Abstract
Systemic sclerosis (SSc) is a rare, chronic, multisystem autoimmune disease clinically characterized by progressive fibrosis of the skin and internal organs. The basic mechanism appears to involve endothelial cell injury, overproduction of extracellular matrix proteins, and aberrant immune activation. So far, there have been a few attempts to find genetic biomarkers for monitoring disease activity or for correlation with certain symptoms. In order to reveal reliable biomarkers, we analyzed the expression of four genes representing three important signaling pathways, TLR7, TLR9, and JAK2-STAT3. Using RT-qPCR technique, we analyzed the expression of TLR7, TLR9, JAK2, and STAT3 genes in peripheral blood mononuclear cells of 50 SSc patients and 13 healthy individuals. We detected significant upregulation of TLR7 gene expression in a group of SSc patients compared to non-SSc group. Receiver operating characteristic (ROC) analysis showed that TLR7 expression efficiently discriminates SSc cases from healthy individuals. High TLR7 expression positively correlated with the late form of disease, active SSc, and the presence of digital ulcers. Decreased levels of TLR9 and JAK2 mRNA were found in the patient’s cohort in comparison to non-SSc individuals, but showed no correlation with specific clinical outcomes. The expression level of the STAT3 gene did not differ between the analyzed groups. This is the first study on the expression of TLR7, TLR9, and STAT3 genes in SSc patients. Our results show that TLR7, TLR9, and JAK2 genes are potential biomarkers for SSc. The results obtained in this study could contribute to better classification, monitoring, and outcome prediction of patients with SSc based on genetics.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Abraham DJ, Varga J (2005) Scleroderma: from cell and molecular mechanisms to disease models. Trends Immunol 26:587–595. https://doi.org/10.1016/j.it.2005.09.004
Affandi AJ, Radstake TR, Marut W (2015) Update on biomarkers in systemic sclerosis: tools for diagnosis and treatment. Semin Immunopathol 37:475–487. https://doi.org/10.1007/s00281-015-0506-4
Barnes J, Mayes MD (2012) Epidemiology of systemic sclerosis: incidence, prevalence, survival, risk factors, malignancy, and environmental triggers. Curr Opin Rheumatol 24:165–170. https://doi.org/10.1097/BOR.0b013e32834ff2e8
Begon E, Michel L, Flageul B, Beaudoin I, Jean-Louis F, Bachelez H, Dubertret L, Musette P (2007) Expression, subcellular localization and cytokinic modulation of toll-like receptors (TLRs) in normal human keratinocytes: TLR2 up-regulation in psoriatic skin. Eur J Dermatol 17:497–506. https://doi.org/10.1684/ejd.2007.0264
Blasius AL, Beutler B (2010) Intracellular toll-like receptors. Immunity 32:305–315. https://doi.org/10.1016/j.immuni.2010.03.012
Budczies J, Klauschen F, Sinn BV, Győrffy B, Schmitt WD, Darb-Esfahani S, Denkert C (2012) Cutoff Finder: a comprehensive and straightforward web application enabling rapid biomarker cutoff optimization. PLoS One 7:e51862. https://doi.org/10.1371/journal.pone.0051862
Celhar T, Fairhurst AM (2014) Toll-like receptors in systemic lupus erythematosus: potential for personalized treatment. Front Pharmacol 5:265. https://doi.org/10.3389/fphar.2014.00265
Chen K, See A, Shumack S (2003) Epidemiology and pathogenesis of scleroderma. Australas J Dermatol 44:1–9
Chifflot H, Fautrel B, Sordet C, Chatelus E, Sibilia J (2008) Incidence and prevalence of systemic sclerosis: a systematic literature review. Semin Arthritis Rheum 37:223–235. https://doi.org/10.1016/j.semarthrit.2007.05.003
