Antioxidants to prevent bovine neutrophil-induced mammary epithelial cell damage

Activated neutrophils are able to produce a large quantity of bactericidal molecules such as reactive oxygen species that have been associated with tissue damage in several inflammation models. The protective effects of antioxidants in a context of neutrophil-induced damage to mammary epithelial cells were first evaluated in vitro using a coculture model of activated bovine neutrophils and a bovine mammary epithelial cell line (MAC-T cells). Cell damage was determined by quantifying the release of lactate dehydrogenase by MAC-T cells in culture medium. Morphological observation of cells stained with acridine orange was used to visualize the extent of cell damage. When incubated with neutrophils activated by lipopolysaccharides and phorbol 12-myristate 13-acetate, MAC-T cells released large amounts of lactate dehydrogenase indicating significant cell damage. The addition of dimethylthiourea or bathocuproine disulfonic acid did not reduce the damage whereas catechin, deferoxamine or glutathione ethyl ester significantly reduced neutrophil-induced cytotoxicity in a dose-dependent manner. The effect of deferoxamine, an iron chelator, on the growth of Escherichia coli and the ability of bovine neutrophils to phagocytose these bacteria were then assessed in vitro. Our data showed that deferoxamine did not interfere with the phagocytic activity of neutrophils but inhibited growth of the bacteria. Overall, our results suggest that antioxidants may be effective tools for protecting mammary tissue against neutrophil-induced oxidative stress during bovine mastitis.     

Deferoxamine reduces tissue damage during endotoxin-induced mastitis in dairy cows

The protective effects of 3 antioxidants on polymorphonuclear neutrophil-induced damage to mammary cells were evaluated in vivo using an endotoxin-induced mastitis model. Fifteen healthy, midlactation cows with no history of clinical Escherichia coli mastitis were randomly assigned to 1 of the 3 treatment groups corresponding to each modulator to be evaluated, that is, deferoxamine, catechin, and glutathione ethyl ester. Each cow had 1 quarter infused with saline and 1 quarter infused with the selected modulator; a third quarter was infused with lipopolysaccharides (LPS), whereas the fourth quarter received a combination of LPS and the modulator. Infusion of LPS caused acute mastitis as determined by visual observations and by large increases in milk somatic cell count, BSA, and proteolytic activity. These parameters were not affected by antioxidant administration. The extent of cell damage was evaluated by measuring milk levels of lactate dehydrogenase and N-acetyl-β-_D-glucosaminidase activity. Levels of these parameters were several times higher after LPS administration. Intramammary infusions of catechin or glutathione ethyl ester did not exert any protective effect, whereas infusion of deferoxamine, a chelator of iron, decreased milk lactate dehydrogenase and NA-Gase activity, suggesting a protective effect against neutrophil-induced damage. The protective effect of deferoxamine was also evidenced by a lower milk level of haptoglobin. The proteolytic activity of mastitic milk was not influenced by the presence of deferoxamine. Overall, our results suggest that local infusion of deferoxamine may be an effective tool to protect mammary tissue against neutrophil-induced oxidative stress during bovine mastitis.     

Induction of nitric oxide production by bovine mammary epithelial cells and blood leukocytes.

A recent study from our laboratory has shown that significant amounts of nitric oxide are released by somatic cells recovered during endotoxin-induced mastitis. The present study was undertaken to investigate which cell type(s) among milk somatic cell population can produce nitric oxide under inflammatory conditions. Nitric oxide release from mammary epithelial cell lines and from bovine neutrophils and monocytes extracted from blood was measured in response to cytokines and Escherichia coli lipopolysaccharides. An epithelial cell line isolated from bovine mammary gland, FbE cells, was found to release nitric oxide after exposure to interleukin-1beta. This nitric oxide production was completely abolished by addition of L-N6-(1-iminoethyl) lysine, a potent inducible nitric oxide synthase inhibitor. Bovine monocytes produced nitric oxide in response to recombinant bovine interferon-gamma alone or in combination with E. coli lipopolysaccharides. In these cells, nitric oxide release was reduced by the addition of inducible nitric oxide synthase inhibitors L-N6-(1-iminoethyl) lysine and aminoguanidine. Lipopolysaccharides and recombinant bovine interferon-gamma increased nitric oxide synthase mRNA in neutrophils, but nitric oxide release could not be detected under any of the experimental conditions used. These results show that bovine epithelial cells and mononuclear phagocytes produce nitric oxide under inflammatory conditions and suggest that these cell populations are responsible for nitric oxide release observed during mastitis.     

