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To observe the effect of borax on Hedgehog signaling pathway protein expression of human hepatocellular carcinoma HepG2 cells. Methods HepG2 cells were randomly divided into two groups. The experimental group was treated with 4. 0 mmol/L borax for 24 h, and the control group was not treated. The cells were collected and the Hedgehog signaling pathway proteins Gli2, Ptchl and Cyclin Dl were detected by real-time quantitative PCR ( qRT-PCR). The Hedgehog signaling pathway proteins Gli2, Ptchl and Cyclin Dl were detected by Western blotting. Results The relative mRN A expression levels of Gli2, Ptchl and Cyclin Dl were 0. 58 ± 0. 05, 0. 64 ± 0. 08, and 0. 35 ± 0. 03, respectively, and the corresponding protein relative expression levels were 1. 14 ± 0. 03, 0. 93 ± 0. 02, and 1. 86 ± 0. 02, respectively, which were lower than those of the control group, with statistically significant differences ( all P < 0. 05). Conclusion Borax may down-regulate the expression of Hedgehog signaling pathway protein in human hepatocellular carcinoma HepG2 cells, which provides a basis for the development and target of new anti-tumor drugs. [ABSTRACT FROM AUTHOR]

We developed a high-efficiency microfluidic chip for extracting exosomes from human plasma. We collected peripheral blood from normal human, designed and fabricated a microfluidic chip based on nanoporous membrane and agarose gel electrophoresis to isolate exosomes. The extracted exosomes were characterized by transmission electron microscopy, nanosight and Western blotting, the morphology, concentration and particle size of exosomes were identified and analyzed. Meanwhile, we used ultracentrifugation and microfluidic chip to isolate exosomes separately. The particle size and concentration of the exosomes extracted by two methods were compared and analyzed, and their respective extraction efficiency was discussed. Finally, the expression level of miRNA-21 in exosomes was analyzed by RT-PCR. The microfluidic chip isolated (in 1 hour) high-purity exosomes with size ranging from 30-200 nm directly from human plasma, allowing downstream exosomal miRNA analysis. By comparing with ultracentrifugation, the isolation yield of microfluidic chip was 3.80 times higher than ultracentrifugation when the volume of plasma sample less than 100 μL. The optimized parameters for exosome isolation by gel electrophoresis microfluidic chip were: voltage: 100 V; concentration of agarose gel: 1.0%; flow rate of injection pump: 0.1 mL/h. The gel electrophoresis microfluidic chips could rapidly and efficiently isolate the exosomes, showing great potential in the research of exosomes and cancer biomarkers.

Lung cancer is a malignant tumor with high incidence and mortality in the world, which seriously threatens people's safety and health. Early diagnosis of lung cancer is the key part in the process of prevention and treatment of lung cancer. It can improve the survival of patients with lung cancer. Exosomes are closely related to the invasion and metastasis process of tumor, it plays an important role in the development of lung cancer. Biomarkers based on exosomes have become a powerful diagnostic tool of lung cancer. Exosomes are lipid bilayer vesicles with uniform size and diameter of 30 nm-200 nm secreted by cells. Exosomes contain different types of nucleic acids and proteins. These nucleic acids and proteins are derived from their parent cells (including parent cancer cells), which have a wide range of physiological functions, including immune regulation, intercellular communication and other physiological activities. Biomacromolecules in exosomes, such as single-stranded RNA, long noncoding RNA, microRNA, protein and lipids, which can provide valuable genetic information for early clinical diagnosis of lung cancer. This review described the origin, structural characteristics, extraction methods, biological characteristics of exosomes and the relationships of exosomes in the early diagnosis of lung cancer.
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Objective To develop and verify a medical microdevice for analyzing the three-dimensional(3D)migration of tumor cells in extracellular matrix. Methods The mold of the microdevice was made by precision machining,and then the medical microdevice based on polydimethylsiloxane(PDMS)-glass was obtained by PDMS casting,moulding,and bonding.During the analysis,the suspension of tumor cells and matrigel were mixed and then added into the migration channel of microdevice,and the controllable migration of tumor cells in matrigel was induced by establishing chemokine concentration gradient on both sides of the migration channel.Meanwhile,the migration process of tumor cells was recorded with the live cell dynamic imaging device. Results Breast cancer cell line MCF-7 was taken as an example to verify the feasibility of the microdevice to control and dynamically monitor the 3D migration process of tumor cells in vitro.Qualitative analysis of imaging data showed that the migration of MCF-7 cell lines in matrigel was determined by the concentration gradient distribution direction of chemokine and presented as the amoeboid-like migration mode.The proportion and migration velocity of MCF-7 cells could be quantified by the quantitative analysis of cell migration process.The inhibition ability of matrix metalloproteinase inhibitors(Batimastat)and adenosine triphosphatase inhibitors(Blebbistation)on the 3D migration behavior of MCF-7 cells was found to be different. Conclusion This device can be used for in-depth analysis of tumor cell migration and its mechanism and for evaluating the efficacy of anti-metastatic drugs.

