Habib S L, Srikanth N S, Scappaticci F A, Faletto M B, Maccubbin A, Farber E, Ghoshal A K, Gurtoo H L
Grace Cancer Drug Center, Roswell Park Cancer Institute, Buffalo, New York 14263.
Toxicol Appl Pharmacol. 1994 Jan;124(1):139-48. doi: 10.1006/taap.1994.1017.
Levels of various cytochrome P450 proteins have been reported to be decreased to varying degrees in chemically induced hepatocyte nodules and following partial hepatectomy (PH). By screening a rat liver lambda ZAP cDNA expression library with antibodies raised against a partially purified preparation of cytochrome P450 isolated from untreated male Fischer 344 rats, we have isolated a 1.1-kb cDNA. This cDNA was sequenced for 139 bases from the 5' end of the sense strand and comparison of the resulting sequence with the sequences in Gene Man DNA data bank revealed 95% homology of the sequenced portion with male-specific rat cytochrome P450 (M-1, CYP IIC11). The 32P-labeled cDNA was used as a hybridization probe on RNA blots (Northern blots) prepared with total RNA from rat livers obtained post PH, from aflatoxin B1(AFB1)-induced rat liver tumors and from rat liver nodules induced with a combination of diethylnitrosamine/acetylaminofluorene/PH (DEN/AFF/PH). At 36 and 72 hr post PH, the mRNA level was decreased by > 93%. Relative to the corresponding control livers, the mRNA level was also decreased by 97% in the liver nodules and by 57% in AFB1-induced liver tumors. The RNA blots derived from the liver nodules and AFB1-induced liver tumors were also probed with a cDNA probe (R17) that recognizes other cytochromes P450 (CYP IIB1/CYP IIB2). The mRNA corresponding to CYP IIB1/CYP IIB2 was also depressed 92% in the nodules and 65% in the tumors. These results clearly indicate that the depression of both CYP IIC11 and IIB1/IIB2 in the hepatic nodules and the tumors is related to the inhibition of transcription and/or enhanced degradation of the mRNA.
据报道，在化学诱导的肝细胞结节以及部分肝切除（PH）后，各种细胞色素P450蛋白的水平会不同程度地降低。通过用针对从未经处理的雄性Fischer 344大鼠中分离出的部分纯化的细胞色素P450制剂制备的抗体筛选大鼠肝脏λZAP cDNA表达文库，我们分离出了一个1.1 kb的cDNA。对该cDNA从有义链的5'端进行了139个碱基的测序，并将所得序列与基因库DNA数据库中的序列进行比较，结果显示测序部分与雄性特异性大鼠细胞色素P450（M-1，CYP IIC11）有95%的同源性。用32P标记的cDNA作为杂交探针，用于对用PH后获得的大鼠肝脏、黄曲霉毒素B1（AFB1）诱导的大鼠肝肿瘤以及用二乙基亚硝胺/乙酰氨基芴/PH（DEN/AFF/PH）联合诱导的大鼠肝结节的总RNA制备的RNA印迹（Northern印迹）进行检测。在PH后36和72小时，mRNA水平降低了>93%。相对于相应的对照肝脏，肝结节中的mRNA水平也降低了97%，AFB1诱导的肝肿瘤中的mRNA水平降低了57%。还使用识别其他细胞色素P450（CYP IIB1/CYP IIB2）的cDNA探针（R17）对来自肝结节和AFB1诱导的肝肿瘤的RNA印迹进行检测。与CYP IIB1/CYP IIB2相对应的mRNA在结节中也降低了92%，在肿瘤中降低了65%。这些结果清楚地表明，肝结节和肿瘤中CYP IIC11和IIB1/IIB2的降低与mRNA转录的抑制和/或增强的降解有关。
