CN116831983A - 一种浓缩生长因子复合凝胶制剂、制备方法及生发应用 - Google Patents
一种浓缩生长因子复合凝胶制剂、制备方法及生发应用 Download PDFInfo
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- CN116831983A CN116831983A CN202311124913.XA CN202311124913A CN116831983A CN 116831983 A CN116831983 A CN 116831983A CN 202311124913 A CN202311124913 A CN 202311124913A CN 116831983 A CN116831983 A CN 116831983A
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- cgf
- growth factor
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- sodium hyaluronate
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Abstract
本发明属生物医用材料技术领域,特别是涉及一种富含自体浓缩生长因子的复合凝胶制剂、制备方法及生发应用,用肝素抗凝管采集自体静脉新鲜外周血,然后将采集的自体静脉新鲜外周血放置在梯度离心机中,使用梯度变速离心法将自体抗凝外周血离心,然后分离上层血浆,取上层血浆的下1/3,获得CGF溶液,向CGF溶液中加入卡波姆凝胶基质液和透明质酸钠溶液,形成CGF凝胶。本发明提供CGF凝胶来源于自体外周血,安全性好,并且可以冷藏长期保存,性价比更高,能够通过外敷的方式进行局部的生发治疗,该CGF凝胶吸水能力强、保湿,并且能够达到缓慢释放和吸收,联合CGF溶液皮下注射的生发治疗效果更好。
Description
技术领域
本发明属于生物医用材料技术领域,特别是涉及一种富含自体浓缩生长因子的复合凝胶制剂、制备方法及生发应用。
背景技术
脱发(Hair loss)是指头发脱落的现象,正常脱落的头发都是处于退行期及休止期的毛发,由于进入退行期与新进入生长期的毛发不断处于动态平衡,故能维持正常数量的头发。然而,快节奏的生活,巨大的精神压力,却使得越来越多的人面临脱发的困扰。脱发不仅仅影响外观,对人的自信心也是一个十分大的打击。脱发治疗已成为越来越多的人迫切需要解决的问题。
目前,对于脱发的治疗手段多种多样,随着对浓缩生长因子(ConcentratedGrowth Factor, CGF)的研究,人们发现CGF在口颌面部软硬组织损伤修复领域的应用,随着基础研究不断深入和临床试验的不断开展,CGF在牙种植、骨缺损、整形美容、创面修复等领域展现出独特的优势和良好的应用市场。现在大多使用CGF液体制剂治疗脱发,治疗效果不好。
发明内容
本发明的目的在于提供一种浓缩生长因子复合凝胶制剂、制备方法及生发应用,以解决上述背景技术中提出的问题。
为达到上述技术目的,本发明的技术方案:
一种浓缩生长因子复合凝胶制剂,所述复合凝胶制剂由以下原料组成:CGF溶液、透明质酸钠溶液和卡波姆凝胶基质液,所述CGF溶液包括转化生长因子-β、血小板源性生长因子、血管内皮生长因子、表皮生长因子、成纤维细胞生长因子和CD34+造血干细胞;
所述透明质酸钠溶液为低分子透明质酸钠的生理盐水溶液,所述低分子透明质酸钠的浓度为130g/L;
所述卡波姆凝胶基质液为卡波姆940溶解于生理盐水中,经搅拌、振荡、加热混合均匀后制成,所述卡波姆940的质量浓度为2%;
所述CGF溶液与卡波姆凝胶基质液的体积比为1:1,所述复合凝胶制剂中透明质酸钠的质量浓度为1.3g/L。
进一步地,所述CGF溶液为采集自体静脉新鲜外周血并梯度变速离心,然后分离上层血浆的下1/3部分制得。
本发明还提供浓缩生长因子复合凝胶制剂的制备方法,包括以下步骤:
S1:CGF溶液的提取
用肝素抗凝管采集自体静脉新鲜外周血并梯度变速离心,然后分离上层血浆的下1/3部分,获得CGF溶液;
S2:CGF复合凝胶体系配制
