Ying-xin Zhang

The protective effect of 17β-estradiol against hydrogen peroxide-induced apoptosis on mesenchymal stem cell.

This study was designed to investigate the function of 17β-estradiol (17β-E2) against oxidative stress on the cell death of mice bone marrow mesenchymal stem cells (BMSCs) induced by hydrogen peroxide (H₂O₂). BMSCs were treated with 17β-E2 for 24h and then treated with 100μM H₂O₂ for 1h. Cell viability, apoptosis, caspase-9 mRNA, JNKs (Jun N-terminal kinases) and c-Jun protein expression in BMSCs were evaluated. Cell apoptosis of BMSCs were increased in a dose-dependent manner after treated with H₂O₂ compared to control group. But pretreatment with 17β-E2 can inhibit apoptosis of BMSCs, preserve the mitochondrial transmembrane potential, decrease caspase-9 mRNA, JNK1/2 and c-Jun protein expression. In conclusion, 17β-E₂ exerts antiapoptotic effects in BMSCs which related to the mitochondria death pathway and JNKs pathway. The study revealed that 17β-E₂ can reduce the donor stem cells damage.

Upregulated ROS production induced by the proteasome inhibitor MG-132 on XBP1 gene expression and cell apoptosis in Tca-8113 cells.

Exposure of Tca-8113 cells to proteasome inhibitor carbobenzoxy-Leu-Leu-leucinal (MG-132) causing apoptosis is associated with endoplasmic reticulum (ER) stress. X-box-binding protein-1 (XBP1) is an important regulator of a subset of genes active during ER stress, which is related to cell survival and is required for tumor growth. The present study is to evaluate the effect of MG-132 on ROS production, XBP1 gene expression, tumor necrosis factor receptor-associated factor 2 (TRAF2), ASK1 and c-jun protein expression in tongue squamous cell carcinoma cell line Tca-8113 cells. ROS production was measured by reactive oxygen species assay. X-box binding protein-1 (XBP1) mRNA was analyzed by real-time-PCR, TRAF2, ASK1 and c-jun protein were investigated by western blot and immunocytochemistry respectively. The result indicated that ROS production, TRAF2, ASK1 and c-jun were elevated in MG-132 treated cells. Giving ROS scavenger N-acetyl-L-cysteine (NAC) largely prevented the effects of MG-132. Furthermore, treating with MG-132 lead to decreased XBP1 mRNA expression but could not completely block the expression of XBP1. Taken together, these findings provide the evidence that MG-132 induced ER stress lead to Tca-8113 cells apoptosis through ROS generation and TRAF2-ASK1-JNK signal pathway activation.

The molecular mechanisms of XBP-1 gene silencing on IRE1α-TRAF2-ASK1-JNK pathways in oral squamous cell carcinoma under endoplasmic reticulum stress

BACKGROUND Proteasome inhibitor Carbobenzoxy-Leu-Leu-leucinal (MG132) induces the unfolded protein response (UPR) in oral squamous cell carcinoma (OSCC). X-box binding protein 1 (XBP1) is a key UPR component that regulates endoplasmic reticulum stress (ER) homeostasis. This study was aimed to investigate the activation of IRE1α-TRAF2-ASK1-JNK pathway by silencing the XBP1 expression in an OSCC cell line. METHODS The XBP1 specific short hairpin RNA (shRNA) plasmid vector was constructed and then transfected into the Tca-8113 cells. The effect of XBP-1 gene silencing on IRE1α-TRAF2-ASK1-JNK pathway under MG132 induced endoplasmic reticulum stress in Tca-8113 were investigated by real-time RT-PCR or western blot. Cell apoptosis was detected by flow cytometry. RESULTS XBP1 expression was reduced in transfected groups and MG132 groups. shRNA-XBP1 induces IRE1α-TRAF2-ASK1 signaling activation to activate pro-apoptotic ASK1-JNK signaling. Moreover, combined shRNA-XBP1 with MG132 further enhanced downregulated XBP1 expression and upregulated activation of ASK1-JNK signaling. CONCLUSIONS Silencing XBP1 expression under MG132 induced ER stress block the XBP1 survival pathway and synergism with MG132 to promote Tca8113 cell apoptosis. These findings provide a therapeutic option in oral squamous cell carcinoma by inhibition of proteasome and XBP1 splicing.

