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Objective To investigate the association between arsenic(+3 oxidation state) methyltransferase (AS3MT) genetic polymorphism and susceptibility to endemic arsenism.Methods Polymerase chain reactionrestriction fragment length polymorphism-single strand conformation polymorphism(PCR-RFLP-SSCP) technology was performed to detect mutations of AS3MT gene intron 8 and exon 9 in genome DNA of the 79 cases and 110 controls.PCR products with abnormal band forms were further sequenced to find the types and sites of mutation.Chi-square test and multivariate Logistic analyses were conducted.Results The incidence of the 9149 base mutation(A→C) in AS3MT gene intron 8(AS3MT-9149) in case group(19.0％,15/79) was lower than that in control group (23.6％,26/110).The incidence of the codon 287 mutation(ATG→AT/CG) in AS3MT gene exon 9(AS3MT-287)in case group(10.1％,8/79) was lower than that in control group (11.8％,13/110).However,statistical analysis indicated no significant difference in both mutations between two groups[AS3MT-9149:odds ratio(OR) =0.59,95％ confidence interval(CI):0.26-1.31,P =0.195; AS3MT-287:OR =0.85,95％ CI:0.32-230,P =0.751].Conclusions There are no significant association between the genetic polymorphisms of AS3MT-9149,AS3MT-287 and the susceptibility to endemic arsenism.Similarly,due to small sample amount,we can not exclude the possibility that these gene polymorphisms are related to susceptibility to endemic arsenism.
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Objective To explore the DNA damage in peripheral blood lymphocytes of rats exposed to sodium arsenite. Methods Thirty-two Wistar rats, weighing 180 - 200 g, equal male and female, were randomly divided into 4 groups, 8 in each group. Sodium arsenite 0(control) ,0.05,0.15,0.45 mg/L were given through drinking water for 30 days. Body weight and drinking water consumption were measured every day. Blood were collected and DNA damage in peripheral blood lymphocytes was examined by single cell gel electrophoresis.Results The increase of body mass[( 121.00 ± 38.57), ( 120.62 ± 42.80), ( 125.38 ± 48.68)g]and water intake [(36.9 ± 6.2), (37.9 ± 5.8), (39.3 ± 4.2)ml/d]in 0.05,0.15,0.45 mg/L sodium arsenite groups were compared with the control group[( 119.25 ± 47.27)g, (38.4 ± 5.1 )ml/d], and the difference were not significant (F = 0.040,0.828, all P ＞ 0.05). The tail ratios[46.25%(185/400) ,57.00%(228/400),64.00%(256/400)], tail lengths [(32.89 ± 17.18), (58.74 ± 36.28), (77.55 ± 35.73 ) μm]and tail moments [(6.29 ± 3.74), ( 11.20 ± 9.64),(17.30 ± 12.60)μm]in 0.05,0.15,0.45 mg/L sodium arsenite groups were significantly higher than those of the control group[39.25%(157/400), (18.73 ± 15.83),(2.61 ± 1.05)μm, all P ＜ 0.01], and the tail ratios,tail lengths and tail moments in lymphocytes increased with increased doses of arsenic concentration. Conclusions Low doses of arsenic exposure can induce DNA damage in peripheral blood lymphocytes of rats.