Ciechomska M, Cant R, Finnigan J, van Laar JM, O’Reilly S (2013) Role of toll-like receptors in systemic sclerosis. Expert Rev Mol Med 15:e9. https://doi.org/10.1017/erm.2013.10
Diebold SS, Kaisho T, Hemmi H, Akira S, Reis e Sousa C (2004) Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA. Science 303:1529–1531. https://doi.org/10.1126/science.1093616
Fang F, Marangoni RG, Zhou X, Yang Y, Ye B, Shangguang A, Qin W, Wang W, Bhattacharyya S, Wei J, Tourtellotte WG, Varga J (2016) Toll-like receptor 9 signaling is augmented in systemic sclerosis and elicits transforming growth factor beta-dependent fibroblast activation. Arthritis Rheumatol 68:1989–2002. https://doi.org/10.1002/art.39655
Ferri C, Valentini G, Cozzi F, Sebastiani M, Michelassi C, La Montagna G, Bullo A, Cazzato M, Tirri E, Storino F, Giuggioli D, Cuomo G, Rosada M, Bombardieri S, Todesco S, Tirri G; Systemic Sclerosis Study Group of the Italian Society of Rheumatology (SIR-GSSSc) (2002) Systemic sclerosis: demographic, clinical, and serologic features and survival in 1,012 Italian patients. Medicine (Baltimore) 81:139–153
Haghikia A, Hoch M, Stapel B, Hilfiker-Kleiner D (2012) STAT3 regulation of and by microRNAs in development and disease. JAKSTAT 1:143–150. https://doi.org/10.4161/jkst.19573
Hamaguchi Y (2010) Autoantibody profiles in systemic sclerosis: predictive value for clinical evaluation and prognosis. J Dermatol 37:42–53. https://doi.org/10.1111/j.1346-8138.2009.00762.x
Heil F, Hemmi H, Hochrein H, Ampenberger F, Kirschning C, Akira S, Lipford G, Wagner H, Bauer S (2004) Species-specific recognition of single-stranded RNA via toll-like receptor 7 and 8. Science 303:1526–1529. https://doi.org/10.1126/science.1093620
Hemmi H, Takeuchi O, Kawai T, Kaisho T, Sato S, Sanjo H, Matsumoto M, Hoshino K, Wagner H, Takeda K, Akira S (2000) A Toll-like receptor recognizes bacterial DNA. Nature 408:740–745. https://doi.org/10.1038/35047123
Hemmi H, Kaisho T, Takeuchi O, Sato S, Sanjo H, Hoshino K, Horiuchi T, Tomizawa H, Takeda K, Akira S (2002) Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway. Nat Immunol 3:196–200. https://doi.org/10.1038/ni758
Ho YY, Lagares D, Tager AM, Kapoor M (2014) Fibrosis—a lethal component of systemic sclerosis. Nat Rev Rheumatol 10:390–402. https://doi.org/10.1038/nrrheum.2014.53
Huang QQ, Pope RM (2009) The role of toll-like receptors in rheumatoid arthritis. Curr Rheumatol Rep 11:357–364
Jimenez SA, Derk CT (2004) Following the molecular pathways toward an understanding of the pathogenesis of systemic sclerosis. Ann Intern Med 140:37–50
Johnson SR, Hinchcliff M, Asano Y (2016) Controversies: molecular vs. clinical systemic sclerosis classification. J Scleroderma Relat Disord 1:277–285
Kamimura D, Ishihara K, Hirano T (2003) IL-6 signal transduction and its physiological roles: the signal orchestration model. Rev Physiol Biochem Pharmacol 149:1–38. https://doi.org/10.1007/s10254-003-0012-2
Kralovics R, Passamonti F, Buser AS, Teo SS, Tiedt R, Passweg JR, Tichelli A, Cazzola M, Skoda RC (2005) A gain-of-function mutation of JAK2 in myeloproliferative disorders. N Engl J Med 352:1779–1790. https://doi.org/10.1056/NEJMoa051113
Lau CM, Broughton C, Tabor AS, Akira S, Flavell RA, Mamula MJ, Christensen SR, Shlomchik MJ, Viglianti GA, Rifkin IR, Marshak-Rothstein A (2005) RNA-associated autoantigens activate B cells by combined B cell antigen receptor/Toll-like receptor 7 engagement. J Exp Med 202:1171–1177. https://doi.org/10.1084/jem.20050630
LeRoy EC, Medsger TA Jr (2001) Criteria for the classification of early systemic sclerosis. J Rheumatol 28:1573–1576
Lövgren T, Eloranta ML, Båve U, Alm GV, Rönnblom L (2004) Induction of interferon-alpha production in plasmacytoid dendritic cells by immune complexes containing nucleic acid released by necrotic or late apoptotic cells and lupus IgG. Arthritis Rheum 50:1861–1872. https://doi.org/10.1002/art.20254
