Myeloid differentiation factor 2 inhibitors exert protective effects on lipopolysaccharides-treated human dental pulp cells via suppression of toll-like receptor 4-mediated signaling

Savitri Vaseenon; Tanida Srisuwan; Guang Liang; Nipon Chattipakorn; Siriporn C Chattipakorn

doi:10.1016/j.jds.2023.04.024

Myeloid differentiation factor 2 inhibitors exert protective effects on lipopolysaccharides-treated human dental pulp cells via suppression of toll-like receptor 4-mediated signaling

J Dent Sci. 2024 Jan;19(1):220-230. doi: 10.1016/j.jds.2023.04.024. Epub 2023 May 5.

Authors

Savitri Vaseenon^{1

2}, Tanida Srisuwan¹, Guang Liang³, Nipon Chattipakorn^{2

4}, Siriporn C Chattipakorn^{2

4

5}

Affiliations

¹ Department of Restorative Dentistry and Periodontology, Faculty of Dentistry, Chiang Mai University, Chiang Mai, Thailand.
² Neurophysiology Unit, Cardiac Electrophysiology Research and Training Center, Faculty of Medicine, Chiang Mai University, Chiang Mai, Thailand.
³ Chemical Biology Research Center, School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang, China.
⁴ Center of Excellence in Cardiac Electrophysiology Research, Chiang Mai University, Chiang Mai, Thailand.
⁵ Department of Oral Biology and Diagnostic Sciences, Faculty of Dentistry, Chiang Mai University, Chiang Mai, Thailand.

Abstract

Background/purpose: The toll-like receptor 4 (TLR4)-myeloid differentiation factor 2 (MD-2) complex is known to have a role in inflammation. Blocking MD-2 can suppress inflammatory process. However, the actual action of MD-2 inhibitors, including MAC28, L6H21, and 2i-10, on the inflamed human dental pulp cells (HDPCs) has never been examined. This study aims to determine the pharmacological effects of these 3 compounds on cell viability, inflammation, and osteo/odontogenic differentiation of lipopolysaccharide (LPS)-treated HDPCs.

Materials and methods: HDPCs were pretreated with 10 μM of MAC28, L6H21, or 2i-10 for 2 h followed by either 20 μg/mL LPS or vehicle for 24 h. Cell viability was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The mRNA and expression of the proteins TLR4, MD-2, tumor necrosis factor alpha (TNF-α), and interleukin 6 (IL-6) were determined using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot analysis. Osteo/odontogenic differentiation was investigated using qRT-PCR and Alizarin Red staining.

Results: LPS did not alter cell viability but significantly increased the expression levels of TLR4, MD-2, TNF-α, and IL-6 in HDPCs while the osteo/odontogenic differentiation ability decreased significantly when compared to the vehicle-treated group. MAC28, L6H21, and 2i-10-pretreatment in LPS-treated HDPCs reduced inflammation and restored osteo/odontogenic differentiation to similar levels as the vehicle-treated group.

Conclusion: MAC28, L6H21, and 2i-10 exhibited equal efficacy in attenuating inflammation through downregulation of TLR4-MD-2 signaling and restored osteo/odontogenic differentiation in LPS-treated HDPCs. These MD-2 inhibitors could be considered as the potential therapeutic supplement for curing inflammation of dental pulp in future studies.

Keywords: Dental pulp; Inflammation; Lipopolysaccharides; Myeloid differentiation factor 2; Toll-like receptor 4.