{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,24]],"date-time":"2026-08-24T20:04:17Z","timestamp":1787601857058,"version":"build-2736575974"},"reference-count":41,"publisher":"Public Library of Science (PLoS)","issue":"1","license":[{"start":{"date-parts":[[2024,1,26]],"date-time":"2024-01-26T00:00:00Z","timestamp":1706227200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:sec id=\"sec001\">\n<jats:title>Purpose<\/jats:title>\n<jats:p>We endeavored to identify objective salivary biomarkers for pain, a subjective sensation with a biological basis, using molecules already described related to pain. The study aimed to analyze inter-individual differences and intersession variability in salivary potential ocular pain biomarkers on healthy subjects, in samples obtained under the influence of controlled environmental conditions.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec002\">\n<jats:title>Methods<\/jats:title>\n<jats:p>Thirty-four healthy subjects, 20 male, 14 female, median age 35.44 years (range 30\u201340) were exposed for 30 minutes under standard environmental conditions (T: 22\u00b0C, 50% relative humidity) in the Controlled Environmental Research Laboratory (CE-Lab, Vision R&amp;D, Valladolid Spain) in two separate visits (V1, V2) at least 24 hours apart. Saliva was collected after the exposure in each of the visits, and cortisol, \u03b1-amylase (sAA), secretory IgA (sIgA), testosterone, and soluble fraction of TNF\u03b1 receptor II (sTNF\u03b1RII) were analyzed by ELISA. Repeatability of inter-subject inter-session measurements was assayed by intraclass correlation coefficient (ICC).<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec003\">\n<jats:title>Results<\/jats:title>\n<jats:p>There were no significant inter-session differences in testosterone (p = 0.2497), sTNF\u03b1RII (p = 0.6451) and sIgA (p = 0.9689) salivary levels. The reproducibility for salivary cortisol, sAA, testosterone, sTNF\u03b1RII and sIgA were 0.98 ng\/ml, 20.58 U\/ml, 21.07 \u03bcg\/ml, 24.68 pg\/ml and 0.19 pg\/ml, respectively. Salivary cortisol, sAA, testosterone, sTNF\u03b1RII and sIgA yielded the following ICCs: 0.506, 0.569, 0.824, 0.870 and 0.4295, respectively; all these ICCs (except that for cortisol and sIgA) were found to be improved compared to those found previously by our group in a previous study in salivary samples obtained from healthy subjects under non-controlled environmental conditions; Cortisol\u00b4s ICC didn\u00b4t improve and was in both cases at the limit of acceptability.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec004\">\n<jats:title>Conclusion<\/jats:title>\n<jats:p>Environmental factors such as temperature and relative humidity affect the reproducibility of measurement of some salivary molecules which have been proposed as potential pain biomarkers. The exposure of subjects to standard controlled environmental conditions before salivary sample obtention would improve the reproducibility of these molecule measures\u2019 as potential biomarkers of chronic ocular pain.<\/jats:p>\n<\/jats:sec>","DOI":"10.1371\/journal.pone.0296764","type":"journal-article","created":{"date-parts":[[2024,1,26]],"date-time":"2024-01-26T18:29:32Z","timestamp":1706293772000},"page":"e0296764","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":2,"title":["Influence of controlled environmental conditions in potential salivary ocular pain biomarkers for enhancing the assessment of ocular pain"],"prefix":"10.1371","volume":"19","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2712-8616","authenticated-orcid":true,"given":"Eva M.","family":"Sobas","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"V\u00e1zquez","family":"Amanda","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5077-4448","authenticated-orcid":true,"given":"Itziar","family":"Fern\u00e1ndez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Roberto","family":"Reinoso","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carmen","family":"Garc\u00eda-V\u00e1zquez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Enrique","family":"Ortega","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amalia","family":"Enr\u00edquez-de-Salamanca","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2024,1,26]]},"reference":[{"key":"pone.0296764.ref001","doi-asserted-by":"crossref","first-page":"1865","DOI":"10.1016\/S0140-6736(99)04088-X","article-title":"Management of chronic pain","volume":"353","author":"MA