Safe levels of cadmium intake to prevent renal toxicity in human subjects.

The present review attempts to provide an update of the scientific knowledge on the renal toxicity which occurs in human subjects as a result of chronic ingestion of low-level dietary Cd. It highlights important features of Cd toxicology and sources of uncertainty in the assessment of health risk due to dietary Cd. It also discusses potential mechanisms for increased susceptibility to Cd toxicity in individuals with diabetes. Exposure assessment on the basis of Cd levels in foodstuffs reveals that vegetables and cereals are the main sources of dietary Cd, although Cd is also found in meat, albeit to a lesser extent. Cd accumulates particularly in the kidney and liver, and hence offal contains relatively high amounts. Fish contains only small quantities of Cd, while crustaceans and molluscs may accumulate larger amounts from the aquatic environment. Data on Cd accumulation in human kidney and liver obtained from autopsy studies are presented, along with results of epidemiological studies showing the relationship between renal tubular dysfunction and kidney Cd burden. These findings suggest that a kidney Cd level of 50 microg/g wet weight is a maximum tolerable level in order to avoid abnormal kidney function. This renal Cd burden corresponds to a urinary Cd excretion of 2 microg/d. Accordingly, safe daily levels of Cd intake should be kept below 30 microg per person. Individual variations in Cd absorption and sensitivity to toxicity predicts that a dietary Cd intake of 30 microg/d may result in a slight renal dysfunction in about 1% of the adult population. The previous guideline for a maximum recommended Cd intake of 1 microg/kg body weight per d is thus shown to be too high to ensure that renal dysfunction does not occur as a result of dietary Cd intake.

Relationships between the synthesis of N-nitrosodimethylamine and immune responses to chronic infection with the carcinogenic parasite, Opisthorchis viverrini, in men.

This study investigated the relationship between immune responses to infection with the liver fluke, Opisthorchis viverrini, and the synthesis of the carcinogen, N-nitrosodimethylamine (NDMA) in humans. It also examined associations between synthesis of nitric oxide (NO) and nitrosation of amines, in vivo. Antibody and T cell responses to fluke antigens and post-alcohol urinary NDMA excretion were assessed among three groups of 40-50 men with no, moderate and heavy liver fluke infection. Markers of NO synthesis (nitrate, nitrite) and nitrosation (nitrosamino acids) were also measured in biological fluids. Assessments were carried out under controlled conditions which minimised intake of exogenous nitrate and nitrite and were carried out at two time points, namely before and 4 months after elimination of the infection with praziquantel treatment. No statistically significant variation was observed in the amount of NDMA excreted between the 3 groups. However, during active infection, a strong negative association was observed between in vitro lymphoproliferative responses to some liver fluke antigens and NDMA excretion. After treatment this association was reduced. Multivariate statistical models revealed a highly significant relationship between NDMA levels and urinary nitrate, stimulation indices for two T cell responses to two parasite antigens (MW 37 kDa and 110 kDa) and gall bladder dimensions. NDMA levels after treatment were best described by the ratio between parasite-specific IgG2 and IgE, background levels of T cell proliferation, a urinary marker of nitrosation (N-nitrosothioproline) and usual level of alcohol consumption. These results suggest that individual background immunologic activity, parasite-specific responses and/or parasite products and NO synthesis are important determinants of endogenous generation of nitrosamines in O. viverrini-infected humans.

Induction of cytochrome P450 2A6 expression in humans by the carcinogenic parasite infection, opisthorchiasis viverrini.

The purpose of this study was to examine in vivo the activity of cytochrome P450 (CYP) 2A6, an enzyme capable of activating carcinogens, including N-nitrosodimethylamine, in humans with the carcinogenic liver fluke infection, opisthorchiasis viverrini, before and after treatment with the antiparasitic agent, praziquantel. Coumarin hydroxylase activity of CYP 2A6 was assessed by administering a probe drug, coumarin, and measuring its metabolite, 7-hydroxycoumarin, in urines collected between 0-2 h and 2-4 h of 106 people with varying intensities of Opisthorchis viverrini infection. Five individuals who did not excrete any detectable 7-hydroxy coumarin (and have a genetic defect probably leading to an absence of catalytic activity of the CYP 2A6 protein) were excluded from analysis. Infected people excreted an average of 22.7 mumol of 7-hydroxycoumarin in the first 2 h after taking the drug, whereas the mean of the uninfected group was 19.4 mumol; this difference did not reach statistical significance (P = 0.10). However, a highly significant increase in CYP 2A6-related activity was observed in infected individuals who also had radiological evidence of biliary fibrosis (28.1 mumol) compared to those without (19.4 mumol; P = 0.01). Reassessments of coumarin hydroxylase activity of CYP 2A6 made 2 months after praziquantel treatment showed highly significant reductions in the amount of 7-hydroxycoumarin excreted among the infected groups but no difference in the uninfected group. These results suggest that expression of CYP 2A6 is induced among chronically infected people who also have fibrosis of the intrahepatic bile duct. As already demonstrated in an animal model and now observed in humans for the first time, this increase in CYP 2A6-related enzyme activity may represent an important mechanistic link between inflammatory products of chronic liver fluke infection (e.g., DNA alkylation damage from endogenously formed N-nitrosamines) and the high risk of cholangiocarcinoma faced by infected individuals.

