Browsing by Subject "DNA fragment"

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Open Access
Common telomerase reverse transcriptase promoter mutations in hepatocellular carcinomas from different geographical locations
(WJG Press, 2015) Cevik, D.; Yildiz G.; Ozturk, M.
AIM: To determine the mutation status of human telomerase reverse transcriptase gene (TERT ) promoter region in hepatocellular carcinoma (HCC) from different geographical regions. METHODS: We analyzed the genomic DNA sequences of 59 HCC samples comprising 15 cell lines and 44 primary tumors, collected from patients living in Asia, Europe and Africa. We amplified a 474 bp DNA fragment of the promoter region of TERT gene including the 1295228 and 1295250 sequence of chromosome 5 by using PCR. Amplicons were then sequenced by Sanger technique and the sequence data were analyzed with by using DNADynamo software in comparison with wild type TERT gene sequence as a reference. RESULTS: The TERT mutations were found highly frequent in HCC. Eight of the fifteen tested cell lines displayed C228T mutation, and one had C250T mutation with a mutation frequency up to 60%. All of the mutations were heterozygous and mutually exclusive. Ten out of forty-four tumors displayed C228T mutation, and additional five tumors had C250T mutation providing evidence for mutation frequency of 34% in primary tumors. Considering the geographic origins of HCC tumors tested, TERT promoter mutation frequencies were higher in African (53%), when compared to non-African (24%) tumors (P = 0.056). There was also a weak inverse correlation between TERT promoter mutations and murine double minute 2 single nucleotide polymorphism 309 TG polymorphism (P = 0.058). Mutation frequency was nearly two times higher in established HCC cell lines (60%) compared to the primary tumors (34%). CONCLUSION: TERT promoter is one of most frequent mutational targets in liver cancer, and hepatocellular carcinogenesis is highly associated with the loss of telomere-dependent cellular senescence control. © The Author(s) 2015.
Open Access
The role of bcl-2 family of genes during kindling
(Blackwell Publication, 2005) Akcali, K. C.; Sahiner, M.; Sahiner, T.
Purpose: Several experimental models of human temporal lobe epilepsy have shown that apoptotic death of neurons is an important part of this degenerative disease. However, the role of apoptotic regulators is not clear during the epileptogenesis. Therefore we investigated the expression pattern of bcl-2 family of genes during the formation of kindling model of epilepsy in rats. Methods: We examined the expression pattern of bax, bcl-2, bcl-xL, mtd, and bcl-w both at messenger RNA (mRNA) and protein level in the brain tissues during the formation of epilepsy with kindling model in adult rats, which has been the most acceptable form of experimental model of human epilepsy. We also assessed the onset of DNA fragmentation by using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. Results: Animals have started to have epileptic discharges after day 10 of kindling model. Recurrent subthreshold electrical stimuli induced not only epileptic foci but also the expression of box, an inducer of apoptosis, in this time period. Conversely, bcl-xL, which is an inhibitor of apoptosis, had an opposite pattern of expression both at mRNA and protein level during the formation of epilepsy. We did not observe DNA fragmentation by TUNEL staining. Conclusions: Our study shows differential expression of Bax and Bcl-x L at the CA1 region during the formation of hippocampal kindling model. The absence of DNA fragmentation during this period suggests that epileptic changes in neurons have the potential to induce DNA fragmentation by altering the expression levels of Bax and Bcl-xL.