Short link N acts as a disease modifying osteoarthritis drug

Osteoarthritis (OA) is a degenerative joint disease characterised by a progressive degradation of articular cartilage and underlaying bone and is associated with pain and disability. Currently, there is no medical treatment to reverse or even retard OA. Based on our previous reports, where we establish the repair potential of short Link N (sLN) in the intervertebral disc, a cartilage-like tissue, we hypothesise that sLN may hold similar promises in the repair of articular cartilage. This study aimed to determine if sLN, could prevent OA disease progression. Skeletally mature New Zealand white rabbits underwent unilateral anterior cruciate ligament transection (ACLT) of their left femorotibial joints to induce joint degeneration typical of OA. Beginning 3 weeks post-operatively, and every three weeks thereafter for 12 weeks, either saline (1 mL) or sLN (100 μg in 1 mL saline) was injected intraarticularly into the operated knee. Six additional rabbits underwent sham surgery but without ACLT or post-operative injections. The effects on gross joint morphology and cartilage histologic changes were evaluated. In the Saline group, prominent erosion of articular cartilage occurred in both femoral condyle compartments and the lateral compartment of the tibial plateau while, sLN treatment reduced the severity of the cartilage damage in these compartments of the knee showing erosion. Furthermore, statistically significant differences were detected between the joint OA score of the saline and sLN treated groups (p = 0.0118). Therefore, periodic intraarticular injection of sLN is a promising nonsurgical treatment for preventing or retarding OA progression, by reducing cartilage degradation

The Potential of N-Rich Plasma-Polymerized Ethylene (PPE:N) Films for Regulating the Phenotype of the Nucleus Pulposus

We recently developed a nitrogen-rich plasma-polymerized biomaterial, designated “PPE:N” (N-doped plasma-polymerized ethylene) that is capable of suppressing cellular hypertrophy while promoting type I collagen and aggrecan expression in mesenchymal stem cells from osteoarthritis patients. We then hypothesized that these surfaces would form an ideal substrate on which the nucleus pulposus (NP) phenotype would be maintained. Recent evidence using microarrays showed that in young rats, the relative mRNA levels of glypican-3 (GPC3) and pleiotrophin binding factor (PTN) were significantly higher in nucleus pulposus (NP) compared to annulus fibrosus (AF) and articular cartilage. Furthermore, vimentin (VIM) mRNA levels were higher in NP versus articular cartilage. In contrast, the levels of expression of cartilage oligomeric matrix protein (COMP) and matrix gla protein precursor (MGP) were lower in NP compared to articular cartilage. The objective of this study was to compare the expression profiles of these genes in NP cells from fetal bovine lumbar discs when cultured on either commercial polystyrene (PS) tissue culture dishes or on PPE:N with time. We found that the expression of these genes varies with the concentration of N ([N]). More specifically, the expression of several genes of NP was sensitive to [N], with a decrease of GPC3, VIM, PTN, and MGP in function of decreasing [N]. The expression of aggrecan, collagen type I, and collagen type II was also studied: no significant differences were observed in the cells on different surfaces with different culture time. The results support the concept that PPE:N may be a suitable scaffold for the culture of NP cells. Further studies are however necessary to better understand their effects on cellular phenotypes

Human MMP28 expression is unresponsive to inflammatory stimuli and does not correlate to the grade of intervertebral disc degeneration

