高强度聚焦超声对SD大鼠感染耻垢分枝杆菌病灶损伤效应的实验研究
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摘要
目的探究高强度聚焦超声(HIFU)对SD大鼠感染耻垢分枝杆菌(MS)结核动物模型的损伤效应。方法 20只SD大鼠臀部皮下分别注射MS悬液1 ml建立感染MS结核动物模型。选取10只成功建立模型的SD大鼠分为HIFU组和对照组,每组5只。HIFU组行HIFU辐照15 s(频率1 MHz,声强6369 W/cm2);对照组行假照15 s(频率1 MHz,声强0 W/cm2)。观察两组辐照前后结节B超灰度变化。对结节组织分别进行HE染色和抗酸染色,光学显微镜下观察结节组织病理变化;激光共聚焦显微镜及扫描电镜下观察HIFU辐照对结节的损伤效应。结果 HIFU组HIFU辐照前后SD大鼠结核结节的B超灰度值比较差异有统计学意义(P=0.004);对照组辐照前后结节的B超灰度值无明显变化。HE染色:对照组光学显微镜下见皮肤组织结构完整,皮肤附件结构正常,未发现明显的炎症浸润;HIFU组光学显微镜下见皮肤组织结构完整,真皮层组织未见损伤,皮肤附件正常,炎性细胞浸润加重,呈急性损伤表现。抗酸染色:对照组中MS聚集成团,HIFU组中MS较分散,呈短棒状。对照组和HIFU组MS存活率分别为93.61%和48.60%。经HIFU辐照后,MS菌体表面变得粗糙,菌体细胞壁破裂。结论一定剂量的HIFU辐照能有效抑制SD大鼠感染MS的结核结节,有望成为治疗结核病的新方法。
Objective To investigate the injury effect of high intensity focused ultrasound(HIFU)on mycobacterium smegmatis(MS)in tuberculous rat models. Methods By subcutaneous injection of 1 ml of MS suspension into the hip of SD rats,10 SD rats were successfully established MS tuberculous infection model and divided into HIFU group and control group,5 rats in each group.HIFU group was irradiated with HIFU for 15 s(frequency of 1 MHz,sound intensity of 6369 W/cm2),while control group was sham for 15 s(frequency of 1 MHz,sound intensity of 0 W/cm2).The changes of B-mode ultrasonic grayscale of nodules before and after irradiation were observed in the two groups. HE staining and acid-fast staining were used to observe the pathological changes of nodule tissues under optical microscope.The effects of HIFU irradiation damage were observed by confocal laser and scanning electron microscope.Results There was statistically significant difference in the B-ultrasound grayscale of the tuberculous nodules before and after irradiation in the HIFU group(P=0.004).There was no significant change in B-ultrasound grayscale of nodules before and after irradiation in the control group.HE staining:under the optical microscope,the skin tissue structure was intact and the skin attachment structure was normal in the control group,no obvious inflammatory infiltration was found. The structure of the skin tissue was intact under the optical microscope of the HIFU group,no damage was observed to the dermal tissue,and the skin appendages were normal,inflammatory cells infiltrated and presented acute injury.Acid-fast staining:MS in the control group was clustered,while MS in the HIFU group was dispersed and appeared as a short rod. Compared with the control group,the survival rate of MS bacteria in HIFU group decreased significantly(93.61% vs.48.60%). After HIFU irradiation,the surface of MS became rough and the cell wall was broken. Conclusion HIFU at a certain dose can effectively inhibit the infected MS tuberculous nodules in SD rats,which is expected to be a new method for the treatment of tuberculosis.
