Bioactive antioxidant mixtures promote proliferation and migration on human oral fibroblasts.
ABSTRACT Antioxidants (AOs) are the first line of defence against free radical damage and are critical for maintaining optimum health and well being. The need for AOs becomes even more critical with increased exposure to free radicals generated by pollution, cigarette smoke, drugs, illness, stress and exercise. Antioxidant supplementation is an excellent way of improving free radical protection. The aim of this study was to provide cytotoxicity, proliferation and migration data on the in vitro effects of bioactive AO mixtures on human oral fibroblasts.
Human oral fibroblasts were obtained from human gingival (HGF) and periodontal (HPDL) tissues. Each of these oral fibroblasts was cultured separately in three concentrations of the bioactive pure polyphenol and turmeric derivative mixtures; resveratrol (R), ferulic acid (F), phloretin (P) and tetrahydrocurcuminoids (T); [(RFT), (PFR), and (PFT)]. Cell viability, proliferation, morphology and migratory behaviour were analysed in vitro using high throughput in vitro 96 well plate wound assay.
RFT decreased (10(-3)M) and increased (10(-5)M) cell number in HGF cells. Three concentrations (10(-3), 10(-4), and 10(-5)M) of PFR and PFT increased DNA synthesis in HGF cells. PFT promoted cell migration but PFR and RFT had no significant change in HGF wound healing rates in a 96 well plate assay monolayer wound. In the HPDL cells, the 10(-4)M concentration of both RFT and PFT increased cell number at 72 h and 96 h whereas the lower concentration 10(-5)M of RFT significantly stimulated cell number at 96 h. PFR (10(-3)M and 10(-5)M) and PFT (10(-3)M) increased DNA synthesis after 48 h treatment in HPDL cells.
High and low concentrations (10(-3)-10(-5)M) of these AOs (RFT, PFR) may have beneficial effects on functional mechanisms regulating fibroblast migration and proliferation during gingival healing or periodontal repair.
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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.Journal of Periodontal Research 08/2002; 37(4):279-85. · 1.99 Impact Factor
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ABSTRACT: BACKGROUND, aims: The habit of betel nut chewing impinges on the daily lives of approximately 200 million people. Betel quid chewers have a higher prevalence of periodontal diseases than non-chewers. This study examined the pathobiological effects of arecoline, a major component of the betel nut alkaloids, on human periodontal ligament fibroblasts (PDLF) in vitro. Cell viability, proliferation, protein synthesis, and cellular thiol levels were used to investigate the effects of human PDLF exposed to arecoline levels of 0 to 200 microg/ml. In addition, nicotine was added to test how it modulated the effects of arecoline. Arecoline significantly inhibited cell proliferation in a dose-dependent manner. At concentrations of 10 and 30 microg/ml, arecoline suppressed the growth of PDLF by 20% and 50% (p < 0.05), respectively. Arecoline also decreased protein synthesis in a dose-dependent manner during a 24-h culture period. A 100 microg/ ml concentration level of arecoline significantly inhibited protein synthesis to only 50% of that in the untreated control (p < 0.05). Moreover, arecoline significantly depleted intracellular thiols in a dose-dependent manner. At concentrations of 25 microg/ml and 100 microg/ml, arecoline depleted about 18% and 56% of thiols (p < 0.05), respectively. This suggests that arecoline itself might augment the destruction of periodontium associated with betel nut use. Furthermore, the addition of nicotine acted with a synergistic effect on the arecoline-induced cytotoxicity. At a concentration of 60 microg/ml, arecoline suppressed the growth of PDLF by about 33% and 5 mM nicotine enhanced the arecoline-induced cytotoxic response to cause about 66% cell death. During thiol depletion, arecoline may render human PDLF more vulnerable to reactive agents within cigarettes. Taken together, people who combine habits of betel nut chewing with cigarette smoking could be more susceptible to periodontium damage than betel nut chewing alone.Journal Of Clinical Periodontology 03/2001; 28(3):277-82. · 3.69 Impact Factor
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ABSTRACT: Reduced expression of proapoptotic and terminal differentiation genes in conjunction with increased levels of the proinflammatory and angiogenesis-inducing enzymes, cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS), correlate with malignant transformation of oral intraepithelial neoplasia (IEN). Accordingly, this study investigated the effects of a 10% (w/w) freeze-dried black raspberry gel on oral IEN histopathology, gene expression profiles, intraepithelial COX-2 and iNOS proteins, and microvascular densities. Our laboratories have shown that freeze-dried black raspberries possess antioxidant properties and also induce keratinocyte apoptosis and terminal differentiation. Oral IEN tissues were hemisected to provide samples for pretreatment diagnoses and establish baseline biochemical and molecular variables. Treatment of the remaining lesional tissue (0.5 g gel applied four times daily for 6 weeks) began 1 week after the initial biopsy. RNA was isolated from snap-frozen IEN lesions for microarray analyses, followed by quantitative reverse transcription-PCR validation. Additional epithelial gene-specific quantitative reverse transcription-PCR analyses facilitated the assessment of target tissue treatment effects. Surface epithelial COX-2 and iNOS protein levels and microvascular densities were determined by image analysis quantified immunohistochemistry. Topical berry gel application uniformly suppressed genes associated with RNA processing, growth factor recycling, and inhibition of apoptosis. Although the majority of participants showed posttreatment decreases in epithelial iNOS and COX-2 proteins, only COX-2 reductions were statistically significant. These data show that berry gel application modulated oral IEN gene expression profiles, ultimately reducing epithelial COX-2 protein. In a patient subset, berry gel application also reduced vascular densities in the superficial connective tissues and induced genes associated with keratinocyte terminal differentiation.Cancer Research 07/2008; 68(12):4945-57. · 8.65 Impact Factor