ABSTRACT
<p><b>OBJECTIVE</b>To study the effects of human telomerase reverse transcriptase (hTERT) RNA interference (RNAi) on biological characteristics of hepatocellular carcinoma cell lines HepG2 and SMMC-7721 and on apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL).</p><p><b>METHODS</b>Small hairpin hTERT (shTERT) sequence was identified by PCR method; hTERT expressions, morphological features, cell proliferation and replicative senescence were respectively determined using RT-PCR, hematoxylin-eosin (HE) staining, growth curve and beta-galactosidase (b-Gal) staining; cell cycle and apoptosis were identified using flow cytometry after propidium iodide (PI) staining and annexin V/PI double staining.</p><p><b>RESULTS</b>shRNA were found in 6/8 HepG2 and 6/6 SMMC-7721 cell clones transformed by the recombined plasmid pSilencer 3.1-H1 neo-shTERT. The interference rates of hTERT on HepG2 and SMMC-7721 were 100% and 43.3% respectively. Cells in G2-M phases increased from 7.1% to 10.6% and from 6.9% to 7.9% respectively; and the percentage of replicative senenscence cells increased from 0 to 20.4% and from 3.6% to 10.0% respectively. The nucleus/cytoplasm ratios of the cells were obviously decreased after hTERT RNAi treatment. Moreover, apoptosis of hepatocellular carcinoma cells and apoptosis induced by TRAIL were strikingly increased by hTERT RNAi (P < 0.05). For example, apoptosis rates were increased from 3.5% to 5.2% in HepG2 cells and from 4.8% to 7.9% in SMMC-7721 cells after hTERT RNAi treatment. Apoptosis rates were increased from 5.3% to 10.4% in HepG 2 cells and from 13.9% to 77.2% in SMMC-7721 cells after being treated by 100 ng/ml TRAIL for 24 h. However, there were no remarkable changes between control cells and untransformed cells.</p><p><b>CONCLUSION</b>hTERT RNAi not only has a significant effect on biological characteristics of hepatocellular carcinoma cells, but also obviously can increase cell apoptosis induced by TRAIL.</p>
Subject(s)
Humans , Apoptosis , Carcinoma, Hepatocellular , Pathology , Liver Neoplasms , Pathology , RNA Interference , RNA, Small Interfering , Genetics , TNF-Related Apoptosis-Inducing Ligand , Pharmacology , Telomerase , Genetics , Tumor Cells, CulturedABSTRACT
<p><b>OBJECTIVE</b>To explore the relationship between methylenetetrahydrofolate reductase (MTHFR) gene polymorphism and microsatellite instability (MSI) in patients with gastric cancer.</p><p><b>METHODS</b>MTHFR gene C677T and A1298C polymorphism were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and MSI was examined with PCR.</p><p><b>RESULTS</b>MTHFR gene C677T and A1298C polymorphisms were analyzed on 122 gastric cancers and 110 normal controls The genotype frequencies of MTHFR 677CC, 677CT and 677TT were 47.5%, 39.3% and 13.1% on patients with gastric cancer, and 48.5%, 42.6%, 8.9% in the controls respectively. There was no significant difference of genotype frequency between the two groups (P > 0.05). The individuals with 677CT genotype, 677TT genotype and 677CT + TT genotype exhibited significantly reduced risk (OR = 0.38,95% CI: 0.15-0.98; OR = 0.26,95% CI: 0.03-2.18 and OR = 0.36,95% CI: 0.07-0.98) of developing gastric cardia cancer compared with those harboring the wild-type(677CC). The individuals with 677TT genotype having a 3.03-fold (95% CI: 1.07-8.65) increased risk of developing gastric corpus cancer. The genotype frequency of MTHFR 1298AA, 1298AC and 1298CC were 59.8%, 36.1% and 4.1% in gastric cancer patients, and 57.4%, 7.6%, 5.0% in the controls, respectively. The distribution of MTHFR A1298C genotype was not significantly different between gastric cancer and controls (P > 0.05). The individuals with 1298CC genotype had a reduced risk of developing gastric antrum cancer (OR = 0.41- fold, 95% CI: 0.03-2.18, 0.05-3.72) when comparing with those having 1298AA genotype. Patients with MSI+ gastric cancer had an increased frequency of the MTHFR 677TT genotype when comparing with those suffering from MSI- gastric cancer (P = 0.009) and with controlled subjects (P = 0.008). There was no significant association found between MTHFR A1298C polymorphism and MSI (P>0.05).