Effects of ginkgo biloba tablets on liver injury in patients with coal-burning-borne arsenism based on DNA damage and repair inhibition / 中华地方病学杂志

Objective:To investigate the role of DNA damage and repair inhibition in the effect of ginkgo biloba on liver injury in patients with coal-burning-borne arsenism.Methods:In March 2017, the investigation was conducted in Jiaole village arsenic poisoning area in Yuzhang Town, Xingren County, Guizhou Province. According to the "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015) and the "Diagnostic Criteria of Occupational Toxic Hepatopathy" (GBZ 59-2010), 52 patients with arsenism were selected as the ginkgo biloba intervention group, and 49 cases of arsenism patients as intervention control group. Ginkgo biloba tablets were given orally for 3 months (1 tablet/time, 3 times/d) according to the commonly used clinical methods, and no other drugs were given to all subjects during the intervention period. The intervention control group was given placebo in the same way as that of ginkgo biloba intervention group. A total of 41 residents who did not burn high arsenic coal 12 km away with no abnormal liver function were selected as normal control group. Physical examinations were performed before the intervention and at the end of the intervention at 3 months. After receiving signed informed consent, morning urine and peripheral venous blood samples were collected to detect urinary arsenic content by inductively coupled plasma mass spectrometry (ICP-MS); liver function biochemical indexes [albumin (ALB), albumin/globulin (A/G), cholinesterase (CHE), total bile acid (TBA)] were determined by automatic biochemical analyzer, DNA damage by single-cell gel electrophoresis assay, and the expression of miR-145 (repair inhibition index) by qRT-PCR.Results:There were 116 subjects, 41 in normal control group, 39 in ginkgo biloba intervention group and 36 in intervention control group. In ginkgo biloba and intervention and intervention control groups, there was no significant difference in age, gender, smoking habits and drinking compared with normal control group ( P > 0.05). Urinary arsenic content, TBA level, DNA damage degree [comet tail DNA percentage (TailDNA%) and olive tail moment (OTM)] and plasma miR-145 expression level [(38.75 ± 19.09) μg/g Cr, (11.13 ± 1.55) μmol/L, 8.50 ± 0.88, 7.43 ± 0.68, 5.78 ± 0.75, respectively] in ginkgo biloba intervention group patients before intervention were higher than those in normal control group [(11.62 ± 5.33) μg/g Cr, (5.36 ± 0.87) μmol/L, 5.24 ± 0.33, 4.71 ± 0.29, 2.05 ± 0.27, respectively], the differences were statistically significant ( P < 0.05); the levels of ALB, A/G and CHE were significantly lower than those in normal control group ( P < 0.05). After the intervention of ginkgo biloba, urinary arsenic content, TBA level, DNA damage degree (TailDNA% and OTM) and plasma miR-145 expression level in patients were significantly lower than those before the intervention ( P < 0.05); the levels of ALB, A/G and CHE were significantly higher than those before the intervention ( P < 0.05). There was no significant difference in the above indexes before and after intervention in the intervention control group ( P > 0.05). The results of correlation analysis between DNA damage degree, miR-145 and liver function indexes after the intervention of ginkgo biloba showed that, DNA damage degree (TailDNA% and OTM) was negatively correlated with the levels of ALB, A/G and CHE ( r = - 0.34, - 0.33, - 0.48, - 0.31, - 0.31, - 0.42, P < 0.05), and positively correlated with the level of TBA ( r = 0.49, 0.48, P < 0.05); miR-145 was negatively correlated with the levels of ALB, A/G and CHE ( r = - 0.26, - 0.23, - 0.38, P < 0.05), which was positively correlated with the level of TBA ( r = 0.32, P < 0.05); and DNA damage degree was positively correlated with the expression of miR-145 ( r = 0.65, 0.52, P < 0.05). Conclusion:Ginkgo biloba tablets can alleviate the liver damage caused by arsenic through coal burning, and the mechanism of this process is related to its inhibition of miR-145 expression and reduction of DNA damage.

Role of LXRα/SREBP-1c in arsenic induced lipid metabolism disorders in rats / 中华地方病学杂志

