Abbreviations: anti-HCV = antibodies to hepatitis C virus, HBsAg = hepatitis B surface antigen, HBV = hepatitis B virus, HCV = hepatitis C virus, OR = odds ratio, CI = confidence interval.
Cirrhosis is the end stage of all chronic disease affecting the liver 5,14,23,32,34). It is a risk factor for hepatocellular carcinoma, although with a variable penetrance 6,13,15,26–29,31,36,54–56,58,59,62–65. About 60%–89% of hepatocellular carcinoma is associated with underlying cirrhosis 26,28,54,56,63–65, while the prevalence of hepatocellular carcinoma complicating cirrhosis is between 2.2% and 73%13,15,26,54,56,65. Although chronic hepatitis B virus (HBV) and hepatitis C virus (HCV) infection have been implicated as the major risk factors for cirrhosis 3,11,12,20,53,54,61,62, some cirrhosis occurs in patients without serologic evidence of HBV/HCV infection 1,5,10,14,16,18,37,38,53,60, suggesting that other viruses, environmental factors, or differences in lifestyle habits are also important. To prevent cirrhosis-associated mortality and morbidity, exploration of risk factors other than HBV/HCV is urgent.
Betel quid chewing is common in 10%–20% of the human population 43, and is part of traditional Taiwanese culture 30. The estimated number of habitual betel quid chewers is about one-tenth of the 22 million inhabitants of Taiwan 30. The betel quid prepared in Taiwan is quite different from that in other parts of the world. It consists of 2 halves of a fresh areca nut, sandwiched with a piece of the betel leaf, and red slaked lime paste. Betel quid chewers generally swallow the saliva completely, thus bathing the epithelial lining of the upper digestive tract with the toxins released during chewing. This increases the possibility of severe toxic effects of betel quid at sites other than the oral cavity.
Chronic inflammation of the liver appears to be a risk factor for cirrhosis regardless of the underlying etiology 5,10,16,25,53,54,61,62. Experimental study has indicated that persistent hepatocellular damage occurs after chronic feeding with betel quid 50,57. Therefore, habitual betel quid chewing may play a role in causing cirrhosis. However, the role of betel quid chewing in the development of cirrhosis, and its interaction with other known risk factors for cirrhosis, have never been explored adequately. In this case-control study we address the independent and interactive roles of habitual betel quid chewing and other known risk factors for cirrhosis.
SUBJECTS AND METHODS
A total of 210 consecutive newly diagnosed cirrhotic patients were enrolled as cases. These patients were hospitalized or visited outpatient clinics at Kaohsiung Medical University Hospital from January 1996 to December 1997. There were 170 males and 40 females. During the same study period, 210 healthy community residents who entered the hospital for routine physicals, matched on age (±5 yr) and sex, were enrolled as the control group. There was no difference in the median age between cases (57 yr; range, 36–82 yr) and controls (58 yr; range, 36–82 yr). Cirrhosis was diagnosed by liver biopsy, abdominal sonography, and biochemical evidence of parenchymal damage plus endoscopic esophageal or gastric varices 61,62. Patients with cirrhosis were classified into the 3 Child-Pugh grades based on their clinical status 45. All healthy controls had normal serum aminotransferase levels and normal abdominal sonography. Informed consent was obtained from each subject studied. The study was approved by the Investigation and Ethics Committee of the hospital.
Structured Questionnaire and Standardized Interview
We designed a structured questionnaire to obtain information on age, sex, educational level, habits of smoking (the quantity of cigarettes smoked per day and the duration of smoking), alcohol drinking (the quantity and duration of drinking, types of alcoholic beverage), and betel quid chewing practice (the duration of habit, daily amount consumed, type of betel quid consumed). A habitual betel quid chewer was defined as chewing 1 quid or more daily for at least 1 year. A habitual cigarette smoker was defined as smoking 1 cigarette or more per day for at least 1 year. A habitual alcohol drinker was defined as drinking an alcohol beverage for more than 4 days a week for a total of at least 1 year. All cirrhotic cases and matched controls were interviewed by interviewers trained in study details and questionnaire contents. All interviews were conducted in person using the structured questionnaire.
