Neuroprotective effects of honey against traumatic brain injury-induced anxiety and motor function impairment in Wistar rats
DOI:
https://doi.org/10.31117/neuroscirn.v7i4.360Keywords:
Anxiety, Antioxidant, Honey, Traumatic brain injury, Motor functionAbstract
Traumatic brain injury (TBI) poses a significant risk to neurological function, primarily attributable to oxidative stress. Our hypothesis suggests that honey, renowned for its high phenol and flavonoid content, exerts neuroprotective effects by mitigating oxidative stress. This study aims to assess honey's potential as an innovative therapeutic agent in a TBI rat model, primarily focusing on its impact on behaviour, lipid peroxidation, and antioxidant enzyme activity. A total of twenty adult male Wistar rats were randomly divided into four groups: Group A (control), Group B (honey-treated), Group C (TBI-induced), and Group D (honey-treated TBI). We conducted comprehensive assessments using the Rotarod and Elevated Plus Maze tests to evaluate behaviour. Additionally, biochemical evaluations included quantifying lipid peroxidation levels and conducting a detailed analysis of antioxidant enzyme status, specifically emphasising the activity of glutathione (GSH) and catalase (CAT). Our findings indicated that the TBI model rats displayed impaired motor coordination, heightened anxiety-like behaviour, and elevated lipid peroxidation levels. Intriguingly, the honey treatment effectively reversed these behavioural deficits while concurrently reducing lipid peroxidation. Notably, honey treatment significantly augmented the activity of antioxidant enzymes (CAT and GSH); there was a significant increase in CAT activity compared to GSH activity in the treated TBI model. This study supports our hypothesis, demonstrating that honey is a potent neuroprotective agent that counteracts TBI-induced oxidative stress. Our investigation elucidates honey's capacity to alleviate neurobehavioral impairments and mitigate oxidative damage within a TBI model. These results underscore honey's significance as an innovative and promising therapeutic approach in TBI management, emphasizing its potential to enhance neurological outcomes and improve the overall well-being of individuals affected by TBI.
References
Abdul‐Muneer, P. M., Chandra, N., & Haorah, J. (2015). Interactions of oxidative stress and neurovascular inflammation in the pathogenesis of traumatic brain injury. Molecular Neurobiology, 51(2), 966–979. https://doi.org/10.1007/s12035-014-8752-3
Ahmad, R. S., Hussain, M. B., Saeed, F., Waheed, M., & Tufail, T. (2017). Phytochemistry, metabolism, and ethnomedical scenario of honey: A concurrent review. International Journal of Food Properties, 20(sup1), S254–S269. https://doi.org/10.1080/10942912.2017.1295257
Akanmu, M. A., Olowookere, T. A., Atunwa, S. A., et al. (2011). Neuropharmacological effects of Nigerian honey in mice. African Journal of Traditional, Complementary and Alternative Medicines, 8(3), 230–249. https://doi.org/10.4314/ajtcam.v8i3.65285
Al-Kader, D. A., Onyechi, C. I., Ikedum, I. V., Fattah, A., Zafar, S., Bhat, S., Malik, M. A., Bheesham, N., Qadar, L. T., & Sajjad Cheema, M. (2022). Depression and Anxiety in Patients with a History of Traumatic Brain Injury: A Case-Control Study. Cureus, 14(8), e27971. https://doi.org/10.7759/cureus.27971
Albani, S., Andrawis, M., Abella, R., Fulgham, J., Vafamand, N., & Dumas, T. (2015). Behaviour in the Elevated Plus Maze Is Differentially Affected by Testing Conditions in Rats Under and Over Three Weeks of Age. Frontiers in Behavioral Neuroscience, 9, 31. https://doi.org/10.3389/fnbeh.2015.00031
Ali, A. M., & Kunugi, H. (2019). Bee honey protects astrocytes against oxidative stress: A preliminary in vitro investigation. Neuropsychopharmacology Reports, 39, 312–314. https://doi.org/10.1002/npr2.12079
Ali, A. M., & Hendawy, A. O. (2018a). So antidepressant drugs have serious adverse effects, but what are the alternatives? Novel Approaches in Drug Designing & Development, 4(3), 555636. https://doi.org/10.19080/NAPDD.2018.04.555636
Ali, A. M., & Hendawy, A. O. (2018b). Bee honey as a potentially effective treatment for depression: A review of clinical and preclinical findings. JOJ Nurse and Healthcare, 9(2), 555764. http://dx.doi.org/10.19080/JOJNHC.2018.09.555764
Ansari, M. A., Roberts, K. N., & Scheff, S. W. (2008). A time course of contusion‐induced oxidative stress and synaptic proteins in cortex in a rat model of TBI. Journal of Neurotrauma, 25(5), 513–526. https://doi.org/10.1089/neu.2007.0451
Assefa, B. T., Gebre, A. K., & Altaye, B. M. (2018). Reactive Astrocytes as Drug Target in Alzheimer’s Disease. BioMed Research International, 2018, 4160247. https://doi.org/10.1155/2018/4160247
Azman, K. F., Zakaria, R., Abdul Aziz, C. B., & Othman, Z. (2019). Tualang Honey Exerts Antidepressant-like Effects and Antioxidant Properties in Stress-exposed Rats. Malaysian Journal of Applied Sciences, 4(1), 15–25.
