动物硒蛋白-磷脂氢谷胱甘肽过氧化物酶

磷脂的过氧化过程与多种疾病有关,包括炎症、动脉粥样硬化、神经退行性变化、衰老等。磷脂氢谷胱甘肽过氧化物酶是目前发现唯一可以直接减少膜系统及脂蛋白上的磷脂氢过氧化物,联合维生素E可抑制磷脂过氧化。磷脂氢谷胱甘肽过氧化物酶是谷胱甘肽过氧化物酶家族中的第2个被鉴定的含硒酶,是相对分子质量为19000(M r)的单体酶,其基因位于人类19号染色体上。磷脂氢谷胱甘肽过氧化物酶曾被认为是一种保护质膜的普通的抗氧化酶,现在认为具有多种功能,与人类多种疾病的发生发展有关。     

p,p'‐DDE damages spermatogenesis via phospholipid hydroperoxide glutathione peroxidase depletion and mitochondria apoptosis pathway

One, 1‐dichloro‐2,2‐bis(p‐chlorophenyl) ethylene (p,p'‐DDE), the major metabolite of 2,2‐bis(4‐chlorophenyl)?1,1,1‐trichloroethane (DDT), is a known persistent organic pollutant (POPs) and male reproductive toxicant. However, the mechanism by which p,p'‐DDE exposure causes male reproductive toxicity remains unknown. The objective of this study was to elucidate some mechanisms involved in this process, including the mitochondria apoptosis pathway and the role of phospholipid hydroperoxide glutathione peroxidase (PHGPx). Puberty male SD rats were given different doses of p,p'‐DDE (0, 20, 60, 100 mg/kg body weight), after the treatment, the semen quality was evaluated. Western blotting was used to detect the PHGPx protein expression. Furthermore, real‐time PCR was used to analyze the genetic expression of PHGPx, Bax, Cytochrom C (Cyt C), Apaf‐1, and caspase‐3 in the testis. Results indicated that after the exposure, sperm malformation rate showed a significant rise compared with the control group, and meanwhile, the sperm density and sperm motility parameters were reduced to some extent in different treated groups. The mitochondria apoptosis pathway was activated. And remarkably, the expression of PHGPx protein was greatly reduced by the exposure. We conclude that p,p'‐DDE can damage spermatogenesis via PHGPx depletion and mitochondria apoptosis pathway. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 593–600, 2016.     

Organic selenium supplementation increases PHGPx but does not improve viability in chilled boar semen

This study evaluated the effects of dietary organic selenium (Se) on viability of chilled boar semen. Twelve boars were divided into three groups: control (CON), 0.3 mg kg^?1 sodium selenite; inorganic (INO), 0.5 mg kg^?1 sodium selenite and organic (ORG), 0.5 mg kg^?1 Se yeast. The experiment was conducted within 10 weeks, and analysis was performed fortnightly, in storage semen by 72 h. No effect was observed on motility; however, straightness and linearity percentages were higher (P < 0.05) in the animals receiving CON diet compared with INO group. Percentages of cells with both plasma and acrosomal intact membranes, lipidic membrane peroxidation and mitochondrial membrane potential were similar on all treatments. Animals receiving CON diet presented higher (P < 0.05) values of ATP when compared with INO group. The PHGPx was higher (P < 0.05) in animals that received ORG in comparison with INO group. In conclusion, organic selenium supplementation increases PHGPx but does not improve chilled semen viability in 72 h.     

PHGPx底物合成及其活力测定方法的研究──Ⅰ．PHGPx活力测定方法的建立

建立了ＰＨＧＰｘ活力测定方法，完成了酶量曲线。底物ＰＣＯＯＨ和ＧＳＨ曲线，并用建立的方法测定了不同硒营养水平大鼠睾丸中ＰＨＧＰｘ活力。     

Failure of phospholipid hydroperoxide glutathione peroxidase expression in oligoasthenozoospermia and mutations in the PHGPx gene

Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a selenoprotein belonging to the family of glutathione peroxidases. PHGPx has long been considered a major antioxidant that, in cooperation with vitamin E, protects biomembranes. To determine the expression pattern of PHGPx mRNA in human, quantitative RT-polymerase chain reaction (PCR) analyses using RNA from different embryonal and adult tissues were performed. A predominant expression was found in testes. In spermatozoa, PHGPx was found to be localized in the mid-piece of spermatozoa. We studied the relationship between spermatozoa PHGPx expression, mutations in PHGPx gene and human oligoasthenozoospermia, a defect in which both the number and the motility of spermatozoa are significantly below normal. Spermatozoa specimens from 45 infertile males were analysed for fertility-related parameters according to World Health Organisation and were classified as suffering from oligoasthenozoospermia. Two patients (4.44%) showed no expression of PHGPx and in nine patients (20.00%), a reduced expression of the enzyme was observed. DNA sequences of various regions of the PHGPx gene (coding, 5'flanking region and intron 1) from these patients and 58 fertile volunteers were analysed for mutations by PCR amplification and direct sequencing. Sequence data revealed no cause/effect relationship for any of the variants. From these data it can be concluded that oligoasthenozoospermia is associated with a decrease in the level of expression of PHGPx in the spermatozoa of some infertile men (24.44%), but is not linked to mutations in PHGPx gene.     

