{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,24]],"date-time":"2026-06-24T11:46:53Z","timestamp":1782301613837,"version":"3.54.5"},"reference-count":60,"publisher":"Wiley","issue":"2","license":[{"start":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T00:00:00Z","timestamp":1774569600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T00:00:00Z","timestamp":1774569600000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["JOR Spine"],"published-print":{"date-parts":[[2026,6]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Intervertebral disc degeneration (IDD) involves complex inflammatory and oxidative stress pathways within nucleus pulposus (NP) cells, leading to catabolic remodeling and loss of disc function. Traditional single\u2010analyte assays capture only a small part of these coordinated networks. Leveraging the multiplex capability of Luminex, this study uses targeted immunoassays to simultaneously quantify 73 extracellular proteins and 21 intracellular signaling molecules in primary human NP cells stimulated with IL\u20101\u03b2, Pam2CSK4, or hydrogen peroxide, enabling comparison of shared and stimulus\u2010specific responses.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Human NP cells isolated from patients undergoing spine surgery were plated in monolayer or encapsulated in alginate and stimulated for 1\u2009week with either: 100\u2009pg\/mL or 1\u2009ng\/mL Interleukin (IL)\u20101\u03b2, or the Toll\u2010Like Receptor (TLR) 2\/6 agonist Pam2CSK4 at 100\u2009ng\/mL, or 50\u2009\u03bcM Hydrogen Peroxide (H\n                      <jats:sub>2<\/jats:sub>\n                      O\n                      <jats:sub>2<\/jats:sub>\n                      ). The role of the stimuli was investigated using mRNA expression analysis and Luminex multiplex immunoassays.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      TLR2\/6 agonist and IL\u20101\u03b2 stimulation significantly up\u2010regulated IL\u20106, IL\u20108, IL\u20101\u03b2, and MMP\u20103 mRNA expression, while H\n                      <jats:sub>2<\/jats:sub>\n                      O\n                      <jats:sub>2<\/jats:sub>\n                      failed to induce any changes. The secretome showed that IL\u20108, IL\u20106, MMP\u20107, Intercellular Adhesion Molecule 1 (ICAM\u20101), and Growth\u2010Regulated Oncogene\u2010\u03b1 (GRO\u2010\u03b1) were the most responsive proteins across treatments and culture conditions. Intracellularly, phosphorylation of Jun Proto\u2010Oncogene AP\u20101 Transcription Factor Subunit (cJUN), Nuclear Factor Kappa B (NF\u2010kB), and Inhibitor of Nuclear Factor Kappa B Alpha (IKBA) was observed following IL\u20101\u03b2 and Pam2CSK4 stimulation.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>Across inflammatory and innate immune activation and under both monolayer and 3D culture, a concise five\u2010marker panel, namely IL\u20106, IL\u20108, GRO\u2010\u03b1 (CXCL\u20101), ICAM\u20101, and MMP\u20107 captured NP cell responses with high consistency, while phosphoproteomics demonstrated convergence on NF\u2010\u03baB\/AP\u20101 signaling. These markers are well established within disc\u2010degeneration pathways (inflammation, innate immunity, and ECM remodeling); thus, they offer practical, reusable readouts for comparative pharmacology, assay qualification, and early biomarker development in translational spine research.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1002\/jsp2.70165","type":"journal-article","created":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T10:26:22Z","timestamp":1774607182000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Secretome and Pathway Analysis of Stress Induced Disc Degeneration In\u00a0Vitro"],"prefix":"10.1002","volume":"9","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2059-1480","authenticated-orcid":false,"given":"Exarchos","family":"Kanelis","sequence":"first","affiliation":[{"name":"Protavio, Demokritos Technology Park  Agia Paraskevi Greece"},{"name":"Biomedical Systems Laboratory, School of Mechanical Engineering National Technical University of Athens  Athens Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0962-8705","authenticated-orcid":false,"given":"Andrea","family":"N\u00fcesch","sequence":"additional","affiliation":[{"name":"Division of Clinical Medicine, School of Medicine and Population Health University of Sheffield  Sheffield UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0200-4596","authenticated-orcid":false,"given":"Joseph","family":"Snuggs","sequence":"additional","affiliation":[{"name":"Division of Clinical Medicine, School of Medicine and Population Health University of Sheffield  Sheffield UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4489-7107","authenticated-orcid":false,"given":"Christine Lyn","family":"Le Maitre","sequence":"additional","affiliation":[{"name":"Division of Clinical Medicine, School of Medicine and Population Health University of Sheffield  Sheffield UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0425-166X","authenticated-orcid":false,"given":"Leonidas G.","family":"Alexopoulos","sequence":"additional","affiliation":[{"name":"Protavio, Demokritos Technology Park  Agia Paraskevi Greece"},{"name":"Biomedical Systems Laboratory, School of Mechanical Engineering National Technical University of Athens  Athens Greece"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2026,3,27]]},"reference":[{"key":"e_1_2_12_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0140-6736(18)32279-7"},{"key":"e_1_2_12_3_1","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2022.924692"},{"key":"e_1_2_12_4_1","doi-asserted-by":"publisher","DOI":"10.3390\/ijms24010208"},{"key":"e_1_2_12_5_1","doi-asserted-by":"crossref","unstructured":"S.Roberts H.Evans J.Trivedi andJ.Menage \u201cHistology and Pathology of the Human Intervertebral Disc \u201d 2006.","DOI":"10.2106\/00004623-200604002-00003"},{"key":"e_1_2_12_6_1","doi-asserted-by":"publisher","DOI":"10.1097\/01.brs.0000231761.73859.2c"},{"key":"e_1_2_12_7_1","doi-asserted-by":"publisher","DOI":"10.1038\/nrrheum.2013.160"},{"key":"e_1_2_12_8_1","doi-asserted-by":"crossref","unstructured":"K. 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