Diagnostic salivary biomarkers in traumatic brain injury: narrative review

Document Type : Review


1 Medical Student, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran

2 Ph.D., Professor, Department of Biological Sciences, 111 Research Drive, Lehigh University, Bethlehem, PA 18015, USA

3 Professor of Cardiac Anesthesia, School of Medicine, Atherosclerosis Research Center, Baqiyatallah University of Medical sciences, Tehran, Iran


Traumatic brain injury (TBI) is a common cause of disability and mortality worldwide. TBI is an acquired brain injury that may be open (penetrating) or closed (non-penetrating) and is be categorized as mild, moderate, or severe, depending on the clinical presentation. Accurate diagnosis at the earliest stages can significantly affect patient discomfort, prognosis, therapeutic intervention, survival rates and recurrence. Whereas traditional CT and MRI techniques for diagnosis are dominant in clinical situations, a promising direction for clinical diagnosis is the use of fluid biomarkers like blood, CSF, urine, and saliva. Fluid biomarkers that may track these injuries and inflammatory processes have been explored for their potential to provide objective measures in TBI assessment. At present, there are limited clinical guidelines available regarding the use of fluid biomarkers in TBI.
In recent years, saliva has received significant attention as a biomarker for TBI in clinical practice due to the non-invasive accessibility, cost-effective collection, and consistent relationship with serum. This review examines the utility of saliva biomarkers such as S100B, noncoding RNAs (ncRNAs), extracellular vesicles (EVs), miRNAs levels, microtubule-associated protein tau, alpha-amylase, cortisol, and oxidative stress in TBI.
The study highlights the current state of salivary diagnostics, future aspirations, and their potential as the preferred route of TBI detection. The newly developed techniques for salivary analysis of these molecules may help to improve outcomes for TBI through rapid detection current unavailable with serum samples. Future studies via salivary biomarkers will help establish consistent strategies for early diagnosis of TBI and improve treatment outcomes of TBI patients.


  1. Rubiano AM, Carney N, Chesnut R, Puyana JC. Global neurotrauma research challenges and opportunities. Nature. 2015; 527:S193-S197. doi:10.1038/nature16035
  2. Cheng Y, Pereira M, Raukar N, Reagan JL, Queseneberry M, Goldberg L, et al. Potential biomarkers to detect traumatic brain injury by the profiling of salivary extracellular vesicles. J Cell Physiol. 2019; 234(8):14377-88. doi:10.1002/jcp.28139 PMid:30644102 PMCid:PMC6478516
  3. Monteleone MC, Billi SC, Viale L, Catoira NP, Frasch AC, Brocco MA. Search of brain-enriched proteins in salivary extracellular vesicles for their use as mental disease biomarkers: a pilot study of the neuronal glycoprotein M6a. J Affect Disord Rep. 2020;1:100003. doi:10.1016/j.jadr.2020.100003
  4. Courtney A, Courtney M. The complexity of biomechanics causing primary blast-induced traumatic brain injury: a review of potential mechanisms. Front Neurol. 2015;6:221. doi:10.3389/fneur.2015.00221
    PMid:26539158 PMCid:PMC4609847
  5. Helmick KM, Spells CA, Malik SZ, Davies CA, Marion DW, Hinds SR. Traumatic brain injury in the US military: epidemiology and key clinical and research programs. Brain Imaging Behav. 2015; 9(3):358-66. doi:10.1007/s11682-015-9399-z PMid:25972118