</p><p><b>CONCLUSION</b>Polymorphism of MTHFR C677T was associated with increased risk on gastric corpus cancer and reduced risk on gastric cardia cancer. The polymorphism of MTHFR A1298C was associated with reduced risk for gastric antrum cancer while MSI pathway was possibly involved in the development of gastric cancer with MTHFR 677TT genotype.</p>
Subject(s)
Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Case-Control Studies , Gene Frequency , Genotype , Methylenetetrahydrofolate Reductase (NADPH2) , Genetics , Microsatellite Instability , Polymorphism, Genetic , Stomach Neoplasms , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To evaluate the relationship between mitochondrial DNA instability (mtMSI) and interleukin-8 (IL-8) activity in gastric mucosa of various lesions.</p><p><b>METHODS</b>IL-8 level in gastric mucosa was assayed using ELISA method. The mtMSI was detected by PCR-SSCP techniques.</p><p><b>RESULTS</b>mtMSI was observed in 11 out of 30 (36.7%) gastric cancers, 2 of 15 (13.3%) intestinal metaplasia, 2 of 10 dysplasia and 1 of 10 chronic atrophic gastritis. IL-8 level in mtMSI+ group [(76.8 +/- 3.8) pg/mg] was significantly higher than that in mtMSI- group [(48.3 +/- 3.6) pg/mg, P < 0.05].</p><p><b>CONCLUSION</b>mtMSI closely correlates with IL-8 level in gastric mucosa and is involved in gastric carcinogenesis.</p>
Subject(s)
Humans , DNA, Mitochondrial , Genetics , Enzyme-Linked Immunosorbent Assay , Gastric Mucosa , Metabolism , Pathology , Gastritis, Atrophic , Genetics , Metabolism , Genomic Instability , Interleukin-8 , Metabolism , Metaplasia , Genetics , Metabolism , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational , Precancerous Conditions , Genetics , Metabolism , Stomach Neoplasms , Genetics , MetabolismABSTRACT
Objective To assess mucin gene expression in Barrett's esophagus.Methods Mucin core protein-MUC1,MUC2,MUC3,MUCSAC and MUC6 were detected by immunohistochemistry.The re- lationship between mucin expression and magnification-endoscopic characteristics,pathohistologic epithelial types of Barrett's esophagus was analyzed.Results Mild expression of MUC1 was predominantly found in the superficial epithelium of both gastric and specialised intestinal metaplasia.In a small number of specimens, mild expression of MUC1 was also noted in glands.Strong MUC2 expression was noted only in the goblet cells in Barrett's oesophagus.MUC3 was expressed in the superficial columnar cells of specialized intestinal metaplasia with or without globlet cells but not in gastric metaplasia of the oesophagus.In some specimens MUC3 was expressed in the vacuolus of the globlet cells and the lumen of gland.Strong staining of MUCSAC was noted in the columnar epithelium of both gastric metaplasia and specialized intestinal metaplasia in Barrett's oesophagus,as well as expressed in the cytoplasm and vacuolus of the globlet cells in some speci- mens.Expression of MUC6 protein was detected at the basement of the crypts in gastric metaplasia and spe- cialised Barrett's glands.Expression of MUC2 and MUC3 protein was found much higher in villous or irregu- lar pit pattern than that in dot or rod pit pattern(P