从埃及纳特龙湖土壤中分离出的两种已鉴定菌株，即茄病镰刀菌和构巢曲霉，对Caco-2（ATCC）细胞系的分子分析及抗癌特性
Molecular analysis and anticancer properties of two identified isolates, Fusarium solani and Emericella nidulans isolated from Wady El-Natron soil in Egypt against Caco-2 (ATCC) cell line.
作者信息
Mohamed Hala F
机构信息
University of Al-Azhar, Faculty of Science (Girls Branch), Department of Botany and Microbiology,Youssef Abbas Street, Nasr City, Cairo, Egypt.
出版信息
Asian Pac J Trop Biomed. 2012 Nov;2(11):863-9. doi: 10.1016/S2221-1691(12)60244-5.
OBJECTIVE
To characterize, identify and investigate the anticancer properties of two new soil fungal isolates, Emericella nidulans and Fusarium solani isolated from Wady El-Natron in Egypt against colon cancer Caco-2 (ATCC) cell line.
METHODS
Soil sample was cultured and two strains were chosen for morphological and phenotypical characterization. Partial sequences of the 18s rRNA gene and the internal transcribed spacer region ITS of the two isolates were amplified by PCR. Phylogenetic tree construction and analysis of the resulted multiple sequences from the two fugal isolates were also carried out. In vitro anticancer activity of the two strains was done against colon Caco-2 cancer cell line. Reverse transcription - PCR was carried out to detect level of expression of p53 in Caco-2 cell line.
RESULTS
HF.1 displayed morphological and genotypic characteristics most similar to that of Fusarium solani while HF.2 was most similar to Emericella nidulans with high similarity of 99% and 97% respectively. The multiple sequence alignment of the two fungal isolates showed that, the maximum identical conserved domains in the 18s rRNA genes were identified with the nucleotide regions of 51st to 399th base pairs, 88th to 525th base pairs respectively. While those in the ITS genes were identified with the nucleotide regions of 88th to 463rd and 51st to 274th. The two isolates showed IC50 value with (6.24±5.21) and (9.84±0.36) µg/mL) concentrations respectively at 28h. Reverse transcription - PCR indicated that these cells showed high level of expression for p53 mRNA.
CONCLUSIONS
The morphology and molecular analysis identified HF.1 and HF.2 to be Fusarium solani and Emericella nidulans; new isolates of anticancer producing fungi from Wady El-Natroon city in Egypt. Treatment with the two isolates caused P53 expression in Caco-2 cell line. These two isolates can be used as an anticancer agents.
目的
对从埃及纳特龙湖分离出的两种新的土壤真菌分离株——构巢曲霉和茄病镰刀菌进行特性鉴定，并研究它们对结肠癌Caco-2（ATCC）细胞系的抗癌特性。
方法
对土壤样本进行培养，选择两个菌株进行形态学和表型特征鉴定。通过聚合酶链反应（PCR）扩增这两个分离株的18s rRNA基因和内部转录间隔区（ITS）的部分序列。还进行了系统发育树构建以及对这两种真菌分离株所得多条序列的分析。对这两个菌株进行了针对结肠癌Caco-2癌细胞系的体外抗癌活性研究。进行逆转录-聚合酶链反应（RT-PCR）以检测Caco-2细胞系中p53的表达水平。
结果
HF.1表现出与茄病镰刀菌最相似的形态学和基因型特征，而HF.2与构巢曲霉最相似，相似度分别高达99%和97%。这两种真菌分离株的多序列比对显示，18s rRNA基因中最大的相同保守结构域分别分别别位于第51至399个碱基对、第88至525个碱基对的核苷酸区域。而ITS基因中的保守结构域分别位于第88至463个以及第51至274个核苷酸区域。这两个分离株在28小时时的半数抑制浓度（IC50）值分别为（6.24±5.21）和（9.84±0.36）μg/mL。逆转录-聚合酶链反应表明这些细胞显示出p53信使核糖核酸（mRNA）的高表达水平。
结论
形态学和分子分析确定HF.1和HF.2分别为茄病镰刀菌和构巢曲霉，是从埃及纳特龙湖市分离出的新的抗癌真菌分离株。用这两个分离株处理导致Caco-2细胞系中P53表达。这两个分离株可作为抗癌剂使用。
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