Red blood cells (RBCs) of the Asian-type DEL phenotype express few RhD proteins and are typed as serologic RhD-negative (D-) in routine testing. RhD-positive (D+) RBC transfusion for Asian-type DEL patients has been proposed but has not been generally adopted due to a lack of direct evidence regarding its safety and underlying mechanism. We performed a single-arm multicenter clinical trial to document the outcome of D+ RBC transfusion in Asian-type DEL patients; none of the recipients (0/42; 95% confidence interval, 0%-8.40%) developed alloanti-D after a median follow-up of 226 days. We conducted a large retrospective study to detect alloanti-D immunization in 4,045 serologic D- pregnant women throughout China; alloanti-D was found only in true D- individuals (2.63%, 79/3,009), but not in those with Asian-type DEL (0/1,032). We further retrospectively examined 127 serologic D- pregnant women who had developed alloanti-D and found none with Asian-type DEL (0/127). Finally, we analyzed RHD transcripts from Asian-type DEL erythroblasts and examined antigen epitopes expressed by various RHD transcripts in vitro, finding a low abundance of full-length RHD transcripts (0.18% of the total) expressing RhD antigens carrying the entire repertoire of epitopes, which could explain the immune tolerance against D+ RBCs. Our results provide multiple lines of evidence that individuals with Asian-type DEL cannot produce alloanti-D when exposed to D+ RBCs following transfusion or pregnancy. Therefore, we recommend considering D+ RBC transfusion and discontinuing anti-D prophylaxis in Asian-type DEL patients, including pregnant women. This clinical trial is registered at www.clinicaltrials.gov as NCT03727230.

[目的]本研究旨在对RhD阳性个体中红细胞特异性表达的RHD基因不同转录本进行分析,并对选择性剪接发生的机制进行初步探讨.[方法]利用体外有核红细胞培养体系分离并培养RhD阳性个体的有核红细胞,使用逆转录聚合酶链反应(RT-PCR)技术对RhD阳性个体RHD基因外显子6至3'非编码区扩增后进行二代测序,分析mRNA转录本,并采用生物信息学方法分析RHD基因所有外显子5'及3'剪接位点的最大熵值.[结果]RHD基因的转录本以正常全长mRNA为主,除此之外还存在其他8种异常转录本.按照表达量从多到少,分别为外显子7缺失、外显子9缺失、外显子8和9缺失、外显子7、8和9缺失、外显子7和9缺失、外显子8和9缺失合并外显子7与10间插入一段170 bp的内含子7片段、全长mRNA但插入170 bp的内含子7片段和外显子9缺失但插入一170 bp的内含子7片段,其中最后3种为新转录本.生物信息学分析提示外显子7和外显子9异常剪接可能与其5'-剪接位点(5'-splice site,5'ss)或3'ss和剪接体结合能力下降有关,而内含子7片段异常表达于转录本中可能与其两端存在类似5'ss和3'ss剪接保守序列有关.[结论]RHD基因存在十分复杂的选择性剪接模式,使RhD阳性个体的RHD基因在mRNA水平上存在多种不同的剪接转录本.