Immunotoxicity of bisphenol a to Carassius auratus Lymphocytes and macrophages following in vitro exposure

Bisphenol A （BPA） is the monomer component of polycarbonate plastics and classified as an endocrine disrupting chemical （EDC）. The reproductive toxicity of BPA has been extensively studied in mammals; however, relatively little information is available on the immunotoxic responses of fish to BPA. In this study, we investigated the effects of BPA on the immune functions of lymphocytes and macrophages in Carassius auratus. The effects of BPA were compared with those of two natural steroid hormones, estradiol and hydrocortisone. Proliferation of the two types of cells in response to PHA was measured using colorimetric MTT assay. Macrophage respiratory burst stimulated by Con A was measured using chemiluminescence assay. Results showed that BPA （0.054-5.4 mg/L）, estradiol （0.0002-2.0 mg/L） and hydrocortisone （5-50 mg/L） significantly induced Carassius auratus lymphocyte proliferation while higher doses of hydrocortisone （500-5000 mg/L） appeared to be inhibitory. BPA （0.005-50 mg/L）, estradiol （0.005-800 mg/L） and hydrocortisone （0.005-500 mg/L） markedly enhanced macrophage proliferation, whereas higher doses of BPA （500-1000 mg/L） appeared to inhibit cell proliferation. Furthermore, higher dosage of BPA （50 mg/L） and hydrocortisone （50 and 500 mg/L） suppressed the macrophages respiratory burst while estradiol is stimulative all the doses tested （0.05-500 mg/L）. In conclusion, BPA could have immunotoxicity to Carassius auratus and functional changes of lymphocyte and macrophage in Carassius auratus may be different between low and high dosages.     

Evaluation of protective effects of sulforaphane on DNA damage caused by exposure to low levels of pesticide mixture using comet assay

The objective of the study was to evaluate the potential risk of DNA damage due to exposure to a mixture of the most widely used pesticides, namely endosulfan, chlorpyriphos and thiram at an environmentally relevant concentration (5 μM each) and the DNA protective capacity of sulforaphane (SFN) (10–30 μg/mL). DNA damage in human lymphocytes was ascertained with Single Cell Gel Electrophoresis (SCGE), also called Comet Assay. For positive control, H₂O₂ at 100 mM was used. The pesticide mixture produced DNA damage at the concentration used in the lymphocytes. SFN was able to offer a statistically significant (P < 0.01), concentration-dependant protection to DNA damage between 10–20 μg/mL in both the pre-incubation and co-incubation strategies. The results indicate that exposure to low levels of these pesticide mixtures can induce DNA damage, and the presence of SFN in diet may reduce the incidence of genetic damage, especially in farm workers. However, it is not clear whether SFN is involved in quenching of the free radicals generated by the pesticide mixture or it is involved in DNA repair mechanism.     

Micronucleus frequency in sheep lymphocytes after in vitro exposure to fungicide tolylfluanid

The fungicide tolylfluanid (N - dichlorofluoromethylthio-N′, N - dimethyl - N - p - tolylsulfamide), was investigated by cytokinesis-block micronucleus assay. Tolylfluanid at the lowest concentration (1 × 10^{? 6}mol L^{? 1})did not influence significantly the frequency of micronuclei in sheep lymphocyte cultures in comparison with control (32.33 ± 3.51/1000 binucleated cells versus 30.33 ± 2.82/1000 binucleated cells in dimethylsulfoxide (DMSO) control, P = 0.44). Higher tolylfluanid concentrations (1 × 10^{? 4} and, 1 × 10^{? 5} mol L^{? 1}) resulted in a significant dose-dependent increase in the number of micronuclei in comparison with control (74.00 ± 13.00/1000 binucleated cells and 52.67 ± 10.12/1000 binucleated cells versus 30.33 ± 2. 82/1000 binucleated cells in DMSO control, P = 0.005 and 0.02, respectively, ANOVA followed by Tukey test P < 0.05). Many of the treated cells also possessed multiple micronuclei. Tolylfluanid did not affect the nuclear division index at all treatment concentrations. Our in vitro results thus demonstrate that tolylfluanid had a significant genotoxic effect at only the highest concentration tested.     

