中国富血小板血浆临床制备方法的研究进展

自体富血小板血浆(PRP)是一种通过离心的方法从自体血中提取出来的血小板浓缩物。近20年来,已经被广泛应用于多种学科,如骨科、口腔颌面外科、整形美容科等。然而,至今,PRP的临床制作方法还没有统一标准,不同的制作方法导致PRP的成分以及临床疗效不同,给临床医务工作者应用PRP造成了诸多困扰。通过检索和分析中国已发表有关PRP制备的文献(中文和英文)发现,国内PRP临床制备方法差异性较大,包括采血量、离心设备、离心次数、离心力、离心时间和PRP制备量等。PRP中血小板富集度和回收率,白细胞浓度等指标也不稳定,亟需更高质量的研究或指南来规范和指导临床PRP的制备。     

Sperm motility and DNA integrity affected by different g‐forces in the preparation of sperm in urine specimens

Retrograde ejaculation, a common type of anejaculation, is attributable to many causes, some of which can be treated with medication and some of which cannot. For infertility treatment, sperm must be collected from the urine of the patients. Our study attempts to ascertain the effects of different g‐forces on sperm motility, morphology and DNA integrity in sperm preparation by the Sil‐Select? density gradient method of isolating sperm from urine specimens. Forty‐seven semen samples with normal semen analyses according to World Health Organisation (WHO) 1999 criteria were included in this study. Semen samples of 1 ml were mixed with 20 ml alkalinised normal urine and then divided equally into tubes A and B. The two samples were prepared by the Sil‐Select? density gradient centrifugation method at 350 g (tube A) and at 700 g (tube B). Total motile sperm after centrifugation at 700 g was significantly higher than after centrifugation at 350 g [6.7 (0.4–23.0) million versus 3.1 (0.1–13.7) million] (P < 0.001). There was no significant difference between the either the percentage of sperm with normal morphology or with DNA damage between centrifugation at 350 g and 700 g (P > 0.05), although centrifugation at 700 g achieves a higher number of total motile sperm compared with Sil‐Select? sperm preparation at 350 g centrifugation.     

一种新型肠上皮干细胞分离方法的建立

目的 探讨密度梯度离心法作为一种新型肠上皮干细胞分离方法的可行性.方法 通过腹腔注射大剂量5-氟尿嘧啶(5-FU)制备肠黏膜严重损伤小鼠模型,取出损伤的肠黏膜消化成单细胞悬液,利用Percoll密度梯度离心法分离细胞体积大小不同的细胞群.HE染色观察各群细胞形态特征,免疫细胞化学及RT-PCR检测各群细胞musashi-1(msi-1)的表达.结果 分离的大部分细胞位于50%和70%密度梯度的Percoll分离液体层,50%密度梯度分离液体层中的细胞符合干细胞的形态特征,免疫细胞化学检测其msi-1表达阳性率约为93%,RT-PCR显示该群细胞msi-1 mRNA表达较强.结论 建立了一种简便、有效的分离具有活性的肠上皮干细胞的方法.     

人重症肌无力胸腺树突状细胞的分离和培养

目的　探讨人重症肌无力胸腺树突状细胞 (TDC)的分离方法和诱导培养的最佳条件 ,为研究胸腺树突状细胞的生理功能及其在重症肌无力抗原提呈中的作用奠定基础。方法　新鲜人胸腺组织经机械破碎 ,梯度离心获得胸腺低密度细胞 ,经粒 巨噬细胞集落刺激因子 (GM CSF)和干细胞因子 (SCF)诱导 ,所获得细胞通过透射电镜、扫描电镜观察 ,并用FITC标记的抗人S10 0b、HLA DR、CD11c抗体进行鉴定。结果　从胸腺组织可获取足够数量胸腺树突状细胞 ,体外培养 10～ 18d细胞生长较为活跃 ,第 14天左右达到最高值 ,维持 3～ 4d开始下降 ,2 0d内细胞数仍能维持在 (1.0～ 2 .5 )× 10 5个 /ml。结论　梯度离心法能成功分离胸腺树突状前体细胞 ,GM CSF和SCF体外诱导可提供胸腺树突状细胞原代培养较为满意的生长条件。     