Lövgren T, Eloranta ML, Kastner B, Wahren-Herlenius M, Alm GV, Rönnblom L (2006) Induction of interferon-alpha by immune complexes or liposomes containing systemic lupus erythematosus autoantigen- and Sjögren’s syndrome autoantigen-associated RNA. Arthritis Rheum 54:1917–1927. https://doi.org/10.1002/art.21893
Lund J, Sato A, Akira S, Medzhitov R, Iwasaki A (2003) Toll-like receptor 9-mediated recognition of Herpes simplex virus-2 by plasmacytoid dendritic cells. J Exp Med 198:513–520. https://doi.org/10.1084/jem.20030162
Lund JM, Alexopoulou L, Sato A, Karow M, Adams NC, Gale NW, Iwasaki A, Flavell RA (2004) Recognition of single-stranded RNA viruses by Toll-like receptor 7. Proc Natl Acad Sci U S A 101:5598–5603. https://doi.org/10.1073/pnas.0400937101
Marshak-Rothstein A (2006) Toll-like receptors in systemic autoimmune disease. Nat Rev Immunol 6:823–835. https://doi.org/10.1038/nri1957
Meier FM, Frommer KW, Dinser R, Walker UA, Czirjak L, Denton CP, Allanore Y, Distler O, Riemekasten G, Valentini G, Müller-Ladner U; EUSTAR Co-authors (2012) Update on the profile of the EUSTAR cohort: an analysis of the EULAR Scleroderma Trials and Research group database. Ann Rheum Dis 71:1355–1360. https://doi.org/10.1136/annrheumdis-2011-200742
Miggin SM, O’Neill LA (2006) New insights into the regulation of TLR signaling. J Leukoc Biol 80:220–226. https://doi.org/10.1189/jlb.1105672
Nikpour M, Stevens WM, Herrick AL, Proudman SM (2010) Epidemiology of systemic sclerosis. Best Pract Res Clin Rheumatol 24:857–869. https://doi.org/10.1016/j.berh.2010.10.007
O’Reilly S, Ciechomska M, Cant R, Hügle T, van Laar JM (2012) Interleukin-6, its role in fibrosing conditions. Cytokine Growth Factor Rev 23:99–107. https://doi.org/10.1016/j.cytogfr.2012.04.003
Ospelt C, Brentano F, Rengel Y, Stanczyk J, Kolling C, Tak PP, Gay RE, Gay S, Kyburz D (2008) Overexpression of toll-like receptors 3 and 4 in synovial tissue from patients with early rheumatoid arthritis: toll-like receptor expression in early and longstanding arthritis. Arthritis Rheum 58:3684–3692. https://doi.org/10.1002/art.24140
Papadimitraki ED, Choulaki C, Koutala E, Bertsias G, Tsatsanis C, Gergianaki I, Raptopoulou A, Kritikos HD, Mamalaki C, Sidiropoulos P, Boumpas DT (2006) Expansion of toll-like receptor 9-expressing B cells in active systemic lupus erythematosus: implications for the induction and maintenance of the autoimmune process. Arthritis Rheum 54:3601–3611. https://doi.org/10.1002/art.22197
Pendergrass SA, Hayes E, Farina G, Lemaire R, Farber HW, Whitfield ML, Lafyatis R (2010) Limited systemic sclerosis patients with pulmonary arterial hypertension show biomarkers of inflammation and vascular injury. PLoS One 5:e12106. https://doi.org/10.1371/journal.pone.0012106
Raja J, Denton CP (2015) Cytokines in the immunopathology of systemic sclerosis. Semin Immunopathol 37:543–557. https://doi.org/10.1007/s00281-015-0511-7
Rawlings JS, Rosler KM, Harrison DA (2004) The JAK/STAT signaling pathway. J Cell Sci 117:1281–1283. https://doi.org/10.1242/jcs.00963
Rifkin IR, Leadbetter EA, Busconi L, Viglianti G, Marshak-Rothstein A (2005) Toll-like receptors, endogenous ligands, and systemic autoimmune disease. Immunol Rev 204:27–42. https://doi.org/10.1111/j.0105-2896.2005.00239.x
Steen VD (2008) The many faces of scleroderma. Rheum Dis Clin North Am 34:1–15. https://doi.org/10.1016/j.rdc.2007.12.001
