晶状体生化的研究:正常和亚硒酸钠诱发白内障大鼠晶状体中与谷胱甘肽代谢有关的三种酶活性的测定

测定了用亚硒酸钠诱发的大鼠白内障晶状体中谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽还原酶(GSSG-R)和谷胱甘肽硫转移酶(GSH-S)的活性,并与正常晶休中这三种酶的活性作了比较。结果表明,核浊浑期晶状体中GSH-Px的活性比正常晶状体的高一倍,但在整个晶状体浑浊时降低,GSSG-R的活性变化与GSH-PX相似,这两种酶在代谢上是相关的。GSH-S的活性在核浑浊期不改变,但在完全浑浊后降低。     

AC1及AC3抗白内障形成中晶状体谷胱甘肽代谢相关酶活性的变化

观察了亚硒酸钠,AC1,AC3对大鼠晶状体中谷胱甘肽过氧化物酶(GSH-Px),谷胱甘肽还原酶(GR)及谷胱甘肽硫转移酶(GST)的影响。结果表明,亚硒酸钠组大鼠的晶状体尚未混浊前已出现GSH-Px活性增高及GR和GST的活性降低。GR活性下降随白内障进展而加重。AC1及AC3均可使亚硒酸钠所致的酶活性变化逆转,但对正常晶状体的酶活性没有影响。     

抗氧化剂与自由基清除剂对硒性白内障形成的影响

本文观察了8种化合物(抗氧化剂及自由基清除剂)对大鼠亚硒酸钠性白内障的影响。实验分为正常对照组,亚硒酸钠组及药物对抗组。亚硒酸钠组系给13日龄大鼠皮下注射亚硒酸钠(6μmoles/kg体重),间日一次,逐次递增1μmole/kg体重,连续5次,药物对抗组则同时腹腔注射抗氧化剂或自由基清除剂,每日观察并记录白内障的发生频率及程度,实验表明,一些抗氧化剂及自由基清除剂能够有效的对抗亚硒酸钠性白内障的发生发展,其中AC1、AC3及AC3的效果尤为明显。本文的结果为探讨白内障形成机理及防治提供了实验依据。     

硒性白内障大鼠模型晶状体中GR和GSH-Px的表达

 为探讨硒性白内障大鼠晶状体中谷胱甘肽过氧化物酶 (GSH Px)和谷胱甘肽还原酶 (GR)的活性调节在硒性白内障形成中的作用及调节方式 ,采用半定量RT PCR方法 ,比较正常晶状体、核中心混浊晶状体 (核白 )和完全混浊晶状体 (全白 )中GSH Px和GR的mRNA水平及酶活性的变化 .研究发现 ,核白晶状体中 2种酶的活性和mRNA水平均升高 ,其中酶活性的升高幅度小于mRNA水平 .随着白内障的发展 ,2种酶的活性和mRNA水平均逐渐下降 .至晶状体全白时 ,2种酶的活性均显著低于正常 ;全白时GR的mRNA水平降至正常 ,GSH Px的mRNA水平则仍高于正常 .结果表明 ,硒性白内障形成与细胞内GSH Px和GR的活性调节密切相关 ,GSH Px和GR的活性调节可能主要发生在转录水平     

亚硒酸钠诱发大鼠白内障晶状体前列腺素生物合成的研究

 用亚硒酸钠诱发大鼠产生白内障后,将晶状体微粒体与外源性花生四烯酸共同孵育,用放射免疫方法测定白内障晶状体前列腺素E_2(PGE_2)及前列腺素F_2α(PG-F_2α)的生物合成情况,并与正常晶状体进行了比较,结果表明大鼠晶状体具有酶促合成PGs的能力。正常晶状体及白内障晶状体合成PGE_2的能力分别为687.75±113.97及1095.00±79.39pg/100mg晶状体湿重/15分钟,PGE_2α则分别为51.45±36.72及158.83±115.94pg/100mg晶状体湿重/15分钟(平均数±S.D.)。这说明大鼠白内障晶状体合成PGs的能力明显增高,与正常晶状体相比有显著性差异(PGE_2P<0.001,PGF_2αP<0.02)。在前2次注射亚硒酸钠后,大鼠白内障晶状体PGs的合成能力逐渐高于正常晶状体,并随注射亚硒酸钠的次数增加和白内障晶状体混浊程度加重,PGs在晶状体内的含量增加。     

