Chinese Journal of Tissue Engineering Research ›› 2018, Vol. 22 ›› Issue (20): 3263-3266.doi: 10.3969/j.issn.2095-4344.0806
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Zhang Tian-jia, Dai Jie-wen, Wang Xu-dong
Received:
2017-11-15
Online:
2018-07-18
Published:
2018-07-18
Contact:
Wang Xu-dong, M.D., Doctoral supervisor, Professor, Chief physician, Department of Oral and Craniomaxillofacial Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai 200011, China
About author:
Zhang Tian-jia, Studying for master’s degree, Department of Oral and Craniomaxillofacial Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai 200011, China
Supported by:
the Shanghai Three-Year Action Plan of Promoting the Clinical Skill and Innovation Abilities of Municipal Hospitals, No. 16CR3019A; the Plan for Promoting Clinical Research of Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, No. JYLJ006; the Medical-Engineering Cross Foundation of Shanghai Jiao Tong University in 2017, No. YG2017ZD03
CLC Number:
Zhang Tian-jia, Dai Jie-wen, Wang Xu-dong. Application of concentrated platelet products in autologous fat grafting: lack of quantitative indicators and application guidelines[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(20): 3263-3266.
2.1 血小板浓缩制品的定义与发展 2.1.1 富血小板血浆 最早是由 Assoian于1984年提取制备的,是自身静脉血经梯度离心分层后位于白细胞下方的富含血小板的血浆部分,含有全血中70%以上的血小板[13]。Marx[14]给富血板血浆定义为含有比自身正常值更高浓度血小板的少量血浆。正常成年人的血浆血小板浓度一般为(7.5-20)×1010 L-1,而在富血小板血浆中,血小板的浓度可高达10×1011 L-1,为正常浓度的4-7 倍[15]。血小板在制备富血小板血浆过程中激活的α颗粒可以释放出多种生长因子,包括:转化生长因子β、骨形成蛋白、血小板衍生生长因子、类胰岛素生长因子、血管内皮生长因子、表皮生长因子和成纤维细胞生长因子等[16-18]。血小板中另一种颗粒成分致密体则为富血小板血浆提供了血清素、组胺、多巴胺、钙离子和腺苷[19]。富血小板血浆的制备过程相对较为复杂,需要在手术开始前采集静脉血以减少因手术造成的血小板凝集[20]。之后需要对静脉血进行2次离心:第1次使用较慢的速度离心,分离后弃去最下方的血细胞成分;保留上方的血浆及中间的富血小板层后,进行第2次速度较快的离心,并最终获得由沉淀的血小板及部分血浆组成的富血小板血浆。但制备富血小板血浆所需要的离心参数及步骤并没有明确的规定。近年来有学者提出了富血小板血浆的制备指南,其中提及到在2次离心过程中,应尽量增加转速并减少离心时间以获得富含更多血小板的富血小板血浆[21-22]。在得到富血小板血浆之后,还需要对其进行活化来达到释放生长因子的目的。最为常用的方法是在富血小板血浆中添加氯化钙溶液、凝血酶成分后通入气泡使其激活[23]。目前为止依然没有一种公认的富血小板血浆制备方法,因而也导致了其在成分和治疗效果上的不稳定性[24]。且激活富血小板血浆过程中添加的外源性物质也在一定程度上增加了产生其他并发症的风险[25]。 2.1.2 富血小板纤维蛋白 Dohan等[26]及Choukroun等[27]在2000年制备了富血小板纤维蛋白,并主要应用于口腔颌面外科手术。相比富血小板血浆,富血小板纤维蛋白因不需要加入抗凝或凝血制品而具有较为明显的优势,且制备过程相对简单:取患者10 mL外周血液后,加入离心管中以3 000 r/min的速度离心10 min后即可在贫血小板血浆层与红细胞碎片层之间获取富含血小板的纤维蛋白凝胶。富血小板纤维蛋白的纤维蛋白作为一种基质为细胞的附着,迁移以及分化提供了有利的微环境,浓缩的血小板在立体网状结构中被活化,并释放出多种生长因子:转化生长因子,血小板衍生生长因子,血管内皮生长因子以及胰岛素样生长因子,这些生长因子可有效调控成骨细胞、脂肪干细胞以及成纤维细胞等与组织修复紧密相关细胞的增殖、分化以及凋亡[28-30]。因为缺少额外添加的抗凝成分,血液在接触离心管壁的瞬间就开始发生凝集的现象,且纤维蛋白原开始发生缓慢的聚合,形成三分子的立体网状结构。富血小板纤维蛋白在离心完成的同时也活化了血小板颗粒并释放出了上述生长因子,但得益于其特有的结构,生长因子并不会因此而失去活性,因此得到了许多医生的青睐[31-32]。 2.1.3 浓缩生长因子 2006年有学者提出了新一代血小板浓缩制品浓缩生长因子的概念,与富血小板纤维蛋白相似,浓缩生长因子由静脉血分离制备而成[33]。但浓缩生长因子的制备相比富血小板纤维蛋白较为复杂,且需要使用专用的Medifuge离心加速机,充分激活血小板α颗粒,释放出更多的各类生长因子。经过12 min的差速离心后,可见离心管中分为3层,上层为血清层,中间为纤维蛋白凝集层以及最下方的红细胞及血小板层。浓缩生长因子中的纤维蛋白凝胶是由大小不等的纤维蛋白形成的一种三维网状立体结构,其中镶嵌着较多数量的白细胞,表明其具有良好的抗感染和抗炎症的能力,在浓缩生长因子中存在着大量的CD34+细胞、血管内皮生长因子和成纤维细胞生长因子等活性物质,其在免疫调节、血管再生、血管新生等方面具有着重要的作用[34] (表1)。 2.2 富血小板血浆辅助脂肪移植的基础研究及临床应用 富血小板血浆辅助自体脂肪移植在一些体内外研究及临床试验中取得了良好的结果,归因于:①为脂肪细胞提供了血清基质中的营养;②促进了脂肪组织中的血管再生;③促进了脂肪干细胞及前体细胞向成熟脂肪细胞的分化。 Por等[35]使用人脂肪与未经活化的人体富血小板血浆按4∶1的比例混合后注射至裸鼠头皮皮下,在4个月后对存留脂肪进行切片观察,结果显示富血小板血浆组并没有提高血管再生率;Dong等[36]则使用了氯化钙和凝血酶活化后的富血小板血浆与人体脂肪以2∶7的比例混合,注射至裸鼠头皮皮下,10周后观察到了更多的血管生成以及更少的纤维性变; Nakamura等[37]采集了大鼠的腹股沟脂肪及氯化钙活化的富血小板血浆以4∶1的比例通过手术而非注射的方式植入异体大鼠的背部皮下,在4个月后观察到了更多的血管生成和脂肪细胞存活;Fraga等[38]将活化的富血小板血浆与脂肪以1∶1的比例注射至兔子耳部皮下,在6个月后观察到了更多的血管生成;Rodriguez-Flores等[39]采用相似的方法,将富血小板血浆与脂肪注射至兔子唇部,3个月后观察到了更轻的局部炎症反应和更少的破碎脂肪与更高的脂肪成活率和完整性。在动物实验的结果中,富血小板血浆整体上是促进了自体或同种异体脂肪移植的效果。但"
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