miR-155-5p increases the sensitivity of liver cancer cells to adriamycin by regulating ATG5-mediated autophagy.

Liver cancer is the sixth most prevalent cancer worldwide and the third leading cause of cancer-related deaths. Adriamycin (ADR) resistance, which often leads to the progression of malignant tumors, is a major treatment obstacle for liver cancer. It has been confirmed that miR-155-5p could reverse drug resistance in human breast cancer. However, the biological function of miR-155-5p in ADR-resistant liver carcinoma (HepG2/ADR) cells remains unclear. miR-155-5p and ATG5 expression was determined by RT-qPCR and western blot. In addition, MTT, flow cytometry, immunofluorescence staining, and western blotting were performed to evaluate the proliferation, apoptosis, and autophagy of liver cancer cells. Finally, the effect of miR-155-5p on the expression of autophagy-related 5 (ATG5) was analyzed by luciferase activity assay, western blot, and RT-qPCR. Our results showed that miR-155-5p was downregulated in HepG2/ADR cells. Increasing the expression of miR-155-5p enhanced the sensitivity of liver carcinoma cells to ADR and promoted apoptosis through inhibition of autophagy in vitro. In addition, the binding site between miR-155-5p and ATG5 was identified, and miR-155-5p could directly regulate ATG5. Finally, ATG5 partially rescued the effect of miR-155-5p on autophagy and the apoptosis of HepG2/ADR cells. In conclusion, our findings showed that miR-155-5p could reverse ADR resistance in liver cancer by targeting ATG5, which may function as a potential target for liver cancer treatment.

miR-34a-5p suppresses the invasion and metastasis of liver cancer by targeting the transcription factor YY1 to mediate MYCT1 upregulation.

BACKGROUND: In recent years, microRNAs (miRNAs) are reported to act as molecular biomarkers for cancer diagnosis, treatment, and prognosis (including liver cancer) and to be involved in the development and progression of cancer and other physiological and pathological changes. However, the role of miR-34a-5p in liver cancer is still largely unknown. METHODS: In our study, the expression of miR-34a-5p in liver cancer tissues and HCC cell lines was detected by qRT-PCR. The CCK-8, scratch wound-healing motility and Transwell assays were used to evaluate the effect on cell proliferation, migration and invasion. The expression of YY1, E-cadherin, N-cadherin and vimentin was analysed by western blotting. The dual luciferase assay was performed to confirm whether YY1 is a target of miR-34a-5p. The combination of YY1 and MYCT1 was detected by chromatin immunoprecipitation (ChIP) assay. RESULTS: The results showed that miR-34a-5p was downregulated in liver cancer tissues and HCC cell lines. Overexpression of miR-34a-5p inhibited the proliferation, migration and invasion of liver cancer cells. YY1 was a direct target of miR-34a-5p, and the expression of YY1 could reverse the influence of miR-34a-5p on the proliferation, migration and invasion of liver cancer cells. YY1 inhibited MYCT1 expression by directly binding to its promoter region, and knockdown of MYCT1 reversed the influence of miR-34a-5p on the proliferation, migration and invasion of liver cancer cells. CONCLUSION: Our results suggest that miR-34a-5p could inhibit the invasion and metastasis of hepatoma cells by targeting YY1-mediated MYCT1 transcriptional repression.

microRNA-329 suppresses epithelial-to-mesenchymal transition and lymph node metastasis in bile duct cancer by inhibiting laminin subunit beta 3.

Bile duct cancer (BDC), also known as cholangiocarcinoma, is a highly desmoplastic cancer with a growth pattern characterized by periductal extension and infiltration. Studies have suggested that microRNAs (miRNAs) play an important role in BDC progression. Here we aim at investigating the effects of miR-329 on BDC development, focusing especially on epithelial-to-mesenchymal transition (EMT) in vitro and lymph node metastasis in vivo. Expression microarrays associated with BDC tissues were collected and differentially expressed genes were analyzed, followed by miRNA target prediction and verification. The role miR-329 played in BDC was examined using gain-of-function and loss-of-function methods. The expressions of miR-329, laminin subunit beta 3 (LAMB3), and EMT markers, in addition to cell proliferation, migration, and invasion were evaluated. Furthermore, nude mice models of BDC were established to observe tumor growth and metastatic lymph nodes. The LAMB3 was identified as an upregulated gene based on the GSE77984 and GSE45001 microarray analysis. LAMB3 was also predicted and confirmed to be a target gene of miR-329 by dual-luciferase reporter assay. Through further cell experiments, the EMT process was reversed, cell proliferation, invasion, and migration were suppressed, when miR-329 was upregulated. Furthermore, in vivo experiments exhibited that the overexpression of miR-329 inhibited tumor growth and the number of metastatic lymph nodes. This study provides in vivo and in vitro evidence that miR-329 inhibits BDC progression through translational repression of LAMB3. Therefore, the obtained results may aid as an experimental basis for improving prognosis of BDC.