黄芩黄酮对硒性白内障晶状体抗氧化酶表达的影响

为探讨黄芩黄酮防治白内障的作用机理 ,采用半定量RT PCR方法比较正常组、白内障组和中药防治组大鼠晶状体中GSH Px、GR和Cu ZnSOD的mRNA水平 .白内障组GSH Px、GR和Cu ZnSOD的mRNA水平在 15d龄时显著高于正常 ,然后下降 ;在 2 7d和 31d龄 ,GR和Cu ZnSOD的mRNA水平下降至与正常无显著差异 ,GSH PxmRNA水平仍略高于正常 .中药防治组晶状体中 ,3种抗氧化酶的mRNA水平在各实验取样点无明显变化 ;其中 ,GR和Cu ZnSOD的mRNA水平一直与正常无显著差异 ,GSH PxmRNA水平略高于正常 .黄芩黄酮可能通过有效清除亚硒酸钠间接产生的活性氧来防止白内障的发生 ,并使亚硒酸钠对晶状体抗氧化酶表达的影响得以消除     

小檗胺对糖尿病性白内障的预防及SOD、CAT和GSH-Px酶活性变化研究

用抗氧化剂及自由基清除剂小檗胺（中药提纯单体化合物）对ＳＴＺ诱发的大鼠糖尿病性白内障进行腹腔注射的预防实验结果显示：１）在白内障出现早期小檗胺给药组晶状体空泡出现时间比未给药的糖尿病组推迟２周．２）小檗胺有对抗诱发动物模型晚期晶状体混浊出现的功能,以３．４８ｍｇ／ｋｇ体重剂量的效果最好．３）ＳＯＤ,ＣＡＴ,ＧＳＨ－Ｐｘ酶活性的动态变化,未给药的糖尿病组第２周即开始出现,两个剂量小檗胺给药组均比糖尿病组延迟２周出现．这与裂隙灯观察结果相吻合,但形态学变化晚于酶活性变化．这证明早期使用抗氧化剂及自由基清除剂小檗胺对动物实验性糖尿病性白内障的发生、发展有明显的预防作用．     

AC1及AC3对白内障形成中晶状体氧化还原物质及硒含量的变化

观察了AC1和AC3对抗亚硒酸钠性白内障形成过程中晶状体的脂类过氧化作用,非蛋白质疏基水平及硒含量。结果表明,亚硒酸钠组大鼠,在晶状体混浊出现前已发生脂类过氧化作用及硒含量的明显增加,非蛋白质巯基含量的显著降低,并持续至核混浊期;而同时接受AC1或AC3的大鼠,晶状体非蛋白质巯基水平初期降低,然后逐渐恢复至正常。AC1可有效的对抗亚硒酸钠所致的脂类过氧化作用增加,而AC3的对抗效应需一定剂量及时程,两者对晶状体硒含量均无明显影响。     

比较不同类型白内障形成过程中非蛋白质巯基与蛋白质巯基的动态变化及关系

本文报道了在亚硒酸钠、平阳霉素及半乳糖诱发大鼠产生白内障过程中晶状体中非蛋白质巯基及蛋白质巯基的动态变化,并探讨了其变化机理及相互关系。在亚硒酸钠诱发白内障过程中,给药24h后晶状体中非蛋白质巯基减少到正常的二分之一,以后又逐渐回升,但始终未达到正常水平,至第7天,非蛋白质巯基又再度减少。在平阳霉素及半乳糖诱发白内障过程中,晶状体中非蛋白质巯基分别在给药后的第7天及第3天开始大量减少,以后继续减少,至第15天时,其含量分别为正常的十分之一及五分之一。在体外,亚硒酸钠有促进还原型谷胱甘肽自氧化的作用,半乳糖对此作用无影响,而平阳霉素可阻止其进行,但能加强亚硒酸钠的促进作用。在三种白内障晶状体中,蛋白质巯基开始减少的时间均较非蛋白质巯基为晚,这表明只有非蛋白质巯基减少到一定程度后蛋白质巯基才会被大量氧化,同时也说明非蛋白质巯基具有保护蛋白质巯基免受氧化的作用。只有这种保护作用减弱后,才会使蛋白质巯基遭受氧化而导致白内障。     

