CN113559238B - 一种含有活性肽并具有抗衰老作用的药品或化妆品 - Google Patents
一种含有活性肽并具有抗衰老作用的药品或化妆品 Download PDFInfo
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- CN113559238B CN113559238B CN202110935776.2A CN202110935776A CN113559238B CN 113559238 B CN113559238 B CN 113559238B CN 202110935776 A CN202110935776 A CN 202110935776A CN 113559238 B CN113559238 B CN 113559238B
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Abstract
本发明涉及一种含有活性肽并具有抗衰老作用的药品或化妆品。本发明通过在培养干细胞时通过添加FGF和高氧条件下制备获得了高活性的干细胞外泌体，将所述外泌体负载具有抗氧化活性的活性肽之后，通过实验证实了所述负载多肽的外泌体具有促进皮肤成纤维细胞抗衰老的特性，通过小鼠实验也证明了所述外泌体能够提高皮肤的抗衰老性能，具有较好的市场应用价值。
Description
技术领域
本申请涉及生物医药领域，更具体的涉及一种含有活性肽并具有抗衰老作用的药品或化妆品。
背景技术
衰老是机体各组织、器官功能随年龄增长而发生退行性变化的过程。衰老可以降低机体面对环境胁迫维持动态平衡的能力，从而增加机体患病和死亡的可能性。衰老与高血压、2型糖尿病、动脉粥样硬化、老年痴呆等疾病密切相关。机体衰老与组织再生性细胞减少、脏腑虚损、机体内自由基增加、机体中毒、饮食无节律等相关，是体内外许多因素(环境污染、精神紧张、遗传等)共同作用的结果。
衰老是不可逆的生命进程，最普遍的抗衰老方式是使用抗衰老药物。合成药物是用化学或生物方法制成的具有预防、治疗和诊断效果的物质，广泛应用于临床。目前临床上使用的延缓衰老药大多是合成药物，如维生素E可促进细胞分裂及抑制氧自由基生成、普鲁卡因制剂能延长细胞寿命、酰胺吡酮可延缓脑衰老等。而阿司匹林通过对抗氧化应激，延缓年龄相关的机体功能下降，从而延长线虫寿命；二甲双胍作为腺苷酸活化蛋白激酶(AMPK)激活剂，也可改善认知障碍，对延缓衰老也有一定的作用。而PAL-12、白藜芦醇类似物等药物的抗衰老作用则在近期颇受青睐。
目前，活性生物多肽具有安全稳定、易溶于水、易吸收、分子量小等优点，同时具有清除自由基、抗氧化、抗衰老等多种生物活性，不仅能够促进皮肤细胞的增殖，还可以为皮肤提供营养，延缓皮肤老化，促进皮肤创伤修复，因此被广泛应用于皮肤化妆品和抗衰老化妆品中。当今，因为社会老龄化程度加重以及消费者对年轻美丽的不断追求，抗衰老护肤品在化妆品市场中所占的比例也在逐年提升。消费者对抗衰老化妆品的安全性和有效性的需求也促进了对抗衰老化妆品的研究和开发技术的不断改进和完善。根据目前市场上已在售的抗衰老产品调查，已有多个国际知名品牌推出了肽类抗衰老化妆品，例如：玉兰油、雅芳、雅诗兰黛、兰蔻、欧莱雅、香奈儿、TheOrdinary、迪奥、SK-Ⅱ等国际一线知名品牌，可谓掀起了肽类美容护肤品的热潮。
除此之外，外泌体用于皮肤抗衰老的研究也越来越多。外泌体(exosomes)是一种由细胞分泌的直径为40～150nm、密度范围为1.09～1.18g/ml的具有双层磷脂膜结构的囊泡样物质。目前已经从各物种、各类型的细胞培养上清中提取出了外泌体，如神经元细胞、间充质干细胞、成纤维细胞、内皮细胞、巨核细胞等。有研究表明，小鼠下丘脑神经干细胞(hypothalamicneuralstemcell，htNSC)的损耗，导致小鼠出现了衰老的症状以及生命周期缩短。向衰老小鼠脑室注射神经干细胞外泌体可以延缓脑衰老速度，延长小鼠寿命。研究还显示，外泌体携带的miRNA能够部分调节htNSC的抗老化效应，并在htNSC进入脑脊髓液后发挥相应作用。在皮肤抗衰老方面，韩国的Kim等研究发现，脐带间充质干细胞来源的外泌体可以轻易穿透角质层到达真皮，促进人皮肤组织Ⅰ型胶原和弹力蛋白的生成，改善皮肤质地。这些研究表明外泌体具有抗衰老应用前景。在早期发现机体衰老迹象方面，外泌体也有其临床应用价值。唾液里有很多分子物质和微生物，可以作为局部或全身的有效生物指标，血液来源的分子会通过跨细胞或细胞旁路进入唾液腺进而影响唾液的组成成分。日本冈山大学研究人员推测唾液外泌体的miRNA可作为衰老的生物标记物。有研究对青年人群和老年人群的唾液外泌体进行miRNA的筛选和验证，发现miR-24-3p的表达有显著差异，从而认为miR-24-3p是一个新的衰老相关的生物标记物。此结果表明，唾液外泌体检测有希望作为一种标记衰老的手段。
