Risk factors for sporadic colorectal cancer in southern Chinese

AIM:To investigate the role of smoking, alcohol drinking, family history of cancer, and body mass index(BMI) in sporadic colorectal cancer in southern Chinese.METHODS:A hospital-based case-control study was conducted from July 2002 to December 2008.There were 706 cases and 723 controls with their sex and age(within 5 years) matched.An unconditional logistic regression model was used to analyze the association between smoking, alcohol drinking, family history of cancer, BMI and sporadic colorectal cancer.RESULTS:No positive association was observed between smoking status and sporadic colorectal cancer risk.Compared with the non alcohol drinkers, the current and former alcohol drinkers had an increased risk of developing sporadic colorectal cancer(CRC)(adjusted OR = 8.61 and 95% CI = 6.15-12.05;adjusted OR = 2.30, 95% CI = 1.27-4.17).Moreover, the increased risk of developing sporadic CRC was significant in those with a positive family history of cancer(adjusted OR = 1.62, 95% CI = 1.12-3.34) and in those with their BMI ≥ 24.0 kg/m2(adjusted OR = 1.39, 95% CI = 1.10-1.75).Stratification analysis showed that the risk of developing both colon and rectal cancers was increased in current alcohol drinkers(adjusted OR = 7.60 and 95% CI = 5.13-11.25;adjusted OR = 7.52 and 95% CI = 5.13-11.01) and in those with their BMI ≥ 24.0 kg/m2(adjusted OR = 1.38 and 95% CI = 1.04-1.83;adjusted OR = 1.35 and 95% CI = 1.02-1.79).The risk of developing colon cancer, but not rectal cancer, was found in former alcohol drinkers and in those with a positive family history of cancer(adjusted OR = 2.51 and 95% CI = 1.24-5.07;adjusted OR = 1.82 and 95% CI = 1.17-2.82).CONCLUSION:Alcohol drinking, high BMI(≥ 24.0 kg/m2) and positive family history of cancer are the independent risk factors for colorectal cancer in southern Chinese.

Alterations of FHIT Gene and P16 Gene in Nickel Transformed Human Bronchial Epithelial Cells

Objective To study the alterations of FHIT gene and P16 gene in malignant transformed human bronchial epithelial cells induced by crystalline nickel sulfide using an immortal human bronchial epithelial cell line, and to explore the molecular mechanism of nickel carcinogenesis. Methods 16HBE cells were treated 6 times with different concentrations of NiS in vitro, and the degree of malignant transformation was determined by assaying the anchorage-independent growth and tumorigenicity. Malignant transformed cells and tumorigenic cells were examined for alterations of FHIT gene and P16 gene using RT-PCR, DNA sequencing, silver staining PCR-SSCP and Western blotting. Results NiS-treated cells exhibited overlapping growth. Compared with that of negative control cells, soft agar colony formation efficiency of NiS-treated cells showed significant increases (P<0.01) and dose-dependent effects. NiS-treated cells could form tumors in nude mice, and a squamous cell carcinoma was confirmed by histopathological examination. No mutation of exon 2 and exons 2-3, no abnormal expression in p16 gene and mutation of FHIT exons 5-8 and exons 1-4 or exons 5-9 were observed in transformed cells and tumorigenic cells. However, aberrant transcripts or loss of expression of the FHIT gene and Fhit protein was observed in transformed cells and tumorigenic cells. One of the aberrant transcripts in the FHIT gene was confirmed to have a deletion of exon 6, exon 7, exon 8, and an insertion of a 36 bp sequence replacing exon 6-8. Conclusions The FHIT gene rather than the P16 gene, plays a definite role in nickel carcinogenesis. Alterations of the FHIT gene induced by crystalline NiS may be a molecular event associated with carcinogen, chromosome fragile site instability and cell malignant transformation. FHIT may be an important target gene activated by nickel and other exotic carcinogens.

Over-expressed Genes Detected by Suppression Subtractive Hybridization in Carcinoma Derived From Transformed 16HBE Cells Induced by BPDE

To screen the over differentially expressed genes in carcinoma induced by BPDE-transformed 16HBE cells （16HBE-C cells）. Methods The suppression subtractive hybridization （SSH） method was performed to profile differentially expressed genes between 16HBE-C cells and 16HBE cells. The cDNA fragments of differentially expressed genes were inserted into TA cloning vector and transformed competent E. coli strain. Positive clones were randomly picked up and identified by the colony PCR method. Dot blot was used to test the same source with the tester. The differentially expressed cDNA fragments were sequenced and compared with known genes and EST database in Genbank. Results Eight known genes were over-expressed in 16HBE-C cells including eukaryotic translation elongation factor 1 alpha 1, HIF-1 responsive RTP801, ribosomal protein L10 （RPL10）, ribosomal protein S29 （RPS29）, mitochondrion related genes, and laminin receptor 1. Three differentially expressed cDNA fragments could not be matched to the known genes but to the EST database. Conclusion The SSH method can detect differentially expressed genes between 16HBE-C and 16HBE cells. BPDE-induced carcinogenesis may be related to alteration of at least eight known genes and three unknown genes. These expression data provide a clue to further cloning novel genes and studying functions in BPDE-induced carcinoma.

