Sisto Margherita, D'Amore Massimo, Lofrumento Dario Domenico, Scagliusi Pasquale, D'Amore Simona, Mitolo Vincenzo, Lisi Sabrina
Department of Human Anatomy and Histology, University of Bari Medical School, Bari, Italy.
Int Immunol. 2009 Mar;21(3):303-11. doi: 10.1093/intimm/dxp001. Epub 2009 Feb 3.
Important changes in acinar and ductal morphology and function, together with pronounced extracellular matrix (ECM) remodelling, are detectable in the labial salivary glands of Sjögren's syndrome (SS) patients. The objective of this work was to determine the effect of treatment with the anti-Ro/SSA auto-antibodies, characterizing SS, on the expression of fibulin-6, a member of the fibulins family of the ECM, in primary human salivary gland epithelial cell (SGEC) cultures established from biopsies of labial minor salivary glands obtained from healthy donors. The induction of cell detachment and anoikis in SGECs treated with anti-Ro/SSA auto-antibodies were also investigated. Changes in fibulin-6 mRNA expression were measured by semi-quantitative reverse transcriptase-PCR and real-time PCR. Fibulin-6 expression in cells treated with anti-Ro/SSA auto-antibodies was evaluated by flow cytometric analysis and confocal laser scanning microscopy. SGECs undergoing death by anoikis were identified and quantified using Calcein blue/YOPRO-1 dyes. Herein, we present the first evidence of fibulin-6 expression in SGEC that results distributed in the cytoplasm surrounding the inner side of the plasma membrane. Fibulin-6 was down-regulated in SGECs treated with anti-Ro/SSA auto-antibodies in which a marked cell detachment and a reduction of cell viability were observed. Notably, a reduction of fibulin-6 expression in SGECs treated with anti-Ro/SSA auto-antibodies corresponds to an increase of anoikis cell death. Our observations demonstrate a dysregulation of fibulin-6 in the pathological processes observed in SS and provide new evidence that disorganization of the ECM environment could damage the architecture and function of the salivary glands.
在干燥综合征（SS）患者的唇唾液腺中，可检测到腺泡和导管形态及功能的重要变化，以及明显的细胞外基质（ECM）重塑。本研究的目的是确定用抗Ro/SSA自身抗体（其为SS的特征性抗体）处理后，对从健康供体的唇小唾液腺活检组织建立的原代人唾液腺上皮细胞（SGEC）培养物中纤连蛋白-6（ECM纤连蛋白家族的成员）表达的影响。还研究了用抗Ro/SSA自身抗体处理的SGEC中细胞脱离和失巢凋亡的诱导情况。通过半定量逆转录聚合酶链反应（RT-PCR）和实时PCR测量纤连蛋白-6 mRNA表达的变化。通过流式细胞术分析和共聚焦激光扫描显微镜评估用抗Ro/SSA自身抗体处理的细胞中纤连蛋白-6的表达。使用钙黄绿素蓝/碘化丙啶（YOPRO-1）染料鉴定和定量经历失巢凋亡死亡的SGEC。在此，我们首次证明了SGEC中纤连蛋白-6的表达，其结果分布在围绕质膜内侧的细胞质中。在用抗Ro/SSA自身抗体处理的SGEC中，纤连蛋白-6表达下调，其中观察到明显的细胞脱离和细胞活力降低。值得注意的是，用抗Ro/SSA自身抗体处理的SGEC中纤连蛋白-6表达的降低与失巢凋亡细胞死亡的增加相对应。我们的观察结果表明，在SS观察到的病理过程中纤连蛋白-6失调，并提供了新的证据，即ECM环境的紊乱可能损害唾液腺的结构和功能。