二氢杨梅素及其酰化衍生物对肝细胞氧化损伤的保护作用

目的：考察二氢杨梅素(DHM)及其衍生物对肝细胞氧化损伤的保护作用。方法：建立L02细胞过氧化氢损伤模型,通过酶法酰化修饰DHM得到不同亲脂性的DHM衍生物,再通过测定活性氧(ROS)、乳酸脱氢酶(LDH)、丙二醛(MDA)、过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)水平、线粒体膜电位和Caspase 3水平评估DHM及其衍生物对肝损伤细胞的保护作用。结果：除3-O-月桂酰化二氢杨梅素外,其余衍生物处理后的肝损伤细胞中活性氧(ROS)、乳酸脱氢酶(LDH)的释放和丙二醛(MDA)水平显著降低,抗氧化酶系水平升高,线粒体膜电位升高,Caspase 3水平降低,其中3-O-辛酰化二氢杨梅素的保护效果最好。结论：中等链长二氢杨梅素衍生物有较好的护肝效果。     

具抗氧化功能益生菌菌株筛选及其对丙烯酰胺诱导肠上皮细胞氧化损伤的保护作用

目的:以1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl,DPPH)自由基清除率为指标,筛选抗氧化能力强的乳杆菌,并研究其对模拟胃肠道环境的耐受性及对丙烯酰胺所致肠上皮细胞氧化损伤的保护作用。方法:以DPPH自由基清除率为指标,筛选得到清除率高的2株乳杆菌GBE17和GBE29,并进行16S rDNA测序鉴定。构建丙烯酰胺诱导的Caco-2氧化损伤模型,通过形态学观察和测定细胞培养上清液、细胞裂解液中的抗氧化物相关物质及酶活,评价其不同处理方式对Caco-2细胞氧化损伤的保护作用,并分别测定其在pH 3.0、2.5及胆盐质量分数为0.05%、0.1%环境下处理1~4 h的活菌数变化。结果:植物乳杆菌GBE17和唾液乳杆菌GBE29发酵上清液的DPPH自由基清除率分别为89.44%、79.24%。GBE17的治疗组与干预组和GBE29的干预组均可降低乳酸脱氢酶的释放,提高细胞内外超氧化物歧化酶、过氧化氢酶活性。2株乳杆菌在pH 3.0和胆盐质量分数0.1%的环境下处理4 h,存活数均高于107 CFU/mL。结论:植物乳杆菌GBE17和唾液乳杆菌GBE29通过提高细胞内抗氧化酶系的活性,有效地降低丙烯酰胺诱导肠上皮细胞的氧化损伤,且具有较好的胁迫耐受能力。     

Intracellular accumulation, subcellular distribution, and efflux of tilmicosin in bovine mammary, blood, and lung cells.

Tilmicosin is a semisynthetic macrolide antibiotic currently approved for veterinary use in cattle and swine to combat respiratory disease. Because the concentrations of tilmicosin are generally low in bovine serum, the interaction of tilmicosin with three types of bovine phagocytes (monocyte-macrophages, macrophages, and neutrophils from blood, lungs, and mammary gland, respectively) and mammary gland epithelial cells was evaluated to provide an understanding of potential clinical efficacy. After incubation with radiolabeled tilmicosin, uptake was determined and expressed as the ratio of the intracellular to the extracellular drug concentration. Accumulation of tilmicosin at 4 h of incubation by the alveolar macrophages (Cc/Ce 193) was 4 to 13 times more than that observed in monocyte-macrophages (Cc/Ce 43), neutrophils, (Cc/Ce 13), or mammary epithelial cells (Cc/Ce 20). Subcellular distribution showed that 70 to 80% of tilmicosin was localized in the lysosomes. Uptake in mammary gland cells was dependent on cell viability, temperature, and pH, but was not influenced by metabolic inhibitors or anaerobiosis. However, lipopolysaccharide exposure increased tilmicosin uptake by the bovine mammary macrophages and epithelial cells. When neutrophils and epithelial cells were incubated in the presence of tilmicosin and extracellular tilmicosin was then removed, 40% of the intracellular tilmicosin remained cell associated after 4 h of incubation (i.e., 60% effluxed), but only 25% remained in macrophages. These in vitro interactions of tilmicosin with bovine phagocytes and epithelial cells suggest an integral role in effecting clinical efficacy.     

