In-situ-hybridization analysis of HTLV-1-related diseases, employing highly biotinylated probes synthesized by polymerase chain reaction

本项目旨在利用聚合酶链反应合成的高度生物素化探针，采用原位杂交（ISH）技术检测HTLV-1前病毒DNA及其在HTLV-1相关疾病，特别是恶性淋巴瘤（MLs）细胞组成中的信号。ISH未能检测到HTLV-1前病毒DNA，但在HTLV-1相关细胞系细胞和人ML细胞的石蜡切片中成功检测到其信号。HTLV-1前病毒DNA pX Tax区在t细胞多形性淋巴瘤（T-Pleo）包括成人t细胞白血病/淋巴瘤（ATLL）中被识别的频率最高。部分低级别t细胞MLs和其他MLs的信号提示除了ATLL外，还存在htlv -1相关的MLs，并提示htlv -1相关的低级别t细胞MLs向ATLL的促进。在MLs中进行HTLV-1 p40tax的抗原检索免疫组化检测，揭示ISH中的信号。但在许多t细胞MLs中，p40tax分布在ML细胞的细胞质中，在少数t细胞MLs中，p40tax分布在细胞核中，这表明直接比较HTLV-1前病毒DNA的信号与宿主细胞基因的激活将无法看到HTLV-1 p40tax的反激活功能。通过抗原检索和ImmunoMax方法可以在ATLL细胞的细胞核中观察到少量的p40tax。这样，在HTLV-1相关疾病中，考虑到HTLV-1 p40tax除反激活外的功能，通过ISH与抗原检索和ImmunoMax相结合的方法，就可以分析HTLV-1前病毒DNA的激活与宿主细胞原的关系。

This project aimed to detect HTLV-1 proviral DNA and its signals in cellular composition of HTLV-1-related diseases, especially malignant lymphomas (MLs), by means of in-situ-hybridization (ISH) employing highly biotinylated probes synthesized by polymerase chain reaction. The ISH failed to detect HTLV-1 proviral DNA but succeeded to detect its signals in paraffin sections of HTLV-1-related cell line cells and human ML cells. The signals of HTLV-1 proviral DNA pX Tax region were recognized the most frequently in T-cell pleomorphic lymphoma (T-Pleo) including adult T-cell leukemia/lymphoma (ATLL). The signals in some of low-grade T-cell MLs and the other MLs suggested the existence of HTLV-1-related MLs other than ATLL and the promotion from HTLV-1-related low-grade T-cell MLs to ATLL.Antigen-retrieval immunohistochemical detection of HTLV-1 p40tax was performed in the MLs revealing the signals in ISH.Bur p40tax distributed in cytoplasm of ML cells in many T-cell MLs and in nuclei in a small number of T-cell MLs, suggesting that the direct comparision of the signals of HTLV-1 proviral DNA and the activation of host cell genes would fail to see the transactivating function of HTLV-1 p40tax. A small amount of p40tax in the nuclei of ATLL cells could be visualized by antigen-retrieval and ImmunoMax method. Then, the relation of the activation of HTLV-1 proviral DNA and host cell gens in HTLV-1-related diseases will be able tp be analyzed by means of the combination of the ISH and the antigen-retrieval and ImmunoMax method, considering the HTLV-1 p40tax's functions other than the transactivation.