Cervical cancer-derived exosomal miR-663b promotes angiogenesis by inhibiting vinculin expression in vascular endothelial cells.

BACKGROUND: Angiogenesis provides essential nutrients and oxygen for tumor growth and has become the main mechanism of tumor invasion and metastasis. Exosomes are nanoscale membrane vesicles containing proteins, lipids, mRNA and microRNA (miRNA), which mediate intercellular communication and play an important role in tumor progression. Accumulated evidence indicates that tumor-derived exosomal miRNAs participate in the tumor microenvironment and promote angiogenesis. METHODS: Bioinformatic target prediction and dual luciferase reporter assays were performed to identify the binding site between miR-663b and the 3'-UTR of vinculin (VCL). VCL overexpression lentivirus and miR-663b overexpression/inhibition lentivirus were used to create a VCL overexpression model and miR-663b overexpression/inhibition model in-vitro. Immunohistochemistry (IHC) assays and western blot assays were used to detect protein expression. Exosome-cell cocultures, wound healing assays, tube formation assays and transwell assays were used to measure the migration and tube formation ability of vascular endothelial cells [human umbilical vein endothelial cells (HUVECs)]. siRNA targeted VCL was used to knockdown VCL. RESULTS: In the present study, we found that miR-663b was elevated in cervical cancer tissue and exosomes. miR-663b could bind the 3'-UTR of VCL and inhibit its expression. VCL is downregulated in cervical cancer, and decreased VCL has a negative correlation with a high level of miR-663b. Further studies demonstrated that exosomes secreted by cervical cancer cells can deliver miR-663b to HUVECs and inhibit the expression of VCL, thereby promoting angiogenesis and tumor growth. CONCLUSIONS: miR-663b derived from cancer cell exosomes acts as a driving factor for angiogenesis and a potential target of antiangiogenic therapy in cervical cancer. Our findings illustrated a new signaling pathway, including exosomes, miRNAs and target genes, which provides potential targets for antiangiogenic therapy.

Uptake of Aß by OATPs might be a new pathophysiological mechanism of Alzheimer disease.

BACKGROUND: The accumulation of neurotoxic amyloid-beta (Aß) in the brain is a characteristic of Alzheimer's disease (AD), at the same time, it is possible alterations of liver function could affect brain Aß levels through changes in blood Aß concentration. Over the last decade, a number of reports have shown that P-glycoprotein (encoded by ABC1B1) actively mediates the efflux transport of Aß peptides. However, the mechanism by which Aß peptides enter the cells is not clear. In the preliminary study, we found that the protein expression of organic anion transporting Polypeptide 1a4 (OATP1B1) in the liver tissue of mice with AD was significantly higher than that in the normal mice. In contrast, the protein expression of Oatp1a4 in the brain significantly decreased in mice with AD. OATP1B1, an important drug transporter might be related to the pathophysiology of AD. RESULTS: In this study, we established an OATP1B1-GFP-HEK293T cell model to confirm the OATP1B1 mediated transport of Aß1-42. Compared to the control group of GFP-HEK293Tcells, the uptake of Aß1-42 protein in the OATP1B1-GFP-HEK293T group increased significantly with the increase in concentration of Aß1-42, and also increased significantly with an increase in the duration of incubation. Similar results were observed in the flow cytometry experiment, and the uptake of Aß1-42in HEK293T-OATP1B1 cells was almost twice that in the control group. These results indicate that OATPs may act as an important "carrier" for the transport of Aß1-42 from the blood to the tissues, including liver and brain. CONCLUSIONS: This is a novel and interesting finding and OATP1B1 can be investigated as a new treatment target for AD.

Management of high-grade squamous intraepithelial lesion patients with positive margin after LEEP conization: A retrospective study.