鲜切果蔬活性氧产生和抗氧化体系代谢的研究进展

新鲜果蔬经切割后,诱导组织产生O2―?、H2O2和?OH等,均可破坏植物组织中正常的活性氧代谢平衡。但是,鲜切处理同时诱导果蔬启动抗氧化系统而主动防御。本文就鲜切果蔬O2―?、H2O2等的产生,超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)等抗氧化关键酶活性以及抗坏血酸、多酚化合物、还原型谷胱甘肽(GSH)等抗氧化物质的研究进展进行综述,以期为鲜切果蔬针对机械伤害的响应机制方面的研究提供一定的参考。     

黑灵芝多糖对丙烯酰胺诱导小肠上皮细胞氧化损伤的保护作用

目的：研究黑灵芝多糖对丙烯酰胺（acrylamide,AA）所致的小肠上皮细胞氧化损伤的保护作用。方法：构建AA诱导小肠上皮细胞（intestinal epithelial cells,IEC）-6氧化损伤型,采用噻唑蓝比色法检测黑灵芝多糖对IEC-6氧化损伤的保护作用;同时测定细胞培养液中乳酸脱氢酶（lactate dehydrogenase,LDH）的漏出量,以及测定细胞中超氧化物歧化酶（superoxide dismutase,SOD）、谷胱甘肽过氧化物酶（glutathione peroxidase,GSH-Px）及丙二醛（malondialdelyde,MDA）水平。结果：与正常对照组相比,不同质量浓度的黑灵芝多糖对细胞增殖影响无显著性差异。与模型（AA）组相比,不同质量浓度的黑灵芝多糖溶液能够明显减轻AA对细胞的毒害作用,提高细胞生存率,降低细胞培养液中LDH的释放;降低MDA含量,提高细胞SOD和GSH-Px活性。结论：黑灵芝多糖可以通过提高细胞内抗氧化酶系的活性和抗氧化物质GSH-Px活性从而有效地减弱AA诱导的小肠上皮细胞氧化损伤。     

Sensitivity of antioxidant-deficient yeast to hypochlorite and chlorite

Sodium hypochlorite and sodium chlorite are commonly used as disinfectants, and understanding the mechanisms of microbial resistance to these compounds is of considerable importance. In this study, the role of oxidative stress and antioxidant enzymes in the sensitivity of the yeast Saccharomyces cerevisiae to hypochlorite and chlorite was studied. Yeast mutants lacking Cu-Zn superoxide dismutase, but not mutants deficient in cytoplasmic and peroxisomal catalase, were hypersensitive to the action of both hypochlorite and chlorite. Both compounds depleted cellular glutathione, induced the production of reactive oxygen species and decreased the viability of the cells. The toxicity of hypochlorite and chlorite was abolished by hypoxic and anoxic conditions and ameliorated by thiol antioxidants and ascorbate. The results demonstrated that the action of hypochlorite and chlorite involves the formation of superoxide and peroxide and that SOD1 is protective, probably by limiting the formation of hydroxyl radicals and damage to proteins.     

