The Damage Of Genetic Material Of Human Bronchial Epithelial Cells And The Effects On The Expression Of Dna Repair Genes Induced By Glycidyl Methacrylate

Glycidyl methacrylate (GMA) is a unique of the family of acrylic monomers. It has two reactive functional groups: a very reactive epoxy group and an acrylic group. The dual functionality of GMA brings about the property of anti-ultraviolet radiation, water proof and heat resisting. So GMA has a wide application in military and civil industries.GMA was first detected in China in 1980s by our research group. And we first reported the mutation of GMA. With the support by National Science Found Committee (NSFC), our group carried out a series research on mutagenicity, carcinogenicity and their mechanism of GMA. The result was that GMA had the ability of covalent bonding with DNA. And GMA could induce mutation of procaryotic organism, chromosome aberration of mammalian cells, DNA damage of lymphocyte of human and rat, inhibition of semiconservation replication, malignant transformation of several types of mammalian or human cells, such as BALB/c 3T3 cell, SHE cell, human lung embryonic fibroblasts (HLEFs). All these results have demonstrated its carcinogenic potential.In vitro cell transformation test is an important method of study on chemical carcinogenesis and widely used in the mechanisms of malignant transformation. 16HBE, an SV40 large T antigen-immortalized human airway epithelial cell line, retains the phenotype of the differentiated lineage of the parent, contact inhibition and rarely produces tumors when injected into nude mice. It provides an ideal target cell for detecting carcinogenic potential of chemical.In this study, we observed the damage of genetic material of human bronchial epithelial cells (16HBE) induced by GMA, which include the examination of the damage of chromosome and mutation of DNA repair genes in the transformation cells. We also used gene chips of "Oligo GEArray(?) Human Genome Stability / DNA Repair Microarray" to detect the effects on the expression of DNA repair genes induced by GMA.The main results obtained were as follows:1. Study on the damage of genetic material of 16HBE induced by GMA1.1 The damage of chromosome induced by GMAChromosome aberration was detected in 16HBE cells after the short time exposure to GMA of various dosages (in 24 hours). And the dose-effect relationship was found. Compared to the group of DMSO, there were significant differences in the group of 16μg/ml and 20μg/ml. GMA could induce several kinds of chromosome structure aberration, such as the broken of chromosome, fragment, ring, double kinetochore, triradial chromosome and so on. And the aberration of chromosome amount was not found.We detected the chromosome aberration of 16HBE treated at the dose of 8μg/ml GMA for different times. It was indicated that the rate of chromosome aberration of the group exposure of GMA for 3 times can remarkably increased compared with the solvent control group (P<0.05). The rate of chromosome aberration increased as the longer of GMA exposure. And the aberration of chromosome amount was not found.There was not significant difference of chromosome structure aberration between the malignant transformation group and the control group (P>0.05). But the malignant transformation cells lack the normal nuclear style. The diploid reduced to 79%, while the hypodiploid and the hyperdiploid increased. There was significant difference of nuclear style between the transformation group and the control group (P<0.05).1.2 Point mutation of repair genes of transformation cells assayPoint mutation of DNA repair genes XRCC1, hMSH2, XPD and XRCC3 were detected with the method PCR-RFLP. We detect the gene type of the location C471A,G322A,IVS12-6 (T>C) and IVS1+9 (C>G) of hMSH2, C26304T,G27466A and G36189A of XRCC1, G23591A,A35931C of XPD, C18067T of XRCC3. And the method of DNA sequencing was used to check the result. The mutation of the location hMSH2 IVS12-6 (T>C) was observed. But the mutations in the other genes were not found.2. The effects on expression of DNA repair genes in 16HBE malignant transformation cells induced by GMAThe expression of DNA repair genes in 30th generation cells was detected with the gene chip of "human genome stability/DNA repair microarray". Eight genes with different expression were screened from 113 genes in the array, 7 of them expressing increasingly and 1 of them expressing decreasingly. According to the function categorization, genes with increased expression were double-strain broken repair gene NBS1, RAD50 and RAD51, base-excision repair gene OGG1, nucleotide excision repair gene XAB2, and gene about genome stabilization TNKS and TOP3A; while gene with decreased expression was base-excision repair gene NEIL2.Above all, the damage of genetic material and the effects on expression of DNA repair genes of 16HBE cells induced by GMA were researched on this study. The results were that GMA affected the function of the genes by damaging the cell chromosome. The defect of DNA repair system induced by GMA and the imbalance of injure- repair led to the cell transformation. The findings of this study were of great importance for further study on carcinogenic potential of mechanism of GMA, and useful for the occupational health monitor in the exposed workers.