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不同声学参数的低强度脉冲超声对软骨细胞活性及培养骨胚生长的影响
Effects of low-intensity pulsed ultrasound of different acoustic parameters on viability of cultured chondrocytes and
ex vivo growth of mouse embryonic metatarsals- 第三军医大学学报 2019年41卷第10期 页码：907-912
- 作者机构：
400042 重庆, 陆军军医大学(第三军医大学)大坪医院野战外科研究所战创伤康复医学研究室, 创伤实验室, 骨代谢与修复中心, 创伤、烧伤与复合伤国家重点实验室, 全军军事训练伤防治与康复实验室
- 作者简介：
[ 杜晓兰，E-mail：DXL_xiaolan@163.com DU Xiaolan, E-mail: DXL_xiaolan@163.com
- 基金信息：国家自然科学基金面上项目(81772306);军事医学创新工程项目(16CXZ2016)
DOI：10.16016/j.1000-5404.201901169
中图分类号： R318.0; R454.4; R684.3收稿：2019-01-15，
修回：2019-02-23，
纸质出版：2019-05-30
- 稿件说明：
移动端阅览
刘汨, 谢杨丽, 黄俊兰, 等. 不同声学参数的低强度脉冲超声对软骨细胞活性及培养骨胚生长的影响[J]. 第三军医大学学报, 2019,41(10):907-912.
Mi LIU, Yangli XIE, Junlan HUANG, et al. Effects of low-intensity pulsed ultrasound of different acoustic parameters on viability of cultured chondrocytes and
ex vivo growth of mouse embryonic metatarsals[J]. Journal of Third Military Medical University, 2019, 41(10): 907-912.刘汨, 谢杨丽, 黄俊兰, 等. 不同声学参数的低强度脉冲超声对软骨细胞活性及培养骨胚生长的影响[J]. 第三军医大学学报, 2019,41(10):907-912. DOI： 10.16016/j.1000-5404.201901169.
Mi LIU, Yangli XIE, Junlan HUANG, et al. Effects of low-intensity pulsed ultrasound of different acoustic parameters on viability of cultured chondrocytes and
ex vivo growth of mouse embryonic metatarsals[J]. Journal of Third Military Medical University, 2019, 41(10): 907-912. DOI： 10.16016/j.1000-5404.201901169.
摘要
目的
2
探讨低强度脉冲超声(low-intensity pulsed ultrasound
LIPUS)对体外培养软骨细胞活性、增殖、分化和凋亡以及骨组织生长的影响。
方法
2
分离新生3~5 d C57BL/6J小鼠的软骨细胞和胚胎期18.5 d小鼠跖骨，分为对照组(未处理组)及3种LIPUS处理组，其声强度和占空比的参数为：LIPUSⅠ，30 mW/cm
2
-20%；LIPUSⅡ，30 mW/cm
2
-40%；LIPUSⅢ，50 mW/cm
2
-20%；其余参数为：中心频率1.5 MHz，脉冲重复频率1 KHz，处理时间20 min/d；分别用CCK-8、EdU掺入及TUNEL检测软骨细胞的活性变化、增殖、凋亡；定量RT-PCR检测蛋白聚糖(Aggrecan，Acan)、Ⅱ型胶原(Collagen Ⅱ，ColⅡ)、Ⅹ型胶原(Collagen Ⅹ，ColⅩ)等软骨基质相关基因表达；采集图像后测量跖骨长度，计算跖骨全长增
长率、软骨增长率。
结果
2
3种参数的LIPUS处理能促进软骨细胞的活性和增殖，抑制软骨细胞凋亡，其中LIPUSⅡ组效果最明显；定量RT-PCR结果显示：各处理组软骨细胞的Acan、ColⅡ的mRNA表达均较对照组明显增高，ColⅩ表达降低(
P
<
0.05)；跖骨长度计算结果显示：LIPUSⅡ、Ⅲ组软骨和全长增长率均显著高于对照组(
P
<
0.01)，而LIPUSⅠ组与对照组无明显差异。
结论
2
LIPUS可能通过促进软骨细胞活性、增殖以及细胞外基质合成，抑制细胞凋亡，加快骨组织生长，且效果与超声参数有关；其中声强度30 mW/cm
2
、占空比40%的LIPUS作用最明显。
Abstract
Objective
2
To investigate the effects of low-intensity pulsed ultrasound (LIPUS) on the viability
proliferation
differentiation and apoptosis of cultured chondrocytes and
ex vivo
development of embryonic mouse metatarsals.
Methods
2
Chondrocytes isolated from neonatal mice (3-5 days old) and metatarsals from mouse embryos (18 days of gestation) were both divided into control group and 3 LIPUS treatment groups for LIPUS exposures with acoustic intensity and duty ratio combinations of 30 mW/cm
2
-20% (LIPUS Ⅰ)
30 mW/cm
2
-40% (LIPUS Ⅱ)
and 50 mW/cm
2
-20% (LIPUS Ⅲ); all the other ultrasound parameters were identical (central frequency of 1.5 MHz
repetition frequency of 1 kHz and exposure time of 20 min per day). After LIPUS treatments
the chondrocytes were examined for changes in cell viability
proliferation and apoptosis using cell counting kit-8
EdU assay and TUNEL assay
respectively. The mRNA expression levels of cartilage matrix-related genes were examined by qRT-PCR. The changes in the total length and cartilage length of the metatarsals were measured after the treatments.
Results
2
Exposure to LIPUS significantly enhanced the viability and proliferation and reduced the apoptotic rate of the chondrocytes
and the effect was the most obvious in LIPUS Ⅱ group. LIPUS treatment significantly up-regulated the mRNA expressions of Col Ⅱ and proteoglycan and reduced the mRNA expression of Col X in the cho
ndrocytes (
P
<
0.05). As for the metatarsals
the growth rates of the total length and cartilage length were significantly greater in LIPUS Ⅱ and LIPUS Ⅲ groups than in the control group (
P
<
0.001)
but no significant difference was found between LIPUS Ⅰ group and control group.
Conclusion
2
LIPUS treatment can promote bone growth by stimulating chondrocyte proliferation and extracellular matrix synthesis while inhibiting cell apoptosis. These effects are related with the acoustic parameters of LIPUS
and the parameter setting with an acoustic intensity of 30 mW/cm
2
and a duty ratio of 40% can be optimal for LIPUS treatment.
关键词
Keywords
references
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