Maturation of dendritic cells and T-cell responses in sentinel lymph nodes from patients with breast carcinomca
Maturation of dendritic cells and T-cell responses in sentinel lymph nodes from patients with breast carcinomca
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作者：
Matsuura, K;Yamaguchi, Y;Toge, T
背景资料。前哨淋巴结(SNS)的识别方法及其临床意义已经建立。分子免疫学的最新进展使分析精确的免疫反应成为可能。本研究的目的是阐明乳腺癌患者树突状细胞(DC)成熟、辅助性T细胞I型(Th-1)和辅助性T细胞(Th-2)反应以及调节性T细胞反应。方法：对70例临床淋巴结阴性(NO)乳腺癌患者采用放射导向和蓝色染料引导的方法鉴定SNS和非SNS。用CD83-异硫氰酸荧光素(FITC)、CD80-藻红蛋白(PE)、CD86-PE、CD40-PE、人类白血病D相关抗原(HLADR)-FITC、CD4-FITC和CD25-PE抗体进行流式细胞仪(FCM)分析。提取SNS和非SNS的总RNA，采用实时定量逆转录聚合酶链式反应(RT-PCR)检测CD83、IL-12p40、干扰素-γ、IL-4、IL-10和Foxp3的表达。关于微转移，进一步分析了SNS的免疫学状态，这些微转移在显微镜下被鉴定为阴性，但根据针对乳巨球蛋白的RT-PCR分析呈阳性。结果：在71例临床上未发现乳腺癌的患者中，70例(98.6%)可检测到SNS。70例患者中有14例(20.0%)有SNS阳性转移。56例转移阴性患者的SNS与非SNS比较，流式细胞仪检测显示SNS中HLADR阳性、CD80阳性、CD86阳性和CD40阳性细胞数均显著减少。RT-PCR分析显示，在44例转移阴性的SNS患者中，SNS组CD83和干扰素-γmRNA的表达水平显著低于非SNS组。对44例转移阴性和14例转移阳性的淋巴结的免疫学参数进行了比较。转移阳性组CD83、IL-12p40、干扰素-γ、IL-10和Foxp3的表达显著高于转移阴性组。结论：在乳腺癌患者中，SNS的细胞免疫反应，从DC成熟到Th-1反应，在转移发生之前，与非SNS相比，似乎不那么活跃。一旦SNS发生转移，DC成熟即被触发，随后Th-1反应上调，这可能反映了SNS中的抗原特异性免疫反应。与SNS的DC成熟和转移后的Th-1反应不同，Th-2的上调和调节性T细胞反应是平行发展的。
BACKGROUND. Methods for identifying sentinel lymph nodes (SNs) and their clinical significance have been established. Recent advances in molecular immunology have enabled the analysis of precise immune responses. The objective of the current study was to clarify the dendritic cell (DC) maturation, T-helper type I (Th-1) and Th-2 responses, and regulatory T-cell responses of SNs in patients with breast carcinoma.METHODS. SNs and non-SNs were identified by radioguided and blue dye-guided methods in 70 consecutive patients with clinically lymph node negative (NO) breast carcinoma. Lymphocytes were collected from SNs and non-SNs and were subjected to flow cytometric analysis (FCM) using antibodies of CD83-fluorescein isothiocyanate (FITC), CD80-phycoerythrin (PE), CD86-PE, CD40-PE, human leukemic D-related antigen (HLA-DR)-FITC, CD4-FITC, and CD25-PE. Total RNA was extracted from SNs and non-SNs, and the expression of CD83, interleukin 12p40 (IL-12p40), interferon gamma (IFN-gamma), IL-4, IL-10, and Foxp3 was evaluated by using quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. The immunologic status of SNs was analyzed further with regard to micrometastases, which were identified as negative microscopically but positive according to an RT-PCR analysis that was specific for mammaglobin.RESULTS. SNs were detectable in 70 of 71 consecutive patients (98.6%) with clinically NO breast carcinoma. Fourteen of 70 patients (20.0%) had positive metastasis in SNs. When SNs were compared with non-SNs in 56 metastasis-negative patients, FCM revealed that HLA-DR-positive, CD80-positive, CD86-positive, and CD40-positive cell populations were decreased significantly in SNs. RT-PCR analysis demonstrated that, among 44 patients with metastasis-negative SNs, the expression levels of CD83 and IFN-gamma mRNA were significantly lower in SNs compared with non-SNs. Immunologic parameters also were compared between 44 metastasis-negative SNs and 14 metastasis-positive SNs. The metastasis-positive SNs demonstrated significantly higher expression of CD83, IL-12p40, IFN-gamma, IL-10, and Foxp3 mRNA than the metastasis-negative SNs. Correction of micrometastasis detected by mammaglobin enhanced these differences consistently.CONCLUSIONS. in patients with breast carcinoma, cellular immune responses, from DC maturation to Th-1 responses, appeared to be less active in SNs compared with non-SNs before metastasis developed. Once metastasis was established in SNs, DC maturation was triggered and was followed by the up-regulation of Th-1 responses, which may reflect antigen-specific immune responses in SNs. Unlike DC maturation and Th-1 responses after metastasis in SNs, up-regulation of Th-2 and regulatory T-cell responses developed in parallel.