling pathway was upregulated in B cells of patients with pSS-associated thrombocytopenia, and activation of the TLR7 pathway led to a thrombocytopenia phenotype in NOD mice.
为了确定 Toll 样受体（TLR）途径的重要性，我们使用 B 细胞高通量测序，并在患者和小鼠模型中探索 TLR7 信号通路在原发性干燥综合征（pSS）相关血小板减少症中的参与。对 9 名患者进行了高通量基因测序和生物信息学分析：3 名 pSS 患者血小板计数正常，3 名 pSS 相关血小板减少症患者，3 名健康对照者。对 24 名 pSS 患者进行了验证。将 24 只非肥胖型糖尿病（NOD）小鼠分为 TLR7 通路抑制剂（CA-4948）、激活剂（Resiquimod）和对照组。收集血清、外周血、骨髓和颌下腺进行血小板减少症和 TLR7 通路分析。鉴定出 7 个在 TLR 通路中富集的枢纽基因。与对照组患者相比，pSS 相关血小板减少症患者 B 细胞中白细胞介素（IL）-8 和 TLR7 通路分子的表达更高。血小板计数与血清 IL-1β 和 IL-8 水平呈负相关。在 NOD 小鼠中，CA-4948/Resiquimod 治疗诱导 TLR7 通路的下调/上调，导致血小板计数一致升高/降低。骨髓中巨核细胞计数在 Resiquimod 组呈上升趋势，裸核增多。与 CA-4948 和对照组相比，Resiquimod 组血清和颌下腺组织中 IL-1β 和 IL-8 水平升高。pSS 相关血小板减少症可能是全身炎症状态的一个亚群，因为在 pSS 相关血小板减少症患者的 B 细胞中 TLR7 信号通路被上调，并且 TLR7 通路的激活导致 NOD 小鼠出现血小板减少症表型。
