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Objective: To investigate the effects of the repair and restitution of ear-shaped cartilage by adipose tissue-derived stem cells(ADSC) and cartilage acellular extracellular matrix. Methods: ADSC were extracted by digesting with collagenase type II from the adipose tissue from 32 patients with adiposity whose fats were drawn, and were cultured and subcultured in vitro. The natural biological scaffolds were prepared by acellular method using porcine ear cartilage, and then the second generation ADSC(5.0×10(7)/ml) were inoculated on the preformed natural bio-scaffold scaffold by culturing in vitro for 3 days to form a cell scaffold complex. 32 New Zealand white rabbits were randomly divided into the experimental groups, the control group A, the control group B and the control group C. All New Zealand white rabbits were modeled by ear cartilage defects. The cell scaffolds composite was implanted into the experimental group of the ear cartilage defects of rabbits, the ADSC were implanted into the control group A, the cartilage acellular extracellular matrix scaffold was implanted into the control group B and the control group C was modeled only by ear cartilage defects. After 16 weeks, the animals were sacrificed and the repair effect was observed by gross appearance and histological examinations including HE, Toluidine blue staining, Safranin O and typeⅡ collagen staining. Its were quantitatively analyzed by positive staining results of type Ⅱ collagen. Ear cartilage tissue elasticity was detected. SPSS 17.0 software was used to analyze the data. Results: The cartilage defects in the experimental group were repaired well by general shape observation and those in the control group was filled in with granulation tissue. There were significant differences between the experimental group and the control group in the wet weight( P< 0.05). HE staining showed that cartilage cavities formed in articular cartilage defects, and only the fibrous tissue was filled with the ear cartilage defect in the control groups. In the repair area, Toluidine blue staining, Safranin O and type Ⅱ collagen staining were positive in the experimental group, and negative in the control groups. There was no significant difference between the experimental group and the normal ear cartilage in the ear cartilage elastic constant detection( P> 0.05). Conclusions: The mechanics and histology of rabbit ear neonatal cartilage constructed by ADSC combined with cartilage acellular matrix are close to normal ear cartilage. Cartilage acellular matrix material combined with adipose-derived stem cells has good repair and reconstruction ability for ear cartilage defects, which possesses potential clinical application value.

ADAM17 is believed to promote tumor development by facilitating both cell proliferation and migration. In this study, we investigated the involvement of ADAM17 and the activation of the integrin pathway in the regulation of the malignant properties of hepatocellular carcinoma cells and tissues. ADAM17 was positively correlated with active integrin β1, which was determined using a human tissue microarray and an N-nitrosodiethylamine-induced HCC mouse model. We found elevated ADAM17 and active integrin β1 levels in HCC tissues compared with adjacent liver tissues, and the active integrin β1 levels were associated with tumor size and TNM grade. High ADAM17 and active integrin β1 levels in tumor tissues were significantly associated with poor survival of HCC patients. RNAi-mediated ADAM17 knockdown and integrin β1 blockade significantly attenuated the migration and invasion of HCC cells, and overexpression of ADAM17 showed the reverse effects. ADAM17 interference attenuated the intrahepatic growth and metastasis of HCC cells in an orthotopic xenograft model. ADAM17-knockdown cells showed diminished levels of active integrin β1, p-FAK, p-AKT, MMP-2 and MMP-9. ADAM17 knockdown significantly attenuated the translocation of the Notch1 intracellular domain into the nucleus, whereas overexpression of the Notch1 intracellular domain rescued the translocation and enhanced the activation of integrin β1. Our data provide evidence for ADAM17 as an important determinant of malignant properties via regulation of integrin β1 activation and Notch1 signaling. Inhibition of ADAM17 may provide viable therapeutic potential for preventing HCC metastasis., (Copyright © 2018 Elsevier Inc. All rights reserved.)

To understand the role of glycosaminoglycans in bacterial cellular invasion, xylosyltransferase-deficient mutants of Chinese hamster ovary (CHO) cells were created using clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR-associated gene 9 (CRISPR-cas9) gene targeting. When these mutants were compared to the pgsA745 cell line, a CHO xylosyltransferase mutant generated previously using chemical mutagenesis, an unexpected result was obtained. Bacterial invasion of pgsA745 cells by group B Streptococcus (GBS), group A Streptococcus, and Staphylococcus aureus was markedly reduced compared to the invasion of wild-type cells, but newly generated CRISPR-cas9 mutants were only resistant to GBS. Invasion of pgsA745 cells was not restored by transfection with xylosyltransferase, suggesting that an additional mutation conferring panresistance to multiple bacteria was present in pgsA745 cells. Whole-genome sequencing and transcriptome sequencing (RNA-Seq) uncovered a deletion in the gene encoding the laminin subunit α2 (Lama2) that eliminated much of domain L4a. Silencing of the long Lama2 isoform in wild-type cells strongly reduced bacterial invasion, whereas transfection with human LAMA2 cDNA significantly enhanced invasion in pgsA745 cells. The addition of exogenous laminin-α2β1γ1/laminin-α2β2γ1 strongly increased bacterial invasion in CHO cells, as well as in human alveolar basal epithelial and human brain microvascular endothelial cells. Thus, the L4a domain in laminin α2 is important for cellular invasion by a number of bacterial pathogens., Importance: Pathogenic bacteria penetrate host cellular barriers by attachment to extracellular matrix molecules, such as proteoglycans, laminins, and collagens, leading to invasion of epithelial and endothelial cells. Here, we show that cellular invasion by the human pathogens group B Streptococcus, group A Streptococcus, and Staphylococcus aureus depends on a specific domain of the laminin α2 subunit. This finding may provide new leads for the molecular pathogenesis of these bacteria and the development of novel antimicrobial drugs., (Copyright © 2017 van Wijk et al.)