Effect of date seeds on oxidative damage and antioxidant status in vivo

BACKGROUND: Date seeds have been shown to contain high amounts of antioxidants. However, in vivo studies on date seeds are lacking. Therefore the purpose of this study was to determine the effect of date seeds on oxidative damage and antioxidant status in vivo. Male Wistar rats were fed a basal diet containing 0, 70 or 140 g kg^?1 date seeds for 30 days. All three diets were isonitrogenous and isocaloric. Indication of oxidative damage was assessed in the liver and serum, and antioxidant status was assessed in the liver. Serum biochemical parameters, including indicators of tissue cellular damage and complete blood count with differential, were also determined. RESULTS: The results showed that date seeds significantly (P < 0.05) reduced liver and serum malondialdehyde (a lipid peroxidative damage product) and serum lactate dehydrogenase and creatine kinase. Liver antioxidants (vitamin E, vitamin C, glutathione, superoxide dismutase, glutathione peroxidase and catalase), complete blood count with differential and other serum biochemical parameters assessed were not significantly altered by date seeds. CONCLUSION: The results obtained suggest a protective effect of date seeds against in vivo oxidative damage, possibly through the action of their bioactive antioxidants. Copyright © 2011 Society of Chemical Industry     

金银花叶黄酮体外抗氧化能力及对H2O2诱导RAW264.7巨噬细胞损伤的保护作用

目的：研究金银花叶黄酮的体外抗氧化能力和对H2O2诱导RAW264.7巨噬细胞损伤的保护作用。方法：金 银花叶粉经提取纯化后得到金银花叶黄酮粉,以抗坏血酸为阳性对照,测定金银花叶黄酮的总还原力,对羟自由 基、超氧阴离子自由基及1,1-二苯基-2-三硝基苯肼自由基的清除能力。体外培养RAW264.7巨噬细胞,实验分为空 白组、模型组、对照组和金银花叶黄酮低、中、高剂量组,用H2O2诱导损伤RAW264.7细胞,噻唑蓝法测定细胞存 活率,试剂盒法测定细胞和细胞培养液中丙二醛、谷胱甘肽含量及超氧化物歧化酶、乳酸脱氢酶活力。结果：金银 花叶黄酮的总还原力及对各自由基的清除能力较强,并在足够质量浓度下等同于对照品抗坏血酸。金银花叶黄酮呈 剂量依赖性保护H2O2引起的RAW264.7细胞的损伤,降低细胞及细胞培养液中丙二醛含量,提高超氧化物歧化酶活力 及谷胱甘肽含量,提高细胞内乳酸脱氢酶活力。结论：金银花叶黄酮抗氧化能力较强,可修复H2O2诱导的RAW264.7 巨噬细胞的损伤,其作用可能与调节细胞氧化还原系统、清除自由基、提高细胞内抗氧化酶系的活力有关。     

巴氏灭活嗜黏蛋白阿克曼氏菌对ox-LDL诱导的HAEC细胞损伤的保护作用及机制初探

目的:本研究以巴氏灭活的嗜黏蛋白阿克曼氏菌（pasteurized Akkermansia muciniphila,PAKK）为研究对象,探究其对氧化低密度脂蛋白（oxidized low density lipoprotein,ox-LDL）诱导的人主动脉内皮细胞（human aortic endothelial cells,HAEC）损伤的保护作用。方法:首先利用MTT法评价PAKK对细胞活性的影响并确定合适的剂量。建立ox-LDL诱导的细胞损伤模型,并通过测定细胞乳酸脱氢酶、活性氧等评价PAKK对细胞损伤的保护作用,最后通过测定抗氧化酶活力以及Nrf2抗氧化通路相关基因及蛋白表达等,探讨PAKK改善HAEC细胞氧化损伤的可能机制。结果:菌数为105和106 CFU/mL的PAKK不影响HAEC细胞的活性;能显著降低ox-LDL诱导HAEC细胞的乳酸脱氢酶释放率;显著降低细胞内的活性氧水平,提高超氧化物歧化酶、过氧化氢酶的活力,以及细胞的总抗氧化能力;显著上调细胞内转录因子核因子-E2相关因子2（nuclear factor erythroid 2-related factor 2,Nrf2）、血红素氧合酶-1（hemeoxygenase-1, HO-1）、谷胱甘肽巯基转移酶（glutathione S-transferase,GST）以及NADPH醌氧化还原酶1 （NADPH quinine oxidoreductase-1,NQO1）的mRNA表达量;显著上调Nrf2、HO-1、NQO1蛋白的表达。结论:本研究表明巴氏灭活的嗜黏蛋白阿克曼氏菌可缓解ox-LDL对HAEC细胞造成的损伤,且其保护作用可能是通过调节Nrf2信号通路从而增强抗氧化相关基因表达。本研究为开发基于嗜黏蛋白阿克曼氏菌的可用于预防动脉粥样硬化的益生菌相关产品奠定理论基础。     