Objective:To investigate the role of liver X-activated receptor (LXRα)/sterol-regulatory element binding protein (SREBP-1c) in arsenic induced lipid metabolism disorders in rats, and to provide a basis for study the mechanism of arsenic induced lipid metabolism disorders.Methods:Twenty-four healthy clean grade Wistar rats, were randomly divided into 4 groups according to body weight (80 - 100 g) by the random number table method, with 6 rats in each group, half male and half female. Rats in control group were given deionized water by gavage. The low, medium and high arsenic dose groups were given 2.5, 5.0 and 10.0 mg·kg -1·d -1 sodium arsenite solution by gavage, respectively. They were exposed to arsenic for 6 days a week for 4 months. At the end of the experiment, blood and liver samples of rats in each group were collected. The hepatic arsenic content was determined by inductively coupled plasma mass spectrometry (ICP-MS); the serum levels of triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were measured by automatic biochemical analyzer. The mRNA expression levels of LXRα and SREBP-1c in liver tissues were determined by real-time PCR; the protein expression levels of LXRα, SREBP-1c, acetyl CoA carboxylase (ACC) and phospho-ACC (pACC) in liver tissues were determined by Western blotting. Results:The hepatic arsenic contents of rats in low, medium and high arsenic dose groups were (61.04 ± 4.98), (62.66 ± 6.71) and (87.86 ± 13.89) μg/g, respectively, which were higher than that in control group [(2.43 ± 0.63) μg/g, P < 0.05], and the hepatic arsenic content of rats in high arsenic dose group was higher than those in low and medium arsenic dose groups ( P < 0.05). The serum TG levels of rats in low, medium and high arsenic dose groups were (0.90 ± 0.17), (1.28 ± 0.24) and (1.82 ± 0.18) mmol/L, respectively, which were higher than that in control group [(0.50 ± 0.12) mmol/L, P < 0.05]; the serum LDL-C levels of rats in low, medium and high arsenic dose groups were (0.54 ± 0.04), (0.63 ± 0.07) and (0.69 ± 0.08) mmol/L, respectively, which were higher than that in control group [(0.27 ± 0.05) mmol/L, P < 0.05]; the serum TC levels of rats in medium and high arsenic dose groups were (1.88 ± 0.23) and (2.10 ± 0.10) mmol/L, respectively, which were higher than that in control group [(1.51 ± 0.14) mmol/L, P < 0.05]; the serum HDL-C levels of rats in medium and high arsenic dose groups were (0.84 ± 0.11) and (0.71 ± 0.14) mmol/L, respectively, which were lower than that in control group [(1.15 ± 0.08) mmol/L, P < 0.05]; and the serum levels of TG and LDL-C in medium and high arsenic dose groups were higher than those in low arsenic dose group ( P < 0.05), and the serum level of TG in high arsenic dose group was higher than that in medium arsenic dose group ( P < 0.05). The mRNA expression level of hepatic LXRα of rats in high arsenic dose group was higher than those in control group and low arsenic dose group ( P < 0.05); there was no significant difference in mRNA expression levels of hepatic SREBP-1c of rats between low, medium and high arsenic dose groups and control group ( P > 0.05). The protein expression levels of hepatic LXRα of rats in medium and high arsenic dose groups were higher than that in control group ( P < 0.05), and high arsenic dose group was higher than low arsenic dose group ( P < 0.05); the protein expression levels of hepatic SREBP-1c and ACC of rats in high arsenic dose group were higher than that in control group ( P < 0.05). There was a positive correlation between hepatic arsenic content in arsenic-exposed rats and the serum levels of TG, TC, LDL-C, the mRNA expression level of hepatic LXRα, the protein expression levels of hepatic LXRα, SREBP-1c and ACC ( r = 0.84, 0.62, 0.89, 0.55, 0.54, 0.64, 0.70, P < 0.05), and the serum level of HDL-C was negatively correlated with the hepatic arsenic content in arsenic-exposed rats ( r = - 0.75, P < 0.001). Conclusion:Sodium arsenite can increase the serum levels of TG, TC and LDL-C, decrease the serum level of HDL-C and increase the protein expression levels of LXRα and SREBP-1c in liver tissues, suggesting that arsenic induced lipid metabolism disorders in rats may be related to the upstream regulation mechanism of LXRα/SREBP-1c.

Correlation between the disease condition and dyslipidemia in patients with coal-burning-borne endemic arsenic poisoning / 中华地方病学杂志