虽然抗衰老机制和抗衰老药物的研究发展迅速，但研究还是不够深入，多数药物抗衰老效果不甚理想。临床直接用于治疗衰老的药物品种不多，还需进一步研究衰老分子机制、发掘抗衰老药物。
发明内容
本发明克服现有技术的缺陷，提供了一种抗衰老的化妆品。
本发明一方面，提供了一种源自高活性骨髓间充质干细胞的外泌体。
另外一方面，本发明还提供所述外泌体的制备方法，具体包括如下步骤：
取骨髓加等量PBS混匀，用针头沿管壁慢慢注入到1.073g/ml的Pereoll淋巴细胞分离液上，使之成为单细胞悬液，2000r/min离心30min。轻轻吸出中间呈白色浑浊状的骨髓间充质干细胞层，放入另一个离心管中，加入PBS，1500r/min离心15min，如此共洗3次。弃PBS，用含10％FCS的L-DMEM培养液制成的细胞悬液进行计数板计数，以1X109/L的细胞数量接种于含青霉素、链霉素各10000U/100ml、含10％FCS的L-DMEM培养液中。培养48h后第一次换液可见贴壁细胞变成梭形，细胞融合达到85％时传代接种于25ml 60Co照射照射量5Gy处理后的细胞培养瓶中，置于5％CO2、37℃二氧化碳培养箱中培养；48h后首次换液，吸弃未贴壁的悬浮细胞，加入新鲜的含10％FCS的L-DMEM培养液继续孵育，每3～4d换液。待贴壁细胞80％融合后，弃掉培养液，PBS冲洗瓶壁，0.25％胰酶消化，镜下观察控制2～3min，用含血清培养液终止消化，反复吹打瓶壁，制成单细胞悬液，按1X 109/L再次传代，至第三代时收获细胞；
将骨髓间充质干细胞使用无血清MSC培养基重悬，接种于底面积25cm2的培养皿中，接种细胞数1×106个，培养基20ml；细胞增殖到90％时用无菌PBS溶液清洗细胞表面，重复2次，加入20ml添加有5mg/L FGF的α-MEM基础培养基在1％O2、3％CO2的培养条件下培养3d后收集培养上清进行外泌体提取；
将收集到细胞上清分别装于50ml离心管中，常温3500rpm，离心25min，去除细胞及其细胞碎片；收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5％的5×PEG 6000试剂，4℃静置过夜；将过夜静置的PEG 6000细胞上清混合液离心，3000rpm离心30min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min。按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000rpm离心20min，弃上清，按照1g:100mL用PBS重悬即获得间充质干细胞外泌体。
本发明通过在培养干细胞时添加FGF以及在1％O2、3％CO2的培养条件下培养时，能够有效的促进外泌体产品的提高，制备得到的外泌体蛋白含量也得到显著的提高。
本发明另外一个方面，发明人前期从鲑鱼鱼皮中通过胶原蛋白水解工艺分离多肽组分鉴定并分离制备得到了2条抗氧化肽分别是DYT-1多肽和DYT-4多肽，所述抗氧化肽具有较好的对DPPH自有基、羟基自由基等的清除能力，具有较好的抗氧化特性。
本发明还有一个方面，提供一种负载所述多肽的外泌体。
具体的，所述负载多肽的外泌体的制备方法为，在1mL 50mM海藻糖PBS溶液中分别加入外泌体100μg、1mg/mL SEQ ID NO:1多肽DMSO溶液20μl；混匀后，加入4mm电穿孔皿中，放入电穿孔仪，按照如下反应条件：电压120V，电容155μF，放电时间1ms，放电次数：2次；穿孔后即可将混悬液放入细胞培养箱中孵育1个小时，继之100000g超速离心两次，每次80min，小心弃尽上清，用PBS液重悬沉淀即为负载多肽外泌体。
本发明另外一方面，还提供一种抗衰老的美容化妆品，所述化妆品中含有负载有多肽的外泌体。
