Objective: The purpose of this study is to explore the effects of microRNA-29b (miR-29b) regulating MAPK/ERK and PI3K/Akt signaling pathways on angiogenesis in endometrial carcinoma (EC) by targeting VEGFA. Methods: Between February 2013 and April 2015, 126 EC patients admitted to the Second Affiliated Hospital of Nanchang University were randomly selected, with 126 EC tissues and the corresponding adjacent normal tissues collected after surgery. The human EC cell lines RL-95-2 and HEC-1-B and human endometrial cells were assigned to the normal group (human endometrial cells), the blank group (untransfected RL-95-2 or HEC-1-B cells), the pMIR-control group (RL-95-2 or HEC-1-B cells transfected with an empty vector), the pMIR-miR-29b group (RL-95-2 or HEC-1-B cells transfected with the miR-29b plasmid), LNA-control group (RL-95-2 or HEC-1-B cells transfected with an oligonucleotide inhibitors control), the LNA-miR-29b inhibitors group (RL-95-2 or HEC-1-B cells transfected with miRCURY LNATM miR-29b inhibitors), the LNA-miR-29b inhibitors + PD98059 group (RL-95-2 or HEC-1-B cells transfected with miRCURY LNATM miR-29b inhibitors and PD98059, an inhibitor of the MAPK/ERK signaling pathway) and the LNA-miR-29b inhibitors + wortmannin group (RL-95-2 or HEC-1-B cells transfected with miRCURY LNATM miR-29b inhibitors and wortmannin, an inhibitor of the PI3K/Akt signaling pathway). qRT-PCR and Western blotting were conducted to detect the miR-29b expression and the mRNA and protein expressions of VEGFA, ERK, Akt, mTOR and Bcl-2. Immunohistochemistry (IHC) was performed to determine the microvessel density (MVD) expression in the EC tissues, adjacent normal tissues and nude-mice. Results: Compared with the adjacent normal tissues, miR-29b expression was down-regulated, the mRNA and protein expressions of VEGFA, ERK, Akt, mTOR and Bcl-2 were up-regulated, and MVD expression was increased in the EC tissues. Compared with the normal group, miR-29b expression was down-regulated, while the mRNA and protein expressions of VEGFA, ERK, Akt, mTOR and Bcl-2 were up-regulated in the other groups. Compared with the blank, pMIR-control and LNA-control groups, miR-29b expression was increased, while mRNA and protein expressions of VEGFA, ERK, Akt, mTOR and Bcl-2 were decreased in the pMIR-miR-29b group. The LNA-miR-29b inhibitors group exhibited elevated miR-29b expression and decreased mRNA and protein expressions of VEGFA, ERK, Akt, mTOR and Bcl-2 (All P < 0.05). Additionally, miR-29b expression was reduced in the LNA-miR-29b inhibitors + PD98059 and LNA-miR-29b inhibitors + wortmannin groups. In comparison to the normal group, MVD expression was elevated in the other groups. Compared with the blank, pMIR-control, LNA-control, LNA-miR-29b inhibitors + PD98059 and LNA-miR-29b inhibitors + wortmannin groups, MVD expression was decreased in the pMIR-miR-29b group but increased in the LNA-miR-29b inhibitors group. Conclusion: Our results indicate that miR-29b negatively mo...
An acquired resistance to platinum-based drugs has emerged as a significant impediment to effective ovarian cancer therapy. The present study explored the anticancer mechanisms of triptolide (TPL) in SKOV3PT platinum-resistant human ovarian cancer cells and observed that TPL activated caspase 3 and induced the dose-dependent apoptosis of the SKOV3PT cells. Furthermore, TPL inhibited complex I of the mitochondrial respiratory chain (MRC) followed by an increase of reactive oxygen species (ROS), which further inhibited nuclear factor (NF)-κB activation and resulted in the downregulation of anti-apoptotic proteins, Bcl-2 and X-linked inhibitor of apoptosis protein (XIAP). Notably, the pre-treatment with N-acetyl-L-cysteine (NAC) abolished the TPL-induced ROS generation, NF-κB inhibition and cell apoptosis, but did not affect the inhibitory effect of TPL on complex I activity. These results suggested that TPL negatively regulated the NF-κB pathway through mitochondria-derived ROS accumulation, promoting the apoptosis of the SKOV3PT cells. Furthermore, TPL synergistically enhanced the cytotoxicity of cisplatin against platinum-resistant ovarian cancer cells. Collectively, these findings suggest that TPL is able to overcome chemoresistance and that it may be an effective treatment for platinum-resistant ovarian cancer, either alone or as an adjuvant therapy.
The present study aimed to investigate the effects of menstrual blood-derived stem cells (MenSCs) on endometrial injury repair. MenSCs were isolated from human menstrual blood and were cultured in vitro. Flow cytometric analysis of cells in the third generation demonstrated that MenSCs exhibited higher expression levels of cluster of differentiation (CD)90 and lower expression levels of CD146, which suggested that the MenSCs were cultured successfully. A mechanical damage model of unilateral (right) endometrium was established in BALB/c nude mice, which were divided into four groups, Normal, negative control (NC), Model and MenSC. MenSCs transfected with adenovirus-enhanced green fluorescent protein were transplanted into the right uterine cavity of mice in the MenSC and NC groups. The protein expression levels of keratin, vimentin, and vascular endothelial growth factor (VEGF) and the average endometrial thickness were measured by immunohistochemistry; the average optical density of vimentin, VEGF and keratin in the MenSC-treated group was significantly higher compared with the untreated Model group. Fertility tests were performed to determine the pregnancy rate of each group; following endometrial damage in BALB/c nude mice, endometrial thickness was decreased in the Model group, whereas model mice treated with MenSC exhibited increased endometrial thickness and increased the pregnancy rates. Therefore, MenSCs may promote the repair of endometrial lesions in mice by promoting the expression of vimentin, VEGF and keratin.