Ashburn","year":"1999","journal-title":"Lancet Lond Engl"},{"key":"pone.0296764.ref002","doi-asserted-by":"crossref","first-page":"3220","DOI":"10.1038\/s41598-018-21131-4","article-title":"Saliva as a medium to detect and measure biomarkers related to pain","volume":"8","author":"H Jasim","year":"2018","journal-title":"Sci Rep"},{"key":"pone.0296764.ref003","doi-asserted-by":"crossref","first-page":"348","DOI":"10.1016\/S0049-0172(98)80014-0","article-title":"A possible role for saliva as a diagnostic fluid in patients with chronic pain","volume":"27","author":"HP Fischer","year":"1998","journal-title":"Semin Arthritis Rheum"},{"key":"pone.0296764.ref004","volume-title":"BEST (Biomarkers, EndpointS, and other Tools) Resource.","author":"FDA-NIH Biomarker Working Group","year":"2016"},{"issue":"9","key":"pone.0296764.ref005","first-page":"23.1","article-title":"Biomarkers-A General Review.","volume":"76","author":"JK Aronson","year":"2017","journal-title":"Curr Protoc Pharmacol"},{"key":"pone.0296764.ref006","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1016\/j.cden.2010.08.004","article-title":"Saliva as a diagnostic fluid.","volume":"55","author":"D. 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Tes\u00f3n","year":"2013","journal-title":"Invest Ophthalmol Vis Sci"},{"key":"pone.0296764.ref010","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1016\/j.jtos.2019.03.009","article-title":"Response profiles to a controlled adverse desiccating environment based on clinical and tear molecule changes.","volume":"17","author":"I Fern\u00e1ndez","year":"2019","journal-title":"Ocul Surf"},{"key":"pone.0296764.ref011","doi-asserted-by":"crossref","first-page":"e0166976","DOI":"10.1371\/journal.pone.0166976","article-title":"Reliability of Potential Pain Biomarkers in the Saliva of Healthy Subjects: Inter-Individual Differences and Intersession Variability.","volume":"11","author":"EM Sobas","year":"2016","journal-title":"PloS One"},{"key":"pone.0296764.ref012","article-title":"Saliva sample results in healthy subjects in the Controlled Environmental Research Laboratory","author":"EM Sobas","year":"2023","journal-title":"Figshare_My 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Epidemiol."},{"key":"pone.0296764.ref017","doi-asserted-by":"crossref","first-page":"744","DOI":"10.1136\/bmj.313.7059.744","article-title":"Measurement error.","volume":"313","author":"JM Bland","year":"1996","journal-title":"BMJ"},{"key":"pone.0296764.ref018","doi-asserted-by":"crossref","first-page":"106","DOI":"10.1136\/bmj.313.7049.106","article-title":"Measurement error proportional to the mean","volume":"313","author":"JM Bland","year":"1996","journal-title":"BMJ"},{"key":"pone.0296764.ref019","doi-asserted-by":"crossref","first-page":"2256","DOI":"10.1016\/j.joms.2012.03.005","article-title":"Utility of a salivary biosensor for objective assessment of surgery-related stress.","volume":"70","author":"TF Robles","year":"2012","journal-title":"J Oral Maxillofac Surg Off J Am Assoc Oral Maxillofac Surg"},{"key":"pone.0296764.ref020","doi-asserted-by":"crossref","first-page":"486","DOI":"10.1016\/j.psyneuen.2009.01.014","article-title":"Salivary alpha-amylase as a non-invasive biomarker for the sympathetic nervous system: current state of research.","volume":"34","author":"UM Nater","year":"2009","journal-title":"Psychoneuroendocrinology"},{"key":"pone.0296764.ref021","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1039\/C7AN90101A","article-title":"Correction: Chemical analysis in saliva and the search for salivary biomarkers\u2014a tutorial review.","volume":"143","author":"K Ngamchuea","year":"2018","journal-title":"The Analyst"},{"key":"pone.0296764.ref022","doi-asserted-by":"crossref","first-page":"2142","DOI":"10.1097\/j.pain.0000000000001305","article-title":"Depression in chronic pain: might opioids be responsible?","volume":"159","author":"G Mazereeuw","year":"2018","journal-title":"Pain"},{"key":"pone.0296764.ref023","doi-asserted-by":"crossref","first-page":"449","DOI":"10.1016\/j.psyneuen.2010.12.019","article-title":"\u03b1-Amylase as a reliable and convenient measure of sympathetic activity: don\u2019t start salivating just 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Bartley","year":"2015","journal-title":"Clin J Pain"},{"key":"pone.0296764.ref027","doi-asserted-by":"crossref","first-page":"231","DOI":"10.1007\/s10508-013-0123-z","article-title":"Measurement of testosterone in human sexuality research: methodological considerations.","volume":"43","author":"SM van Anders","year":"2014","journal-title":"Arch Sex Behav"},{"key":"pone.0296764.ref028","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1580\/07-WEME-OR-123R2.1","article-title":"Effects of cold environment exposure and cold acclimatization on exercise-induced salivary cortisol response.","volume":"20","author":"S Izawa","year":"2009","journal-title":"Wilderness Environ Med."