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Objective To observe the sub-chronic effects of low doses of arsenic poisoning in rabbits exposed to different periods on some of the serum enzymes and biochemical indicators, and to provide the basis for screening of meaningful hematologic indicators for early diagnosis of arsenic poisoning. Methods Twelve adult rabbits,weighing 2.0 - 3.5 kg, were randomly divided into four groups, 3 in each group, and they were fed with drinking water containing sodium arsenite 0(control),0.01,0.05,0.25 mg/L, respectively. Serum alanine aminotransferase (ALT), aspartate amino transferase (AST), alkaline phosphatase (ALP), γ-glutamyl transacylase (y-GT), total protein(TP), albumin(ALB), globulin(GLP), and ALB/GLP of rabbit were measured by SYSMEX-180 automated biochemistry analyzer after 8 weeks and 12 weeks exposure. Results The results showed that ALT in 0.05 mg/Lgroup of 12 week[(60.00 ± 4.14)U/L]increased significantly compared with the control[(41.50 ± 2.12)U/L, P ＜0.05];AST in 0.25 mg/L group of 8 week and 12 week[(46.50 ± 3.21 ), (52.33 ± 3.81 )U/L]increased significantly compared with the control[(21.33 ± 3.53), (29.50 ± 3.23 )U/L, all P ＜ 0.05];ALP in 0.05 mg/L and 0.25 mg/L group of 12 week [(78.68 ± 4.85 ), ( 103.00 ± 7.83 ) U / L]increased significantly compared with the control [(45.50 ± 5.50)U/L, all P ＜ 0.05];γ-GT in 0.05 mg/L group of 12 week[(19.33 ± 7.50)U/L]increased significantly compared with the contro1[(8.50 ± 3.53)U/L, P＜ 0.05]. TP, ALB, GLP, ALB/GLP of different groups of 8 week and 12 week were not significantly different statistically(F= 0.77,0.02,0.16,3.14 and 0.51,0.29,0.41,0.52, all P ＞ 0.05). Conclusions Zero point zero five mg/L and higher doses of sub-chronic arsenic exposure has some major damage to the liver. Compared with other serum enzymes and the biochemical indexes, serum AST is a early sensitive indicator of liver injury of the arsenic poisoning.
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Objective To observe the effect of low arsenic sub-chronic exposure on blood routine test index in rabbits to pave a way for screening early injury of the low arsenic exposure. Methods Twelve healthy male rabbits were randomly divided into four groups. They were administrated with As at the concentration of 0(control), 10, 50 and 250 μg/L in the drinking water. Blood samples were collected from the vein of the ear edge in 8 weeks, and blood test routine including leukocyte (WBC), lymphocyte (LYM), lymphocyte percentage (LYM%), neutrophil (GRA), neutrophil percentage (GRA%), monocyte (MON), monocyte percentage (MON%), red blood cell (RBC), hemoglobin (HGB), hematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular hemoglobin content (MCH), mean corpuscular hemoglobin concentration(MCHC), RDW, platelets(PLT), mean platelet volume(MPV), platelet hematocrit(PCT) and platelet distribution width (PDW), were detected by the ABX-60 hemocyte analyzer. Results Compared with the control group, the WBC, GRA and GRA% increased in 0, 50 and 250 μg/L groups, but there was no significance(P > 0.05). PLT and MPV had a statistical significance in 4 groups(F = 4.07,4.20, all P < 0.05). Compared with the control group[(292.00±16.97)×109/L, (7.10±0.99)fL], PLT decreased in the 250 μg/L group [(221.33±22.50)×109/L] and MPV decreased in the 50μg/L group [(5.57±0.46)fL] significantly (P < 0.05). The other index didn't change obviously. Conclusions Sub-chronic low level arsenic exposure may lead to the change in the blood system. The blood routine test may be considered for early injury of the arsenic poisoning.
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Objective To explore the effect of gene mutations of arsenic transport proteins-muhidrug resistance-associated proteins(MRP1 and MRP2)on phenotype of endemic arsenic poisoning.Methods Two hundreds and thirty-nine rural residents in 3 villages of Shuocheng Region,Shanxi Province were interviewed and examined by simple random sampling who had been lived there for 20 yearn at least.All the objects were divided into two groups on the basis of clinical examination with"The Standard Diagnosis of Endemic Arsenic Poisoning" (WT/S 211-2001):subjectives with skin lesion as a arsenic poisoning group and without skin lesion as a control group. One hundred and ninety-three blood samples were collected from each participanL Seventy-five arsenic poisoning cases and 118 controls were detected the gene mutations in the 2,17,23 exons of M RPI and the 10,18,31 exons of MRP2 by PCR-single strand conformation polymorphism (PCR-SSCP) and compared by multivariate Logistic regression model. Results