Objective:To understand the relationship between the disease condition of patients with coal-burning-borne endemic arsenic poisoning (abbreviated as coal-burning-borne arsenic poisoning) and serum lipid metabolism indicators.Methods:Using a case-control study method, in the coal-burning-borne arsenic poisoning village of Yuzhang Town, Qianxinan Prefecture, Guizhou Province, 204 patients with arsenic poisoning diagnosed according to the standard of "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015) were included in case group, including 87 males and 117 females, aged(53.37 ± 8.06) years old; and they were divided into mild arsenic poisoning group (71 cases), moderate arsenic poisoning group (59 cases) and severe arsenic poisoning group (74 cases) according to the clinical grading. Another 63 residents were selected into control group in a non-arsenic-exposed village about 12 km away from the diseased village, including 23 males and 40 females, aged (53.78 ± 9.10) years old. A face-to-face questionnaire survey was conducted for each group of people, including basic information such as general demographic characteristics, smoking status, and drinking status; fasting peripheral blood was collected, and an automatic biochemical analyzer was used to detect serum total cholesterol (TC) and triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) levels.Results:There were significant differences of serum TC [(4.94 ± 1.00), (5.00 ± 0.99), (5.27 ± 0.94), (5.57 ± 1.07) mmol/L], TG [(2.17 ± 0.90), (2.25 ± 1.31), (2.66 ± 1.43), (2.78 ± 1.40) mmol/L], LDL-C [(2.51 ± 0.79), (2.74 ± 0.64), (2.97 ± 0.66), (3.15 ± 0.80) mmol/L], and HDL-C levels [(1.57 ± 0.55), (1.42 ± 0.43), (1.36 ± 0.42), (1.30 ± 0.38) mmol/L] in control group, mild, moderate and severe arsenic poisoning groups ( F = 5.83, 3.64, 9.72, 4.41, P < 0.01 or < 0.05). Among them, the serum TC level in severe arsenic poisoning group, serum TG and LDL-C levels in moderate and severe arsenic poisoning groups were significantly higher than those in control group ( P < 0.05); the serum HDL-C level in moderate and severe arsenic poisoning groups were lower than that in control group ( P < 0.05); the serum TC, TG and LDL-C levels in severe arsenic poisoning group were significantly higher than those in mild arsenic poisoning group ( P < 0.05). After linear trend test, serum TC, TG and LDL-C levels all showed an upward trend with the degree of arsenic poisoning ( Ftrend = 15.77, 10.14, 29.15, P < 0.05), and serum HDL-C level showed a downward trend with the degree of arsenic poisoning ( Ftrend = 12.75, P < 0.05). There were significant differences in the abnormal rates of serum TC, TG and LDL-C levels among control group and mild, moderate and severe arsenic poisoning groups (χ 2 = 21.16, 16.60, 8.29, P < 0.01 or < 0.05). Among them, the serum TC and TG levels abnormal rates in moderate and severe arsenic poisoning groups and serum LDL-C level abnormal rate in severe arsenic poisoning group were higher than those in control group ( P < 0.05), the serum TC, TG and LDL-C levels abnormal rates in severe arsenic poisoning group were higher than those in mild arsenic poisoning group ( P < 0.05). There was no significant difference of the serum HDL-C level abnormal rate among four groups (χ 2 = 2.11 , P > 0.05). The results of trend chisquare analysis showed that the abnormal rates of serum TC, TG and LDL-C levels presented an increasing trend with the degree of arsenic poisoning (χ 2trend = 19.90, 15.25, 7.63, P<0.05). The results of logistic regression analysis showed that the risk of abnormal serum TC level in patients with severe arsenic poisoning was 2.90 times that in control group [odds ratio ( OR) = 2.90, 95% confidence interval ( CI): 1.43 - 5.91], and the risk of abnormal serum LDL-C level in patients with severe arsenic poisoning was 2.87 times that in control group ( OR = 2.87, 95% CI: 1.22 - 6.71). Conclusion:There is a correlation between the disease condition of patients with coal-burning-borne arsenic poisoning and their dyslipidemia.

Prevalence of diabetes among residents in coal-burning arsenic poisoning area of Yuzhang Town Guizhou Province / 中华地方病学杂志

Objective:Comparative analysis of diabetes was carried out in coal-burning arsenic poisoning areas and non-arsenic exposed villages of Yuzhang Town, so as to explore the relationship between arsenic exposure and diabetes.Methods:Data of basic information of 594 people who were diagnosed and included in the diabetes management in Central Health Center of Yuzhang Town in Qianxinan Prefecture Guizhou Province in 2018 were collected. According to the "Standards for the Determination and Classification of Endemic Arsenic Poisoning Areas" (WS 277-2007), 11 administrative villages in the town were divided into 5 arsenic poisoning villages and 6 non-arsenic exposure villages. The prevalence (%) was used for statistical description.Results:In 2018, the prevalence of diabetes in Yuzhang Town was 1.74% (594/34 218), 1.38% (243/17 665) for men and 2.12% (351/16 553) for women, the gender difference was statistically significant (χ 2=27.794, P < 0.05). The prevalence of standardized diabetes in arsenic poisoning villages was 3.38%; the prevalence of standardized diabetes in non-arsenic exposure villages was 3.13%. After sex stratification analysis, the non-arsenic exposed villages were used as reference. The OR and 95% CI of diabetic patients in arsenic poisoning villages were 0.65 (0.50-0.81) for males and 1.35 (1.09-1.67) for females. Conclusions:The association between arsenic exposure and diabetes is related to gender. The risk of diabetes mellitus in women is higher than that in men.

Association of endemic arsenic poisoning with liver injury: Meta-analysis / 中华地方病学杂志

