CN114854667B - 一种猕猴桃来源的植物纳米囊泡及其应用 - Google Patents
一种猕猴桃来源的植物纳米囊泡及其应用 Download PDFInfo
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Abstract
本发明公开了一种猕猴桃来源的植物纳米囊泡及其应用,其通过以下步骤制得:(1)将猕猴桃通过低速螺旋挤压榨出原浆;(2)将(1)中所得原浆通过筛网过滤除杂,收集滤液;(3)将(2)中得到的滤液依次进行低速、中速、高速和超速离心,每次离心后弃去沉淀,收集上清液进行下一次离心,其中最后一次离心,收集沉淀;(4)将(3)中所述最后一次离心收集的沉淀用缓冲液重悬后超速离心,收集沉淀;所述沉淀用缓冲液再次重悬后高速离心,收集上清液;所述上清液通过滤膜即得猕猴桃来源的植物纳米囊泡。所述猕猴桃来源的植物纳米囊泡制备方法简单,成本低,产量大,稳定性好。
Description
技术领域
本发明涉及一种纳米囊泡,具体涉及一种猕猴桃来源的植物纳米囊泡及其应用。
背景技术
肺癌是全世界最常见的癌症死亡原因,估计每年有160万人死亡。大约85%的患者有一组组织学亚型统称为非小细胞肺癌(NSCLC),其中肺腺癌和肺鳞癌是最常见的亚型。在过去的二十年里,非小细胞肺癌的治疗取得了重要进展,加深了我们对疾病生物学和肿瘤进展机制的了解,促进了早期发现和多模式护理。小分子酪氨酸激酶抑制剂的使用和免疫治疗在选定的患者中带来了前所未有的生存益处。然而,NSCLC的总体治愈率和存活率仍然很低,特别是在转移性疾病中。因此,需要继续研究新药和联合疗法,以便将临床益处扩大到更广泛的患者群体,并改善非小细胞肺癌的预后。
目前药物载体是一个高度热议的话题,其他的药物载体有脂质载体、聚合物载体、病毒型载体、无机分子载体等等。但是这些载体有热动稳定性低、定位靶向能力弱、非靶向的体内传递引起的高毒性、被机体生物代谢率低等问题。尤其是非靶向的体内传递大大局限了这些载体在新一代抗肿瘤药物中的应用。细胞外囊泡是由细胞分泌的含有RNA/DNA,脂质和蛋白质等的小泡。几乎所有的细胞都能够产生囊泡,因此囊泡来源广泛,能够进行大量生产。囊泡是天然的细胞间成分交流和传递的载体,其生物安全性高、免疫原性低、容易被细胞吸收,同时,所运输的药物在细胞内能够有效释放,不易在细胞内被降解。其众多特点使之成为一个非常有应用价值的药物载体。而且植物来源的囊泡比动物来源的囊泡更加具有上述优势。
猕猴桃,为猕猴桃科猕猴桃属植物猕猴桃Actinidia chinensis Planch.的果实,是一款深受人民喜爱的水果,但很少人知道它还是一味药食同源的中药。其药性较为平和,具有解热,止渴,健胃,通淋之功效,适合大多数人使用。现代药理研究表明猕猴桃具有抗肿瘤、降糖降脂、抗氧化、免疫调节等作用。
发明内容
发明目的:本发明的目的在于提供制备天然来源、安全的药物递送载体。猕猴桃来源的植物纳米囊泡是一种新型安全的药物载体,替代传统具有毒性的脂质体药物载体。
技术方案:本发明所述的猕猴桃来源的植物纳米囊泡,通过以下步骤制得:
(1)将猕猴桃通过低速螺旋挤压榨出原浆;
(2)将(1)中所得原浆通过筛网过滤除杂,收集滤液;
(3)将(2)中得到的滤液依次进行低速、中速、高速和超速离心,每次离心后弃去沉淀,收集上清液进行下一次离心,其中最后一次离心,收集沉淀;
(4)将(3)中所述最后一次离心收集的沉淀用缓冲液重悬后超速离心,收集沉淀;所述沉淀用缓冲液再次重悬后高速离心,收集上清液;所述上清液通过滤膜即得猕猴桃来源的植物纳米囊泡。
优选的,以上操作在4~25摄氏度环境中进行。
所述的猕猴桃来源的植物纳米囊泡,步骤(1)中所述低速螺旋挤压的转速为30~60转/分钟。
所述的猕猴桃来源的植物纳米囊泡,步骤(2)中所述筛网为200~2000目。
