研究發現輸血相關急性肺損傷的新觀點

作者：Roxanne Nelson  
出處：WebMD醫學新聞

　　December 21, 2009 (紐澳良) — 輸血相關急性肺損傷(Transfusion-related acute lung injury[TRALI])是輸血相關發病率和死亡率的主因，特徵包括急性非心因性肺水腫以及呼吸狀態不佳。不過，TRALI的病因還未充分瞭解，許多TRALI案例可能是血液成分中抗白血球抗原的抗體導致。
　　
　　發表於美國血液協會第51屆年會的這篇新研究，有助於減少這個嚴重的輸血相關併發症。
　　
　　加州大學舊金山分校血液腫瘤科副教授、Lawrence Berkeley國家實驗室科學研究員Joel Anne Chasis醫師解釋，現在已經知道TRALI是由人類嗜中性白血球抗原(human neutrophil antigen[HNA])-3a-陰性特定抗體引起，一般人約4%到5%有此風險，因為他們是HNA-3a-陰性。
　　
　　未參與該研究的Chasis醫師向Medscape Oncology表示，這個新研究描述的基因圖調查發現，一個單一核苷酸多型性(single-nucleotide polymorphism[SNP])與HNS-3a-陰性顯型有關。Chasis醫師解釋，希望本研究最終可以導出篩檢個人血液和捐贈者血液中的HNS-3a-抗體之策略；HNS-3a-抗體可能會引起TRALI。
　　
　　【急性肺損傷】
　　根據研究第一作者、密爾瓦基州威斯康辛血液中心血小板與嗜中性白血球免疫實驗室技術主任Brian R. Curtis、MT(ASCP)SBB解釋，TRALI之定義為輸血期間或輸血後6小時內發生的急性肺損傷，通常是捐血製品中的白血球抗體導致。白血球抗體被認為與接受者的嗜中性白血球發生交互作用，導致肺微血管活化與凝集，釋出局部生物反應調節子，導致微血管裂縫與肺損傷。
　　
　　於最新摘要小組會議中發表此發現的Curtis先生表示，這是通報給美國食品藥物管理局之輸血相關死因的第一名，與TRALI有關聯的抗體包括捐贈者的第一類人類白血球抗原(human leukocyte antigen[HLA])、第二類HLA、和/或嗜中性白血球特定抗體，即使有許多嗜中性白血球特定抗體曾被報導，最常見的是與HNA-3a抗原直接有關。
　　
　　他指出，即使HNA-3a/b抗原在45年前發現，抗原本身的蛋白質載體和分子組成依舊未知，Curtis先生表示，但是，經確認HNA-3a-特定抗體特別容易引起嚴重且經常致命的TRALI，定義HNA-3a/b抗原的重要性益發必要。
　　
　　【CTL2 有關聯】
　　本研究中，Curtis先生等人試圖確認HNA-3a的蛋白質載體，對8個HNA-3a陰性者與107個無關正常人的DNA進行基因圖SNP掃描。他們觀察發現，每個HNA-3a陰性者位於SLC44A2 基因、代表膽鹼類運輸子蛋白質2(choline transporter-like protein[CTL]2)的nt542之SNP (G>A)都是同型合子(AA) (P< .000001、校正調查的SNP數量)。相反地，正常對照組只有5個如此。
　　
　　他表示，這個特別的SNP (rs2288904)是SLC44A2上面唯一與HNA-3a陰性顯型有關者，預測成熟蛋白質的細胞外胺基酸取代(R>Q154)。
　　
　　為了確定他們的SNP分析，研究者直接定序代表CTL2的DNA，無意外的，發現nt542G和nt542A只與HNA-3a和HNA-3b顯型有關。
　　
　　接著，他們進行十二烷基硫酸鈉聚丙烯醯胺膠體電泳(sodium dodecyl sulfate polyacrylamide gel electrophoresis[SDS-PAGE])，且將可能的區段進行質譜分析，以獲得CTL2帶有HNA-3a的直接證據，三次獨立分析之後，他們觀察發現，衍生自CTL2的胜肽超過蛋白質的整個長度，因此，他們結論表示，SLC44A2基因的nt542，以G代替A所產生的HNA-3a抗原，導致在CTL2第一細胞外環的154位置處插入R取代Q。
　　
　　雖然CTL2的功能依舊不知道，研究者指出，嗜中性白血球上抗HNA-3a抗原之交聯，誘發其細胞的爆炸性活化。這個活化表示CTL2在這類細胞內的功能性，因此提供在CTL-2特定抗體病患輸血時觀察發現之嚴重TRALI的可能解釋。
　　
　　Curtis先生結論表示，說明HNA-3a/b的分子基礎有助於我們針對重組蛋白質或蛋白質片段之捐贈者血液中抗體的偵測能力，我們目前可以定出個人的基因型，迅速確定HNA-3a陰性者，作為發生TRALI時診斷評估的一部份，也可確定有風險的病患。
　　
　　他指出，這將可以降低未來捐血時的此一併發症風險。
　　
　　作者們皆宣告沒有相關財務關係。
　　
　　美國血液協會(ASH)第51屆年會：摘要LBA 4。發表於2009年12月8日。


Study Reveals New Insight Into Transfusion-Related Acute Lung Injury

By Roxanne Nelson
Medscape Medical News

December 21, 2009 (New Orleans, Louisiana) — Transfusion-related acute lung injury (TRALI) is a leading cause of transfusion-related morbidity and mortality, and characterized by acute noncardiogenic pulmonary edema and compromised respiratory status. Although the etiology of TRALI is not fully understood, many cases of TRALI are likely caused by antibodies to leukocyte antigens in blood components.