Hepatitis B surface antigen (HBsAg) and antibodies to hepatitis C virus (anti-HCV) were detected by Ausria-II and second generation ABBOTT HCV EIA (Abbott Laboratories, North Chicago, IL), respectively. For anti-HCV, reactive specimens were retested. Only repeatedly reactive specimens were interpreted as anti-HCV positive. Conventional liver function tests were tested by an autoanalyzer (Hitachi, Model 736, Japan).
The Mann-Whitney U test was used to compare the difference between medians of continuous variables. The chi-square test with the Yates correction or the Fisher exact test was used to compare differences between proportions when appropriate. Mantel extension test for trend was used to examine the dose-response relationship for the risk estimates of various combinations of risk factors. Odds ratio (OR) with 95% confidence interval (95% CI) was used to estimate causal relations between risk factors and exposure. A conditional logistic regression analysis was used for multivariate analysis about risk factors for cirrhosis. Unconditional stepwise logistic regression analysis was used for risk factors for habitual betel quid chewing in cirrhotic patients. Adjusted odds ratios and 95% CI were derived from logistic regression coefficients to provide an estimate of the statistical association between a given variable and the disease (cirrhosis) with the other variables held constant. Synergy index was used to estimate the interactive effect among risk factors for cirrhosis 49.
To calculate the population-attributable risk for factors significantly associated with cirrhosis development in multivariate analysis, the frequency distribution of these risk factors in the control group was used to represent the proportion of persons exposed to the factor in the general population. Two-tailed p values and 95% CI were given when appropriate. An alpha of 0.05 was considered as the indicator of statistical significance.
Demographic Characteristics of Cases and Controls
The frequency distribution of sex, age, status of HBsAg and anti-HCV, and Child-Pugh grades of the study population is shown in Table 1. More than 80% of cases and controls were between 40 and 69 years old. At least 1 marker of HBsAg or anti-HCV was found in 89.05% of cirrhotic patients. The frequency distributions of Child-Pugh grades A, B, and C in cirrhotic patients were 59.04%, 21.90%, and 19.04%, respectively.
Among patients with cirrhosis, we compared the ages of beginning the habit of chewing betel quid, alcohol drinking, and smoking. There was no significant difference among median age of beginning betel quid chewing (median, 34 yr; range, 11–60 yr), alcohol drinking (median, 33 yr; range, 16–63 yr), and smoking (median, 34 yr; range, 11–56 yr).
Among controls, there was a significant difference in the frequency of habitual alcohol drinkers between those chewing betel quid (5/11, 45.45%) and those not chewing betel quid (19/199; 9.54%; p = 0.001). The frequency of habitual smokers in betel quid chewers (10/11, 90.91%) was also higher than that in betel quid non-users (66/199; 33.16%; p = 0.0002).
Univariate and Multivariate Analyses of Risk Factors for Cirrhosis
The prevalence of habitual betel quid chewing, HBsAg, and anti-HCV in cases (16.19%, 66.19%, and 28.57%, respectively) was significantly higher than that in controls (5.23%, 18.57%, and 4.28%, respectively; each p < 0.001). As shown in Table 2, univariate analysis indicated that betel quid chewing, HBsAg-positivity, anti-HCV-positivity, alcohol drinking, and smoking were significant risk factors for cirrhosis, whereas older age (>50 yr), low education level (<10 yr), and history of previous surgery and blood transfusion were not risk factors for cirrhosis. By a conditional logistic regression analysis, only betel quid chewing, HBsAg-positivity, and anti-HCV-positivity were significant independent risk factors for cirrhosis (Table 3). The estimated population-attributable risks for subjects with anti-HCV alone, subjects with HBsAg alone, subjects positive for both anti-HCV and HBsAg, and all betel quid chewers were 20.57%, 51.80%, 4.53%, and 11.60%, respectively.
Interactive Effect of Betel Quid Chewing and Chronic HBV/HCV Infection on Risk of Cirrhosis
By using HBsAg-negative, anti-HCV-negative betel quid non-users as a reference group, the risk for cirrhosis increased significantly in subjects with HBsAg alone or subjects with anti-HCV alone, or in patients coinfected with HBV/HCV infection (Table 4). It is noteworthy that betel quid chewing alone also conferred a significantly higher risk for developing cirrhosis (OR 5.45; 95% CI 1.36–21.23). Among habitual betel quid chewers, the risk for cirrhosis in subjects with HBsAg alone was higher than in those without HBV/HCV infection (OR 14.4; 95% CI 1.82–95.69). Betel quid chewers with anti-HCV alone also had a higher risk for developing cirrhosis compared with subjects without HBV/HCV infection (p = 0.024, Fishers exact test).