所述的猕猴桃来源的植物纳米囊泡,步骤(3)中所述低速离心的离心力为100~800×g,离心时间为5~10分钟;所述中速离心的离心力为1000~8000×g,离心时间为10~30分钟;所述高速离心的离心力为8000~12000×g,离心时间为30~60分钟;所述超速离心的离心力为100000~200000×g,离心时间为60~120分钟。
所述的猕猴桃来源的植物纳米囊泡,所述低速离心的离心力为200~500×g;所述中速离心的离心力为2000~5000×g;所述高速离心的离心力为100000~12000×g;所述超速离心的离心力为100000~120000×g。
所述的猕猴桃来源的植物纳米囊泡,步骤(4)中,所述缓冲液为磷酸盐缓冲液,所述缓冲液的pH范围为pH7.2~7.4;所述超速离心的离心力为100000~200000×g,离心时间为60~120分钟;所述高速离心的离心力为8000~12000×g,离心时间为30~60分钟。
所述的猕猴桃来源的植物纳米囊泡,所述滤膜的孔径为0.45μm。
所述的猕猴桃来源的植物纳米囊泡在制备药物载体中的应用。
所述的猕猴桃来源的植物纳米囊泡在制备抗肿瘤药物中的应用。
纳米囊泡RNA纳米颗粒复合物,由所述的猕猴桃来源的植物纳米囊泡作为载体,包裹siRNA。
有益效果:与现有技术相比,本发明具有如下优点:(1)所述猕猴桃来源的植物纳米囊泡制备方法简单。(2)成本低,产量大。(3)稳定性好(见图3)。(4)将猕猴桃作为植物来源的纳米囊泡作为药物载体安全性优。对囊泡进行修饰,赋予囊泡原本更多额外的功能,以实现特定部位药物递送以及体内成像和跟踪,是药物递送研究中的一个新兴领域。纳米囊泡RNA纳米颗粒复合物将拥有定位靶向能力高、无毒、热稳定性高、易被人体代谢降解、免疫原性低等等优势。
附图说明
图1为猕猴桃来源的植物纳米囊泡(KEVs)的制备过程图;
图2为电镜观察KEVs的形态;
图3为马尔文粒径仪分析的猕猴桃来源的纳米颗粒的粒径大小及分布;
图4为EGFRapt-KEVs-siRNA的稳定性实验;
图5为非小细胞肺癌肿瘤细胞吞噬KEVs;
图6为电镜观察KEVs装载siRNA(EGFRapt-KEVs-siRNA)后的形态;
图7为共聚焦显微镜观察EGFRapt-KEVs-siRNA的空间分布;
图8为EGFRapt-KEVs-siRNA体外将药物递送到非小细胞肺癌细胞的共聚焦图像;
图9为EGFRapt-KEVs-siRNA治疗非小细胞肺癌在体实验示意图;
图10为各组小鼠实验期间体重曲线;
图11为各组小鼠实验期间肿瘤生长曲线;
图12为各组小鼠肿瘤最终重量;
图13为各组小鼠肿瘤最终大小;
图14为各组小鼠肿瘤组织中STAT3蛋白表达情况;
图15为各组小鼠肿瘤组织中Ki67蛋白表达情况;
图16为各组小鼠肿瘤组织中凋亡情况;
图17为KEVs与常规脂质体安全性对比实验。
具体实施方式
根据下述实施例,可以更好地理解本发明。然而,本领域的技术人员容易理解,实施例所描述的内容仅用于说明本发明,而不应当也不会限制权利要求书中所详细描述的本发明。
实施例1
猕猴桃来源的纳米颗粒的制备
(1)将新鲜猕猴桃用清水洗净,在20℃环境中通过低速螺旋挤压技术榨出原浆;
(2)将榨出的原浆在20℃环境中经500目筛网过滤除杂,收集滤液;
(3)将收集的滤液先以200×g,离心10分钟,弃沉淀,收集上清;再将上清液以2000×g,离心30分钟,弃沉淀,收集上清;再将上清液以10000×g,离心60分钟,弃沉淀,收集上清;最后将上清液以120000×g,离心60分钟,收集沉淀;
(4)将沉淀用pH7.2的磷酸盐缓冲液重悬后,以120000×g,离心60分钟,收集沉淀;将沉淀再次用pH7.2的磷酸盐缓冲液重悬后,以10000×g,离心60分钟,收集上清液;最后通过孔径为0.45μm的除菌级滤膜后即获得纳米级颗粒。保存于-20℃~-80℃中。具体步骤见图1。
实施例2
(1)KEVs-siRNA的制备
