Immunohistochemical Staining for Nerve Fibers in Normal Tissues with Frozen Sections: Structure and Function Rela-tionship
Main Article Content
Abstract
Small nerve fibers are difficult to immunostain with routinely formalin-fixed and paraffin-embedded sections. Normal tissues except central nervous system are not densely innervated compared to densely distributed lymphatic and blood vessels and nerve fibers are not properly preserved in the routinely formalin-fixed and paraffin-embedded tissues. Nerve density technique with skin punch biopsy is an established method using 50 µm tissue sections, which are initially preserved in fixative. The fixed tissues are frozen sectioned and floating tissue sections are immunostained for nerve fiber markers. We had developed an alternative method using unfixed frozen sections mounted on glass slides. We had previously used this frozen section method for lymphatic and blood vessel immunohistochemistry and extended to nerve fiber immunostaining using neurofilament and CD56 as nerve fiber markers. Immunostained tissues with frozen sections were compared to formalin-fixed and paraffin-embedded colon and kidney sections. The normal tissues from rhesus monkey included heart, intestines, diaphragm, pancreas, spleen, kidney, thyroid, urinary bladder and others. Frozen section immunostained sections clearly depicted much more fine nerve fibers especially using CD56 as a nerve marker than with paraffin-embedded tissues. This technique is more labor-intensive but provides more precise distribution of fine nerve fibers than with routinely paraffin-embedded tissues. This immunostaining with frozen sections validated usefulness to depict more detailed nerve distribution and will shed light on normal histology and histopathology.
Keywords:
CD56, frozen sections, immunohistochemistry, nerve fibers, neurofilament, normal rhesus monkey tissues
Article Details
How to Cite
TOMITA, Tatsuo.
Immunohistochemical Staining for Nerve Fibers in Normal Tissues with Frozen Sections: Structure and Function Rela-tionship.
Medical Research Archives, [S.l.], v. 11, n. 6, july 2023.
ISSN 2375-1924.
Available at: <https://esmed.org/MRA/mra/article/view/3884>. Date accessed: 03 dec. 2024.
doi: https://doi.org/10.18103/mra.v11i6.3884.
Section
Research Articles
The Medical Research Archives grants authors the right to publish and reproduce the unrevised contribution in whole or in part at any time and in any form for any scholarly non-commercial purpose with the condition that all publications of the contribution include a full citation to the journal as published by the Medical Research Archives.
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4. Tokushige, N, Marham, R, Russell, P, Fraser, IS: High density of small nerve fibers in the func-tional layer of the endometrium in women with endometriosis. Hum Reprod 2006, 21(3);782-787.
5. Tokushige, N, Markham, R, Russell, P, Fraser, IS: Different types of small nerve fibers in eu-topic endometrium and myometrium in women with endometriosis. Fert Steril 2007, 88(4);795-803.
6. Boker, A, Kyama, CM, Vercruysse, L, Fass-bender, Gevaert, O et al: Density of small di-ameter sensory nerve fibers in endometrium: a semi-invasive diagnostic test for minimal to mild endometriosis. Human Reprod 2009, 24(12);3025-3032.
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8. McArthur, JC, Stocks, EA, Hauer, P, Cornblath, DR, Griffin, JW: Epidermal nerve fibers densi-ty. Arch Neurol 1998, 55(12);1513-1520.
9. Ebenzer, GS, Hauer, P, Gibbons, C, McArthur, JC, Polydefkis, M: Assessment of epidermal fi-bers: A new diagnostic and predictive for pe-ripheral neuropathies. J Neuropath Exp Nerol 2007, 66(12);1059-1073.
10. Saperstein, DS, Leveine, TD, Hank, N: Useful-ness of skin biopsy in the evaluation and man-agement of patients with suspected small nerve neuropathy fiber neuropathy. Int J Neu-rosci 2013,123(1);38-41.
11. Grayston, R, Czanner, E, Elhadd, K, Goebel, A, Frank, B et al: A systematic review and meta-analysis of the prevalence of small fiber in pa-thology in fibromyalgia of a new paradigm in pathology in fibromyalgia etiogenesis. Sem Arthritis Rheumat 2019, 48(5);933-949.
12. Tomita, T, Mah, K: Cyclic changes of lymphatic and venous vessels in human endometrium. Open J Pathol 2014, 4(1);5-12.
13. Tomita, T, Mah, K: Cyclic changes of nerve fibers in human endometrium. Open J Pathol 2014, 4(1);68-78.
14. Tomita, T: Immunohistochemical staining for lymphatic and blood vessels in normal tissues: comparison between routinely paraffin-embedded tissues and frozen sections. Acta Medica Acad 2120, 50(1);13-28.
15. Thompson, RS, Doran, JF, Jackson, P, Dhillon, AP, Rode, J: PGP9.5 -A new marker for verte-brate neurons and neuroendocrine cells. Brain Res 1983, 278(1-2);224-228.
16. Wilkinson, KD, Lee, KM, Deschpande, S, Deschpande-Hughs, P, Boss, JM: The neuron specific protein PGP9.5 is ubiquitin carboxy-terminal hydroxylase . Science 1989, 246(4930);670-673.