ABSTRACT: To explore the optimal way to manage patients with high-grade squamous intraepithelial lesion (HSIL) and positive margin by identifying the risk factors for its recurrence and residue.A retrospective study was conducted on 267 cases of a pathologically confirmed HSIL with positive margin following conization by loop electrosurgical excisional procedure (LEEP) between January 2010 and December 2015. One hundred two cases were selected for regular follow-up every 6 months, and 165 cases were selected for a second surgery (repeat cervical conization or hysterectomy) within 3 months of initial LEEP. We analyzed the association between recurrent or residual diseases and these factors: age, menopausal status, ThinPrep cytologic test (TCT) results, high-risk human papillomavirus (HR-HPV) infection, pathological grades of the margin, number of involved margins, and glandular involvement.The recurrence rate among 102 cases who underwent follow-up was 17.6% (18/102). The factors: atypical squamous cells of undetermined significance cannot exclude HSIL (ASC-H) or higher lesions in the pre-LEEP TCT (P = .038), persistent HR-HPV infection at the 6th month post-LEEP (P = .03), HSIL-positive margin (P = .003), and multifocal-involved margin (P = .002) were significantly associated with recurrent disease, while age, menopause, and pre-LEEP HR-HPV infection were not associated with recurrent disease (P > .05). The residual rate among 165 patients who underwent a second surgery was 45.5% (75/165), of which 15 cases were residual cervical cancer. The factors: menopause (P = .02), ≥ASC-H in pre-LEEP TCT (P = .04), pre-LEEP HR-HPV infection (P = .04), ≥HSIL-positive margin (P < .001), and multifocal-involved margin (P < .001) significantly increased the risk of residual disease. No correlation existed between residual disease and age or glandular involvement (P > .05).For patients with a positive margin after LEEP, regular follow-up or second surgery should be selected according to fertility requirement and pathological characteristics of the positive margin, as well as TCT and HR-HPV infection condition.

Exosomal miR­663b exposed to TGF­ß1 promotes cervical cancer metastasis and epithelial­mesenchymal transition by targeting MGAT3.

Transforming growth factor (TGF)­ß1 is a key cytokine affecting the pathogenesis and progression of cervical cancer. Tumor­derived exosomes contain microRNAs (miRNAs/miRs) that interact with cancer and stromal cells, thereby contributing to tissue remodeling in the tumor microenvironment (TME). The present study was designed to clarify how TGF­ß1 affects tumor biological functions through exosomes released by cervical cancer cells. Deep RNA sequencing found that TGF­ß1 stimulated cervical cancer cells to secrete more miR­663b­containing exosomes, which could be transferred into new target cells to promote metastasis. Further studies have shown that miR­663b directly targets the 3'-untranslated regions (3'­UTR) of mannoside acetylglucosaminyltransferase 3 (MGAT3) and is involved in the epithelial­mesenchymal transition (EMT) process. Remarkably, the overexpression of MGAT3 suppressed cervical cancer cell metastasis promoted by exosomal miR­663b, causing increased expression of epithelial differentiation marker E­cadherin and decreased expression of mesenchymal markers N­cadherin and ß­catenin. Throughout our study, online bioinformation tools and dual luciferase reporter assay were applied to identify MGAT3 as a novel direct target of miR­663b. Exosome PKH67­labeling experiment verified that exosomal miR­663b could be endocytosed by cervical cancer cells and subsequently influence its migration and invasion functions which were measured by wound healing and Transwell assays. The expression of miR­663b and MGAT3 and the regulation of the EMT pathway caused by MGAT3 were detected by quantitative real­time transcription­polymerase chain reaction (qPCR) and western blot analysis. These results, thus, provide evidence that cancer cell­derived exosomal miR­663b is endocytosed by cervical cancer cells adjacent or distant after TGF­ß1 exposure and inhibits the expression of MGAT3, thereby accelerating the EMT process and ultimately promoting local and distant metastasis.

Potential risk and control strategy of biofilm pretreatment process treating raw water.

An enhanced lab-scale biofilm pretreatment process treating raw water obtained from eutrophicated water bodies was established and started up with a novel strategy of low-level nutrients addition and effluent recirculation. Results showed that the startup strategy was useful for biofilm formation and pollutants removal, but it had the risks of increasing substrate affinity constant (Ks) and biofilm decay in treating raw water. Fortunately, the increased Ks value did not affected the NH4(+)-N removal performance via keeping the NH4(+)-N loading rate larger than 6.29 mg L(-1)d(-1). In addition, lower hydraulic retention time (HRT) favored the removal of organic matters, and the maximum TOC removal rate of 76.5 mg L(-1)d(-1) were achieved at HRT of 2h. After long-term acclimatization at oligotrophic niche, the decrease of Ks value and increase of biomass, extracellular polymeric substances, bioactivity were achieved. Finally, the stable operation of biofilm pretreatment process was realized in treating polluted raw water.