五种抗真菌药物对马尔尼菲青霉菌的体外抗菌活性观察

目的 了解两性霉素B(AMB)、酮康唑(KET)、氟康唑(FCA)、5-氟胞嘧啶(5-Fc)和伊曲康唑(ICA)5种抗真菌药物对马尔尼菲青霉菌体外抗菌活性,为临床用药提供参考依据.方法 采用浓度梯度法(E-test)测试AMB、KET、FCA、5-Fc、ICA 5种抗真菌药物对52株从不同AIDS患者骨髓、血液、皮肤损害标本中分离的马尔尼菲青霉菌酵母相和菌丝相的体外抗菌活性.数据采用U检验.结暴 AMB、KET、FCA、5-Fc、ICA的90%酵母相马尔尼菲青霉菌的最低抑制浓度(MIC90)分别为0.250、0.160、24.000、4.000和0.006 mg/L,最低抑制浓度(MIC)范围分别为0.004～0.500、00002～0.016、1.000～256.000、0.002～32.000和0.002～0.008 mg/L;对90%菌丝相马尔尼菲青霉菌的MIC90分别为1.500、0.125、256.000、24.000和0.012 mg/L,MIC范围分别为0.064～4.000、0.006～0.940、1.000～256.000、0.125～32.000和0.002～0.064 mg/L.不同抗真菌药对双相马尔尼菲青霉菌的体外抗菌活性不同,以ICA最强,其次为KET.酵母相和菌丝相的马尔尼菲青霉菌对同一药物的MIC比较差异有统计学意义(AMB、KET、FCA、5-Fc和ICA的U值分别为4.221 9、1.912、28.798、6.43、7.21,均P＜0.05).结论 进行马尔尼菲青霉菌体外抗真菌药物敏感实验对临床有重要参考意义.     

Computational simulation of the blood separation process

The aim of this work is to construct a computational fluid dynamics model capable of simulating the quasitransient process of apheresis. To this end a Lagrangian-Eulerian model has been developed which tracks the blood particles within a delineated two-dimensional flow domain. Within the Eulerian method, the fluid flow conservation equations within the separator are solved. Taking the calculated values of the flow field and using a Lagrangian method, the displacement of the blood particles is calculated. Thus, the local blood density within the separator at a given time step is known. Subsequently, the flow field in the separator is recalculated. This process continues until a quasisteady behavior is reached. The simulations show good agreement with experimental results. They shows a complete separation of plasma and red blood cells, as well as nearly complete separation of red blood cells and platelets. The white blood cells build clusters in the low concentrate cell bed.     

Comparison of the fatigue characteristics of centrifuged and uncentrifuged Simplex P bone cement

Fatigue test specimens of Simplex P bone cement (Howmedica Inc., Rutherford, NJ) prepared according to the manufacturer's instructions and specimens of Simplex P prepared by centrifuging the cement immediately after mixing were subjected to fully reversed tension-compression fatigue tests at initial strain levels ranging from 0.01 to 0.001. S:N curves for both cement preparations were generated and compared on the basis of regression analysis, Weibull analysis, and Student's t tests. Centrifuged Simplex P was able to withstand significantly more fatigue cycles than uncentrifuged Simplex P at all strain levels tested. Importantly, at the more physiologic strain levels of 2,000 and 1,000 microstrain, the centrifuged cement demonstrated superior fatigue life. Specifically, at the 1,000-microstrain level eight of the 11 uncentrifuged specimens fractured before undergoing 10 million cycles. Of those that failed, the average number of cycles to failure was 1.8 million with a range of 560,000 to 4 million cycles. In contrast, all 11 centrifuged specimens tested at 1,000 microstrain remained intact at 10 million cycles.     

自体树突状细胞体外诱导扩增方法的安全性改进

目的 :改进从外周血获得自体树突状细胞的方法的临床安全性。方法 :密度梯度离心法分离鼻咽癌患儿外周血单个核细胞 (PBMC) ,贴壁法获得单核细胞 ,以IL 4、GM CSF、TNF α和人血浆进行诱导扩增树突细胞(DC)并进行表面分子鉴定 ,同时检测其对自体外周血T细胞的刺激作用。结果 :成功培养并扩增出成熟度较高的DC ,平均从每 1mL外周血扩增出的DC数量达到 ( 3 2± 1 1)× 10 5,这些DC具备很强的刺激T细胞增殖的能力。结论 :本法可从外周血扩增大量自体树突状细胞 ,有很强的刺激自体T细胞增殖的活性 ,并增加临床应用的安全性。     

骨髓基质细胞不同培养方法效果比较

目的：观察全骨髓培养法与密度梯度离心法对培养骨髓基质细胞的效果差异,建立简单、经济、实用分离骨髓间充质干细胞方法。方法：体外提取骨髓基质细胞,分别采用全骨髓培养法与密度梯度离心法培养骨髓基质细胞,观察细胞形态学、细胞活力、细胞周期及细胞表面分子表达差异。结果：两种方法均能有效培养出骨髓间充质干细胞,各细胞周期的细胞检出率无显著差异（P〉0．05）,细胞形态学无差异,细胞表面分子CD34、SH1、SH2、SH3表达差异不显著（P〉0．05）。结论：全骨髓培养法在分离培养骨髓基质细胞方面与密度梯度离心法具有类似效果。