ANTITRANSFORMING ACTIVITY OF CHLOROPHYLLIN AGAINST SELECTED CARCINOGENS AND COMPLEX MIXTURES

Chlorophyllin,the sodium and copper salt of chlo-rophyll,has been shown by several investigators tobe an antimutagenic agent.In a previous study wefound that it is a potent inhibitor of the mutagenicactivity of dietary and environmental complex mixtu-res.No information is available,however。

Recombinant GrB and PFP Co-expression in Hep-2 Cells

Objective： To achieve the co-expression of GrB and PFP in Hep-2 cells and analyze the growth inhibiting effects on Hep-2 cells. Methods： Lymphocytes were separated from human laryngeal carcinoma tissue, complete Exon fragments of GrB and PFP were amplified by RT-PCR via extracting lymphocytes total RNA, and they were recombined to the downstream of T7 promoter in the vector pVAX1. The recombinant plasmid pVAX1-PIG was transfected into Hep-2 cells with Lipofectamine 2000. The expression of proteins was identified by RT-PCR, MTT and western blot assay. Results： The gene sequence of the RT-PCR products of GrB and PFP were consistent with the data of GenBank by DNA sequencing analysis. The GrB and PFP cDNA fragment were cloned into the vector of pVAX1 in the right direction and the open reading fragment of GrB and PFP were maintained. The target proteins were detected in the transfected Hep-2 cells, and the inhibitive effect of PFP and GrB on Hep-2 cells growth were studied by thiazolyl blue （MTT） test. Conclusion： The pVAX1-PFP-IRES-GrB plasmid was successfully constructed and expressed, and the expression of PFP and GrB could inhibit the growth of Hep-2 cells.

Expression Silence of DNA Repair Gene hMGMT Induced by RNA Interference

Objective： MGMT protein expression has been associated with tumor resistance to alkylating agents. The objective of this paper is to construct the RNA interference vector which can specifically induce the expression silence of human DNA repair gene hMGMT. Methods： The hMGMT specific siRNA expression cassette was made by two steps PCR, linked with pUCI 9 to get pU6-MGMTi, co-transfected with pEGFP-CI into 16HBE and screened by G418. The MGMT mRNA and protein levels were detected by RT-PCR and Western Blot respectively. Results： hMGMT specific RNA interfere vector pU6-MGMTi was constructed successfully. In transfected 16HBE cells MGMT mRNA level could hardly be detected and the protein level was only 10% of control. Conclusion： MGMT specific RNAi expression cassette can effectively inhibit MGMT expression. MGMT silence cell line was built by co-transfection technology, which offered condition for studying the gene function of MGMT.

Abnormal expression of c-Myc in human bronchial epithelial cells malignantly transformed by anti-BPDE

Anti-benzo[a]pyrene-7,8-diol-9,10-epoxide(anti-BPDE)is a metabolite of benzo[a]pyrene(B[a]P)and acts as a potent mutagen in mammalian systems.However,molecular mechanisms related to anti-BPDE-induced carcinogenesis are poorly understood.Here,we investigated the expression of proto-oncogene c-myc in human bronchial epithelial cells(16HBE-T)transformed by exposure to anti-BPDE.The levels of mRNA and pro-tein of c-Myc were examined in the 16HBE-T and vehicle-treated control cells(16HBE-N)by using different meth-ods respectively,including reverse transcriptase-polymer-ase chain reaction(RT-PCR),quantitative real-time PCR(Q-PCR),western blot and immunocytochemical meth-ods.The level of c-myc mRNA appeared to be signifi-cantly increased in 16HBE-T,as compared with those of the 16HBE-N.Likewise,the expression of c-Myc protein was significantly enhanced as compared with those of the control cells.Moreover,the localization of c-Myc protein shows mainly nuclear staining in 16HBE-T.In conclu-sion,the abnormal expression of c-Myc was present in anti-BPDE malignantly transformed 16HBE cells,which may be involved in the carcinogenesis molecular mech-anism of anti-BPDE.