OBJECTIVETo investigate the effects and related mechanisms of hepatitis B virus X (HBx) protein on cell cycle and growth in hepatocellular carcinoma.

METHODSA human hepatocyte HepG2 cell line stably expressing a green fluorescent protein (GFP)-tagged HBx (HepG2/GFP-HBx cells) was used for the experiment, and HepG2 parental and HepG2/GFP cells was used as the controls. Effect of HBx on cell growth was evaluated by the MTT cell proliferation assay and on cell cycle progression by flow cytometry analysis of cells with or without treatment with 5-aza-2'-deoxycytidine (5-Aza-CdR; 5 pmol/L). Effect of HBx expression on promoter methylation status of the p16INK4A tumor-suppressor gene was detected by methylation-specific polymerase chain reaction and on p16 protein level was analyzed with western blotting.

RESULTSThe HepG2/GFP-HBx cells showed significantly higher cell proliferation at 72 hrs of culture (3.225+/-0.038 A490) than either control (HepG2: 2.012+/-0.022 A490, t = -46.86, P less than 0.001; HepG2/GFP: 2.038+/-0.029 A490, t = 42.51, P less than 0.001). The HepG2/GFP-HBx cells also showed significantly lower proportion of cells in the G0/G1 phase (16.45%+/-0.45%) than either control (HepG2: 44.81%+/-1.36%, t = -34.202, P less than 0.001; HepG2/GFP: 42.76%+/-1.58%, t = -28.88, P less than 0.001). However, 5-Aza-CdR treatment did lead to a significant amount of HepG2/GFP-HBx cells being arrested in the G0/G1 phase (33.25%+/-0.79%, t = 31.85, P less than 0.001). The p16INK4A promoter was methylated in the HepG2/GFP-HBx cells, and became demethylation after treatment with 5-Aza-CdR. However, no methylation of p16INK4A promoter was observed in both HepG2 and HepG2/GFP cells. The p16 protein level was significantly lower in the HepG2/GFP-HBx (vs. HepG2 and HepG2/GFP cells) and this level increased after treatment with 5-Aza-CdR.

CONCLUSIONHBx protein promotes hepatocellular carcinoma cell cycle progression and growth by shortening the G0/G1 phase, and the underlying mechanism may involve inducing p16INK4A promoter methylation and downregulating p16 protein expression.

Carcinoma, Hepatocellular ; metabolism ; pathology ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cyclin-Dependent Kinase Inhibitor p16 ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Genes, p16 ; Hep G2 Cells ; Hepatitis B virus ; metabolism ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Promoter Regions, Genetic ; Trans-Activators ; pharmacology

OBJECTIVETo investigate the anti-tumor effect of small interfering RNA targeting to HBV X gene (X-siRNA) and 5-aza-2'-deoxycytidine (5-aza-dC) on HBV-related hepatocellular carcinoma.

METHODSX-siRNA and control siRNA were synthesized. HepG2/GFP-HBx cells were treated with X-siRNA, and the levels of HBV X mRNA were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Nude mice were inoculated with HepG2/GFP and HepG2/GFP-HBx cells subcutaneous respectively to establish implant models of hepatocellular carcinoma, and were treated with X-siRNA, 5-aza-dC alone or in combination, and tumor growth was observed. The methylation of p16 gene promoter was detected by methylation specific polymerase chain reaction (MSP).

RESULTSRT-PCR showed the expression of HBV X mRNA in HepG2/GFP-HBx cells was inhibited markedly by X-siRNA. The nude mice experiment showed that the gross tumor volume was much bigger in HepG2/GFP-HBx group than that in HepG2/GFP group (P < 0.05). The growth of palpable tumors in X-siRNA or 5-aza-dC treatment group notably decreased (P < 0.05). MSP analysis showed that p16 gene methylation was observed in HepG2/ GFP-HBx-caused palpable tumors, while no methylation was detected in HepG2/GFP group. However, after treatment with X-siRNA or 5-aza-dC, p16 gene methylation reduced.