Cross-sectional patterns of hepatobiliary abnormalities and possible precursor conditions of cholangiocarcinoma associated with Opisthorchis viverrini infection in humans.

The liver fluke, Opisthorchis viverrini, is both highly prevalent and closely associated with cholangio-carcinoma in northeast Thailand. This study measured associations between intensity of liver fluke infection and nonmalignant hepatobiliary disease diagnosed by ultrasonography among 1, 807 largely asymptomatic adult residents drawn from endemic communities. Abnormalities significantly associated with intensity of infection included gallbladder enlargerment in all dimensions, presence of sludge, irregular gallbladder wall, liver enlargement, and enhanced portal vein radicle echoes. While gallbladder enlargement was not sex-specific, the prevalence odds of the other abnormalities were 2-3 times higher among males compared with females. Those recently treated with the anthelmintic praziquantel had higher odds of these abnormalities compared with others with the same infection status who were untreated. The low prevalence of gallstones suggests that this impairment of gallbladder structure and function does not frequently stimulate gallstone formation. However, gallbladder disturbances, together with chronic inflammation and fibrosis of the bile ducts, which are visualized as enhanced portal vein radicle echoes, may contribute to the strikingly enhanced susceptibility to cholangiocarcinoma among people, especially males, with heavy liver fluke infection.

Thiocyanate-independent nitrosation in humans with carcinogenic parasite infection.

Infection with the liver fluke, Opisthorchis viverrini, is a causative agent of cholangiocarcinoma. One possible contributing factor in this carcinogenesis is the chronic, local generation of nitric oxide by inflammatory cells expressing inducible nitric oxide synthase and the production of N-nitroso compounds via the reaction between amines and nitrosating agents derived from nitric oxide. Our previous studies provided evidence that nitric oxide synthesis is elevated during human liver fluke infection. Here we present data on the same sample of men which definitively demonstrates increased nitrosation of proline and thioproline (thiazolidine-4-carboxylic acid) among infected men compared to uninfected control subjects on a low nitrate diet. This difference was specifically abolished by co-administration of ascorbic acid with proline and by elimination of parasites by praziquantel treatment. Multivariate statistical models demonstrate the importance of salivary thiocyanate levels to variation in the nitrosation of proline among uninfected individuals, but not among those with current fluke infection. This suggests that considerable generation of nitrosating agents (N203/N204) in infected people may be occurring via oxidation of arginine by nitric oxide synthase in inflamed tissue which is thiocyanate insensitive. Analyses revealed positive associations between N-nitrosoproline excretion and nitrate/nitrite levels in urine, plasma and saliva and with usual alcohol intake; with variation in these trends between groups. In conclusion, we have confirmed the relationship between O.viverrini infection and enhanced endogenous nitrosation, showing evidence of its extragastric site. New information is also provided on the determinants of N-nitrosamino acid excretion in men on a controlled low nitrate diet without smoking, conditions which reduce exogenous sources of nitrosating agents.

Cross-sectional study of Opisthorchis viverrini infection and cholangiocarcinoma in communities within a high-risk area in northeast Thailand.