进一步的，所述化妆品是凝胶形式，具体的，所述凝胶的制备方法是按照如下配方采用本领域常规的制备方法来制备：0.65％的丙烯酸(酯)类/C10-30烷醇丙烯酸酯交联聚合物(U-21)，0.05％的EDTA二钠，3％的甘油，3％丁二醇，0.25％PHENONIP，1％负载有多肽的外泌体，4％分子透明质酸钠，然后用氢氧化钾10％溶液调节pH为5.6，余量为水。
在一些情况下，化妆品可以是那些用于治疗或护理，或以某种方式保湿，改善，加速更新，保护，防止损伤或清洁皮肤的产品。
一种化妆品可作为乳膏、外用贴剂，水凝胶贴剂，透皮贴剂，软膏，凝胶，液体，粉末，洗剂，血清，乳剂，油，粘土，保湿剂，泡沫，面膜，摩丝，气雾剂，喷雾剂，清洁剂，调色剂或洗发剂。
在一些情况下，化妆品可以是粘合剂的形式，一种绷带，剥落剂，牙膏，保湿剂，洗剂，底漆，唇膏，唇膏，无水封闭保湿剂，止汗剂，除臭剂，个人清洁产品，封闭药物递送贴剂，指甲油，粉末，纸巾，擦拭物，护发素或剃须膏。
在一些情况下，化妆品可包含皮肤调理剂(例如湿润剂，剥落剂，润肤剂或水合物)。在一些情况下，润肤剂是防止水分流失并对皮肤具有软化和舒缓作用的试剂。在一些实施例中，润肤剂可包含植物油中的至少一种，矿物油，牛油树脂，可可脂，凡士林，脂肪酸(动物油，包括EMU，水貂和羊毛脂)，甘油三酯，苯甲酸酯，肉豆蔻酸酯，棕榈酸酯，硬脂酸酯，糖脂，磷脂，角鲨烯，甘油，玫瑰果油，ANDIROBA油，葡萄籽油，鳄梨油，李子油，Pracaxi油，蓝藻油，杏仁油，Argan油，辛酸/癸酸甘油三酯，荷荷巴油，荷荷巴油，Spectrastat G2，神经酰胺和藻类提取物。
在一些情况下，化妆品还额外包括但不限于甘油，角鲨烯，山梨醇，透明质酸，透明质酸衍生物，透明质酸钠，透明质酸钠交联聚合物。
有益效果
本发明通过在培养干细胞时通过添加FGF和高氧条件下制备获得了高活性的干细胞外泌体，将所述外泌体负载具有抗氧化活性的活性肽之后，通过实验证实了所述负载多肽的外泌体具有促进皮肤成纤维细胞抗衰老的特性，通过小鼠实验也证明了所述外泌体能够提高皮肤的抗衰老性能，具有较好的市场应用价值。
附图说明
图1抗氧化能力检测结果图
图2细胞增殖结果图
具体实施方式
下面结合具体实施例对本发明进行进一步描述，但本发明的保护范围并不仅限于此：下述实施例中所用材料、试剂等，如无特别说明，均可从商业途径获得。
实施例1高活性干细胞的培养
取骨髓1ml加等量PBS混匀，用针头沿管壁慢慢注入到1.073g/ml的Pereoll淋巴细胞分离液上，使之成为单细胞悬液，2000r/min离心30min。轻轻吸出中间呈白色浑浊状的骨髓间充质干细胞层，放入另一个离心管中，加入PBS，1500r/min离心15min，如此共洗3次。弃PBS，用含10％FCS的L-DMEM培养液制成的细胞悬液进行计数板计数，以1X109/L的细胞数量接种于含青霉素、链霉素各10000U/100ml、含10％FCS的L-DMEM培养液中。培养48h后第一次换液可见贴壁细胞变成梭形，细胞融合达到85％时传代接种于25ml 60Co照射(照射量5Gy)处理后的细胞培养瓶中，置于5％CO2、37℃二氧化碳培养箱中培养。48h后首次换液，吸弃未贴壁的悬浮细胞，加入新鲜的含10％FCS的L-DMEM培养液继续孵育，每3～4d换液。待贴壁细胞80％融合后，弃掉培养液，PBS冲洗瓶壁，0.25％胰酶消化，镜下观察控制2～3min，用含血清培养液终止消化，反复吹打瓶壁，制成单细胞悬液，按1X 109/L再次传代，至第三代时收获细胞。
细胞鉴定：(1)将经过预处理的盖玻片预先铺在培养板底部，以制备细胞爬片。取生长良好的第3代细胞，1×104/ml接种于16孔培养板内。(2)培养2d后，细胞融合70％时去除培养液。(3)PBS(0.01M，pH7.4)冲洗5minX3。(4)10％甲醛溶液固定10min。(5)PBS冲洗5min×3。(6)3％过氧化氢室温孵育10min以消除内源性过氧化物酶。(7)PBS冲洗5min×3。(8)分别加人CD29、CD44、CD34、CD45单克隆抗体，阴性对照组加入PBS。(9)4℃孵育过夜。(10)次日取出，去除细胞培养板孔内一抗，PBS冲洗5minX3。(11)加入辣根过氧化物酶标记羊抗兔二抗。(12)4℃孵育30min。(13)PBS冲洗5min X 3，加入DAB显色：滴加适当比例稀释的DAB显色剂，显微镜下观察显色情况，自来水充分冲洗终止显色。
观察传至第3代的MSCs形态与原代细胞样仍呈长梭形，免疫细胞化学对细胞进行染色，CD34、CD45表达为阴性，CD29、CD44表达为阳性，CD29阳性率为(97.8±2.9)％、CD44阳性率为(99.1±3.1)％，细胞均一性高，表明制备的干细胞较好。
