Cellular energetics and glutathione status in NRK-52E cells: toxicological implications

Cellular energetics and redox status were evaluated in NRK-52E cells, a stable cell line derived from rat proximal tubules. To assess toxicological implications of these properties, susceptibility to apoptosis induced by S-(1,2-dichlorovinyl)-L-cysteine (DCVC), a well-known mitochondrial and renal cytotoxicant, was studied. Cells exhibited high activities of several glutathione (GSH)-dependent enzymes, including gamma-glutamylcysteine synthetase, GSH peroxidase, glutathione disulfide reductase, and GSH S-transferase, but very low activities of gamma-glutamyltransferase and alkaline phosphatase, consistent with a low content of brush-border microvilli. Uptake and total cellular accumulation of [14C]alpha-methylglucose was significantly higher when cells were exposed at the basolateral as compared to the brush-border membrane. Similarly, uptake of GSH was nearly 2-fold higher across the basolateral than the brush-border membrane. High activities of (Na(+)+K(+))-ATPase and malic dehydrogenase, but low activities of other mitochondrial enzymes, respiration, and transport of GSH and dicarboxylates into mitochondria were observed. Examination of mitochondrial density by confocal microscopy, using a fluorescent marker (MitoTracker Orange), indicated that NRK-52E cells contain a much lower content of mitochondria than rat renal proximal tubules in vivo. Incubation of cells with DCVC caused time- and concentration-dependent ATP depletion that was largely dependent on transport and bioactivation, as observed in the rat, on induction of apoptosis, and on morphological damage. Comparison with primary cultures of rat and human proximal tubular cells suggests that the NRK-52E cells are modestly less sensitive to DCVC. In most respects, however, NRK-52E cells exhibited functions similar to those of the rat renal proximal tubule in vivo.     

二硫化碳作业工人的神经肌电图及大脑诱发电位研究

本文对96名二硫化碳作业工人进行了神经肌电图测定,并对其中部分工人作了胫后神经体感诱发电位、视觉诱发电位和脑干听觉诱发电位检查。结果发现,二硫化碳作业工人的神经肌电图改变,符合周围神经远端轴索病之特点。胫后神经体感诱发电位结果显示,脊髓传导时间延长,脊髓传导速度减慢,颈髓以上中枢传导时间与对照组相比,无显著差异。视觉诱发电位和脑干听觉诱发电位无明显改变。     

职业接触二硫化碳生物接触限值的探讨

目的 探讨我国职业接触二硫化碳(CS2)的生物接触限值.方法 用高效液相色谱法测定工人班末尿中2-硫代噻唑烷-4-羧酸(TTCA)含量,用气相色谱法测定接触CS2工人作业场所空气中CS2浓度,探讨二者的相关关系,比较生物接触限值和PC-TWA判定结果.结果 CS2作业工人班末尿中TTCA含量与其接触的工作场所空气中CS2浓度呈正相关,回归方程式Y=0.265X-0.165 (r=0.91,P＜0.01).基于本次研究的回归方程,根据GBZ 2.1-2007《工作场所有害因素职业接触限值第1部分:化学有害因素》规定的CS2的PC-TWA 5 mg/m3推算,CS2接触工人班末尿中TTCA浓度生物限值为1.2 mgTTCA/g Cr.结论 建议我国CS2生物接触限值修订为1.2 mg TTCA/g Cr.     

二烯丙基三硫对胃癌MGC-803细胞的生长抑制作用

目的　研究二烯丙基三硫 (DATS)在体外对胃癌MGC 80 3细胞的生长抑制作用。方法　采用MTT法、集落形成率及生长曲线绘制分别观察不同浓度DATS对胃癌MGC 80 3细胞生长的影响。结果　DATS处理后 ,培养的MGC 80 3细胞增殖抑制而稀少。不同浓度DATS对胃癌MGC 80 3细胞的作用 ,4、8、12、16、2 4mg·L-1的细胞生长抑制率分别为 2 6 %、4 6 %、6 5 %、76 %、89% ,其半数抑制浓度(IC50 )为 8 2mg·L-1。 8、12、16、2 4mg·L-1的细胞集落形成率和集落形成相对数各为 32 4 %和 5 8 7%、2 4 8%和4 2 5 %、19%和 33 5 %、8 8%和 15 1% ,皆具有明显的量效关系 ,且随着浓度增高 ,细胞生长曲线趋于低平。结论　DATS在体外对胃癌MGC 80 3细胞具有良好的抑制作用     

