Wakamatsu Ei, Matsumoto Isao, Yasukochi Takanori, Naito Yusuke, Goto Daisuke, Mamura Mizuko, Ito Satoshi, Tsutsumi Akito, Sumida Takayuki
University of Tsukuba, Tsukuba City, Ibaraki 305-8575, Japan.
Arthritis Rheum. 2006 Nov;54(11):3476-84. doi: 10.1002/art.22176.
To clarify the molecular mechanisms of Sjögren's syndrome (SS), we analyzed the functional role of the STAT-1 gene, one of the interferon-gamma (IFNgamma)-inducible genes, in labial salivary glands (LSGs) from SS patients.
The expression of STAT-1 messenger RNA (mRNA) was examined by real-time polymerase chain reaction (PCR) analysis, and the phosphorylation of STAT-1 protein (Tyr(701) and Ser(727) pSTAT-1) was investigated by Western blot and immunohistochemical analyses. The expression of IFNgamma-inducible 10-kd protein (IP-10), IFN regulatory factor 1 (IRF-1), and Fas was also examined by real-time PCR and immunohistochemical analyses.
STAT-1alpha and STAT-1beta mRNA were highly expressed in LSGs from SS patients. The level of STAT-1alpha protein in SS LSGs was higher than that in 3 control LSGs, whereas STAT-1beta protein was not clearly detected by Western blot analysis. Moreover, Tyr(701) and Ser(727) pSTAT-1alpha proteins were specifically detected in SS LSGs. Immunohistochemical analysis showed localization of Tyr(701) pSTAT-1 in infiltrating lymphocytes and the adjacent ductal epithelium from SS patients. Ser(727) pSTAT-1 was localized only in the ductal epithelium of SS LSGs. The STAT-1-inducible genes IP-10 and IRF-1 and the Fas genes were highly expressed in SS LSGs and were colocalized with Ser(727) pSTAT-1-positive, but not Tyr(701) pSTAT-1-positive, cells.
We found evidence of the up-regulation of STAT-1alpha mRNA and protein in LSGs from SS patients, as well as the presence of pSTAT-1alpha in ductal epithelium from SS patients. Our findings suggest that STAT-1alpha, especially Ser(727) pSTAT-1, may function as a key molecule in the pathogenesis of SS.
为阐明干燥综合征（SS）的分子机制，我们分析了信号转导子和转录激活子1（STAT-1）基因（一种γ干扰素（IFNγ）诱导基因）在SS患者唇腺（LSG）中的功能作用。
通过实时聚合酶链反应（PCR）分析检测STAT-1信使核糖核酸（mRNA）的表达，并通过蛋白质印迹法和免疫组织化学分析研究STAT-1蛋白（酪氨酸（Tyr）701和丝氨酸（Ser）727磷酸化STAT-1）的磷酸化情况。还通过实时PCR和免疫组织化学分析检测IFNγ诱导的10千道尔顿蛋白（IP-10）、IFN调节因子1（IRF-1）和Fas的表达。
STAT-1α和STAT-1β mRNA在SS患者的LSG中高表达。SS患者LSG中STAT-1α蛋白水平高于3个对照LSG中的水平，而蛋白质印迹分析未清楚检测到STAT-1β蛋白。此外，在SS患者的LSG中特异性检测到酪氨酸（Tyr）701和丝氨酸（Ser）727磷酸化STAT-1α蛋白。免疫组织化学分析显示，酪氨酸（Tyr）701磷酸化STAT-1定位于SS患者浸润淋巴细胞及相邻导管上皮中。丝氨酸（Ser）727磷酸化STAT-1仅定位于SS患者LSG的导管上皮中。STAT-1诱导基因IP-10和IRF-1以及Fas基因在SS患者的LSG中高表达，且与丝氨酸（Ser）727磷酸化STAT-1阳性细胞共定位，但不与酪氨酸（Tyr）701磷酸化STAT-1阳性细胞共定位。
我们发现了SS患者LSG中STAT-1α mRNA和蛋白上调的证据，以及SS患者导管上皮中磷酸化STAT-1α的存在。我们的研究结果表明，STAT-1α，尤其是丝氨酸（Ser）727磷酸化STAT-1，可能在SS发病机制中起关键分子的作用。