实施例2干细胞外泌体的制备
将实施例1鉴定的骨髓干细胞，使用无血清MSC培养基重悬，接种于底面积25cm2的培养皿中，接种细胞数1×106个，培养基20ml。细胞增殖到90％时用无菌PBS溶液清洗细胞表面，重复2次，加入20ml添加有5mg/L FGF的α-MEM基础培养基在1％O2、3％CO2的培养条件下培养3d后收集培养上清进行外泌体提取。
细胞计数后将收集到细胞上清分别装于50ml离心管中，常温3500rpm，离心25min，去除细胞及其细胞碎片。收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5％的5×PEG 6000试剂，4℃静置过夜。将过夜静置的PEG 6000细胞上清混合液离心，3 000rpm离心30min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min。按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000rpm离心20min，弃上清，按照1g:100mL用PBS重悬即获得外泌体溶液。通过投射电镜检测外泌体大体为圆形的小囊泡，直径在50-110nm左右。根据BCA试剂盒说明书测定提取的外泌体所含总蛋白量，测定得到外泌体中总蛋白的含量是(916.9±47.2)μg/L×107个细胞。采用流式细胞学检测外泌体中标志物的表达情况，其中高表达CD63和CD44，说明分离的是间充质干细胞外泌体。
实施例3对照干细胞外泌体的制备
将实施例1鉴定的骨髓干细胞，使用无血清MSC培养基重悬，接种于底面积25cm2的培养皿中，接种细胞数1×106个，培养基20ml。细胞增殖到90％时用无菌PBS溶液清洗细胞表面，重复2次，加入20mlα-MEM基础培养基在3％CO2的培养条件下培养3d后收集培养上清进行外泌体提取。
细胞计数后将收集到细胞上清分别装于50ml离心管中，常温3500rpm，离心25min，去除细胞及其细胞碎片。收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5％的5×PEG 6000试剂，4℃静置过夜。将过夜静置的PEG 6000细胞上清混合液离心，3 000rpm离心30min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min。按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000rpm离心20min，弃上清，按照1g:100mL用PBS重悬即获得外泌体溶液。通过投射电镜检测外泌体大体为圆形的小囊泡，直径在50-110nm左右。根据BCA试剂盒说明书测定提取的外泌体所含总蛋白量，测定得到外泌体中总蛋白的含量是(803.4±53.7)μg/L×107个细胞。采用流式细胞学检测外泌体中标志物的表达情况，其中高表达CD63和CD44，说明分离的是间充质干细胞外泌体。从结果可以看出，外泌体中总蛋白含量比实施例2的低。
实施例4负载多肽纳米材料的外泌体制备
在1mL 50mM海藻糖PBS溶液中分别加入实施例2或实施例3的外泌体100μg、1mg/mLSEQ ID NO:1DYT-4多肽DMSO溶液20μl；混匀后，加入4mm(容积750μl)电穿孔皿中，放入电穿孔仪，按照如下反应条件：电压120V，电容155μF，放电时间1ms，放电次数：2次。穿孔后即可将混悬液放入细胞培养箱中孵育1个小时，继之100000g超速离心两次，每次80min，小心弃尽上清，用PBS液重悬沉淀即为负载多肽外泌体，4℃储存备用。
实施例5抗氧化能力检测
量取不同浓度的多肽、外泌体以及负载多肽的外泌体样品溶液以Vc为对照，添加8mmol/L的硫酸亚铁溶液0.3mL、20mmol/L的水杨酸溶液1mL置于离心管中混合，使用37℃的温水进行水浴，随后取出离心管冷却，添加0.45mL蒸馏水，定容至3mL，使用转速为3000r/min条件离心10min，在510nm测量溶液上清液的吸光度。每一个溶液样品制作3组平行，样品对羟自由基的清除率按下式计算。羟基自由基清除率(％)＝((A0－(A1－A2))/A0)×100式中：A0为不加样品溶液的吸光度；A1为加入样品溶液、硫酸亚铁、水杨酸-乙醇、过氧化氢溶液的吸光度；A2为不加过氧化氢溶液的吸光度。结果如图1所示。