Becker, R. E., Kapogiannis, D., & Greig, N. H. (2018). Does traumatic brain injury hold the key to the Alzheimer's disease puzzle? Alzheimer's & Dementia, 14(4), 431–443. https://doi.org/10.1016/j.jalz.2017.11.007
Biedermann, S. V., Biedermann, D. G., Wenzlaff, F., Kurjack, T., & Nouri, S. A. (2017). An elevated plus maze in mixed reality for studying human anxiety related behavior. Behavioral Neuroscience, 15(1), 125. https://doi.org/10.1186/s12915-017-0463-6
Boyko, M., Gruenbaum, B. F., Shelef, I., Zvenigorodsky, V., Severynovska, O., Binyamin, Y., Knyazer, B., Frenkel, A., Frank, D., & Zlotnik, A. (2022). Traumatic brain injury-induced submissive behavior in rats: link to depression and anxiety. Translational Psychiatry, 12(1), 239. https://doi.org/10.1038/s41398-022-01991-1
Can, Z., Yildiz, O., Sahin, H., Akyuz Turumtay, E., Silici, S., & Kolayli, S. (2015). An investigation of Turkish honeys: their physico-chemical properties, antioxidant capacities and phenolic profiles. Food Chemistry, 180, 133–141. https://doi.org/10.1016/j.foodchem.2015.02.024
Çetin, A., & Deveci, E. (2019). Expression of vascular endothelial growth factor and glial fibrillary acidic protein in a rat model of traumatic brain injury treated with honokiol: a biochemical and immunohistochemical study. Folia Morphologica, 78(4), 684–694. https://doi.org/10.5603/FM.a2019.0029
Che Mohd Nassir, C. M. N., Abdul Hamid, H., Hambali, A., Abd Manan, N., Mehat, M. Z., Ismail, N. I. & Mustapha, M. (2022). Neuroprotective Potentials of Honey for Cerebral Small Vessel Disease. OBM Neurobiology; 6(4), 144. https://doi.org/10.21926/obm.neurobiol.2204144
Chen, X., Wang, H., Zhou, M., Li, X., Fang, Z., Gao, H., Li, Y., & Hu, W. (2018). Valproic acid attenuates traumatic brain injury-induced inflammation in vivo: Involvement of autophagy and the Nrf2/ARE signaling pathway. Frontiers in Molecular Neuroscience, 11, 117. https://doi.org/10.3389/fnmol.2018.00117
Chou, A., Krukowski, K., Jopson, T., Zhu, P. J., Costa-Mattioli, M., Walter, P., & Rosi, S. (2017). Inhibition of the integrated stress response reverses cognitive deficits after traumatic brain injury. Proceedings of the National Academy of Sciences, 114(31), E6420-E6426. https://doi.org/10.1073/pnas.1707661114
Ciulu, M., Spano, N., Pilo, M. I., & Sanna, G. (2016). Recent Advances in the Analysis of Phenolic Compounds in Unifloral Honeys. Molecules, 21(4), 451. https://doi.org/10.3390/molecules21040451
Cole, J. T., Yarnell, A., Kean, W. S., Gold, E., Lewis, B., Ren, M., McMullen, D. C., Jacobowitz, D. M., Pollard, H. B., O'Neill, J. T., Grunberg, N. E., Dalgard, C. L., Frank, J. A., & Watson, W. D. (2011). Craniotomy: true sham for traumatic brain injury, or a sham of a sham? Journal of Neurotrauma, 28(3), 359–369. https://doi.org/10.1089/neu.2010.1427
Corps, K. N., Roth, T. L., & McGavern, D. B. (2015). Inflammation and neuroprotection in traumatic brain injury. JAMA Neurology, 72(3), 355–362. https://doi.org/10.1001/jamaneurol.2014.3558
Dennis, D., Memudu, A. E., Solomon, A., & Adebayo, G. (2022). Honey-induced elevation of superoxide dismutase declined lipid peroxidation and attenuates neurodegeneration in weight drop model of traumatic brain injury in adult Wistar rats. Acta Scientific Anatomy, 1(7), 02-11.