New translational research presented here at the American Society of Hematology 51st Annual Meeting might help lead to a reduction of this serious transfusion-related complication.

TRALI is now known to be caused by human neutrophil antigen (HNA)-3a-negative specific antibodies, and about 4% to 5% of the general population is at risk because they are HNA-3a-negative, explained Joel Anne Chasis, MD, associate professor of hematology/oncology at the University of California in San Francisco, and a staff scientist at the Lawrence Berkeley National Laboratory.

Dr. Chasis, who was not involved in the study, told Medscape Oncology that this new research describes a genome-wide survey that found a single-nucleotide polymorphism (SNP) that correlates with the HNS-3a-negative phenotype. The hope is that this research will eventually lead to strategies to screen both individuals and donated blood for presence of the HNA-3a-antibodies, which are capable of causing TRALI, Dr. Chasis explained.

Acute Lung Injury

TRALI is defined as an acute lung injury occurring during or within 6 hours of a transfusion, and is most often caused by leukocyte antibodies that are present in the blood-donor product. The leukocyte antibodies are thought to interact with recipient neutrophils, resulting in the activation and aggregation in pulmonary capillaries, the release of local biologic response modifiers causing capillary leak, and lung injury, according to the study's lead author, Brian R. Curtis, MS, MT(ASCP)SBB, technical director of the Platelet Neutrophil Immunology Laboratory, BloodCenter of Wisconsin, in Milwaukee.

"It continues to be the number?1 cause of transfusion-related mortality reported to the [US Food and Drug Administration]]," said Mr. Curtis, who presented the findings during a late-breaking abstract session. Antibodies that have been implicated in TRALI include donor human leukocyte antigen (HLA) class?I, HLA class?II, and/or neutrophil-specific antibodies, and even though a number of neutrophil-specific antibodies have been reported, the most common is directed at the HNA-3a antigen.

He pointed out that even though HNA-3a/b antigens were discovered 45 years ago, the protein carrier and the molecular composition of the antigens themselves remains unknown. But with the recognition that HNA-3a-specific antibodies are especially prone to cause severe and often fatal TRALI, the importance of defining the HNA-3a/b antigens has become imperative, said Mr. Curtis.

CTL2 Implicated

In this study, Mr. Curtis and colleagues attempted to determine the protein carrier for HNA-3a, and conducted a genome-wide SNP scanning of the DNA from 8 HNA-3a-negative people and 107 unrelated normal people. They observed that each of the HNA-3a-negative people were homozygous (AA) for a SNP (G>A) located at nt542 of the SLC44A2 gene encoding the choline transporter-like protein (CTL)2 (P?< .000001 corrected for the number of SNPs surveyed). Conversely, this was true for only 5 of the normal individuals.

This particular SNP (rs2288904) was the only one in SLC44A2 that correlated with the HNA-3a-negative phenotype, and predicted an extracellular amino acid substitution (R>Q154) in the mature protein, he said.

To validate their SNP analyses, the researchers directly sequenced DNA encoding CTL2 and found that, without exception, nt542G and nt542A correlated only with the phenotypes HNA-3a and HNA-3b.

Next, they performed sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and subjected the candidate bands to mass spectrometric analysis to obtain direct evidence that HNA-3a is carried on CTL2. After 3 independent assays, they observed that peptides derived from CTL2 spanned the entire length of the protein. Therefore, they concluded that the "HNA-3a antigen results from the substitution of G for A at nt542 of the SLC44A2 gene, leading to the insertion of R for Q at position 154 in the first extracellular loop of CTL2."

Although the function of CTL2 remains unknown, the researchers noted that "cross-linking of the antigen by anti-HNA-3a in neutrophils triggers an explosive activation of primed cells." This action indicates that CTL2 is functional within this type of cell, and thus provides a possible explanation for the severe TRALI reactions that have been observed in patients transfused with CTL2-specific antibodies.

"Elucidation of the molecular basis of HNA-3a/b should facilitate our ability to detect the antibodies in blood donors using recombinant proteins or fragments of the protein that target the antibody," concluded Mr. Curtis. "We can currently genotype individuals now and rapidly identify HNA-3-negative individuals as part of a diagnostic assessment of TRALI in cases that occur, and also identify persons at risk."

This could potentially reduce the risk for this complication in future blood donation, he added.

The authors have disclosed no relevant financial relationships.

American Society of Hematology (ASH) 51st Annual Meeting: Abstract LBA 4. Presented December 8, 2009.