},{"key":"pone.0296764.ref029","doi-asserted-by":"crossref","first-page":"81","DOI":"10.1580\/PR30-04.1","article-title":"Autonomic nervous system and adrenal response to cold in man at Antarctica.","volume":"16","author":"K Harinath","year":"2005","journal-title":"Wilderness Environ Med"},{"key":"pone.0296764.ref030","doi-asserted-by":"crossref","first-page":"127","DOI":"10.1007\/BF00964127","article-title":"Thyroid function during a prolonged stay in Antarctica.","volume":"72","author":"RC Sawhney","year":"1995","journal-title":"Eur J Appl Physiol"},{"key":"pone.0296764.ref031","doi-asserted-by":"crossref","first-page":"118","DOI":"10.1111\/j.1469-8986.2011.01280.x","article-title":"Salivary cortisol and soluble tumor necrosis factor-\u03b1 receptor II responses to multiple experimental modalities of acute pain","volume":"49","author":"BR Goodin","year":"2012","journal-title":"Psychophysiology"},{"key":"pone.0296764.ref032","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1097\/AJP.0b013e31822cf542","article-title":"Enhanced cortisol increase upon awakening is associated with greater pain ratings but not salivary cortisol or soluble tumor necrosis factor-\u03b1 receptor II responses to acute pain","volume":"28","author":"BR Goodin","year":"2012","journal-title":"Clin J Pain"},{"key":"pone.0296764.ref033","doi-asserted-by":"crossref","first-page":"17","DOI":"10.2165\/00007256-199826010-00002","article-title":"Saliva composition and exercise.","volume":"26","author":"JL Chicharro","year":"1998","journal-title":"Sports Med Auckl NZ"},{"key":"pone.0296764.ref034","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1016\/S0165-5728(02)00466-6","article-title":"Salivary secretion of immunoglobulin A by submandibular glands in response to autonomimetic infusions in anaesthetised rats","volume":"136","author":"GB Proctor","year":"2003","journal-title":"J Neuroimmunol"},{"key":"pone.0296764.ref035","doi-asserted-by":"crossref","first-page":"1179","DOI":"10.1007\/s00421-018-3847-6","article-title":"Exercise intensity and its impact on relationships between salivary immunoglobulin A, saliva flow rate and plasma cortisol concentration","volume":"118","author":"CA Leicht","year":"2018","journal-title":"Eur J Appl Physiol"},{"key":"pone.0296764.ref036","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1016\/j.physbeh.2010.05.016","article-title":"Salivary flow and alpha-amylase: collection technique, duration, and oral fluid type","volume":"101","author":"EK Beltzer","year":"2010","journal-title":"Physiol Behav"},{"key":"pone.0296764.ref037","doi-asserted-by":"crossref","first-page":"1744","DOI":"10.12688\/f1000research.20099.1","article-title":"Salivary nitric oxide, Simplified Oral Hygiene Index, and salivary flow rate in smokers and non-smokers: a cross-sectional study.","volume":"8","author":"EW Bachtiar","year":"2019","journal-title":"F1000Research"},{"key":"pone.0296764.ref038","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1097\/IIO.0000000000000168","article-title":"Effects of the External Environment on Dry Eye Disease.","volume":"57","author":"M Calonge","year":"2017","journal-title":"Int Ophthalmol Clin."},{"key":"pone.0296764.ref039","doi-asserted-by":"crossref","first-page":"445","DOI":"10.1080\/02713683.2017.1420197","article-title":"Controlled Adverse Environment Chambers in Dry Eye Research","volume":"43","author":"M Calonge","year":"2018","journal-title":"Curr Eye Res"},{"key":"pone.0296764.ref040","first-page":"331","article-title":"Heat acclimation does not negatively affect salivary immunoglobulin-A and self-reported illness symptoms and wellness in recreational athletes.","volume":"9","author":"P Alkemade","journal-title":"Temp Multidiscip Biomed J"},{"key":"pone.0296764.ref041","doi-asserted-by":"crossref","first-page":"143","DOI":"10.1016\/S1542-0124(12)70196-9","article-title":"Methodologies for the study of ocular surface disease.","volume":"3","author":"GW Ousler","year":"2005","journal-title":"Ocul Surf."}],"container-title":["PLOS 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