  6. Buttram SD, Garcia-Filion P, Miller J, Youssfi M, Brown SD, Dalton HJ, et al. Computed tomography vs magnetic resonance imaging for identifying acute lesions in pediatric traumatic brain injury. Hosp Pediatr. 2015;5(2):79-84. doi:10.1542/hpeds.2014-0094 PMid:25646200
  7. de Almeida PD, Gregio AM, Machado MA, De Lima AA, Azevedo LR. Saliva composition and functions: a comprehensive review. J Contemp Dent Pract. 2008;9(3):72-80. doi:10.5005/jcdp-9-3-72 PMid:18335122
  8. Tiwari M. Science behind human saliva. J Nat Sci Biol Med. 2011; 2(1):53. doi:10.4103/0976-9668.82322 PMid:22470235 PMCid:PMC3312700
  9. Munro CL, Grap MJ, Jablonski R, Boyle A. Oral health measurement in nursing research: state of the science. Biol Res Nurs. 2006;8(1):35-42. doi:10.1177/1099800406289343 PMid:16766627 PMCid:PMC2213421
  10. Altin KT, Topcuoglu N, Duman G, Unsal M, Celik A, Kuvvetli SS, et al. Antibacterial effects of saliva substitutes containing lysozyme or lactoferrin against Streptococcus mutans. Arch Oral Biol. 2021:105183. doi:10.1016/j.archoralbio.2021.105183 PMid:34091207
  11. Janigro D, Kawata K, Silverman E, Marchi N, Diaz-Arrastia R. Is salivary S100B a biomarker of traumatic brain injury? A pilot study. Front Neurol. 2020;11:528. doi:10.3389/fneur.2020.00528 PMid:32595592 PMCid:PMC7303321
  12. Menetski JP, Hoffmann SC, Cush SS, Kamphaus TN, Austin CP, Herrling PL, et al. The Foundation for the National Institutes of Health Biomarkers Consortium: past accomplishments and new strategic direction. Clin Pharmacol Ther. 2019; 105 (4):829-43. doi:10.1002/cpt.1362 PMid:30648736 PMCid:PMC6593617
  13. Michetti F, Bruschettini M, Frigiola A, Abella R, Giamberti A, Marchese N, et al. Saliva S100B in professional sportsmen: high levels at resting conditions and increased after vigorous physical activity. Clin Biochem. 2011;44(2-3):245-7. doi:10.1016/j.clinbiochem.2010.10.007 PMid:20970414
  14. Ostendorp T, Leclerc E, Galichet A, Koch M, Demling N, Weigle B, et al. Structural and functional insights into RAGE activation by multimeric S100B. EMBO J. 2007;26(16):3868-78. doi:10.1038/sj.emboj.7601805 PMid:17660747 PMCid:PMC1952220
  15. Koh SX, Lee JK. S100B as a marker for brain damage and blood-brain barrier disruption following exercise. Sports Med. 2014; 44 (3):369-85. doi:10.1007/s40279-013-0119-9 PMid:24194479
  16. Michetti F, D'Ambrosi N, Toesca A, Puglisi MA, Serrano A, Marchese E, et al. The S100B story: from biomarker to active factor in neural injury. J Neurochem. 2019;148(2):168-87. doi:10.1111/jnc.14574PMid:30144068
  17. Sone M, Hayashi T, Tarui H, Agata K, Takeichi M, Nakagawa S. The mRNA-like noncoding RNA Gomafucons titutes a novel nuclear domain in a subset of neurons. J Cell Sci. 2007;120 (15): 2498-506 doi:10.1242/jcs.009357 PMid:17623775
  18. Tano K, Mizuno R, Okada T, Rakwal R, Shibato J, Masuo Y, et al. MALAT‐1 enhances cell motility of lungadenocarcinoma cells by influencing the expression of motility‐related genes. FEBS Lett. 2010;584(22):4575-80 doi:10.1016/j.febslet.2010.10.008 PMid:20937273
  19. Mateen BA, Hill CS, Biddie SC, Menon DK. DNA methylation: basic biology and application to traumatic brain injury. J Neurotrauma. 2017;34(16):2379-88. doi:10.1089/neu.2017.5007 PMid:28482743
