Salimi Mahboobeh, Shojaee Saeedeh, Keshavarz Hossein, Mohebali Mehdi
Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran; Center for Research of Endemic Parasites of Iran (CREPI), Tehran University of Medical Sciences, Tehran, Iran.
Iran J Parasitol. 2016 Jan-Mar;11(1):81-5.
This study was performed to induce conversion of RH strain tachyzoites of Toxoplasma gondii to bradyzoites by pH changing of the culture medium.
HeLa cell monolayers were infected at a 1:1 tachyzoite to cell ratio. Four hours after infection, the culture medium was removed and replaced with culture medium and 5% FCS, adjusted to pH 8 with NaOH. The culture was maintained at 37 °C without CO2 until the end of the experiment. Cyst-like structures were collected and stained with periodic acid schiff (PAS) staining method. The soluble antigens of cyst-like structures of RH strain, in addition to RH tachyzoite, bradyzoites of avirulent Tehran strain and uninfected HeLa cells were electrophoresed on 12.5% polyacrylamide gel. The gel was stained by coomassie brilliant blue R-250.
Four days after infection of HeLa cells with tachyzoites of T. gondii, RH strain, cyst- like structures were noticed and stained with PAS. In the SDS-PAGE, protein bands of these structures had some differences with tachyzoites of RH strain, but there was quite similarity between protein bands of these structures and tissue cysts (bradyzoites) of Tehran strains. P34 and P36 (bradyzoite-specific proteins) were observed only in T. gondii bradyzoites of RH (cyst like structures) and bradyzoites of Tehran strains.
Alkalization of culture medium to pH 8 induced expression of bradyzoite- specific proteins and production of RH cysts in cell culture.
本研究旨在通过改变培养基的pH值诱导刚地弓形虫RH株速殖子转化为缓殖子。
以1:1的速殖子与细胞比例感染HeLa细胞单层。感染4小时后，去除培养基，用含5%胎牛血清的培养基替换，并用氢氧化钠将pH值调至8。将培养物在37℃无二氧化碳条件下维持至实验结束。收集囊样结构并用过碘酸希夫（PAS）染色法染色。将RH株囊样结构的可溶性抗原，以及RH速殖子、无毒力德黑兰株缓殖子和未感染的HeLa细胞在12.5%聚丙烯酰胺凝胶上进行电泳。凝胶用考马斯亮蓝R-250染色。
用刚地弓形虫RH株速殖子感染HeLa细胞4天后，观察到囊样结构并用PAS染色。在SDS-PAGE中，这些结构的蛋白条带与RH株速殖子有一些差异，但这些结构的蛋白条带与德黑兰株的组织囊肿（缓殖子）相当相似。仅在RH株的刚地弓形虫缓殖子（囊样结构）和德黑兰株缓殖子中观察到P34和P36（缓殖子特异性蛋白）。
将培养基碱化至pH 8可诱导缓殖子特异性蛋白的表达并在细胞培养中产生RH囊肿。