Seventy-five cases and 164 controls underwent questionnaires. Age[ (58.85±11.26) vs (45.73±11.92),OR = 3.378,P ＜ 0.05],gender[male,57.3%(43/75)vs 27.4%(45/164),OR = 3.553,P＜ 0.01 ],smoking[46.7%(35/75) vs 21.3%(35/164),OR = 3.225,P ＜ 0.01 ],drinking[ 17.3%(13/75) vs 8.5% (14/164),OR = 1.836,P ＞ 0.05],vegetable and fruit intake[5.3%(4/75) vs 9.1%(15/164),OR = 0.560,P ＞ 0.05],egg and meat intake[34.7%(26/75) vs 30.5%(50/164),OR = 1.210,P ＞ 0.05],exposure of pesticide [41.3%(31/75) vs 29.3%(48/164),OR = 1.864,P ＜ 0.05] were tested by Logistic regression model. There was no gene mutation detected in the 23 exon of MRP1 and the 18 exon of MRP2. The gene mutations frequencies of the 2 exons of MRP1 in arsenic poisoning and control groups were 8.00% (6/75) and 5.93% (7/118),respectively;they were 13.33%(10/75) and 8.47%(10/118) of the 17 exons of MRP1,respectively;they were 22.67%(17/75) and 18.64%(22/118) of the 10 exons of MRP2,respectively;they were 5.33%(4/75) and 2.54%(3/118) of the 31 exons of MRP2,respectively. There was no significant difference between two groups(x2 = 0.312,1.165,0.460, 2.794,respectively,all P ＞ 0.05). After age,gender,smoking,drinking,nutritional level and exposure of pesticide being adjusted by multivariate Logistic regression model,there was no significant difference between two groups (OR = 0.803,1.892,2.388,1.098,respectively,all P ＞ 0.05). Conclusions The gene mutations of 2,17,23 exons of MRPI and the 10,18,31 exons of MRP2 may have no effect on the phenotype of endemic arsenic poisoning.
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Objective To study the change and the significance of T-lymphocyte immune function in peripheral blood in population living in arsenic-contaminated area. Methods Fifty-three cases of patients with arsenism symptoms were selected into experimental group, inhabitants who had no chronic arsenism symptoms into control group in the endemic area of Shuocheng District, Shuozhou City, Shanxi Province in 2006. Vein blood samples were taken and analyzed with SAP assay to measure the percentage of CD3+ ,CD4+ and CD8+ T-cells. Results It was found that the percentage of CD3+, CD4+, and CD4+/CD8+ [(41.89 ± 11.58)%, (25.60 ± 9.05)% and 1.02 ± 0.41] in the experimental group was lower than that in the control group [(68.38 ± 7.23)%, (39.17± 4.28)% ,1.69 ± 0.56, t = 13.61,18.72,14.79, all P < 0.05], while there was no statistical differences of CD8+ [(25.30 ± 6.85)%] compared to the control group[(23.54 ± 8.35)%,t = 3.07,P > 0.05]. The gender-related effect of arsenic on CD4+ and CD8+ was found by multiple linear step regression analysis(t = - 3.05, - 4.30, all P < 0.05). In case group, there were no statistical differences in CD3+, CD4+, CD8+ and CD4+/CD8+[(40.65±10.06)%, (24.48 ± 6.29)%, (24.52 ± 8.16)%,0.98 ± 0.25] between males and females [(43.07±12.96)%, (26.77±3.12)%, (26.50 ±9.32)%, 1.07 ±0.41, t = - 0.76,3.05,0.30,2.10, all P > 0.05]. Conclusions The immune function of T-lymphocytes of patients with chronic arsenism has been suppressed. It is of active significance to detect T-lymphocyte subpopulation in peripheral vein in patients with chronic arsenism aiming at estimating the function of cell immune and providing early diagnosis index.
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Objective To test whether sodium arsenite can induce in vitro reverse mutation of Salmonella typhimurium histamine-auxotroph mutant. Methods Ames test was carded out with Salmonella typhimurium strains TA97,TA98,TA100 and TA102 by standard method with or without the liver microsomal enzyme activation system (+S9,-S9). Results At concentrations of sodium arsenite from 500.00 to 5000.00 μg/plate, no colonies were seen on the plates of TA97,TA98,TA100 or TA102, with or without the presence of S9. At concentrations of sodium arsenite of 0.01,0.10,10.00 μg/plate and with the presence of S9, twice as many colonies grew on the plates of TA102 as the negative control(P<0.05). Without S9 activation,twice as many colonies grew on the plates of TA100 as the negative control(P<0.05)at concentrations of sodium arsenite of 1.00,10.00 μg/plate(P<0.05). The reverse mutation colonies induced by sodium arsenite in TA98 strain were twice as many as negative control group at concentrations of 0.01,0.10 μg/plate(P<0.05). There was no obvious increase of the strain clones in the other(P0.05). Conclusions With and without S9 activation, the doses of 500.00,5000.00 μg/plate sodium arsenite resulted in a toxic effect and a reduction of the revertants among the strain. At concentrations of 0.01～10.00 μg/plate, sodium arsenite exhibited mutngenesis effects.
