Chen Ji-de, Zhou Tao-li, Xu Wen-yue, Ding Yan, Huang Fu-sheng
Department of Pathogenic Biology, The Third Military Medical University, Chongqing 400038, China.
Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi. 2009 Feb 28;27(1):1-5.
To study the role of cytidine-phosphate-guanosine oligodeoxynucleotide (CpG ODN) on the development of Plasmodium liver stage.
Plasmodium yoelii BY265 18S rRNA was cloned, and the TaqMan real-time PCR was established on P. yoelii BY265 18S rRNA and mouse GAPDH as quantitative analysis model. The model was tested by the level of liver Plasmodium load with the liver cDNA in BALB/c mice infected by salivary gland sporozoites for 42 hours. Twelve BALB/c mice were randomly divided into CpG group, CpG control group and PBS control group which were injected respectively by ODN1826 30 microg, ODN1826 control 30 microg and 0.01 mol/L PBS 200 microl via vena caudalis. Twenty-four hours later, each mouse was inoculated with 100 sporozoites. Mice were sacrificed in 42 hours after infection, and the liver load of Plasmodium was analyzed by TaqMan real-time PCR.
The cloned Py BY265 18S rRNA gene showed 98% similarity to Py 17XNL. The quantitative analysis model consisted by 18S rRNA and GAPDH showed positive correlation between the level of liver Plasmodium load and the sporozoite inoculation dose to mice. The Plasmodium load in CpG ODN pre-treated mice was reduced to one fifth of the control group (0.28/1.33) (P<0.05).
The quantitative analysis model of TaqMan RT-PCR can detect the liver load of Plasmodium parasites, and CpG ODN can inhibit the development of its pre-erythrocytic stage.
研究胞苷-磷酸-鸟苷寡脱氧核苷酸(CpG ODN)在疟原虫肝期发育中的作用。
克隆约氏疟原虫BY265 18S rRNA,建立以约氏疟原虫BY265 18S rRNA和小鼠甘油醛-3-磷酸脱氢酶(GAPDH)为定量分析模型的TaqMan实时荧光定量PCR。用感染唾液腺子孢子42小时的BALB/c小鼠肝脏cDNA,以肝脏疟原虫载量水平对该模型进行验证。12只BALB/c小鼠随机分为CpG组、CpG对照组和PBS对照组,分别经尾静脉注射ODN1826 30 μg、ODN1826对照30 μg和0.01 mol/L PBS 200 μl。24小时后,每只小鼠接种100个子孢子。感染后42小时处死小鼠,用TaqMan实时荧光定量PCR分析肝脏疟原虫载量。
克隆的Py BY265 18S rRNA基因与Py 17XNL的相似性为98%。由18S rRNA和GAPDH组成的定量分析模型显示,肝脏疟原虫载量水平与小鼠子孢子接种剂量呈正相关。CpG ODN预处理小鼠的疟原虫载量降至对照组的五分之一(0.28/1.33)(P<0.05)。
TaqMan RT-PCR定量分析模型可检测疟原虫肝脏载量,CpG ODN可抑制其红细胞前期发育。