Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors.
Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors.
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DOI：
10.1186/scrt218
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发表时间：
2013-06-07
影响因子：
7.5
通讯作者：
Borojevic R
中科院分区：
文献类型：
--
作者：
Amable PR;Carias RB;Teixeira MV;da Cruz Pacheco I;Corrêa do Amaral RJ;Granjeiro JM;Borojevic R
富血小板血浆（PRP）作为促进组织再生的生长因子库，目前已广泛应用于骨科、眼科和愈合治疗等不同临床场景。对其临床疗效的研究尚无定论，主要原因之一是使用不同的PRP制剂，引起的反应不同，无法进行比较。血小板定量和生长因子含量的定义必须明确，以便了解PRP再生强度背后的分子机制。因此，迫切需要PRP制剂的标准化。通过改变相对离心力、温度和时间来离心制备PRP。在对血小板回收率和产率进行量化后，选择了产量最高的两步法并进行了进一步分析。细胞因子的含量测定在整个离心过程中获得的不同部分。我们的方法在应用于不同的献血者时具有可重复性。我们恢复了46.9 ~ 69.5%的总初始血小板，该过程导致血小板浓度增加5.4 ~ 7.3倍（1.4 × 106 ~ 1.9 × 106血小板/μl）。血小板是高度纯化的，因为在最终的PRP制备中只有<0.3%的红细胞和白细胞存在。我们还量化了经钙和钙/凝血酶活化的浓缩血小板分泌的生长因子、细胞因子和趋化因子。分泌高浓度的血小板源性生长因子、内皮生长因子、转化生长因子（TGF），同时分泌抗炎、促炎细胞因子白介素(IL)-4、IL-8、IL-13、IL-17、肿瘤坏死因子(TNF)-α、干扰素(IFN)-α。血小板活化前无细胞因子分泌。未检测到TGF-β3和IFNγ。血小板凝固后的凝块保留了高浓度的几种生长因子，包括血小板源性生长因子和TGF。我们的研究产生了一种一致的PRP制备方法，该方法从不同的供体中获得细胞因子和生长因子库，具有高重复性。这些发现支持了PRP在组织再生治疗中的应用，其内容表征将使我们能够理解和改善临床结果。
Platelet-rich plasma (PRP) is nowadays widely applied in different clinical scenarios, such as orthopedics, ophthalmology and healing therapies, as a growth factor pool for improving tissue regeneration. Studies into its clinical efficiency are not conclusive and one of the main reasons for this is that different PRP preparations are used, eliciting different responses that cannot be compared. Platelet quantification and the growth factor content definition must be defined in order to understand molecular mechanisms behind PRP regenerative strength. Standardization of PRP preparations is thus urgently needed. PRP was prepared by centrifugation varying the relative centrifugal force, temperature, and time. Having quantified platelet recovery and yield, the two-step procedure that rendered the highest output was chosen and further analyzed. Cytokine content was determined in different fractions obtained throughout the whole centrifugation procedure. Our method showed reproducibility when applied to different blood donors. We recovered 46.9 to 69.5% of total initial platelets and the procedure resulted in a 5.4-fold to 7.3-fold increase in platelet concentration (1.4 × 106 to 1.9 × 106 platelets/μl). Platelets were highly purified, because only <0.3% from the initial red blood cells and leukocytes was present in the final PRP preparation. We also quantified growth factors, cytokines and chemokines secreted by the concentrated platelets after activation with calcium and calcium/thrombin. High concentrations of platelet-derived growth factor, endothelial growth factor and transforming growth factor (TGF) were secreted, together with the anti-inflammatory and proinflammatory cytokines interleukin (IL)-4, IL-8, IL-13, IL-17, tumor necrosis factor (TNF)-α and interferon (IFN)-α. No cytokines were secreted before platelet activation. TGF-β3 and IFNγ were not detected in any studied fraction. Clots obtained after platelet coagulation retained a high concentration of several growth factors, including platelet-derived growth factor and TGF. Our study resulted in a consistent PRP preparation method that yielded a cytokine and growth factor pool from different donors with high reproducibility. These findings support the use of PRP in therapies aiming for tissue regeneration, and its content characterization will allow us to understand and improve the clinical outcomes.
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DOI：
10.3343/kjlm.2011.31.3.212
发表时间：
2011-07
期刊：
The Korean journal of laboratory medicine
影响因子：
--
作者：
Cho HS;Song IH;Park SY;Sung MC;Ahn MW;Song KE
通讯作者：
Song KE
影响因子：
4.4
作者：
Freire, Vanesa;Andollo, Noelia;Morales, Maria-Celia
通讯作者：
Morales, Maria-Celia
影响因子：
4.8
作者：
Jo, Chris Hyunchul;Kim, Ji Eun;Shin, Sue
通讯作者：
Shin, Sue
影响因子：
3.6
作者：
Kakudo, Natsuko;Minakata, Tatsuya;Kusumoto, Kenji
通讯作者：
Kusumoto, Kenji
影响因子：
4.8
作者：
de Mos, Marieke;van der Windt, Anna E.;van Osch, Gerjo J. V. M.
通讯作者：
van Osch, Gerjo J. V. M.