烟曲霉分生孢子表面形态改变及纤连蛋白受体分布状态  

Change of Surface Shape and Distribution of Fibronectin Receptors on Aspergillus fumigatus Conidia

作　　者：徐赤宇[1] 温海[1] 吴建华[1] 

机构地区：[1]第二军医大学长征医院皮肤科,上海200003

出　　处：《中华皮肤科杂志》2002年第5期361-363,共3页Chinese Journal of Dermatology

基　　金：国家自然科学基金资助项目(39770002)

摘　　要：目的观察烟曲霉分生孢子静止期及其在膨胀生发过程中表面形态的改变。观察纤连蛋白受体在孢子表面分布状态及其在膨胀生发过程中的变化。方法取培养成熟的烟曲霉,磷酸盐缓冲液冲洗后,通过3.5～4.0μm的钢丝网过滤提取制备纯净分生孢子悬液(108细胞/mL)。加入细胞培养液(RPMI1640+10%FBS),分别在0、2、4、6h取材固定,进行扫描电镜观察。另取定量分生孢子悬液(108细胞/mL)孵育在包被有定量纤连蛋白(50μg/mL)的微量测定细胞培养板中,37℃3h;用磷酸盐缓冲液冲洗3次,再分别加入兔抗人-纤连蛋白抗体和羊抗兔IgG-FITC孵育(37℃1h),胰酶消化后冲洗固定立即送共距焦免疫荧光电镜检查。结果静止期分生孢子表面有特征性的小棘状层形式,即有许多刺猬毛状的突起分布在其表面。孢子膨胀生发过程中,刺猬毛样小棘状层结构逐渐降解脱落,大约4h后完全降解脱落。孢子变成由条索状纤维质构成的光滑细胞壁球体;通过共距焦免疫荧光电镜发现FITC均匀地分布粘附在分生孢子表面刺猬毛样棘状突起上。结论静止期分生孢子的表面可表达出高水平纤连蛋白受体。带有放射荧光标记的纤连蛋白配体-受体均匀地分布在静止期分生孢子最外面小棘状层。孢子膨胀过程中表面小棘状层逐渐脱落。Objective To observe the changes of the outer ce ll wall in resting phase and during sw elling process of conidia of Aspergillus fumigatus,and the distribution of fibronectin receptors on conidia surface.Methods After being harvested with 3.5～4.0μm steel filters,the conidia were resuspended in RPMI 1640+10%FBS medium.The specimens were observed by the scanning electron microscopy after incubation for 0,2,4,6hours a t37℃respectively.The same amount of conidia was resuspendesd in PBS buffer,and put the suspension on 50μg /ml fibronectin coated microtitre plate,after incubation for 3hrs at 37℃,t he conidia were washed three times wi th PBS.Then rabbit antifibronectin antibody and fluorescein isothiocyanate(FITC)conjugated goat anti-rabbit immunoglob-ulin antibody were added,respectiv ely.After digesting with trypsin,t he conidia were washed again with PBS and examined by immuno-confocal microscopy.Results Acharacteristic echinulate pattern of protrusions was shown in the cell wall surface of resting coni dia.During swelling process,the ec hinulate layer of swollen conidia wa s gradually disintegrated,and almost complete ly shed 4hr later.Then,a smooth cell wall layer covered by some strands of fib-rillar material was shown.Binding o f fibronectin to the surface of conid ia was revealed by immuno-confocal m i-croscopy.FITC was homogeneously ad hered to the protrusions on the conid ial outer cell wall layer.Conclusion In resting phase the conidia of A.fumigatus express high levels of fibronectin r eceptors on their surface,they are h o-mogeneously distributed over the ex ternal echinulate layer of the conid ia,and shed gradually during the swe lling process.

关 键 词：烟曲霉分生孢子 纤连蛋白受体 烟曲霉 宿主细胞 细胞粘附 致病性 超微结构 

分 类 号：R379[医药卫生—病原生物学]