Thiamine as a peripheral neuro-protective agent in comparison with N-acetyl cysteine in axotomized rats

Document Type : Original Article


1 Biology Department, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran

2 Immunology Research Center, Inflammation and Inflammatory Diseases Division, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran


Objective(s): In this study, the impact of thiamine (Thi), N-acetyl cysteine (NAC), and dexamethasone (DEX) were investigated in axotomized rats, as a model for neural injury.
Materials and Methods: Sixty-five axotomized rats were divided into two different experimental approaches, the first experiments included five study groups (n=5): intrathecal Thi (, intraperitoneal (Thi), NAC, DEX, and control. Cell survival was assessed in L5DRG in the 4th week by histological assessment. In the second study, 40 animals were engaged to assess Bcl-2, Bax, IL-6, and TNF-α expression in L4-L5DRG in the 1st and 2nd weeks after sural nerve axotomy under treatment of these agents (n=10).
Results: Ghost cells were observed in morphological assessment of L5DRG sections, and following stereological analysis, the volume and neuronal cell counts significantly were improved in the NAC and groups in the 4th week (P<0.05). Although Bcl-2 expression did not show significant differences, Bax was reduced in the Thi group (P=0.01); and the Bcl-2/Bax ratio increased in the NAC group (1st week, P<0.01). Furthermore, the IL-6 and TNF-α expression decreased in the Thi and NAC groups, on the 1st week of treatment (P≤0.05 and P<0.01). However, in the 2nd week, the IL-6 expression in both Thi and NAC groups (P<0.01), and the TNF-α expression in the DEX group (P=0.05) were significantly decreased. 
Conclusion: The findings may classify Thi in the category of peripheral neuroprotective agents, in combination with routine medications. Furthermore, it had strong cell survival effects as it could interfere with the destructive effects of TNF-α by increasing Bax.


