[科学文献]无创性产前诊断进展 (二)

母亲血浆中的胎儿DNA及RNA

最近从母亲血浆中发现数量易于检测的胎儿DNA为无创性产前检测提供新的可能性。游离的胎儿DNA随孕期不断增多,估计占母亲血浆总DNA的3%到6%,在血清中数量较少[23、24]。因此,直接分析游离胎儿DNA在技术上比识别和分离胎儿细胞容易。而且,分娩后胎儿DNA从母亲循环中迅速清除,在分娩后两小时内就检测不到[25]。除DNA外,母血循环中出现的胎儿RNA非常稳定,能在循环中存在至少24小时而不被降解,因为这些RNA通过与特殊物质结合而受保护[26]。尽管母亲血浆中游离胎儿DNA及RNA的来源还未确定,但一些指征表明其来源于滋养细胞破裂。人绒毛膜促性腺激素及胎盘促乳激素序列的RNA转录物的出现表明这些RNA至少部分来源于胎盘[27]。有趣的是出现21三体及13三体等非整倍体时,胎儿游离DNA及胎儿细胞均增加[28,29],可能是因为这些情况下胎盘结构出现异常,使得胎儿细胞及DNA渗漏到母亲循环。

 

无创性产前诊断的应用

母亲循环中出现胎儿细胞提供了一种检测胎儿某些基因位点是否出现或缺失的方法,这在Y染色体探查中首次得到证实。由于通过Y染色体识别男性胎儿,该技术被迅速应用于诊断女性胎儿以排除X连锁疾病。类似的技术如FISH能检测非整倍体,例如21三体等。最近,PCR分析游离胎儿DNA已被应用于探查怀有RHD阳性胎儿的RHD阴性母亲血浆中的RHD基因位点[31,32]。这种检测已经成为国际血型参照实验室(http://www.bloodnet.nbs.nhs.uk/ibgrl)常规临床应用的方法。

应用这些技术对单基因病(例如错义或物义突变引起的)产前诊断发展缓慢,因为技术的复杂性导致难以进行大规模的筛查,只能有选择的应用于有阳性家族史、符合孟德尔遗传规律及基因位点明确的疾病再发风险的病例。虽然如此,通过分析分离到的有限胎儿细胞,已经在临床上应用于镰状细胞性贫血和地中海贫血等疾病的产前诊断[32]。在激光捕获显微切割等显微操作系统的辅助下,可以从母亲血液中分析到被染色的单个胎儿细胞,然后使用单细胞PCR技术寻找已知突变。

对游离胎儿DNA直接测序以检测胎儿特异的突变,能避开分离胎儿细胞的问题。但分析胎儿特异的DNA突变的主要问题在于,胎儿DNA中来源于母亲的等位基因总是被循环中母亲自身的序列所掩蔽,导致无法分辨只有父源突变的杂合子携带者和复合杂合子患病胎儿。虽然如此,以下策略可能有用:首先筛查胎儿DNA的父源突变,假如出现突变,再应用另外其他方法诊断胎儿。这些方法加以改进很可能将父源显性突变和常染色体隐性遗传病的复合杂合子的父源突变这两种突变区分开来。研究表明利用这些方法探查囊性纤维化、软骨发育不全、先天性肾上腺过度增生等的突变基因位点是可行的[33-35],还可以在后续妊娠中监测非整倍体中父源性染色体结构重排[36]。

小结

在母亲循环中发现的胎儿细胞及游离DNA为无创性产前诊断的发展提供了基础。无创性检测能避免绒毛采样或羊水穿刺等侵入性技术导致的流产风险增高等并发症,也使得产前诊断的时间尽量提前,从而为父母亲和医务人员提供有关胎儿基因型的更准确和及时的信息。

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Opinion TRENDS in Molecular Medicine,2003,9(8):339-343.(黄捷译)
(文章来源:<基因诊断>杂志http://www.daangene.com )