  20. DiStefano JK. The emerging role of long noncoding RNAs in human disease. Dis Gene Identif. 2018:91-110.
    doi:10.1007/978-1-4939-7471-9_6 PMid:29423795
  21. Fuller CW, Fuller GW, Kemp SP, Raftery M. Evaluation of World Rugby's concussion management process: results from Rugby World Cup 2015. Br J Sports Med. 2017;51(1):64-9. doi:10.1136/bjsports-2016-096461 PMid:27587799
  22. Xie BS, Wang YQ, Lin Y, Zhao CC, Mao Q, Feng JF, et al. Circular RNA expression profiles alter significantly after traumatic brain injury in rats. J Neurotrauma. 2018;35(14):1659-66. doi:10.1089/neu.2017.5468 PMid:29357736
  23. Fedorchak G, Rangnekar A, Onks C, Loeffert AC, Loeffert J, Olympia RP, et al. Saliva RNA biomarkers predict concussion duration and detect symptom recovery: a comparison with balance and cognitive testing. J Neurol. 2021:1-3. doi:10.1007/s00415-021-10566-x PMid:34028616 PMCid:PMC8505318
  24. Lee RC, Feinbaum RL, Ambros V. The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell. 1993;75(5):843-54. doi:10.1016/0092-8674(93)90529-Y
  25. Rodriguez A, Griffiths-Jones S, Ashurst JL, Bradley A. Identification of mammalian microRNA host genes and transcription units. Genome Res. 2004;14(10a):1902-10. doi:10.1101/gr.2722704 PMid:15364901 PMCid:PMC524413
  26. Adlakha YK, Saini N. Brain microRNAs and insights into biological functions and therapeutic potential of brain enriched miRNA-128. Mol Cancer. 2014;13(1):1-8. doi:10.1186/1476-4598-13-33 PMid:24555688 PMCid:PMC3936914
  27. Wittmann J, Jäck HM. Serum microRNAs as powerful cancer biomarkers. Biochim Biophys Acta Bioenerg. 2010;1806(2):200-7. doi:10.1016/j.bbcan.2010.07.002 PMid:20637263
  28. Hicks SD, Johnson J, Carney MC, Bramley H, Olympia RP, Loeffert AC, et al. Overlapping microRNA expression in saliva and cerebrospinal fluid accurately identifies pediatric traumatic brain injury. J Neurotrauma. 2018;35(1):64-72. doi:10.1089/neu.2017.5111 PMid:28762893 PMCid:PMC7227420
  29. LaRocca D, Barns S, Hicks SD, Brindle A, Williams J, Uhlig R, et al. Comparison of serum and saliva miRNAs for identification and characterization of mTBI in adult mixed martial arts fighters. PloS one. 2019;14(1):e0207785. doi:10.1371/journal.pone.0207785 PMid:30601825 PMCid:PMC6314626
  30. Camussi G, Deregibus MC, Bruno S, Cantaluppi V, Biancone L. Exosomes/microvesicles as a mechanism of cell-to-cell communication. Kidney Int. 2010;78(9):838-48. doi:10.1038/ki.2010.278 PMid:20703216
  31. Santiago-Dieppa DR, Steinberg J, Gonda D, Cheung VJ, Carter BS, Chen CC. Extracellular vesicles as a platform for 'liquid biopsy'in glioblastoma patients. Expert Rev Mol Diagn. 2014; 14 (7):819-25. doi:10.1586/14737159.2014.943193 PMid:25136839 PMCid:PMC4436244
  32. Hu T, Wolfram J, Srivastava S. Extracellular vesicles in cancer detection: hopes and hypes. Trends Cancer. 2020. doi:10.1016/j.trecan.2020.09.003 PMid:33008796
  33. Dabrowska S, Andrzejewska A, Janowski M, Lukomska B. Immunomodulatory and Regenerative Effects of Mesenchymal Stem Cells and Extracellular Vesicles: Therapeutic Outlook for Inflammatory and Degenerative Diseases. Front Immunol. 2021; 11:3809. doi:10.3389/fimmu.2020.591065 PMid:33613514 PMCid:PMC7893976
  34. Massa M, Croce S, Campanelli R, Abbà C, Lenta E, Valsecchi C, Avanzini MA. Clinical applications of mesenchymal stem/stromal cell derived extracellular vesicles: therapeutic potential of an acellular product. Diagnostics. 2020;10(12):999. doi:10.3390/diagnostics10120999
    PMid:33255416 PMCid:PMC7760121
  35. Cheng Y, Pereira M, Raukar NP, Reagan JL, Quesenberry M, Goldberg L, et al. Inflammation-related gene expression profiles of salivary extracellular vesicles in patients with head trauma. Neural Regen Res. 2020;15(4):676. doi:10.4103/1673-5374.266924 PMid:31638091 PMCid:PMC6975135
