Transcriptome profile in INT cells following Cryptosporidium parvum infection. Homo sapiens

Cryptosporidium parvum is an important opportunistic parasite pathogen for immunocompromised individuals and a common cause of diarrhea in young children in developing countries. Certain parasite molecules can be delivered into host epithelial cells and may act as effector molecules for parasite intracellular development. This study aims to measure the impact of transfection of a parasite low-protein coding potential RNA transcript, Cdg7_FLc_0990, on the transcriptome profile in intestinal epithelial cells. Human intestinal epithelial INT (FHs 74 Int) cells were grown to 80% confluence and transfected with the Cdg7_FLc_0990 full-length or the empty vector for 48h. Total RNA was collected for the genome-wide analysis. RNA from INT cells following C. parvum infection for 48h and from cell of the non-infected control was also collected for the analysis. The Agilent SurePrint G3 Human Gene Expression Microarray (G4851B) was used for the genome-wide analysis, which provides full coverage of genes and transcripts with the most up-to-date content, including mRNAs and lincRNAs (http://www.chem.agilent.com/store/en_US/Prod-G4851B/G4851B) Overall design: The FHs 74 Int were grown to 80% confluence for four groups: control (Group 00-C), C. parvum infection (Group 00-D, cells treated with C. parvum), pcDNA3.1(+) empty vector (Group CP-A, cells transfected with pcDNA3.1(+) empty vector), and pcDNA3.1(+)_Cdg7_FLc_0990 full-length (Group CP-D, cells transfected with pcDNA3.1(+)_Cdg7_FLc_0990 full-length ). 48h later, total RNAs were prepared with the RNeasy Mini kit (Qiagen) according to the manufacturer’s instruction.

微小隐孢子虫（Cryptosporidium parvum）是一类重要的机会致病性寄生虫病原体，可感染免疫功能低下人群，同时也是发展中国家幼儿腹泻的常见病因。部分寄生虫分子可被递送至宿主上皮细胞内，并可作为效应分子参与寄生虫的胞内发育过程。本研究旨在探究寄生虫低蛋白编码潜能RNA转录本Cdg7_FLc_0990的转染对肠上皮细胞转录组图谱的影响。将人肠上皮INT（FHs 74 Int）细胞培养至汇合度达80%，分别用Cdg7_FLc_0990全长转录本或空载体进行转染，培养48小时后收集总RNA用于全基因组分析；同时收集经C. parvum感染48小时的INT细胞，以及未感染的对照组细胞的RNA，同样用于后续分析。本研究采用安捷伦SurePrint G3人类基因表达微阵列（G4851B）开展全基因组分析，该芯片可通过最新的探针内容实现对基因及转录本的全面覆盖，涵盖信使RNA（mRNA）及基因间长链非编码RNA（lincRNA），详情可参见：http://www.chem.agilent.com/store/en_US/Prod-G4851B/G4851B。实验设计如下：将FHs 74 Int细胞培养至汇合度80%，分为四组：对照组（Group 00-C）、小隐孢子虫感染组（Group 00-D，经C. parvum感染处理）、空载体转染组（Group CP-A，转染pcDNA3.1(+)空载体）、Cdg7_FLc_0990全长转录本转染组（Group CP-D，转染pcDNA3.1(+)_Cdg7_FLc_0990全长转录本）。转染48小时后，按照制造商说明书操作，使用RNeasy Mini试剂盒（Qiagen）提取总RNA。