Characterization of Streptococcus lutetiensis isolated from clinical mastitis of dairy cows

Streptococcus lutetiensis, previously termed Streptococcus bovis type II/1, has rarely been associated with bovine mastitis. The objectives of this work were to characterize the molecular diversity, antimicrobial resistance profiles, virulence genes of Strep. lutetiensis (n = 37) isolated from bovine clinical mastitis, as well as its pathogenic effects in a murine mastitis model. Genetic relationships of isolates were determined by random amplified polymorphic DNA (RAPD)-PCR, virulence genes were detected by PCR. Antimicrobial susceptibility testing was carried out by broth microdilution technique. The pathogenic effects of Strep. lutetiensis were studied with 2 infection models: bovine mammary epithelial cells cultured in vitro and murine mammary infection in vivo. Streptococcus lutetiensis isolates were clustered into 5 RAPD-types (A–E), with a dominant type A representing 84% of isolates. Eighteen (49%), 16 (43%), and 9 (24%) isolates were resistant to ceftiofur, tetracycline, and erythromycin, respectively. Prevalence of multidrug resistance (resistant to ≥3 classes of antimicrobials) was 24% (9/37). The most prevalent virulence genes were bca (100%), speG (100%), hly (97%), scpB (95%), and ssa (95%). There was no difference between isolates from mild and moderate cases of bovine mastitis in prevalence of virulence genes. Streptococcus lutetiensis rapidly adhered to and subsequently invaded (1 and 3 h after infection, respectively) bovine mammary epithelial cells, resulting in elevated lactate dehydrogenase release (4 h after infection). Edema and hyperemia were observed in challenged mammary glands and bacteria were consistently isolated at 12, 24, and 48 h after infection. In addition, numerous neutrophils migrated into gland alveoli and interstitium of infected mammary tissue. We concluded that Strep. lutetiensis had potential to spread within a dairy herd and good adaptive ability in bovine mammary cells or tissue, which are generally characteristics of a contagious mastitis pathogen.     

川皮苷体外抗氧化活性及对肿瘤细胞生长抑制作用

目的：研究川皮苷的抗氧化活性及对肿瘤细胞生长抑制作用。方法：在体外测定川皮苷对1,1-二苯基-2-三硝基苯肼（1,1-diphenyl-2-picrylhydrazyl,DPPH）自由基、羟自由基（•OH）和超氧阴离子自由基（O2－•）的清除能力;通过四甲基偶氮唑蓝（methyl thiazolyl tetrazolium,MTT）法筛选对川皮苷作用敏感的肿瘤细胞株,并用倒置显微镜和透射电镜观察敏感细胞株在川皮苷作用之后细胞结构的变化。结果：川皮苷在体外对DPPH自由基、•OH和O2－•都具有清除能力,其中对•OH的清除能力突出。同时,人胰腺癌细胞株PANC-1对川皮苷作用敏感,即川皮苷可抑制PANC-1细胞的生长;透射电镜观察发现川皮苷可使PANC-1细胞的细胞核膜破裂,胞质溶出,形成凋亡小体。结论：川皮苷具有显著的体外抗氧化活性,并且可以抑制PANC-1细胞的增殖,在功能食品等领域具有良好的应用前景。     