Table 5 displays the interactive effect between betel quid chewing and HCV infection. By using anti-HCV-negative betel quid non-users as a reference group, either betel quid chewing or presence of anti-HCV was an independent risk factor for cirrhosis. The highest OR was found in anti-HCV-positive betel quid chewers. Calculation of synergy index indicated that there was an additive interaction between betel quid chewing and chronic HCV infection. Similarly, the risk for developing cirrhosis was strongly associated with the presence of HBsAg and chewing betel quid (Table 6). Moreover, HBsAg-positive betel quid chewers had the highest OR, and a synergy index of 2.76. These results indicate an additive interaction between betel quid chewing and chronic HBV/HCV infection on risk of cirrhosis.
Characteristics of Betel Quid Chewing in Patients and Controls
All betel quid chewers chewed areca nut. Chewing with betel leaf or unripe betel fruit was strongly associated with increased risk for cirrhosis (Table 7). Chewing betel quid for a duration of more than 20 years was an independent risk factor for developing cirrhosis (OR 6.21; 95% CI 1.97–17.23). Moreover, the longer the duration of betel quid chewing, the higher the risk for developing cirrhosis (p for trend <0.0001; see Table 7).
The median number of total betel quids consumed in cirrhotic patients (204,400 quids; range, 21,900–849,000 quids) was higher than that in controls (54,750 quids; range, 10,950–548,000 quids) (p = 0.0001). There was an increased risk for developing cirrhosis in subjects who consumed more than 100,000 quids (OR 6.10; 95% CI 2.17–18.48). There was a positive linear trend between betel quids consumed and the risk for cirrhosis (p for trend <0.0001; see Table 7).
Characteristics of Cirrhotic Patients by Betel Quid Chewing Status
As shown in Table 8, cirrhotic patients who chewed betel quid were predominantly male (OR 9.39; 95% CI 1.24–70.88; p = 0.004) and tended to be Child-Pugh grade B or C (OR 5.23; 95% CI 2.30–11.92; p = 0.0001). Betel quid chewers frequently had habits of smoking (OR 16.02; 95% CI 4.44–44.12; p = 0.0001) and alcohol drinking (OR 8.73; 95% CI 3.88–19.63; p = 0.0001). It is noteworthy that among the 5 betel quid chewers who had cirrhosis without hepatitis, 3 were habitual alcohol drinkers and 4 were habitual smokers. Multivariate analysis with stepwise logistic regression indicated that habitual smoking (OR 13.52; 95% CI 3.57–51.16; p = 0.0001), alcohol drinking (OR 4.29; 95% CI 1.63–11.29; p = 0.003), and Child-Pugh grade B or C (OR 5.53; 95% CI 2.14–14.25; p = 0.0001) were independent risk factors for habitual betel quid chewing.
To our knowledge, this is the first case-control study to show the association between betel quid chewing and risk for cirrhosis (see Tables 2 and 3). However, the risk for cirrhosis in betel quid chewers is still significantly lower than that for subjects with single HBV or HCV infection alone (see Table 4). The estimated population-attributable risk also shows this trend. Based on this observation and our previous study 61,62, although betel quid chewing is a risk factor for cirrhosis, chronic HBV/HCV infection is still the most important risk factor for cirrhosis in this area hyperendemic for hepatotropic viruses (see Table 4). In betel quid chewers without HBV/HCV infection, the association between betel quid chewing and cirrhosis might not be strong. Although the odds ratio was greater than 1, the small number of subjects (5 patients and 8 controls) reduces the certainty of this association (see Table 4).
Since betel quid chewing has never before been shown to be a risk factor for cirrhosis, it is important to validate that our finding is not due to confounding bias. Betel quid chewing is an island-wide, popular habit of Taiwanese. The prevalence of betel quid chewing in our controls was 5.2% (see Table 2). Based on data estimated from 1,299 subjects collected from the same community as our study, the prevalence of betel quid chewers was 6% (95% CI 4.6–7.1; inclusive of all ages and sexes) 30. Another report from the same community indicated that there was a prevalence rate of 6.5% for daily chewers among 511 men, whereas none of 651 women chewed quid daily 8. The prevalence of betel quid chewing in our control group fell within the 95% CI values of these 2 reports. The prevalence of HBsAg (18.6%) and anti-HCV (4.3%) in our healthy controls showed no significant difference from those in volunteer blood donors 66 or community controls in the same area 61–64,67. Therefore, our controls might be representative of the general population in this community. On the other hand, previous reports have indicated that male gender and older age were associated with a higher prevalence of betel quid chewing 8,30. To control for the possible confounding effect of age and sex, we enrolled study subjects by matching and adjusted our results by multivariate analysis. In the current study, cirrhotic patients who chewed betel quid were male predominant. Moreover, there was a close association between habitual betel quid chewing and habitual smoking, alcohol drinking, and more severe liver damage in patients with cirrhosis (see Table 8).
