63. Study on the deletions of the FHIT gene mRNA in murine lung cancer induced by Coal Tar Pitch

Epidemiological studies have shown that people who were exposed to Coal Tar Pitch (CTP) had a higher incidence of lung cancer than control population. In addition, CTP could induce two kinds of lung cancer in rats and mice. Although CTP is widely considered as a definite carcinogen, the mechanism of carcinogenesis is still unknown. Ohta discovered a new gene-FHIT (Fragile histidine triad) by exon-capture technique in 1996. It was reported that this gene was closely associated with the environmental carcinogen. FHIT gene is a focus for the study of environmental carcinogen acted upon. In our experiment, CTP was heated directly to produce fume to set an environment simulating workplace. Experimental mice were forced to inhale fume of CTP to induce model of lung cancer. Through series killing, the deletion of FHIT gene during tumor genesis was studied. The purpose of this study is to investigate whether the molecular genetic event can be used to detect primary lung cancer. It can also provide useful information to infer the FHIT changes in human lung tumor genesis. Methods and materials: 64 Kunming mice were provided by Henan Laboratory Animal Center. They were 4 weeks old and half were male. All the mice were randomly divided into experimental and control groups. Each group had 32 mice. Experimental material was Coal tar pitch, which was produced by Beijing Certain Steel Factory. Experimental mice were put into 50 liter sealed vessel, which the fume was blown into by circulating system, the CTP was heated by electric hot plate at 400℃ until CTP did not produce fume. During experimental period, mouse's diet habit and activities were recorded. The mice in experimental group were exposed to CTP fume for 12 weeks, then half of the mice were executed after stopping inhale CTP fume, the rest were executed in 24th weeks. The wet weight of lung and liver were measured. The specimen of lung was divided into two parts. One part was fixed by 10% formalin. The other was frozen in liquid nitrogen. The murine cDNA sequence was found in GenBank. The primers were devised by Primer Devising software. The targeted DNA fragment was amplified by nested-PCR. The expected length of product was 694bp. Results: ①The diet habit and activities of mice in experimental group were normal, the weights of the mice in the experimental groups were 32.83±5.40 gram and 38.34±5.47 gram in 12th weeks and 24th weeks respectively. There were significant loss compared to the control group mice. The index of lung and body in experimental group was increased significantly compared to control group. ②The main histology type of lung cancer induced by CTP fume was carcinoid (87.5%). In experimental group, there were 2 cases of carcinoid occurred (2/16) in 12th week, whereas there were 5 cases and 1 case of adenocarcinoma occurred in 24th weeks. However, there was no any cancer occurred in control group. ③Deletions of FHIT transcripts were found in the tissue of precancerous lesion, tumor lesion and its adjacent tissues. Nested-PCR results show that 52% PCR products exhibited aberrant bands in the exposure group. Among 8 mice with tumors, aberrant bands were shown in 4 carcinoids mice, 5 of 7 precancerous specimens showed aberrant bands. Except for the wild-type 694bp band, there were one or two aberrant bands, which were shorter than normal band. The length of deletions was 100bp or so. Aberrant transcriptions were not detected in control group. Conclusions: ①The main histological type of lung cancer induced by CTP was carcinoid. This finding indicates that the CTP fume can specifically cause mouse carcinoid. ②Aberrant FHIT transcripts were detected consistently, from precancerous lesion to tumor specimen, which suggests that abnormality of FHIT gene transcript is an early and frequent molecular genetic event and this change can last to the occurrence of tumor. The finding is valuable for early diagnosis.