Du, L., Empey, P. E., Ji, J., Chao, H., Kochanek, P. M., Bayır, H., & Clark, R. S. (2016). Probenecid and N-Acetylcysteine Prevent Loss of Intracellular Glutathione and Inhibit Neuronal Death after Mechanical Stretch Injury In Vitro. Journal of Neurotrauma, 33(20), 1913–1917. https://doi.org/10.1089/neu.2015.4342
Erejuwa, O. O., Sulaiman, S. A., & Ab Wahab, M. S. (2012). Honey: A novel antioxidant. Molecules, 17(4), 4400–4423. https://doi.org/10.3390/molecules17044400
Eteraf-Oskouei, T., & Najafi, M. (2013). Traditional and modern uses of natural honey in human diseases: A review. Iranian Journal of Basic Medical Sciences, 16(6), 731–742.
Fadzil, M. A. M., Mustar, S., & Rashed, A. A. (2023). The Potential Use of Honey as a Neuroprotective Agent for the Management of Neurodegenerative Diseases. Nutrients, 15(7), 1558. https://doi.org/10.3390/nu15071558
Filippone, A., Cucinotta, L., Bova, V., Lanza, M., Casili, G., Paterniti, I., Campolo, M., Cuzzocrea, S., & Esposito, E. (2023). Inhibition of LRRK2 Attenuates Depression-Related Symptoms in Mice with Moderate Traumatic Brain Injury. Cells, 12(7), 1040. https://doi.org/10.3390/cells12071040
Flores, M. S. R., Escuredo, O., & Seijo, M. C. (2015). Assessment of physicochemical and antioxidant characteristics of Quercus pyrenaica honeydew honeys. Food Chemistry, 166, 101–106. https://doi.org/10.1016/j.foodchem.2014.06.005
Gholami, M. R., Abbaszadeh, A., Anbari, K., Khaksarian, M., Shabooni, F., Khayat, Z. K., Khorramabadi, R. M., & Gharravi, A. M. (2020). Protective Effects of Honey, Apis mellifera Meda Skorikov, on Ischemia-Reperfusion Induced Muscle Injury. International Journal of Morphology, 38(3), 804–810.
Gholami, M., Khayat, Z. K., Anbari, K., Obidavi, Z., Varzi, A., Boroujeni, M. B., Alipour, M., Niapoor, A., & Gharravi, A. M. (2017). Quercetin ameliorates peripheral nerve ischemia-reperfusion injury through the NF-kappa B pathway. Anatomical Science International, 92(3), 330–337. https://doi.org/10.1007/s12565-016-0336-z
Gruenbaum, S. E., Zlotnik, A., Gruenbaum, B. F., Hersey, D., & Bilotta, F. (2016). Pharmacologic Neuroprotection for Functional Outcomes After Traumatic Brain Injury: A Systematic Review of the Clinical Literature. CNS Drugs, 30(9), 791–806. https://doi.org/10.1007/s40263-016-0355-2
Guo, C., Sun, L., Chen, X., & Zhang, D. (2013). Oxidative stress, mitochondrial damage and neurodegenerative diseases. Neural Regeneration Research, 8(21), 2003–2014. https://doi.org/10.3969/j.issn.1673-5374.2013.21.009
Hampshire, A., & Sharp, D. J. (2015). Contrasting network and modular perspectives on inhibitory control. Trends in Cognitive Sciences, 19, 445–452. https://doi.org/10.1016/j.tics.2015.06.006
Hasenan, S. M., Sahhugi, Z., & Jubri, Z. (2017). Gelam honey increases antioxidant enzyme activity in young rat cardiac mitochondria. International Journal of Biomedical and Advance Research, 8(2), 44-49.
