Nucleosome Positions and Differential Methylation Status of Various Regions within MLH1 CpG Island

Objective： To determine the relationship between nucleosome positions and formation of differential methylation of the reported region A, B, C, and D within the MLH1 CpG island. Methods： Methylation of the MLH1 promoter was analyzed by combined of bisulfite restriction assay. Chromatin of RKO and MGC803 cells were extracted and digested by MNase. Mononucleosomal DNA fragment was isolated and used as templates for detection of nucleosomal distribution by a battery of quantitative PCRs covering the full MLH1 promoter region. Results： The MLH1 was methylated in RKO and unmethylated in MGC803. At the region B, where methylation of CpG sites did not correlated with transcription of this gene well, qPCR product of the M-3 （-599nt ～ -475nt） fragment was amplified in both RKO and MGC803 cells. However, at the region C and D within the core promoter, where methylation of CpG sites correlated with loss of MLH1 transcription well, the M-7 （-257nt ～ -153nt） and M-8 （-189nt ～ -71nt） fragments were amplified remarkably only in RKO cells. Conclusion： Nucleosome may be the basic unit for both CpG methylation and methylation-related regulation of gene transcription. Methylation status of CpG sites within the same nucleosome may be homogeneous; between different nucleosomes, homogeneous or heterogeneous.

DNA COPY PROFILE IN NASOPHARYNGEAL CARCINOMA AND ITS CORRELATION WITH CLINICAL STAGING

Objective: To detect genetic alterations in nasopharyngeal carcinoma (NPC) in Cantonese, the population with the highest incidence of NPC, and to correlate the findings with clinical staging. Methods: Comparative genomic hybridization (CGH) was performed on 35 primary nasopharyngeal carcinomas and a nonparametric x2 test was used to analyze relationship between chromosome changes and clinical staging. Results: The identified common chromosomal alterations in NPC included gain of chromosomes 12q (21 cases, 60%), 4q (19cases, 43%), 3q (18 cases, 51%), 1q (15 cases, 43%),8q (14 cases, 40%), and 2q (12 cases, 30%). The most frequently detected loss of chromosomal materials involved chromosome 1p (24 cases, 69%), chromosome 3p (21 cases, 60%), 11q (20 cases, 57%), 14q (18 cases, 51%), 16q (14 cases, 40%), 13(12 cases, 34%), and 9p(11 cases, 31%). The high frequency (>50%) 4q gain and 1p loss were novel findings. Compared by nonparametric x2 test, gains on 12q and 8q were found mainly in stages III/IV and there were significant differences between two clinical stage groups (stages I/II vs stages III/IV). Conclusions: Current analysis has revealed a comprehensive profile of the chromosomal regions showing DNA copy number changes, which may harbor oncogenes or tumor suppressor genes involved in the development of primary NPC.

The therapeutic effect of Lamivudine on HBV-DNA in PBMC and its inducement effect against cytokine

The purpose of this investigation was to study the therapeutic effect of Lamivudine on HBVDNA in peripheral blood mononudear cells （PBMC） and serum, and the level of cytokines in serum of the patients with chronic hepatitis B. The patients were divided into two groups （A = 47, B = 34）, and treated by Lamivudine, routine medicine, respectively. The levels of HBV-DNA in PBMC and serum and eytokines were all detected before and after treatment. After the treatment of Lamivndine for 36 weeks, the total conversion negative rates of HBV-DNA in PBMC and serum of the patients treated with Lamivudine were 55.32% （26/47） and 61.70% （29/47）, respectively. The total negative conversion rates of HBV-DNA in PBMC and serum of the patients treated by routine medicine were 26.47 % （9/34） and 32.35% （11/34）, respectively. There was significant difference between Lamivudine group and routine medicine group （ P 〈 0.01 ）. The negative conversion rates of HBeAg in serum of the patients were 46.81% （22/47） and 68.09% （32/47） at the end of 24 weeks and 36 weeks, and were higher than those of routine medicine group （ P 〈 0.05 and P 〈 0.01 ）. The levels of alanine aminotransferase （ALT）, aspartate aminotransferase （AST）, ALT/AST in serum of the patients after being treated by Lamivudine, routine medicine were down-regulated to （30.1 ± 9.6） U/ml, （32.3 ±10.7） U/ml, 0.9 ±0.1 and （48.4±10.7） U/ml, （44.7 ±11.0） U/ml, 1.1 ±0.2. After the analysis of variance, the high significant difference was obvious between the two groups （ P 〈 0.01）. It was due to the high levels of IL-6, IL-8 and TNF-α in chronic hepatitis B which could be down-regulated to （250.5 ±33.3） pg/ml, （153.4 ±22.2） pg/ml, （232.6 ±21.2） pg/ml by Lamivudine, which was more obvious than that of routine medicine （ P 〈 0.01）. Lamivudine has high therapeutic effect on the treatment of HBVDNA in PBMC and serum and has better therapeutic effect than that of routine therapy. Lamivudine may also have higher down-regtdated inflananatory infdtration and secretion in local site caused by chemotactic cytokines and produce promotional effect on the recovery of liver function.