Chemical synthesis of β‐defensins and LEAP‐1/hepcidin

Abstract: A large and steadily growing subfamily of antimicrobially active peptides of animals and plants is formed by the defensins, which are highly disulfide‐bonded, cationic peptides with a molecular mass of about 4 kDa. The synthesis of the human β‐defensins 1 and 2 (hBD‐1, hBD‐2) as well as of the novel murine β‐defensins 7 and 8 (mBD‐7 and mBD‐8) is reported. The peptides were synthesized by solid‐phase peptide synthesis using fluorenylmethoxycarbonyl chemistry. The linear products were oxidized in the presence of the cysteine/cystine redox system to the biologically active molecules. The correct disulfide connectivity of the resulting cyclic products was partly verified by mass spectrometry and sequence analysis of the fragments obtained after tryptic cleavage. In addition, the recently discovered antimicrobially active human peptide LEAP‐1/hepcidin, which contains four disulfide bonds, was successfully synthesized and subsequently oxidized. For Liver‐expressed anti microbial peptide (LEAP)‐1/hepcidin and hBD‐1, the identity of native and synthetic peptides was demonstrated by high‐pressure liquid chromatography and capillary electrophoretic analysis. The general synthetic procedure is suitable to rapidly perform the total chemical synthesis of novel fully bioactive defensins, which are expected to be identified soon, as well as of structurally modified analogs.     

Biological monitoring of workers exposed to carbon disulfide (CS2) in a viscose rayon fibers factory

The exposure-excretion relationship to carbon disulfide (CS₂) vapor in 407 exposed workers was studied during the second half of the working week. Carbon disulfide concentrations were also determined in 50 nonexposed subjects. The geometric mean value for CS₂ in urine samples from the latter was: 0.23 μg/l (95% upper limit = 0.52 μg/l) when log-normal distribution was assumed. Among the exposed workers, the CS₂ level in urine samples collected after the first half shift exceeded the 95% upper limit of nonexposed subjects in every case. The time-weighted average intensity of exposure to CS₂ vapor was measured using personal diffusive samplers (in which carbon cloth served as an adsorbent). CS₂ concentrations in urine were determined in samples collected at the end of the first half shift from the 407 exposed cases as well as from 50 nonexposed controls. There was a significant correlation (p < 0.0001) between the exposure to CS₂ vapor at concentrations of up to 64 mg/m³ and the levels of CS₂ measured in the urine samples after four hours of exposure. The correlation indicated that a mean level of 15.5 μg CS₂/l urine (95% confidence range, 13.8–17.1 μg/l) was excreted following an exposure to CS₂ at 31 mg/m³ (the current occupational exposure limit). Am. J. Ind. Med. 33:478–484, 1998. © 1998 Wiley-Liss, Inc.     

慢性低浓度二硫化碳接触作业工人血脂的变化

对某化纤厂99名长期接触低浓度二硫化碳（CS2）作业工人和28名同厂非CS2作业工人的血脂进行流行病学横断面调查。结果发现：除接触组脂蛋白（a）［Lp（a）］浓度明显高于对照组外,其它血脂指标差别均无统计学意义;进一步通过多元逐步回归分析发现,长期低浓度CS2接触对Lp（a）的影响最大,二者呈正相关（r＝0．43,P＜0．01）。提示Lp（a）可能在CS2致脂质代谢障碍毒作用机制中起着重要作用。     

混合二烷基二硫溶硫性能的评定及溶硫机理的初探

分析我国某油田Merox法所得二硫化物,发现是一组二烷基二硫混合物(Ⅰ),其烷基的含碳数为1—5个。Ⅰ和二乙胺混合(重量比100∶7.1或100∶30)后,室温溶硫量超过传统的溶硫剂二硫化碳。溶硫机理是:Ⅰ在二乙胺催化下和硫发生反应,耗去一部分硫,生成的二烷基多硫(Ⅱ),通过物理作用继续溶解硫。Ⅱ在质谱中的开裂规则是逐个脱下硫,直到成为Ⅰ为止,然后以Ⅰ的开裂方式开裂。     

