背景与目的：BNIP3L（Bel-2/ElB 19 kDa-iteracting protein 3-like）基因是从人胎肝cDNA文库中克隆得到的具有抑瘤活性的基因，定位于8p21肺癌高频杂合性丢失（loss of heterozygosity, LOH）区；BNIP3L蛋白能与Bel-2、Bel-xL、E1819K等抗凋亡蛋白相互作用，促进细胞凋亡。本研究旨在探讨BNIP3L基因表达、结构异常与肺癌发生、发展的关系。方法：采用免疫组化和免疫印迹技术、半定量逆转录聚合酶链反应（reverse transcnption-PCR.RT-PCR）、PCR-单链构象多态性（PCR-single straincon formation polymorphism, PCR-SSCP）分析等方法检测4种肺癌细胞株NCI-H520、A549、NCI-H460、NCI-H446和30例肺癌组织中BNIP3L基因的表达及结构异常。结果：（1）除NCI-H520细胞外，A549、NCI-H460和NCI-H446细胞中均无BNIP3L蛋白表达，NCI-H520细胞中BNIP3L蛋白表达水平也略低于水生化支气管上皮细胞HBE4-E6/E7。30例肺癌组织中BNIP3L蛋白阳性率为46.7％（14／30），12例正常肺组织中阳性率为100％（12/12），二者差异有统计学意义（P<0.05）。（2）4种肺癌细胞株中均存在BNIP3L mRNA表达，其表达水平与HBE4-E6/E7细胞相比无明显差异。30例肺癌组织中8例（26 7％）存在BNIP3L mRNA表达缺失或明显减弱，肺癌组织中BNIP3L mRNA平均表达量为0.404±0.070，配对正常肺组织为0.575±0.065，二者差异有显著性（P<0.05）。所有表达BNIP3L mRNA的肺癌细胞和组织，其RT-PCR产物（包括编码区全长）大小与对照样本一致，未检测到缺失、重排、剪切异常等BNIP3L基因结构改变。（3）4种肺癌细胞和30例肺癌组织中BNIP3L基因6个外显子均未检测到点突变。结论：BNIP3L蛋白在肺癌中存在表达下调，可能参与肺癌的发生、发展；BNIP3L蛋白在肺癌中表达下调部分是由转录下调造成的；基因结构异常可能不是BNIP3L在肺癌中失活的机制。

Two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of human lung squamous carcinoma tissue and paired surrounding normal bronchial epithelial tissue were compared. Selected differential protein-spots were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both the tumor and the normal tissues were acquired. The average deviations of spot position were 0.8730.125mm in IEF direction and 1.0250.213mm in SDS–PAGE direction, respectively. For the tumor tissues, a total of 134967 spots were detected and 123548 spots were matched with an average matching rate of 91.5%. For the corresponding normal tissues, a total of 129773 spots were detected and 118356 spots were matched with an average matching rate of 91.2%. A total of 106945 spots were matched between the tumor and the normal tissues. Forty differential proteins between tumor and normal tissues were characterized. Some proteins were the products of oncogenes and others were involved in the regulation of cell cycle and signal transduction. These data are valuable for mass identification of differentially expressed proteins involved in lung carcinogenesis, establishing human lung cancer proteome database and screening molecular marker to further study human lung squamous carcinoma.

蛋白质组学一经出现，人们便开始尝试将其用于疾病特定分子标志物的研究当中，比如通过比较疾病与正常生理状况下蛋白质表达谱的差异来寻找与疾病密切相关的蛋白质。但采用该方法有一明显的缺点，即需要分析大量同一肿瘤组织的二维凝胶电泳（two-dimensional gel electrophoresis, 22DE）图谱，否则难以得到有统计学意义的差异蛋白质。另外，由于细胞内的蛋白质表达是不均匀的，这就导致了高表达和易溶解的蛋白质远较其他低表达、难溶解的蛋白质容易出现在22DE图谱中，因而明显地降低了计算机匹配分析发现有生物学意义的差异蛋白质的灵敏性。

毒蝇碱型乙酰胆碱受体（muscarinic acetylcholine receptor, mAChR）和Bcl22家族蛋白均具有调控神经细胞凋亡和生存的作用，然而mAChR 和Bcl22 家族蛋白之间的内在联系即信号转导通路仍然不清楚。为此，对mAChR 调控神经母细胞瘤SH2SY5Y细胞生存蛋白Bcl22 和磷酸化Bad 的信号转导通路进行了研究。结果显示：（1）mAChR激动剂卡巴可（carbachol）不仅活化SH2SY5Y细胞的MEK/ERK21/2，而且上调Bcl22和磷酸化Bad的表达；（2）mAChR拮抗剂阿托品、MEK抑制剂PD98059、PKC抑制剂bisindolymaleimide2I和Src抑制剂PP1均能完全阻断或显著减弱卡巴可的上述作用，但G蛋白脱偶联剂百日咳毒素和PI23激酶抑制剂wortmannin对卡巴可的上述作用无明显影响；（3）显性负突变Ras和Raf均能阻断卡巴可上调转染至SH2SY5Y细胞内的Bcl22启动子的转录调控活性。结果表明：mAChR通过Gq/11、PKC和Src依赖的Ras2ERK21/2信号转导通路上调SH2SY5Y细胞Bcl22和磷酸化Bad蛋白表达。这一研究将有助于揭示神经递质、神经营养因子和神经营养药物等抑制神经细胞凋亡、促进神经细胞生存的分子机制。

Differential proteome profiles of human lung squamous carcinoma tissue compared to paired tumor-adjacent normal bronchial epithelial tissue were established and analyzed by means of immobilized pH gradient-based two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). The results showed that well-resolved, reproducible 2-DE patterns of human lung squamous carcinoma and adjacent normal bronchial epithelial tissues were obtained under the condition of 0.75-rug protein-load. The average deviation of spot position was 0.733