siRNA的序列:
sense:5′-CCGUGGAACCAUACACAAATT-3′
antisense:5′-UUUGUGUAUGGUUCCACGGTT-3′
将6.67μM siRNA、0.34μg/μL KEVs与Exo-Fect混合,37℃条件下孵育10min,以100000×g,离心60分钟,沉淀用50μL PBS重悬。
(2)EGFRapt-KEVs-siRNA的制备
取上一步所得的KEVs-siRNA,接着加入0.5nM EGFRapt,37℃条件下孵育45min,然后在4℃条件下孵育60min,-80℃条件下保存备用,制得EGFRapt-KEVs-siRNA。
EGFRapt由以下四种序列构成(小写字母表示2′-OME核苷酸):
(1)3WJ-A-胆固醇:5′-uuG ccAuGu GuAuG GGG/3CholTEG/-3′
(2)3WJ-B:5′-ccc AcA uAc uuu Full GAu cc-3′
(3)3WJ-B-EGFR:5′-ccc AcA uAc uuu Guu GAu ccG ccu uAG uAA cGu Gcu uuGAuG ucG Au cGA cAG GAG Gc-3′;
(4)3WJ-C-AF647:5′-/5Alex647N/GGA ucA Auc AuG GcA A-3′
实施例3
电镜观察KEVs及EGFRapt-KEVs-siRNA的形态
将制备好的KEVs和EGFRapt-KEVs-siRNA 120000×g,60分钟超速离心后使其沉淀并压缩紧密,弃上清,沉淀用2.5%(v/v)戊二醛固定,送电镜室进行处理,上镜观察KEVs和EGFRapt-KEVs-siRNA的形态。如图2所示:电镜下可见KEVs和EGFRapt-KEVs-siRNA均为具有膜结构的纳米颗粒,大小在150-500nm之间,证实有效募集了猕猴桃来源的纳米级微体,即猕猴桃来源的植物纳米囊泡。
实施例4
马尔文粒径仪分析的猕猴桃来源的纳米颗粒的粒径大小及分布
将提取好的KEVs通过马尔文粒径仪进行粒径检测,如图3所示,KEVs的峰值均一性较好。
实施例5
EGFRapt-KEVs-siRNA的稳定性实验
将构建好的EGFRapt-KEVs-siRNA保存在4℃条件下,在第0天、第7天和第14天取出,使用NTA对其粒径进行测定,结果如图4所示。
实施例6
KEVs体外被非小细胞肺癌细胞摄取
人类非小细胞肺癌细胞系PC9-GR4-AZD1由Miguel Angel Molina和RafaelRosell(Pangaea,Barcelona,Spain)提供。
1.非小细胞肺癌细胞对KEVs的摄取
①将非小细胞肺癌细胞加入带有爬片的培养皿中,使其贴壁生长。
②将100μL KEVs用稀释液稀释至250μL。
③将1μL DiI加入250μL稀释液中,作为染料。
④将稀释的KEVs加入到染料中,快速混合。
⑤25℃孵育的2-5分钟,定时轻轻颠倒离心管保证在25℃充分混匀。
⑥加入等量血清中止染色反应孵育1分钟。
⑦120000×g,60分钟超速离心。
⑧吸弃上清,将染色的KEVs用100μL PBS重悬。
⑨以20μg/mL的浓度将KEVs加入培养的非小细胞肺癌细胞中,培养24小时。
⑩取出爬片,加入DAPI染色8分钟,激光共聚焦观察非小细胞肺癌细胞对KEVs的吞噬。
如图5所示:DAPI是一种细胞核燃料;DiI是一种红色的膜染料,能够与KEVs表面的膜结合。在激光共聚焦显微镜下,可以明显的观察到非小细胞肺癌细胞胞内出现了多个红色颗粒,表明非小细胞肺癌细胞能够有效的吞噬KEVs。
实施例7
EGFRapt-KEVs-siRNA空间分布观察
①将构建好的EGFRapt-KEVs-siRNA直接电镜和共聚焦显微镜下观测,如图6和7所示:KEVs将siRNA包裹在其内部的空腔。
实施例8
EGFRapt-KEVs-siRNA体外将siRNA递送到非小细胞肺癌细胞内