红毛藻多糖对H2O2诱导的Caco-2细胞氧化损伤的保护作用

目的：研究红毛藻多糖（Bangia fusco-purpurea polysaccharide,BFP）对H2O2诱导的人结肠腺癌（Caco-2）细胞氧化应激损伤的保护作用。方法：构建过氧化氢（H2O2）诱导的Caco-2细胞氧化损伤模型,通过形态学观察和测定细胞裂解液中的抗氧化物水平及相关酶活力,评价BFP对Caco-2细胞氧化损伤的保护作用。结果：BFP可明显提高H2O2诱导氧化损伤的Caco-2细胞活力,降低细胞内活性氧生成水平,提高细胞内超氧化物歧化酶和过氧化氢酶活力以及还原型谷胱甘肽水平,减少丙二醛生成,并通过抑制Caspase-3和Caspase-9活性来改善氧化应激损伤引起的细胞凋亡。结论：BFP对H2O2诱导氧化应激损伤的Caco-2细胞具有保护作用,可通过增强细胞内源性抗氧化酶活力、提高非酶类抗氧化物水平和抑制细胞凋亡明显缓解H2O2诱导的Caco-2细胞氧化应激损伤。     

Evidence for a local effect of leptin in bovine mammary gland

On average, high-energy diets promoting body growth rates above 1 kg/d before puberty impair mammary development by 15 to 20% in cattle. We hypothesized that leptin, a protein produced by adipocytes, mediates the inhibitory effect of high-energy diets on mammary development. Therefore, our objectives were to determine the effect of leptin on mammary epithelial cell proliferation, and the distribution of mRNA for two leptin receptor isoforms in prepubertal bovine mammary glands and other peripheral tissues. Addition of leptin to culture media containing either 5 ng/ml of insulin-like growth factor-I (IGF-I) or 1% fetal bovine serum decreased DNA synthesis of a bovine mammary epithelial cell line (MAC-T) in a dose-dependent manner. The minimal doses of leptin that decreased IGF-I- and fetal bovine serum-stimulated cell proliferation were 64 and 1 ng/ml, respectively. In addition, we determined that MAC-T cells and isolated bovine mammary epithelial cells express the long form of leptin receptor (Ob-Rb) mRNA. Ob-Rb mRNA was detected in all bovine tissues examined. In contrast with reports on other species, mRNA expression of the short form of leptin receptor (Ob-Ra) was detected only in bovine liver, pituitary body, and spleen. These results support the concept that leptin mediates the inhibitory effect of high-energy diets on mammary development.     

In vitro differentiation of a cloned bovine mammary epithelial cell

The aim of the study was to establish in vitro a bovine mammary epithelial cell (MEC) clone, able to respond to mitogenic growth factors and to lactogenic hormones. Mammary tissue from a 200-d pregnant Holstein cow was used as a source of MEC, from which a clone was established through a process of limiting dilution. When plated on plastic, the cells assumed a monolayer, cobblestone, epithelial-like morphology, with close contact between cells. Inclusion of IGF-1 and EGF in the media significantly increased the number of cells 5 d after plating. All cells stained strongly for cytokeratin and moderately for vimentin at young and old passage stages, indicating the epithelial nature of this cell clone. When the cells were plated at a high density on a thin layer of a commercial extracellular matrix preparation (Matrigel), lobular, alveoli-like structures developed within approximately 5 d, with a clearly visible lumen. When cells were plated onto Matrigel in differentiation media (containing lactogenic hormones), detectable quantities of alpha-casein were present in the media and particularly on the lumen side of the structures. Omission of one of the lactogenic hormones (insulin, prolactin or hydrocortisone) reduced alpha-casein release to the limit of detection of the assay used. Lactoferrin was also produced when the cells were plated on Matrigel, again principally on the lumen side of the lobules, though this was independent of the lactogenic hormones. By passage 40, the cells had senesced, and it was not possible to induce alpha-casein or lactoferrin production. This study notes the establishment of a functional bovine mammary epithelial cell clone, which is responsive to mitogenic and lactogenic hormones and an extracellular matrix.     

