Rao U R, Salinas G, Mehta K, Klei T R
Department of Microbiology and Parasitology, School of Veterinary Medicine, Louisiana State University, Baton Rouge, USA.
Parasitol Res. 2000 Nov;86(11):908-15. doi: 10.1007/s004360000255.
Brugia filarial nematodes are pathogenic lymphatic-dwelling parasites that, like other helminths, may modify the host's defense mechanisms by a major detoxification process involving glutathione-binding proteins such as glutathione S-transferases (GSTs). In the present study, soluble extracts of third-stage larvae, adult male and female worms, microfilariae of either B. pahangi or B. malayi or the adult worm excretory-secretory products of B. malayi were used to determine GST activity. These extracts and affinity-purified fractions of B. pahangi adult worms had a specific enzymatic activity when 1-chloro-2,4-dinitrobenzene was used as a substrate. The observance of this enzyme in all life cycle stages of Brugia spp. demonstrates its ubiquitous nature. Lavage of intraperitoneally infected jirds, but not that of uninfected jirds, also showed increased enzymatic activity, suggesting that GST is secreted in vivo. Soluble proteins of both Brugia spp. were strongly recognized by antibodies in sera from rabbits immunized with affinity-purified native GST of Onchocerca volvulus. Immunohistochemical studies localized these proteins in adult worms, demonstrating cross-reactivity between the GST of these two filarial nematodes. The effect of this enzyme on the motility and viability of adult worms, microfilariae, and larvae was tested in vitro using a battery of known GST inhibitors. Of all those tested, ethacrynic acid, N-ethylmalemide, 4-nitropyridine-oxide, or 1-chloro-2,4-dinitrobenzene at micromolar concentrations reduced the viability and motility of microfilariae, third-stage larvae, and adult worms. These results suggest that Brugia GSTs are major metabolic enzymes and may play an important role in the parasite's survival.
布鲁氏丝虫线虫是寄生于淋巴系统的致病性寄生虫，与其他蠕虫一样，它们可能通过涉及谷胱甘肽结合蛋白（如谷胱甘肽S-转移酶，GSTs）的主要解毒过程来改变宿主的防御机制。在本研究中，使用了第三期幼虫、成年雄虫和雌虫、帕氏布鲁线虫或马来布鲁线虫的微丝蚴的可溶性提取物，或马来布鲁线虫成年虫体的排泄-分泌产物来测定GST活性。当以1-氯-2,4-二硝基苯为底物时，这些提取物和帕氏布鲁线虫成年虫体的亲和纯化级分具有特定的酶活性。在布鲁氏丝虫属的所有生命周期阶段都观察到这种酶，这表明了它的普遍存在性。对经腹腔感染的沙鼠进行灌洗，但未感染的沙鼠则未出现这种情况，灌洗结果也显示酶活性增加，这表明GST在体内会分泌。两种布鲁氏丝虫属的可溶性蛋白都能被用盘尾丝虫亲和纯化天然GST免疫的兔血清中的抗体强烈识别。免疫组织化学研究将这些蛋白定位在成年虫体中，证明了这两种丝虫线虫的GST之间存在交叉反应性。使用一系列已知的GST抑制剂在体外测试了这种酶对成年虫体、微丝蚴和幼虫的运动性和活力的影响。在所有测试的抑制剂中，微摩尔浓度的依他尼酸、N-乙基马来酰胺、4-硝基吡啶氧化物或1-氯-2,4-二硝基苯降低了微丝蚴、第三期幼虫和成年虫体的活力和运动性。这些结果表明，布鲁氏丝虫GSTs是主要的代谢酶，可能在寄生虫的生存中发挥重要作用。