松胞素B对人血淋巴细胞和CHL细胞微核率的影响

研究了不同浓度的松胞素Ｂ对人血淋巴细胞和中国仓鼠肺成纤维细胞（ＣＨＬ）分裂吉周期和微核率的影响。结果表明,松胞素Ｂ浓度增高过高能引起细胞微核率增高。在本研究中,对ＣＨＬ细胞微核率试验以３μｇ／ｍＬ松胞素Ｂ较适合;而对人血淋巴细胞微核试验,松胞素Ｂ浓度以２－４μｇ／ｍＬ为宜,松胞表Ｂ浓度过高会提高微核背景值,降低试验的灵敏度和精确性。     

Genotoxic effects of metals on human salivary gland tissue and lymphocytes as detected by the Comet assay

The etiology of salivary gland malignancies still remains unclear. Metal compounds are of special interest since they show ubiquitous presence in the environment, are present in many working places, and are accepted (co-)carcinogens in some other malignancies. Metals enter the body as xenobiotics by inhalation or ingestion. This study investigated the genotoxic potential of sodium dichromate (Na₂Cr₂O₇), nickel sulfate (NiSO₄), cadmium sulfate (CdSO₄) and zinc chloride (ZnCl₂) on human salivary gland cells and lymphocytes. Macroscopically healthy tissue of salivary glands was harvested from 46 patients during surgery and isolated to single cells by enzymatic digestion. The cells were incubated with Na₂Cr₂O₇, NiSO₄, CdSO₄ or ZnCl₂. Na₂Cr₂O₇ was also incubated in combination with the other metal compounds listed. Carcinogenic and co-carcinogenic effects of cadmium were tested by incubation with Na₂Cr₂O₇ and consecutive repair intervals. DNA damage and repair were evaluated by the Comet assay, determining DNA-strand breaks. The extent of damage was quantified using a digital analysis system. Na₂Cr₂O₇ produced significantly enhanced DNA-strand breaks in human salivary gland tissue and lymphocytes. All other metal compounds exerted no damaging effect on both cell types. Co-incubation of Na₂Cr₂O₇ with the other metals revealed a significant additive effect only for CdSO₄. Specific analysis of the influence of cadmium showed a reduction of DNA-repair after Na₂Cr₂O₇-induced strand breaks in salivary gland cells. This study provides evidence that exposure to distinct metals may significantly contribute to malignant salivary gland tumors. In consequence, further studies as epidemiological and toxicological data are warranted to determine the role of distinct metals as potential (co-) carcinogens.     

铝对体外培养鸡脾淋巴细胞的免疫毒性

以鸡脾脏淋巴细胞为试验对象,通过短期体外培养的方法研究了铝暴露对鸡免疫细胞的毒性作用.将不同浓度的AlCl3添加到培养液中(终浓度分别为0、0.2、0.4、0.6、0.8mg·mL-1),对鸡脾淋巴细胞原代培养24h,检测培养液上清中乳酸脱氢酶(LDH)活性,淋巴细胞增殖、白细胞介素-2(IL-2)和肿瘤坏死因子-α(TNF-α)含量.结果表明,与对照组相比,各暴露组培养液上清中LDH活性均显著升高(p<0.05,p<0.01),且随铝暴露浓度的增加,LDH活性逐渐升高,呈明显的剂量-反应关系,表明铝暴露能够破坏鸡脾淋巴细胞膜,导致细胞活性下降;与对照组相比,各暴露组淋巴细胞增殖能力、培养液上清中IL-2和TNF-α含量均显著降低(p<0.05,p<0.01),且随铝暴露浓度的增加,3者均逐渐降低,呈明显的剂量-反应关系,表明铝对体外培养鸡脾淋巴细胞具有一定的免疫毒性作用。     

