Covalently closed circular DNA(cccDNA)in hepatocytes is the replication intermediate of hepa-titis B virus(HBV),which is closely related to HBV replication.Meanwhile,it is the transcriptional template of pregenomic RNA(pgRNA),and most of the HBV RNA in serum is derived from unreversed transcribed pgRNA.In recent years,many studies have demonstrated that HBV RNA has an important role in monitoring disease progression and predicting the prognosis of chronic HBV-infected patients,and is a potential biomarker of chronic viral hepatitis B.This article provides an overview of the molecular characterisation of HBV RNA,detection methods and research progress in clinical applications.

OBJECTIVE: To investigate the role of autophagy in lipopolysaccharide (LPS)-induced apoptosis of murine odontoblasts. METHODS: Murine odontoblasts (mDPC-23 cells) were treated with 5 μg/mL LPS for 6, 12 and 24 h, and the changes in cell viability was examined using CCK8 kit and cell apoptosis was detected by TUNEL staining. The changes in the protein levels of LC3, Beclin1, Atg5, AKT, p-AKT, mTOR and p-mTOR were detected using Western blotting. The effect of 3-MA treatment for 24 h on LPS-induced apoptosis of mDPC-23 cells was evaluated by detecting the expressions of apoptosis-related proteins caspase-3 and Bax using Western blotting. RESULTS: Stimulation with LPS for 6 and 12 h did not cause significant changes in the proliferation or apoptosis of mDPC-23 cells, but LPS treatment for 24 h significantly suppressed cell proliferation ( CONCLUSIONS LPS stimulation induces autophagy to promote apoptosis of mDPC-23 cells, and suppression of autophagy attenuates LPS-induced apoptosis. Autophagy may play an important role in the injury of inflamed pulp tissues.
Animals ; Apoptosis ; Autophagy ; Lipopolysaccharides/pharmacology* ; Mice ; Odontoblasts/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Signal Transduction

In recent years, many researchers have devoted themselves to the application of photodynamic therapy (PDT) in root canal disinfection, as conventional root canal disinfection methods have failed to achieve the optimal effect. Some clinicians have also applied PDT to root canal disinfection. PDT is expected to have a better effect than traditional root canal disinfection. This paper reviews the research progress on the mechanism, effect, influencing factors and limitations of PDT in root canal disinfection. Current research suggests that differences in the type and status of the bacteria, photosensitizers, light sources, operating environment and methods all affect the efficacy of root canal disinfection of PDT. Most of the research into PDT for root canal disinfection finds that it is effective, nontoxic, advantageous to dental pulp regeneration and comfortable for the patient, as well as lacking an excitant; however, its bactericidal effect is inferior to that of sodium hypochlorite. At present, it cannot replace traditional chemical washing but is a promising auxiliary method. The design of the photosensitizer, the energy dose of the light source and the optimal irradiation time need to be determined by further experiments, and more clinical verification is needed before its application in root canal therapy.

OBJECTIVE: To investigate the effects of autophagy on osteogenic differentiation of stem cells from the apical papilla (SCAPs) in the presence of tumor necrosis factor- (TNF-) stimulation . METHODS: SCAPs treated with TNF- (0, 5, and 10 ng/mL) with or without 5 mmol/L 3-MA were examined for the expression of autophagy marker LC3-Ⅱ using Western blotting. The cells were transfected with GFP-LC3 plasmid and fluorescence microscopy was used for quantitative analysis of intracellular GFP-LC3; AO staining was used to detect the acidic vesicles in the cells. The cell viability was assessed with CCK-8 assays and the cell apoptosis rate was analyzed using flow cytometry. The cells treated with TNF- or with TNF- and 3-MA were cultured in osteogenic differentiation medium for 3 to 14 days, and real- time PCR was used to detect the mRNA expressions of osteogenesis-related genes (ALP, BSP, and OCN) for evaluating the cell differentiation. RESULTS: TNF- induced activation of autophagy in cultured SCAPs. Pharmacological inhibition of TNF--induced autophagy by 3-MA significantly decreased the cell viability and increased the apoptosis rate of SCAPs ( < 0.05). Compared with the cells treated with TNF- alone, the cells treated with both TNF- and 3-MA exhibited decreased expressions of the ALP and BSP mRNA on days 3, 7 and 14 during osteogenic induction ( < 0.05) and decreased expression of OCN mRNA on days 3 and 7 during the induction ( < 0.05). CONCLUSIONS Autophagy may play an important role during the osteogenic differentiation of SCAPs in the presence of TNF- stimulation.
Autophagy ; drug effects ; physiology ; Cell Differentiation ; drug effects ; physiology ; Cell Survival ; drug effects ; Cells, Cultured ; Dental Papilla ; cytology ; Green Fluorescent Proteins ; Humans ; Osteogenesis ; physiology ; Stem Cells ; drug effects ; physiology ; Transfection ; Tumor Necrosis Factor-alpha ; administration & dosage ; antagonists & inhibitors ; pharmacology