Our results indicate that a person who chewed betel quid was usually also a habitual smoker and alcohol drinker (see Table 8). It is reasonable that betel quid chewing might indeed be merely a marker for greater alcohol or cigarette consumption.
Betel quid chewing is popular island-wide in Taiwan 30. The habit of betel quid chewing appears to be acquired usually at junior high school between the ages of 12 and 15 years 8. In the current study, our patients usually began chewing betel quid many years before cirrhosis developed. They began chewing betel quid at a median age of 34 years (range, 11–60 yr). Moreover, although none of our patients began chewing betel quid after the diagnosis of cirrhosis (data not shown), it may be that cirrhotic patients are more likely to chew betel quid.
In the current study, there was no significant difference among the ages when patients with cirrhosis began chewing betel quid, drinking alcohol, or smoking. A report from Taiwan 9 indicated that betel quid chewers were more likely than others to engage in high-risk behaviors that could lead to HBV/HCV infection. It is possible there are other confounders for cirrhosis that we did not study.
Although there was a significant difference in the total amount of betel quid chewing in cirrhotic patients and in controls, there was a large overlap between them. As shown in Table 7, the type of betel quid ingredients, the duration of betel quid chewing, and probably underlying liver disease or genetic predisposition might explain why healthy people and cirrhotic patients could consume varying quantities of betel quids with such different outcomes.
The pathogenic mechanisms for the association between habitual betel quid chewing and the risk of cirrhosis are largely unknown. Areca nut consumption may modulate the function of the hepatic detoxification system and increase the risk of toxic hepatitis 50,57. Chronic betel quid feeding in animals causes persistent hepatocyte necroinflammation 50. In this study, habitual betel quid chewers had more severe liver damage, as evidenced by their significantly higher frequency of being Child-Pugh grade B or C (see Table 8). This observation confirms that episodic necroinflammation with subsequent persistent liver injury is important in inducing cirrhosis 26. Alternatively, the immunosuppressive action of areca nut may facilitate progression of HBV/HCV-associated chronic liver disease 24,51,52. On the other hand, fibrosis is an almost invariable part of chronic liver disease regardless of etiology 19,44. It is the fibrous scarring that leads to architectural distortion and cirrhosis. Concurrent hepatic insult by more than 1 agent, such as HCV and alcohol 4,44 or coinfection with other viruses 19, is synergistic for the progression of fibrosis. Moreover, oxygen-derived free radicals and other reactive oxygen species have been implicated as important mediators of hepatic fibrogenesis in liver injury 22,68. These compounds can be found in inflammatory byproducts derived from habitual betel quid chewing 7,35,39–41, alcoholic liver disease, or chronic HBV/HCV infection 17,21. Taken together, persistent necroinflammation and progressive hepatic fibrogenesis may contribute, at least in part, to the additive interaction between habitual betel quid chewing and chronic HBV/ HCV infection. It would be instructive to know how long the betel quid chewers had been infected with HBV/HCV before developing cirrhosis. It is possible that betel quid accelerates progression of cirrhosis.
