Long-Term, Low-Dose Exposure to Microcystin-LR Does not Cause or Increase the Severity of Liver Disease in Rodents.

BACKGROUND: Acute exposure to high concentrations of microcystin-LR (MC-LR) can cause significant hepatocyte injury. AIM: To document the effects of long-term, low-dose MC-LR exposure on hepatic inflammation and fibrosis in mice with healthy and diseased livers. MATERIAL AND METHODS: Male CD1 mice (N = 20/group) were exposed to 1.0 µg/L of MC-LR in drinking water; 1.0 µg/L MC-LR plus 300 mg/L of the hepatotoxin thioacetamide (MC-LR/TAA); or 300 mg/L TAA alone for 28 weeks. Liver biochemistry and histology were documented at the end of the study period. In addition, hepatic stellate cells (HSCs), were exposed in vitro to MC-LR (0.1-10,000 µg/L) and monitored for changes in cell metabolism, proliferation and activation. RESULTS: Liver biochemistry and histology were essentially normal in MC-LR alone exposed mice. MC-LR/TAA and TAA alone exposed mice had significant hepatic inflammation and fibrosis but the extent of the changes were similar in the two groups. In vitro, MC-LR had no effect on HSC metabolism, proliferation or activation. CONCLUSION: Long-term, low-dose exposure to MC-LR is unlikely to lead to chronic liver disease in the setting of a normal liver or exacerbate existing liver disease in the setting of ongoing hepatitis.

Long-term, low-dose exposure to microcystin toxin does not increase the risk of liver tumor development or growth in mice.

AIM: Cyanotoxins are biological toxins produced by cyanobacteria (blue green algae) that have been implicated in the pathogenesis of liver tumors. Based on acute toxicity studies, the World Health Organization has designated 1.0 µg/L of cyanotoxin-contaminated drinking water as the safe allowable limit for daily oral consumption. The aim of this study was to determine whether long-term exposure to this concentration of cyanotoxins is capable of initiating or promoting the growth of liver tumors. METHODS: In the present study, four groups of adult, male CD-1 mice (n = 20/group) were exposed to either drinking water alone (water group), drinking water containing 1.0 µg/L of microcystin-LR (MC-LR group), MC-LR plus the tumor promoter thioacetamide (MC-LR/TAA group) or thioacetamide alone (TAA group). Following 28 weeks of exposure, mice were killed and the livers examined for tumor number and size. RESULTS: No tumors were present in the water or MC-LR alone groups while five mice in the MC-LR/TAA group and four in the TAA alone group developed liver tumors. The mean size of the tumors in the MC-LR/TAA and TAA alone groups were similar as were the results of Ki-67 staining, number of atypical mitoses and liver cancer gene expression profiles. In vitro MC-LR (0.1-1000 µg/L) exposure did not induce malignant transformation of WB-F344 hepatic stem cells or increase the proliferative activity or invasiveness of PLC/PRF/5 malignant hepatocytes. CONCLUSION: The results of this study suggest that long-term, low dose cyanotoxin exposure is unlikely to result in liver tumor development or enhance existing tumor growth.

A lack of association between increases in gastrointestinal, liver, and urologic cancers and cyanobacterial contamination of fresh water lakes in Manitoba.

Cyanobacteria or blue-green algae are becoming increasingly abundant in North American fresh water lakes. Toxins produced by cyanobacteria have been associated with gastrointestinal injury, liver failure, and nephrotoxicity. They have also been implicated in the pathogenesis of gastrointestinal and liver cancers. The purpose of the present study was to determine whether the incidence rates of gastrointestinal, liver, and urologic cancers are increasing in the province of Manitoba and, if so, whether these increases spatially and/or temporally correlate with areas where cyanobacterial contamination of fresh water lakes have been identified. Cancer incidence data were obtained from the Manitoba Cancer Registry. Cyanobacterial contamination data, as reflected by microcystin toxin concentrations, were available from the Manitoba Water Stewardship. ArcGIS mapping was employed to document spatial and temporal relationships between cancer incidence and cyanobacterial data. The results revealed that although the incidence rates for all three cancers have increased over the past 20-25 years, these increases were not disproportionally higher in zones with high microcystin toxin determinations. The results of this study argue against increased exposure to cyanotoxins as an explanation for the increase in gastrointestinal, liver and urologic cancers in Manitoba.