从图1可以看出，与对照Vc相比，本发明的DYT-4多肽和外泌体均具有消除羟基自由基能力，虽然相同浓度下没有Vc的清除能力强，但是当将多肽负载到外泌体后，其羟基自由基的清除能力得到显著的提高，在250ug/mL的浓度下，最高能达到97％的清除羟基自由基的能力。
实施例6人衰老皮肤成纤维细胞实验
皮肤标本按照“植块法原代培养人衰老皮肤成纤维细胞”中所述的植块法原代培养，建立衰老的人皮肤成纤维细胞系。实验选用第4代细胞。分为0、50、100、200mg/L多肽组、外泌体组以及负载多肽的外泌体组，Vc组为对照，处理细胞72h后，四甲基偶氮唑盐(MTT)比色法检测细胞增殖情况；结果如图2所示。
从图2的结果可以看出，培养72h各组细胞在添加了相应的药物后，均相对于不添加的0mg/L的条件下细胞均得到促进生长的效果，相对于对照组，负载多肽的外泌体在200mg/L的浓度下作用72h后吸光度可以达到0.82，具有较好的促进衰老细胞增殖作用。
对培养72h的细胞，针对空白没有给药的人衰老皮肤成纤维细胞以及药物浓度为200mg/L处理的各组细胞进行抗氧化酶活性测定：分别收集各组细胞，超声破碎.离心取上清液，按试剂盒说明书中的方法测定SOD、CAT、GSH-Px活性。结果如表1所示。
表1 SOD、CAT、GSH-Px检测结果
从表1的结果可以看出，多肽组、外泌体组、负载多肽的外泌体组和Vc对照组均相对于空白组能够提高SOD、CAT、GSH-Px的活性，相对于对照组，负载多肽的外泌体组能够更明显的提高相应的各组分的SOD、CAT、GSH-Px活性，SOD为(24.23±2.23)KU/L，CAT为(2.74±0.88)KU/L，GSH-Px为(22.17±2.89)KU/L，具有较好的效果。
实施例7凝胶抗衰老化妆品的制备及验证
凝胶的制备按照如下配方采用本领域常规的制备方法来制备：0.65％的丙烯酸(酯)类/C10-30烷醇丙烯酸酯交联聚合物(U-21)，0.05％的EDTA二钠，3％的甘油，3％丁二醇，0.25％PHENONIP，1％多肽负载外泌体，4％分子透明质酸钠，然后用氢氧化钾10％溶液调节pH为5.6，余量为水。
首先将雌性小鼠适应性饲养7d，保证小鼠适应现环境后开始进行实验。将3雌性小鼠按照随机数字表分组的方法分成5组，其中4组小鼠作为造模组，皮下注射D-半乳糖1.0g·kg-1·d-1，一共注射30d；余下一组小鼠作为空白组，每天注射同体积生理盐水。30d后造模组和空白组皮肤外观比对，模型组明显皮肤松弛，出现细密皱纹，而空白组则相反。将4组皮肤衰老模型小鼠分为：模型组、多肽组、外泌体组和负载多肽的外泌体组。用剪刀将小鼠背部毛发剪短，再用剃须刀剃毛备用。选取小鼠背部中央位置4cm×4cm的区域，将各组凝胶涂抹在对应各组小鼠选取的皮肤区域表面，每只每天涂抹0.2g，24h后清洁皮肤并再次涂抹，连续涂抹21d，期间还需手工脱毛4次。
取涂药部位皮肤组织0.5g，用冰浴预冷的生理盐水进行漂洗，滤纸擦干，加入冰浴预冷的生理盐水，使用玻璃匀浆器研磨成浓度为10％的匀浆液。将得到的匀浆液在0℃下3000r/min离心10min，取上清液。按照HYP试剂盒说明书所示方法，使用超微量微孔板分光光度计测定OD值，计算各实验组小鼠皮肤HYP含量。
表2小鼠皮肤组织HYP含量结果
| 组别 | 药物浓度(g) | HYP(μg/mg) |
| 空白组 | 0 | 4.16±0.04 |
| 多肽组 | 0.2g/d | 4.03±0.11 |
| 外泌体组 | 0.2g/d | 3.82±0.14 |
| 负载多肽的外泌体组 | 0.2g/d | 4.29±0.24 |
| 模型组 | 0 | 3.61±0.14 |
从表2的结果可以看出，模型组小鼠皮肤组织HYP含量显著低于空白组，说明小鼠的皮肤衰老模型造模成功。多肽组和外泌体组以及负载多肽的外泌体组均能够相对于模型组具有提高的HYP效果，特别是负载多肽的外泌体组HYP达到了(4.29±0.24)μg/mg，具有较好的效果。
应当理解，以上所述的仅是本发明的一些实施方式，应当指出，对于本领域的普通技术人员来说，在不脱离本发明的创造构思的前提下，还可以做出其它变形和改进，这些都属于本发明的保护范围。
序列表
<110> 北京戴域生物技术有限公司
<120> 一种含有活性肽并具有抗衰老作用的药品或化妆品