①将非小细胞肺癌细胞加入带有爬片的培养皿中,使其贴壁生长。
②加入EGFRapt-KEVs-siRNA(20μg/mL),24小时后,DAPI染色,共聚焦观察。
如图8所示:EGFRapt-KEVs-siRNA能够将siRNA有效的递送到细胞内。
实施例9
EGFRapt-KEVs-siRNA体内将siRNA递送到肿瘤内部,从而抑制肿瘤的生长。
EGFRapt-KEVs-siScramble是KEVs装载siScramble小干扰RNA,再在表面修饰有EGFRapt,具体制法同实施例2。
EGFRapt-KEVs-siSTAT3是KEVs装载siSTAT3制得的,即KEVs装载是实施例2所述siRNA,再在表面修饰有EGFRapt,具体制法同实施例2。
①雌性裸鼠,体重18-20克,购于江苏常州卡文斯实验动物有限公司。适应性生长1周。
②小鼠右侧腋下皮下接种人非小细胞肺癌细胞PC9-GR4-AZD1(2.0×106个/只),逐日观察。
③种瘤14天后,小鼠移植瘤大小平均值达到80-120mm3,将荷瘤小鼠随机分为3组:荷瘤模型组(尾静脉注射PBS)、EGFRapt-KEVs-siScramble组(EGFRapt-KEVs-siScramble 150μg/只)、EGFRapt-KEVs-siSTAT3组(EGFRapt-KEVs-siSTAT3 150μg/只),治疗间隔时间为6天(图9)。
④逐日观察各组小鼠生长状态,每2天测小鼠体重及肿瘤体积(肿瘤体积计算=长×宽2/2)
⑤治疗24天后,测体重后眼眶采血,引颈处死小鼠,剥离肿瘤组织测量体积并称重,其余主要脏器分离后甲醛固定。
结果显示:EGFRapt-KEVs-siSTAT3的尾静脉注射治疗及腹腔注射治疗能抑制人非小细胞肺癌生长(图10~13);通过对各组小鼠肿瘤组织组化切片的分析,发现EGFRapt-KEVs-siSTAT3的尾静脉注射治疗肿瘤组织中,STAT3蛋白表达降低(图14);同时,Ki67蛋白的表达水平也降低(图15),TUNEL水平上升(图16)。这些结果提示我们:EGFRapt-KEVs-siSTAT3在体内能够有效地将肿瘤组织中STAT3的表达,抑制肿瘤的生长。
实施例10
KEVs与常规脂质体安全性对比实验
使用商品化脂质体liposomes(YEASEN,China)40802ES02,用PBS稀释成不同浓度。
用PBS将KEVs稀释成不同浓度。
将上述两种载体与IC21和HEK293细胞进行相容性实验。在96孔板中每孔接种5000个细胞,在培养箱中过夜培养。加入不同浓度的KEVs和脂质体,置于培养箱中培养24h。每孔加入10μL的CCK-8试剂,置于培养箱孵育2-4h,用酶标仪检测吸光度。
结果显示,猕猴桃来源的植物纳米囊泡具有更高的生物相容性,对细胞活力的影响较小,具体结果见图17。
序列表
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Claims (6)
1.一种猕猴桃来源的植物纳米囊泡的制备方法,其特征在于,包括以下步骤:
(1)将猕猴桃通过低速螺旋挤压榨出原浆;
(2)将(1)中所得原浆通过筛网过滤除杂,收集滤液;
(3)将(2)中得到的滤液依次进行低速、中速、高速和超速离心,每次离心后弃去沉淀,收集上清液进行下一次离心,其中最后一次离心,收集沉淀;所述低速离心的离心力为100~800×g,离心时间为5~10分钟;所述中速离心的离心力为1000~8000×g,离心时间为10~30分钟;所述高速离心的离心力为8000~12000×g,离心时间为30~60分钟;所述超速离心的离心力为100000~200000×g,离心时间为60~120分钟;