Huang, C., Sakry, D., Menzel, L., et al. (2016). Lack of NG2 exacerbates neurological outcome and modulates glial responses after traumatic brain injury. Glia, 64, 507–523. https://doi.org/10.1002/glia.22944
Huang, Y., Long, X., Tang, J., Li, X., Zhang, X., Luo, C., Zhou, Y., & Zhang, P. (2020). The Attenuation of Traumatic Brain Injury via Inhibition of Oxidative Stress and Apoptosis by Tanshinone IIA. Oxidative Medicine and Cellular Longevity, 2020, 4170156. https://doi.org/10.1155/2020/4170156
Iftikhar, A., Nausheen, R., Muzaffar, H., Naeem, M. A., Farooq, M., Khurshid, M., Almatroudi, A., Alrumaihi, F., Allemailem, K. S., & Anwar, H. (2022). Potential Therapeutic Benefits of Honey in Neurological Disorders: The Role of Polyphenols. Molecules, 27(10), 3297. https://doi.org/10.3390/molecules2710329
Jatana, M., Giri, S., Ansari, M. A., Elango, C., Singh, A. K., Singh, I., & Khan, M. (2006). Inhibition of NF-kB activation by 5-lipoxygenase inhibitors protects brain against injury in a rat model of focal cerebral ischemia. Journal of Neuroinflammation, 3, 12. https://doi.org/10.1186/1742-2094-3-12
Johnson, V. E., Meaney, D. F., Cullen, D. K., & Smith, D. H. (2015). Animal models of traumatic brain injury. Handbook of Clinical Neurology, 127, 115–128. https://doi.org/10.1016/B978-0-444-52892-6.00008-8
Karve, I. P., Taylor, J. M., & Crack, P. J. (2016). The contribution of astrocytes and microglia to traumatic brain injury. British Journal of Pharmacology, 173(4), 692–702. https://doi.org/10.1111/bph.13125
Khan, M., Sakakima, H., Dhammu, T. S., Shunmugavel, A., Im, Y.-B., Gilg, A. G., Singh, A. K., & Singh, I. (2011). S-Nitrosoglutathione reduces oxidative injury and promotes mechanisms of neuro repair following traumatic brain injury in rats. Journal of Neuroinflammation, 8(78), 1–17. https://doi.org/10.1186/1742-2094-8-78
Kraeuter, A. K., Guest, P. C., & Sarnyai, Z. (2019). The Elevated Plus Maze Test for Measuring Anxiety-Like Behavior in Rodents. Methods in Molecular Biology, 1916, 69–74. https://doi.org/10.1007/978-1-4939-8994-2_4
Krämer, T., Grob, T., Menzel, L., Hirnet, T., Griemert, E., Radyushkin, K., Thal, S. C., Methner, A., & Schaefer, M. K. E. (2017). Dimethyl fumarate treatment after traumatic brain injury prevents depletion of antioxidative brain glutathione and confers neuroprotection. Journal of Neurochemistry, 143(5), 523–533. https://doi.org/10.1111/jnc.14220
Lecca, D., Bader, M., Tweedie, D., Hoffman, A. F., Jung, Y. J., Hsueh, S.-C., Hoffer, B. J., Becker, R. E., Pick, C. G., Lupica, C. R., & Greig, N. H. (2019). (-)-Phenserine and the prevention of pre-programmed cell death and neuroinflammation in mild traumatic brain injury and Alzheimer's disease challenged mice. Neurobiology of Disease, 130, 1-15. https://doi.org/10.1016/j.nbd.2019.104528
Liang, J., Wu, S., Xie, W., & He, H. (2018). Ketamine ameliorates oxidative stress-induced apoptosis in experimental traumatic brain injury via the Nrf2 pathway. Drug Design, Development and Therapy, 12, 845–853. https://doi.org/10.2147/DDDT.S160046
Luchese, R. H., Prudêncio, E. R., & Guerra, A. F. (2017). Honey as a Functional Food. In Vagner de Alencar Arnaut de Toledo (Ed.), Honey Analysis. IntechOpen. https://doi.org/10.5772/67020
Mahboob, A., Farhat, S. M., Iqbal, G., Babar, M. M., Zaidi, N. U., Nabavi, S. M., et al. (2016). Alpha-lipoic acid-mediated activation of muscarinic receptors improves hippocampus- and amygdala-dependent memory. Brain Research Bulletin, 122, 19–28. https://doi.org/10.1016/j.brainresbull.2016.02.014
