Thymosin Beta-4 Cytoprotective In Human Gingival Fibroblasts

Reti, Robert, Michelle Wheater, and Gabriel Sosne

Thymosin beta 4 (Tß4) is a highly conserved protein found in all tissues, except erythrocytes. Until recently it was thought to function solely as the major G-actin sequestering molecule. Current investigations have identified it to be an endothelial cell chemoattractant, an angiogenic stimulant, and an anti-apoptotic and anti-inflammatory agent most notably in ocular, skin and heart tissue. OBJECTIVES: The aim of the current investigation was to examine the cytoprotective role of Tß4 in oral tissue using human gingival fibroblasts. METHODS: The human gingival fibroblast cell line HGF-1 (ATCC CRL-2014) was cultured in medium containing 10% serum and then serum-starved. In one set of experiments gingival fibroblasts were treated with TNF-α or chlorhexedine with or without Tß4. Apoptosis was assayed colorimetrically using the ApoPercentage assay. Apoptotic cells were counted in digitally captured images. Apoptosis data was determined as mean number of apoptotic cells +/- SEM. In a second set of experiments gingival fibroblasts were cultured in 24-well plates and treated with either TNF-α, chlorhexidine digluconate, Peridex, or carbamide peroxide with or without Tß4. Cytotoxicity was assayed for lactate dehydrogenase (LDH) activity using a colorimetric LDH cytotoxicity assay kit. Statistical analysis was performed using the student's t-test with significance value of p < 0.05. RESULTS: Tß4 was able to protect gingival fibroblasts from apoptosis induced by TNF-α or by chlorhexidine. Additionally, Tß4 was able to protect gingival fibroblasts against the cytotoxic effects of common dental chemotherapeutics such as chlorhexidine digluconate, Peridex, and carbamide peroxide. CONCLUSION: Because of its multifunctional roles in protecting cells against damage, Tß4 has significant potential for use in a mouth rinse with its combined antimicrobial, anti-inflammatory, anti-apoptotic, and cytoprotective properties.