Effectiveness and safety of chemotherapy combined with cytokine-induced killer cell /dendritic cell-cytokine-induced killer cell therapy for treatment of gastric cancer in China: A systematic review and meta-analysis

BACKGROUND Currently, cytokine-induced killer cells (CIK)/dendritic cell (DC)-CIK-mediated immunotherapy is widely used to treat gastric cancer. However, limited information regarding clinical trials on CIK/DC-CIK therapy is available. Therefore, systemic evaluation of the efficacy and safety of the combination therapy is necessary. METHODS A meta-analysis involving 1735 patients with gastric cancer was conducted. Before analysis, the study quality and heterogeneity were evaluated. The effects of chemotherapy combined with CIK/DC-CIK on gastric cancer were compared with the effects observed when chemotherapy alone was used. Pooled analysis was performed using RevMan version 5.2 from random or fixed-effect models. RESULTS Seventeen trials were included. First, the analysis showed that the combination therapy significantly increased the overall survival rate and disease-free survival rate compared with those in patients treated using chemotherapy alone. The overall response rate (P = 0.002), disease control rate (P = 0.0007), and quality of life improved rate (P = 0.0008) were significantly improved in patients who received combined treatment than in patients who received chemotherapy alone. Second, the percentage of lymphocyte subsets (CD3(+), CD4(+) and CD3(-)CD56(+), CD3(+)CD56(+); P <0.01) and the levels of interleukin-12 and interferon-γ, which reflect immune function, were significantly increased (P <0.05) after the CIK/DC-CIK therapy. Further, carbohydrate antigen tumor markers were significantly reduced compared with the pre-therapy levels. Immunotherapy with CIK/DC-CIK obviously alleviated the adverse events caused by chemotherapy. CONCLUSION The combination of CIK/DC-CIK therapy and chemotherapy was superior in prolonging the survival time, enhancing immune function and alleviating the adverse events caused by chemotherapy.

The stimulatory effects of eNOS/F92A-Cav1 on NO production and angiogenesis in BMSCs.

BACKGROUND Nitric oxide (NO) is generated in endothelial cells by endothelial nitric oxide synthase (eNOS). Caveolin-1 (Cav1) inhibits eNOS function and NO production. Modifying Cav1 scaffold domain, in particular Phenylalanine at position 92 (F92) is critical for the inhibitory actions of Cav1 toward eNOS. The aims of this study were to investigate the effect of enhanced NO production in term of in vitro angiogenesis on rat bone marrow derived mesenchymal stem cells (BMSCs) transduced with a novel bicistronic lentiviral vector co-expressing eNOS and mutant Cav1 (F92A). METHODS A bicistronic eNOS/F92-Cav1 lentiviral vector was constructed, and used to transduce rat BMSCs. The expression of eNOS and VEGF protein were confirmed by western-blot. NO production was detected by the greiss assay and in vitro angiogenesis was assessed by matrigel assisted capillary tube formation. The cell viability was evaluated using a Cell Counting Kit (CCK)-8. RESULTS The bicistronic eNOS/F92A-Cav1 lentiviral vector increased eNOS and VEGF protein expression, NO production compared to controls. In vitro capillary formation was increased in eNOS-F92A transduced cells and cell viability was not affected by transduction. CONCLUSION Transduction of rat BMSCs with an eNOS-F92A-Cav1 lentiviral vector can increase NO production by enhancing eNOS protein expression. The increased NO production did not reduce cell viability. This study demonstrates that genetic modification of BMSCs to enhance NO producton could be applied in stem cell based therapeutic approaches to treat diseases such as pulmonary arterial hypertension (PAH) which is characterized by decreased endothelial NO release.