Objective To investigate the injury effect of high intensity focused ultrasound(HIFU)on mycobacterium smegmatis(MS)in tuberculous rat models. Methods By subcutaneous injection of 1 ml of MS suspension into the hip of SD rats,10 SD rats were successfully established MS tuberculous infection model and divided into HIFU group and control group,5 rats in each group.HIFU group was irradiated with HIFU for 15 s(frequency of 1 MHz,sound intensity of 6369 W/cm2),while control group was sham for 15 s(frequency of 1 MHz,sound intensity of 0 W/cm2).The changes of B-mode ultrasonic grayscale of nodules before and after irradiation were observed in the two groups. HE staining and acid-fast staining were used to observe the pathological changes of nodule tissues under optical microscope.The effects of HIFU irradiation damage were observed by confocal laser and scanning electron microscope.Results There was statistically significant difference in the B-ultrasound grayscale of the tuberculous nodules before and after irradiation in the HIFU group(P=0.004).There was no significant change in B-ultrasound grayscale of nodules before and after irradiation in the control group.HE staining:under the optical microscope,the skin tissue structure was intact and the skin attachment structure was normal in the control group,no obvious inflammatory infiltration was found. The structure of the skin tissue was intact under the optical microscope of the HIFU group,no damage was observed to the dermal tissue,and the skin appendages were normal,inflammatory cells infiltrated and presented acute injury.Acid-fast staining:MS in the control group was clustered,while MS in the HIFU group was dispersed and appeared as a short rod. Compared with the control group,the survival rate of MS bacteria in HIFU group decreased significantly(93.61% vs.48.60%). After HIFU irradiation,the surface of MS became rough and the cell wall was broken. Conclusion HIFU at a certain dose can effectively inhibit the infected MS tuberculous nodules in SD rats,which is expected to be a new method for the treatment of tuberculosis.
引文
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[6]谢霜,杨敏,杜永洪,等.低频低强度超声对巨噬细胞内耻垢分枝杆菌的损伤效应[J].第三军医大学学报,2018,40(16):1476-1482.
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[10]石廷玉,王园媛,向宇波,等.结核分枝杆菌生物被膜基质及潜在的药物靶点和治疗策略[J].中国科学:生命科学,2018,48(7):760-771.
[11]杜永洪,陈畅,陈飞,等.聚焦超声致早孕小鼠子宫平滑肌及胚胎损伤效应的观察[J].临床超声医学杂志,2013,15(1):1-4.
[12]向洋,李帅,叶合敏,等.低频低强度超声对白色念珠菌急性损伤效应的实验研究[J].临床超声医学杂志,2016,18(9):577-579.
[2] Reyrat JM,Kahn D.Mycobacterium smegmatis:an absurd model for tuberculosis?[J].Trends Microbiol,2001,9(10):472-474.
[3] Bohsali A,Abdalla H,Velmurugan K,et al. The non-pathogenic mycobacteria M.smegmatis and M.fortuitum induce rapid host cell apoptosis via a caspase-3 and TNF dependent pathway[J]. BMC Microbiol,2010,10(10):237.
[4]董源,李哲,唐筱潇,等.低频超声对BCG急性损伤效应的实验研究[J].中国超声医学杂志,2014,30(2):175-178.
[5]杜永洪,王陈美依,彭从峰,等.聚焦超声对卡介苗急性损伤效应的实验研究[J].中国超声医学杂志,2012,28(10):880-883.
[6]谢霜,杨敏,杜永洪,等.低频低强度超声对巨噬细胞内耻垢分枝杆菌的损伤效应[J].第三军医大学学报,2018,40(16):1476-1482.
[7] Zumla A,George A,Sharma V,et al. The WHO 2014 global tuberculosis report——further to go[J].Lancet Glob Health,2015,3(1):10-12.
[8] Donald PR,van Helden PD.The global burden of tuberculosis——combating drug resistance in difficult times[J].N Engl J Med,2009,360(23):2393-2395.
[9] Sotgiu G,Dara M,Centis R,et al.Breaking the barriers:migrants and tuberculosis[J].Presse Med,2017,46(2 Pt 2):5-11.
[10]石廷玉,王园媛,向宇波,等.结核分枝杆菌生物被膜基质及潜在的药物靶点和治疗策略[J].中国科学:生命科学,2018,48(7):760-771.
[11]杜永洪,陈畅,陈飞,等.聚焦超声致早孕小鼠子宫平滑肌及胚胎损伤效应的观察[J].临床超声医学杂志,2013,15(1):1-4.
[12]向洋,李帅,叶合敏,等.低频低强度超声对白色念珠菌急性损伤效应的实验研究[J].临床超声医学杂志,2016,18(9):577-579.
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