Objective@#To systematically review the correlation between endemic arsenism and liver injury.@*Methods@#Pertinent studies were identified by searching Pubmed, Embase, China National Knowledge Infrastructure (CNKI) and Wanfang Data databases through November 2018. Studies that reported endemic arsenism and liver injury at home and abroad were collected. All of the Meta-analysis were performed by using Review Manager 5.3 software. The odds ratios (OR) and standardized mean differences (SMD) were used to compare continuous and dichotomous variables. The I2 was used to assess heterogeneity among studies, and the fixed-effects model or the random-effects model was chosen for the quantitative analysis. Subgroup analysis were performed to discuss sources of heterogeneity. The publication bias was evaluated by inverted funnel plot.@*Results@#Totally 16 documents were included, including 7 in English and 9 in Chinese. With liver injury (including the results of hepatomegaly epidemic, abnormal Bultrasound or pathological examination of liver, cirrhosis with definite diagnosis, etc.), different serum liver function and liver fibrosis indicators as the outcome, 7 274 cases, 410-820 cases and 255 cases were included in the arsenic exposure group, and 5 078 cases, 134-327 cases and 164 cases in the control group, respectively. The Meta-analysis showed that arsenic exposure was associated with increased risk of liver injury [OR value and 95% confidence interval (CI) was 2.40 (1.43-4.05), Z=3.30, P < 0.01]. Arsenic exposure had promoted the levels of serum aspartate aminotransferase (AST), alkaline phosphatase (ALP), glutamyltransferase (γ-GT) and total bile acid [TBA, SMD and 95%CI were 1.71 (0.18-3.23), 1.73 (0.42-3.03), 0.25 (0.09-0.41) and 0.55 (0.35-0.75), respectively, P < 0.05 or < 0.01], while had suppressed the level of serum albumin [ALB, SMD and 95%CI were-2.37 (-4.01 -- 0.73), P < 0.01]. Among the serum indicators for liver fibrosis, the levels of serum procollagen type Ⅲ (PC-Ⅲ) and collagen type Ⅳ(Ⅳ-C) were significantly increased in arsenic exposed patients [SMD and 95%CI were 0.98 (0.05-1.91), 1.60 (1.38-1.83), respectively, P < 0.05 or < 0.01]. Subgroup analysis showed that arsenic exposure had been stopped or not at the time of study, study design (including cross-sectional study and cohort study) had a significant effect on the association between arsenic exposure and liver injury (χ2=17.17, 11.85, P < 0.01).@*Conclusions@#Both drinking water-borne arsenicosis and coal burning-borne arsenicosis can cause high incidence of liver injury. ALP, ALB, γ-GT and TBA can be used as serum biomarkers for arsenic exposure induced liver injury. After blocking of arsenic exposure by adopting comprehensive prevention and control measures such as changing water sources and installation of ventilated stoves, the liver injury of patients could be effectively improved.

Changes and significance of the ratio of T helper 17 and regulatory T cell in peripheral blood of patients with coal-burning-borne arsenic poisoning / 中华地方病学杂志

Objective To observe the change of T helper 17 (Th17),regulatory T cell (Treg) as a percentage of lymphocytes and the Th17/Treg ratio in peripheral blood of patients with coal-burning-borne arsenic poisoning,and to explore the role of Th17 cells and Treg cells balance in arsenic-induced immune injury.Methods A case-control study was conducted to investigate 149 cases of arsenic poisoning in Yuzhang arsenic poisoning area in the southwestern Gnizhou Province,and the age was (50.69 ± 6.14) years old,including 94 males and 55 females.The diagnosis was based on the "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015),and the cases were divided into mild arsenic poisoning group (39 cases),moderate arsenic poisoning group (54 cases) and severe arsenic poisoning group (56 cases);forty--one residents of non-arsenic exposed villages about 12 km away from the diseased area were collected as control group,the age was (45.76 ± 7.88) years old,including 12 males and 29 females.Hair samples and peripheral blood were collected from the subjects.The content of hair arsenic was detected by inductively coupled plasma mass spectrometry (ICP-MS).The percentages of Th17 cells and Treg cells in peripheral blood lymphocytes were detected by flow cytometry,and changes in the ratio of Th17/Treg in each group were analyzed.Results The hair arsenic contents in control,mild,moderate,and severe arsenic poisoning groups [median (quartile)] were 0.12 (0.08-0.18),0.20 (0.16-0.33),0.25 (0.18-0.41),0.28 (0.21-0.50) μg/g,and the differences were statistically significant between groups (H =52.22,P ＜ 0.01),and the hair arsenic content in each arsenic poisoning group was higher than that of the control group (P ＜ 0.05).The percentages of Th17 cells in peripheral blood lymphocytes of moderate and severe arsenic poisoning groups [(0.42 ± 0.21)％,(0.41 ± 0.23)％] were higher than that of the control group [(0.29 ± 0.16)％,P ＜ 0.05].The percentages of Treg cells in peripheral blood lymphocytes of mild,moderate and severe arsenic poisoning groups [(0.37 ± 0.18)％,(0.31 ± 0.19)％,(0.27 ± 0.18)％] were lower than that of the control group [(0.71 ± 0.20)％,P ＜ 0.05];with respect to the mild arsenic poisoning group,the percentage of Treg cells in severe arsenic poisoning group was reduced (P ＜ 0.05).The ratios of Th17/Treg in mild,moderate and severe arsenic poisoning groups (1.17 ± 0.63,1.56 ± 0.69,1.83 ± 0.85) were higher than that of the control group (0.43 ± 0.22,P ＜ 0.05);compared with mild arsenic poisoning group,the ratio of Th17/Treg in severe arsenic poisoning group was elevated (P ＜ 0.05).Correlation analysis showed that the hair arsenic content was positively correlated with the percentage of Th17 cells in peripheral blood lymphocytes and the ratio of Th17/Treg (r =0.323,0.608,P ＜ 0.05),and negatively correlated with the percentage of Treg cells in peripheral blood lymphocytes (r =-0.486,P ＜ 0.05).Conclusion Coal-burning-borne arsenic poisoning can cause the proportion of Th17 cells in the peripheral blood of patients to increase in lymphocytes,and the proportion of Treg cells in lymphocytes to decrease,which in turn changes the balance of Th17/Treg,resulting in weakened immune tolerance and disorder the regulation of inflammatory response,thus participates in the occurrence and development of arsenic-induced immune damage.

