Microcystin-leucine arginine (MC-LR), a potentially carcinogenic toxin, is produced by Cyanobacteria such as Microcystis and Ananabacteria during water bloom. Increasing evidence demonstrated that MC-LR induces male reproductive toxicity, mainly by inducing germ cell apoptosis, destroying cell cytoskeleton, interfering with DNA damage repair pathway, and damaging blood-testicular barrier (BTB), which eventually lead to male sterility. Testicular Sertoli cells are the somatic cells that directly contact with spermatogenic cells in seminiferous tubules. They not only regulate immune response to maintain testicular immune homeostasis by secreting a variety of cytokines and immunosuppressive factors, but also provide the protective effects of spermatogenic cells by forming BTB. MC-LR induces inflammation and apoptosis of Sertoli cells, and destroys the integrity of the BTB, and then causes spermatogenesis dysfunction.
Male ; Humans ; Sertoli Cells ; Leucine/pharmacology* ; Arginine/pharmacology* ; Microcystins/metabolism* ; Immunity

Microcystin is one of the monocyclic heptapeptides produced primarily by microcystis aeruginosa. Recent studies suggest that microcystin can induce cell apoptosis, as well as oxidative stress and mitochondrial alteration. Studies also indicate that Bcl-2 family and p53 may play an important role in the apoptosis induced by microcystin.
Animals ; Apoptosis ; physiology ; Humans ; Microcystins ; toxicity ; Microcystis ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVETo investigate the influence of microcystin-LR (MC-LR) on monocytes and lymphocytes in blood of mice and to find a sensitive index of toxic effects.

METHODSSpecific pathogen free Kunming male mice, aging 1 month-old,were randomly divided into 5 groups by weights, 7 mice for each group. The mice in 5 groups were exposed to MC-LR through intraperitoneal injection at 0, 3.125,6.250, 12.500 and 25.000 µg/kg respectively for 7 days. Then cytokine levels in the serum were measured by radioimmunoassay, DNA-protein crosslinks (DPC) was measured by the SDS/KCl precipitation technique, and the phagocytosis and ROS of leukocytes were detected by flow cytometry.

RESULTSThe levels of interleukin 6 in the 6.250, 12.500 and 25.000 µg·kg(-1)·d(-1) dose groups were (346.837 ± 25.536), (360.847 ± 37.886) and (434.245 ± 35.858)pg/ml respectively, which were significantly higher than those in the control group which the value was (232.775 ± 32.816) pg/ml (t values were -7.258, -6.760 and -10.966 respectively, P values were all < 0.05).While the level of tumor necrosis factor-alpha was(10.782 ± 0.966) fmol/ml in 25 µg·kg(-1)·d(-1) dose group was statistically lower than it in the control group which the value was (16.878 ± 3.378) fmol/ml (t value was 4.591, P < 0.05). The DPC levels of lymphocytes in 6.250, 12.500 µg·kg(-1)·d(-1) dose group were (242.576 ± 7.545),(241.472 ± 2.793) ng/ml,higher than it in the control group while the value was (228.657 ± 4.130) ng/ml (t value was -4.282, -6.801, P values were all <0.05). The fluorescence intensity of DCF in lymphocytes in the 4 treated groups were separately 3299.37 ± 120.54, 3281.38 ± 58.34, 3308.06 ± 136.12 and 3346.92 ± 108.69, all significantly lower than 3770.81 ± 131.39 in the control group (t values were 6.995, 9.007, 6.472 and 6.577 respectively, and P values were all <0.05). The fluorescence intensity of DCF in monocytes in the 4 treated groups (3271.51 ± 140.79, 3270.05 ± 117.92, 3326.90 ± 114.39 and 3292.49 ± 145.97 respectively) were also significantly lower than the value in the control group was 3841.72 ± 130.92 (t values were 7.847, 8.584, 7.835 and 7.411 respectively, P values were all <0.05). There was no significant difference in other index among the four experiment groups and the control group.