Main Subjects

1. Maripuu A, Björkman A, Björkman-Burtscher IM, Mannfolk P, Andersson G, Dahlin LB. Reconstruction of sciatic nerve after traumatic injury in humans-factors influencing outcome as related to neurobiological knowledge from animal research. J Brachial Plex Peripher Nerve Inj 2012;7:1-13. 
2. Gaudet AD, Popovich PG, Ramer MS. Wallerian degeneration: Gaining perspective on inflammatory events after peripheral nerve injury. J Neuroinflammation 2011;8:1-3.
3. Welin D, Novikova LN, Wiberg M, Kellerth J-O, Novikov LN. Survival and regeneration of cutaneous and muscular afferent neurons after peripheral nerve injury in adult rats. Ex Brain Res 2008;186:315-323. 
4. Hart AM, Brannstrom T, Wiberg M, Terenghi G. Primary sensory neurons and satellite cells after peripheral axotomy in the adult rat. Ex Brain Res 2002;142: 308-318. 
5. Rock KL. Pathobiology of inflammation to cell death. Biology of blood and marrow transplantation: Biol Blood Marrow Transplant 2008;15(1 Suppl):137. 
6. Martin SL, Reid AJ, Verkhratsky A, Magnaghi V, Faroni A. Gene expression changes in dorsal root ganglia following peripheral nerve injury: Roles in inflammation, cell death and nociception.  Neural Regen Res 2019;14:939. 
7. Abe N, Cavalli V. Nerve injury signaling. Cur Opin  Neurobiol 2008;18:276-283. 
8. Li S, Xue C, Yuan Y, Zhang R, Wang Y, Wang Y, et al. The transcriptional landscape of dorsal root ganglia after sciatic nerve transection. Sci Rep 2015:16888. 
9. Chandran V, Coppola G, Nawabi H, Omura T, Versano R, Huebner EA, et al. A systems-level analysis of the peripheral nerve intrinsic axonal growth program. Neuron. 2016;89:956-970. 
10. Gillardon F, Klimaschewski L, Wickert H, Krajewski S, Reed J, Zimmermann M. Expression pattern of candidate cell death effector proteins Bax, Bcl-2, Bcl-X, and c-Jun in sensory and motor neurons following sciatic nerve transection in the rat. Brain Res 1996;739:244-250. 
11. Dubový P. Wallerian degeneration and peripheral nerve conditions for both axonal regeneration and neuropathic pain induction. Ann Anat 2011;193:267-275. 
12. Dhir S, Tarasenko M, Napoli E, Giulivi C. Neurological, psychiatric, and biochemical aspects of thiamine deficiency in children and adults. Front Psychiatry 2019:-15. 
13. Tallaksen C, Taubøll E. Excitatory effect of thiamin on CA1 pyramidal neurones in rat hippocampal slices in vitro. Eur Neurol 2000;7:693-698. 
14. Manzetti S, Zhang J, van der Spoel D. Thiamin function, metabolism, uptake, and transport. Biochem. 2014;53:821- 35. 
15. Gibson GE, Zhang H. Interactions of oxidative stress with thiamine homeostasis promote neurodegeneration. Neurochem Int 2002;40:493-504. 
16. McGrath J, Drummond G, McLachlan E, Kilkenny C, Wainwright C. Guidelines for reporting experiments involving animals: The ARRIVE guidelines. Br J Pharmacol 2010;160:1573-1576. 
17. Terenghi G, Hart A, Wiberg M. The nerve injury and the dying neurons: Diagnosis and prevention. J Hand Surg Eur 2011;36:730-4. 
18. Earp JC, Pyszczynski NA,  Molano DS,    Jusko WJ.      Pharmacokinetics of dexamethasone in a rat model of rheumatoid arthritis. 
Biopharm. Drug Dispos 2008;29:366-372. 
19. Balk L, Hägerroth P-Å, Åkerman G, Hanson M, Tjärnlund U, Hansson T, et al. Wild birds of declining European species are dying from a thiamine deficiency syndrome. Proc Natl Acad Sci 2009;106:12001-12006. 
20. Størkson RV, Kjørsvik A, Tjølsen A, Hole K. Lumbar catheterization of the spinal subarachnoid space in the rat. 
J Neurosci  Methods 1996;65:167-172. 
21. Rosenling ATI. Proteomic screening of cerebrospinal fluid: Candidate proteomic biomarkers for sample stability and experimental autoimmune encephalomyelitis: University of Groningen; 2010. 
22. Tandrup T. Unbiased estimates of number and size of rat dorsal root ganglion cells in studies of structure and cell survival. J Neurophysio 2004;33:173-192. 
23. Tarokhian H, Rahimi H, Mosavat A, Shirdel A, Rafatpanah H, Akbarin MM, et al. HTLV-1-host interactions on the development of adult T cell leukemia/lymphoma: Virus and host gene expressions. BMC Cancer 2018;18:1-12. 
24. Terenghi G. Peripheral nerve regeneration and neurotrophic factors. J Anat 1999;194:1-14. 
25. Hart A, Terenghi G, Kellerth J-O, Wiberg M. Sensory neuroprotection, mitochondrial preservation, and therapeutic potential of N-acetyl-cysteine after nerve injury. Neuroscience 2004;125:91-101. 
26. Yuan J, Yankner BA. Apoptosis in the nervous system. Nature 2000;407:802-809. 
27. Chang L, Putcha G, Deshmukh M, Johnson Jr E. Mitochondrial involvement in the point of no return in neuronal apoptosis. Biochimie 2002;84:223-231. 
28. Lundborg G. A 25-year perspective of peripheral nerve surgery: Evolving neuroscientific concepts and clinical significance. J Hand Surg. 2000;25:391-414. 
29. Katare R, Caporali A, Emanueli C, Madeddu P. Benfotiamine improves functional recovery of the infarcted heart via activation of pro-survival G6PD/Akt signaling pathway and modulation of neurohormonal response. J Mol Cell Cardiol 2010;49:625-638. 
30. Yadav UC, Kalariya NM, Srivastava SK, Ramana KV. Protective role of benfotiamine, a fat-soluble vitamin B1 analogue, in lipopolysaccharide-induced cytotoxic signals in murine macrophages. Free Radic Biol Med 2010;48:1423-1434. 
31. Beltramo E, Berrone E, Tarallo S, Porta M. Different apoptotic responses of human and bovine pericytes to fluctuating glucose levels and protective role of thiamine. Diabetes Metab Res Rev 2009;25:566-576. 