Malkoç, M., Özer Yaman, S., Imamoğlu, Y., İnce, İ., Vanizor Kural, B., Mungan, S., Livaoglu, M., Yıldız, O., & Kolaylı, S. (2020). Anti-inflammatory, antioxidant and wound-healing effects of mad honey in streptozotocin-induced diabetic rats. International Journal of Experimental Pathology, 59(4). https://doi.org/10.1080/00218839.2019.1689036
McKee, C. A., & Lukens, J. R. (2016). Emerging roles for the immune system in traumatic brain injury. Frontiers in Immunology, 7, 556. https://doi.org/10.3389/fimmu.2016.00556
McKee, C., & Robinson, E. (2014). Military-related traumatic brain injury and neurodegeneration. Alzheimer's & Dementia, 10(5), 242–253. https://doi.org/10.1016/j.jalz.2014.04.003
Mijanur Rahman, M., Gan, S. H., & Khalil, M. I. (2014). Neurological effects of honey: current and future prospects. Evidence-based Complementary and Alternative Medicine, 2014, 958721. https://doi.org/10.1155/2014/958721
Mojtahedzadeh, M., Ahmadi, A., Mahmoodpoor, A., Beigmohammadi, M. T., Abdollahi, M., Khazaeipour, Z., Shaki, F., Kuochaki, B., & Hendouei, N. (2014). Hypertonic saline solution reduces the oxidative stress responses in traumatic brain injury patients. Journal of Research in Medical, 19(9), 867–874.
Morales, N., Popovic, N., & Popovic, M. (2018). Time of day and age impact of memory in elevated plus maze test in rats. Frontiers in Behavioral Neuroscience, 12, 20. https://doi.org/10.3389/fnbeh.2018.00304
Nathan, C., & Cunningham-Bussel, A. (2013). Beyond oxidative stress: An immunologist's guide to reactive oxygen species. Nature Reviews Immunology, 13, 349–361. https://doi.org/10.1038/nri3423
National Research Council. 2011. Guide for the Care and Use of Laboratory Animals: Eighth Edition. Washington, DC: The National Academies Press. https://doi.org/10.17226/12910
O'leary, R. A., & Nichol, A. D. (2018). Pathophysiology of severe traumatic brain injury. Journal of Neurosurgical Sciences, 62(5), 542–548. https://doi.org/10.23736/S0390-5616.18.04501-0
Oyekunle, O. A., Akanmu, M. A., & Ogundeji, T. P. (2010). Evaluation of anxiolytic and novelty induced behaviours following bee-honey consumption in rats. Journal of Neuroscience and Behavioral Health, 2(4), 38–43.
Özevren, H., Deveci, E., & Tuncer, M. C. (2018). Histopathological changes in the choroid plexus after traumatic brain injury in rats: A histologic and immunohistochemical study. Folia Morphologica, 77(4), 642-648. https://doi.org/10.5603/fm.a2018.0029
Pandey, D. K., Yadav, S. K., Mahesh, R., & Rajkumar, R. (2009). Depression-like and anxiety-like behavioral aftermaths of impact accelerated traumatic brain injury in rats: A model of comorbid depression and anxiety? Behavioural Brain Research, 205(2), 436–442. https://doi.org/10.1016/j.bbr.2009.07.027
Popovitz, J., Mysore, S. P., & Adwanikar, H. (2019). Long-Term Effects of Traumatic Brain Injury on Anxiety-Like Behaviors in Mice: Behavioral and Neural Correlates. Frontiers in Behavioral Neuroscience, 13, 6. https://doi.org/10.3389/fnbeh.2019.00006
Quillinan, N., Herson, P. S., & Traystman, R. J. (2016). Neuropathophysiology of brain injury. Anesthesiology Clinics, 34, 453-464. https://doi.org/10.1016/j.anclin.2016.04.011
Rao, P. V., Krishnan, K. T., Salleh, N., & Gan, S. H. (2016). Biological and therapeutic effects of honey produced by honeybees and stingless bees: A comparative review. Brazilan Journal of Pharmacognosy, 26(5), 657–664. https://doi.org/10.1016/j.bjp.2016.01.012
Rodríguez-Rodríguez, A., Egea-Guerrero, J. J., Murillo-Cabezas, F., & Carrillo-Vico, A. (2014). Oxidative stress in traumatic brain injury. Current Medicinal Chemistry, 21, 1201–1211. https://doi.org/10.2174/0929867321666131217153310
Solumsmoen, S., Lilja-Cyron, A., Buch, K. F., & Kelsen, J. (2018). [Traumatic penetrating brain injury]. Ugeskrift for Læger, 180(51), V03180201.
