Cell transformation induced by bovine papillomavirus DNA as an assay for tumor promoters and chemopreventive agents.

An in vitro assay was designed to examine and quantitate the action of chemical promoters and chemopreventive agents on papillomavirus DNA-carrying cells. Cultured C3H/10T1/2 cells transfected with bovine papillomavirus type 1 DNA (plasmid pdBPV-1) were used as targets, and the frequency of transformed foci was used as an endpoint. The development of foci with a transformed phenotype was greatly enhanced by tumor promoters (e.g., mezerein, 12-O-tetradecanoyl-phorbol-13-acetate, teleocidin, and okadaic acid) and complex mixtures such as extracts of the areca nut, which is an integral part of a betel quid and is linked to oral cancers among chewers. The degree of promotion depended on the length of exposure, the type of promoter, and the time of application after transfection with BPV DNA. The inhibitory effect of chemopreventive agents on transformation can be tested either directly on BPV DNA transfected cells (promoter-independent transformation), or on transfected cells that were exposed to tumor promoters (promoter-dependent transformation). Retinol, and to a lesser degree beta-carotene, exerted an inhibitory effect on promoter-dependent and promoter-independent transformation of BPV DNA transfected cells. The inhibitory effect was conveyed either by the addition of retinol simultaneously with promoters, or after exposure to the promoting agents was completed. The significance of this short-term in vitro assay for the design of chemopreventive trials is discussed.     

Oral lesions, genotoxicity and nitrosamines in betel quid chewers with no obvious increase in oral cancer risk

A link between the generation of areca nut-related N-nitrosamines in the saliva, the induction of genotoxic damage in the oral mucosa, as judged by an increase in micronucleated exfoliated cells (MEC), and a low incidence of oral cancer was studied in 2 population groups characterized by their habit of chewing quids without tobacco: Guamanians, who chew areca nuts (Areca catechu) with or without the addition of betel leaf (Piper betle); Taiwanese, who use areca nut, betel leaf or inference and slaked lime. The levels of N-nitrosoguvacoline (NG) in the saliva of chewers of fresh green areca nuts were very high (70.8 ng/ml) as compared to those reported for individuals using the more complex Indian betel quids (0.91 ng/ml or 5.6 ng/ml). None of the other areca nut-related nitrosamines (N-nitrosoguvacine (NGC), 3-(methylnitrosamino)propionitrile (MNPN) and 3-(methylnitrosamino)propionaldehyde (MNPA)) were detected in the saliva of Taiwanese betel quid chewers. The addition of slaked lime to the areca nut enhances the formation of NG during a chewing session. The frequency of MEC did not increase in the oral mucosa of areca nut chewers who do not use slaked lime, but showed a small but significant elevation in individuals using lime-containing quids. The elevation of MEC in Taiwanese, who are at low risk for oral cancer, is relatively small as compared to that found in chewers of Indian betel quids (pan), who show a highly elevated oral cancer risk. The results seem to suggest that NG may play only a minor role, if any, in the etiology of oral cancer among betel quid chewers.     

Role of areca nut consumption in the cause of oral cancers. A cytogenetic assessment.

BACKGROUND. Cytogenetic studies, framed to assess the possible genomic damage caused by areca nut consumption (without tobacco and not as a component of betel quid), were performed among areca nut chewers, which included normal people who chew areca nuts, patients with oral submucous fibrosis, and patients with oral cancer, and healthy nonchewing controls. RESULTS. The analysis showed statistically significant increases in the frequencies of sister chromatid exchanges and chromosome aberrations in peripheral blood lymphocytes and the percentage of micronucleated cells in exfoliated cells of buccal mucosa among all three groups of chewers when compared with those of the controls. CONCLUSIONS. The current data, the first of this type among only areca nut chewers, highlight that this popular masticatory is erroneously considered "safe" and that it increases the genomic damage even when chewed without tobacco. The data also signify that, henceforth, in cytogenetic biomonitoring, areca nut consumption also should be considered as one of the confounding factors.     