(4)将(3)中所述最后一次离心收集的沉淀用缓冲液重悬后超速离心,收集沉淀;所述沉淀用缓冲液再次重悬后高速离心,收集上清液;所述上清液通过滤膜即得猕猴桃来源的植物纳米囊泡;所述缓冲液为磷酸盐缓冲液,所述缓冲液的pH范围为pH7.2~7.4;所述超速离心的离心力为100000~200000×g,离心时间为60~120分钟;所述高速离心的离心力为8000~12000×g,离心时间为30~60分钟;所述滤膜的孔径为0.45μm。
2.根据权利要求1所述的猕猴桃来源的植物纳米囊泡的制备方法,其特征在于,步骤(1)中所述低速螺旋挤压的转速为30~60转/分钟。
3.根据权利要求1所述的猕猴桃来源的植物纳米囊泡的制备方法,其特征在于,步骤(2)中所述筛网为200~2000目。
4.根据权利要求1所述的猕猴桃来源的植物纳米囊泡的制备方法,其特征在于,步骤(3)中所述低速离心的离心力为200~500×g;所述中速离心的离心力为2000~5000×g;所述高速离心的离心力为10000~12000×g;所述超速离心的离心力为100000~120000×g。
5.一种纳米囊泡RNA纳米颗粒复合物,其特征在于,由权利要求1所述的制备方法制得的猕猴桃来源的植物纳米囊泡作为载体,包裹siRNA,并与EGFR适配体复合形成EGFRapt-KEVs-siRNA。
6.权利要求5所述的纳米囊泡RNA纳米颗粒复合物在制备抗非小细胞肺癌药物中的应用。
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Citations (11)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009120950A2 (en) * | 2008-03-28 | 2009-10-01 | Donald Danforth Plant Science Center | Alteration of phospholipase de (plde) or phospholipase da3 (pld a3) expression in plants |
CN103202810A (zh) * | 2013-04-01 | 2013-07-17 | 浙江中医药大学 | 一种猕猴桃根多糖脂质体及其制备方法 |
CN103479597A (zh) * | 2012-06-14 | 2014-01-01 | 苏州恒宇生物科技有限公司 | 一种葡萄来源活性成分—纳米级膜性囊泡的制备方法及其用途 |
CN103479682A (zh) * | 2012-06-14 | 2014-01-01 | 苏州恒宇生物科技有限公司 | 一种植物来源活性成分-纳米级膜性囊泡的制备方法 |
CN103705550A (zh) * | 2012-10-08 | 2014-04-09 | 上海医药工业研究院 | 一种猕猴桃素及其制备方法和应用 |
WO2016097331A1 (en) * | 2014-12-19 | 2016-06-23 | MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. | Plant protection from a pest or pathogen by expression of double-stranded rnas in the plastid |
CN107375234A (zh) * | 2017-07-10 | 2017-11-24 | 武汉大学 | 一种基于体液中细胞源性囊泡的多功能载体及制备方法和应用 |
CN112654722A (zh) * | 2018-12-10 | 2021-04-13 | Md保健株式会社 | 来源于鞘氨醇单胞菌属细菌的纳米囊泡及其用途 |
CN113425849A (zh) * | 2021-06-18 | 2021-09-24 | 中国科学院苏州纳米技术与纳米仿生研究所 | siRNA递送载体及其制备方法与应用 |