磷脂过氧化氢物谷胱甘肽过氧化物酶与精子成熟

磷脂过氧化氢物谷胱甘肽过氧化物酶 (PHGPx)基因位于人类 19号染色体上 ,是第二个被鉴定的细胞内硒依赖的谷胱甘肽过氧化物酶 ,是一种相对分子质量 (Mr)为 190 0 0的单体酶 ,它通过催化脂质过氧化物的减少来保护细胞不受脂质过氧化物介导的损害。PHGPx曾被认为是一种保护质膜的普通的抗氧化酶 ,现在认为其具有多种功能 ,如抑制脂肪氧化酶和在人类精子成熟过程中变成一种线粒体内囊的无酶活性的结构蛋白成分 ,即PHGPx在精子细胞内以可溶性过氧化物酶的形式存在 ,而在成熟精子以无活性、被氧化交叉结合、不可溶性的结构蛋白的形式持续存在     

Phospholipid hydroperoxide glutathione peroxidase is involved in the maintenance of male fertility under cryptorchidism in mice

Severe oxidative stress by cryptorchidism leads to infertility. To assess the functional significance of phospholipid hydroperoxidase glutathione peroxidase (PHGPx) under cryptorchidism, PHGPx expression was spatiotemporally analyzed in testes and epididymis excised at 1, 4, 7, 14, 21, and 28 days after experimental bilateral cryptorchidism in adult mice. In testes, while apoptosis-related caspase 3 and Bcl-xL mRNAs were significantly changed after 14 days, 3 beta-hydroxysteroid dehydrogenase mRNA was greatly reduced immediately after cryptorchidism. Under cryptorchidism, PHGPx was significantly decreased in both organs after 21 days, while its mRNA was greatly reduced in testes after 14 days and in epididymis after 4 days. However, PHGPx was upregulated in degenerative spermatids, multinucleated giant cells, and Leydig cells in testes and desquamous spermatids in epididymis until 21 days, but was weakly detected in the spermatids at 28 days. These findings suggest that PHGPx is necessary for maintenance of male fertility under cryptorchidism in testes.     

1周速度训练对大鼠骨骼肌PHGPx基因表达的影响

目的 :采用RT -PCR技术测定和分析不同类型骨骼肌中谷胱甘肽磷脂氢过氧化物酶(PHGPx)基因表达水平以及速度训练前后PHGPx基因的表达 ,以探讨PHGPx在骨骼肌抗氧化体系中的作用。方法 :雄性SD大鼠 2 0只 ,随机分为训练组 (T ,n =10 )和对照组 (C ,n =10 )。在动物跑台上进行为期 1周的速度训练 ,采用RT -PCR方法测定比目鱼肌、腓肠肌红肌和白肌PHGPxmRNA含量。结果 :对照组腓肠肌红肌和白肌PHGPx基因表达无显著差异 ,但均显著高于比目鱼肌 (P <0 0 5 ) ;1周速度训练后腓肠肌白肌PHGPx基因表达增加 (P <0 0 5 ) ,红肌PHGPx基因表达下降 (P<0 0 5 )。结论 :PHGPx基因表达水平存在肌纤维类型特异性 ,IIa和IIb型肌纤维中的基因表达高于I型肌纤维 ;腓肠肌PHGPx基因表达水平受运动训练的调节 ,速度训练使PHGPx基因在Ⅱb型肌纤维中表达升高 ,而在Ⅱa型肌纤维中表达降低。     

Polyunsaturated fatty acids of germ cell membranes,glutathione and blutathione-dependent enzyme-PHGPx: from basic to clinic

The lipid metabolism in sperm cells is important both for energy production and for cell structure. A special composition of membrane phospholipids, rich in polyunsaturated fatty acids (PUFA), and the different composition of sperm and immature germ cell membrane are described and discussed. Testis germ cells as well as epididymal maturing spermatozoa are endowed with enzymatic and non-enzymatic scavenger systems to prevent lipoperoxidative damage. Catalase, superoxide dismutase, and glutathione-dependent oxidoreductases are present in variable amounts in the different developmental stages. Phospholipid hydroperoxide glutathione peroxidase (PHGPx) activity and roles in caput and cauda epididymal sperm cells are discussed. Also seminal plasma has a highly specialized scavenger system that defends the sperm membrane against lipoperoxidation and the degree of PUFA insaturation acts to achieve the same goal. Systemic predisposition and a number of pathologies can lead to an anti-oxidant/pro-oxidant disequilibrium. Scavengers, such as glutathione can be used to treat these cases as they can restore the physiological constitution of PUFA in the cell membrane.     

Enzymatic and immunochemical evaluation of phospholipid hydroperoxide glutathione peroxidase (PHGPx) in testes and epididymal spermatozoa of rats of different ages

Selenium (Se) and selenoproteins such as glutathione peroxidases are necessary for the proper development and fertilizing capacity of sperm cells. Phospholipid hydroperoxide glutathione peroxidase (PHGPx, E.C. 1.11.1.12) is a monomeric seleno-enzyme present in different mammalian tissues in soluble and bound form. Its function, like the other glutathione peroxidases, was originally viewed as a protective role against hydroperoxides, but direct and indirect evidence indicates that it has additional regulatory roles. PHGPx is present in testis cells and sperm cells, and its appearance is hormone regulated. We present here biochemical data, which clearly indicate that the enzyme specific activity in rat is age-dependent during the life-span monitored (from 36 to 365 days), with a maximum at 3 months of age in the testis germ cells and at 6 months of age in the isolated epididymal sperm cells. Western blotting and immunocytochemical analysis by means of anti-PHGPx antibodies show the different distribution and the strong binding of PHGPx in the testes and sperm cell subcellular compartments (nucleus, acrosome, mitochondria and residual bodies) of rats of different age. The presence of the protein exhibits in the testis cells a pattern different from that of the catalytic activity, with a maximum at 6 months of age. The subcellular distribution of PHGPx is qualitatively, but not quantitatively, unchanged during ageing. These different behaviours are compared and discussed.