O6-methylguanine-DNA methyltransferase activity in human buccal mucosal tissue and cell cultures. Complex mixtures related to habitual use of tobacco and betel quid inhibit the activity in vitro

Extracts prepared from tissue specimens of normal, non-tumourous human buccal mucosa, and cultured buccal epithelial cells and fibroblasts, exhibited O6-methylguanine-DNA methyltransferase (MGMT) activity by catalysing the repair of the premutagenic O6-methylguanine lesion in isolated DNA with rates of 0.2 to 0.3 pmol/mg protein. An SV40 T antigen-immortalized buccal epithelial cell line termed SVpgC2a and a buccal squamous carcinoma line termed SqCC/Y1, both of which lack normal tumour suppressor gene p53 function, exhibited about 50 and 10% of the MGMT activity of normal cells, respectively. The normal, experimentally transformed and tumourous buccal cell types showed MGMT mRNA levels which correlated with their respective levels of MGMT activity. Exposure of buccal cell cultures to various organic or water- based extracts of products related to the use of tobacco and betel quid, decreased both cell survival (measured by reduction of tetrazolium dye) and MGMT activity (measured subsequently to the exposures in cellular extracts). Organic extracts of bidi smoke condensate and betel leaf showed higher potency than those of tobacco and snuff. An aqueous snuff extract also decreased both parameters, whereas an aqueous areca nut extract was without effect. The well- established sulph-hydryl-reactive agent Hg2+, a corrosion product of dental amalgam, served as a positive control and decreased MGMT activity following treatment of cells within a range of 1-10 microM. Taken together, significant MGMT activities were demonstrated in buccal tissue specimens and in the major buccal mucosal cell types in vitro. Lower than normal MGMT activity in two transformed buccal epithelial cell lines correlated with decreased MGMT mRNA and lack of functional p53. Finally, in vitro experiments suggested the potential inhibition of buccal mucosal MGMT activity by complex mixtures present in the saliva of tobacco and betel nut chewers.      

Chromosome-damaging activity of saliva of betel nut and tobacco chewers

Saliva of volunteers chewing betel quid, cured betel nut (Areca catechu), betel leaves (Piper betle), a mixture of quid ingredients (dried betel nut flakes, catechu, cardamon, lime, copra and menthol) and Indian tobacco was collected and examined for its genotoxic activity. Chromosome aberrations (chromatid breaks and chromatid exchanges) in Chinese hamster ovary (CHO) cells were used to estimate the genotoxic effect. No detectable levels of clastogenic activity were observed in the saliva of non-chewing individuals. After 5 min of chewing betel quid, betel nut, betel leaves, quid ingredients and Indian tobacco, the saliva samples showed relatively potent clastogenic activities. The addition of transition metals Mn²⁺ and Cu²⁺ to the saliva samples of betel nut and Indian tobacco chewers enhanced their clastogenic activities, whereas Fe³⁺ increased the clastogenicity of the betel nut saliva but decreased the genotoxic effect of the saliva of Indian tobacco chewers. After removal of the betel quid or its components from the mouth, the clastogenic activity disappeared within 5 min. The western-type chewing tobacco did not produce a genotoxic activity in the saliva of chewers. A possible association between the genotoxicity in the saliva of betel quid chewers and the development of oral, pharyngeal and esophageal carcinomas is discussed.     

Cytotoxic Effects of Betel Quid and Areca Nut Aqueous Extracts on Mouse Fibroblast,Human Mouth-Ordinary-Epithelium 1 and Human Oral Squamous Cell Carcinoma Cell Lines