Differential expression and significance of Foxp3, TGF-β1 and IL-2 in peripheral blood of patients with coal-burning arsenic poisoning / 中华地方病学杂志

Objective To investigate the expression of transcription factor forkhead/winged helix transcription factor 3 (Foxp3),immune factor transforming growth factor-beta 1 (TGF-β1),and T-lymphocyte activation related factor interleukin-2 (IL-2) in peripheral blood of patients with coal-burning arsenic poisoning,and to analyze the effects of arsenic exposure on immune function.Methods A case-control study was conducted to investigate 149 cases [94 males and 55 females,(50.69 ± 6.14) years old] of arsenic poisoning in Yuzhang coalburning arsenic poisoning area,southwestern Guizhou Province,and the cases were diagnosed based on the "Diagnosis of Endemic Arsenicosis" (WS/T 211-2015) and confirmed by clinical review.According to skin damage,the patients were divided into mild arsenic poisoning group (39 cases),moderate arsenic poisoning group (54 cases) and severe arsenic poisoning group (56 cases);and 41 cases [12 males and 29 females,(45.76 ± 7.88) years old] of non-arsenic exposed residents from 12 km of Yuzhang coal-burning area were selected as control group.Morning urine and peripheral blood samples were collected with informed consent.Urine arsenic content was measured by inductively coupled plasma mass spectrometry (ICP-MS).Urine arsenic was corrected by creatinine (Cr).Detection of regulatory T cell (Treg)-specific transcription factor Foxp3 gene expression in human peripheral blood was done by real-time fluorescence quantitative PCR,and the levels of Treg-related immune factor TGF-β1 and IL-2 in serum were detected by enzyme linked immunosorbent assay (ELISA).Results The urinary arsenic contents [median (quartile):29.13 (19.75-54.50),31.81 (17.52-53.31),30.51 (18.35-45.76) μg/g Cr] in each arsenic poisoning group were higher than that in the control group [21.62 (17.65-28.44) μg/g Cr,P ＜ 0.05].The expression levels of Foxp3 mRNA in peripheral blood of each arsenic poisoning group [median (quartile):0.58 (0.17-1.27),0.32 (0.17-0.61),0.33 (0.13-0.62)] were significantly lower than that in the control group [0.87 (0.64-1.50),P ＜ 0.05];compared with mild arsenic poisoning group,the expression of Foxp3 mRNA in peripheral blood of moderate and severe arsenic poisoning groups decreased (P ＜ 0.05).The contents of serum TGF-β1 [(13.14 ± 5.19),(12.85 ± 5.51),(12.78 ± 4.95) μg/L] in each arsenic poisoning group were significantly higher than that in the control group [(3.90 ± 1.53) μg/L,P ＜ 0.05].The levels of IL-2 in serum of each arsenic poisoning group [(9.85 ± 5.38),(11.64 ± 6.40),(12.27 ± 6.19) ng/L] were lower than that in the control group [(34.30 ± 4.84) ng/L,P ＜ 0.05];the serum level of IL-2 in severe arsenic poisoning group was higher than that in mild arsenic poisoning group (P ＜ 0.05).Conclusions Arsenic exposure can cause abnormal changes of Treg-specific transcription factor Foxp3 and related immune factors TGF-β1 and IL-2 in peripheral blood of patients.It is suggested that Treg dysfunction may be related to arsenic poisoning.

Association of endemic arsenic poisoning with liver injury: Meta-analysis / 中华地方病学杂志