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 1
Trp Pro Arg Ser Gly Asp Leu Ser Lys Met Glu Asn Met Gln
1 5 10
Claims (7)
1.一种抗衰老的药物，其特征在于：所述药物含有负载有多肽的外泌体，并且是凝胶形式；其中外泌体的制备方法为将骨髓间充质干细胞使用无血清MSC培养基重悬，接种于底面积25 cm2的培养皿中，接种细胞数1×106个，培养基20 ml；细胞增殖到90%时用无菌PBS溶液清洗细胞表面，重复2次，加入20 ml 添加有5mg/L FGF的α-MEM基础培养基在1%O2、3%CO2的培养条件下培养3 d后收集培养上清进行外泌体提取；将收集到细胞上清分别装于50 ml离心管中，常温3500 rpm，离心25min，去除细胞及其细胞碎片；收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5%的5×PEG 6000试剂，4 ℃静置过夜；将过夜静置的PEG 6000细胞上清混合液离心，3 000 rpm离心30 min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min，按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000 rpm离心20 min，弃上清，按照1g:100mL用PBS重悬即获得间充质干细胞外泌体；
所述负载多肽的外泌体的制备方法为，在1mL 50mM海藻糖PBS溶液中分别加入外泌体100μg、1mg/mL SEQ ID NO:1 DYT-4多肽DMSO溶液20μl；混匀后，加入4mm电穿孔皿中，放入电穿孔仪，按照如下反应条件：电压120V，电容155μF，放电时间1ms，放电次数：2次；穿孔后即可将混悬液放入细胞培养箱中孵育1个小时，采用100000g超速离心两次，每次80min，小心弃尽上清，用PBS液重悬沉淀即为负载多肽外泌体；
将所述负载多肽外泌体制备凝胶，所述凝胶的制备方法是按照如下配方采用本领域常规的制备方法来制备：0.65％的丙烯酸(酯)类／C10-30烷醇丙烯酸酯交联聚合物，0.05％的EDTA二钠，3％的甘油，3％丁二醇，0．25％PHENONIP，1％负载有多肽的外泌体，4％分子透明质酸钠，然后用氢氧化钾10％溶液调节pH为5.6，余量为水。
2.一种抗衰老的化妆品，其特征在于：所述化妆品含有负载有多肽的外泌体，并且是凝胶形式；其中外泌体的制备方法为将骨髓间充质干细胞使用无血清MSC培养基重悬，接种于底面积25 cm2的培养皿中，接种细胞数1×106个，培养基20 ml；细胞增殖到90%时用无菌PBS溶液清洗细胞表面，重复2次，加入20 ml 添加有5mg/L FGF的α-MEM基础培养基在1%O2、3%CO2的培养条件下培养3 d后收集培养上清进行外泌体提取；将收集到细胞上清分别装于50ml离心管中，常温3500 rpm，离心25min，去除细胞及其细胞碎片；收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5%的5×PEG 6000试剂，4 ℃静置过夜；将过夜静置的PEG 6000细胞上清混合液离心，3 000 rpm离心30 min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min，按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000 rpm离心20 min，弃上清，按照1g:100mL用PBS重悬即获得间充质干细胞外泌体；
所述负载多肽的外泌体的制备方法为，在1mL 50mM海藻糖PBS溶液中分别加入外泌体100μg、1mg/mL SEQ ID NO:1 DYT-4多肽DMSO溶液20μl；混匀后，加入4mm电穿孔皿中，放入电穿孔仪，按照如下反应条件：电压120V，电容155μF，放电时间1ms，放电次数：2次；穿孔后即可将混悬液放入细胞培养箱中孵育1个小时，采用100000g超速离心两次，每次80min，小心弃尽上清，用PBS液重悬沉淀即为负载多肽外泌体；