CONCLUSIONThe MC-LR administered via intraperitoneal injection to mice induced the alterations of some cytokines of monocytes and lymphocytes in blood. By comparison, the ROS of leukocyte was the most sensitive index.

Animals ; Cytokines ; metabolism ; Lymphocytes ; drug effects ; Male ; Mice ; Mice, Inbred Strains ; Microcystins ; pharmacology ; Monocytes ; drug effects ; Reactive Oxygen Species ; metabolism

To evaluate the public health risk of exposure to microcystins in fish food in China, the distribution pattern of microcystin-LR and microcystin-RR in various organs (liver, intestine, kidney, muscle and lipid) of the dominant freshwater phytoplanktivorous fish Hypophthalmichthys molitrix in Hangzhou, China's Tiesha River was investigated with the method of HPLC-ESI-MS analysis. The distribution of microcystins was different in the fish organs and the major total microcystins (microcystin-LR and microcystin-RR) were present in the intestines (6.49 micro g/g fresh weight), followed by the livers (4.52 micro g/g fresh weight) and the muscles (2.86 micro g/g fresh weight). Microcystins were detected in kidneys (1.35 micro g/g fresh weight), but not detected in lipid. The results suggested that the mean daily intake from fish was 0.03 micro g/kg body weight which was very close to the recommended WHO tolerable daily intake (TDI) level of 0.04 micro g/kg body weight per day, and local people were warned they may have health risk if they consumed fish from the river.
Animals ; Carps ; metabolism ; parasitology ; Fresh Water ; analysis ; parasitology ; Microcystins ; metabolism ; Organ Specificity ; Phytoplankton ; metabolism ; Risk Assessment ; methods ; Risk Factors ; Tissue Distribution ; Water Pollutants, Chemical ; analysis

OBJECTIVETo evaluate the effects of antagonistic action of epigallocatechin-3-gallate (EGCG) on microcystin LR (MC-LR) induced oxidative damage on mice and the expression of cytochrome P450 2E1 (CYP2E1) which was one of phase Iota detoxification enzymes.

METHODSA total of 24 specific pathogen free (SPF) male BALB/c mice were randomly divided into four groups, including control group, MC-LR group, low concentration EGCG group, and high concentration EGCG group. Mice were sacrificed on the 15th day, body weight, and the relative organ weight, liver antioxidant enzyme level and lipid peroxidation product, liver histopathology and CYP2E1 gene and protein expression were detected and analyzed respectively.

RESULTS(1) EGCG could antagonise the liver injury which had been damaged by MC-LR. (2) The malonaldehyde (MDA) level ((2.87 +/- 0.03) nmol/mg prot) and superoxide dismutase (SOD) level ((168.18 +/- 2.86) U/mg prot) in MC-LR group were significantly different when compared with the two EGCG treatment groups (the MDA values of the low and high concentration EGCG group were (2.37 +/- 0.05) nmol/mg prot and (1.44 +/- 0.05) nmol/mg prot, F = 906.63, P < 0.01; the SOD values were (176.55 +/- 2.98) U/mg prot and (184.89 +/- 1.53) U/mg prot, F = 32.32, P < 0.01). (3) MC-LR up-regulated the mRNA and protein expression of CYP2E1 (the mRNA values of MC-LR group and control were 1.41 +/- 0.26, 0.86 +/- 0.13, t = -4.22, P = 0.003; the protein values of MC-LR group and control were 0.24 +/- 0.03, 0.12 +/- 0.02, t = -9.21, P < 0.05). EGCG down-regulated the mRNA (the values of the low and high concentration EGCG group were 1.09 +/- 0.08, 0.99 +/- 0.09, F = 9.03, P = 0.004) and protein expression (the values of the low and high concentration EGCG group were 0.21 +/- 0.03, 0.14 +/- 0.02, F = 24.76, P < 0.05) of CYP2E1 which activated by MC-LR.

CONCLUSIONThe up-regulation of CYP2E1 which induced by MC-LR was inhibited by EGCG intervention. EGCG might antagonize the oxidation damage of hepatocytes in a certain degree.