Decrease of miR-622 expression promoted the proliferation, migration and invasion of cholangiocarcinoma cells by targeting regulation of c-Myc.

OBJECTIVE: To explore the mechanism of miR-622 in regulating the proliferation, migration and invasion of cholangiocarcinoma (CCA) cells. MATERIALS AND METHODS: Quantitative real-time PCR was conducted to measure the expression of miR-622 and c-Myc in CCA tissues and cell lines. Protein level of c-Myc was measured by Western blot. The effect of miR-622 on cell proliferation, migration and invasion was analyzed by MTT assay and Transwell chamber migration assay. Luciferase reporter assay was performed to measure the effect of miR-622 on c-Myc. RESULTS: miR-622 expression was downregulated in both CCA tissues and cell lines, while c-Myc expression was uregulated. Overexpression of miR-622 in CCA cells was statistically correlated with a decrease of cell proliferation, migration and invasion, while inhibition of miR-622 made an inverse result. We also proved c-Myc was identified as a target gene of miR-622 in CCA. Moreover, we found overexpression of c-Myc can strengthen the effects of miR-622 on the proliferation, migration and invasion of CCA cells. CONCLUSION: Decrease of miR-622 promotes the proliferation, migration and invasion of CCA cells by directly targeting c-Myc.

Glutathione-s-transferase A 4 (GSTA4) suppresses tumor growth and metastasis of human hepatocellular carcinoma by targeting AKT pathway.

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies of cancers and its prognosis remains dismal due to the paucity of effective therapeutic targets. Up-regulation of glutathione-s-transferase A 4 (GSTA4) is associated with poor prognosis of HCC, but its functional mechanism in HCC remains unclear. In this study, we investigated the roles of GSTA4 in tumor growth and metastasis of HCC and found that GSTA4 was frequently up-regulated in HCC tissues. Through gain- and loss-of-function studies, GSTA4 was demonstrated to significantly regulate cell proliferation, migration, and invasion in vitro. Furthermore, GSTA4 overexpressing significantly promoted the tumorigenicity and metastasis of HCC cells in nude mice models bearing human HCC, whereas silencing endogenous GSTA4 caused an opposite outcome. Moreover, we demonstrated that GSTA4 enhanced HCC aggressiveness by activating protein kinase B (AKT) signaling. In multivariate analysis, our results GSTA4 overexpression promotes the progression of hepatocellular carcinoma and might represent a novel therapeutic target for its treatment.

[Preliminary study of wireless biofeedback therapy for treatment of bruxism].

OBJECTIVE: To access the effect of wireless biofeedback therapy on bruxism. METHODS: Ten voluntary bruxers (seven female and three male, mean age 26.1 years) were invited to participate in this clinical research. An electric resistance strain gauge was embedded in the position of canine of a maxillary splint for monitoring the abnormal clenching or grinding movement of teeth during sleep. The relevant details of bruxism events, including value of relative force, occurring time and duration were recorded and analyzed by the receiver device and monitoring program respectively. Meanwhile, for the purpose of nerve system and muscle relaxation, a watch-style device around the patient's wrist will vibrate to alert the patient of teeth grinding or clenching if the value of biting force and duration exceed the threshold. Total average episodes of bruxism and duration was observed during eight hours sleep, and was analyzed with one-way analysis of variance in SPSS 19.0 by the end of 6th week and three months following biofeedback therapy. RESULTS: The average episodes of bruxism has declined dramatically from (9.8 ± 2.2) times to (3.0 ± 1.2) times during one night (P < 0.05), and the average duration of bruxism events was reduced from (20.7 ± 12.2) s to (10.0 ± 3.4) s (P < 0.05) after six weeks biofeedback therapy. By the end of three months, the average episodes declined to (2.9 ± 1.2) times (P < 0.05), and the average duration decline to (9.2 ± 2.9) s (P < 0.05) with contrast to preliminary night. CONCLUSIONS: The pressure-based wireless biofeedback device is able to monitoring clenching and grinding of bruxism. The results suggest that biofeedback therapy may be an effective, novel and convenient measure for treatment of bruxism according to several months therapy.

