The effect of insulin-loaded chitosan particle-aggregated scaffolds in chondrogenic differentiation

Osteochondral defect repair requires a tissue engineering approach that aims at mimicking the physiological properties and structure of two different tissues (cartilage and bone) using a scaffold–cell construct. One ideal approach would be to engineer in vitro a hybrid material using a single-cell source. For that purpose, the scaffold should be able to provide the adequate biochemical cues to promote the selective but simultaneous differentiation of both tissues. In this work, attention was paid primarily to the chondrogenic differentiation by focusing on the development of polymeric systems that provide biomolecules release to induce chondrogenic differentiation. For that, different formulations of insulin-loaded chitosan particle–aggregated scaffolds were developed as a potential model system for cartilage and osteochondral tissue engineering applications using insulin as a potent bioactive substance known to induce chondrogenic differentiation. The insulin encapsulation efficiency was shown to be high with values of 70.37!0.8%, 84.26!1.76%, and 87.23!1.58% for loadings of 0.05%, 0.5%, and 5%, respectively. The in vitro release profiles were assessed in physiological conditions mimicking the cell culture procedures and quantified by Micro-BCA! protein assay. Different release profiles were obtained that showed to be dependent on the initial insulin-loading percentage. Further, the effect on prechondrogenic ATDC5 cells was investigated for periods up to 4 weeks by studying the influence of these release systems on cell morphology, DNA and glycosaminoglycan content, histology, and gene expression of collagen types I and II, Sox-9, and aggrecan assessed by real-time polymerase chain reaction. When compared with control conditions (unloaded scaffolds cultured with the standard chondrogenic-inducing medium), insulin-loaded scaffolds upregulated the Sox-9 and aggrecan expression after 4 weeks of culture. From the overall results, it is reasonable to conclude that the developed loaded scaffolds when seeded with ATDC5 can provide biochemical cues for chondrogenic differentiation. Among the tested formulations, the higher insulin-loaded system (5%) was the most effective in promoting chondrogenic differentiation.The authors would like to acknowledge the Portuguese Foundation for Science and Technology for the Ph. D. Grant to Patricia B. Malafaya (SFRH/BD/11155/2002). This work was partially supported and carried out under the scope of the European STREP Project HIPPOCRATES (NMP3-CT-2003-505758) and European NoE EXPERTISSUES (NMP3CT-2004-500283). The authors also like to acknowledge the Life and Health Sciences Research Institute (ICVS), University of Minho, for the use of their facilities, namely, to Luis Martins for histological sections slicing and H&E stain processing

Orthopaedic regenerative tissue engineering en route to the holy grail: disequilibrium between the demand and the supply in the operating room

Orthopaedic disorders are very frequent, globally found and often partially unresolved despite the substantial advances in science and medicine. Their surgical intervention is multifarious and the most favourable treatment is chosen by the orthopaedic surgeon on a case-by-case basis depending on a number of factors related with the patient and the lesion. Numerous regenerative tissue engineering strategies have been developed and studied extensively in laboratory through in vitro experiments and preclinical in vivo trials with various established animal models, while a small proportion of them reached the operating room. However, based on the available literature, the current strategies have not yet achieved to fully solve the clinical problems. Thus, the gold standards, if existing, remain unchanged in the clinics, notwithstanding the known limitations and drawbacks. Herein, the involvement of regenerative tissue engineering in the clinical orthopaedics is reviewed. The current challenges are indicated and discussed in order to describe the current disequilibrium between the needs and solutions made available in the operating room. Regenerative tissue engineering is a very dynamic field that has a high growth rate and a great openness and ability to incorporate new technologies with passion to edge towards the Holy Grail that is functional tissue regeneration. Thus, the future of clinical solutions making use of regenerative tissue engineering principles for the management of orthopaedic disorders is firmly supported by the clinical need.FROnTHERA (NORTE-01-0145-FEDER-000023), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). I. F. Cengiz thanks the Portuguese Foundation for Science and Technology (FCT) for the Ph.D. scholarship (SFRH/BD/99555/2014). J. M. Oliveira also thanks the FCT for the funds provided under the program Investigador FCT 2012 and 2015 (IF/00423/2012 and IF/01285/2015)info:eu-repo/semantics/publishedVersio

The microRNA-29 family in cartilage homeostasis and osteoarthritis

MicroRNAs have been shown to function in cartilage development and homeostasis, as well as in progression of osteoarthritis. The objective of the current study was to identify microRNAs involved in the onset or early progression of osteoarthritis and characterise their function in chondrocytes. MicroRNA expression in mouse knee joints post-DMM surgery was measured over 7 days. Expression of miR-29b-3p was increased at day 1 and regulated in the opposite direction to its potential targets. In a mouse model of cartilage injury and in end-stage human OA cartilage, the miR-29 family were also regulated. SOX9 repressed expression of miR-29a-3p and miR-29b-3p via the 29a/b1 promoter. TGFβ1 decreased expression of miR-29a, b and c (3p) in primary chondrocytes, whilst IL-1β increased (but LPS decreased) their expression. The miR-29 family negatively regulated Smad, NFκB and canonical WNT signalling pathways. Expression profiles revealed regulation of new WNT-related genes. Amongst these, FZD3, FZD5, DVL3, FRAT2, CK2A2 were validated as direct targets of the miR-29 family. These data identify the miR-29 family as microRNAs acting across development and progression of OA. They are regulated by factors which are important in OA and impact on relevant signalling pathways

Current trends in tendinopathy: consensus of the ESSKA basic science committee : Part I: biology, biomechanics, anatomy and an exercise-based approach

Chronic tendinopathies represent a major problem in the clinical practice of sports orthopaedic surgeons, sports doctors and other health professionals involved in the treatment of athletes and patients that perform repetitive actions. The lack of consensus relative to the diagnostic tools and treatment modalities represents a management dilemma for these professionals. With this review, the purpose of the ESSKA Basic Science Committee is to establish guidelines for understanding, diagnosing and treating this complex pathology

Mechanism of Splicing Regulation of Spinal Muscular Atrophy Genes

Spinal muscular atrophy (SMA) is one of the major genetic disorders associated with infant mortality. More than 90% cases of SMA result from deletions or mutations of Survival Motor Neuron 1 (SMN1) gene. SMN2, a nearly identical copy of SMN1, does not compensate for the loss of SMN1due to predominant skipping of exon 7. However, correction of SMN2 exon 7 splicing has proven to confer therapeutic benefits in SMA patients. The only approved drug for SMA is an antisense oligonucleotide (Spinraza™/Nusinersen), which corrects SMN2 exon 7 splicing by blocking intronic splicing silencer N1 (ISS-N1) located immediately downstream of exon 7. ISS-N1 is a complex regulatory element encompassing overlapping negative motifs and sequestering a cryptic splice site. More than 40 protein factors have been implicated in the regulation of SMN exon 7 splicing. There is evidence to support that multiple exons of SMN are alternatively spliced during oxidative stress, which is associated with a growing number of pathological conditions. Here, we provide the most up to date account of the mechanism of splicing regulation of the SMN genes

Web-based mini-games for language learning that support spoken interaction

Contains fulltext : 150810.pdf (publisher's version ) (Open Access)SLaTE 2015. Workshop on Speech and Language Technology in Education, 04 september 201