MECCANISMO TRASFORMANTE DEI VIRUS ONCOGENI HPV IN CELLULE EPITELIALI UMANE DELLA CERVICE UTERINA

The aims of this study are: (i) to set up a culture protocol to derive pathological and normal keratinocytes from small human cervical intraneoplasia (CIN) biopsies and normal uterine cervix (NUC) tissues; (ii) to investigate the expression prfile induced by transforming mechanisms due to human papillomavirus (HPV) in CIN keratinocytes clones from different CIN grades, CIN1-3/CIS, using the microarray technique. In this study a new approach to establishing CIN and NUC keratinocyte cultures from small tissue fragments was developed. CIN specimens and corresponding normal tissues, which were used as controls, were digested with collagenase. Tissue-derived fibroblasts and keratinocytes were co-cultured in calcium serum medium conditions. Primary co-cultures were subsequently sub-cultured. Single keratinocyte clones from primary co-cultures were expanded using a culture medium which was optimized in our laboratory. Primary clones from CIN and normal tissues, as well as expanded clones, were tested by immunofluorescence for epithelial and cervical markers such as 5-, 14-, 17- and 19-keratins, and p63. Our results indicate that primary CIN, as well as normal keratinocyte clones, can be obtained in a co-culture system with live human fibroblasts in calcium and serum medium conditions. The clone number varied depending on the grade of CIN lesions from which clones originated. CIN and normal keratinocytes from single clones, when cultured with our new medium, grew at a high rate with uniform morphology. The second objective of our study was to identify genes that are related to progression and transformation inducted by HPV in CIN lesions using the microarray technique. One clone of CIN1; two clones of CIN2 and corresponding normal clones; one clone of CIN3 and corresponding normal; one clone of in situ cervical cancer (CIS), were investigated with microarray technique for analysis of differential gene expression profile. The expression of ~ 41,000 genes were compared and 598 genes resulted differentially expressed between CIN and NUC. Among 598 genes, 152 genes were upregulated and 262 were down-regulated. We selected some genes with a greater than 2 fold difference between CIN and NUC for validation of microarray results with RTqPCR. Some genes were selected as possible candidates involved in progression from CIN1 to CIN3/CIS. Among up-regulated we identified PFKFB3, FOXD2, HOXB3, HOXB4, HOXA3, HOXA4, HOXA5, EMX2, WNT4 and among down-regulated APOBEC3B, APOBEC3F, PTPN3, CLDN11, S1PR5, IL1B. These results will need to be validated in a larger series of dysplasia to identify whether our selected genes could play a role in cervical tumorigenesis. In conclusion, our study reports, for the first time, a co-culture system of keratinocytes from minute CIN and normal cervix biopsies and offers a potential of developing newer diagnostic markers and therapeutic targets for the prevention and treatment of the cervical carcinoma

Carbon nanotubes for organ regeneration: an electrifying performance

Tissue regeneration research is advancing rapidly, thanks to the innovation potential of stem cells and nanomaterials. In particular, carbon nanotubes (CNTs) have shown an unmatched performance in conductive tissue regeneration. When grown in contact with CNTs, conducting cells become \u201celectrified\u201d, i.e., electrically more active, mature, and better interconnected. The challenges inherent in translating these concepts into 3D printing of whole organs for biomedical use are being addressed worldwide

The Glitter of Carbon Nanostructures in Hybrid/Composite Hydrogels for Medicinal Use

In recent years, we have witnessed to fast developments in the medicinal field of hydrogels containing various forms of integrated nanostructured carbon that adds interesting mechanical, thermal, and electronic properties. Besides key advances in tissue engineering (especially for conductive tissue, such as for the brain and the heart), there has been innovation also in the area of drug delivery on-demand, with engineered hydrogels capable of repeated response to light, thermal, or electric stimuli. This mini-review focusses on the most promising developments as applied to the gelation of protein/ peptide (including self-assembling amino acids and low-molecular-weight gelators), polysaccharide, and/or synthetic polymer components in medicine. The emerging field of graphene-only hydrogels is also briefly discussed, to give the reader a full flavor of the rising new paradigms in medicine that are made possible through the integration of nanostructured carbon (e.g., carbon nanotubes, nanohorns, nanodiamonds, fullerene, etc.). Nanocarbons are offering great opportunities to bring on a revolution in therapy that the modern medicinal chemist needs to master, to realise their full potential into powerful therapeutic solutions for the patient

Finite Element Analysis to Study Percutaneous Heart Valves

Communications engineering / telecommunication

Computational Fluid Dynamic Analysis of the Left Atrial Appendage to Predict Thrombosis Risk

During Atrial Fibrillation (AF) more than 90% of the left atrial thrombi responsible for thromboembolic events originate in the left atrial appendage (LAA), a complex small sac protruding from the left atrium (LA). Current available treatments to prevent thromboembolic events are oral anticoagulation, surgical LAA exclusion, or percutaneous LAA occlusion. However, the mechanism behind thrombus formation in the LAA is poorly understood. The aim of this work is to analyse the hemodynamic behaviour in four typical LAA morphologies - “Chicken wing”, “Cactus”, “Windsock” and “Cauliflower” - to identify potential relationships between the different shapes and the risk of thrombotic events. Computerised tomography (CT) images from four patients with no LA pathology were segmented to derive the 3D anatomical shape of LAA and LA. Computational Fluid Dynamic (CFD) analyses based on the patient-specific anatomies were carried out imposing both healthy and AF flow conditions. Velocity and shear strain rate (SSR) were analysed for all cases. Residence time in the different LAA regions was estimated with a virtual contrast agent washing out. CFD results indicate that both velocity and SSR decrease along the LAA, from the ostium to the tip, at each instant in the cardiac cycle, thus making the LAA tip more prone to fluid stagnation, and therefore to thrombus formation. Velocity and SSR also decrease from normal to AF conditions. After four cardiac cycles, the lowest washout of contrast agent was observed for the Cauliflower morphology (3.27% of residual contrast in AF), and the highest for the Windsock (0.56% of residual contrast in AF). This suggests that the former is expected to be associated with a higher risk of thrombosis, in agreement with clinical reports in the literature. The presented computational models highlight the major role played by the LAA morphology on the hemodynamics, both in normal and AF conditions, revealing the potential support that numerical analyses can provide in the stratification of patients under risk of thrombus formation, towards personalised patient care