Valentini G, Della Rossa A, Bombardieri S, Bencivelli W, Silman AJ, D’Angelo S, Cerinic MM, Belch JF, Black CM, Bruhlmann P, Czirják L, De Luca A, Drosos AA, Ferri C, Gabrielli A, Giacomelli R, Hayem G, Inanc M, McHugh NJ, Nielsen H, Rosada M, Scorza R, Stork J, Sysa A, van den Hoogen FH, Vlachoyiannopoulos PJ (2001) European multicentre study to define disease activity criteria for systemic sclerosis. II. Identification of disease activity variables and development of preliminary activity indexes. Ann Rheum Dis 60:592–598
Valentino L, Pierre J (2006) JAK/STAT signal transduction: regulators and implication in hematological malignancies. Biochem Pharmacol 71:713–721. https://doi.org/10.1016/j.bcp.2005.12.017
van den Hoogen F, Khanna D, Fransen J, Johnson SR, Baron M, Tyndall A, Matucci-Cerinic M, Naden RP, Medsger TA Jr, Carreira PE, Riemekasten G, Clements PJ, Denton CP, Distler O, Allanore Y, Furst DE, Gabrielli A, Mayes MD, van Laar JM, Seibold JR, Czirjak L, Steen VD, Inanc M, Kowal-Bielecka O, Müller-Ladner U, Valentini G, Veale DJ, Vonk MC, Walker UA, Chung L, Collier DH, Ellen Csuka M, Fessler BJ, Guiducci S, Herrick A, Hsu VM, Jimenez S, Kahaleh B, Merkel PA, Sierakowski S, Silver RM, Simms RW, Varga J, Pope JE (2013) 2013 classification criteria for systemic sclerosis: an American college of rheumatology/European league against rheumatism collaborative initiative. Ann Rheum Dis 72:1747–1755. https://doi.org/10.1136/annrheumdis-2013-204424
Varga J, Abraham D (2007) Systemic sclerosis: a prototypic multisystem fibrotic disorder. J Clin Invest 117:557–567. https://doi.org/10.1172/jci31139
Vollmer J, Tluk S, Schmitz C, Hamm S, Jurk M, Forsbach A, Akira S, Kelly KM, Reeves WH, Bauer S, Krieg AM (2005) Immune stimulation mediated by autoantigen binding sites within small nuclear RNAs involves toll-like receptors 7 and 8. J Exp Med 202:1575–1585. https://doi.org/10.1084/jem.20051696
Wang T, Niu G, Kortylewski M, Burdelya L, Shain K, Zhang S, Bhattacharya R, Gabrilovich D, Heller R, Coppola D, Dalton W, Jove R, Pardoll D, Yu H (2004) Regulation of the innate and adaptive immune responses by Stat-3 signaling in tumor cells. Nat Med 10:48–54. https://doi.org/10.1038/nm976
Wei J, Bhattacharyya S, Tourtellotte WG, Varga J (2011) Fibrosis in systemic sclerosis: emerging concepts and implications for targeted therapy. Autoimmun Rev 10:267–275. https://doi.org/10.1016/j.autrev.2010.09.015
Wu P, Wu J, Liu S, Han X, Lu J, Shi Y, Wang J, Lu L, Cao X (2008) TLR9/TLR7-triggered downregulation of BDCA2 expression on human plasmacytoid dendritic cells from healthy individuals and lupus patients. Clin Immunol 129:40–48. https://doi.org/10.1016/j.clim.2008.06.004
Yu H, Kortylewski M, Pardoll D (2007) Crosstalk between cancer and immune cells: role of STAT3 in the tumour microenvironment. Nat Rev Immunol 7:41–51. https://doi.org/10.1038/nri1995
Acknowledgements
This work was supported by the Ministry of Education, Science and Technological Development, Republic of Serbia (grant no. III41004).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors do not have any conflict of interest or financial support.
Ethical approval
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
Informed consent
Informed consent was obtained from all individual participants included in the study.
Additional information
Communicated by: Michal Witt
Electronic supplementary material
Below is the link to the electronic supplementary material.
ESM 1
(DOCX 13 kb)
Rights and permissions
About this article
Cite this article
Vreća, M., Zeković, A., Damjanov, N. et al. Expression of TLR7, TLR9, JAK2, and STAT3 genes in peripheral blood mononuclear cells from patients with systemic sclerosis. J Appl Genetics 59, 59–66 (2018). https://doi.org/10.1007/s13353-017-0415-4
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s13353-017-0415-4