Objective: To study the immuno-tolerance mechanism of the third-party bone marrow-derived mesenchymal stem cells (BMSCs) in the allogeneic transplantation., Methods: Forty female C57BL/6 mice and forty male BALB/C mice were respectively used as donors and recipients in skin allogenic graft model. Forty male BALB/C mice were divided randomly into 4 groups: blank control group, CP group, BMSCs group , CP + BMSCs group, with 10 mice in each group. Before skin graft, high-dose abdominal injection of cyclophosphamide (200 mg/kg, 2 d, q. d.) was performed in recipient mice in CP and CP + BMSCs groups. On the transplantation day, a bonus of 2 x 10(6) BMSCs from the SD rat (SD-BMSCs) were injected through the tail vein in the BMSCs and CP + BMSCs groups. The observation and HE staining of skin grafts were used. The expressions of CD29, CD34, CD45 and CD90 of cells were analyzed by using flow cytometry in order to identify BMSCs. The CD4+, CD25+, Foxp3 and Treg cells of spleen were detected by flow cytometry. Cytokine in peripheral blood of recipient mice were measured by ELISA, including TGF-beta, IL-10 and IFN-gamma. T cells were co-cultured with 60Co-irradiated bone marrow MSCs from different individuals. The proliferative activity of T cells were evaluated with MTT assay., Results: The skin graft survival time was significantly prolonged in the CP + BMSCs group, as compared with that in the blank control group, the CP group, the BMSCs group, respectively. Cells cultured by whole bone marrow adherent cultivation showed CD29 (99.7%), CD44+ (96.7%), CD34- (1.6%), CD45- (1.3%). Compared with the control group and CP group, the ratio of the CD4+, CD25+, Foxp3+ and Treg cells significantly increased in the SD-BMSCs group and CP + BMSCs group (P < 0.05). Analysis of peripheral blood by ELISA showed significant high level of TGF-beta, IL-10 and low level of IFN-gamma in BMSCs group and CP group,compared with that in control group. When co-cultured with BMSCs from different individuals, T- lymphocytes proliferation decreased apparently in SD-BMSCs group and C57-BMSCs group (P < 0.05), but there was no significant difference between SD-BMSCs group and C57-BMSCs group (P > 0.05)., Conclusions: The immunotolerance mechanism of the third-party bone marrow-derived mesenchymal stem cells in the allogeneic transplantation might be associated with its effect on the proliferation of Treg cells and increasing expression of TGF-beta and IL-10, decreasing expression of IFN-gamma.

Objective: To investigate the effect of the third-party bone marrow-derived mesenchymal stem cells (BMSCs) on the allogeneic skin transplantation., Methods: 40 female C57BL/6 mice and 50 male BALB/C mice were respectively used as donors and recipients of skin transplantation. 50 BALB/C mice were divided randomly into 5 groups: Blank control group, Cyclophosphamide group BMSCs group, Cyclophosphamide + BMSCs group and CM-DiI staining group, with 10 mice in each group. Before skin transplantation, high-dose abdominal injection of Cyclophosphamide (200 mg/kg, 2 d) was performed in recipient mice. On the transplantation day, a bonus of 1 x 10(5) BMSCs of the SD rat (SD-BMSCs) were injected through the tail vein. The observation of skin grafts, mixed lymphocyte culture (MLC), HE staining, the observation of CM-DiI-labeled SD-BMSCs and FACS were used., Results: The skin graft survival time was significantly prolonged in the Cyclophosphamide + BMSCs group, as compared with the blank control group, the Cyclophosphamide group, the BMSCs group respectively. When BMSC and lymphocyte mixed at the ratio of 1:1 and 1:10, rat BMSCs inhibited T lymphocyte proliferation. More angiogenesis and less lymphocyte infiltration were found in the experimental group than them in other groups. Red fluorescent cells were found in CM-DiI staining group under long-term observation. The SD-BMSCs can he detected by flow cytometry in the cell group and the Cyclophosphamide + BMSCs group., Conclusions: BMSCs can survive in the heterogeneous recipient body; the third-party BMSCs transplantation can prolong skin graft survival time; BMSCs can inhibit T lymphocyte activation and proliferation.