Selenomethionine stimulates expression of glutathione peroxidase 1 and 3 and growth of bovine mammary epithelial cells in primary culture

This study examined the localization of cellular glutathione peroxidase (GPx1) and extracellular glutathione peroxidase (GPx3) in lactating mammary tissue and in primary cultures of bovine mammary epithelial cells (BMEC). The effect of selenium as selenomethionine (SeMet) on the growth and viability of BMEC and GPx protein expression and activity were also studied. Single mammary epithelial cells were recovered by serial collagenase/hyaluronidase digestion from lactating bovine mammary tissue and cultured in a low-serum collagen gel system enriched with lactogenic hormones and 0, 10, 20, or 50 nM SeMet. Positive immunostaining with anti-cytokeratin and bovine anti-casein confirmed the epithelial nature and differentiated state of BMEC. Addition of SeMet to media facilitated rapid confluence of BMEC and formation of dome structures. Immunohistochemical and immunocytochemical staining revealed that both GPx1 and GPx3 are synthesized by BMEC and localized in the cytoplasm and nucleus. Up to 50 nM SeMet linearly increased BMEC number and viability over 5 d of culture. Bovine mammary epithelial cells cultured in SeMet-supplemented medium also exhibited markedly elevated GPx activity and linear increases in abundance of GPx1 and GPx3 proteins. It is apparent that SeMet degradation to release Se for synthesis of selenoproteins is carried out by BMEC. Results indicate that bovine mammary epithelial cells express GPx1 and GPx3 in vivo and in vitro; SeMet enhances expression of these selenoproteins in vitro and the growth and viability of BMEC.     

Modulation of the cytotoxicity and genotoxicity of the drinking water disinfection byproduct lodoacetic acid by suppressors of oxidative stress

Drinking water disinfection byproducts (DBPs) are generated by the chemical disinfection of water and may pose a hazard to the public health. Previously we demonstrated that iodoacetic acid was the most cytotoxic and genotoxic DBP analyzed in a mammalian cell system. Little is known of the mechanisms of its genotoxicity. The involvement of oxidative stress in the toxicity of iodoacetic acid was analyzed with the antioxidants catalase and butylated hydroxyanisole (BHA). lodoacetic acid toxicity was quantitatively measured with and without antioxidants in Salmonella typhimurium strain TA100 and with Chinese hamster ovary (CHO) cells. The endpoints included cytotoxicity in S. typhimurium or in CHO cells, mutagenicity in S. typhimurium, and genotoxicity in CHO cells. Neither catalase nor BHA reduced the level of iodoacetic acid induced cytotoxicity in S. typhimurium. In CHO cells neither antioxidant caused a significant reduction in iodoacetic acid induced cytotoxicity. However, in S. typhimurium, BHA or catalase reduced the mutagenicity of iodoacetic acid by 33.5 and 26.8%, respectively. Likewise, BHA or catalase reduced iodoacetic acid induced genomic DNA damage by 86.5 and 42%, respectively. These results support the hypothesis that oxidative stress is involved in the induction of genotoxicity and mutagenicity by iodoacetic acid.     

Mammary epithelial proliferation and estrogen receptor alpha expression in prepubertal heifers: effects of ovariectomy and growth hormone

The objectives of this study were to determine the effects of ovariectomy and growth hormone on mammary epithelial cell proliferation and estrogen receptor alpha (ER alpha) expression within the bovine mammary gland. Two experiments were performed. In the first experiment, eight Holstein heifer calves aged between 1 and 3 mo were ovariectomized, while six calves served as controls. At 6 mo of age, calves were treated with bromodeoxyuridine (BrdU) to label proliferating cells and sacrificed 2 h later. Coinciding with reduced mammary mass (304 +/- 25 vs. 130 +/- 21 g), proliferation of mammary epithelial cells was significantly lower in ovariectomized heifers compared to control heifers (2.24 vs. 0.25%). ER alpha expression was restricted to mammary epithelial cells and was not observed within intra-lobular stroma of parenchymal tissue. The proportion of ER alpha positive cells was significantly higher in ovariectomized heifers than in controls (36.1% +/- 2.2 vs. 46.7% +/- 2.4). In the second experiment, mammary biopsies were taken from five 6-mo-old heifers, immediately preceding and 7 d following a single injection of bovine growth hormone. Mammary epithelial cell proliferation (assessed by incorporation of 3H-thymidine) was increased by growth hormone. The proportion of ER alpha positive mammary epithelial cells was not increased by growth hormone. In conclusion, reduced mammary epithelial cell proliferation following ovariectomy was associated with an increase in ER alpha expression, whereas increased proliferation caused by bovine growth hormone was not associated with changes in the proportion of ER alpha positive cells.