Kakourou Georgia, Vrettou Christina, Kattamis Antonis, Destouni Aspasia, Poulou Myrto, Moutafi Maria, Kokkali Georgia, Pantos Konstantinos, Davies Stephen, Kitsiou-Tzeli Sophia, Kanavakis Emmanuel, Traeger-Synodinos Joanne
a Department of Medical Genetics , National and Kapodistrian University of Athens, Choremio Research Laboratory .
b First Department of Pediatrics , Athens University Medical School .
Syst Biol Reprod Med. 2016;62(1):69-76. doi: 10.3109/19396368.2015.1100692. Epub 2015 Dec 4.
Preimplantation genetic diagnosis (PGD) to select histocompatible siblings to facilitate curative haematopoeitic stem-cell transplantation (HSCT) is now an acceptable option in the absence of an available human leukocyte antigen (HLA) compatible donor. We describe a case where the couple who requested HLA-PGD, were both carriers of two serious haematological diseases, beta-thalassaemia and sideroblastic anaemia. Their daughter, affected with sideroblastic anaemia, was programmed to have HSCT. A multiplex-fluorescent-touchdown-PCR protocol was optimized for the simultaneous amplification of: the two HBB-gene mutated regions (c.118C> T, c.25-26delAA), four short tandem repeats (STRs) in chr11p15.5 linked to the HBB gene, the SLC25A38 gene mutation (c.726C > T), two STRs in chr3p22.1 linked to the SLC25A38 gene, plus eleven informative STRs for HLA-haplotyping (chr6p22.1-21.3). This was followed by real-time nested PCR and high-resolution melting analysis (HRMA) for the detection of HBB and SLC25A38 gene mutations, as well as the analysis of all STRs on an automatic genetic analyzer (sequencer). The couple completed four clinical in vitro fertilization (IVF)/PGD cycles. At least one matched unaffected embryo was identified and transferred in each cycle. A twin pregnancy was established in the fourth PGD cycle and genotyping results at all loci were confirmed by prenatal diagnosis. Two healthy baby girls were delivered at week 38 of pregnancy. The need to exclude two familial disorders for HLA-PGD is rarely encountered. The methodological approach described here is fast, accurate, clinically-validated, and of relatively low cost.
在没有可用的人类白细胞抗原（HLA）匹配供体的情况下，通过植入前基因诊断（PGD）选择组织相容性同胞以促进治愈性造血干细胞移植（HSCT）现在是一种可接受的选择。我们描述了一对请求进行HLA-PGD的夫妇的案例，他们都是两种严重血液疾病——β地中海贫血和铁粒幼细胞贫血的携带者。他们患有铁粒幼细胞贫血的女儿计划接受HSCT。优化了一种多重荧光降落式PCR方案，用于同时扩增：两个HBB基因突变区域（c.118C>T，c.25 - 26delAA）、11号染色体p15.5上与HBB基因连锁的四个短串联重复序列（STR）、SLC25A38基因突变（c.726C>T）、3号染色体p22.1上与SLC25A38基因连锁的两个STR，以及用于HLA单倍型分型的11个信息性STR（6号染色体p22.1 - 21.3）。随后进行实时巢式PCR和高分辨率熔解分析（HRMA）以检测HBB和SLC25A38基因突变，并在自动基因分析仪（测序仪）上分析所有STR。这对夫妇完成了四个临床体外受精（IVF）/PGD周期。在每个周期中至少鉴定并移植了一个匹配的未受影响胚胎。在第四个PGD周期中确定为双胎妊娠，并且所有位点的基因分型结果通过产前诊断得到证实。在妊娠第38周时分娩了两个健康女婴。为进行HLA-PGD而需要排除两种家族性疾病的情况很少见。这里描述的方法学方法快速、准确、经过临床验证且成本相对较低。