Effect of short hairpin RNA-induced CXCR4 silence on ovarian cancer cell.

This study was aimed to investigate the effect of down-regulating the CXC chemokine receptor-4 (CXCR4) expression on cell proliferation, invasion and migration of human ovarian cancer cell line SW626. The CXCR4 specific short hairpin RNA (shRNA) plasmid vector was constructed and then transfected into the SW626 cells. The expression of CXCR4 mRNA and protein was detected by real-time RT-PCR and western blot respectively. Cell proliferation was detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Cell invasion and migration was assayed in Biocoat Matrigel invasion chambers. The expression level of CXCR4 in SW626 cell transfected with CXCR4-siRNA was inhibited, leading to significant decrease in SW626 cell proliferation, invasion and migration. We conclude that CXCR4 is essential for tumor cell proliferation and invasion. The CXCR4 molecule is a potential therapeutic target to control ovarian cancer cell growth or metastasis.

Apoptotic effect of MG-132 on human tongue squamous cell carcinoma

The aim of this study was to investigate the apoptotic effect of a proteasome inhibitor MG-132 on Tca-8113, a cell line of human tongue squamous cell carcinoma. Tca-8113 cells were treated with 10, 20, and 30μM of MG-132, or 5μM thapsigargin. Apoptosis rate was determined with annexin V/propidium iodide double staining. Expression of E3ubiquitin-protein ligase was determined by ELISA, and Grp78 and caspase-12 mRNA, and Grp78 and caspase-12 protein was assessed by RT-PCR and Western blot, respectively. Apoptosis was observed 18h after MG-132 treatment. The apoptotic rate in the 10, 20, and 30μM MG-132 group was 13.5, 19.6 and 34.7%, respectively, which was higher than in the thapsigargin (8.5%, P<0.01) or control group (0.5%, P<0.01). The expression of E3 ubiquitin-protein ligase in the 10, 20, and 30μM MG-132 group was 28.75±2.28, 18.16±0.65, 8.85±0.72, respectively, which was lower than in the thapsigargin (38.96±0.33, P<0.05 or 0.01) or control (40.88±4.52, P<0.05 or 0.01) group. The levels of Grp78 and capase-12 mRNA, Grp78 and caspase-12 protein in the MG-132 groups were higher than in the control group (P<0.01). In conclusion, MG-132 induces apoptosis in Tca-8113 cells in a concentration-dependent manner. The MG-132-induced apoptosis may involve downregulation of E3 ubiquitin ligase, and upregulation of Grp78 and caspase-12.

Cytokine-induced killer cells/dendritic cells-cytokine induced killer cells immunotherapy combined with chemotherapy for treatment of colorectal cancer in China: a meta-analysis of 29 trials involving 2,610 patients

Purpose To systematically evaluate the efficacy and safety of Cytokine-induced killer cells/dendritic cells-cytokine induced killer cells (CIK/DC-CIK) immunotherapy in treating advanced colorectal cancer (CRC) patients. Results 29 trials including 2,610 CRC patients were evolved. Compared with chemotherapy alone, the combination of chemotherapy with CIK/DC-CIK immunotherapy significantly prolonged the overall survival rate (OS) and disease-free survival rate (DFS) (1–5 year OS, P < 0.01; 1-, 2-, 3- and 5-year DFS, P < 0.01). The combined therapy also improved patients’ overall response, disease control rate and life quality (P < 0.05). After immunotherapy, lymphocyte subsets percentages of CD3+, CD3−CD56+, CD3+CD56+ and CD16+CD56+ (P < 0.01) and cytokines levels of IL-2 and IFN-γ (P < 0.05) were increased, while CD4+, CD8+ and CD4+CD25+ and IL-6 and TNF-α did not show significant change (P > 0.05). Materials and Methods Clinical trials reporting response or safety of CIK/DC-CIK immunotherapy treating advanced CRC patients and published before September 2016 were searched in Cochrane Library, EMBASE, PubMed, Wanfang and CNKI database. Research quality and heterogeneity were evaluated before analysis. Pooled analyses were performed using random or fixed-effect models. Conclusions The combination of CIK/DC-CIK immunotherapy and chemotherapy prolong CRC patients’ survival time, enhanced patients’ immune function and alleviates the adverse effects caused by chemotherapy.