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<p><b>OBJECTIVE</b>To investigate the role of multidrug resistant protein 2 (MRP2) and glutathione (GSH) cotransport system in hepatic arsenic metabolism in rats.</p><p><b>METHODS</b>Thirty healthy Wistar rats were divided randomizedly into five groups. The first group was the control group and the rats in this group were administered with normal saline. In the second, third and fourth group the rats were administered with 4, 10 and 20 mg As(+)3/kg BW of sodium arsenite respectively every other day for two weeks. The fifth group was the benzene-soluble organics (BSO) intervention group and in this group the rats were administered with 2 mmol/kg BW BSO intraperitoneally every day three days before the end of the experiment. The other treatment was the same as in other groups. All rats were sacrificed two weeks after the treatments. Arsenic contents in bile, liver and blood were detected by atomic absorption spectroscopy (AAS), and the expression of MRP2 in the membrane of hepatocyte was determined by Western-blot analysis.</p><p><b>RESULTS</b>The level of total arsenic (including organic arsenic and inorganic arsenic) in bile, liver and blood in all three different dose groups was higher than those in the control groups (P < 0.05). Arsenic levels of bile and liver were increased with intragastric arsenic dose. Blood arsenic levels were not significantly different in three different dose groups. Expression of hepatic MRP2 was increased with intragastric arsenic concentration. A positive correlation between biliary arsenic concentration and MRP2 levels was found in liver (r = 0.986, P < 0.05). For the rats pretreated with BSO, the biliary arsenic was significantly higher than that in the control group but lower than that in the high dose group; the liver and blood arsenic was higher than that in the control group and in the high dose group. Expression of MRP2 pretreated with BSO was decreased.</p><p><b>CONCLUSION</b>Sodium arsenite can induce expression of MRP2 and the up-regulation of MRP2 may play an important role in the bile secretion of arsenite and its metabolites. The function of MRP2 for transportation of arsenic and its metabolites is associated with the intracellular GSH level. BSO inhibits the synthesis of GSH, which weakens the function of the MRP2-GSH cotransport system and makes the liver arsenic increased.</p>
Subject(s)
Animals , Female , Male , Rats , Arsenic , Pharmacokinetics , Arsenic Poisoning , Metabolism , Bile , Metabolism , Glutathione , Liver , Metabolism , Membrane Transport Proteins , Multidrug Resistance-Associated Proteins , Random Allocation , Up-RegulationABSTRACT
<p><b>OBJECTIVE</b>To investigate the role of multidrug resistance-associated protein 2 (MRP2) in the hepatic cell membrane of rats.</p><p><b>METHODS</b>Thirty healthy Wistar rats were divided randomly into six groups based on time of administration (2 w, 4 w, 6 w) of 20 mg/kg of sodium arsenite, and their corresponding control groups. Animals were administered every other day. Arsenic content in blood and bile were detected by atomic absorption spectroscopy (AAS), and the expression of MRP2 in the membrane of hepatocyte by Western blotting was determined.</p><p><b>RESULTS</b>Total arsenic levels (including organic arsenic and inorganic arsenic) in blood and bile were significantly higher than control groups (P < 0.05) at all three different time points, especially in 2 w and 4 w group (16.8 and 13.8 fold greater than that in control). The expression of MRP2 increased 36.61%, 32.36%, 12.73% more respectively in 2 w, 4 w, 6 w groups than those in control groups (P < 0.05). The expression of MRP2 was correlated with total arsenic content in bile (r = 0.713, P < 0.05).</p><p><b>CONCLUSIONS</b>Bile is one of the major routes for the excretion of arsenite and its metabolites, and the overexpression of MRP2 may play an important role in the bile excretion of them at early stage.</p>