摘要
背景:课题组前期研究发现鹿角胶小分子肽能够促进骨生长,对于治疗骨类疾病具有良好的应用前景。然而鹿角胶如何在体内发挥作用以及原理尚未明确。目的:荧光标记及活体示踪技术研究鹿角胶体内靶器官分布动向。方法:使用异硫氰酸荧光素(fluorescein isothiocyanate,FITC)对鹿角胶进行荧光标记,通过荧光成像、紫外光谱扫描检测标记结果。将标记成功的鹿角胶,通过灌胃注入小鼠体内,通过激光共聚焦显微镜检测鹿角胶吸收入血情况,应用小动物活体成像仪检测鹿角胶在小鼠体内的分布情况。在血清及骨内荧光最强时间点取材,对血清及骨组织蛋白液进行凝胶电泳,通过二级质谱检测确定吸收入靶器官的鹿角胶成分。结果与结论:①葡聚糖凝胶色谱成功分离荧光标志物,荧光成像及紫外光谱扫描证明标记成功,并测得FITC-鹿角胶的荧光取代度为0.953%;②入血成分荧光强度检测结果显示口服2 h后鹿角胶在血清中分布最多;③不同时间小鼠的双侧股骨与胫骨荧光成像显示相同,证明鹿角胶可通过入骨发挥作用;④二级质谱检测对比UniProt数据库发现该肽段为核心蛋白聚糖的特征片段,证明鹿角胶中的核心蛋白聚糖可进入骨内发挥疗效。
BACKGROUND:Our previous studies found that the polypeptide of Cornus cervi Colla can promote bone growth,which has a good application prospect in the treatment of bone diseases.However,how Cornus cervi Colla works in the body and the principle are not clear.OBJECTIVE:To study the in vivo distribution and tracing of Cornus cervi Colla using fluorescence labeling and tracer technique.METHODS:Cornus cervi Colla was fluorescently labeled using fluorescein isothiocyanate,and the labeling results were detected by fluorescence imaging and UV spectral scanning.Successfully labeled Cornus cervi Colla was injected into mice by gavage,and the absorption of Cornus cervi Colla into blood was detected by laser confocal microscopy,and the distribution of Cornus cervi Colla in mice was detected by small animal in vivo imager.The distribution of Cornus cervi Colla in the mice was detected by laser confocal microscopy.Samples were taken from serum and bone at the time of the strongest fluorescence,and gel electrophoresis was carried out on serum and bone tissue protein solutions,and the components of Cornus cervi Colla absorbed into target organs were determined by secondary mass spectrometry.RESULTS AND CONCLUSION:The fluorescent markers were successfully separated by dextran gel chromatography,and the fluorescence imaging and ultraviolet spectrum scanning proved that the labeling was successful,and the fluorescence substitution degree of FITC-labeled Cornus cervi Colla was 0.953%.The fluorescence intensity of the components of Cornus cervi Colla in the blood showed that Cornus cervi Colla was most distributed in serum after oral administration for 2 hours.The fluorescence images of mice at different times were the same as those of bilateral femur and tibia,indicating that Cornus cervi Colla could play a role by entering the bone.Compared with UniProt database,secondary mass spectrometry showed that the peptide was a characteristic fragment of decorin.It is proved that decorin in Cornus cervi Colla can enter the bone to play a therapeutic role.
作者
胡亚楠
杜海涛
于洋
董丽敏
景田园
殷武
王平
Hu Yanan;Du Haitao;Yu Yang;Dong Limin;Jing Tianyuan;Yin Wu;Wang Ping(Shandong Hongjitang Pharmaceutical Group Co.,Ltd.,Jinan 250103,Shandong Province,China;School of Pharmaceutical Sciences,Shandong University of Traditional Chinese Medicine,Jinan 250355,Shandong Province,China;Shandong Academy of Chinese Medicine,Jinan 250014,Shandong Province,China;Advanced Medical Research Institute,Shandong University,Jinan 250012,Shandong Province,China;Nanjing University School of Life Science,Nanjing 210093,Jiangsu Province,China)
出处
《中国组织工程研究》
CAS
北大核心
2024年第28期4441-4446,共6页
Chinese Journal of Tissue Engineering Research
基金
山东省自然科学基金(ZR2020MH386),项目负责人:王平
中央引导地方科技发展专项基金(YDZX2021117),项目负责人:王平
山东省中央引导地方科技发展资金项目(YDZX20203700002055),项目负责人:王平
山东省重点研发计划(科技示范工程)(2021SFGC1205),项目负责人:王平。
关键词
鹿角胶
荧光示踪
入血成分
入骨成分
核心蛋白聚糖
Cornus cervi Colla
fluorescent tracer
components that enter the blood
components that enter the bone
decorin