Overexpression of tumor suppressor gene ZNF750 inhibits oral squamous cell carcinoma metastasis

Zinc-finger protein 750 (ZNF750) encodes a putative C2H2 zinc finger protein and is typically mutated or deleted in squamous cell carcinoma. The role of ZNF750 in oral squamous cell carcinoma (OSCC) remains unknown. The aim of the present study was to investigate the effects of ZNF750 overexpression in CAL-27 cells. Cell viability, and the expression of genes associated with proliferation, differentiation and the epithelial-mesenchymal transition were investigated in CAL-27 cells following ZNF750 overexpression, using Cell Counting kit-8, reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. In addition, scratch wound, invasion and migration assays were performed. Cell viability, matrix metalloproteinase 28 expression, cyclin B1 expression and mesenchymal marker neural cadherin expression were decreased following ZNF750 overexpression compared with the control groups. ZNF750 overexpression induced the differentiation-associated genes late cornified envelope 3A and small proline-rich protein 1A and upregulated the expression of late epidermal differentiation factor Kruppel-like factor 4. Overexpression of ZNF750 in CAL-27 cells resulted in inhibition of cell invasion and migration. Taken together, these data suggest that ZNF750 may inhibit the metastasis of OSCC.

Cytokine-induced killer cells/dendritic cells and cytokine-induced killer cells immunotherapy for the treatment of esophageal cancer in China: a meta-analysis

Background Immunotherapy based on cytokine-induced killer cells or combination of dendritic cells and cytokine-induced killer cells (CIK/DC-CIK) showed promising clinical outcomes for treating esophageal cancer (EC). However, the clinical benefit varies among previous studies. Therefore, it is necessary to systematically evaluate the curative efficacy and safety of CIK/DC-CIK immunotherapy as an adjuvant therapy for conventional therapeutic strategies in the treatment of EC. Materials and methods Clinical trials published before October 2016 and reporting CIK/DC-CIK immunotherapy treatment responses or safety for EC were searched in Cochrane Library, EMBASE, PubMed, Wanfang and China National Knowledge Internet databases. Research quality and heterogeneity were evaluated before analysis, and pooled analyses were performed using random- or fixed-effect models. Results This research covered 11 trials including 994 EC patients. Results of this meta-analysis indicated that compared with conventional therapy, the combination of conventional therapy with CIK/DC-CIK immunotherapy significantly prolonged the 1-year overall survival (OS) rate, overall response rate (ORR) and disease control rate (DCR) (1-year OS: P=0.0005; ORR and DCR: P<0.00001). Patients with combination therapy also showed significantly improved quality of life (QoL) (P=0.02). After CIK/DC-CIK immunotherapy, lymphocyte percentages of CD3+ and CD3−CD56+ subsets (P<0.01) and cytokines levels of IFN-γ, -2, TNF-α and IL-12 (P<0.00001) were significantly increased, and the percentage of cluster of differentiation (CD)4+CD25+CD127− subset was significantly decreased, whereas analysis of CD4+, CD8+, CD4+/CD8+ and CD3+CD56+ did not show significant difference (P>0.05). Conclusion The combination of CIK/DC-CIK immunotherapy and conventional therapy is safe and markedly prolongs survival time, enhances immune function and improves the treatment efficacy for EC.