The change of HBV nucleotide sequence during Adefovir dipivoxil treatment / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the change of HBV nucleotide sequence during Adefovir dipivoxil treatment.</p><p><b>METHODS</b>Serum samples were collected from 4 patients (numbers 228, 229, 230 and 233) with chronic hepatitis B (CHB) treated with PMEA. PCR amplification of full-length genomes, total genome clone, and sequencing and linkage of the HBV viruses were done with the serum samples collected before the therapy, and again on the 28th week, 52nd week and on the 92nd week of the therapy. Gene types and serum types were studied according to the HBV DNA sequence. The related biological information was analyzed, including the homogeneity and heterogeneity of the HBV DNA sequence before and during PMEA therapy, and variance of amino acids coded by HBV sequence.</p><p><b>RESULTS</b>The gene types and serum types of HBV of the 4 patients were determined. Patient 228 was gene type C/serum type adrq+. Patient 229 was complex gene type B and C/complex serum type adw2 and adrq+. Patients 230 and 233 were both serum type adw2. The homogeneity of total gene sequence was 97.5% - 99.8% in one group of patients and 90.6% - 100% between groups in variant patients. The base mutation quantity of 52W was significantly higher than that of 28W and 92W. There was a common site of hotspot congregating mutation that existed in the P region. The target of PMEA was located in the P region encoding reverse transcriptase, which was generally very conservative. Therefore the mutations in this region were very significant. This might affect the activity of reverse transcriptase, leading to resistance to PMEA.</p><p><b>CONCLUSION</b>We discovered some unusual aminophenol variations in the HBV genomes, which most probably are related to the HBV mutation that resulted in the developing resistance to PMEA.</p>

SHP-1 gene's methylation status of Daudi lymphoma cell and the demethylation effect of 5-aza-2'-deoxycytidine / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the transcription regulation of 5-aza-2'-deoxycytidine(5-Aza-CdR) on SHP-1 gene and its effects on Daudi cell line growth.</p><p><b>METHODS</b>MTT method and flow cytometry were used to detect the growth and apoptosis of Daudi cells after treated with different dosage of 5-Aza-CdIR. Bisulfite sequencing PCR ( BSP) , T-A cloning and sequence analysis were evaluated for methylation status. The SHP-I mRNA and protein were determined by reverse transcription polymerase chain reaction (RT-PCR) ,immunohistochemistry.</p><p><b>RESULTS</b>(1)After 7 d treatment with 2. 00 micromol/L of 5-Aza-CdR, all cytosines (C) in Daudi cells genome DNA were converted to thymidine, and SHP-1 mRNA and protein expressed again in the cells while those Cs in CpG dinucleotides in untreated Daudi cells remained Cs; (2)5-Aza-CdR inhibited the cell growth,The effects within certain extent were dose and time dependent:after 72 h treatment with 5-Aza-CdR at 200. 00, 20. 00, 2. 00 and 0. 20 micromol/L, the inhibitive rates were 72. 0% , 65. 1%, 51. 5%, 28.8% ,23.4% respectively; (3) 5-Aza-CdR increased apoptosis rate of tumor cells with a dose and times dependent manner within certain extent, too:at the 1,3,5 d treatment with 5-Aza-CdR 2. 00 micromol/L,the apoptosis rates were 2. 3% ,10. 8 % and 17. 1% ; respectively. (4) 5-Aza-CdR also changed cell cycle of tumor cells: at 24 h treatment with 5-Aza-CdR 2.00 micromol/L,92. 7% tumor cells stopped at S phase and G, phase cells were increased gradually with time.</p><p><b>CONCLUSION</b>DNA promoter hypermethylation is associated with SHP-1 gene silence in Daudi lymphoma cell line. 5-Aza-CdR could effectively cause demethylation and inhibit the growth of tumor cell by reactivating the gene transcription.</p>