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标题: 德国不反对临床用MSC采用含有FBS的培养基，但需适当控制量 [打印本页]

作者: xylophone 时间: 2012-2-12 17:23 标题: 德国不反对临床用MSC采用含有FBS的培养基，但需适当控制量

回复编委：采用牛血清用于细胞培养是安全的，
Hum Gene Ther. 2011 Jun;22(6):775; author reply 776.
Safety of bovine sera for production of mesenchymal stem cells for therapeutic use
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To the Editor:: W& z" i4 I% b% n
IN THE PUBLICATION “Good Manufacturing Practices Production of Mesenchymal Stem/Stromal Cells” by Luc Sensebé, Philippe Bourin, and Karin Tarte, published in the 2011 January issue of Human Gene Therapy (Sensebé et al., 2011), the statement is made that the Paul-Ehrlich-Institut (PEI) in Germany is prohibiting the use of fetal bovine serum (FBS) for the GMP-compliant production of mesenchymal stem cells (MSCs). This is not correct. Use of adequately controlled bovine sera is allowed.
Experience from production of vaccines and other biotechnical medicinal products has shown that bovine sera may be a source of virus contamination. Therefore, the PEI discourages the use of FBS when possible, and welcomes the development of serum-free media. However, it is well recognized that many cell culture systems require bovine sera and the use of adequately controlled bovine sera is allowed. Basic requirements for the quality of bovine sera used in the manufacture of medicinal products have been outlined in the European Pharmacopoeia (European Pharmacopoeia. 7th Ed. Monograph 01/2008:2262 Bovine Serum. Council of Europe, Strasbourg). This includes the aspects of virus testing and virus inactivation. A process for virus inactivation (e.g., γ radiation at a dose of ≥30 kGy) should be applied unless otherwise justified. When the use of noninactivated serum is essential, a detailed demonstration of the need for such sera is expected and, in addition, virus safety measures should be discussed. Another safety aspect that needs to be addressed concerns transmissible spongiform encephalopathies (TSEs). The safety of bovine sera with respect to TSE can be conveniently demonstrated by providing a certificate from the European Directorate for the Quality of Medicines & HealthCare (EDQM, Strasbourg, France). A list of TSE-certified animal sera can be found online in the EDQM database of certified substances (www.edqm.eu/site/Databases-10.html). Unfortunately, there is no equivalent certification system for virus safety. Therefore, all information regarding virus safety of bovine sera must be provided by the bovine serum supplier.
In addition, we wish to point out that heterologous human-derived substances, such as human AB serum or platelet lysates, cannot be regarded as “more secure” with respect to virus safety than bovine sera. These human-derived substances are usually not subjected to virus inactivation. Despite comprehensive selection and testing of blood donors for specific viruses according to the official regulations for blood donation, there is always an underlying residual risk of contamination with human pathogenic viruses.4 a+ w+ t$ c( d$ c( d* p
Furthermore, the development of defined serum-free media based on cocktails of growth factors and other substances such as fibronectin is welcomed with respect to pathogen safety as well as avoidance of potential problems related to batch-to-batch inconsistency from human or animal sera. Commercial availability of such media is not a regulatory requirement. Such media can be used in clinical trials or for the production of medicinal products, provided that adequate quality and safety can be demonstrated for each component

作者: liuzongcai800 时间: 2012-2-13 09:00

“临床用MSCs制备中使用血清的安全性”

作者: uslzhang 时间: 2012-2-13 10:24

使用platelet lysates规模化培养细胞存在资源有限的问题。

作者: youndie 时间: 2012-2-13 16:07

platelet lysates最大的问题在于异体来源

作者: xylophone 时间: 2012-2-15 14:00

采用血小板裂解液应该是一个不错的选择，与常规采用FCS培养MSC 同样都会涉及到异源物质的问题，5 r4 c$ @, X- Z. r$ R
异源物质引入的安全问题虽然还在争论，但是如果能够获得足够纯的血小板裂解液也将会是一个培养MSC不错途径' o, ~3 j' H/ |
异源的距离也能由动物拉近到人类了！
附相关报道：《南方医科大学学报》 2011年08期加入收藏获取最新

人血小板裂解液替代胎牛血清促进骨髓间质干细胞的增殖许茹夏文杰戎霞叶欣邵媛王敏罗广平付涌水
【摘要】：目的探讨人血小板裂解液(human platelet lysates,HPL)对骨髓间质干细胞(mesenchymal stem cells,MSCs)增殖以及生物学特性的影响。方法通过对机采血小板反复冻融获得HPL,采用密度梯度离心法从骨髓中分离MSCs,分别用10%胎牛血清(feotal calf serum,FCS)和LD-DMEM添加不同浓度的HPL进行培养,以确定最佳HPL浓度。分离6株人源MSCs,从原代开始分别用添加胎牛血清和最佳HPL浓度的LD-DMEM进行培养,对两种培养体系下6株不同来源的MSCs的增殖能力,表面标记表达量以及细胞周期进行研究,并利用t-test进行统计学分析,P0.05视为有统计学差异。结果研究发现HPL中含有MSCs生长所必需的4种生长因子,PDGF-AB(人血小板衍生生长因子AB),bFGF(成纤维细胞生长因子),IGF-1(胰岛素样生长因子1),TGF-β1(转化生长因子β1),浓度分别为

0.53±0.06)ng/ml,(37.5±4.31)pg/ml,(0.15±0.06)mg/ml,(5150±463)pg/ml。适合MSCs增殖的最佳HPL浓度为7.5%。两种培养条件下的细胞形态无明显区别,均为长梭形。8代以前,细胞增殖能力没有显著性差异(P0.05),两种培养条件下的MSCs均表达CD105,CD106以及粘附分子CD29和CD44,不表达CD34,CD45,且其表达量没有显著性差异(P0.05)。两种培养体系下绝大多数细胞处于G0-G1期,FCS培养的MSCs其G0-G1期占(96.97±0.95)%,7.5%的HPL培养的MSCs其G0-G1期占(94.58±1.56)%,两种培养条件下不同周期没有显著性差异(P0.05)。结论本研究适合MSCs增殖的最佳HPL浓度为7.5%,HPL可以替代胎牛血清体外培养扩增人骨髓间质干细胞,并保持其生物学特性基本不变。7 x& |. k- ^4 w# L; Z/ I
【作者单位】：广州血液中心输血研究所;
【关键词】：骨髓间质干细胞血小板裂解液增殖表面标记细胞周期 : c' r2 f4 I9 N
【基金】：广州市医药卫生科技项目(2008-Zdi-10)

作者: Eul_er 时间: 2012-2-25 11:09

现在无血清培养已经很成熟了。就安全性而言，还是该提倡无血清培养。

作者: uslzhang 时间: 2013-5-21 11:00

楼上“现在无血清培养已经很成熟了。就安全性而言，还是该提倡无血清培养。”的结论从何而来？无血清培养基都是人工配制的，毫无“自然”可言，无法知道培养过程中细胞发生了什么变化，只能从表面看到那几个细胞指标“没有发生变化”，实际上已经报道了无血清培养基与牛血清培养的干细胞存在许多不同点，实质上未知的太多。牛血清至少是天然来源的。

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