Background: Betel quid chewing is more common among the older generation in rural areas of Malaysia. Oral cancer in Asia has been associated with the habit of chewing betel quid and areca nut. Objective:  This study aims to investigate the cytotoxic effects of betel quid and areca nut extracts on the fibroblast (L929), mouth-ordinary-epithelium 1 (MOE1) and oral squamous cell carcinoma (HSC-2) cell lines. Methods: L929, MOE1 and HSC-2 cells were treated with 0.1, 0.2 and 0.4 g/ml of betel quid and areca nut extracts for 24, 48 and 72 h. MTT assay was performed to assess the cell viability. Results: Both extracts, regardless of concentration, significantly reduced the cell viability of L929 compared with the control (P<0.05). Cell viability of MOE1 was significantly enhanced by all betel quid concentrations compared with the control (P<0.05). By contrast, 0.4 g/ml of areca nut extract significantly reduced the cell viability of MOE1 at 48 and 72 h of incubation. Cell viability of HSC-2 was significantly lowered by all areca nut extracts, but 0.4 g/ml of betel quid significantly increased the cell viability of HSC-2 (P<0.05). Conclusion: Areca nut extract is cytotoxic to L929 and HSC-2, whereas the lower concentrations of areca nut extract significantly increased the cell viability of MOE1 compared to the higher concentration and control group. Although betel quid extract is cytotoxic to L929, the same effect is not observed in MOE1 and HSC-2 cell lines. Further investigations are needed to clarify the mechanism of action.     

Tobacco (Kretek) Smoking,Betel Quid Chewing and Risk of Oral Cancer in a Selected Jakarta Population

Purpose: This study aimed to determine the association between tobacco consumption (kretek) and betel quidchewing with oral cancer risk. Materials and Methods: A total of 81 cases of oral cancers were matched with162 controls in this hospital-based study. Information on sociodemographic characteristics and details of riskhabits (duration, frequency and type of tobacco consumption and betel quid chewing) were collected. Associationbetween smoking and betel quid chewing with oral cancer were analysed using conditional logistic regression.Results: Slightly more than half of the cases (55.6%) were smokers where 88.9% of them smoked kretek. Afteradjusting for confounders, smokers have two fold increased risk, while the risk for kretek consumers and thosesmoking for more than 10 years was increased to almost three-fold. Prevalence of betel quid chewing among casesand controls was low (7.4% and 1.9% respectively). Chewing of at least one quid per day, and quid combinationof betel leaf, areca nut, lime and tobacco conferred a 5-6 fold increased risk. Conclusions: Smoking is positivelyassociated with oral cancer risk. A similar direct association was also seen among betel quid chewers.     

Use of the micronucleus test to monitor the effect of vitamin A, beta-carotene and canthaxanthin on the buccal mucosa of betel nut/tobacco chewers

The frequency of exfoliated cells with micronuclei in buccal swabs was used to estimate the protective effect of vitamin A, beta-carotene and canthaxanthin (4,4'-diketo-beta-carotene) on the buccal mucosa of betel (areca) nut/tobacco chewers. Micronuclei were scored on exfoliated cells taken by swabbing and stained with the Feulgen reaction and fast green. The betel (areca) nut/tobacco chewers served as their own controls. Prior to the administration of vitamin A and beta-carotene, the examined betel quid chewers had elevated frequencies of micronucleated buccal mucosa cells, averaging 4.03% +/- 1.24 SD (n = 26) and 3.43% +/- 1.22 SD (n = 25), respectively. The frequency of micronucleated buccal mucosa cells in non-chewers and non-smokers was 0.51% (n = 52). Following a 9-week ingestion of vitamin A (150,000 IU/week) and beta-carotene (180 mg/week in 6 capsules), the frequency of micronucleated cells decreased significantly (p less than 0.001) to 1.70% and 1.16%, respectively. No significant shift in the frequencies of micronucleated cells was observed following the intake of canthaxanthin (180 mg/week in 6 capsules) for 9 weeks or that of a placebo. The lack of protective activity of canthaxanthin, which is a good trapper of oxygen singlets but cannot be converted into vitamin A, suggests that vitamin A and beta-carotene exert their inhibitory effect on the formation of micronuclei by a mechanism not involving the scavenging of free radicals. The efficacy of beta-carotene as an inhibitor of micronucleated cell formation, the lack of toxicity, and its availability from a multitude of dietary sources should focus attention on this carotenoid as a promising chemopreventive agent.     

