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Toll-like Receptor in Salivary Glands from Patients with Sjögren's Syndrome: Functional Analysis by Human Salivary Gland Cell Line
ATSUSHI KAWAKAMI, KOTO NAKASHIMA, MAMI TAMAI, HIDEKI NAKAMURA, NOZOMI IWANAGA, KEITA FUJIKAWA, TOSHIYUKI ARAMAKI, KAZUHIKO ARIMA, NAOKI IWAMOTO, KUNIHIRO ICHINOSE, MAKOTO KAMACHI, HIROAKI IDA, TOMOKI ORIGUCHI, and KATSUMI EGUCHI ABSTRACT. Objective. We investigated expression of toll-like receptor (TLR) in labial salivary glands of patients with Sjögren's syndrome (SS) and functional TLR expression in the cultured salivary gland cell line. Methods. Expression of TLR2, TLR3, TLR4, and myeloid differentiation factor 88 (MyD88) in labial salivary glands was examined by immunohistochemistry. Human salivary gland (HSG) cell-line cells were cultured with TLR ligands [peptidoglycan, poly (I:C) and lipopolysaccharide], and CD54 expression and interleukin 6 (IL-6) production was studied. Phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, and Akt was examined by Western blotting. Activation of nuclear factor-kB (NF-kB) p65 in HSG cells was studied by NF-kB p65 nuclear translocation by microscopic immunofluorescence or chemiluminescent electrophoretic mobility shift assay and detection of NF-kB p65 phosphorylation. Results. TLR2, TLR3, TLR4, and MyD88 were more strongly expressed in the labial salivary glands of SS patients (n =12) than in control subjects (n = 4), and were found in salivary-infiltrating mononuclear cells as well as acinar cells and ductal epithelial cells. In cultured HSG cells, a similar expression pattern was observed, and TLR ligands stimulated CD54 expression and IL-6 production. TLR ligands induced phosphorylation of ERK, JNK, and p38 in HSG cells, but not Akt phosphorylation or activation of NF-kB p65. Conclusion. Although the putative ligands remain to be determined, our study indicated the activation of the TLR-mediated immune response in SS, and suggested that the TLR effect is mediated through the mitogen-activated protein kinase pathway. (First Release April 1 2007; J Rheumatol 2007;34:1019–26) Key Indexing Terms: SJÖGREN'S SYNDROME From the First Department of Internal Medicine, Graduate School of Biomedical Sciences, Nagasaki University; and Nagasaki University of Health Sciences, Nagasaki University, Nagasaki, Japan. Dr. Kawakami and Dr. Nakashima contributed equally to this report. A. Kawakami, MD, PhD; K. Nakashima, MD, PhD; M. Tamai, MD, PhD; H. Nakamura, MD, PhD; N. Iwanaga, MD, PhD; K. Fujikawa, MD; T. Aramaki, MD; K. Arima, MD, PhD; N. Iwamoto, MD; K. Ichinose, MD; M. Kamachi, MD, PhD; H. Ida, MD, PhD, First Department of Internal Medicine, Graduate School of Biomedical Sciences; T. Origuchi, MD, PhD, Nagasaki University of Health Sciences; K. Eguchi, MD, PhD, First Department of Internal Medicine, Graduate School of Biomedical Sciences. Address reprint requests to Dr. A. Kawakami, First Department of Internal Medicine, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8501, Japan. E-mail: atsushik@net.nagasaki-u.ac.jp Accepted for publication January 22, 2007.
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