亚硒酸钠对小麦幼苗中谷胱甘肽过氧化物酶和谷胱甘肽转硫酶活性以及谷胱甘肽含量的影响

用1.0 mg·L-1的亚硒酸钠根施小麦幼苗,测定亚硒酸钠对谷胱甘肽过氧化物酶和谷胱甘肽转硫酶活性以及还原性谷胱甘肽含量的结果表明,外源亚硒酸钠对麦苗地上部的谷胱甘肽过氧化物酶和谷胱甘肽转硫酶活性均有诱导作用,使麦苗体内的谷胱甘肽含量水平增加.     

A comparative study on effect of dietary selenium and vitamin E on some antioxidant enzyme activities of liver and brain tissues

Since selenium and vitamin E have been increasingly recognized as an essential element in biology and medicine, current research activities in the field of human medicine and nutrition are devoted to the possibilities of using these antioxidants for the prevention or treatment of many diseases. The present study was aimed at investigating and comparing the effects of dietary antioxidants on glutathione reductase and glutathione peroxidase activities as well as free and protein-bound sulfhydryl contents of rat liver and brain tissues. For 12–14 wk, both sex of weanling rats were fed a standardized selenium-deficient and vitamin E-deficient diet, a selenium-excess diet, or a control diet. It is observed that glutathione reductase and glutathione peroxidase activities of both tissues of the rats fed with a selenium-deficient or excess diet were significantly lower than the values of the control group. It is also shown that free and bound sulfhydryl concentrations of these tissues of both experimental groups were significantly lower than the control group. The percentage of glutathione reductase and glutathione peroxidase activities of the deficient group with respect to the control were 50% and 47% in liver and 66% and 61% in the brain, respectively; while these values in excess group were 51% and 69% in liver and 55% and 80% in brain, respectively. Free sulfhydryl contents of the tissues in both experimental groups showed a parallel decrease. Furthermore, the decrease in protein-bound sulfhydryl values of brain tissues were more pronounced than the values found for liver. It seems that not only liver but also the brain is an important target organ to the alteration in antioxidant system through either a deficiency of both selenium and vitamin E or an excess of selenium alone in the diet.     

Glutathione metabolism in primate lenses: A phylogenetic study of glutathione synthesis and glutathione redox cycle enzyme activities

Lens wet weights, soluble protein, and activities of γ-glutiamylcysteine synthetase, glutathione synthetase, glutathione peroxidase, and glutathione reductase were determined in primate lenses. The primary sources of lenses were middle-aged adult animals. The Primates, from 23 genera, were categorized into six superfamilies: hominoids (five species), Old World monkeys (seven species), New World monkeys (five species), tarsiers (two species), lemurs (six species), and lorisids (three species). Significant differences between various groups or combinations of groups were noted for γ-glutamylcysteine synthetase, glutathione peroxidase, and glutathione reductase activities. Lenticular γ-glutamylcysteine synthetase activity was very low in the Old World simian lenses and highest in the prosimians. Glutathione peroxidase activity was extraordinarily high in lenses of Old World monkeys. Glutathione reductase activity was low in all the prosimians but tenfold higher in hominoid lenses with intermediate values in monkeys of both the Old World and New World. Glutathione synthetase activity was variable, and no clear pattern which might be useful for primate classification was noted. Lenticular activity ratios of glutathione synthetase:γ-glutamylcysteine synthetase were highest in the Old World simians and lowest in the prosimians. These data with emphasis upon Aotus and the tarsiers were examined with regard to phylogenetic relationships. © 1994 Wiley-Liss, Inc.     

Effect of selenium on the system of superoxide anion radical formation and detoxication in hepatocarcinogenesis

It is established that the introduction of selenium in combination with diethylnitrosamine into rat organisms has a preventive influence on the tumour formation. The intensity of superoxide radicals formation by the liver cell microsomes in this case decreases, while the activity of superoxide dismutase, glutathione peroxidase I, glutathione reductase and concentration of selenium in microsomes increases. The anticarcinogenic action of selenium is considered as a result of an increase in the activity of superoxide dismutase, glutathione peroxidase I and glutathione reductase. This increase induces detoxication of superoxide radicals forming in considerable amounts in rat liver cells under the effect of carcinogen.     