Tobacco-specific and betel nut-specific N-nitroso compounds: occurrence in saliva and urine of betel quid chewers and formation in vitro by nitrosation of betel quid

In order to evaluate exposure of betel quid chewers to N-nitrosocompounds, saliva and urine samples were collected from chewersof betel quid with or without tobacco, from tobacco chewers,from cigarette smokers and from people with no such habit, andwere analysed for the presence of N-nitrosamines by gas chromatographycoupled with Thermal Energy Analyzer and alkaloids derived frombetel nut and tobacco by capillary gas chromatography fittedwith nitrogen-phosphorous selective detector. The levels ofthe betel nut-specific nitrosamines, N-nitrosoguvacoline andN-nitrososoguvacine (the latter being detected for the firsttime in saliva), ranged from 0 to 7.1 and 0 to 30.4 ng/ml, respectively.High levels of tobacco-specific nitrosamines were detected inthe saliva of chewers of betel quid with tobacco and in thatof chewers of tobacco, ranging from 1.6 to 59.7 (N'-nitrosonornicotine),1.0 to 51.7 (N'-nitrosoanatabine) and 0 to 2.3 [4-(methyl-nitrosamino)-1-(3-pyridyl)-l-butanone]ng/ml. Urinary concentrations of certain N-nitrosamino acids,including N-nitrosoproline, were determined as a possible indexof exposure to nitroso compounds and their precursors in thestudy groups: no clear difference was observed. The betel nut-specificalkaloid, arecoline, was present at high levels in the salivaof betel quid chewers with or without tobacco. Nicotine andcotinine were also detected in saliva and urine of chewers oftobacco and of betel quid with tobacco. In order to assess whetherN-nitroso compounds are formed in vivo in the oral cavity duringchewing or in the stomach after swallowing the quids, the levelsof N-nitroso compounds in betel quid extracts were determinedbefore and after nitrosation at pH 7.4 and 2.1. The resultsindicate that N-nitroso compounds could easily be formed invivo. The possible role of N-nitroso compounds in the causationof cancer of the upper alimentary tract in betel quid chewersis discussed.     

Effect of lime composition on the formation of reactive oxygen species from areca nut extract in vitro

Lime, representative of that used by betel quid chewers, was collected in a region of Papua New Guinea where the incidence of oral cancer is high. The free calcium hydroxide content and pH of 25 lime samples were highly correlated with the generation of reactive oxygen species from areca nut extract in vitro, and DNA damage in vitro, measured as 8-hydroxy-2'-deoxyguanosine. Fe2+ and Mg2+ levels in the lime samples were too low to modify formation of reactive oxygen species, but hydrogen peroxide formation was almost entirely inhibited by addition of Mg2+ to the reaction mixture. These results suggest that the calcium hydroxide content of lime in the presence of areca nut is primarily responsible for the formation of reactive oxygen species which might cause oxidative damage in the DNA of buccal mucosa cells of betel quid chewers.     

Influence of alpha-tocopherol and ascorbic acid on pan masala induced genomic damage. An in vitro experiment

Pan masala is a dry complex mixture of areca nut, catechu, lime, cardamon, unspecified flavouring agents etc., with (PMT) or without tobacco (pm). We have previously reported genotoxic potential of tobacco, areca nut and pan masala per se. An antigenotoxic effect of alpha-tocopherol (AT) and ascorbic acid (AA) against the PM/PMT induced genotoxic on Chinese hamster ovary (CHO) cells have been studied using chromosone aberration (CA) assay. AT and AA, per se, had no effect on CA frequency at the concentrations used in the present study. The short-term treatment of AT with aqueous extracts of PM/PMT yielded lower frequencies of CA as compared to the cultures treated with aqueous extracts of PM/PMT alone. However, a statistically significant reduction in CA frequency was observed with continuous treatment only. AA had no statistically significant protective effect except for continuous treatment with 10 ug/ml AA against the aqueous extract of PMT. The results indicate the possible use of AT to reduce the risk of oral cancer among PM/PMT chewers.     

