Lactobacilli Attenuate the Effect of Aggregatibacter actinomycetemcomitans Infection in Gingival Epithelial Cells

Author(s)

Bueno, Manuela R. ; Ishikawa, Karin H. ; Almeida-Santos, Gislane ; Ando-Suguimoto, Ellen S. ; Shimabukuro, Natali ; Kawamoto, Dione ; Mayer, Marcia P. A.

• Document type: Journal article
• Source: FRONTIERS IN MICROBIOLOGY; v. 13, p. 11-pg., 2022-05-04.

Abstract

Probiotics may be considered as an additional strategy to achieve a balanced microbiome in periodontitis. However, the mechanisms underlying the use of probiotics in the prevention or control of periodontitis are still not fully elucidated. This in vitro study aimed to evaluate the effect of two commercially available strains of lactobacilli on gingival epithelial cells (GECs) challenged by Aggregatibacter actinomycetemcomitans. OBA-9 GECs were infected with A. actinomycetemcomitans strain JP2 at an MOI of 1:100 and/or co-infected with Lactobacillus acidophilus La5 (La5) or Lacticaseibacillus rhamnosus Lr32 (Lr32) at an MOI of 1:10 for 2 and 24 h. The number of adherent/internalized bacteria to GECs was determined by qPCR. Production of inflammatory mediators (CXCL-8, IL-1 beta, GM-CSF, and IL-10) by GECs was determined by ELISA, and the expression of genes encoding cell receptors and involved in apoptosis was determined by RT-qPCR. Apoptosis was also analyzed by Annexin V staining. There was a slight loss in OBA-9 cell viability after infection with A. actinomycetemcomitans or the tested probiotics after 2 h, which was magnified after 24-h co-infection. Adherence of A. actinomycetemcomitans to GECs was 1.8 x 10(7) (+/- 1.2 x 10(6)) cells/well in the mono-infection but reduced to 1.2 x 10(7) (+/- 1.5 x 10(6)) in the co-infection with Lr32 and to 6 x 10(6) (+/- 1 x 10(6)) in the co-infection with La5 (p < 0.05). GECs mono-infected with A. actinomycetemcomitans produced CXCL-8, GM-CSF, and IL-1 beta, and the co-infection with both probiotic strains altered this profile. While the co-infection of A. actinomycetemcomitans with La5 resulted in reduced levels of all mediators, the co-infection with Lr32 promoted reduced levels of CXCL-8 and GM-CSF but increased the production of IL-1 beta. The probiotics upregulated the expression of TLR2 and downregulated TLR4 in cells co-infected with A. actinomycetemcomitans. A. actinomycetemcomitans-induced the upregulation of NRLP3 was attenuated by La5 but increased by Lr32. Furthermore, the transcription of the anti-apoptotic gene BCL-2 was upregulated, whereas the pro-apoptotic BAX was downregulated in cells co-infected with A. actinomycetemcomitans and the probiotics. Infection with A. actinomycetemcomitans induced apoptosis in GECs, whereas the co-infection with lactobacilli attenuated the apoptotic phenotype. Both tested lactobacilli may interfere in A. actinomycetemcomitans colonization of the oral cavity by reducing its ability to interact with gingival epithelial cells and modulating cells response. However, L. acidophilus La5 properties suggest that this strain has a higher potential to control A. actinomycetemcomitans-associated periodontitis than L. rhamnosus Lr32. (AU)

• FAPESP's process: 16/13159-6 - Supra and subgingival microbiota diversity analysis and protein profile of saliva and enamel film of individuals with chronic periodontitis and microbial succession
• Grantee: Dione Kawamoto
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 17/16377-7 - EVALUATION OF THE EFFECT OF PROBIOTIC BACTERIA ON THE IMUNE RESPONSE TO A.ACTINOMYCETEMCOMITANS: A STUDY IN GINGIVAL EPITHELIAL CELLS AND IN EXPERIMENTAL MODEL OF INFECTION IN VIVO.
• Grantee: Manuela Rocha Bueno
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 16/13156-7 - Direct and immunomodulatory effects of probiotic bacteria on periodontal pathogens
• Grantee: Karin Hitomi Ishikawa
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 16/14687-6 - Evaluation of immunomodulatory potential of probiotics in periodontal disease
• Grantee: Emmanuel Albuquerque de Souza
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 15/18273-9 - New strategies for the control of periodontitis
• Grantee: Marcia Pinto Alves Mayer
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 17/22345-0 - Effect of Bifidobacterium probiotics on the modulation of immune response and alveolar bone loss in an experimental periodontitis model on normoglycemic and diabetic mice
• Grantee: Natali Shimabukuro
• Support Opportunities: Scholarships in Brazil - Master