Animals ; Catechin ; analogs & derivatives ; pharmacology ; Cytochrome P-450 CYP2E1 ; metabolism ; Hepatocytes ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Microcystins ; adverse effects ; Oxidative Stress ; drug effects

Animals ; Gene Expression ; Genes, p53 ; Liver ; drug effects ; metabolism ; pathology ; Liver Neoplasms ; metabolism ; pathology ; Male ; Microcystins ; toxicity ; Mutation ; Rats ; Rats, Wistar

OBJECTIVETo investigate the hispathological characteristics and antioxidant responses in liver of silver carp after intraperitoneal administration of microcystins (MCs) for further understanding hepatic intoxication and antioxidation mechanism in fish.

METHODSPhytoplanktivorous silver carp was injected intraperitoneally (i.p.) with extracted hepatotoxic microcystins (mainly MC-RR and -LR) at a dose of 1000 microg MC-LReq./kg body weight, and liver histopathological changes and antioxidant responses were studied at 1, 3, 12, 24, and 48 h, respectively, after injection.

RESULTSThe damage to liver structure and the activities of hepatic antioxidant enzymes including catalase (CAT), superoxide dismutase (SOD), and glutathione peroxide (GPX) were increased in a time-dependent manner.

CONCLUSIONIn terms of clinical and histological signs of intoxication and LD50 (i.p.) dose of MC-LR, silver carp appears rather resistant to MCs exposure than other fishes. Also, the significantly increased SOD activity in the liver of silver carp suggests a higher degree of response to MCs exposure than CAT and GPX.

Animals ; Antioxidants ; metabolism ; Carps ; metabolism ; Catalase ; metabolism ; Glutathione Peroxidase ; metabolism ; Injections, Intraperitoneal ; Liver ; drug effects ; enzymology ; pathology ; Microcystins ; administration & dosage ; isolation & purification ; pharmacology ; Phytoplankton ; physiology ; Reactive Oxygen Species ; metabolism ; Superoxide Dismutase ; metabolism ; Survival Analysis ; Time Factors

OBJECTIVETo evaluate the antagonism effects of green tea (GT) against microcystin LR (MC-LR) induced hepatotoxicity and nephrotoxicity in mice.

METHODSAll 40 male mice were randomly divided into four groups. Mice in group III and IV were pretreated with green tea for free drink at doses of 2 g/L and 12 g/L prior to MC-LR intoxication, for consecutively 18 days. The toxin treatment mice were administered continually intraperitoneal injections of MC-LR at a dose of 10 microg x kg(-1) x d(-1) bw from day 6th till sacrifice, continually 13 days. Mice were sacrificed and immediately subjected to necropsy, and the body weight, relative organ weight, serum biochemical parameters, antioxidant enzyme levels (SOD and GSH), lipid peroxidation products (MDA) and histopathology were systematically evaluated.

RESULTSMC-LR exposure led to increase the oxidative stress and organ injury was significantly observed through biochemical parameters and microscopic evaluation. However, high dose of GT pretreatment caused a significant elevation in serum GSH and SOD levels, and a decrease of serum MDA level as compared with MC-LR control. The mean values of GSH and SOD activities were separately 467.29 mg/L and 139.22 U/ml in group IV. Subsequently, GT pretreatment obviously diminished the serum ALT, AST and Cr activities. Those pathological damages in liver and kidney, were to a certain extent, lessened in GT pretreatment mice in correlation with the biochemical parameters.

CONCLUSIONGT might elevate antioxidant defense system, clean up free radicals, lessen oxidative damages and protect liver and kidney against MC-LR induced toxicity.

Animals ; Antioxidants ; pharmacology ; Chemical and Drug Induced Liver Injury ; Free Radicals ; metabolism ; Kidney Diseases ; chemically induced ; metabolism ; pathology ; Liver Diseases ; metabolism ; pathology ; Male ; Mice ; Mice, Inbred Strains ; Microcystins ; toxicity ; Oxidative Stress ; Tea

OBJECTIVETo deeply explore the effects of microcystins (MC-LR) on Bax and Bcl-2 during the course of MC-LR promoting liver tumor.