Objective To systematically review the correlation between endemic arsenism and liver injury. Methods Pertinent studies were identified by searching Pubmed, Embase, China National Knowledge Infrastructure (CNKI) and Wanfang Data databases through November 2018. Studies that reported endemic arsenism and liver injury at home and abroad were collected. All of the Meta-analysis were performed by using Review Manager 5.3 software. The odds ratios (OR) and standardized mean differences (SMD) were used to compare continuous and dichotomous variables. The I2 was used to assess heterogeneity among studies, and the fixed-effects model or the random-effects model was chosen for the quantitative analysis. Subgroup analysis were performed to discuss sources of heterogeneity. The publication bias was evaluated by inverted funnel plot. Results Totally 16 documents were included, including 7 in English and 9 in Chinese. With liver injury (including the results of hepatomegaly epidemic, abnormal B -ultrasound or pathological examination of liver, cirrhosis with definite diagnosis, etc.), different serum liver function and liver fibrosis indicators as the outcome, 7274 cases, 410 - 820 cases and 255 cases were included in the arsenic exposure group, and 5078 cases, 134 - 327 cases and 164 cases in the control group, respectively. The Meta-analysis showed that arsenic exposure was associated with increased risk of liver injury [OR value and 95％ confidence interval (CI) was 2.40 (1.43 - 4.05), Z = 3.30, P < 0.01]. Arsenic exposure had promoted the levels of serum aspartate aminotransferase (AST), alkaline phosphatase (ALP), glutamyltransferase (γ-GT) and total bile acid [TBA, SMD and 95％CI were 1.71 (0.18 - 3.23), 1.73 (0.42 - 3.03), 0.25 (0.09 - 0.41) and 0.55 (0.35 - 0.75), respectively, P < 0.05 or < 0.01], while had suppressed the level of serum albumin [ALB, SMD and 95％CI were - 2.37 ( - 4.01 - - 0.73), P < 0.01]. Among the serum indicators for liver fibrosis, the levels of serum procollagen type Ⅲ (PC-Ⅲ) and collagen type Ⅳ (Ⅳ-C) were significantly increased in arsenic exposed patients [SMD and 95％CI were 0.98 (0.05 - 1.91), 1.60 (1.38 - 1.83), respectively, P < 0.05 or < 0.01]. Subgroup analysis showed that arsenic exposure had been stopped or not at the time of study, study design (including cross-sectional study and cohort study) had a significant effect on the association between arsenic exposure and liver injury (χ2 = 17.17, 11.85, P < 0.01). Conclusions Both drinking water-borne arsenicosis and coal burning-borne arsenicosis can cause high incidence of liver injury. ALP, ALB, γ-GT and TBA can be used as serum biomarkers for arsenic exposure induced liver injury. After blocking of arsenic exposure by adopting comprehensive prevention and control measures such as changing water sources and installation of ventilated stoves, the liver injury of patients could be effectively improved.

Roles of T helper 17 and regulatory T cell infiltration in hepatic injury induced by arsenic in rats / 中华地方病学杂志

Objective To investigate the infiltration of T helper 17 (Th17) and regulatory T cells (Treg) in the liver of rats exposed to arsenic,and to investigate the roles of Th17 and Treg in infiltration of liver injury induced by arsenic.Methods Thirty-two Wistar rats,half male and half female,were randomly divided into control,low,medium and high arsenic dose groups by body weight via the random number table method,8 rats per group.Rats in control group were given oral garage of deionized water,while other groups were given oral gavage doses of 2.00 g/L sodium arsenite (NaAsO2) according to their body weight for 6 days every week,the concentrations of NaAsO2 were 1.25,2.50 and 5.00 ml/kg,respectively.After 4 months,liver tissue samples of rats were collected,the content of arsenic in liver was determined by inductively coupled plasma mass spectrometry (ICP-MS);Hematoxylin-eosin staining (HE) method was used to observe the morphological changes of liver tissue in rats;the protein expressions of interleukin-17A (IL-17A,the imflammatory factor secreted by Th17 cells) and Forkhead Box P3 (Foxp3,the lineage-specific transcription factor of Treg cells) were measured with immunohistochemistry.Results ① Arsenic content in liver of low,medium,and high arsenic exposed groups [63.83 (52.79-80.26),59.16 (51.38-76.58),79.26 (69.59-107.44) μg/g] were higher than those of the control group [2.86 (1.76-3.56)μg/g,P ＜ 0.05],and the high arsenic dose group was higher than the medium arsenic dose group (P ＜ 0.05).② With increasing doses of arsenic,the numbers of inflammatory cells in the liver tissue of rats were increased,and the liver tissue of the high arsenic dose group showed vacuolar degeneration and pathological changes in some areas.③ Compared with the control group,low and medium arsenic dose groups (0.001 + 0.001,0.010 ± 0.020,0.030 ± 0.080),the expression of IL-17A protein in the liver in high arsenic dose group were significantly increased (0.220 ± 0.130,P ＜ 0.05),the differences were statistically significant between groups (F =14.776,P ＜0.05).The expressions of Foxp3 protein in the liver in low,medium,and high arsenic dose groups were significantly higher (0.270 ± 0.050,0.330 ± 0.040,0.320 ± 0.070) than that in the control group (0.070 ± 0.020),the differences were statistically significant between groups (F =56.990,P ＜ 0.05).④ There was a positive correlation between hepatic arsenic levels and protein levels of IL-17A and Foxp3 in liver (r =0.48,0.81,P ＜ 0.05).Conclusion Arsenic exposure can increase the content of arsenic in liver tissue of rats,which causes the changes of infiltration of Th17 and Treg cells,leading to the change of immune status,suggesting that Thl7 and Treg cells play an important role in the development of arsenic-induced immune injury.

