ROLE OF HUMAN T LYMPHOTROPIC VIRUS TYPE I ON PATHOGENESIS OF SJOGREN'S SYNDROME

人类T细胞嗜淋巴病毒I型（HTLV-I）是一种人类逆转录病毒，最初被确定为成人T细胞白血病的病原，并与HTLV-I相关的脊髓病/热带痉挛性截瘫（HAM/TSP）密切相关。我们之前的研究表明，干燥综合征（SS）患者HTLV-I的血清阳性率为23.0%，显著高于正常献血者（3.4%）。根据Mantel-Haenszel方法估计，SS患者与正常献血者相比，经年龄调整的HTLV-I感染的总优势比为3.1。此外，病因部分，SS患者归因于HTLV-I感染的比例估计为17.6%。首先，我们调查了SS是否合并了HAM患者。对14例HAM患者的临床特点、组织学表现及血清自身抗体的流行情况进行了分析。所有患者在组织学上均表现为单个核细胞浸润于阴唇…更多唾液腺。根据欧共体提出的SS初步标准，确诊SS 8例，疑似SS 4例。8例确诊性SS患者、htlv - i血清阳性和-血清阴性SS患者的自身抗体包括类风湿因子、抗核、抗SS- a （Ro）和抗SS- b （La）抗体的患病率无显著差异。CD4^+ T细胞优先浸润到HAM患者的唾液腺，以及htlv - i血清阳性和-血清阴性患者的唾液腺。接下来，研究htlv - 1血清阳性和-血清阴性SS患者唇唾液腺的凋亡情况。在唇唾液腺上皮细胞（腺泡和导管）中发现凋亡细胞死亡。在唇唾液腺上皮细胞上发现Fas抗原。浸润淋巴细胞染色Fas配体。血清htlv - 1阳性与血清- 1阴性的SS患者在上述结果上无显著差异。我们采用巢式两步PCR检测7例HTLV-I血清阳性SS患者外周血淋巴细胞和唇唾液腺HTLV-I前病毒DNA的存在。HTLV-I基因组LTR、gag、pol和pX 4个独立区域均可从所有血清阳性患者外周血淋巴细胞和组织DNA中扩增出来。接下来，我们用半定量PCR方法研究HTLV-I在每个样本DNA中的拷贝数。组织拷贝数与淋巴细胞拷贝数之比不同，但无HAM基因的4例SS患者中有3例拷贝数非常高。这些发现提示HTLV-I感染的细胞可能在这些患者的唾液腺中积聚。从以上结果来看，可能是HAM患者外周血T细胞优先浸润到唾液腺，然后浸润的T细胞引起SS外分泌器官的病理改变。因此，我们研究了来自HAM患者的循环T细胞通过重组基底膜的迁移活性。在Transwell细胞培养室中测定淋巴细胞的移行活性。我们将HAM患者和正常人的循环CD4^+细胞分别应用于Transwell细胞培养室，然后计算迁移细胞的数量。HAM患者的CD4^+细胞转移率明显高于正常人。此外，在HAM患者转移的CD4^+细胞中量化的HTLV-I前病毒DNA比未转移的CD4^+细胞高2至8倍。这些结果表明HTLV-I感染的HAM患者的CD4^+细胞通过重组基底膜的迁移活性上调。总之，我们的研究结果有力地支持了HTLV-I参与流行地区部分SS患者发病机制的观点。少

Human T-cell lymphotropic virus type I (HTLV-I) is a human retrovirus, originally identified as an aetiological agent of adult T cell leukemia, and closely associated with HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). We previously demonstrated that the HTLV-I seroprevalence in Sjogren's syndrome (SS) patients was 23.0%, which was significantly higher than those of the normal blood donors (3.4%). The age-adjusted summary odds ratio of HTLV-I infection in SS patients as compared with normal blood donors, estimated by the Mantel-Haenszel procedure, was 3.1. Furthermore, etiologic fraction, the proportion of SS patients attributable to HTLV-I infection, was estimated to be 17.6%.At first, we investigated whether SS was complicated with HAM patients. The clinical feature and histological findings of SS and the prevalence of serum autoantibodies were examined 14 patients with HAM.All patients with HAM histologically showed a mononuclear cell infiltration in the labial … More salivary glands. According to the preliminary criteria for SS proposed by the European Community, definitive SS was diagnosed in 8 patients and probable SS in 4 patients. There was no significant difference in the prevalence of autoantibodies including rheumatoid factor, anti-nuclear, anti-SS-A (Ro) and anti-SS-B (La) antibodies among 8 HAM patients with definitive SS,HTLV-I-seropositive and- seronegative SS patients. The CD4^+ T cells preferentially infiltrated into the salivary glands in HAM patients as well as the salivary glands of patients with HTLV-I-seropositive and -seronegative patients.Next, apoptosis in labial salivary glands from HTLV-I-seropositive and -seronegative SS patients was studied. Apoptotic cell death was detected in epithelial cells (acinus and ducts) of labial salivary glands. Fas antigen was found on epithelial cells in labial salivary glands. Fas ligand was stained in the infiltrated lymphocytes. There was no significant difference in above findings between HTLV-I-seropositive and -seronegative SS patients.We examined the presence of HTLV-I provirus DNA of both peripheral blood lymphocytes and labial salivary glands form 7 HTLV-I seropositive patients with SS by nested two-step PCR.All 4 independent regions of the HTLV-I genomic, including LTR,gag, pol and pX,were readily amplified from DNA of peripheral blood lymphocytes and tissues of all the seropositive patients. Next, we studied the copy number of HTLV-I in each sample DNA by semiquantitative PCR.The ratio of the copy number between tissue and lymphocytes was variable, but 3 out of 4 SS patients without HAM gene very high values. These findings suggest that HTLV-I infected cells may accumulate in the salivary gland of these patients. From the above findings, it seems likely that peripheral blood T cells from HAM patients preferentially infiltrate into the salivary glands, and then the infiltrated T cells induce SS the pathological changes in the exocrine organs. We studied, therefore, transmigratory activity of circulating T cells from HAM patients through reconstituted basement membrane. Transmigratory activity of lymphocytes was assayd in Transwell cell-culture chambers. We applied the circulating CD4^+ cells from patients with HAM and normal subjects on Transwell cell-culture chambers, and then counted the number of transmigrated cells. The percentage of transmigrated CD4^+ cells from HAM patients was significantly higher than that from normal subjects. Furthermore, HTLV-I proviral DNA quantified in the transmigrated CD4^+ cells from HAM patients were 2 to 8 fold more frequent than that in the non-transmigrated CD4^+ cells. These findings indicate that HTLV-I infected CD4^+ cells of HAM patients have up-regulated transmigratory activity through reconstituted basement membrane.In summary, our findings strongly support the idea that HTLV-I is involved in the pathogenesis of the disease in a subset of patients with SS in endemic areas. Less