Platelet lysate maintains chondrogenic potential and promotes cartilage regeneration

cartilage. We report the biological effect of the platelet lysate (PL), a PRP derivative, on primary human articular chondrocytes (HAC) cultured under both physiological and inflammatory condition. Added to the culture medium, PL induced a strong mitogenic response in the chondrocytes. The in vitro expanded cell population maintained a chondrogenic re-­‐differentiation potential as revealed by micromass culture in vitro as well as in vivo as demonstrated by ectopic cartilage formation in nude mice. Furthermore, in chondrocytes cultured in the presence of the pro-­‐inflammatory cytokine IL-­‐1α, the PL induced a drastic enhancement of the synthesis of the cytokines IL-­‐6 and IL-­‐8 and of NGAL, a lipocalin expressed in cells of the chondrogenic lineage. These events were controlled by the p38 MAP kinase and NF-­‐κΒ pathways. The pro-­‐inflammatory effect of the PL was a transient phenomenon. In fact, after an initial up regulation, we observed a significant reduction of the NF-­‐κΒ activity together with the repression of the inflammatory enzyme ciclooxygenase-­‐2 (COX-­‐2). Moreover, the medium of chondrocytes cultured in the contemporary presence of PL and IL-­‐1α, showed a significant enhancement of the chemoattractant activity versus untreated chondrocytes. On the whole, our findings support the concept that the platelet products have a direct beneficial effect on articular chondrocytes and at the same time could drive in sequence a trans

Encapsulation of human articular chondrocytes into 3D hydrogel : phenotype and genotype characterization

This chapter is intended to provide a summary of the current materials used in cell encapsulation technology as well as methods for evaluating the performance of cells encapsulated in a polymeric matrix. In particular, it describes the experimental procedure to prepare a hydrogel matrix based on natural polymers for encapsulating and culturing human articular chondrocytes with the interest in cartilage regeneration. Protocols to evaluate the viability, proliferation, differentiation, and matrix production of embedded cells are also described and include standard protocols such as the MTT and [3H] Thymidine assays, reverse transcription polymerase chain reaction (RT-PCR) technique, histology, and immunohistochemistry analysis. The assessment of cell distribution within the 3D hydrogel construct is also described using APoTome analysis.(undefined

Kernel Methods for Document Filtering

This paper describes the algorithms implemented by the KerMIT consortium for its participation in the Trec 2002 Filtering track. The consortium submitted runs for the routing task using a linear SVM, for the batch task using the same SVM in combination with an innovation threshold-selection mechanism, and for the adaptive task using both a second-order perceptron and a combination of SVM and perceptron with uneven margin. Results seem to indicate that these algorithm performed relatively well on the extensive TREC benchmark

Novel injectable gel (system) as a vehicle for human articular chondrocytes in cartilage tissue regeneration

We developed a novel injectable carrageenan/fibrin/hyaluronic acid-based hydrogel with in situ gelling properties to be seeded with chondrogenic cells and used for cartilage tissue engineering applications. We first analysed the distribution within the hydrogel construct and the phenotype of human articular chondrocytes (HACs) cultured for 3 weeks in vitro. We observed a statistically significant increase in the cell number during the first 2 weeks and maintenance of cell viability throughout the cell culture, together with the deposition/formation of a cartilage-specific extracellular matrix (ECM). Taking advantage of a new in vivo model that allows the integration between newly formed and preexisting cartilage in immunodeficient mice to be investigated, we showed that injectable hydrogel seeded with human articular chondrocytes was able to regenerate and repair an experimentally made lesion in bovine articular cartilage, thus demonstrating the potential of this novel cell delivery system for cartilage tissue engineering.The authors are grateful to Recco orthopaedic staff members for the collaboration and patients for bioptic material donation as well as to Mrs Daniela Marubbi for histological assistance. This work was supported by funds from the Italian MUR (FIRB-Tissuenet project), the European Union-funded STREP project, HIPPOCRATES (Grant No. NMP3-CT-2003-505758) and the European NoE EXPERTISSUES project (Grant No. NMP3-CT-2004-500283)

A preliminary approach to the multilabel classification problem of Portuguese juridical documents

Portuguese juridical documents from Supreme Courts and the Attorney General’s Office are manually classified by juridical experts into a set of classes belonging to a taxonomy of concepts. In this paper, a preliminary approach to develop techniques to automat- ically classify these juridical documents, is proposed. As basic strategy, the integration of natural language processing techniques with machine learning ones is used. Support Vector Machines (SVM) are used as learn- ing algorithm and the obtained results are presented and compared with other approaches, such as C4.5 and Naive Bayes

A cartilage tissue engineering approach combining starch-polycaprolactone fibre mesh scaffolds with bovine articular chondrocytes