Differential expression and significance of CD4+CD25+Foxp3+regulatory T cell in liver of arsenic-exposed rats / 中华地方病学杂志

Objective To observe the differential expression level of CD4+CD25+Foxp3+regulatory T cells (Treg) in liver of arsenic-exposed rats, explore the regulatory mechanisms on immunological of hepatic injury induced by arsenic, and provide a basis for prevention and treatment of the disease. Methods Thirty-two healthy Wistar rats were selected and randomly divided into control,low,medium and high arsenic dose groups by weight,8 rats per group. Rats in control group were given oral gavage of deionized water, while the other groups were given oral gavage doses of 2.00 g/L sodium arsenite(NaAsO2) according to their body weight for 6 days every week, the concentrations were 1.25, 2.50 and 5.00 ml/kg. After 4 months, liver tissue samples of rats were collected, the content of arsenic in liver was detected by inductively coupled plasma mass spectrometry (ICP-MS);the expression of Treg cells in liver was detected by immunohistochemistry; enzyme-linked immunosorbent assay (ELISA) was applied to detect the levels of interloukin-10 (IL-10),transforming growth factor beta 1 (TGF-β1), IL-6, IL-17 and IL-2. Results Compared with the control group [28.57 (17.64 - 35.64)μg/g], the content of arsenic in liver in low,medium and high arsenic exposed groups[M(P25-P75):638.30(527.91-802.58),591.64(513.82-723.16),792.55 (695.93 - 1 074.41) μg/g] increased, the differences were statistically significant(P < 0.05). Compared with low arsenic group, the content of arsenic in liver in high arsenic group increased, the difference was statistically significant (P < 0.05). Numerical density on area (NA) of positive Treg cells in medium,high arsenic exposed groups [(2.25 ± 0.50),(4.00 ± 2.16)A/cm2]was higher than that of the control group[(0.60 ± 0.54)A/cm2,P<0.05];NA of positive Treg cells in high arsenic exposed group was higher than that of the low arsenic exposed group[(1.50 ± 0.58) A/cm2, P < 0.05]. The expressions of the IL-10 in low, medium and high arsenic exposed groups [(5.58 ± 1.70), (6.78 ± 1.09),(7.18 ± 0.53)μg/L]were higher than that of the control group[(2.32 ± 0.83) μg/L,P<0.05];compared with low arsenic group, the expression of IL-10 in high arsenic group increased (P < 0.05); compared with control group [(1.46 ± 0.65) μg/L], the expression of TGF-β1 in high arsenic exposed group increased[(9.06 ± 3.60)μg/L, P<0.05];compared with control group [(2.33 ± 0.66)μg/L], the expression of IL-6 in high arsenic exposed group increased [(5.03 ± 1.39) μg/L, P < 0.05], compared with low arsenic exposed group [(2.46 ± 1.71) μg/L], the expressions of IL-6 in high arsenic exposed group increased, the difference was statistically significant (P < 0.05);the expression of IL-17 among control, low, medium and high arsenic exposed groups[(4.87 ± 1.64),(7.50 ± 2.74), (6.21 ± 1.47),(7.23 ± 2.68)μg/L]were not statistically significant (F = 1.429, P > 0.05); compared with control group [(16.30 ± 3.98) μg/L], the expression of IL-2 in high arsenic exposed group decreased[(9.93 ± 2.65) μg/L, P <0.05]. The content of arsenic in liver was positively correlated with the expression of IL-10, TGF-β1, IL-17, IL-6 (rs=0.696,0.463,0.632,0.502,P<0.05),and negatively correlated with the expression of IL-2(rs=-0.522,P<0.05). Conclusion With increasing of arsenic exposure level, the content of arsenic in liver and the expression of CD4+CD25+Foxp3+Treg have increased,the cytokines are secreted abnormally,liver immunological micro environment is disordered,immune tolerance is formed,and immune clearance is inhibited,which may play an important role in the occur and development of immunological liver damage induced by arsenic in rat.

Effects of sodium arsenite exposure on activation and extracellular matrix secretion of human hepatic stellate cells / 中华预防医学杂志