  36. Chiabotto G, Gai C, Deregibus MC, Camussi G. Salivary extracellular vesicle-associated exRNA as cancer biomarker. Cancers. 2019;11(7):891. doi:10.3390/cancers11070891
    PMid:31247906 PMCid:PMC6679099
  37. Matuk R, Pereira M, Baird J, Dooner M, Cheng Y, Wen S, et al. The role of salivary vesicles as a potential inflammatory biomarker to detect traumatic brain injury in mixed martial artists. Sci Rep. 2021 ;11(1). doi:10.1038/s41598-021-87180-4 PMid:33854105 PMCid:PMC8047010
  38. Spillantini MG, Goedert M. Tau pathology and neurodegeneration. Lancet Neurol. 2013;12(6):609-22. doi:10.1016/S1474-4422(13)70090-5
  39. Rubenstein R, Chang B, Davies P, Wagner AK, Robertson CS, Wang KK. A novel, ultrasensitive assay for tau: potential for assessing traumatic brain injury in tissues and biofluids. J Neurotrauma. 2015;32(5):342-52. doi:10.1089/neu.2014.3548 PMid:25177776 PMCid:PMC4348038
  40. Olczak M, Niderla-Bielińska J, Kwiatkowska M, Samojłowicz D, Tarka S, Wierzba-Bobrowicz T. Tau protein (MAPT) as a possible biochemical marker of traumatic brain injury in postmortem examination. Forensic Sci Int. 2017;280:1-7. doi:10.1016/j.forsciint.2017.09.008 PMid:28942078   
  41. Blomberg M, Jensen M, Basun H, Lannfelt L, Wahlund LO. Cerebrospinal fluid tau levels increase with age in healthy individuals. Dement Geriatr Cogn Disord. 2001;12(2):127-32. doi:10.1159/000051246 PMid:11173885
  42. Olczak M, Poniatowski ŁA, Niderla-Bielińska J, Kwiatkowska M, Chutorański D, Tarka S, et al. Concentration of microtubule associated protein tau (MAPT) in urine and saliva as a potential biomarker of traumatic brain injury in relationship with blood-brain barrier disruption in postmortem examination. Forensic Sci Int. 2019;301:28-36. doi:10.1016/j.forsciint.2019.05.010 PMid:31128406
  43. Jashnani KD, Kale SA, Rupani AB. Vitreous humor: biochemical constituents in estimation of postmortem interval. J Forensic Sci. 2010;55(6):1523-7. doi:10.1111/j.1556-4029.2010.01501.x PMid:20666922
  44. Farah R, Haraty H, Salame Z, Fares Y, Ojcius DM, Sadier NS. Salivary biomarkers for the diagnosis and monitoring of neurological diseases. Biomed J. 2018;41(2):63-87. doi:10.1016/j.bj.2018.03.004 PMid:29866603 PMCid:PMC6138769         
  45. Dadas A, Janigro D. The role and diagnostic significance of cellular barriers after concussive head trauma. Concussion. 2018;3(1):CNC53. doi:10.2217/cnc-2017-0019
    PMid:30202595 PMCid:PMC6093276
  46. Ali N, Nater UM. Salivary alpha-amylase as a biomarker of stress in behavioral medicine. Int J Behav Med. 2020;27(3):337-42. doi:10.1007/s12529-019-09843-x PMid:31900867 PMCid:PMC7250801
  47. Ewing-Cobbs L, Prasad MR, Cox Jr CS, Granger DA, Duque G, Swank PR. Altered stress system reactivity after pediatric injury: Relation with post-traumatic stress symptoms. Psychoneuroendocrinology. 2017;84:66-75. doi:10.1016/j.psyneuen.2017.06.003 PMid:28667938 PMCid:PMC5555029
  48. Yoon SA, Weierich MR. Salivary biomarkers of neural hypervigilance in trauma-exposed women. Psychoneuroendocrinology. 2016;63:17-25. doi:10.1016/j.psyneuen.2015.09.007 PMid:26398002 PMCid:PMC4695293
  49. Kirschbaum C, Hellhammer DH. Salivary cortisol. Encyclopedia of stress. 2000;3(379-383).
  50. Garde AH, Hansen ÅM. Long‐term stability of salivary cortisol. Scandinavian J Clin Lab Investig. 2005; 65(5):433-6. doi:10.1080/00365510510025773 PMid:16081365
  51. Dimopoulou I, Tsagarakis S, Theodorakopoulou M, Douka E, Zervou M, Kouyialis AT, et al. Endocrine abnormalities in critical care patients with moderate-to-severe head trauma: incidence, pattern and predisposing factors. Intensive Care Med. 2004;30(6):1051-7. doi:10.1007/s00134-004-2257-x PMid:15069597
  52. Apilux A, Rengpipat S, Suwanjang W, Chailapakul O. Development of competitive lateral flow immunoassay coupled with silver enhancement for simple and sensitive salivary cortisol detection. EXCLI J. 2018;17:1198.