Arsenic-induced hepatic mitochondrial toxicity in rats and its amelioration by diallyl trisulfide

The present investigation was aimed to investigate the possible protective role of diallyl trisulfide (DATS) against arsenic (As)-induced hepatic mitochondrial toxicity in rats. Mitochondria were isolated from the liver tissue of rats from all the groups. Lipid profile, lipid peroxidation, antioxidant enzyme activities, hepatic function enzymes, mitochondrial swelling, cytochrome c oxidase activity, mitochondrial Ca⁺-ATPase and Na⁺/K⁺-ATPase activity, mitochondrial calcium content and mitochondrial enzyme activities were measured. Short-term As exposure (5?mg/kg?bw/d for 28?d) caused liver damage as evidenced by changes in activities of liver enzymes. The effects of As were coupled with enhanced reactive oxygen species generation, mitochondrial swelling, inhibition of cytochrome c oxidase, complex I-mediated electron transfer, decreased Ca²⁺-ATPase and Na⁺/K⁺-ATPase activity, a reduction in mitochondrial calcium content, changes in indices of hepatic mitochondrial oxidative stress, significant increase in mitochondrial lipid peroxidation products and alterations in mitochondrial lipid profile. Significant decreases in mitochondrial antioxidants and tricarboxylic acid cycle enzymes were also found in the liver mitochondria of As-induced hepatic mitochondrial toxicity in rats. As also increased hepatic caspase-3 activity and DNA fragmentation. All these apoptosis-related molecular changes caused by As could be alleviated by supplementation with DATS, which likely suggests a protective role against As-induced hepatotoxic changes and hepatic mitochondrial toxicity. The protective effect of DATS on the liver mitochondria was evidenced by altering all the changes induced by As. Free radical scavenging and metal chelating activities of DATS may be the mechanism, responsible for the protective action against As-induced mitochondrial damage in liver.     

Preparation of protected peptidyl thioester intermediates for native chemical ligation by Nα‐9‐fluorenylmethoxycarbonyl (Fmoc) chemistry: considerations of side‐chain and backbone anchoring strategies,and compatible protection for N‐terminal cysteine*,†

Abstract: Native chemical ligation has proven to be a powerful method for the synthesis of small proteins and the semisynthesis of larger ones. The essential synthetic intermediates, which are C‐terminal peptide thioesters, cannot survive the repetitive piperidine deprotection steps of N^α‐9‐fluorenylmethoxycarbonyl (Fmoc) chemistry. Therefore, peptide scientists who prefer to not use N^α‐t‐butyloxycarbonyl (Boc) chemistry need to adopt more esoteric strategies and tactics in order to integrate ligation approaches with Fmoc chemistry. In the present work, side‐chain and backbone anchoring strategies have been used to prepare the required suitably (partially) protected and/or activated peptide intermediates spanning the length of bovine pancreatic trypsin inhibitor (BPTI). Three separate strategies for managing the critical N‐terminal cysteine residue have been developed: (i) incorporation of N^α‐9‐fluorenylmethoxycarbonyl‐S‐(N‐methyl‐N‐phenylcarbamoyl)sulfenylcysteine [Fmoc‐Cys(Snm)‐OH], allowing creation of an otherwise fully protected resin‐bound intermediate with N‐terminal free Cys; (ii) incorporation of N^α‐9‐fluorenylmethoxycarbonyl‐S‐triphenylmethylcysteine [Fmoc‐Cys(Trt)‐OH], generating a stable Fmoc‐Cys(H)‐peptide upon acidolytic cleavage; and (iii) incorporation of N^α‐t‐butyloxycarbonyl‐S‐fluorenylmethylcysteine [Boc‐Cys(Fm)‐OH], generating a stable H‐Cys(Fm)‐peptide upon cleavage. In separate stages of these strategies, thioesters are established at the C‐termini by selective deprotection and coupling steps carried out while peptides remain bound to the supports. Pilot native chemical ligations were pursued directly on‐resin, as well as in solution after cleavage/purification.