Clonorchis sinensis legumain promotes migration and invasion of cholangiocarcinoma cells via regulating tumor-related molecules.

BACKGROUND: Clonorchis sinensis infection causes serious pathological changes in the bile duct and is highly correlated with cholangiocarcinoma. The excretory-secretory products (ESP) of C. sinensis play a critical role in the oncogenesis and progression of cholangiocarcinoma, while the components and precise mechanism remain unclear. Here, we evaluated the function of C. sinensis legumain (Cslegumain) in promoting the invasion and migration of cholangiocarcinoma cells and the mechanism involved. METHODS: The structural and molecular characteristics of Cslegumain were predicted and analyzed using the online program Phyre2. Quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical staining were performed to test the transcriptional level of Cslegumain and its localization in the adult. Native Cslegumain was detected by western blotting assay. The effects of Cslegumain on the proliferation, invasion and migration of cholangiocarcinoma cells were checked using CCK-8 assay, Matrigel transwell assay and scratch wound healing assay. Expression levels of tumor-related molecules regulated by Cslegumain were evaluated by qRT-PCR and western blotting assay. RESULTS: Cslegumain showed high similarity with human legumain in the secondary and tertiary structures and displayed higher transcriptional levels in the adult worm than in the metacercariae. Native Cslegumain was detected in a catalytic form and was localized mainly in the intestine of the C. sinensis adult and epithelial cells of the intrahepatic bile duct. After transfection into RBE cells, Cslegumain showed high ability in promoting the invasion and migration but not the proliferation of cholangiocarcinoma RBE cells. Furthermore, the expression levels of some molecules including E-cadherin and N-cadherin were downregulated, while the levels of α-actinin 4, ß-catenin and inducible nitric oxide synthase (iNOS) were upregulated. CONCLUSIONS: Our findings indicated that Cslegumain showed very similar structures as those of human legumain and could promote the invasion and migration of cholangiocarcinoma cells by regulating some tumor-related molecules.

MicroRNA-26a Promotes Regulatory T cells and Suppresses Autoimmune Diabetes in Mice.

Type-1 diabetes (TID) is an autoimmune disease in which the body's own immune cells attack islet ß cells, the cells in the pancreas that produce and release the hormone insulin. Mir-26a has been reported to play functions in cellular differentiation, cell growth, cell apoptosis, and metastasis. However, the role of microRNA-26a (Mir-26a) in autoimmune TID has never been investigated. In our current study, we found that pre-Mir-26a (LV-26a)-treated mice had significantly longer normoglycemic time and lower frequency of autoreactive IFN-γ-producing CD4(+) cells compared with an empty lentiviral vector (LV-Con)-treated non-obese diabetic (NOD) mice. Mir-26a suppresses autoreactive T cells and expands Tregs in vivo and in vitro. Furthermore, in our adoptive transfer study, the groups receiving whole splenocytes and CD25-depleted splenocytes from LV-Con-treated diabetic NOD mice develop diabetes at 3 to 4 weeks of age. In comparison, mice injected with undepleted splenocytes obtained from LV-26a-treated reversal NOD mice develop diabetes after 6-8 weeks. And depletion of CD25(+) cells in the splenocytes of reversed mice abrogates the delay in diabetes onset. In conclusion, Mir-26a suppresses autoimmune diabetes in NOD mice in part through promoted regulatory T cells (Tregs) expression.

The HIF-1α/CXCR4 pathway supports hypoxia-induced metastasis of human osteosarcoma cells.

HIF-1α mediates hypoxia-induced expression of the chemokine receptor CXCR4 and contributes to metastasis in many different cancers. We have previously shown that hypoxia promotes migration of human osteosarcoma cells by activating the HIF-1α/CXCR4 pathway. Here, immunohistochemical analysis showed that unlike control osteochondroma samples, osteosarcoma specimens were characterized by elevated expression levels of HIF-1α and CXCR4. Moreover, we found that hypoxia-induced invasiveness was more pronounced in high metastatic potential F5M2 osteosarcoma cells than in low metastatic potential F4 cells, and that this induction was sensitive to treatment with the CXCR4 antagonist AMD3100 and the HIF-1α inhibitor KC7F2. Interestingly, hypoxia-induced CXCR4 expression persisted after cultured osteosarcoma cells were returned to normoxic conditions. These observations were confirmed by experiments in a mouse model of osteosarcoma lung metastasis showing that hypoxia stimulation of pulmonary metastasis was greater in F5M2 than in F4 cells, and was sensitive to treatment with AMD3100. Our study provides further evidence of the contributions of hypoxia and the HIF-1α/CXCR4 pathway to the progression of osteosarcoma, and suggests that this axis might be efficiently leveraged in the development of novel osteosarcoma therapeutics.

In vivo molecular imaging of gastric cancer in human-murine xenograft models with confocal laser endomicroscopy using a tumor vascular homing peptide.

The early detection of premalignant lesions and cancers are very important for improving the survival of patients with gastric malignancies. Confocal laser endomicroscopy (CLE) is a novel imaging tool for achieving real-time microscopy during the ongoing endoscopy at subcellular resolution. In the present study, to evaluate the feasibility of real-time molecular imaging of GEBP11 by CLE in gastric cancer, CLE was performed on two types of tumor-bearing mice models, as well as surgical specimens of patients with gastric cancer, after the application of GEBP11. A whole-body fluorescent imaging device was first used to screen for the strongest specific fluorescent signal in xenograft models. Next, the tumor sites, as well as human tissues, were scanned with CLE. After this, targeted specimens were obtained for fluorescence microscopy and histology. We confirmed that GEBP11 could specifically bind to co-HUVECs by means of CLE in cell experiments. Thereafter, a specific signal was observed in both subcutaneous and orthotopic xenograft models in vivo after the injection of FITC-GEBP11 via tail vein, whereas the group injected with FITC-URP showed no fluorescent signals. In human tissues, a specific signal of GEBP11 was observed in 26/28 neoplastic specimens and in 8/28 samples of non-neoplastic specimens from the patients (p < 0.01). The findings from ex vivo immunofluorescence microscopy of cryostat sections correlated well with that obtained by CLE. These findings indicate that the peptide, GEBP11, might be a potential candidate for the molecular imaging of gastric cancer.

Evaluation of the right ventricular function in pneumoconiosis patients using volume-time curves obtained by real-time three-dimensional echocardiography.

The study was aimed to evaluate the right ventricular function in pneumoconiosis patients by real-time three-dimensional echocardiography. A total of 80 individuals including 44 consecutive pneumoconiosis patients and 36 age- and gender-matched healthy volunteers as controls were prospectively recruited for the study. All the patients underwent two- and three-dimensional echocardiography. Measurements of the right ventricle included tricuspid regurgitation pressure (TRPG), anterior and posterior wall thickness and range of motion (TH1, TH2, M1, M2), right end-diastolic volume and end-systolic volume. The right ventricular ejection fraction (RVEF) was also calculated. The RVEF of healthy volunteers ranged from 50 to 78 %, whereas that of the pneumoconiosis patients varied from 29 to 73 %. An increase in TRPG caused a significant (p = 0.006) decrease in RVEF (by 77.3 %), suggesting the two variables were negatively correlated (r = -0.643, p < 0.01). In comparison with normal, the volume-time curves of the pneumoconiosis patients showed a lower trough. Use of real-time three-dimensional echocardiography provides with added clinical information needed to evaluate right ventricular function in pneumoconiosis patients.