METHODSapplied to set up the animal model, and the effect of MC-LR promoting liver tumor was evaluated by the Albertgamma-GT methods. And then, the immunohistochemical technique, RT-PCR and image analysis were used to study the expression of the Bcl-2 and Bax during the course of promoting tumor.

RESULTS(1) MC-LR might enhance the positive reaction rate of GGT. The positive reaction rate of GGT in DEN + pure toxin group was 100%, it was significantly higher than the DEN control group 22.22% (P < 0.05). (2) The intension and areas of the protein expression of Bcl-2 in DEN + pure toxin group were 0.0977 and 0.0315, and in DEN control group were 0.0460 and 0.0205, respectively. The expression level of Bcl-2 protein in DEN + pure toxin group were significantly higher than in DEN control group (P < 0.05). Simultaneously, the protein expression of Bax was significantly decreased by MC-LR (P < 0.05). The intension and areas of the expression of Bax in DEN + pure toxin group were 0.0283 and 0.0073, and in DEN control group were 0.0655 and 0.0244 respectively. (3) The mRNA expression of Bcl-2 was significantly increased by MC-LR. The intension of Bcl-2 mRNA expression in DEN + pure toxin group was 2.244, being significantly higher than in the other groups (P < 0.05). However, the mRNA expression of Bax showed no significant difference between DEN + pure toxin and the other groups.

CONCLUSIONThe expression change of Bcl-2 and Bax should possibly play an important role in the course of MC-LR promoting liver tumor.

Animals ; Apoptosis ; Carcinogens ; toxicity ; Hepatocytes ; metabolism ; Liver Neoplasms ; chemically induced ; metabolism ; Male ; Microcystins ; toxicity ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; bcl-2-Associated X Protein ; biosynthesis ; bcl-Associated Death Protein ; biosynthesis

OBJECTIVETo study the molecular mechanism of microcystin (MC) induced liver tumorigenesis in rats.

METHODSThe two-stage-medium-term tumorigenesis theory was applied to establish the animal model, and the effect of MC in liver tumor formation was evaluated by the Albert gamma-GT methods, and then, the immunohistochemical technique and image analysis were used to study the expression of the bcl-2 and bax genes during tumorigenesis.

RESULTS(1) MC enhanced the formation of gamma-GT foci in liver (100%), which was significantly higher than the diethylnitrosamine (DEN) control group (22.22%) (P < 0.05). (2) MC decreased the expression of bax gene. The intensity and area of bax gene expression in the pure MC toxin group were 0.028 3 AODV and 0.007 3 ( micro m(2)/ micro m(2)) and in the DEN control group were 0.065 5 AODV and 0.024 4 ( micro m(2)/ micro m(2)), respectively. The intensity and areas of bax gene expression in the pure MC toxin group were significantly lower than those in the DEN control group (P < 0.05). (3) MC increased the expression of bcl-2 gene. The intensity and area of bcl-2 gene expression in the pure MC toxin group wee 0.097 7 AODV and 0.031 5 ( micro m(2)/ micro m(2)), respectively, and in the DEN control group were 0.046 0 AODV and 0.020 5 ( micro m(2)/ micro m(2)) respectively (P < 0.05).

CONCLUSION(1) MC can strongly promote liver tumorigenesis. (2) The changes of bcl-2 and bax gene expression possibly play an important role in the MC induced liver tumor formation.

Animals ; Carcinogens ; toxicity ; Immunohistochemistry ; Liver ; drug effects ; metabolism ; pathology ; Liver Neoplasms ; chemically induced ; metabolism ; pathology ; Mice ; Microcystins ; Peptides, Cyclic ; toxicity ; Proto-Oncogene Proteins ; biosynthesis ; Proto-Oncogene Proteins c-bcl-2 ; biosynthesis ; Rats ; Rats, Wistar ; bcl-2-Associated X Protein