Objective: To investigate the effects of leptin on the proliferation of stem cells from human stem cells from the apical papilla (hSCAPs) and the expression of osteogenic/dentinogenic genes in vitro to provide an experimental basis for the sustainable development of young permanent teeth. Methods : The tissue block method was used to isolate and culture hSCAPs from the apical papilla of the immature third permanent molar. The expression of leptin and OBRb in hSCAPs was detected using immunocytofluorescence staining, western blotting and reverse transcription polymerase chain reaction (RT-PCR) analysis. The hSCAPs was treated with 0.1 μg/mL of leptin (0.1 μg/mL group) or 1.5 μg/mL of leptin (1.5 μg/mL group) at different time points. The control group was treated with alpha-MEM medium. Cell proliferation was measured using the CCK8 assay and cell cycle analysis. QRT-PCR was used to detect the expression of related osteoblast/odontogenic genes for alkaline phosphatase (ALP), dentin matrix protein -1 (DMP-1), dentin sialophosphoprotein (DSPP), and osteocalcin (OCN) mRNA. The differences between the treatment groups and the control group were analyzed statistically using one-way ANOVA followed by Bonferroni analysis. Results: The expression of both leptin and OBRb were found in hSCAPs. Compared with the control group, the cell proliferation capacity and S phase cells in the treatment groups were higher than those in the control group, with the 1.5 μg /mL group displaying higher levels than 0.1 μg /mL group, and the treated hSCAPs demonstrated a higher proliferation rate and a higher expression of ALP, DSPP, and DMP-1 from day 3 to day 7, with the 1.5 μg /mL group displaying higher levels than 0.1 μg /mL group , and the difference was statistically significant (P < 0.05), at day 7. The treated hSCAPs demonstrated a lower expression of ALP, DSPP, and DMP-1. Compared with the control group, the treated hSCAPs demonstrated a higher expression of OCN from day 7 to day 14, with significantly higher expression in the 1.5 μg /mL group compared to the 0.1 μg /mL group. Conclusion Leptin may promote cell proliferation and upregulate the expression of relative osteogenic/dentinogenic genes.

Objective To explore the change and clinical significance of the serum carcinoembryonic antigen (CEA)and carbohydrate antigen 724(CA724)in patients with rectal cancer before and after neoadjuvant ra-diotherapy and chemotherapy.Methods The serum levels of CEA and CA724 of 30 patients with rectal carci-noma were detected by electrochemiluminescence method and were compared with those in 30 healthy people. Results The serum levels of CEA and CA724 in rectal carcinoma patients before neoadjuvant radiotherapy and chemotherapy were significantly higher than those in the healthy people,the difference was statistically significant(P<0.05).The serum levels in complete remission and partial remission patients after eight-week neoadjuvant radiotherapy and chemotherapy were significantly lower than those before treatment,the differ-ence was statistically significant(P<0.05),and the reduction levels were obviously higher than those in the stable group,the difference was statistically significant(P< 0.05).The CEA and CA724 levels in the stable treatment patients were also lower than those of before neoadjuvant radiotherapy and chemotherapy,the difference was statistically significant(P<0.05).The serum levels in the progression group were higher than those of before treatment(P<0.05).The serum levels of CEA and CA724 before neoadjuvant radiotherapy and chemotherapy existed a positive correlation(r=0.862,P=0.000).Conclusion The serum levels of CEA and CA724 in rectal cancer are highly expressed,suggesting that both of them are closely related to the occur-rence and development of rectal carcinoma,the measurement of the serum level changes of CEA and CA724 in patients with rectal cancer before and after treatment contributes to estimate the efficacy of neoadjuvant radio-therapy and chemotherapy and tumor progression.

