Decalcification-free technique on the analysis of dental pulp tissue: histological and immunohistochemical analysis
Abstract
Background: Histological techniques are essential for the microscopic study and investigation of the human dental pulp. The aim of this study was to evaluate the impact of decalcification-free technique by examining dental pulp morphology by histological staining with haematoxylin and eosin and immunohistochemistry. Materials and methods: The sample consisted of 30 healthy third molars extracted for orthodontic indication, the pulp tissue was obtained by removing the mineralized tissues, separating the enamel and dentine and by marking with a flexible diamond disc on the coronal surface and longitudinal axis of the root. These guides made it possible to separate the fragments and obtain the pulp tissue for fixation and staining with H&E and subsequent immunohistochemistry with CD34 and S-100 antibodies. Results: The technique showed preservation of pulp morphology with adequate preservation of microscopic structures. No alterations in tissue viability were observed. The staining allowed an accurate assessment of vascular and nervous components by means of CD34 and S-100 markers, respectively. Conclusions: This technique allows preservation of pulp tissue, maintaining viable tissue for histological analysis and immunohistochemistry tests, as well as reducing sample processing time.
Keywords: CD34histologydental pulpimmunohistochemistryS-100
References
- Beniash E, Stifler CA, Sun CY, et al. The hidden structure of human enamel. Nat Commun. 2019; 10(1): 4383.
- Couve E, Lovera M, Suzuki K, et al. Schwann cell phenotype changes in aging human dental pulp. J Dent Res. 2018; 97(3): 347–355.
- Daud S, Nambiar P, Hossain M, et al. Removal of the apical one-third of the root improves the fixation process of the dental pulp in teeth. J Histotechnol. 2016; 39(3): 81–87.
- Feldman AT, Wolfe D. Tissue processing and hematoxylin and eosin staining. Methods Mol Biol. 2014; 1180: 31–43.
- Frank RM, Nalbandian J. Structure and ultrastructure of the dental pulp. Teeth. 1989: 249–307.
- Gaudin A, Renard E, Hill M, et al. Phenotypic analysis of immunocompetent cells in healthy human dental pulp. J Endod. 2015; 41(5): 621–627.
- Gheorghe AG, Mercuţ V, Popescu SM, et al. Histological and immunohistochemical aspects of the atrophic dental pulp modifications of abutment teeth. Rom J Morphol Embryol. 2019; 60(3): 889–894.
- Goldberg M, Lasfargues JJ. Pulpo-dentinal complex revisited. J Dent. 1995; 23(1): 15–20.
- Gupta S, Jawanda MK, Sm M, et al. Qualitative histological evaluation of hard and soft tissue components of human permanent teeth using various decalcifying agents - a comparative study. J Clin Diagn Res. 2014; 8(9): ZC69–ZC72.
- Johansson CB, Röser K, Jimbo R, Hammarström JP, Larsson L, Östberg AK. Histological analysis. In: Comprehensive Biomaterials II, 2 ed. Elsevier, London 2017: 335–363.
- Kawashima N, Okiji T. Odontoblasts: specialized hard-tissue-forming cells in the dentin-pulp complex. Congenit Anom (Kyoto). 2016; 56(4): 144–153.
- Keklikoglu N, Akinci S. Comparison of three different techniques for histological tooth preparation. Folia Histochem Cytobiol. 2013; 51(4): 286–291.
- Kleinert A, Kleinert L, Ozimirska M, et al. Endodontium — together or separately? Folia Morphol. 2018; 77(3): 409–415.
- Ramos-Vara JA. Principles and methods of immunohistochemistry. Methods Mol Biol. 2017; 1641: 115–128.
- Ricucci D, Amantea M, Girone C, et al. Atypically grown large periradicular cyst affecting adjacent teeth and leading to confounding diagnosis of non-endodontic pathology. Aust Endod J. 2020; 46(2): 272–281.
- Ricucci D, Loghin S, Niu LN, et al. Changes in the radicular pulp-dentine complex in healthy intact teeth and in response to deep caries or restorations: A histological and histobacteriological study. J Dent. 2018; 73: 76–90.
- Ricucci D, Siqueira JF, Abdelsayed RA, et al. Bacterial invasion of pulp blood vessels in teeth with symptomatic irreversible pulpitis. J Endod. 2021; 47(12): 1854–1864.
- Sarna-Boś K, Boguta P, Szymańska-Chargot M, et al. Insight into structural and chemical profile/composition of powdered enamel and dentine in different types of permanent human teeth. Micron. 2024; 179: 103608.
- Shetty H, Kakade A, Shetty S, et al. Immunohistochemical characterization of stem cell and differentiation markers of the dental pulp of human natal teeth. Future Sci OA. 2018; 4(10): FSO342.
- Singh H, Bishen K, Garg D, et al. Fixation and fixatives: roles and functions — a short review. Dent J Adv Stud. 2019; 07(02): 051–055.
- Slaoui M, Bauchet AL, Fiette L. Tissue Sampling and Processing for Histopathology Evaluation. Methods Mol Biol. 2017; 1641: 101–114.
- Titford M. Progress in the development of microscopical techniques for diagnostic pathology. J Histotechnol. 2013; 32(1): 9–19.
- Tjäderhane L, Paju S. Dentin‐pulp and periodontal anatomy and physiology. In: Orstavik D. ed. Essential Endodontology (3rd ed). Wiley-Blackwell, Oxford 2020: 11–58.
- Tomaszewska JM, Miskowiak B, Matthews-Brzozowska T, et al. Characteristics of dental pulp in human upper first premolar teeth based on immunohistochemical and morphometric examinations. Folia Histochem Cytobiol. 2013; 51(2): 149–155.
- Widbiller M, Rothmaier C, Saliter D, et al. Histology of human teeth: standard and specific staining methods revisited. Arch Oral Biol. 2021; 127: 105136.