Global profiling of viral and cellular non-coding RNAs in Epstein-Barr virus-induced lymphoblastoid cell lines and released exosome cargos.

Abstract The human EBV-transformed lymphoblastoid cell line (LCL), obtained by infecting peripheral blood monocular cells with Epstein–Barr Virus, has been extensively used for human genetic, pharmacogenomic, and immunologic studies. Recently, the role of exosomes has also been indicated as crucial in the crosstalk between EBV and the host microenvironment. Because the role that the LCL and LCL exosomal cargo might play in maintaining persistent infection, and since little is known regarding the non-coding RNAs of LCL, the aim of our work was the comprehensive characterization of this class of RNA, cellular and viral miRNAs, and cellular lncRNAs, in LCL compared with PBMC derived from the same donors. In this study, we have demonstrated, for the first time, that all the viral miRNAs expressed by LCL are also packaged in the exosomes, and we found that two miRNAs, ebv-miR-BART3 and ebv-miR-BHRF1-1, are more abundant in the exosomes, suggesting a microvescicular viral microRNA transfer. In addition, lncRNA profiling revealed that LCLs were enriched in lncRNA H19 and H19 antisense, and released these through exosomes, suggesting a leading role in the regulation of the tumor microenvironment

Numerical model of a valvuloplasty balloon: in vitro validation in a rapid‑prototyped phantom

Background Patient-specific simulations can provide insight into the mechanics of cardiovascular procedures. Amongst cardiovascular devices, non-compliant balloons are used in several minimally invasive procedures, such as balloon aortic valvuloplasty. Although these balloons are often included in the computer simulations of these procedures, validation of the balloon behaviour is often lacking. We therefore aim to create and validate a computational model of a valvuloplasty balloon. Methods A finite element (FE) model of a valvuloplasty balloon (Edwards 9350BC23) was designed, including balloon geometry and material properties from tensile testing. Young’s Modulus and distensibility of different rapid prototyping (RP) rubber-like materials were evaluated to identify the most suitable compound to reproduce the mechanical properties of calcified arteries in which such balloons are likely to be employed clinically. A cylindrical, simplified implantation site was 3D printed using the selected material and the balloon was inflated inside it. The FE model of balloon inflation alone and its interaction with the cylinder were validated by comparison with experimental Pressure–Volume (P–V) and diameter–Volume (d–V) curves. Results Root mean square errors (RMSE) of pressure and diameter were RMSE P = 161.98 mmHg (3.8 % of the maximum pressure) and RMSE d = 0.12 mm (<0.5 mm, within the acquisition system resolution) for the balloon alone, and RMSE P = 94.87 mmHg (1.9 % of the maximum pressure) and RMSE d = 0.49 mm for the balloon inflated inside the simplified implantation site, respectively. Conclusions This validated computational model could be used to virtually simulate more realistic valvuloplasty interventions

Numerical model of a valvuloplasty balloon:in vitro validation in a rapid-prototyped phantom

BACKGROUND: Patient-specific simulations can provide insight into the mechanics of cardiovascular procedures. Amongst cardiovascular devices, non-compliant balloons are used in several minimally invasive procedures, such as balloon aortic valvuloplasty. Although these balloons are often included in the computer simulations of these procedures, validation of the balloon behaviour is often lacking. We therefore aim to create and validate a computational model of a valvuloplasty balloon. METHODS: A finite element (FE) model of a valvuloplasty balloon (Edwards 9350BC23) was designed, including balloon geometry and material properties from tensile testing. Young’s Modulus and distensibility of different rapid prototyping (RP) rubber-like materials were evaluated to identify the most suitable compound to reproduce the mechanical properties of calcified arteries in which such balloons are likely to be employed clinically. A cylindrical, simplified implantation site was 3D printed using the selected material and the balloon was inflated inside it. The FE model of balloon inflation alone and its interaction with the cylinder were validated by comparison with experimental Pressure–Volume (P–V) and diameter–Volume (d–V) curves. RESULTS: Root mean square errors (RMSE) of pressure and diameter were RMSE(P) = 161.98 mmHg (3.8 % of the maximum pressure) and RMSE(d) = 0.12 mm (<0.5 mm, within the acquisition system resolution) for the balloon alone, and RMSE(P) = 94.87 mmHg (1.9 % of the maximum pressure) and RMSE(d) = 0.49 mm for the balloon inflated inside the simplified implantation site, respectively. CONCLUSIONS: This validated computational model could be used to virtually simulate more realistic valvuloplasty interventions

Increase in FOXP3+ Regulatory T Cells in GVHD Skin Biopsies Is Associated with Lower Disease Severity and Treatment Response

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