Table_1_Differential Expression of Immune Response Genes in Asymptomatic Chronic Chagas Disease Patients Versus Healthy Subjects.docx

Infection by the Trypanosoma cruzi parasite causes Chagas disease and triggers multiple immune mechanisms in the host to combat the pathogen. Chagas disease has a variable clinical presentation and progression, producing in the chronic phase a fragile balance between the host immune response and parasite replication that keeps patients in a clinically silent asymptomatic stage for years. Since the parasite is intracellular and replicates within cells, the cell-mediated response of the host adaptive immunity plays a critical role. This function is mainly orchestrated by T lymphocytes, which recognize parasite antigens and promote specific functions to control the infection. However, little is known about the immunological markers associated with this asymptomatic stage of the disease. In this large-scale analysis, the differential expression of 106 immune system-related genes has been analyzed using high-throughput qPCR in T. cruzi antigen-stimulated PBMC from chronic Chagas disease patients with indeterminate form (IND) and healthy donors (HD) from endemic and non-endemic areas of Chagas disease. This analysis revealed that there were no differences in the expression level of most genes under study between healthy donors from endemic and non-endemic areas determined by PCA and differential gene expression analysis. Instead, PCA revealed the existence of different expression profiles between IND patients and HD (p < 0.0001), dependent on the 32 genes included in PC1. Differential gene expression analysis also revealed 23 upregulated genes (expression fold change > 2) and 11 downregulated genes (expression fold change < 0.5) in IND patients versus HD. Enrichment analysis showed that several upregulated genes in IND patients participate in relevant immunological pathways such as antigen-dependent B cell activation, stress induction of HSP regulation, NO2-dependent IL12 pathway in NK cells, and cytokine-inflammatory response. The antigen-specific differential gene expression profile detected in these patients and the relevant immunological pathways that seem to be activated could represent potential biomarkers of the asymptomatic form of Chagas disease, helpful to diagnosis and infection control.

克氏锥虫（Trypanosoma cruzi）感染可引发恰加斯病（Chagas disease），并触发宿主多重免疫机制以对抗病原体。恰加斯病的临床表现与病程存在显著异质性，在慢性阶段，宿主免疫应答与寄生虫复制之间维持着脆弱的动态平衡，使得患者可在数年时间内处于临床无症状的隐匿感染状态。由于该寄生虫为胞内寄生生物，可在宿主细胞内增殖，宿主适应性免疫的细胞介导应答发挥着核心调控作用。这一免疫功能主要由T淋巴细胞介导，其可识别寄生虫抗原并激活特异性免疫通路以控制感染进程。然而，目前学界对与该病无症状阶段相关的免疫学标志物仍知之甚少。本研究开展了一项大规模分析：针对恰加斯病流行区与非流行区的健康供体（Healthy Donors, HD），以及慢性恰加斯病隐匿型（Indeterminate form, IND）患者，通过高通量定量PCR（quantitative PCR, qPCR）检测其经克氏锥虫抗原刺激后的外周血单个核细胞（Peripheral Blood Mononuclear Cell, PBMC）中106个免疫相关基因的差异表达情况。分析结果显示：经主成分分析（Principal Component Analysis, PCA）与差异基因表达分析验证，流行区与非流行区的健康供体之间，绝大多数待测基因的表达水平无显著差异。与之相反，PCA分析揭示隐匿型患者与健康供体的基因表达谱存在显著分化（p < 0.0001），该差异由PC1所包含的32个基因主导。差异基因表达分析同样发现，相较于健康供体，隐匿型患者体内存在23个上调基因（表达倍数变化>2）与11个下调基因（表达倍数变化<0.5）。富集分析结果表明，隐匿型患者体内的多个上调基因参与了多条关键免疫学通路，包括抗原依赖性B细胞活化、热休克蛋白（Heat Shock Protein, HSP）调控的应激诱导、NK细胞中NO2依赖性IL12通路，以及细胞因子-炎症应答。本研究在患者中检测到的抗原特异性差异基因表达谱，以及其所涉及的激活型免疫学通路，或可作为恰加斯病无症状型的潜在生物标志物，为该病的临床诊断与感染防控提供新的辅助手段。