17. Karanth, SS, Springall, DR, Kuhn, DM, Levine, MM, Pollak, JM: An immunocytochemical study of neuropeptides and protein gene 9.5 in man and common employed laboratory animals. Am J Anat, 1991, 191(4);369-383.
18. Schlepfer, WW: Neurofilaments: structure, metabolism and implications in disease. J Neu-ropathol Exp Neurol 1987, 46(2):117-129.
19. Yuan, A, Rao, MV, Veeranna, Nixon, RA: Neu-rofilaments at a glance. J Cell Sci 2012, 125(14);3257-3263.
20. Hall, J: Guyton and Hall Textbook of Medical Physiology, 12th ed., Saunders/Elsevier, Phila-delphia, PA, 2011, 563-564.
21. Nagi, SS, Marshall, AG, Makdani, MA, Jaro-cha, E, Lifjencranz, J et al: An ultrafast system for signaling mechanical pain in human skin. Sci Adv 2019, 5(7);1297.
22. Mechtersheimer, G, Stauder, M, Moller, P : Expression of the natural killer cell--associated antigen CD56 and CD57 in human neural and striated muscle cells and their tumors. Cancer Res 1991, 51(5);1300-1307.
23. Jensen,M, Berthold, F: Targeting the neural cell adhesion in molecules in cancer. Cancer Lett 2007, 258(1);9-21.
24. Enriquez-Barreto, L, Palazzetti, C, Brenonann, LH, Maness, PF, Fairen, A: Neural cell adhesion in molecule, NCAM, regulates thalamocortical axon pathfinding and the organization of cor-tical somatosensory regeneration in mouse. Fron Molec Neurosci 2012, 00076.
25. Leshcynska, I, Liew, HT, Shephard, C, Halliday, GM, Stevens, CH: Aβ-derived reduction of MCA M2 -mediated synaptic adhesion con-tributes to synaptic loss in Alzheimer’s disease. Nature Common 2015, 6;8836.
26. Lajoix, AD, Reggio, H, Chardes, T, Peraldi-Roux, S, Tribillac, E et al: A neural isoform of nitric oxide synthase expression in pancreatic β-cells control insulin secretion. Diabetes 2001, 5(6);1311-1323.
27. Bahadoran, Z, Caristyrom, M, Mirmiran, P, Ghasemi, A: Nitric oxide: To be not to be en-docrine hormone? Acta Physiolo 2020,229(1);e13443.
28. Wheeler, T, Watkins, PS: Cardiac denervation in diabetes. Br J Med 1973, 4;584-586.
29. Marron, K, Wharton, J, Shepard, MN, Fagan, D, Royston, D et al: Distribution, morphology, and neurochemistry of endocardial and epi-cardial nerve terminal arborization in the hu-man heart. Circulation 1995, 92(8);2343-2351.
30. Armour, JA, Murphy, DA, Yuan, BX, MacDon-ald, S, Hopkins, DA: Gross and microscopic anatomy of the human intrinsic cardiac nerv-ous system. Anat Rec 1999, 247(2);289-298.
31. Michell, GA: Innervation of the heart. Br Heart J, 1953,15(2);159-171.
32. Cavallotti, C, Bruzzone, P, Mancone, M: Cate-cholaminergic nerve fibers and beta-adrenergic receptors in the human heart and coronary vessels. Heart Valves 2002, 17(1);30-35.
33. Hanna, P, Rajendran, PS, Ajiola, O, Vaseghi, M, Andrew, J et al: Cardiac neuroanatomy-imaging nerves to define functional control. Auton Neurosci 2017,207;48-58.
34. Christensen, AH, Nygaard, S, Rolid, K, Ny-troen, K, Gullestad, L et al: Early signs of sino-atrial reinnervation in the transplanted heart. Transplantation 2021, 105(9);2086-2096.
35. Grupper, E, Gewirtz, H, Kushwaha, S: Reinner-vation post-transplantation. Eur J Heart, 2018, 39(20);1799-1806.
36. Crick, SJ, Wharlton, J, Sheppard, MN, Royton, D, Yacoub, MH et al: Innervation of the human cardiac conduction system: A quantitative im-munohistochemical and histochemical study. Circulation 1994, 89(4);1697-1708.
37. Sparrow, MP, Weichselbaum, M, McCry, B Jr: Development of the innervation and airway smooth muscle in human fetal lung. Am J Respir Cell Mol Biol 1999, 20(4);550-560.
38. Lauwryns, JM, Van Ranst, L: Protein gene product 9.5 expression in the lungs of humans and other mammals. Immunocytochemical de-tection in neuroepithelial bodies, neuroendo-crine cells and nerves. Neurosci 1988, 85(3);311-316.
39. Laitinen, A: Autonomic innervation of the hu-man respiratory tract as revealed by histo-chemical and ultrastructural method. Eur J Respir Dis 1985, 140(Suppl);1-42.
40. Fisher, A, Hoffmann, B: Nitric oxide synthase in neurons and nerve fibers of lower airways and in vagal sensory ganglia of man. Correlations with neuropeptides. Am J Respir Crit Med 1996, 154(1);209-216.
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