Souzaki Ryota, Tajiri Tatsuro, Higashi Mayumi, Kinoshita Yoshiaki, Tanaka Sakura, Kohashi Kenichi, Tsuneyoshi Masazumi, Taguchi Tomoaki
Department of Pediatric Surgery, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
Pediatr Surg Int. 2008 Oct;24(10):1095-100. doi: 10.1007/s00383-008-2228-3.
Recently, determining the MYCN status in neuroblastoma (NB) using the quantitative PCR (Q-PCR) and FISH instead of the Southern blotting (SB) has been recommended. In order to assess the implications of the gene dosage of MYCN in NB, the MYCN status was evaluated using Q-PCR on DNA extracted from small areas of NB specimens obtained using laser capture microdissection (LCM).
MYCN gene dosages (MYCN/NAGK) were determined in 63 primary NB block samples, as well as in 243 microdissected tissues from 63 samples using Q-PCR. In 23 of 63 cases, the MYCN gene status was evaluated using FISH.
Nine block samples with the amplification of MYCN based on SB showed a remarkable increase of the MYCN gene dosage using Q-PCR. Twelve of 54 block samples with no amplification of MYCN based on SB showed a slight increase of the MYCN gene dosage (3.56 > or == MYCN/NAGK > 1.84), and 8 of these 12 cases were in the advanced stage. Among these 12 cases, 1 case had several LCM areas with a high copy number of MYCN and several LCM areas which showed no increase of MYCN gene. Another case showed a slight increase in the MYCN gene dosage (3.65 < or == MYCN/NAGK < or == 4.82) in all LCM areas. In addition, a large number of cells with the MYCN gain were found using FISH in the block sample. In 2 other cases of 12 cases, although no LCM areas showed an increased gene dosage of MYCN, a small number of cells with MYCN amplification were found using FISH were found in the block sample.
A slight increase in the gene dosage of MYCN detected by Q-PCR may indicate that the NB tissue contains a small number of cells with the MYCN amplification or a large number of cells with the MYCN gain, which are associated with the aggressive progression of NB.
最近，有人建议使用定量聚合酶链反应（Q-PCR）和荧光原位杂交（FISH）而非Southern印迹法（SB）来确定神经母细胞瘤（NB）中的MYCN状态。为了评估MYCN基因剂量在NB中的意义，我们使用Q-PCR对通过激光捕获显微切割（LCM）从NB标本小区域提取的DNA进行了MYCN状态评估。
使用Q-PCR测定了63个原发性NB块状样本以及来自63个样本的243个显微切割组织中的MYCN基因剂量（MYCN/NAGK）。在63例中的23例中，使用FISH评估了MYCN基因状态。
基于SB显示MYCN扩增的9个块状样本，使用Q-PCR显示MYCN基因剂量显著增加。基于SB显示无MYCN扩增的54个块状样本中的12个显示MYCN基因剂量略有增加（3.56≥MYCN/NAGK＞1.84），这12例中的8例处于晚期。在这12例中，1例有几个MYCN拷贝数高的LCM区域和几个未显示MYCN基因增加的LCM区域。另一例在所有LCM区域中MYCN基因剂量略有增加（3.65≤MYCN/NAGK≤4.82）。此外，在块状样本中使用FISH发现大量具有MYCN增加的细胞。在12例中的另外2例中，虽然没有LCM区域显示MYCN基因剂量增加，但在块状样本中使用FISH发现少量具有MYCN扩增的细胞。
Q-PCR检测到的MYCN基因剂量略有增加可能表明NB组织含有少量具有MYCN扩增的细胞或大量具有MYCN增加的细胞，这与NB的侵袭性进展相关。
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