Mechanisms of cytotoxicity of nicotine in human periodontal ligament fibroblast cultures in vitro

Abstract

The use of tobacco products significantly contributes to the progression of periodontal disease and poor response to healing following periodontal therapy. The purpose of this study was to determine the effects of nicotine, a major component of cigarette smoking, on human periodontal ligament fibroblast (PDLF) growth, proliferation, and protein synthesis to elucidate its role in periodontal destruction associated with its use. Human PDLFs were derived from three healthy individuals undergoing extraction for orthodontic reasons. At a concentration higher than 2.5 mM, nicotine was found to be cytotoxic to human PDLFs (P < 0.05). Nicotine also significantly inhibited cell proliferation and decreased protein synthesis in a dose-dependent manner. At concentrations of 50 and 200 microM, nicotine suppressed the growth of PDLFs by 48% and 86% (P < 0.05), respectively. A 10-mM concentration level of nicotine significantly inhibited the protein synthesis to only 44% of these in the untreated control (P < 0.05). Furthermore, the effects of antioxidants (superoxide dismutase (SOD); catalase and 2-oxothiazolidine-4-carboxylic acid (OTZ) and buthionine sulfoximine (BSO) were added to search for the possible mechanism of action, as well as a method for the prevention, of cigarette smoking-associated periodontal diseases. The addition of OTZ, a precursor of cysteine that metabolically promotes GSH synthesis, acted as a protective effect on the nicotine-induced cytotoxicity. However, SOD and catalase did not decrease the nicotine-induced cytotoxicity. In contrast, the addition of BSO, a cellular GSH synthesis inhibitor, enhanced the nicotine-induced cytotoxicity. These results indicate that thiol depletion could be the mechanism for nicotine cytotoxicity. The levels of nicotine tested inhibited cell growth, proliferation, and protein synthesis on human PDLFs. This suggests that nicotine itself might augment the destruction of periodontium associated with cigarette smoking. In addition, these inhibitory effects were associated with intracellular thiol levels. Factors that induce glutathione synthesis of human PDLF may be used for further chemoprevention of cigarette smoking-related periodontal diseases.

Nicotine inhibits myofibroblast differentiation in human gingival fibroblasts

“…In in vitro experiments using human gingival fibroblasts (HGFs), it was found that nicotine treatment inhibited HGF proliferation in doses higher than 7.8 mM, and disrupted the microtubule and vimentin cytoskeleton in doses higher than 3.9 mM [Alpar et al, 1998]. Whereas in studies using human periodontal ligament fibroblasts, it was demonstrated that nicotine (≥ 25 μM) inhibited cell proliferation and protein synthesis (≥ 5 mM) in a dose-dependent manner [Chang et al, 2002]. In our study, HGFs were subconfluent or confluent before exposure to nicotine; therefore, the effect of nicotine on cell proliferation was not evaluated.…”

Poor patient awareness of the relationship between smoking and periodontal diseases

“…10 Nicotine has cytotoxic effects on periodontal ligament fibroblasts. 11 It also has inhibitory effects on periodontal cell proliferation and protein synthesis which result in impaired wound healing. 11 Smoking results in changes to vascular, 12 inflammatory, 13,14 immune 15 and healing responses.…”

“…11 It also has inhibitory effects on periodontal cell proliferation and protein synthesis which result in impaired wound healing. 11 Smoking results in changes to vascular, 12 inflammatory, 13,14 immune 15 and healing responses. 10,11 There is considerable scientific evidence of its harmful long term effects on periodontal diseases.…”

“…11 Smoking results in changes to vascular, 12 inflammatory, 13,14 immune 15 and healing responses. 10,11 There is considerable scientific evidence of its harmful long term effects on periodontal diseases. 16 Grossi et al 17,18 stated that the odds ratio of smokers developing adult periodontitis was greater than the relationship between periodontitis and the gram-negative anaerobic bacteria that causes this.…”

Deliberations on Non-Surgical Periodontal Therapy

“…According to Johnson and Guthmiller it increases the odds of developing periodontal disease 2 to 8 -fold, depending on the definition of disease severity and smoking dose. The exact mechanism of smoking action is still unknown however several mechanisms for negative periodontal effects of smoking are proposed which include decreased Ig G2 production [14], chronic reduction in blood flow and vascularity [15], increased prevalence of potential periodontal pathogens [16], shift in neutrophil function towards destructive activities [17], negative effects on cytokine and growth factor production [18], and inhibition of fibroblast growth, attachment and collagen production [19]. These mechanisms in one way or another, makes individuals more susceptible to periodontal disease and compromise the healing response.…”