  53. Ebrecht M, Hextall J, Kirtley LG, Taylor A, Dyson M, Weinman J. Perceived stress and cortisol levels predict speed of wound healing in healthy male adults. Psychoneuroendocrinology. 2004; 29 (6): 798-809. doi:10.1016/S0306-4530(03)00144-6
  54. Hanafy KA, Selim MH. Antioxidant strategies in neurocritical care. Neurotherapeutics. 2012;9(1):44-55. doi:10.1007/s13311-011-0085-6 PMid:22135010 PMCid:PMC3271156
  55. Yoshikawa T, Naito Y. What is oxidative stress?. Japan med Assoc J. 2002;45(7):271-6.
  56. Tavazzi B, Signoretti S, Lazzarino G, Amorini AM, Delfini R, Cimatti M, et al. Cerebral oxidative stress and depression of energy metabolism correlate with severity of diffuse brain injury in rats. Neurosurgery. 2005;56(3):582-9. doi:10.1227/01.NEU.0000156715.04900.E6 PMid:15730584
  57. Al Nimer F, Ström M, Lindblom R, Aeinehband S, Bellander BM, Nyengaard JR, et al. Naturally occurring variation in the glutathione-S-transferase 4 gene determines neurodegeneration after traumatic brain injury. Antioxid Redox Signal. 2013; 18(7): 784-94. doi:10.1089/ars.2011.4440 PMid:22881716 PMCid:PMC3555113
  58. Gümüş P, Emingil G, Öztürk VÖ, Belibasakis GN, Bostanci N. Oxidative stress markers in saliva and periodontal disease status: modulation during pregnancy and postpartum. BMC Infect Dis. 2015;15(1):1-9. doi:10.1186/s12879-015-1003-z PMid:26152310 PMCid:PMC4495776
  59. Khurshid Z, Zafar MS, Khan RS, Najeeb S, Slowey PD, Rehman IU. Role of salivary biomarkers in oral cancer detection. Adv Clin Chem. 2018;86:23-70. doi:10.1016/bs.acc.2018.05.002 PMid:30144841
  60. Walt DR, Blicharz TM, Hayman RB, Rissin DM, Bowden M, Siqueira WL, et al. Microsensor arrays for saliva diagnostics. Ann N Y Acad Sci. 2007;1098(1):389. doi:10.1196/annals.1384.031 PMid:17435144 PMCid:PMC7168095
  61. McMahon PJ, Panczykowski DM, Yue JK, Puccio AM, Inoue T, Sorani MD, et al. Measurement of the glial fibrillary acidic protein and its breakdown products GFAP-BDP biomarker for the detection of traumatic brain injury compared to computed tomography and magnetic resonance imaging. J Neurotrauma. 2015;32 (8):527-33. doi:10.1089/neu.2014.3635 PMid:25264814 PMCid:PMC4394160
  62. Mondello S, Palmio J, Streeter J, Hayes RL, Peltola J, Jeromin A. Ubiquitin carboxy-terminal hydrolase L1 (UCH-L1) is increased in cerebrospinal fluid and plasma of patients after epileptic seizure. BMC Neurol. 2012;12(1):1-7. doi:10.1186/1471-2377-12-85 PMid:22931063 PMCid:PMC3500207
  63. Mayo S, Benito-León J, Peña-Bautista C, Baquero M, Cháfer-Pericás C. Recent Evidence in Epigenomics and Proteomics Biomarkers for Early and Minimally Invasive Diagnosis of Alzheimer's and Parkinson's Diseases. Curr Neuropharmacol. 2021; 19 (8): 1273-303. doi:10.2174/1570159X19666201223154009 PMid:33357195 PMCid:PMC8719284