[Gene therapy for tongue carcinoma mediated by oral administration of attenuated Salmonella harboring tip30 and IFN-γ genes in nude mouse model].

PURPOSE: To observe the effect of gene therapy mediated by oral administration of attenuated Salmonella carrying tip30 and IFN-γ genes in human tongue carcinoma nude mouse model. METHODS: 25 four-week-old BALB/C male nude mice were divided randomly into 5 groups based on the differently harboring genes in the recombinant attenuated Salmonella typhimuriums SL7207, which included blank group (PBS), control group (SL7207), IFN treatments group (SL7207-pCI-IFN), tip30 treatment group (SL7207-pCI-tip30) and the combination of IFN and tip30 treatment group (SL7207-pCI-tip30/IFN). On 10d after submandibular subcutaneous injection of Tca8113 cells into the mice, the recombinant attenuated Salmonella typhimuriums were orally administrated 3 times at 1 week interval. The treatment effect indicators included the growth curve, the tumor inhabitation rate, the survival rate, the apoptosis typical DNA ladder and the protein expressions of tip30 and IFN-γ in tumor cells. Statistical analysis was performed with SPSS15.0 software package. RESULTS: The growth curve, the tumor inhabitation rate and the survival rate indicated that the tumor in the PBS group constantly grew up and induced the animal death in earlier time, while the SL7207 treatment had a slight inhibiting effect compared with PBS group. However, the SL7207-pCI-IFN group and the SL7207-pCI-tip30 had a moderate inhibiting effect compared with the SL7207 group, while the combination of IFN and tip30 had the strongest inhibiting effect. The protein expressions of tip30 and IFN-γ were detected in tumor cells while the typical DNA ladders of apoptosis were observed only in the tip30 gene transfer groups (SL7207-pCI-tip30 and SL7207-pCI-tip30/IFN). CONCLUSIONS: The recombinant attenuated Salmonella typhimurium possesses a capacity of gene transfer in vivo through oral administration. The combination of the harboring tip30 and IFN-γ genes have synergistic inhibiting tumor effect for tongue carcinoma.

[Regulation of PTHrP in proliferation and differentiation of chondrocytes of condyle in fetal mouse].

OBJECTIVE: To investigate the regulatory mechanism of parathyroid hormone-related protein (PTHrP) in proliferation and differentiation of chondrocytes of condyle in fetal mouse. METHODS: Chondrocytes of condyle in fetal mouse were separated and cultured in vitro, the influence of PTHrP on proliferation and differentiation was observed. RESULTS: After two weeks' culture in 0.01 nmol/L, 0.1 nmol/L, 1 nmol/L, 10 nmol/L human PTHrP, there was significant difference in the number of cartilage nodule formed between experiment group and control group (P<0.05), and there was no significant difference in 0.01 nmol/L group (P>0.05). Alkaline phosphatase (ALP) activity was significantly intensified in experiment group and control group (P<0.05). Meanwhile, it was found that this function of promotion was lessened after anti-PTHR antibody used. CONCLUSION: It can be seen that PTHrP, via its receptor, can promote proliferation and differentiation of chondrocytes of condyle, which resemble its modulation mechanism in epiphyseal growth plate cartilage intramembrane in mandibule.

[A study of electric information monitoring in the treatment of bruxism with occlusal splint].

OBJECTIVE: An electric monitor of bruxism had been invented in order to evaluate the curative effect of cuspid occlusal upheaving splint and stabilization splint. METHODS: 20 patients with bruxism were randomly divided into two groups. A cuspid occlusal upheaving splint or a stabilization splint was fabricated respectively for patients. The vertical dimension for each splint was 0.5 mm lower than mandibular postural position. Sleeping time, bruxist time and times of bruxism were recorded with bruxism monitor that was invented for studying bruxing. RESULTS: The bruxist time and the times of bruxism were decreased obviously in patients with cuspid occlusal upheaving splint, while no significant difference was shown before and after using the stabilization splint. CONCLUSIONS: The bruxism monitor can automatically measure and record the data of bruxism with splint, which is valuable for clinic. The curative effect of cuspid occlusal upheaving splint is better than that of stabilization splint for treating bruxism.