We describe an innovative strategy to quantify risk of cancer associated with varying levels of exposure to chronic parasitic infection through the identification of asymptomatic cases of cholangiocarcinoma within a population-based survey of Opisthorchis viverrini infection. Stool samples from 12,311 adults over age 24 years from 85 villages in northeast Thailand were examined for intensity of liver fluke infection. People from varying egg count categories were selected for ultrasound examination to identify hepatobiliary disease. Fifteen preclinical cases of cholangiocarcinoma were diagnosed from a total of 1,807 people based on ultrasonographic evidence with confirmation by endoscopy where possible. The prevalence odds of the diagnosis of cholangiocarcinoma increased gradually within the light and moderate intensity groups. In contrast, sharply elevated prevalence odds [age-, sex- and locality-adjusted prevalence odds ratio (POR) 14.1, p < 0.05] were observed within the most heavily liver fluke-infected group compared with the uninfected group. Males were more frequently affected than females (crude POR 4.5), but after controlling for intensity of infection, age and locality, the magnitude and significance of this measurement was reduced. Our data clearly demonstrate a significant relationship between intensity of liver fluke infection and cholangiocarcinoma and a strikingly high prevalence of the disease among heavily infected males.

Parasite-associated morbidity: liver fluke infection and bile duct cancer in northeast Thailand.

Infection with the liver fluke, Opisthorchis viverrini, remains a major public health problem in Northeast Thailand, where approximately one-third of the population is infected. The northeast region is largely populated by Laos-descendent Thais who enjoy eating raw fish, which harbour the infective stage of the fluke. The parasite has maintained its presence in the population despite the widespread use of praziquantel and dissemination of health education material throughout the region by vigorous government-sponsored programs in recent years. The most severe consequence of liver fluke infection is cholangiocarcinoma, i.e. cancer of the bile duct epithelium. Although mortality due to the parasites alone appears to be uncommon, cholangiocarcinoma arising as a result of infection is one of the leading causes of death in the region. This paper reviews the pathogenesis of infection and the geographic, hospital-based and community studies which demonstrate the close relationship between infection and cancer. In addition, data from the Cancer Registry of Khon Kaen, Northeast Thailand and population-based studies using ultrasonography to visualize early tumours which illuminate the very high frequency of the cancer among heavily infected individuals and communities are discussed. Finally, the paper will close with a brief commentary on the prospects for control of the parasite and its likely impact on the frequency of cancer given the current epidemiological situation of liver fluke infection.

Liver fluke infection and cholangiocarcinoma: model of endogenous nitric oxide and extragastric nitrosation in human carcinogenesis.

Cancers arising during bacterial, viral and parasitic infection provide useful models to investigate the link between inflammation and carcinogenesis. Because the inflammatory agent is known, relationships between immune responses, the production of DNA-damaging agents, such as nitric oxide, oxygen radicles and N-nitroso compounds, and cancer risk can be explored. This paper first describes the close relationship between infection with the liver fluke, Opisthorchis viverrini, and cholangiocarcinoma in humans. Data are then presented which demonstrate an elevation in levels of salivary nitrite and urinary and plasma nitrate among men with moderate and heavy liver fluke infections compared to uninfected controls which was absent 4 months after the parasites were cleared with praziquantel. Because of the strict control over subject selection and dietary intake plus the absence of the increase following treatment, we conclude that the higher levels of nitrate and nitrite reflect endogenous generation of nitric oxide resulting from liver fluke infection. Excess nitric oxide generation in the inflamed tissue is likely to lead directly to the formation of N-nitroso compounds mediated by activated macrophages. Further work will attempt to demonstrate a link between this increase and both parasite-specific immune responses and the risk of cancer.

Reversal of biliary tract abnormalities associated with Opisthorchis viverrini infection following praziquantel treatment.

We recently demonstrated a number of biliary tract abnormalities associated with moderate to heavy Opisthorchis viverrini infection among 95 selected village residents in north-east Thailand, who were treated with praziquantel immediately after initial examination and re-examined by abdominal ultrasonography 10 months later. Dramatic improvement in the gall-bladder status of previously infected individuals was observed, including reduction of gall-bladder length and width and regained contractibility of the organ following fatty meal consumption. The frequency of severe enhanced portal vein radicle echoes and gall-bladder sludge and wall irregularities also decreased following treatment, but persisted in many individuals. Two masses were observed in the right hepatic lobe of one individual who was previously considered normal. The findings suggest that gall-bladder abnormalities are reversible following elimination of liver fluke infection, but malignancies, once initiated, are not likely to be affected by treatment.

Opisthorchis viverrini and cholangiocarcinoma in Northeast Thailand.