Urine of tobaccolareca nut chewers causes genomic damage in Chinese hamster ovary cells

The chromosome-damaging effects of urine concentrates (UCs)from tobacco plus areca nut (T/AN) chewers (a highly popularhabit and a major risk factor for oral cancer in India) wereevaluated on Chinese hamster ovary (CHO) cells employing twocytogenetic end-points, namely chromosome aberration (CA) andsister chromatid exchange (SCE) frequencies. Urine creatininelevels were comparable between controls and T/AN chewers. CAand SCE frequencies in CHO cells were found to be elevated significantly(P < 0.001) following treatment with UCs prepared from T/ANchewers (UC-T/AN chewers) as well as with UCs of non-chewercontrols (UC-control subjects). Moreover, elevation of thesetwo parameters by UC-T/AN chewers was significantly higher incomparison to that of UC-controls. The results of the presentstudy indicated that besides the oral cavity, which is a targetorgan for T/AN chewers, mutagen/scarcinogens in tobacco andareca nut might be playing a causative role in cancer of theurinary bladder as well.     

Elevated frequency of micronucleated cells in the buccal mucosa of individuals at high risk for oral cancer: betel quid chewers

The micronucleus test was applied to buccal mucosa cells of 2 population groups at high risk for oral cancer: Khasis of the northeastern hill region of India, who eat raw betel nuts together with betel leaves and lime, and residents of the state of Orissa (India), who chew betel quids consisting mainly of perfumed tobacco, dried betel nut, betel leaf, lime and several spices. Micronuclei were scored on Feulgen/fast green-stained smear preparations of exfoliated cells obtained by scraping the surface of the buccal mucosa. All 17 raw betel nut eaters and all 20 chewers of betel quids had significantly elevated frequencies of micronucleated mucosa cells over nonchewing controls of comparable ethnic background and dietary habits. The frequencies of micronucleated exfoliated cells were higher at the site within the oral cavity where the quid was kept compared to those at the opposite buccal wall. The micronuclei frequency was lower among individuals chewing a raw betel nut, betel leaf and lime mixture compared to those using tobacco,-betel nut-, lime- and betel leaf-containing quids. Micronuclei frequencies in exfoliated human cells seem to represent a useful 'internal dosimeter' for estimating exposure to genotoxic, and by implication, carcinogenic agents in the tissue from which cancers will develop.     

Evaluation of Successfulness of Capacity Building Programmes on Smokeless Tobacco and Areca Nut Cessation

Background: Prevalence of smoking in Sri Lanka has shown a gradual reduction whilst the use of smokeless tobacco and areca nut exhibits an increasing trend. At present, only a few well-structured smokeless tobacco (SLT)/areca nut (AN) cessation programs have been conducted in Sri Lanka, which is a gross underachievement as betel chewing-related oral squamous cell carcinoma is the most common cancer in Sri Lankan males. As General Dental Practitioners (GDP) do not contribute significantly to SLT/AN cessation activities at present, capacity building programs on SLT/AN control were carried out. The study evaluated the knowledge, attitude and practices  imparted on SLT/AN control among dental surgeons. Methods: Following a single day capacity building program on smokeless tobacco / areca nut control, two self-administered questionnaires were used to assess the improvement of knowledge and change of attitudes among 663 GDPs. Results: Majority had a good knowledge on harmful effects of SLT but not on areca nut. Knowledge of the current legislation on SLT control in Sri Lanka and carcinogenicity of areca nut was not satisfactory. Almost all agreed that proper counseling leads to patient quitting the habit, a formal training is necessary to conduct tobacco control activities and it should be a part of the regular treatment modalities. More than 80% of the participants support strict legislation. Most important factors leading to poor involvement in tobacco cessation activities were lack of expertise and inadequate educational material and not breach of patient privacy and lack of financial incentives. 20.1% dental surgeons had consumed smokeless tobacco / areca nut products in the past and only a few were current users of tobacco and/or areca nut. Conclusions: Well planned workshops are efficient in improving knowledge, practices and attitudes of dental surgeons towards SLT/AN cessation.     