Notch3 and pS6 play important roles in tumor angiogenesis. To assess the expression of Notch3 and pS6 in Chinese ovarian epithelial cancer patients, a ten-year follow-up study was performed in ovarian epithelial cancer tissues from 120 specimens of human ovarian epithelial cancer, 30 specimens from benign ovarian tumors, and 30 samples from healthy ovaries by immunohistochemistry. The results indicate that the expression of Notch3 and pS6 was higher in ovarian epithelial cancer than in normal ovary tissues and in benign ovarian tumor tissues (p < 0.01). In tumor tissues, Notch3 expression and pS6 expression were negatively associated with age (p > 0.05) but positively associated with clinical stage, pathological grading, histologic type, lymph node metastasis, and ascites (p < 0.05 or p < 0.01). A follow-up survey of 64 patients with ovarian epithelial cancer showed that patients with high Notch3 and pS6 expression had a shorter survival time (p < 0.01), in which the clinical stage (p < 0.05) and Notch3 expression (p < 0.01) played important roles. In conclusion, Notch3 and pS6 are significantly related to ovarian epithelial cancer development and prognosis, and their combination represents a potential biomarker and therapeutic target in ovarian tumor angiogenesis.
Introduction: Advanced ovarian cancer is the main cause of ovarian cancer deaths, and it is important to seek safe and effective phytochemicals to suppress cancer or lower the chemotherapy resistance of ovarian cancer.Methods: This study evaluated the effect of Triptolide (TPL) on the proliferation, cycle distribution, apoptosis, and ultra-structure of COC1/DDP cells in vitro, as well as the anti-cancer effect and sensibilisation effect of TPL in vivo.Results: The results indicated that TPL could significantly inhibit the growth of COC1/DDP cells (P<0.05), and 3 ng/ml TPL and 50 ng/ml TPL made COC1/DDP cells present obvious apoptosis characteristics and arrest 35% and 55% of COC/DDP cells in the G0/G1 phase, respectively (P<0.05). The animal experiments also indicated that 0.1mg/kg.d TPL significantly reduced the tumour weight and the spleen cell transformation rate (SI), and it lowered the inflammatory factors IL-2 and TNF-a in rat serum (P<0.05). Moreover, the significant reduction of p-Akt and p-GSK3β made the TPL+DDP possess the highest apoptosis rate [(51.13±3.325)%] in COC1/DDP cells.Conclusions: TPL used in combination with DDP may produce a synergistic anti-cancer effect that warrants further investigation for its potential clinical applications in the treatment of epithelial ovarian cancer.
Chemotherapy resistance of advanced ovarian cancers is responsible for death of most cancer patients, so it is necessary to seek safe and effective natural ingredients to lower the chemotherapy resistance of ovarian cancer. In the present study, we studied the anticancer effects of triptolide (TPL) and TPL + cisplatin (DDP) in vitro and in vivo using SKOV3/DDP cell line and a mouse model. In vitro results showed that TPL and TPL + DDP inhibited cellular invasion and migration of SKOV3/DDP cells (P<0.05), and significantly reduced the expression of adhesion-related proteins integrin β1 (ITGβ1) and apoptosis-inhibiting proteins survivin, matrix metalloproteinase 2 (MMP-2) and MMP-9 (P<0.05). Animal results demonstrated that TPL and TPL + DDP had significantly enhanced the inflammatory factor-2 (IL-2) and tumor necrosis factor-α (TNF-α) in serum of mice, and significantly increased the NK cell-related protein levels of CD16 and CD56, while significantly inhibited the production of vascular endothelial growth factor (VEGF) related protein clusters of differentiation 31 (CD31) and CD105. Collectively, the combination of TPL and DDP may produce a synergistic anticancer effect on epithelial ovarian cancer (EOC).
Endometriosis (EMS) is a disease characterized by estrogen-dependent, chronic inflammatory, and annoying symptoms, which inflicts about 10% reproductive-age women. The diagnosis of endometriosis mainly depends on pathological examination after surgical resection while the pathogenesis of EMS is not clear enough. Surgical resection and drug therapy (including painkillers and hormone therapy, especially gonadotropin-releasing hormone analogs, GnRH-a) are widely used, but they are expensive and have many side effects. There are few studies on vaginal microorganisms in women with endometriosis. We collected vaginal secretions from women with EMS confirmed by pathology and demonstrated that they were different from that of healthy women by 16s rRNA high-throughput sequencing. Additionally, we established the EMS model in female mice by intraperitoneally injecting fragments from donor mice (3-week growth). Then, the mice were treated with mixed antibiotics (vagina) and NF-κB signaling pathway inhibitors (intraperitoneal injection), respectively. The result suggested that the ectopic lesions were inhibited. In addition, inflammatory cytokines IL-1β, IL-6, and TNF-α in peritoneal fluid, cell proliferation marker ki-67, and macrophage marker Iba-1 in ectopic lesions decreased significantly from that of mock mice. We also observed similar results as above by vaginal microbiota transplantation (VMT) and subcutaneous injection of leuprorelin acetate (LA, one of GnRH-a) for mice with EMS. These results showed that vaginal use of antibiotics or VMT is helpful to treat endometriosis in mice. However, due to the great difference between human and mouse vaginal microbiota, its mechanism and clinical transformation application still need to be further studied in the future.
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