Population-based research is beginning to show clearly the devastating impact that Opisthorchis viverrini infection has on the Laos-descendent population of Northeast Thailand who love to eat uncooked meat and fish, one of which being the parasite's intermediate host: cyprinoid fish. Here, Melissa Haswell-Elkins, Paiboon Sithithaworn and David Elkins discuss the parasite's life cycle and epidemiology, highlighting the close relationship between this infection and bile duct carcinoma, which is a major cause of death among adults in rural Northeast Thailand.

Phaneropsolus spinicirrus n. sp. (Digenea: Lecithodendriidae), a human parasite in Thailand.

Phaneropsolus spinicirrus n. sp., collected in a postpraziquantel treatment stool of a 44-yr-old woman from Kalasin Province in northeastern Thailand, is described. It is the second species of the genus Phaneropsolus Looss, 1899, found to parasitize a human host. It differs from the first species, Phaneropsolus bonnei Lie, 1951, in the presence of a short spinose cirrus and the structure and distribution of tegumental spines. The new species is most similar to Phaneropsolus perodictici Goodman and Panesar, 1986, isolated from the potto, Perodicticus potto, in Uganda because digeneans of both species have numerous spines on the cirrus. However, the new species differs from P. perodictici in having more conspicuous spines, the genital pore at the posterior border of the oral sucker or lateral to the pharynx, a body and internal organs of larger size, conical spines on the tegument, an ovoid to bilobed ovary and a large V-shaped excretory bladder. Phaneropsolus spinicirrus is distinguishable from other previously reported Phaneropsolus by possessing a spinose cirrus.

Immune responsiveness and parasite-specific antibody levels in human hepatobiliary disease associated with Opisthorchis viverrini infection.

Opisthorchis viverrini infection is associated with human hepatobiliary disease and cholangiocarcinoma, but the role of the immune response in the pathogenesis of infection is unclear. Here ultrasonography was used to examine the biliary tracts of residents from an endemic community. Delayed-type hypersensitivity responses to unrelated antigens, and fluke-specific IgG and IgA levels in serum of this group were also examined. Relationships between immunological parameters, intensity of infection and radiologically measured variables are reported. Immune responsiveness to unrelated antigens did not vary with intensity of parasite infection or disease status. Of all the variables, IgG levels were most markedly elevated in disease cases compared with normal subjects and were closely associated with gall bladder size and dysfunction. This is consistent with the hypothesis that an immunopathologic mechanism is involved in opisthorchiasis and suggests that antibody levels may be useful in screening populations for fluke-associated hepatobiliary disease.

Cholangiocarcinoma in north east Thailand. A hospital-based study.

The population of Northeast Thailand has one of the highest known rates of cholangiocarcinoma (cancer of the bile ducts). We sought firstly to obtain an estimate of the frequency of hospital-based diagnosis of cholangiocarcinoma in the northeastern province of Khon Kaen. All pathology, ultrasound and in-patient and out-patient records were searched for cases diagnosed in tertiary hospitals in a random calendar month in 1988. A total of 203 persons from various parts of northeastern Thailand were newly-diagnosed in the two hospitals, 75 of whom resided in Khon Kaen province. We estimated the minimal age-standardised annual incidence rate in this province to be 135.4 per 100,000 among males and 43.0 per 100,000 among females (world standard). Truncated standardised incidence rates (ages 35-64 years only) were 334.2 and 104.3 per 100,000 for males and females respectively. Secondly we aimed to assess the feasibility of secondary prevention by early clinical diagnosis. However, the symptoms and signs observed were typically non-specific and associated with advanced disease in patients with cholangiocarcinoma who presented to hospital. Primary preventive measures appear to be the key to decreasing the mortality from this disease.

A high frequency of hepatobiliary disease and suspected cholangiocarcinoma associated with heavy Opisthorchis viverrini infection in a small community in north-east Thailand.

A group of 87 adults from a small village in north-east Thailand was chosen to undergo ultrasound investigation based on their intensity of infection with the liver fluke, Opisthorchis viverrini, or clinical status (history of jaundice, current hepatomegaly). From this group, 8 cases of suspected early cholangiocarcinoma were found, and the diagnostic features of 6 of the 8 were confirmed by computerized tomography scan and endoscopic retrograde cholangiopancreatography. In addition, several cases of mild gall-bladder disease, chronic cholecystitis, cholelithiasis and parenchymal liver disease were detected. A highly significant positive relationship between the intensity of liver fluke worm burden and the severity of biliary tract disease within individuals is reported. These results indicate that Opisthorchis is associated with moderate to severe hepatobiliary disease in a considerable proportion of infected individuals.