Search for maternal cells in human umbilical cord blood by polymerase chain reaction amplification of two minisatellite sequences.
Search for maternal cells in human umbilical cord blood by polymerase chain reaction amplification of two minisatellite sequences.
复制标题
通过聚合酶链反应扩增两个小卫星序列来寻找人脐带血中的母体细胞。
作者：
G. Socié;E. Gluckman;E. Carosella;Y. Brossard;C. Lafon;O. Brison
自1989年首次报告以来，已报告了26例脐带血移植。此外，全世界正在进行大规模使用库存胎盘血进行无关受体骨髓重建可行性的系统研究。然而，早在1989年，就有人指出，脐带血的使用可能会受到新生儿血液与母体细胞污染的阻碍，这些细胞会导致不可接受的移植物抗宿主病（GVHD）。在本研究中，我们使用聚合酶链反应（PCR）扩增2个小卫星序列（33.6和MS 51）来解决这个问题。已知小卫星序列的PCR扩增的灵敏度为1%至0.1%。在本研究中证实了这种灵敏度，其中对进行细胞分离的每个实验进行了稀释分析。在这些分析中纳入稀释实验使我们能够估计污染母体细胞的相对量（如果有的话）。在47例病例中（31例全血分析，10例梯度分离和6例亚群分离），2个小卫星序列的共扩增允许在42例（89%）中区分母亲和新生儿等位基因。在一个案例中，我们能够在母亲的全血样本中检测到儿童特异性等位基因，从而验证了我们在脐带血中寻找母体细胞的方法。在另一个病例中，我们能够在脐带血样本中检测到母亲特异性等位基因。在全血、多形核细胞和淋巴细胞组分中检测到该母体特异性等位基因。将这3份脐带血样本获得的信号强度与同一分析中进行的稀释实验结果进行比较，得出多形核细胞组分中母体细胞的估计值为1 - 5%，全血和淋巴细胞组分中母体细胞的估计值为0.1%-1%。总之，我们的研究表明，母体细胞很少存在于出生时收集的脐带血中，因为我们只在47例病例中的1例中检测到它们。更重要的是，当检测时，它们在淋巴细胞部分中的水平非常低（0.1%至1%）。然而，尽管细胞量很小，但这种细胞量可能导致易感受体的GVHD。因为我们使用的方法允许从10（4）个有核细胞中检测脐带血中的母体细胞，因此它在脐带血库的角度是有意义的。
Since our first report in 1989, 26 transplants by means of umbilical cord blood have been reported. Furthermore, systematic studies of the feasibility of using banked placental blood for bone marrow reconstitution of unrelated recipients on a large scale are in progress worldwide. However, already by 1989, it was pointed out that the use of cord blood might be hampered by contamination of neonatal blood with maternal cells contributing unacceptably to graft-versus-host disease (GVHD). In the present study, we used the polymerase chain reaction (PCR) amplification of 2 minisatellite sequences (33.6 and MS 51) to address this question. The sensitivity of PCR amplification of minisatellite sequences is known to be of 1% to 0.1%. This sensitivity has been confirmed in the present study, in which a dilution analysis was performed for each experiment in which cell separation was performed. The inclusion of the dilution experiment in these analyses allowed us to estimate the relative amount of contaminating maternal cells, if any. Among 47 cases (31 whole blood analyses, 10 gradient separations, and 6 subpopulation separations), the coamplification of the 2 minisatellites sequences allowed the discrimination of maternal and neonate alleles in 42 cases (89%). In 1 case, we were able to detect a child-specific allele in a mother's whole blood sample, thus validating our approach to search for maternal cells in cord blood. In a single other case, we were able to detect a maternal-specific allele in the cord blood sample. This maternal specific allele was detected in the whole blood, polymorphonuclear cell, and lymphocyte fractions. Comparison of the signal intensity obtained with these 3 cord blood samples to the result of the dilution experiment performed in the same analysis led to an estimate of 1 to 5% maternal cells in the polymorphonuclear cell fraction and 0.1% to 1% maternal cells in the whole blood and lymphocyte cell fractions. In conclusion, our study indicates that maternal cells are very rarely present in the cord blood collected at birth because we detected them in only 1 of 47 cases. More importantly, when detected, they were present at very low level in the lymphocyte cell fraction (0.1% to 1%). However, although small, this amount of cells may result in GVHD in a susceptible recipient. Because the method we used allows the detection of maternal cells within cord blood from 10(4) nucleated cells, it would thus be of interest in a cord blood banking perspective.
DOI：
--
发表时间：
1991
期刊：
Blood cells
影响因子：
--
作者：
H. Broxmeyer;J. Kurtzberg;E. Gluckman;A. Auerbach;G. Douglas;S. Cooper;J. Falkenburg;J. Bard;E. A. Boyse
通讯作者：
H. Broxmeyer;J. Kurtzberg;E. Gluckman;A. Auerbach;G. Douglas;S. Cooper;J. Falkenburg;J. Bard;E. A. Boyse
影响因子：
158.5
作者：
GLUCKMAN, E;BROXMEYER, HE;BOYSE, EA
通讯作者：
BOYSE, EA
DOI：
10.1073/pnas.86.10.3828
发表时间：
1989-05-01
影响因子：
11.1
作者：
BROXMEYER, HE;DOUGLAS, GW;BOYSE, EA
通讯作者：
BOYSE, EA