Characterisation of P-glycoprotein-9.1 in Haemonchus contortus.
Characterisation of P-glycoprotein-9.1 in Haemonchus contortus.
复制标题
DOI：
10.1186/s13071-016-1317-8
复制
发表时间：
2016-01-28
影响因子：
3.2
通讯作者：
Prichard RK
中科院分区：
文献类型：
--
作者：
Godoy P;Che H;Beech RN;Prichard RK
存在的兽医重要线虫，如捻转血矛线虫抗驱虫药，包括大环内酯，已成为动物健康的主要问题。H.大环内酯抗性。捻转虫似乎是多基因的，包括这些药物的主动外排的P-糖蛋白，ABC转运蛋白家族的成员，存在于这种寄生虫。本工作的目的是确定H.捻转虫P-糖蛋白9.1（Hco-PGP-9.1）及其与阿维菌素、伊维菌素、阿维菌素以及米尔倍霉素、莫昔克丁的相互作用。此外，在蠕虫中寻找Hco-PGP-9.1的定位。克隆Hco-Pgp-9.1并在哺乳动物细胞中表达，并分别通过qRT-PCR和Western-blot在转录和蛋白质水平上确定其表达谱。使用示踪染料罗丹明123评估线虫运输活性。不同大环内酯和罗丹明123之间的配体竞争试验用于确定Hco-PGP-9.1与阿维菌素（阿维菌素和伊维菌素）或莫昔克丁之间是否存在相互作用。此外，进行免疫染色以定位Hco-PGP-9.1在转基因细胞和成年雌性寄生虫中的表达。Hco-PGP-9.1在转染的宿主细胞的细胞膜中表达，并且能够挤出罗丹明123。伊维菌素和阿维菌素，但不是莫昔克丁，Hco-PGP-9.1运输罗丹明123的能力有显着的抑制作用。针对Hco-PGP-9.1表位产生的抗体定位于成年女性H.扭曲这些结果表明阿维菌素与Hco-PGP-9.1的强相互作用。然而，可能由于其理化性质，莫昔克丁对Hco-PGP-9.1的影响显著较小。由于阿维菌素与该转运蛋白的相互作用大于莫昔克丁，因此在H.扭曲Hco-PGP-9.1在抗性蠕虫的雌性生殖系统中可能的过度表达可以减少大环内酯引起的子宫麻痹，从而允许药物激发的抗性蠕虫中继续释放卵。本文的在线版本（doi：10.1186/s13071-016-1317-8）包含补充材料，可供授权用户使用。
The existence nematodes of veterinary importance such as Haemonchus contortus resistant to anthelmintic drugs, including the macrocyclic lactones, has become a major concern in animal health. Macrocyclic lactone resistance in H. contortus seems to be multigenic including the active efflux of these drugs by P-glycoproteins, members of the ABC transporter family, present in this parasite. The goals of the present work were to determine the activity of H. contortus P-glycoprotein 9.1 (Hco-PGP-9.1) and its interaction with the avermectins, ivermectin, abamectin, and also the milbemycin, moxidectin. Additionally, the localisation of Hco-PGP-9.1 was sought in adult worms. Hco-Pgp-9.1 was cloned and expressed in mammalian cells and its expression profile was determined at the transcriptional and protein level by qRT-PCR and Western-blot, respectively. The nematode transport activity was assessed using the tracer dye Rhodamine 123. A ligand competition assay between different macrocyclic lactones and Rhodamine 123 was used to establish whether or not there was interaction between Hco-PGP-9.1 and the avermectins (abamectin and ivermectin) or moxidectin. In addition, immunostaining was carried out to localise Hco-PGP-9.1 expression in the transgenic cells and in adult female parasites. Hco-PGP-9.1 was expressed in the cell membrane of the transfected host cells and was able to extrude Rhodamine 123. Ivermectin and abamectin, but not moxidectin, had a pronounced inhibitory effect on the ability of Hco-PGP-9.1 to transport Rhodamine 123. Antibodies raised against Hco-PGP-9.1 epitopes localised to the uterus of adult female H. contortus. These results suggest a strong interaction of the avermectins with Hco-PGP-9.1. However, possibly due to its physico-chemical properties, moxidectin had markedly less effect on Hco-PGP-9.1. Because of the greater interaction of the avermectins than moxidectin with this transporter, it is more likely to contribute to avermectin resistance than to moxidectin resistance in H. contortus. Possible over expression of Hco-PGP-9.1 in the female reproductive system in resistant worms could reduce paralysis of the uterus by macrocyclic lactones, allowing continued egg release in drug challenged resistant worms. The online version of this article (doi:10.1186/s13071-016-1317-8) contains supplementary material, which is available to authorized users.