CONCLUSIONSHBV X-siRNA and methylation inhibitor can inhibit the growth of hepatoma cells via reversing p16 methylation.

Animals ; Antimetabolites, Antineoplastic ; pharmacology ; Azacitidine ; analogs & derivatives ; pharmacology ; DNA Methylation ; Genes, p16 ; Hep G2 Cells ; Humans ; Liver Neoplasms, Experimental ; therapy ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; RNA, Small Interfering ; Trans-Activators ; antagonists & inhibitors ; genetics

BACKGROUND: Unicompartment knee replacement is more popular for small trauma, rapid recovery, low less complications and almost normal knee mechanics, and has been used to repair unicompartmental knee diseases. At abroad, unicompartmental knee arthroplasty for advanced spontaneous osteonecrosis of knee (SONK) has obtained satisfactory outcomes, but its long-term efficacy and safety are not known in China. OBJECTIVE: To explore the short-term effectiveness of unicompartmental knee arthroplasty for advanced SONK. METHODS: Clinical data of 12 SONK patients (12 knees) admitted between January and August 2015 were analyzed retrospectively. Unicompartmental knee arthroplasty was operated by the same surgical team using the 3rdgeneration of Oxford?Unicompartmental Knee. The Visual Analogue Scale, femorotibial angle, range of motion of the knee and Hospital for Special Surgery scores were used to evaluate the curative efficacy at 3, 6, 12, and 18 months postoperatively. RESULTS AND CONCLUSION: (1) All patients were followed up for 12-18 months. The incision in all patients achieved primary union, and no infection, lower limb venous thrombosis or fracture occurred. (2) At the end of follow-up, the Visual Analogue Scale scores were significantly reduced from preoperative (6.67±0.78) to (1.75±0.97); the Hospital for Special Surgery scores were significantly increased from preoperative (63.92±7.27) to (91.67±2.87); the femorotibial angle changed from preoperative (178.28±3.38)° to (176.82±2.37)°(All P < 0.05). But the range of motion of the knee joint did not differ significantly before and after surgery. (3) That is to say, unicompartmental knee arthroplasty obtains satisfactory short-term efficacy in the treatment of advanced SONK.

ObjectiveTo explore the preventive effects of chest compression， abdominal compression and combined thoracoabdominal compression on propofol-induced respiratory depression during gastroscopy. MethodsA total of 544 patients underwent propofol sedation during gastroscopy in our hospital were randomly divided into 4 groups （n=136 each）： Group C （control group）， Group T （chest compression group）， Group A （abdominal compression group） and T&A （combined thoracoabdominal compression group）. Altogether 20 chest， abdominal and combined thoracoabdominal compressions were started respectively on patients in Group T， A and T&A before performing gastroscopy and after unconsciousness at a rate of 30 compressions per minute with a compression depth of 2~3 cm. The incidence of oxygen desaturation （SpO₂2<95%） and hypoxemia （SpO₂<90%） were compared among the groups. Incidence of gastroscope withdrawal and B-mode ultrasound assessment of diaphragm activity were observed. ResultsThere were statistical differences in the incidence of oxygen desaturation and hypoxemia among the four groups （P<0.05）. The incidence of oxygen desaturation in Group T and A were significantly lower than that in Group C （P<0.017）. B-ultrasonography revealed that the diaphragm activity in Group T and T&A were significantly higher than that in Group C （P＜0.01） and the patients in Group T had the highest diaphragm activity. ConclusionBoth chest compression and abdominal compression can reduce the incidence of respiratory depression induced by propofol in patients during gastroscopy， and chest compression is more effective.