硝基芳烃化合物诱发细胞微核的比较研究

利用蚕豆根法细胞和人在淋巴细胞的微核试验对６种硝基芳烃化合物的致突变性进行比较研究，结果表明，邻二硝基苯、间二硝基苯、２，４－二硝基甲苯，２，６－二硝基甲苯，对硝基氯苯和对硝基溴苯均能诱发两咱细胞的微核率增加，除了对硝基溴苯外均具有明显剂量一效应关系。引进标准剂量概念比较５种硝基芳烃人合物的致突变性，结果表明，致突变性强弱依次为２，４－二硝基甲苯〉对硝基氯苯〉间二硝基苯〉２，６－二硝基甲苯〉邻二硝     

镉对体外培养鸡脾淋巴细胞钙稳态的影响

为了探讨钙稳态在镉对禽类免疫抑制中的作用,以终浓度为10 pμmol·L-1的CdCI:对急性分离的鸡脾淋巴细胞进行染毒,分别在培养12、24、36、48 h和72 h时收集细胞,采用Eura-2/AM荧光标记、半定量RT-PCR和酶试剂盒比色的方法分别研究了细胞内游离钙离子浓度、CaMmRNA的表达以及钙泵的活性.结果表明,随着镉作用时间的延长,淋巴细胞内游离钙离子浓度不断升高.CaM mRNA表达水平和钙泵活性不断下降,试验组与对照组差异显著(p<0.05或P<0.01),提示镉可使淋巴细胞内钙稳态失调而发挥免疫毒性作用.     

二氧化硫对人体血淋巴细胞的遗传毒理效应

本文对某硫酸厂接触二氧化硫（ＳＯ＿２）４０名工人的外周血淋巴细胞染色体畸变（ＣＡ）、姊妹染色单体互换（ＳＣＥ）、及微核（ＭＮ）进行了分析。结果表明，接触ＳＯ＿２的工人其ＣＡ、ＳＣＥ及ＭＮ均显著高于对照组。本文也分析了ＳＯ＿２在体内的代谢产物──亚硫酸氢钠对淋巴细胞的遗传毒理效应。结果指出，亚硫酸氢钠可诱发人血淋巴细胞ＣＡ，ＳＣＥ及ＭＮ，且有明确的剂量效应关系。这些研究证明，ＳＯ＿２确是人血淋巴细胞染色体断裂剂和基因毒性因子；ＳＯ＿２与癌症的关系值得进一步研究。     

Reduction of Pain in Reflex Sympathetic Dystrophy (RSD) Associated with Objective Biochemical Changes in Circulating Lymphocytes

Summary In an effort to explain the benefit of therapeutic use of electromagnetic fields (EMF) a systemic effect has been proposed by us. To assess the efficacy of this approach, an objective biochemical approach was developed. Ten patients with Reflex Sympathetic Dystrophy (RSD) were clinically evaluated, and lymphocytes were isolated from their blood samples obtained before and after exposure of the uninvolved limb to an EMF. Utilizing a SpectraCell method that includes radio labeled molecules and protein-free media for culturing the lymphocytes, an elevation of the content of fructose, serine, glycine, and calcium cellular metabolic uptake were found following in the culture to EMF in comparison with non-exposed lymphocytes (p < 0.01). In addition, the pain level was determined by a conventional visual analogue scale (VAS) before and after EMF exposure, evidencing a significant pain relief. Specifically, after exposure to 120 pps semi sinewave, 1500Gauss EMF, generated by a THERAMAG^∘ledR device, an improvement in the flexibility of the limb and a reduction in swelling of the affected extremity were detected clinically. These findings are in concert with the new hypothesis that, with relief of pain, lymphocytes are predominately altered in their cell cycle from M phase to S phases associated with increased structuring of intracellular water. A consideration of the basic understanding of the role lymphocytes may be inferred from this preliminary study. An extensive review of the literature on the basic science and therapeutic use of magnetic fields in humans is provided in an attempt to understand the relevance of magnetic therapy of specific pain syndromes.