Steg Adam, Vickers Selwyn M, Eloubeidi Mohamad, Wang Wenquan, Eltoum Isam A, Grizzle William E, Saif M Wasif, Lobuglio Al F, Frost Andra R, Johnson Martin R
Department of Pharmacology and Toxicology, Division of Clinical Pharmacology, University of Alabama at Birmingham, Birmingham, AL, USA.
Diagn Mol Pathol. 2007 Dec;16(4):229-37. doi: 10.1097/PDM.0b013e31811edc7e.
Recent studies suggest that hedgehog (HH)-pathway signaling is required for the initiation and continued growth of pancreatic adenocarcinoma (PAC). Definitive gene expression analysis of PAC remains difficult, owing to the host desmoplastic stromal interaction and subsequent tumor heterogeneity. The primary goal of this study was to evaluate the effect of heterogeneity within a series (n=5) of matched clinical PAC biopsies [snap-frozen, formalin-fixed paraffin-embedded (FPE), endoscopic ultrasound-guided fine-needle aspirate (EUS-FNA)]. Differential expressions, specific to tumor cells, were evaluated by comparisons of uninvolved pancreas (n=9), EUS-FNA (n=14), and macrodissected (tumor-cell-enriched) biopsies (n=16). To determine whether treatment modulates gene expression, a unique (independent) set of synchronous EUS-FNA samples (n=4) was obtained before, and 2 weeks after, chemoradiation. mRNA levels were evaluated using real-time quantitative polymerase chain reaction formatted in a TaqMan low-density array, which was capable of simultaneously quantifying 46 independent genes in the HH pathway. Protein levels for Patched, Smoothened, and glioma-associated oncogene 1 (Gli-1) in FPE tissues were determined, using immunohistochemistry. A significant concordance (P<0.0001) was observed in the HH-pathway mRNA levels between matched surgically resected (both snap-frozen and FPE) and EUS-FNA biopsies. HH-pathway mRNA levels changed (increased) only after macrodissection, suggesting localization to tumor cells. Immunohistochemical staining for Patched, Smoothened, and Gli-1 confirmed the increased (P<0.001) levels of protein in the PAC cells, compared with cells from uninvolved pancreas. EUS-FNA biopsies that were obtained before and during chemoradiation demonstrated no significant changes in HH-pathway gene expression. Collectively, these studies demonstrate presence of HH-pathway expression in all the clinical PAC biopsies examined, suggesting that this is a significant tumor-associated target and offering the possibility that specific molecular profiling might be attempted from these heterogeneous tissues.
近期研究表明，胰腺癌（PAC）的起始和持续生长需要刺猬（HH）信号通路传导。由于宿主促结缔组织增生性基质相互作用以及随后的肿瘤异质性，对PAC进行确切的基因表达分析仍然困难重重。本研究的主要目的是评估一系列（n = 5）匹配的临床PAC活检样本[速冻、福尔马林固定石蜡包埋（FPE）、内镜超声引导下细针穿刺抽吸（EUS-FNA）]中的异质性影响。通过比较未受累胰腺（n = 9）、EUS-FNA（n = 14）和宏观解剖（富含肿瘤细胞）活检样本（n = 16），评估肿瘤细胞特有的差异表达。为了确定治疗是否会调节基因表达，在放化疗前和放化疗后2周获取了一组独特（独立）的同步EUS-FNA样本（n = 4）。使用TaqMan低密度阵列格式的实时定量聚合酶链反应评估mRNA水平，该阵列能够同时定量HH信号通路中的46个独立基因。使用免疫组织化学法测定FPE组织中Patched、Smoothened和胶质瘤相关癌基因1（Gli-1）的蛋白水平。在手术切除的匹配样本（速冻和FPE）与EUS-FNA活检样本之间，观察到HH信号通路mRNA水平存在显著一致性（P < 0.0001）。HH信号通路mRNA水平仅在宏观解剖后发生变化（升高），表明其定位于肿瘤细胞。与未受累胰腺的细胞相比，对Patched、Smoothened和Gli-1进行的免疫组织化学染色证实PAC细胞中的蛋白水平升高（P < 0.001）。在放化疗前和放化疗期间获取的EUS-FNA活检样本显示HH信号通路基因表达无显著变化。总体而言，这些研究表明在所检查的所有临床PAC活检样本中均存在HH信号通路表达，这表明其是一个重要的肿瘤相关靶点，并提供了从这些异质性组织中尝试进行特定分子谱分析的可能性。