向步骤S1的CGF溶液中加入卡波姆凝胶基质液和透明质酸钠溶液,混合均匀后形成CGF复合凝胶体系,将CGF复合凝胶体系置于4℃冷藏存储。
进一步地,所述步骤S2中,所述透明质酸钠溶液的配制步骤如下:
取低分子透明质酸钠粉末,用生理盐水溶解,通过滤器过滤除菌,配制成130g/L的无菌透明质酸钠溶液,将无菌透明质酸钠溶液于4℃储存。
进一步地,所述步骤S2中,所述卡波姆凝胶基质液的配制步骤如下:
取卡波姆940,加入生理盐水溶解,边加边搅拌,然后放入摇床振荡器中,60℃加热2h,混合均匀后,制成质量浓度2%的卡波姆凝胶基质液,将卡波姆凝胶基质液进行灭菌,冷却后于4℃储存。
进一步地,所述透明质酸钠溶液使用0.2µm的滤器过滤除菌。
进一步地,所述透明质酸钠溶液和卡波姆凝胶基质液配制过程中均使用生理盐水,所述生理盐水的质量浓度为0.9%,配制过程中所使用的配制器具在使用前均进行高压蒸汽灭菌,使用前进行紫外线灭菌30min,在超净台内进行配制。
进一步地,步骤S1中,按照脱发面积占整个头部毛发附着部位的1/3-1比例,采集20-60ml外周血。
本发明还提供浓缩生长因子复合凝胶制剂的生发应用,所述生发应用为浓缩生长因子复合凝胶制剂与CGF皮下注射液的联合应用。
由于采用上述技术方案,本发明的有益效果:
本发明提供的一种浓缩生长因子复合凝胶制剂,作用于损伤的皮肤组织、细胞,发挥局部抗炎和免疫调节作用,促进其修复和毛发再生,并且CGF凝胶中存在高浓度的CD34+造血干细胞,对细胞代谢、生长、修复以及毛细血管增生等起到至关重要的作用,相比现有的CGF液体制剂,适用于外敷,能够通过外敷的方式进行局部的生发治疗,该CGF凝胶吸水能力强、保湿,并且能够促进浓缩生长因子缓慢深入皮肤的表皮层,能够达到缓慢释放和吸收,治疗效果更好。
本发明提供的浓缩生长因子复合凝胶制剂,原料种类少,制备方法简单,一次采血,多次制备,并且可以冷藏长期保存,性价比更高,经济实惠,可减轻患者的经济负担。
本发明将自体血来源的CGF溶液利用物理改性制剂卡波姆940和水溶性低分子透明质酸钠制备成CGF凝胶,CGF来自于自体血液,安全性更高。
本发明制备的CGF凝胶以透明质酸钠为生物活性基质,透明质酸钠作为人体天然成分,易于吸收,无毒、无刺激性、且生物相容性良好。低分子透明质酸钠作为活性分子,能够促进血液微循环、调节皮肤代谢、改善局部营养、加快皮肤损伤修复,并且能够促进CGF渗透到真皮层,进一步提高了CGF凝胶的治疗效果,透明质酸钠制剂在皮肤表层形成黏多糖样网状结构,CGF活性组分可穿过网格结构,缓慢吸收进入皮肤表层,直至真皮层,具有缓释CGF活性组分的作用。此外,由于透明质酸钠作为抗衰、美容的主流药物,与生物制剂CGF一起给药,可协同发挥生物学作用,低分子透明质酸钠辅助CGF更好地渗透、缓慢吸收进入皮肤表层,直至真皮层,并且,透明质酸钠形成的网状结构,能够实现CGF活性组分的缓慢释放,进而协同促进局部血液微循环、调节皮肤代谢、改善局部营养,加快皮肤组织损伤修复和毛发再生,真正达到1+1>2的效果。
本发明中低分子透明质酸钠及卡波姆940有效融合,卡波姆940作为流变学改性剂,粘附在透明质酸钠形成的网状结构上,能够使CGF在局部皮肤形成一层透明膜,更加有效促进CGF缓慢吸收到达皮肤真皮层,达到促进皮肤组织损伤修复和毛发再生的治疗效果,用药一次能在皮肤局部停留更长时间,长效发挥作用,故本品每日使用次数少,每日使用2次即可达到较好的治疗效果。
本复合外用凝胶制剂不含苯扎溴铵等防腐剂,对皮肤无刺激、损伤作用,自体血液来源,更安全,且使用方便,患者用药依从性高。
本复合外用凝胶联合CGF皮下注射治疗,患者用药依从性好,临床生发治疗效果更佳。
附图说明
为了更清楚地说明本发明实施例或现有技术中的技术方案,下面将对实施例中所需要使用的附图作简单地介绍,显而易见地,下面描述中的附图仅仅是本发明的一些实施例,对于本领域普通技术人员来讲,在不付出创造性劳动的前提下,还可以根据这些附图获得其他的附图。
图1是自体浓缩生长因子复合凝胶制剂的制备流程图;
图2是CGF抑制巨噬细胞炎症性细胞因子IL-6、TNF-α的效果图;
图3是复合CGF凝胶联合CGF溶液皮下注射治疗全脱发病人的治疗效果图;
图4是单独使用CGF溶液皮下注射治疗全脱发病人的治疗效果图。
具体实施方式