CN114009772A (zh) * | 2020-09-09 | 2022-02-08 | 陕西佰瑞衡健康科技有限公司 | 一种植物外泌体微囊包埋冻干粉 |
CN114774348A (zh) * | 2022-04-26 | 2022-07-22 | 上海海洋大学 | 猕猴桃胞外囊泡及其在药物载体中的应用 |
-
2022
- 2022-06-14 CN CN202210670706.3A patent/CN114854667B/zh active Active
Patent Citations (11)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009120950A2 (en) * | 2008-03-28 | 2009-10-01 | Donald Danforth Plant Science Center | Alteration of phospholipase de (plde) or phospholipase da3 (pld a3) expression in plants |
CN103479597A (zh) * | 2012-06-14 | 2014-01-01 | 苏州恒宇生物科技有限公司 | 一种葡萄来源活性成分—纳米级膜性囊泡的制备方法及其用途 |
CN103479682A (zh) * | 2012-06-14 | 2014-01-01 | 苏州恒宇生物科技有限公司 | 一种植物来源活性成分-纳米级膜性囊泡的制备方法 |
CN103705550A (zh) * | 2012-10-08 | 2014-04-09 | 上海医药工业研究院 | 一种猕猴桃素及其制备方法和应用 |
CN103202810A (zh) * | 2013-04-01 | 2013-07-17 | 浙江中医药大学 | 一种猕猴桃根多糖脂质体及其制备方法 |
WO2016097331A1 (en) * | 2014-12-19 | 2016-06-23 | MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. | Plant protection from a pest or pathogen by expression of double-stranded rnas in the plastid |
CN107375234A (zh) * | 2017-07-10 | 2017-11-24 | 武汉大学 | 一种基于体液中细胞源性囊泡的多功能载体及制备方法和应用 |
CN112654722A (zh) * | 2018-12-10 | 2021-04-13 | Md保健株式会社 | 来源于鞘氨醇单胞菌属细菌的纳米囊泡及其用途 |
CN114009772A (zh) * | 2020-09-09 | 2022-02-08 | 陕西佰瑞衡健康科技有限公司 | 一种植物外泌体微囊包埋冻干粉 |
CN113425849A (zh) * | 2021-06-18 | 2021-09-24 | 中国科学院苏州纳米技术与纳米仿生研究所 | siRNA递送载体及其制备方法与应用 |
CN114774348A (zh) * | 2022-04-26 | 2022-07-22 | 上海海洋大学 | 猕猴桃胞外囊泡及其在药物载体中的应用 |
Non-Patent Citations (1)
Title |
---|
Li等.Arrowtail RNA for Ligand Display on Ginger Exosome-like Nanovesicles to Systemic Deliver siRNA for Cancer Suppression.Scientific Reports.2018,(第8期),14644. * |
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