Aflatoxins are mycotoxins produced by the fungus Aspergillus flavus. This fungus contaminates inappropriately processed and stored food, such as corn, peanuts, and rice, which are stored for prolonged periods under hot, humid conditions. This is a significant problem in less-developed countries within the boundaries of the tropical latitudes including Taiwan 47. Aflatoxin B1 is the most common aflatoxin contaminant and has been found to be the most hepatotoxic 2,33,47. It was observed that 37.5% of betel nut samples were infested with Aspergillus flavus46. Habitual betel quid chewing may be 1 source of aflatoxin B1 ingestion in humans. Since the recognition of the hepatotoxic potential of aflatoxins, initially in poultry and other animals in 1960, aflatoxins have been implicated in the genesis of various forms of liver injury. Acute fatty liver 47 and toxic hepatitis 2,33,48 are some of the more acute variants of presentation, whereas cirrhosis and hepatocellular carcinoma are more chronic manifestations. Aflatoxin B1-induced acute hepatotoxicity in humans has followed the ingestion of contaminated foods. Cirrhosis has been induced in some animals after repeated exposure to aflatoxins 42. The role of aflatoxin in the production of cirrhosis in humans remains inconclusive, although suspected in some settings. Unexplained cryptogenic cirrhosis and childhood cirrhosis in developing countries may be attributed in part to aflatoxin toxicity 69.
Based on the above discussion, we hypothesize that repetitive liver cell injury from oxygen-derived free radicals and other reactive oxygen species generated by habitual betel quid chewing may increase the risk of cirrhosis. This may explain the additive interaction between habitual betel quid chewing and chronic HBV/HCV infection. In conclusion, both betel quid chewing and chronic HBV/HCV infection are independent risk factors for cirrhosis. There is an additive interaction between betel quid chewing and chronic HBV/HCV infection on the risk of cirrhosis. Whether abstinence of betel quid chewing in subjects with chronic HBV/HCV infection decreases the risk for cirrhosis warrants further study.
1. Becker U, Gronbaek M, Johansen D, Sorensen TI. Lower risk for alcohol-induced cirrhosis in wine drinkers. Hepatology. 2002; 35:868–875.
2. Bhat RV. Aflatoxin and liver injury. Trop Gastroenterol. 1996; 17:153–160.
3. Bortolotti F, Jara P, Crivellaro C, Hierro L, Cadrobbi P, Frauca E, Camarena C, De La Vega A, Diaz C, De Moliner L, Noventa F. Outcome of chronic hepatitis B in Caucasian children during a 20-year observation period. J Hepatol. 1998; 29:184–190.
4. Brechot C, Nalpas B, Feitelson MA. Interaction between alcohol and hepatitis viruses in the liver. Clin Lab Med. 1996; 16:273–287.
5. Caldwell SH, Oelsner DH, Iezzoni JC, Hespenheide EE, Battle EH, Driscoll CJ. Cryptogenic cirrhosis: Clinical characterization and risk factors for underlying disease. Hepatology. 1999; 29:664–669.
6. Chan HL, Leung NW, Hussain M, Wong ML, Lok AS. Hepatitis B e antigen-negative chronic hepatitis B in Hong Kong. Hepatology. 2000; 31:763–768.
7. Chen CL, Chi CW, Liu TY. Hydroxyl radical formation and oxidative DNA damage induced by areca quid in vivo. J Toxicol Environ Health A. 2002; 65:327–336.
8. Chen JW, Shaw JH. A study on betel quid chewing behavior among Kaohsiung residents aged 15 years and above. J Oral Pathol Med. 1996; 25:140–143.
9. Chen KT, Chen CJ, Fagot-Campagna A, Narayan KM. Tobacco, betel quid, alcohol, and illicit drug use among 13- to 35-year-olds in ILan, rural Taiwan: Prevalence and risk factors. Am J Public Health. 2001; 91:1130–1134.
10. Corrao G, Torchio P, Zambon A, D’Amicis A, Lepore AR, di Orio F. Alcohol consumption and micronutrient intake as risk factors for liver cirrhosis: A case-control study. Ann Epidemiol. 1998; 8:154–159.
11. Corrao G, Torchio P, Zambon A, Ferrari P, Arico S, di Orio F. Exploring the combined action of lifetime alcohol intake and chronic hepatotropic virus infections on the risk of symptomatic liver cirrhosis. Collaborative Groups for the Study of Liver Diseases in Italy. Eur J Epidemiol. 1998; 14:447–456.
12. Corrao G, Zambon A, Torchio P, Arico S, La Vecchia C, di Orio F. Attributable risk for symptomatic liver cirrhosis in Italy. Collaborative Groups for the Study of Liver Diseases in Italy. J Hepatol. 1998; 28:608–614.
13. del Olmo JA, Serra MA, Rodriguez F, Escudero A, Gilabert S, Rodrigo JM. Incidence and risk factors for hepatocellular carcinoma in 967 patients with cirrhosis. J Cancer Res Clin Oncol. 1998; 124:560–564.