现详细说明本发明的多种示例性实施方式,该详细说明不应认为是对本发明的限制,而应理解为是对本发明的某些方面、特性和实施方案的更详细的描述。
应理解本发明中所述的术语仅仅是为描述特别的实施方式,并非用于限制本发明。另外,对于本发明中的数值范围,应理解为还具体公开了该范围的上限和下限之间的每个中间值。在任何陈述值或陈述范围内的中间值,以及任何其他陈述值或在所述范围内的中间值之间的每个较小的范围也包括在本发明内。这些较小范围的上限和下限可独立地包括或排除在范围内。
除非另有说明,否则本文使用的所有技术和科学术语具有本发明所述领域的常规技术人员通常理解的相同含义。虽然本发明仅描述了优选的方法和材料,但是在本发明的实施或测试中也可以使用与本文所述相似或等同的任何方法和材料。本说明书中提到的所有文献通过引用并入,用以公开和描述与所述文献相关的方法和/或材料。在与任何并入的文献冲突时,以本说明书的内容为准。
在不背离本发明的范围或精神的情况下,可对本发明说明书的具体实施方式做多种改进和变化,这对本领域技术人员而言是显而易见的。由本发明的说明书得到的其他实施方式对技术人员而言是显而易见得的。本发明说明书和实施例仅是示例性的。
关于本文中所使用的“包含”、“包括”、“具有”、“含有”等等,均为开放性的用语,即意指包含但不限于。
实施例1
一种浓缩生长因子复合凝胶制剂,所述复合凝胶制剂由以下原料组成:CGF溶液、透明质酸钠溶液和卡波姆凝胶基质液,所述CGF溶液采集自体静脉新鲜外周血并梯度变速离心,然后分离上层血浆的下1/3部分制得;
透明质酸钠溶液为低分子透明质酸钠的生理盐水溶液,低分子透明质酸钠的浓度为130g/L;
卡波姆凝胶基质液为卡波姆940溶解于生理盐水中,经搅拌、振荡、加热混合均匀后制成,卡波姆940的质量浓度为2%;
CGF溶液与卡波姆凝胶基质液的体积比为1:1,复合凝胶制剂中透明质酸钠的质量浓度为1.3g/L。
具体地,CGF溶液包括转化生长因子-β、血小板源性生长因子、血管内皮生长因子、表皮生长因子、成纤维细胞生长因子和CD34+造血干细胞。
实施例2
浓缩生长因子复合凝胶制剂的制备方法,包括以下步骤:
S1:透明质酸钠溶液的配制:取低分子透明质酸钠粉末13g,用0.9%的生理盐水溶解,定容至100ml,通过0.2µm的滤器过滤除菌,配制成130g/L的透明质酸钠溶液,将透明质酸钠溶液于4℃储存;
S2:卡波姆凝胶基质液的配制:取卡波姆940 2g,加入0.9%、100ml的生理盐水中,边加边搅拌,然后放入摇床振动器中,60℃加热2h,混合均匀后,制成2%的卡波姆凝胶基质液,将卡波姆凝胶基质液进行高压蒸汽 灭菌,冷却后于4℃储存;
S3:CGF溶液的提取:按照患者脱发面积占整个头部毛发附着部位的1/3-1比例,采集20-60ml外周血,根据具体患者脱发情况,本实施例采集48ml外周血,用肝素抗凝管采集自体静脉新鲜外周血48ml,然后将采集的自体静脉新鲜外周血放置在梯度离心机中,使用梯度变速离心法将自体抗凝外周血离心,然后分离上层血浆,取上层血浆的下1/3,获得8ml的CGF溶液;
S4:将锥形瓶和搅拌棒进行高压蒸汽灭菌备用,向锥形瓶中加入4ml上述CGF溶液、4ml于4℃储存的卡波姆凝胶基质液和80uL于4℃储存的透明质酸钠溶液,用搅拌棒搅拌混合均匀,形成CGF复合凝胶,将CGF复合凝胶置于4℃冷藏存储。
实施例2中的浓缩生长因子复合凝胶制剂的制备方法,在实践中根据每个人的实际情况,即根据患者的脱发面积不同,核定采血量。
CGF抑制巨噬细胞炎症性细胞因子IL-6、TNF-α的实验研究,实验方法如下:
分别用无血清培养基、IFN-γ+LPS共刺激、IL-4刺激、含有10%CGF和含有20%CGF的细胞培养基,与THP-1巨噬细胞共同孵育12h、24h、48h,其中无血清培养基、IFN-γ+LPS共刺激、IL-4刺激、含有10%CGF和含有20%CGF的细胞培养基分别标记为M0组、M1组、M2组、10%CGF+M0组和20%CGF+M0组,然后用流式细胞仪检测培养上清中细胞因子IL-6、TNF-α的生成情况。
实验结果如图2所示,与IFN-γ+LPS刺激的M1组和IL-4刺激的M2组相比,10%和20%的自体CGF在体外刺激巨噬细胞时,均能够明显抑制细胞培养上清中炎性因子IL-6、TNF-α的产生,进而抑制炎症的产生,对脱发具有治疗效果。其中,CGF对IL-6的抑制作用在24h时最明显,对TNF-α的抑制作用在12h时最显著。
