Analyzing Loricrin in Healthy and Periodontally Diseased Gingival Epithelium

• Author / Creator

  Roy, Christopher B

• ABSTRACT

  Introduction: Periodontitis is an inflammatory disease of the periodontium that leads to irreversible loss of periodontal ligament, cementum, and alveolar bone. Stage 3 and stage 4 grade C periodontitis (P3C/P4C) are the most severe forms of periodontitis according to the 2017 Classification of Periodontal and Peri-Implant Diseases and Conditions (Papapanou et al., 2018). Loricrin is an integral structural protein comprising more than 70% of the total protein mass in the outer keratinized layer of the epithelium (Nithya et al., 2015). Downregulated and/or mutant phenotypes of loricrin have been linked to inflammatory epithelial related disorders of the skin (Catunda et al., 2019). This suggests an important role of this protein in maintaining barrier function.
  Hypothesis: If histological and immunohistological differences exist in the epithelium of Healthy versus P3C/P4C samples then this may suggest a pathogen-mediated epithelial barrier breach due to loss of loricrin.
  Methods: A total of 15 gingival epithelium samples were collected during the study period. Discard tissue samples were collected from patients who underwent various periodontal procedures at the University of Alberta Periodontology Clinic. The samples were derived from 5 patients with gingival health, 5 patients with generalized periodontitis stage III grade C, and 5 patients generalized periodontitis IV grade C. The samples were immediately fixed in paraformaldehyde (PFA) and then embedded in paraffin. Each sample was then serially sectioned and mounted on slides for histological and immunohistological analysis. Hematoxylin and Eosin (H&E) staining was completed for a comprehensive expression of the gingival epithelium and connective tissue. Immunofluorescence analysis was completed using antibodies for Loricrin (Lor), Cytokeratin 1 (CK1), and Cytokeratin 14 (CK14). Immunostaining results from these assays were compared and documented. Using ImageJ software [National Institutes of Health (NIH), Bethesda, MD, USA], the immunostaining intensity of loricrin was semi-quantitatively analyzed and compared amongst the tissue samples by two investigators independently.
  Results: For Healthy, P3C, and P4C gingival samples, immunofluorescence assay expressed loricrin, CK1, and CK14 in a similar pattern. Loricrin was expressed predominantly in the stratum granulosum and corneum. Healthy gingival samples had a relatively wider loricrin expression. CK1 was observed in the stratum spinosum and granulosum. A general uniform immunofluorescent signaling trend was observed in the Healthy gingival samples relative to a more interrupted immunofluorescent signal for both P3C and P4C gingival samples. CK14 presented in the strata basale for all gingival sample types. A general uniformed immunofluorescent trend was present for CK14 with no observable distinction amongst the three types of samples.
  Significance: The findings from this study suggest that the inflammatory condition of P3C and P4C epithelium may be attributable to a weaker loricrin phenotypic expression. Studies on the pathogenesis of periodontitis tend to focus on the host immune response and the microbiology of the disease. A weakened epithelial barrier and its role in the pathogenesis of periodontal disease necessitates further study.

• Subjects / Keywords

  • Loricrin
  • Gingival
  • Immunofluorescence
  • Epithelium
  • Healthy
  • Diseased
  • Human
  • Immunohistological

• Graduation date

  Fall 2022

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/r3-k9y2-k104

• License

  This thesis is made available by the University of Alberta Library with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.