Objective To investigate the expression of circulating tumor cells (CTC) in peripheral blood of patients with different stages of colorectal cancer (CRC), and to evaluate its significance in early diagnosis of colorectal cancer metastasis. Methods Sixty patients with CRC (42 inⅡ-Ⅲstage and 18 in IV stage ) and 30 patients with benign rectal disease were recruited from January 2014 to December 2015. The CTC in peripheral blood was purified with Immunomagnetic Separation Technologie, and detected by immunofluorescence in situ hybridization (imFISH). The serum levels of CEA were detected by electrochemiluminescence method meanwhile. The correlation between CTC and CEA was analyzed. Results CTC positive rates in CRC patients were significantly higher than those in benign rectal disease controls. CTC positive rates inⅣstage were significantly higher than those inⅡ-Ⅲ stage. The expression of CTC was significantly correlated with CEA (r = 0.6652, P < 0.01). Conclusions The expression of CTC in CRC patients is significantly higher than that in benign rectal disease control group. It is closely related to clinical stages. Detection of peripheral blood CTC has important clinical significance in the early diagnosis of CRC metastasis.

OBJECTIVETo investigate the effect and its possible mechanism of interleukin-17 (IL-17) on invasion and metastasis of human colon cancer cells.

METHODSIL-17 was added into the culture media of human colon cancer cells SW480 and LOVO. Cells were divided into 4 groups: SW480 control group (SW480 cells), LOVO control group (LOVO cells), SW480 experiment group (50 μg/L IL-17+SW480 cells), and LOVO experiment group (50 μg/L IL-17+LOVO cells). Cell growth was measured by CCK-8 assay. The proliferation rate(%)=[(Aexperiment group-Ablank)/(Acontrol group-Ablank)]×100%). The ability of cell invasion and migration was measured by transwell assay. Real time-PCR was used to detect mRNA expression of VEGF and MMP-9. Western blot was performed to detect protein expression of STAT3, p-STAT3, VEGF and MMP-9. Enzyme-linked immunosorbent assay (ELISA) was applied to measure the protein content of VEGF and MMP-9 in the supernatant.

RESULTSAfter cultivation for 24, 48 and 72 hours, CCK-8 assay revealed that the proliferation rate of SW480 was 1.18%±0.07%, 1.42%±0.09%, and 1.62%±0.08%; the proliferation rate of LOVO was 1.13%±0.02%, 1.32%±0.05% and 1.73%±0.02% in experiment group. Transwell experiments showed that after cultivation with IL-17 for 24 hours, number of invasion cell in experimental groups (SW480: 34.00±0.45, LOVO: 41.60±0.51) was higher as compared to corresponding control groups (SW480: 4.53±0.14; LOVO: 3.67±0.33) with significant differences (SW480: t=-76.026, P=0.001; LOVO: t=-81.580, P=0.005). The number of migration cell in experimental groups (SW480: 36.40±0.51, LOVO: 46.40±0.68) was higher as compared to corresponding control groups (SW480: 7.83±0.69; LOVO: 6.67±0.48) with significant differences (SW480: t=-51.542, P=0.003; LOVO: t=-49.265, P=0.005). Real-time PCR results revealed that after cultivation with IL-17 for 24 hours, VEGF and MMP-9 mRNA relative expression levels in experimental groups (SW480: VEGF:1.53±0.12, MMP-9: 2.44±0.23; LOVO: VEGF: 2.96±0.35, MMP-9: 3.38±0.55) were higher than those in control groups (both 1) with significant differences (VEGF: t=3.799, P=0.043; MMP-9: t=5.254, P=0.039). Western blot illustrated that after cultivation with IL-17 for 24 hours, STAT3, p-STAT3, VEGF and MMP-9 proteins relative expression levels in experimental groups were significantly higher that those in control groups (SW480:STAT3: t=3.233, P=0.023; p-STAT3: t=3.954, P=0.032; VEGF: t=3.201, P=0.025; MMP-9: t=3.154, P=0.029; LOVO: STAT3: t=3.788, P=0.012; p-STAT3: t=2.662, P=0.040; VEGF: t=4.118, P=0.035; MMP-9: t=4.268, P=0.030). ELISA indicated that content of VEGF and MMP-9 in the supernatant of experimental groups (SW480: VEGF 5 491.41±63.22, MMP-9: 21.43±1.35. LOVO: VEGF: 8 631.46±129.59, MMP-9: 178.32±3.20) were higher than those in control groups (SW480: VEGF:4 456.32±87.56, MMP-9:18.57±2.44. LOVO: VEGF: 8 122.38±108.66, MMP-9: 163.22±6.89) with significant differences (SW480: VEGF: t=6.993, P=0.037; MMP-9: t=5.587, P=0.040. LOVO: VEGF: t=7.013, P=0.044; MMP-9: t=6.762, P=0.043).

