| Background & Object Genomic imprinting is an epigeneticmodification in the germ line leading to preferential express one of the two parental alleles. The correct contribution of imprinted genes is required for normal embryonic and fetal development, cell differentiation and proliferation. Studies have shown evidences that genomic imprinting play an important role in carcinogenesis and could be regard as molecular marker for some malignant tumor. IGF2 is one of the imprinted genes in human genome, expressing from paternal allele in most tissues of normal adults. Rainier discovered loss of imprinting (LOI) result in IGF2 bi-allelic express and expression level increased many times in Wilms' tumor, which facilitating tumor progress. This suggests loss of imprinting is new mechanism in cancer occurrence and progress. Resent studies have shown LOI of IGF2 presence in colon cancer, bladder carcinoma, and ovarian tumor and so on. In present study, we will detect IGF2 gene allelic expression status in human primary hepatocellular carcinomas and matched non-cancerous liver tissues, to analyze the alternation of IGF2 geneimprinting status, and to find out their significance in hepatocarcinogensis, in molecular diagnosis, gene therapy of human primary HCC as well.Methods Surgery resected cancerous, and non-cancerous livertissue of 38 primary hepatocellular carcinoma patients were collected. Genomic DNA and total RNA were extracted from tissue specimens. Heterozygote of IGF2 was detected by Polymerase chain reaction ( PCR ) and restriction fragment length polymorphism ( RFLP ) with an Apa I polymorphism site of exon 9. Allelic expression was studied by semi-quantitative reverse transcription-polymerase chain reaction and RFLP. Data of heterozygote and allelic expression of IGF2 were statistically analyzed.Results 47.4% ( 18/38 ) of the specimens were IGF2 geneheterozygote, which could be used to allelic expression analysis. 50% (19/38) were homozygote. One case show heterozygote in cancerous tissue while homozygote in non-cancerous tissue in human primary hepatocarcinoma.10 cases of the 18 heterozygotes show IGF2 mRNA mono-allelic expression or mainly express from one allele, suggesting these cancerous tissues exhibit a gain of imprinting (GOI). 6 (60%) matched non-cancerous cirrhosis tissues among them also showed an expression from one allele suggesting they have a gain of imprinting, while other 4 matched normal liver tissues exhibiting bi-allelic expression. 6 (33.3%) cases of the 18 heterozygotes displayingbi-allelic expression of IGF2 mRNA both in cancerous and non-cancerous tissues, while 2 cases show bi-allelic expression in cancerous but mono-allelic expression in non-cancerous tissues.IGF2 mRNA expression level was analyzed using semi-quantitive RT-PCR, which displaying variation: IGF2 mRNA expression level of cancerous is obviously lower than that of non-cancerous in 4 cases (4/18, 22.2%); IGF2 mRNA expressions level in the cancerous and non-cancerous tissues were coincident in 6 cases (33.3%); And overexpression in cancerous tissues in 8 cases (44.4%). The average level of IGF2 expression in cancerous was (1.6104 ± 1.7991) Amax, 1.4 times higher than that in non-cancerous tissues, which was (1.1208 + 0.9159) Amax . But there is no statistically significant relation between expression level and imprinting status alternation in IGF2.Conclusion Gain of imprinting of 1GF2 is one of importantphenomena in cancerous and cirrhosis tissues of human primary hepatocarcinoma suggesting it may play a critical role in human hepatocarcinogensis. The alternation of IGF2 gene imprinting status dose not influences expression level directly. The mechanism of gain of imprinting of IGF2 and its significance in early molecular diagnosis and gene therapy of human primary HCC were to be elucidated by further investigation. |
PDF Full Text Request