Overexpression of p53 protein in betel- and tobacco-related human oral dysplasia and squamous-cell carcinoma in India

The aetiological factors for oral cancer are not the same in India and in Western countries. Epidemiological studies have shown a correlation between high incidence of oral cancer and heavy consumption of betel and/or tobacco in the Indian population, while this stud/ indicates an association with a genetic change. The p53 tumour-suppressor gene is the most commonly identified mutated gene in human malignancies. Expression of p53 protein was examined in premalignant and malignant oral lesions from Indian patients who were consumers of betel, areca nut and/or tobacco, using anti-p53 monoclonal antibodies PAb 1801 and PAb 421. Cryosections from normal, premalignant or malignant oral mucosa were used for immunostaining and the observations were confirmed by immunoprecipitation. P53 protein was detected in 55% (15/27) premalignant oral lesions (leukoplakia). Strong p53-positive staining was detected in 75% (24/32) of oral squamous-cell carcinomas. Normal oral mucosa did not show positive p53 staining (0/24). The detection of p53 protein in premalignant oral lesions suggests that p53 aberrations are an early event in the development of oral cancer in India. The high incidence of p53 positivity in leukoplakia may be due to differences in aetiological factors. p53 overexpression in premalignant oral lesions is important in view of the significantly earlier onset of leukoplakia in the Indian population compared to the development of oral malignancy, and may be helpful in identifying lesions that are more likely to progress to malignancy. The frequency of p53 protein overexpression was high in premalignant and malignant oral lesions of patients who were heavy consumers of betel, areca nut and tobacco.     

Piper betel Linn (betel vine), the maligned Southeast Asian medicinal plant possesses cancer preventive effects: time to reconsider the wronged opinion

Since antiquity, Piper betel Linn (betel vine; family Piperaceae) has been an important medicinal agent in the various traditional and folk systems of medicine in Southeast Asia countries. The leaves are the most valued plant part and in the past were routinely used as a chewing agent to prevent halitosis. The leaves are also supposed to harden the gum, conserve the teeth and to prevent indigestion, bronchitis, constipation, congestion, coughs and asthma. Innumerable scientific studies have validated the ethnomedicinal claims. Betel leaves are an integral component of the betel quid that consists of areca nut (Areca catechu Linn.), tobacco (Nicotiana tabacum L) and slaked lime; a highly abused agent with carcinogenic properties. Regular chewing of betel quid is associated mainly with oral cancer and detail studies with individual constituents of the quid have shown that both tobacco and areca nut are carcinogenic, while slaked lime is shown to promote the process of carcinogenesis. However unlike other constituents of the betel quid, the betel leaves devoid carcinogenic effects and on the contrary possesses cancer preventive effects including against the carcinogens present in tobacco. This review for the first time provides information on cancer preventive effects and also addresses the various mechanisms which might be involved.     

Betel quid not containing tobacco and oral cancer: a report on a case-control study in Papua New Guinea and a meta-analysis of current evidence

Smoking and betel quid chewing are associated with increased risk of oral cancer but few studies have reported on associations in populations where betel quid does not contain tobacco. We conducted a case-control study in Papua New Guinea and a systematic review. Our case-control study recruited 143 cases with oral cancer and 477 controls. We collected information on smoking and betel quid chewing. Current smoking was associated with an increased risk of oral cancer with an adjusted odds ratio (OR) for daily smokers of 2.63 (95% confidence intervals (95% CI) 1.32, 5.22) and amongst heaviest smokers of 4.63 (95% CI 2.07, 10.36) compared to never-smokers. Betel chewing was associated with increased risk of oral cancer with an adjusted OR for current chewers of 2.03 (95% CI 1.01, 4.09) and in the heaviest chewers of 2.47 (95% CI 1.13, 5.40) compared to nonchewers. The OR in those who both smoked tobacco and chewed betel quid was 4.85 (95% 1.10, 22.25), relative to those who neither smoked nor chewed. The systematic review identified 10 previous studies that examined risk of oral cancer associated with betel quid chewing that controlled for smoking in populations where betel quid did not contain tobacco. In studies that reported results for non-smokers the combined OR was 2.14 (95% CI 1.06, 4.32) in betel quid chewers and in studies that adjusted for smoking the combined OR was 3.50 (95% CI 2.16, 5.65) in betel quid chewers. Preventive efforts should discourage betel quid chewing as well as smoking.     

