巨噬细胞对新生隐球菌主要毒性基因表达的影响

目的研究巨噬细胞对新生隐球菌B3501标准株的主要毒性基因表达影响。方法将对数生长期的J774．16巨噬细胞分别与新生隐球菌野生株B3501共孵育4h，收集被J774．16吞噬的B3501作为实验组，提取实验组和在37℃条件下5％二氧化碳单独培养的对照组B3501的RNA，采用实时荧光定量PCR技术，检测J774．16细胞内和对照组B3501的CNLAC1、CAP60、URE1、NMT表达的差异。结果实验组中新生隐球菌的CAP6，CNLAC1，NMT及URE1基因的mRNA在每百万看家基因（GAPDH基因）中的平均含量分别为（2．698±0．084）×10^4，（1．806±0．322）×10^4，（2．267±0．074）×10^4和（4．041±0．271）×10^4；而对照组这4种毒性因子基因含量分别为：（1．139±0．183）×10^6，（9．324±5．028）×10^3，（1．326±0．028）×10^6和（1．307±0．001）×10^6，均远比实验组高，其中以NMT最为明显。结论新生隐球菌被巨噬细胞吞噬后，主要的毒性因子基因表达下降，其中以NMT最为明显，而CNLAC1下降幅度最小。

Influence of macrophage on virulence factors mRNA expression of Cryptococcus neoformans

Objective To investigate the difference of main virulence genes mRNA level in Cryptococcus neoformans wild strain B3501 after phagocytosis by macrophage J774. 16. Method Co-cultured the logarithmic growing J774. 16 with Cryptococcus noeformans wild strain B3501. Collected the macrophage J774.16 challenged by yeast at co-cultured 4 hours and control group （ solely cultured at 37℃ ,5% CO2） yeast. Then their total RNA was extracted for screening the difference of main virulence factor gene CNLAC1 , CAP60, URE and NMT mRNA expression by real-time PCR. Results The production of the four virulence factor genes in the test group （ ingested yeast） （ CAP60, CNLAC 1, NMT and URE 1 ） （2. 698 ＋ 0. 084）×10^4, （ 1. 806 ± 0. 322） ×10^3, （2. 267 ± 0. 074） ×10^4 and （4.041 ±0.271 ） ×10^4 per million house keeper gene GAPDH, respectively. And those of the control group were （ 1. 139 ± 0.183 ）×10^6, （9. 324 ± 5. 028 ）×10^3, （ 1. 326 ＋ 0. 028 ） ×10^6 and （ 1. 307 ± 0. 001 ） ×10^6, respectively. The production of the four genes of the ingested Cryptococcus neoformans strain B3501 were 5-58 times less than those of the control group. Conclu- sion The four main virulence factor gene all expressed decreasingly after Cryptococcus neoformans was ingested by the maerophage. Among them, changement of N-myristoyltransferase gene （NMT） was the most significantly.