Objective: To investigate the dynamic features of angiogenesis in residual tumors after high intensity focused ultrasound (HIFU),and to determine the temporal effect and mechanism of hypoxia inducible factor-2 alpha (HIF-2a) in the angiogenic process of residual tumors., Methods: Xenograft tumors of HepG2 cells were generated by subcutaneously inoculating athymic BALB/c nu/nu mice with the hepatoma cells.About 30 days after inoculation,all mice (except in the control group) were treated by HIFU and assigned randomly to the following 7 groups according to various time intervals post-treatment:1st,3rd,5th day and 1st,2nd,3rd,4th week when the residual tumor tissues were obtained from the experimental groups.Protein levels of HIF-2a and vascular growth factor A (VEGF-A) were quantified by immunohistochemistry and western blotting,and mRNA levels were measured by (real-time quantitative) qPCR. Microvascular density (MVD) was calculated by counting the CD31-positive vascular endothelial cells identified by means of an immunohistochemical staining method., Results: Compared with results from the control group,the protein and mRNA levels of HIF-2a expression reached the highest level in the experimental mice at the 2nd week (P=0.000 and P < 0.01 respectively),and were decreased thereafter(3rd week and 4th week, P=0.000 and P < 0.05).VEGF-A expression in the residual tumor tissues group that received HIFU was significantly decreased,compared with the control group,at all time points uPto 1 week (all P=0.000 and P < 0.01),but the levels increased compared to controls in the 2nd through 4th week (all P=0.000, P < 0.05). Similar results were obtained for MVD., Conclusion: HIFU treatment can inhibit angiogenesis in residual hepatoma tissues in the short-term (1 to 2 weeks post-treatment) in mice with hepatocellular carcinoma,but can promote angiogenesis overtime (2 to 4 weeks post-treatment); the angiogenic process may involve the HIF-2α/VEGFA pathway.

Objective: To study the changes in hypoxia-inducible factor (HIF1α, HIF2α) in the residual tumor cells in nude mice bearing hepatocellular carcinoma (HCC) following treatment with high-intensity focused ultrasound (HIFU)., Methods: Thirty nude mice bearing human HCC received treatment with HIFU. At 1, 3, and 5 days and 1 and 2 weeks after the treatment, the mice were examined for pathological changes of the residual tumor with HE staining; SP immunohistochemistry, Western blotting and real-time quantitative PCR were used to detect the protein and mRNA expressions of HIF1α and HIF2α in the tumor., Results: HE staining revealed the presence of residual tumor cells and large necrotic areas after the treatment. Immunohistochemistry showed a gradual increment of HIF1α protein and mRNA expressions after the treatment, reaching the peak level at 3 days (P<0.05) followed by progressive reduction at 5 days and 1 and 2 weeks. HIF2α expressions at either the protein or mRNA levels exhibited no significant changes within 3 days after the treatment (P>0.05) but increased significantly at 5 days and 1 and 2 weeks (P<0.05)., Conclusion: The changes of HIF1α and HIF2α in the residual tumor after HIFU treatment in nude mice bearing HCC can be associated with tumor cell apoptosis and angiogenesis after the treatment.