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In Vitro and In Vivo Approaches to Determine Intestinal Epithelial Cell Permeability
Published on: October 19, 2018
Prevotella melaninogenica disrupted oral epithelial barrier function via myosin light chain kinase
Yiting Guo1, Wenhao Han2, Yuan He1
1Department of Oral Medicine, Stomatology Hospital and Dental School of Tongji University, Shanghai Engineering Research Center of Tooth Restoration and Regeneration, Shanghai, China.
Objective:
Our previous studies have found that the composition ratio of Prevotella melaninogenica (Pm) on buccal mucosa surface of oral lichen planus (OLP) patients increased significantly compared with control. Furthermore, Pm could invade the epithelium of OLP patients. This study aimed to further explore the impact of Pm on oral keratinocytes.
Materials And Methods:
The Pm-human oral keratinocyte (HOK) co-culture model was established to detect monolayer permeability, zona occludens-1 (ZO-1) expression, and intracellular survival of Pm. We performed RNA-seq followed by identification of differentially expressed genes (DEGs) and Gene Ontology (GO) analysis, with a particular focus on myosin light chain kinase (MLCK). An MLCK inhibitor ML-7 was utilized in Pm-HOK co-culture model to assess its effects on monolayer permeability and ZO-1 expression.
Results:
HOK monolayer permeability was increased, and ZO-1 expression was decreased after co-culture (p < 0.05). Pm could survive in HOK cells. RNA-seq revealed MLCK was an upregulated common DEG. The expression of MLCK in the Pm-HOK co-culture model was upregulated. Inhibition of MLCK rescued the increased epithelial permeability, and ZO-1 expression was upregulated (p < 0.05).
Conclusion:
MLCK may be involved in disrupting epithelial barrier function by Pm.
Insights
Prevotella melaninogenica (Pm) increases oral keratinocyte permeability and disrupts the epithelial barrier by upregulating myosin light chain kinase (MLCK). Inhibiting MLCK restores barrier function, suggesting a role for Pm in oral lichen planus pathogenesis.
Area of Science:
- Oral microbiology
- Epithelial biology
- Immunodermatology
Background:
- Oral lichen planus (OLP) is associated with increased Prevotella melaninogenica (Pm) on the buccal mucosa.
- Pm has the ability to invade the epithelium of OLP patients.
- The specific impact of Pm on oral keratinocytes remains to be fully elucidated.
Purpose of the Study:
- To investigate the effects of Prevotella melaninogenica (Pm) on human oral keratinocytes (HOKs).
- To identify molecular mechanisms underlying Pm-induced epithelial barrier dysfunction.
- To explore the potential role of myosin light chain kinase (MLCK) in this process.
Main Methods:
- Established a co-culture model of Pm and HOKs.
- Assessed HOK monolayer permeability and zona occludens-1 (ZO-1) expression.
- Utilized RNA-sequencing (RNA-seq) to identify differentially expressed genes (DEGs), focusing on MLCK.
- Administered an MLCK inhibitor (ML-7) to evaluate its impact on epithelial barrier function.
Main Results:
- Co-culture with Pm increased HOK monolayer permeability and decreased ZO-1 expression.
- Pm demonstrated intracellular survival within HOKs.
- RNA-seq identified MLCK as a significantly upregulated DEG in the Pm-HOK model.
- MLCK inhibition reversed the increased permeability and restored ZO-1 expression.
Conclusions:
- Prevotella melaninogenica (Pm) disrupts the oral epithelial barrier function.
- Myosin light chain kinase (MLCK) is implicated in Pm-mediated epithelial barrier disruption.
- Targeting MLCK may offer a therapeutic strategy for OLP by restoring epithelial integrity.
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