Objective@#To explore the effects of sodium arsenite (NaAsO2) exposure on the activation and extracellular matrix secretion of human hepatic stellate cells, and to provide a theoretical basis for the mechanism study of arsenic induced hepatic fibrosis.@*Methods@#Different doses of NaAsO2 (0.0, 0.1, 1.0, 10.0, 50.0, 100.0 μmol/L) were exposed to human hepatic stellate cell line (Lx-2) for 24, 48 and 72 huors. CCK-8 assay was used to measure cell viability and IC50 of NaAsO2 on Lx-2 was then calculated; According to IC50 results, 0.000, 1.875, 3.750, 7.500, and 15.000 μmol/L of NaAsO2 were exposed to Lx-2 cells for 24 hours, besides, 7.500 μmol/L of NaAsO2 was exposed to Lx-2 cells for 0, 12, 24, 48, and 72 hours, then collected cells and culture supernatant; HSC activation-related protein, including α-smooth muscle actin (α-SMA), and transforming growth factor-β1 (TGF-β1) expression levels were detected by Western blot analysis, the main extracellular matrix including laminin (LN) , hyaluronic acid (HA), collagen Ⅳ (COL-Ⅳ) and procollagen Ⅲ(P Ⅲ NP) secretion level was detected by Elisa assay.@*Results@#CCK-8 assay showed that the cell viability of Lx-2 cells were increased obviously at low doses (≤1.0 μmol/L) of arsenic exposure, especially at 48 and 72 h. In contrast, with the increasing doses of arsenic exposure, the survival rate of Lx-2 cell was decreased gradually, and the survival rate of the high-dose (50, 100 μmol/L) arsenic exposure group at 24, 48 and 72 h were significantly lower than 0.0 μmol/L group, P<0.05. The IC50 of NaAsO2 on Lx-2 cells at 24, 48, 72 h were calculated as 72.75, 48.19 and 29.95 μmol/L, respectively; The expression levels of HSC activation-related protein showed that, after treated with 1.875, 3.750, 7.500, 15.000 μmol/L NaAsO2 for 24 h, α-SMA and TGF-β1 protein level were higher than 0.000 μmol/L group. The increased expression of α-SMA and TGF-β1 protein were most significant in 7.500 μmol/L NaAsO2 group (P<0.05). In addition, the expression levels of α-SMA and TGF-β1 also showed a time-dependent increasing in Lx-2 cells after treated with 7.500 μmol/L NaAsO2 for 0, 12, 24, 48 and 72 h; Elisa assay showed that after treated with 1.875, 3.750, 7.500, 15.000 μmol/L NaAsO2 for 24 h, the secretion levels of HA, LN, COL-Ⅳ and PⅢNP were obvious higher than 0.000 μmol/L group (P<0.05). Moreover, the secretion levels of HA, LN, COL-Ⅳ and P Ⅲ NP also showed a time-dependent increased manner in Lx-2 cells after exposed to 7.500 μmol/L NaAsO2 for 0, 12, 24, 48 and 72 h (P<0.05).@*Conclusion@#NaAsO2 exposure to Lx-2 cells can upregulate the expression level of HSC activation-related proteins, induce its further activation, then increase ECM secretion level.

The differential expression and significance of regulatory T cell and T helper 17 related immunologic factors in peripheral blood of arsenic-exposed rats / 中华地方病学杂志

Objective To investigate the differential expression levels and significance of regulatory T cell (Treg) and T helper 17 (Th17) related immunologic factors in peripheral blood of arsenic-exposed rats.Methods Thirty-two Wistar rats were numbered by weight,randomly divided into four groups [control,low (1.25 ml/kg),medium (2.50 ml/kg),and high (5.00 ml/kg)],8 rats per group.Rats in control group were given oral gavage of deionized water,and low,medium and high arsenic exposed groups were given oral gavage doses of 2.00 g/L sodium arsenite (NaAsO2) according to their body weight for 6 days every week.After 4 months,the urine and peripheral blood samples of rats were collected,urinary arsenic (uAs) was detected by inductively coupled plasma-mass spectrometry (ICP-MS),the results were shown in [median (minimum and maximum)],uAs was corrected by urinary creatinine (uCr),the unit was μg/g Cr;enzyme-linked immune-sorbent assay (ELISA) was applied to detect the levels of Treg,Th17,T lymphocytes activation related immunologic factors [interleukin-10 (IL-10),transforming growth factor beta1 (TGF-31),IL-17,IL-6,IL-2],the results were shown in (x) ± s.Results The uAs of the rats was compared between control,low,medium,and high arsenic exposed groups [7.50 (3.16-9.81),72.34 (62.34-106.63),209.15 (154.41-232.20),369.73 (289.50-516.55) μg/g Cr],the differences were statistically significant (F =337.55,P ＜ 0.05).IL-10 [(85.03 ± 7.11),(93.96 ± 8.14),(97.48 ± 6.23),(93.47 ± 4.41) ng/L],TGF-β1 [(72.88 ± 2.96),(81.45 ± 8.15),(86.08 ± 7.55),(90.29 ± 5.35) ng/L],IL-17 [(18.15 ± 3.66),(25.54 ± 5.59),(31.48 ± 5.74),(37.25 ± 7.36) ng/L],IL-6 [(83.68 ± 8.48),(85.14 ± 7.11),(89.78 ± 5.36),(93.48 ± 5.77) ng/L],and IL-2 [(80.65 ± 6.90),(73.86 ± 8.00),(69.93 ± 7.77),(62.06 ± 9.82) ng/L] of the rats were compared between control,low,medium,and high arsenic exposed groups,the differences were statistically significant (F =5.094,11.054,16.249,3.474,5.119,all P ＜ 0.05).There were positive correlations between uAs and TGF-β1,IL-17 concentration (r =0.723,0.605,all P ＜ 0.01),while IL-2 showed a negative correlation (r =-0.484,P ＜ 0.05).Concltsion Arsenic exposure could affect the secretion of Treg and Th17 related immunologic factors,so as to the imbalance of anti-inflammatory and pro-inflammatory,which may play a role in the formation and development of arsenic-related immune injury.