In the present work we originally tested the suitability of corn starch-polycaprolactone (SPCL) scaffolds for pursuing a cartilage tissue engineering approach. Bovine articular chondrocytes were seeded on SPCL scaffolds under dynamic conditions using spinner flasks (total of 4 scaffolds per spinner flask using cell suspensions of 0.5×106 cells/ml) and cultured under orbital agitation for a total of 6 weeks. Poly(glycolic acid) (PGA) non-woven scaffolds and bovine native articular cartilage were used as standard controls for the conducted experiments. PGA is a kind of standard in tissue engineering approaches and it was used as a control in that sense. The tissue engineered constructs were characterized at different time periods by scanning electron microscopy (SEM), hematoxylin-eosin (H&E) and toluidine blue stainings, immunolocalisation of collagen types I and II, and dimethylmethylene blue (DMB) assay for glycosaminoglycans (GAG) quantification assay. SEM results for SPCL constructs showed that the chondrocytes presented normal morphological features, with extensive cells presence at the surface of the support structures, and penetrating the scaffolds pores. These observations were further corroborated by H&E staining. Toluidine blue and immunohistochemistry exhibited extracellular matrix deposition throughout the 3D structure. Glycosaminoglycans, and collagen types I and II were detected. However, stronger staining for collagen type II was observed when compared to collagen type I. The PGA constructs presented similar features toSPCLat the end of the 6 weeks. PGA constructs exhibited higher amounts of matrix glycosaminoglycans when compared to the SPCL scaffolds. However, we also observed a lack of tissue in the central area of the PGA scaffolds. Reasons for these occurrences may include inefficient cells penetration, necrosis due to high cell densities, or necrosis related with acidic by-products degradation. Such situation was not detected in the SPCL scaffolds, indicating the much better biocompatibility of the starch based scaffolds

The anti-bacterial iron-restriction defence mechanisms of egg white; the potential role of three lipocalin-like proteins in resistance against Salmonella

Salmonella enterica serovar Enteritidis (SE) is the most frequently-detected Salmonella in foodborne outbreaks in the European Union. Among such outbreaks, egg and egg products were identified as the most common vehicles of infection. Possibly, the major antibacterial property of egg white is iron restriction, which results from the presence of the iron-binding protein, ovotransferrin. To circumvent iron restriction, SE synthesise catecholate siderophores (i.e. enterobactin and salmochelin) that can chelate iron from host iron-binding proteins. Here, we highlight the role of lipocalin-like proteins found in egg white that could enhance egg-white iron restriction through sequestration of certain siderophores, including enterobactin. Indeed, it is now apparent that the egg-white lipocalin, Ex-FABP, can inhibit bacterial growth via its siderophore-binding capacity in vitro. However, it remains unclear whether ex-FABP performs such a function in egg white or during bird infection. Regarding the two other lipocalins of egg white (Cal-γ and α-1-glycoprotein), there is currently no evidence to indicate that they sequester siderophores

Electrospun silk fibroin fibers for storage and controlled release of human platelet lysate

Human platelet lysate (hPL) is a pool of growth factors and cytokines able to induce regeneration of different tissues. Despite its good potentiality as therapeutic tool for regenerative medicine applications, hPL has been only moderately exploited in this field. A more widespread adoption has been limited because of its rapid degradation at room temperature that decreases its functionality. Another limiting factor for its extensive use is the difficulty of handling the hPL gels. In this work, silk fibroin-based patches were developed to address several points: improving the handling of hPL, enabling their delivery in a controlled manner and facilitating their storage by creating a device ready to use with expanded shelf life. Patches of fibroin loaded with hPL were synthesized by electrospinning to take advantage of the fibrous morphology. The release kinetics of the material was characterized and tuned through the control of fibroin crystallinity. Cell viability assays, performed with primary human dermal fibroblasts, demonstrated that fibroin is able to preserve the hPL biological activity and prolong its shelf-life. The strategy of storing and preserving small active molecules within a naturally-derived, protein-based fibrous scaffold was successfully implemented, leading to the design of a biocompatible device, which can potentially simplify the storage and the application of the hPL on a human patient, undergoing medical procedures such as surgery and wound care. Statement of Significance: Human platelets lysate (hPL) is a mixture of growth factors and cytokines able to induce the regeneration of damaged tissues. This study aims at enclosing hPL in a silk fibroin electrospun matrix to expand its utilization. Silk fibroin showed the ability to preserve the hPL activity at temperature up to 60 \ub0C and the manipulation of fibroin's crystallinity provided a tool to modulate the hPL release kinetic. This entails the possibility to fabricate the hPL silk fibroin patches in advance and store them, resulting in an easy and fast accessibility and an expanded use of hPL for wound healing