Humans ; Retrospective Studies ; Tetanus/epidemiology* ; China/epidemiology*

Objective: To estimate the incidence of HIV-1 infection in men who have sex with men (MSM) in key areas of China through HIV-1 limiting antigen avidity enzyme immunoassay (LAg-Avidity EIA), analyze the deviation from the actual results and identify influencing factors, and provided reference for improving the accuracy of estimation results. Methods: Based on the principle of the cohort randomized study design, 20 cities were selected in China based on population size and the number of HIV-positive MSM. The sample size was estimated to be 700 according to the HIV-1 infection rate in MSM. MSM mobile phone app. was used to establish a detection appointment and questionnaire system, and the baseline cross-sectional survey was conducted from April to November 2019. LAg-Avidity EIA was used to identify the recent infected samples. The incidence of HIV-1 infection was calculated and then adjusted based on the estimation formula designed by WHO. The influencing factors were identified by analyzing the sample collection and detection processes. Results: Among the 10 650 blood samples from the participants, 799 were HIV-positive in initial screening, in which 198 samples (24.78%) missed during confirmation test. Only 621 samples were received by the laboratory. After excluding misreported samples, 520 samples were qualified for testing. A total of 155 samples were eventually determined as recent infection through LAg-Avidity EIA; Based on the estimation formula , the incidence of HIV-1 infection in MSM in 20 cities was 4.06% (95%CI:3.27%-4.85%), it increased to 5.53% (95%CI: 4.45%-6.60%)after the adjusting for sample missing rate. When the sample missing rate and misreporting rate were both adjusted, the incidence of HIV-1 infection in the MSM increased to 5.66% (95%CI:4.67%-6.65%). The actual incidence of HIV-1 infection in MSM in the 20 cities might be between 4.06% and 5.66%. Conclusions: Sample missing and misreporting might cause the deviation of the estimation of HIV-1 infection incidence. It is important to ensure the sample source and the quality of sample collection and detection to reduce the deviation in the estimation of HIV-1 infection incidence.
Cross-Sectional Studies ; HIV Infections/epidemiology* ; HIV-1 ; Homosexuality, Male ; Humans ; Immunoenzyme Techniques ; Incidence ; Male ; Sexual and Gender Minorities

Objective: To investigate the distribution of HIV-1 genetic subtypes and pretreatment drug resistance (PDR) among men who have sex with men (MSM) from 19 cities of 6 provinces in China. Methods: From April to November 2019, 574 plasma samples of ART-naive HIV-1 infected MSM were collected from 19 cities in Hebei, Shandong, Jiangsu, Zhejiang, Fujian, and Guangdong provinces, total ribonucleic acid (RNA) was extracted and amplified the HIV-1 pol gene region by nested polymerase chain reaction (PCR) after reverse transcription. Then sequences were used to construct a phylogenetic tree to determine genetic subtypes and submitted to the Stanford drug resistance database for drug resistance analysis. Results: A total of 479 samples were successfully amplified by PCR. The HIV-1 genetic subtypes included CRF01_AE, CRF07_BC, B, CRF55_01B, CRF59_01B, CRF65_cpx, CRF103_01B, CRF67_01B, CRF68_01B and unrecognized subtype, which accounted for 43.4%, 36.3%, 6.3%, 5.9%, 0.8%, 0.8%, 0.4%, 0.4%, 0.2% and 5.5%, respectively. The distribution of genetic subtypes among provinces is statistically different (χ²=44.141, P<0.001). The overall PDR rate was 4.6% (22/479), the drug resistance rate of non-nucleoside reverse transcriptase inhibitors, nucleoside reverse transcriptase inhibitors, and protease inhibitors were 3.5% (17/479), 0.8% (4/479) and 0.2% (1/479), respectively. The PDR rate of recent infections was significantly higher than that of long-term infections (χ²=4.634, P=0.031). Conclusions: The HIV-1 genetic subtypes among MSM infected with HIV-1 from 19 cities of 6 provinces in China are diverse, and the distribution of subtypes is different among provinces. The overall PDR rate is low, while the PDR rate of recent infections was significantly higher than that of long-term infections, suggesting the surveillance of PDR in recent infections should be strengthened.
China/epidemiology* ; Cities ; Drug Resistance ; Drug Resistance, Viral/genetics* ; Female ; Genotype ; HIV Infections/epidemiology* ; HIV Seropositivity/drug therapy* ; HIV-1/genetics* ; Homosexuality, Male ; Humans ; Male ; Phylogeny ; Reverse Transcriptase Inhibitors/therapeutic use* ; Sexual and Gender Minorities