Selenium supplementation sensitizes renca cells to tert-butylhydroperoxide induced loss of viability

Mouse renal carcinoma (renca) cells growing exponentially in foetal bovine serum (1%) supplemented with selenium (1 microM, sodium selenite) were exposed to oxidative insult. It was found that glutathione peroxidase activity increased (44%), while the activities of catalase, glutathione disulfide reductase, and level of total glutathione did not change due to selenium supplementation. Selenium supplementation made renca cells susceptible to tert-butylhydroperoxide induced cell death, while it did not affect the viability when the cells were exposed to hydrogen peroxide. It suggested that the contribution of glutathione peroxidase in antioxidant defense mechanism of renca cells was possibly not crucial and the function of catalase might be important especially against hydrogen peroxide.     

Biological effects of a nano red elemental selenium.

A novel selenium form, nano red elemental selenium (Nano-Se) was prepared by adding bovine serum albumin to the redox system of selenite and glutathione. Nano-Se has a 7-fold lower acute toxicity than sodium selenite in mice (LD(50) 113 and 15 mg Se/kg body weight respectively). In Se-deficient rat, both Nano-Se and selenite can increase tissue selenium and GPx activity. The biological activities of Nano-Se and selenite were compared in terms of cell proliferation, enzyme induction and protection against free racial-mediated damage in human hepatoma HepG2 cells. Nano-Se and selenite are similarly cell growth inhibited and stimulated synthesis of glutathione peroxidase (GPx), phospholipid hydroperoxide glutathione peroxidase (PHGPx) and thioredoxin reductase (TR). When HepG2 cells were co-treated with selenium and glutathione, Nano-Se showed less pro-oxidative effects than selenite, as measured by cell growth. These results demonstrate that Nano-Se has a similar bioavailability in the rat and antioxidant effects on cells.     

Prevention of Selenite-Induced Cataractogenesis by an Ethanolic Extract of Cineraria maritima: An Experimental Evaluation of the Traditional Eye Medication

In the present study, the antioxidant potential of an ethanolic extract of Cineraria maritima and its efficacy in preventing selenite-induced cataractogenesis were assessed in vitro and in vivo. In the in vitro phase of the study, lenses dissected out from the eyes of Wistar rats were incubated for 24 h at 37°C in Dulbecco’s modified Eagle medium (DMEM) alone (group I), in DMEM containing 100 μM of selenite only (group II), or in DMEM containing 100 μM of selenite and 300 μg/ml C. maritima extract added at the same time (group III). Gross morphological examination of the lenses revealed dense opacification in group II, minimal opacification in group III, and no opacification in group I lenses. The mean activities of the antioxidant enzymes catalase, glutathione peroxidase, and superoxide dismutase were significantly lower in group II than in group I or group III lenses, while malondialdehyde concentration was significantly higher in group II lenses than in group I and group III lenses. In the in vivo phase of the study, dense opacification of lenses was noted in all rat pups (100%) that had received a single subcutaneous injection of sodium selenite alone (19 μM/kg body weight) on postpartum day 10, whereas cataract formation occurred in only 33.3% of rat pups that had received selenite as well as an intraperitoneal injection of the extract of C. maritima (350 mg/kg body weight) for five consecutive days. These observations suggest that the ethanolic extract of C. maritima may prevent experimental selenite-induced cataractogenesis.     

Prevention of selenite induced oxidative stress and cataractogenesis by luteolin isolated from Vitex negundo