The Geographic Distribution of Liver Cancer in Canada Does Not Associate with Cyanobacterial Toxin Exposure.

BACKGROUND: The incidence of liver cancer has been increasing in Canada over the past decade, as has cyanobacterial contamination of Canadian freshwater lakes and drinking water sources. Cyanotoxins released by cyanobacteria have been implicated in the pathogenesis of liver cancer. OBJECTIVE: To determine whether a geographic association exists between liver cancer and surrogate markers of cyanobacterial contamination of freshwater lakes in Canada. METHODS: A negative binomial regression model was employed based on previously identified risk factors for liver cancer. RESULTS: No association existed between the geographic distribution of liver cancer and surrogate markers of cyanobacterial contamination. As predicted, significant associations existed in areas with a high prevalence of hepatitis B virus infection, large immigrant populations and urban residences. DISCUSSION AND CONCLUSIONS: The results of this study suggest that cyanobacterial contamination of freshwater lakes does not play an important role in the increasing incidence of liver cancer in Canada.

Activation of ß-adrenergic receptors increases the in vitro migration of malignant hepatocytes.

AIM: Activation of adrenergic receptors (AR) has been reported to enhance the growth and invasion of various malignancies. The effects of AR agonists on malignant hepatocyte proliferation and migration have yet to be determined. METHODS: PLC/PRF/5 (PLC) and Huh-7 cells were exposed to a wide range of concentrations of the AR agonists noradrenaline (NA) and isoprenaline. Cell proliferation, migration, intracellular cyclic adenosine monophosphate (cAMP), protein kinase A (PKA) and C (PKC), matrix metalloproteinases (MMP)-2, -3, -7 and -9, and α(1) -, ß(1) - and ß(2) -AR expression were documented in both cell lines. RESULTS: Cell proliferative activity was unaltered following exposure to physiological and stress-related concentrations of AR agonists but migration was accelerated, an effect that was inhibited by the nonselective ß-AR antagonist labetalol. cAMP, PKA, PKC or MMP expression remained unchanged. Although α(1) - and ß(1) -AR expressions were abundant, ß(2) -AR expression was limited in both cell lines. CONCLUSION: Unlike other malignancies studied to date, in this study, the proliferative activity of malignant hepatocytes was not increased by exposure to AR agonists, a finding that could be explained by downregulation of ß(2) -AR expression. The increase in malignant hepatocyte migration observed remains unexplained but does not appear to involve adenyl cyclase or MMP signaling pathways.

Interaction of human immunodeficiency virus type 1 integrase with cellular nuclear import receptor importin 7 and its impact on viral replication.

Similar to all other viruses, human immunodeficiency virus type 1 (HIV-1) depends heavily on cellular factors for its successful replication. In this study we have investigated the interaction of HIV-1 integrase (IN) with several host nuclear import factors using co-immunoprecipitation assays. Our results indicate that IN interacts specifically with host importin 7 (Imp7) in vivo, but does not interact with importin 8 (Imp8) or importin alpha (Rch1). In contrast, another HIV-1 karyophilic protein MAp17, which is capable of binding Rch1, fails to interact with Imp7, suggesting that IN and Map17 may interact with different cellular pathways during HIV-1 replication. Genetic analysis revealed that the C-terminal domain of IN is the region responsible for interaction between IN with Imp7, and an IN mutant (K240A,K244A/R263A,K264A) disrupted the Imp7 binding ability of the protein, indicating that both regions ((235)WKGPAKLLWKG and (262)RRKAK) within the C-terminal domain of IN are required for efficient IN/Imp7 interaction. Using a vesicular stomatitis virus G glycoprotein pseudotyped HIV single-cycle replication system, we showed that the IN/Imp7 interaction-deficient mutant was unable to mediate viral replication and displayed impairment at both viral reverse transcription and nuclear import steps. Moreover, transient knockdown of Imp7 in both HIV-1 producing and target cells resulted in a 2.5-3.5-fold inhibition of HIV infection. Altogether, our results indicate that HIV-1 IN specifically interacts with Imp7, and this viral/cellular protein interaction contributes to efficient HIV-1 infection.