32. Ishaque A, Al-Rubeai M. Role of vitamins in determining apoptosis and extent of suppression by bcl-2 during hybridoma cell culture. Apoptosis. 2002;7:231-239. 
33. West CA, Hart AM, Terenghi G, Wiberg M. Analysis of the dose-response of N-acetylcysteine in the prevention of sensory neuronal loss after peripheral nerve injury. How to Improve the Results of Peripheral Nerve Surgery: Springer; 2007. 29-31. 
34. Hart AM, Terenghi G, Wiberg M. Neuronal death after peripheral nerve injury and experimental strategies for neuroprotection. Neurol Res 2008;30: 999-1011. 
35. Chang T-C, Hung M-W, Jiang S-Y, Chu J-T, Chu L-L, Tsai L-C. Dexamethasone suppresses apoptosis in a human gastric cancer cell line through modulation of bcl-x gene expression. FEBS lett 1997;415:11-15. 
36. Lotem J, Sachs L. Regulation of bcl-2, bcl-X~ L and bax in the control of apoptosis by hematopoietic cytokines and dexamethasone. Cell Growth Differ 1995;6:647-654. 
37. Laane E, Panaretakis T, Pokrovskaja K, Buentke E, Corcoran M, Söderhäll S, et al. Dexamethasone-induced apoptosis in acute lymphoblastic leukemia involves differential regulation of Bcl-2 family members. Hematol 2007;92:1460-1469. 
38. Zaman F, Chrysis D, Huntjens K, Chagin A, Takigawa M, Fadeel B, et al. Dexamethasone differentially regulates Bcl-2 family proteins in human proliferative chondrocytes: Role of pro-apoptotic Bid. Toxicol Lett 2014;224:196-200.
39. Abdoul-Azize S, Dubus I, Vannier J-P. Improvement of dexamethasone sensitivity by chelation of intracellular Ca2+ in pediatric acute lymphoblastic leukemia cells through the prosurvival kinase ERK1/2 deactivation. Oncotarget. 2017;8:27339. 
40. Bradley J. The role of nuclear organization in cancer. J Pathol 2008;214:149-160. 
41. Madsen PM, Motti D, Karmally S, Szymkowski DE, Lambertsen KL, Bethea JR, et al. Oligodendroglial TNFR2 mediates membrane TNF-dependent repair in experimental autoimmune encephalomyelitis by promoting oligodendrocyte differentiation and remyelination. J Neurosci 2016;36:5128-5143. 
42. Ohtori S, Takahashi K, Moriya H, Myers RR. TNF-α and TNF-α receptor type 1 upregulation in glia and neurons after peripheral nerve injury: Studies in murine DRG and spinal cord. Spine 2004;29:1082-1088. 
43. Sacerdote P, Franchi S, Trovato AE, Valsecchi AE, Panerai AE, Colleoni M. Transient early expression of TNF-α in sciatic nerve and dorsal root ganglia in a mouse model of painful peripheral neuropathy. Neurosci Lett 2008;436:210-213. 
44. Üçeyler N, Tscharke A, Sommer C. Early cytokine expression in mouse sciatic nerve after chronic constriction nerve injury depends on calpain. Brain Behav Immun 2007;21:553-650. 
45. Dubový P, Jančálek R, Klusáková I, Svíženská I, Pejchalová K. Intra-and extraneuronal changes of immunofluorescence staining for TNF-and TNFR1 in the dorsal root ganglia of rat peripheral neuropathic pain models. Cell  Mol Neurobiol 2006;26:1203-1215. 
46. Murphy P, Grondin J, Altares M, Richardson P. Induction of interleukin-6 in axotomized sensory neurons. J Neurosci 1995;15:5130-5138. 
47. Yang P, Wen H, Ou S, Cui J, Fan D. IL-6 promotes regeneration and functional recovery after cortical spinal tract injury by reactivating intrinsic growth program of neurons and enhancing synapse formation. Exp Neurol 2012;236:19-27. 
48. Vega-Avelaira D, Géranton SM, Fitzgerald M. Differential regulation of immune responses and macrophage/neuron interactions in the dorsal root ganglion in young and adult rats following nerve injury. Mol Pain 2009;5:1-17. 
49. Dubový P, Klusáková I, Svízenská I, Brázda V. Satellite glial cells express IL-6 and corresponding signal-transducing receptors in the dorsal root ganglia of rat neuropathic pain model. Neuron Glia Biol 2010;6:73-83. 
50. Bozic I, Savic D, Laketa D, Bjelobaba I, Milenkovic I, Pekovic S, et al. Benfotiamine attenuates inflammatory response in LPS stimulated BV-2 microglia. PLoS One 2015;10:e0118372 
51. Spinas E, Saggini A, Kritas S, Cerulli G, Caraffa A, Antinolfi P, et al. Crosstalk between vitamin B and immunity. J Biol Regul Homeost Agents 2015;29:283-288. 
52. Victor VM, Rocha M, De La Fuente M. N-acetylcysteine protects mice from lethal endotoxemia by regulating the redox state of immune cells. Free Radic Res 2003;37:919-929. 
53. de la Fuente M, Victor VM. Ascorbic acid and N-acetylcysteine improve in vitro the function of lymphocytes from mice with endotoxin-induced oxidative stress. Free Radic Res 2001;35:73-84. 
54. Aghai ZH, Kumar S, Farhath S, Kumar MA, Saslow J, Nakhla T, et al. Dexamethasone suppresses expression of Nuclear Factor-kappaB in the cells of tracheobronchial lavage fluid in premature neonates with respiratory distress. Pediatr Res 2006;59:811-815. 
55. Chang CK, Llanes S, Schumer W. Effect of dexamethasone on NF-kB activation, tumor necrosis factor formation, and glucose dyshomeostasis in septic rats. J Surg Res 1997;72:141-145. 
56. Ramesh G, Didier PJ, England JD, Santana-Gould L, Doyle- Meyers LA, Martin DS, et al. Inflammation in the pathogenesisof Lyme neuroborreliosis. Am J Clin Pathol 2015;185:1344-1360. 
57. Uysal HB, Dağlı B, Yılmaz M, Kahyaoğlu F, Gökçimen A, Ömürlü İK, et al. Biochemical and histological effects of thiamine pyrophosphate against acetaminophen‐induced hepatotoxicity. Basic Clin Pharmacol Toxicol 2016;118:70-76. 
58. Kisaoglu A, Ozogul B, Turan MI, Yilmaz I, Demiryilmaz I, Atamanalp SS, et al. Damage induced by paracetamol compared with N-acetylcysteine. J Chin Med Assoc 2014;77:463-468.