Walf, A. A., & Frye, C. A. (2013). The use of elevated plus maze as an assay of anxiety-related behaviors in rodents. Nature Protocols, 2(2), 322–328. https://doi.org/10.1038/nprot.2007.44
Wagner, J. M., Sichler, M. E., Schleicher, E. M., Franke, T. N., Irwin, C., Löw, M. J., Beindorff, N., Bouter, C., Bayer, T. A., & Bouter, Y. (2019). Analysis of Motor Function in the Tg4-42 Mouse Model of Alzheimer's Disease. Frontiers in Behavioral Neuroscience, 13, 107. https://doi.org/10.3389/fnbeh.2019.00107
Wang, H. C., Lin, Y. J., Shih, F. Y., et al. (2016). The role of serial oxidative stress levels in acute traumatic brain injury and as predictors of outcome. World Neurosurgery, 87, 463–470. https://doi.org/10.1016/j.wneu.2015.10.010
Wang, S., Wu, S., & Bais, S. (2018). Ameliorative potential of Lagenaria sicereria extract as anti-anxiety drug in various models of anxiety in rats. International Journal of Pharmacology, 14(8), 1179–1187. https://doi.org/10.3923/ijp.2018.1179.1187
Xu, J., Shi, J., Zhang, J., & Zhang, Y. (2018). Vorinostat: A histone deacetylase (HDAC) inhibitor ameliorates traumatic brain injury by inducing iNOS/Nrf2/ARE pathway. Folia Neuropathologica, 56(3), 179–186. https://doi.org/10.5114/fn.2018.78697
Yang, Y., Wang, H., Li, L., Li, X., Wang, Q., Ding, H., et al. (2016). Sinomenine provides neuroprotection in model of traumatic brain injury via the Nrf2-ARE pathway. Frontiers in Neuroscience, 10, 580. https://doi.org/10.3389/fnins.2016.00580
Yates, N. J., Lydiard, S., Fehily, B., Weir, G., Chin, A., Bartlett, C. A., Alderson, J., & Fitzgerald, M. (2017). Repeated mild traumatic brain injury in female rats increases lipid peroxidation in neurons. Experimental Brain Research, 235(7), 2133–2149. https://doi.org/10.1007/s00221-017-4958-8
Zakaria, F. H., Samhani, I., Mustafa, M. Z., & Shafin, N. (2022). Pathophysiology of Depression: Stingless Bee Honey Promising as an Antidepressant. Molecules, 27(16), 5091. https://doi.org/10.3390/molecules27165091
Zamri, N. A., Ghani, N., Ismail, C. A. N., Zakaria, R., & Shafin, N. (2023). Honey on brain health: A promising brain booster. Frontiers in Aging Neuroscience, 14, 1092596. https://doi.org/10.3389/fnagi.2022.1092596
Zulkifli, N. A., Hassan, Z., Mustafa, M. Z., Azman, W. N. W., Hadie, S. N. H., Ghani, N., & Mat Zin, A. A. (2023). The potential neuroprotective effects of stingless bee honey. Frontiers in Aging Neuroscience, 14, 1048028. https://doi.org/10.3389/fnagi.2022.1048028
Downloads
Published
How to Cite
Issue
Section
Categories
License
Copyright (c) 2024 Adejoke Elizabeth Memudu, Deloraine Dennis, Indranath Chatterjee
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
The observations and associated materials published or posted by NeurosciRN are licensed by the authors for use and distribution in accord with the Creative Commons Attribution license CC BY-NC 4.0 international, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