14. Dieter HH, Schimmelpfennig W, Meyer E, Tabert M. Early childhood cirrhoses (ECC) in Germany between 1982 and 1994 with special consideration of copper etiology. Eur J Med Res. 1999; 4:233–242.
15. Di Marco V, Lo Iacono O, Camma C, Vaccaro A, Giunta M, Martorana G, Fuschi P, Almasio PL, Craxi A. The long-term course of chronic hepatitis B. Hepatology. 1999; 30:257–264.
16. Du CL, Wang JD. Increased morbidity odds ratio of primary liver cancer and cirrhosis of the liver among vinyl chloride monomer workers. Occupation Environ Med. 1998; 55:528–532.
17. Farinati F, Cardin R, Degan P, De Maria N, Floyd RA, Van Thiel DH, Naccarato R. Oxidative DNA damage in circulating leukocytes occurs as an early event in chronic HCV infection. Free Radic Biol Med. 1999; 27:1284–1291.
18. Fletcher LM, Dixon JL, Purdie DM, Powell LW, Crawford DH. Excess alcohol greatly increases the prevalence of cirrhosis in hereditary hemochromatosis. Gastroenterology. 2002; 122:281–289.
19. Friedman SL. Hepatitis fibrosis. In: Schiff ER, Sorrell MF, Maddrey WC, eds. Schiff’s disease of the liver, 8th ed. Philadelphia: Lippincott-Raven, 1999. pp 371–385.
20. Guido M, Fagiuoli S, Tessari G, Burra P, Leandro G, Boccagni P, Cillo U, Naccarato R, Rugge M. Histology predicts cirrhotic evolution of post transplant hepatitis C. Gut. 2002; 50:697–700.
21. Hagen TM, Huang S, Curnutte J, Fowler P, Martinez V, Wehr CM, Ames BN, Chisari FV. Extensive oxidative DNA damage in hepatocytes of transgenic mice with chronic active hepatitis destined to develop hepatocellular carcinoma. Proc Natl Acad Sci U S A. 1994; 911:12808–12812.
22. Houglum K, Filip M, Witztum JL, Chojkier M. Malondiadehyde and 4-hydroxynonenal protein adducts in plasma and liver of rats with iron overload. J Clin Invest. 1990; 86:1991–1995.
23. Hsu YS, Chien RN, Yeh CT, Sheen IS, Chiou HY, Chu CM, Liaw YF. Long-term outcome after spontaneous HBeAg seroconversion in patients with chronic hepatitis B. Hepatology. 2002; 35:1522–1527.
24. Hung SL, Chen YL, Wan HC, Liu TY, Chen YT, Ling LJ. Effects of areca nut extracts on the functions of human neutrophils in vitro. J Periodontal Res. 2000; 35:186–193.
25. Huo TI, Wu JC, Lee PC, Tsay SH, Chang FY, Lee SD. Diabetes mellitus as a risk factor of liver cirrhosis in patients with chronic hepatitis B virus infection. J Clin Gastroenterol. 2000; 30:250–254.
26. Idilman R, De Maria N, Colantoni A, Van Thiel DH. Pathogenesis of hepatitis B and C-induced hepatocellular carcinoma. J Viral Hepatitis. 1998; 5:285–299.
27. Ikeda K, Saitoh S, Suzuki Y, Kobayashi M, Tsubota A, Koida I, Arase Y, Fukuda M, Chayama K, Murashima N, Kumada H. Disease progression and hepatocellular carcinogenesis in patients with chronic viral hepatitis: A prospective observation of 2215 patients. J Hepatol. 1998; 28:930–938.
28. Jeffers L. Hepatocellular carcinoma: An emerging problem with hepatitis C. J Natl Med Assoc. 2000; 92:369–371.
29. Kato Y, Hamasaki K, Aritomi T, Nakao K, Nakata K, Eguchi K. Most of the patients with cirrhosis in Japan die from hepatocellular carcinoma. Oncol Rep. 1999; 6:1273–1276.
30. Ko YC, Chiang TA, Chang SJ, Hsieh SF. Prevalence of betel quid chewing habit in Taiwan and related sociodemographic factors. J Oral Pathol Med. 1992; 21:261–264.