Betel quid chewing and oral cancer: experimental studies on hamsters

Betel quid ingredients--betel nut, betel leaf, lime, catechu and tobacco--were tested separately and in various combinations for carcinogenicity, using hamster cheek pouch as the experimental site. The four modes of administration used were (1) tri-weekly painting of the cheek pouch with aqueous extracts of test materials, (2) deposition of replaceable wax pellets containing the test material, (3) gelatin capsules containing the powdered material and (4) insertion of natural material into the pouch for trauma and direct exposure. Untreated controls and standard carcinogen DMBA-treated controls were also maintained. A total of 317 young adult golden Syrian hamsters (Mesocricetus auratus) used for the experiments were killed in two age groups: 6-12 months and 13-24 months, only when signs of general debility were observed. In the untreated controls, animals were free of any malignancy. In the experimental series, various betel quid ingredient combinations under test induced both oral and gastric lesions ranging from massive atypia and precancerous lesions to frank carcinomas. Maximum lesions were observed in the groups receiving betel nut, lime and tobacco combinations and in the polyphenol fraction of betel nut containing major tannins. The mode of administration of test material resulted in distinct differences; tri-weekly paintings giving oral lesions in the range of 22-23% and gastric lesions 39-48%; the same material given either through the replaceable gelatin capsule or in natural form induced 69% oral lesions and 63 to 82% gastric lesions. Overall evaluation of the data of all the four series confirms the potent carcinogenicity of betel nut, particularly its tannin-containing polyphenolic fraction and its combination with lime and tobacco. Maximum oral lesions induced in the hamsters by continuous exposure to capsules and natural material, highlight the direct relationship of frequency of chewing in habitual chewers with oral carcinogenesis. The high incidence of gastric (forestomach) lesions invites special attention.     

Cytotoxic and genotoxic effects of areca nut-related compounds in cultured human buccal epithelial cells

Because betel quid chewing has been linked to the development of oral cancer, pathobiological effects of an aqueous areca nut extract, four areca nut alkaloids (arecoline, guvacoline, guvacine, and arecaidine), and four nitrosated derivatives [N-nitrosoguvacoline, N-nitrosoguvacine, 3-(N-nitrosomethylamino)propionaldehyde and 3-(N-nitrosomethylamino)propionitrile] have been investigated using cultured human buccal epithelial cells. Areca nut extract in a dose-dependent manner decreases cell survival, vital dye accumulation, and membrane integrity, and it causes formation of both DNA single strand breaks and DNA protein cross-links. Depletion of cellular free low-molecular-weight thiols also occurs, albeit at quite toxic concentrations. Comparisons of the areca nut-related N-nitroso compounds and their precursor alkaloids, at concentrations up to 5 mM, indicate that 3-(N-nitrosomethylamino)propionaldehyde is the most potent on a molar basis to decrease both survival and thiol content and to cause significant formation of DNA single strand breaks. Arecoline, guvacoline, or N-nitrosoguvacoline decreases survival and cellular thiols, whereas arecaidine, guvacine, N-nitrosoguvacine, and 3-(N-nitrosomethylamino)propionitrile have only minor effects on these variables. Taken together, the present studies indicate that aqueous extract and, in particular, one N-nitroso compound related to areca nut, i.e., 3-(N-nitrosomethylamino)propionaldehyde, are highly cytotoxic and genotoxic to cultured human buccal epithelial cells, of potential importance in the induction of tumors in betel quid chewers.     

Cultural and dietary risk factors of oral cancer and precancer--a brief overview

This is an update on cultural and dietary risk factors for oral precancer and cancer. It is an overview on ethnic differences (where possible) and socio-cultural risk factors (tobacco/areca nut/betel quid, alcohol use and dietary factors) in relation to oral precancer and cancer. While studies were from Western countries, India and China, this update also attempts to include and highlight some studies conducted in the Asia-Pacific region.