Free radical mediated oxidative stress plays a crucial role in the pathogenesis of cataract and the present study was to determine the efficacy of luteolin in preventing selenite induced oxidative stress and cataractogenesis in vitro. Luteolin is a bioactive flavonoid, isolated and characterized from the leaves of Vitex negundo. Lenses were extracted from Sprague-Dawley strain rats and were organ cultured in DMEM medium. They were divided into three groups with eight lenses in each group as follows: lenses cultured in normal medium (G I), supplemented with 0.1mM sodium selenite (G II) and sodium selenite and 2 μg/ml luteolin (G III). Treatment was from the second to fifth day, while selenite administration was done on the third day. After the experimental period, lenses were taken out and various parameters were studied. The antioxidant potential of luteolin was assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. In the selenite induced group, morphological examination of the lenses showed dense cortical opacification and vacuolization. Biochemical examinations revealed a significant decrease in activities of antioxidant enzymes and enzymes of the glutathione system. Additionally decreased glutathione level and increased reactive oxygen species (ROS) and thiobarbituric acid reactive substances (TBARS) were observed. Luteolin treatment abated selenite induced oxidative stress and cataractogenesis by maintaining antioxidant status, reducing ROS generation and lipid peroxidation in the lens. These finding demonstrated the anticataractogenic effect of luteolin by virtue of its antioxidant property, which has been reported in this paper for the first time.     

Developmental study of rat brain glutathione peroxidase and glutathione reductase

Glutathione peroxidase and glutathione reductase activities were measured in whole rat brains at selected ages from birth to adulthood. On a wet weight basis glutathione peroxidase activity increased 70% during development and glutathione reductase activity increased 160%. On a protein basis glutathione peroxidase declined slightly in activity during the first two weeks of life and then maintained the 14-day activity into adulthood while glutathione reductase showed a 30% increase in activity. While less than the developmental changes in many enzymes involved in aerobic glycolysis or catecholamine metabolism, these increases do suggest a role in CNS metabolism.     

Effects of dietary selenium on glutathione peroxidase and thioredoxin reductase activity and recovery from cardiac ischemia-reperfusion.

Glutathione peroxidase and thioredoxin reductase are selenocysteine-dependent enzymes that protect against oxidative injury. This study examined the effects of dietary selenium on the activity of these two enzymes in rats, and investigated the ability of selenium to modulate myocardial function post ischemia-reperfusion. Male wistar rats were fed diets containing 0, 50, 240 and 1000 microg/kg sodium selenite for 5 weeks. Langendorff perfused hearts isolated from these rats were subjected to 22.5 min global ischemia and 45 min reperfusion, with functional recovery assessed. Liver samples were collected at the time of sacrifice, and heart and liver tissues assayed for thioredoxin reductase and glutathione peroxidase activity. Selenium deficiency reduced the activity of both glutathione peroxidase and thioredoxin reductase systemically. Hearts from selenium deficient animals were more susceptible to ischemia-reperfusion injury when compared to normal controls (38% recovery of rate pressure product (RPP) vs. 47% recovery of RPP). Selenium supplementation increased the endogenous activity of thioredoxin reductase and glutathione peroxidase and resulted in improved recovery of cardiac function post ischemia reperfusion (57% recovery of RPP). Endogenous activity of glutathione peroxidase and thioredoxin reductase is dependent on an adequate supply of the micronutrient selenium. Reduced activity of these antioxidant enzymes is associated with significant reductions in myocardial function post ischemia-reperfusion.     

Peroxide-metabolizing systems of the crystalline lens

The ability of transparent and cataractous human, rabbit and mice lenses to metabolize hydrogen peroxide in the surrounding medium was evaluated. Using a chemiluminescence method in a system of luminol-horseradish peroxidase and a photometric technique, the temperature-dependent kinetics of H₂O₂ decomposition by lenses were measured. The ability of opaque human lenses to catalyze the decomposition of 10^?4 M H₂O₂ was significantly decreased. However, this was reserved by the addition of GSH to the incubation medium. Incubation of the mice lenses with the initial concentration H₂O₂ 10^?4 M led to partial depletion of GSH in normal and cataractous lenses. Human cataractous lenses showed decreased activities of glutathione reductase, glutathione peroxidase (catalyzing reduction of organic hydroperoxides including hydroperoxides of lipids), superoxide dismutase, but no signs of depletion in activities of catalase or glutathione peroxidase (utilizing H₂O₂). The findings indicated an impairment in peroxide metabolism of the mature cataractous lenses compared to normal lenses to be resulted from a deficiency of GSH. An oxidative stress induced by accumulation of lipid peroxidation products in the lens membranes during cataract progression could be considered as a primary cause of GSH deficiency and disturbance of the redox balance in the lens.