31. Kobayashi M, Ikeda K, Saitoh S, Suzuki F, Tsubota A, Suzuki Y, Arase Y, Murashima N, Chayama K, Kumada H. Incidence of primary cholangiocellular carcinoma of the liver in Japanese patients with hepatitis C virus-related cirrhosis. Cancer. 2000; 88:2471–2477.
32. Kondili LA, Tosti ME, Szklo M, Costantino A, Cotichini R, Resuli B, Rapicetta M, Mele A. The relationships of chronic hepatitis and cirrhosis to alcohol intake, hepatitis B and C, and delta virus infection: A case-control study in Albania. Epidemiol Infect. 1998; 121:391–395.
33. Krishnamachari KA, Bhat RV, Nagarajan V, Tilak TB. Hepatitis due to aflatoxicosis: An outbreak in western India. Lancet. 1975; 1:1061–1063.
34. Law MG. Modelling the hepatitis C virus epidemic in Australia. Hepatitis C Virus Projections Working Group. J Gastroenterol Hepatol. 1999; 14:1100–1107.
35. Liu TY, Chen CL, Chi CW. Oxidative damage to DNA induced by areca nut extract. Mutat Res. 1996; 367:25–31.
36. Muir AJ. The natural history of hepatitis C viral infection. Semin Gastrointest Dis. 2000; 11:54–61.
37. Muller T, Schafer H, Rodeck B, Haupt G, Koch H, Bosse H, Welling P, Lange H, Krech R, Feist D, Muhlendahl KE, Bramswig J, Feichtinger H, Muller W. Familial clustering of infantile cirrhosis in Northern Germany: A clue to the etiology of idiopathic copper toxicosis. J Pediatr. 1999; 135:189–196.
38. Muretto P, Angelucci E, Lucarelli G. Reversibility of cirrhosis in patients cured of thalassemia by bone marrow transplantation. Ann Intern Med. 2002; 136:667–672.
39. Nair UJ, Floyd RA, Nair J, Bussachini V, Friesen M, Bartsch H. Formation of reactive oxygen species and of 8-hydroxydeoxyguanosine in DNA in vitro with betel quid ingredients. Chem Biol Interact. 1987; 63:157–169.
40. Nair UJ, Friesen M, Richard I, MacLennan R, Thomas S, Bartsch H. Effect of lime composition on the formation of reactive oxygen species from areca nut extract in vitro. Carcinogenesis. 1990; 11:2145–2148.
41. Nair UJ, Nair J, Friesen MD, Bartsch H, Ohshima H. Ortho- and meta-tyrosine formation from phenylalanine in human saliva as a marker of hydroxyl radical generation during betel quid chewing. Carcinogenesis. 1995; 16:1195–1198.
42. Newberne PM, Butler WH. Acute and chronic effects of aflatoxin on the liver of domestic and laboratory animals: A review. Cancer Res. 1969; 29:236–250.
43. Norton SA. Betel: Consumption and consequences. J Am Acad Derm. 1998; 38:81–88.
44. Poynard T, Bedossa P, Opolon P. Nature history of liver fibrosis progression in patients with chronic hepatitis C. Lancet. 1997; 349:825–832.
45. Pugh RN, Murray-Lyon IM, Dawson JL, Peitroni MC, Williams R. Transection of the esophagus for bleeding esophageal varices. Br J Surg. 1973; 60:646–649.
46. Raisuddin S, Misra JK. Aflatoxin in betel nut and its control by use of food preservatives. Food Add Contam. 1991; 8:707–712.
47. Ramkumar D, LaBrecque DR. Drug-induced liver disease and environmental toxins. In: Zakim D, Boyer TD, eds. Hepatology: A textbook of liver disease, 4th ed. Philadelphia: Saunders, 2003. pp 755–832.
48. Rastogi R, Srivastava AK, Srivastava M, Rastogi AK. Hepatocurative effect of picroliv and silymarin against aflatoxin B1 induced hepatotoxicity in rats. Planta Med. 2000; 66:709–713.
49. Rothman KJ. Modern epidemiology. Boston: Little, Brown, 1986. pp 311–326.
50. Sarma AB, Chakrabarti J, Chakrabarti A, Banerjee TS, Roy D, Mukherjee D, Mukherjee A. Evaluation of pan masala for toxic effects on liver and other organs. Food Chem Toxicol. 1992; 30:161–163.