本发明的典型病例如下:
病例1:患者姜某,女性,15岁,全脱发患者,患者头皮皮下多点注射自体CGF溶液,每月一次,然后注射后进行外敷实施例1中的CGF凝胶,每天一次,每天外敷30min以上,如图3所示,该患者在治疗30天后,头发明显再生,但相对较稀疏,在治疗60天后,头发达到正常人的发量,治疗过程中无不良反应。
病例2:患者,男性,28岁,全脱发患者,患者头皮皮下多点注射自体CGF溶液,每月一次,如图4所示,该患者在治疗30天后,头发有再生,但仍很稀疏,在治疗60天后,头发明显再生,但仍未达到正常人的发量,治疗过程中未发现不良反应。
CGF溶液注射联合CGF复合凝胶制剂外敷治疗,其生发效果明显优于单独应用CGF溶液皮下注射治疗方法。
以上所述本发明的具体实施方式,并不构成对本发明保护范围的限定。任何根据本发明的技术构思所做出的各种其他相应的改变与变形,均应包含在本发明权利要求的保护范围内。
Claims (9)
1.一种浓缩生长因子复合凝胶制剂,其特征在于,所述复合凝胶制剂由以下原料组成:CGF溶液、透明质酸钠溶液和卡波姆凝胶基质液,所述CGF溶液包括转化生长因子-β、血小板源性生长因子、血管内皮生长因子、表皮生长因子、成纤维细胞生长因子和CD34+造血干细胞;
所述透明质酸钠溶液为低分子透明质酸钠的生理盐水溶液,所述低分子透明质酸钠的浓度为130g/L;
所述卡波姆凝胶基质液为卡波姆940溶解于生理盐水中,经搅拌、振荡、加热混合均匀后制成,所述卡波姆940的质量浓度为2%;
所述CGF溶液与卡波姆凝胶基质液的体积比为1:1,所述复合凝胶制剂中透明质酸钠的质量浓度为1.3g/L。
2.根据权利要求1所述的浓缩生长因子复合凝胶制剂,其特征在于,所述CGF溶液为采集自体静脉新鲜外周血并梯度变速离心,然后分离上层血浆的下1/3部分制得。
3.权利要求1或2所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,包括以下步骤:
S1:CGF溶液的提取
用肝素抗凝管采集自体静脉新鲜外周血并梯度变速离心,然后分离上层血浆的下1/3部分,获得CGF溶液;
S2:CGF复合凝胶体系配制
向步骤S1的CGF溶液中加入卡波姆凝胶基质液和透明质酸钠溶液,混合均匀后形成CGF复合凝胶体系,将CGF复合凝胶体系置于4℃冷藏存储。
4.根据权利要求3所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,所述步骤S2中,所述透明质酸钠溶液的配制步骤如下:
取低分子透明质酸钠粉末,用生理盐水溶解,通过滤器过滤除菌,配制成130g/L的无菌透明质酸钠溶液,将无菌透明质酸钠溶液于4℃储存。
5.根据权利要求3所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,所述步骤S2中,所述卡波姆凝胶基质液的配制步骤如下:
取卡波姆940,加入生理盐水溶解,边加边搅拌,然后放入摇床振荡器中,60℃加热2h,混合均匀后,制成质量浓度2%的卡波姆凝胶基质液,将卡波姆凝胶基质液进行灭菌,冷却后于4℃储存。
6.根据权利要求4所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,所述透明质酸钠溶液使用0.2µm的滤器过滤除菌。
7.根据权利要求3所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,所述透明质酸钠溶液和卡波姆凝胶基质液配制过程中均使用生理盐水,所述生理盐水的质量浓度为0.9%,配制过程中所使用的配制器具在使用前均进行高压蒸汽灭菌,使用前进行紫外线灭菌30min,在超净台内进行配制。
8.根据权利要求3所述的浓缩生长因子复合凝胶制剂的制备方法,其特征在于,步骤S1中,按照脱发面积占整个头部毛发附着部位的1/3-1比例,采集20-60ml外周血。
9.权利要求1所述的浓缩生长因子复合凝胶制剂的生发应用,其特征在于,所述生发应用为浓缩生长因子复合凝胶制剂与CGF皮下注射液的联合应用。
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