CONCLUSIONIL-17 may be able to activate STAT3 signal transduction pathway in vitro through up-regulation of VEGF and MMP-9 expression, thereby enhancing the invasion and migration of colon cancer SW480 and LOVO cells.

Cell Cycle ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Colonic Neoplasms ; pathology ; Humans ; Interleukin-17 ; pharmacology ; Matrix Metalloproteinase 9 ; metabolism ; Neoplasm Invasiveness ; Real-Time Polymerase Chain Reaction ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; Up-Regulation ; Vascular Endothelial Growth Factor A ; metabolism

Objective To investiagate cell apoptosis and expressions of Bax,Bcl-2 andCaspase-3 in gambogic acid-treated colorectal cancer cells. Methods SW480/LOVO colorectal cancer cells were treated by gambogic acid. Cell Counting Kit-8 assay (CCK-8) was used to test cell proliferation. Microscopy was used to check the morphological changes. Immunofluorescence staining technique was used to detect cell apoptosis. Expressions of Bax,Bcl-2 and Caspase-3 protein were detected by Western blot assay. Results Gambogic acid inhibited the proliferation of SW480/LOVO in a dose and time-dependent manner. Gambogic acid could induce cell apoptosis. Gambogic acid increased expressions of Caspase-3 and Bax, increased the ratio of Bax/Bcl-2, and decreased Bcl-2 protein expression. Conclusion Gambogic acid can inhibit proliferation and induce apoptosis of SW480LOVO cells, with the mechanism of up-regulation of Bax/Bcl-2 and activation of Caspase-3.

OBJECTIVETo observe the effect of revascularization for treatment of immature teeth with endodontic infection mediated by calcium hydroxide.

METHODSSeventeen pediatric patients with endodontic infections of the permanent teeth were treated with routine root canal and pulp cavity irrigation and disinfection followed by application of calcium hydroxide paste to the root canal orifice to induce revascularization. Another 17 patients received conventional apexification procedures to serve as the control group. The patients were followed up to observe the therapeutic effect of the treatments.

RESULTSIn the revascularization treatment group, 4 cases showed healed periapical lesions 6 to 18 months after the surgery with thickened root canal walls and closure of the apical foramen; in 10 cases, the periapical lesions healed 12 to 18 months postoperatively with lengthened root, thickened root canal wall, and narrowed apical foramen. One patient reported pain and swelling at 2 months, and 2 patients showed the formation of gum fistula and ceased development of the roots at 7 and 8 months. In the control group, the periapical lesions healed in 1 cases at 12 months postoperatively with apical foramen closure; in 11 cases, hard tissues formed in the root apex without obviously lengthened roots 6 to 8 months after the surgery; in 5 cases, no apical barrier formed even 12 to 18 months after the surgery. The overall effective rates were similar between the two groups (P>0.05).

CONCLUSIONSRevascularization by calcium hydroxide sealing can promote root development of immature permanent teeth with pulpitis or periradicular periodontitis.

Adolescent ; Calcium Hydroxide ; therapeutic use ; Child ; Dental Pulp ; blood supply ; Dental Pulp Diseases ; therapy ; Dentition, Permanent ; Guided Tissue Regeneration, Periodontal ; Humans ; Neovascularization, Physiologic ; drug effects ; Treatment Outcome