51. Selvan RS, Selvakkumaran M, Rao AR. Influence of arecoline on immune system. II. Suppression of thymus-dependent immune responses and parameter of non-specific resistance after short-term exposure. Immunopharmacol Immunotoxicol. 1991; 13:281–309.
52. Shahabuddin S, Rao AR. Effect of arecoline on the humoral and cell mediated immune response in mice. Indian J Exp Biol. 1980; 18:1493–1494.
53. Sherlock S, Dooley J. Hepatic cirrhosis. In: Disease of the liver and biliary system. Oxford: Blackwell, 2002. pp 365–80.
54. Sherlock S, Dooley J. Malignant liver tumors. In: Disease of the liver and biliary system. Oxford: Blackwell, 2002. pp 537–562.
55. Shindo M, Ken A, Okuno T. Varying incidence of cirrhosis and hepatocellular carcinoma in patients with chronic hepatitis C responding differently to interferon therapy. Cancer. 1999; 85:1943–1950.
56. Simonetti RG, Camma C, Fiorello F, Politi F, D’Amico G, Pagliaro L. Hepatocellular carcinoma: A worldwide problem and the major risk factors. Dig Dis Sci. 1991; 36:962–972.
57. Singh A, Rao AR. Evaluation of the modifying influence of arecanut on the garlic-modulated hepatic detoxication system enzymes, sulfhydryl content, and lipid peroxidation in mice. Teratog Carcinog Mutagen. 1995; 15:127–134.
58. Sorensen HT, Friis S, Olsen JH, Thulstrup AM, Mellemkjaer L, Linet M, Trichopoulos D, Vilstrup H, Olsen J. Risk of liver and other types of cancer in patients with cirrhosis: a nationwide cohort study in Denmark. Hepatology. 1998; 28:921–925.
59. Tanaka K, Sakai H, Hashizume M, Hirohata T. A long-term followup study on risk factors for hepatocellular carcinoma among Japanese patients with liver cirrhosis. Jpn J Cancer Res. 1998; 89:1241–1250.
60. Tanner MS. Role of copper in Indian childhood cirrhosis. Am J Clin Nutr. 1998; 67(Suppl 5):1074S–81S.
61. Tsai JF, Chang WY, Jeng JE, Ho MS, Wang LY, Hsieh MY, Chen SC, Tsai JH. Hepatitis C virus infection as a risk factor for nonalcoholic liver cirrhosis in Taiwan. J Med Virol. 1993; 41:296–300.
62. Tsai JF, Chang WY, Jeng JE, Ho MS, Lin ZY, Tsai JH. Hepatitis B and C virus infection as risk factors for liver cirrhosis and cirrhotic hepatocellular carcinoma: A case-control study. Liver. 1994; 14:98–102.
63. Tsai JF, Jeng JE, Ho MS, Chang WY, Lin ZY, Tsai JH. Hepatitis B and C virus infection as risk factors for hepatocellular carcinoma in Chinese: A case-control study. Int J Cancer. 1994; 56:619–621.
64. Tsai JF, Jeng JE, Ho MS, Chang WY, Hsieh MY, Lin ZY, Tsai JH. Additive effect modification of hepatitis B surface antigen and e antigen on the development of hepatocellular carcinoma. Br J Cancer. 1996; 73:1498–1502.
65. Tsai JF, Jeng JE, Ho MS, Chang WY, Hsieh MY, Lin ZY, Tsai JH. Effect of hepatitis C and B virus infection on risk of hepatocellular carcinoma: A prospective study. Br J Cancer. 1997; 76:968–974.
66. Tsai JF, Jeng JE, Ho MS, Wang CS, Chang WY, Hsieh MY, Lin ZY, Tsai JH. Serum alanine aminotransferase level in relation to hepatitis B and C virus infections among blood donors. Liver. 1997; 17:24–29.
67. Tsai JF, Jeng JE, Ho MS, Chang WY, Lin ZY, Tsai JH. Independent and additive effect modification of hepatitis C and B viruses infection on the development of chronic hepatitis. J Hepatol. 1996; 24:271–276.
68. Tsukamoto H. Oxidative stress, antioxidants, and alcoholic liver fibrogenesis. Alcohol. 1993; 10:465–467.
69. Zimmerman HJ. Drug-induced liver disease. In: Schiff ER, Sorrelle MF, Maddrey WC, eds. Schiff’s disease of the liver, 8th ed. Philadelphia: Lippincott-Raven, 1999. pp 973–1064.