将所述负载多肽外泌体制备凝胶，所述凝胶的制备方法是按照如下配方采用本领域常规的制备方法来制备：0.65％的丙烯酸(酯)类／C10-30烷醇丙烯酸酯交联聚合物，0.05％的EDTA二钠，3％的甘油，3％丁二醇，0．25％PHENONIP，1％负载有多肽的外泌体，4％分子透明质酸钠，然后用氢氧化钾10％溶液调节pH为5.6，余量为水。
3.如权利要求2所述的化妆品，其中，所述骨髓间充质干细胞的制备方法为：取骨髓加等量PBS混匀，用针头沿管壁慢慢注入到1.073g／ml的Pereoll淋巴细胞分离液上，使之成为单细胞悬液，2000r/min离心30min，轻轻吸出中间呈白色浑浊状的骨髓间充质干细胞层，放入另一个离心管中，加入PBS，1500r／min离心15min，如此共洗3次；弃PBS，用含10％FCS的L-DMEM培养液制成的细胞悬液进行计数板计数，以1×109／L的细胞数量接种于含青霉素、链霉素各10000U／100ml、含10％FCS的L-DMEM培养液中，培养48h后第一次换液可见贴壁细胞变成梭形，细胞融合达到85%时传代接种于25 ml 60Co照射照射量5 Gy处理后的细胞培养瓶中，置于5％CO2、37℃二氧化碳培养箱中培养；48 h后首次换液，吸弃未贴壁的悬浮细胞，加入新鲜的含10％FCS的L-DMEM培养液继续孵育，每3～4 d换液；待贴壁细胞80％融合后，弃掉培养液，PBS冲洗瓶壁，0.25％胰酶消化，镜下观察控制2～3 min，用含血清培养液终止消化，反复吹打瓶壁，制成单细胞悬液，按1×109/L再次传代，至第三代时收获细胞，即为骨髓间充质干细胞。
4.如权利要求2所述的化妆品，其特征在于化妆品中还含有湿润剂，剥落剂，润肤剂或水合物。
5.如权利要求4所述的化妆品，其特征在于润肤剂包含植物油中的至少一种。
6.如权利要求4所述的化妆品，其特征在于润肤剂中含有矿物油， 牛油树脂， 可可脂，凡士林， 脂肪酸，甘油三酯，苯甲酸酯，肉豆蔻酸酯，棕榈酸酯，硬脂酸酯，糖脂，磷脂，角鲨烯，甘油，玫瑰果油，ANDIROBA油，葡萄籽油，鳄梨油，李子油，Pracaxi油，蓝藻油，杏仁油，Argan油，荷荷巴油中的一种。
7.负载有多肽的外泌体在制备抗衰老的药物或化妆品中的用途；其中，所述负载多肽的外泌体的制备方法为，在1mL 50mM海藻糖PBS溶液中分别加入外泌体100μg、1mg/mL SEQID NO:1 DYT-4多肽DMSO溶液20μl；混匀后，加入4mm电穿孔皿中，放入电穿孔仪，按照如下反应条件：电压120V，电容155μF，放电时间1ms，放电次数：2次；穿孔后即可将混悬液放入细胞培养箱中孵育1个小时，采用100000g超速离心两次，每次80min，小心弃尽上清，用PBS液重悬沉淀即为负载多肽外泌体；
其中外泌体的制备方法为：将骨髓间充质干细胞使用无血清MSC培养基重悬，接种于底面积25 cm2的培养皿中，接种细胞数1×106个，培养基20 ml；细胞增殖到90%时用无菌PBS溶液清洗细胞表面，重复2次，加入20 ml 添加有5mg/L FGF的α-MEM基础培养基在1%O2、3%CO2的培养条件下培养3 d后收集培养上清进行外泌体提取；将收集到细胞上清分别装于50 ml离心管中，常温3500 rpm，离心25min，去除细胞及其细胞碎片；收集离心后的上清，过0.22μm的滤过膜除去大的颗粒及囊泡，加入终浓度为5%的5×PEG 6000试剂，4 ℃静置过夜；将过夜静置的PEG 6000细胞上清混合液离心，3 000 rpm离心30 min，可见离心管底及壁上有白色絮状沉淀，将离心管上清弃去，并将离心管倒扣于吸水纸上，风干15min，按照1g:200mL的重量/体积比加入适量PBS重悬，再次5000 rpm离心20 min，弃上清，按照1g:100mL用PBS重悬即获得间充质干细胞外泌体。
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108118077A (zh) * | 2016-11-30 | 2018-06-05 | 中南大学 | 一种酶法水解三文鱼胶原制备抗氧化肽与抗冻肽的工艺 |
| CN110195038A (zh) * | 2019-05-08 | 2019-09-03 | 广州市番禺区中心医院(广州市番禺区人民医院、广州市番禺区心血管疾病研究所) | 一种提高间充质干细胞外泌体产量的制备方法 |
| CN111110699A (zh) * | 2020-04-01 | 2020-05-08 | 北京岳昊科技发展有限公司 | 多肽偶联物与表皮干细胞外泌体的组合在药品和化妆品中的应用 |
| CN111700911A (zh) * | 2020-05-09 | 2020-09-25 | 河南省遗传资源细胞库有限公司 | 一种采用干细胞外泌体对皮肤紧致功效的研究方法 |
| CN111840513A (zh) * | 2020-06-12 | 2020-10-30 | 广东工业大学 | 一种负载促肿瘤凋亡蛋白和抗癌小分子的复合外泌体及其制备方法和应用 |
| CN112280734A (zh) * | 2018-05-17 | 2021-01-29 | 广东芙金干细胞再生医学有限公司 | 一种外泌体的制备方法及含有外泌体的干细胞增殖试剂 |
| CN113041208A (zh) * | 2021-04-02 | 2021-06-29 | 北京岳昊科技发展有限公司 | 胚胎干细胞外泌体在制备美白及抗衰老药物或者化妆品中的用途 |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20210186831A1 (en) * | 2019-12-19 | 2021-06-24 | North Carolina State University | Therapeutic compositions and methods using exosomes derived from human dermal fibroblasts |
-
2021
- 2021-08-16 CN CN202110935776.2A patent/CN113559238B/zh active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108118077A (zh) * | 2016-11-30 | 2018-06-05 | 中南大学 | 一种酶法水解三文鱼胶原制备抗氧化肽与抗冻肽的工艺 |
| CN112280734A (zh) * | 2018-05-17 | 2021-01-29 | 广东芙金干细胞再生医学有限公司 | 一种外泌体的制备方法及含有外泌体的干细胞增殖试剂 |
| CN110195038A (zh) * | 2019-05-08 | 2019-09-03 | 广州市番禺区中心医院(广州市番禺区人民医院、广州市番禺区心血管疾病研究所) | 一种提高间充质干细胞外泌体产量的制备方法 |
| CN111110699A (zh) * | 2020-04-01 | 2020-05-08 | 北京岳昊科技发展有限公司 | 多肽偶联物与表皮干细胞外泌体的组合在药品和化妆品中的应用 |
| CN111700911A (zh) * | 2020-05-09 | 2020-09-25 | 河南省遗传资源细胞库有限公司 | 一种采用干细胞外泌体对皮肤紧致功效的研究方法 |
| CN111840513A (zh) * | 2020-06-12 | 2020-10-30 | 广东工业大学 | 一种负载促肿瘤凋亡蛋白和抗癌小分子的复合外泌体及其制备方法和应用 |
| CN113041208A (zh) * | 2021-04-02 | 2021-06-29 | 北京岳昊科技发展有限公司 | 胚胎干细胞外泌体在制备美白及抗衰老药物或者化妆品中的用途 |
Non-Patent Citations (4)
| Title |
|---|
| PEG 6000快速提取间充质干细胞外泌体;梁亚会等;《空军医学杂志》;20171231;第33卷(第3期);第76-79页 * |
| Preparation of Antioxidant Peptides from Salmon Byproducts with Bacterial Extracellular Proteases;Ribang Wu et al;《Mar Drugs.》;20170131;第15卷(第1期);第1-19页 * |
| 三文鱼胶原肽体外抗氧化活性研究;刘文颖等;《食品科技》;20101231;第35卷(第12期);第86-89页 * |
| 鲑鱼鱼皮胶原蛋白小分子肽的功效性研究;孔惠等;《第十一届中国化妆品学术研讨会论文集》;20161231;第89-94页 * |
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