BACKGROUND: MMP28 (epilysin) is a recently discovered member of the MMP (matrix metalloproteinase) family that is, amongst others, expressed in osteoarthritic cartilage and intervertebral disc (IVD) tissue. In this study the hypothesis that increased expression of MMP28 correlates with higher grades of degeneration and is stimulated by the presence of proinflammatory molecules was tested. Gene expression levels of MMP28 were investigated in traumatic and degenerative human IVD tissue and correlated to the type of disease and the degree of degeneration (Thompson grade). Quantification of MMP28 gene expression in human IVD tissue or in isolated cells after stimulation with the inflammatory mediators lipopolysaccharide (LPS), interleukin (IL)-1β, tumor necrosis factor (TNF)-α or the histondeacetylase inhibitor trichostatin A was performed by real-time RT PCR. RESULTS: While MMP28 expression was increased in individual cases with trauma or disc degeneration, there was no significant correlation between the grade of disease and MMP28 expression. Stimulation with LPS, IL-1β, TNF-α or trichostatin A did not alter MMP28 gene expression at any investigated time point or any concentration. CONCLUSIONS: Our results demonstrate that gene expression of MMP28 in the IVD is not regulated by inflammatory mechanisms, is donor-dependent and cannot be positively or negatively linked to the grade of degeneration and only weakly to the occurrence of trauma. New hypotheses and future studies are needed to find the role of MMP28 in the intervertebral disc

Anatomical heterogeneity of tendon: Fascicular and interfascicular tendon compartments have distinct proteomic composition

This study was funded by the BBSRC (BB/K008412/1)

Local deformation in a hydrogel induced by an external magnetic field

The aim of this study is to prove the feasibility of a system able to apply local mechanical loading on cells seeded in a hydrogel for tissue engineering applications. This experimental study is based on a previously developed artificial cartilage model with different concentrations of poly(vinyl alcohol) (PVA) that simulates the cartilage extracellular matrix (ECM). Poly(l-lactic acid) (PLLA) microspheres with dispersed magnetic nanoparticles (MNPs) were produced with an emulsion method. These microspheres were embedded in aqueous PVA solutions with varying concentration to resemble increased viscosity of growing tissue during regeneration. The ability to induce a local deformation in the ECM was assessed by applying a steady or an oscillatory magnetic field gradient to different PVA solutions containing the magnetic microparticles, similarly as in ferrogels. PLLA microparticle motion was recorded, and the images were analyzed. Besides, PVA gels and PLLA microparticles were introduced into the pores of a polycaprolactone scaffold, and the microparticle distribution and the mechanical properties of the construct were evaluated. The results of this experimental model show that the dispersion of PLLA microparticles containing MNPs, together with cells in a supporting gel, will allow applying local mechanical stimuli to cells during tissue regeneration. This local stimulation can have a positive effect on the differentiation of seeded cells and improve tissue regeneration.The authors gratefully acknowledge the financial support from the Spanish Ministry of Economy and Competitiveness through the MAT2013-46467-C4-1-R project, including the Feder funds. CIBER-BBN is an initiative funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program. CIBER Actions are financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. The authors thank "Servicio de Microscopia Electronica" of Universitat Politecnica de Valencia for their invaluable help. The translation of this paper was funded by the Universitat Politecnica de Valencia, Spain.Vikingsson, L.; Vinals Guitart, Á.; Valera Martínez, A.; Riera Guasp, J.; Vidaurre Garayo, AJ.; Gallego Ferrer, G.; Gómez Ribelles, JL. (2016). Local deformation in a hydrogel induced by an external magnetic field. Journal of Materials Science. 51(22):9979-9990. https://doi.org/10.1007/s10853-016-0226-8S997999905122Eyre D (2002) Collagen of articular cartilage. Arthritis Res 4:30–35Roughley PJ, Lee ER (1994) Cartilage proteoglycans: structure and potential functions. Microsc Res Tech 28:385–397Gillard GC, Reilly HC, Bell-Booth PG, Flint MH (1979) The influence of mechanical forces on the glycosaminoglycan content of the rabbit flexor digitorum profundus tendon. 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Cross-Sample Validation Provides Enhanced Proteome Coverage in Rat Vocal Fold Mucosa

The vocal fold mucosa is a biomechanically unique tissue comprised of a densely cellular epithelium, superficial to an extracellular matrix (ECM)-rich lamina propria. Such ECM-rich tissues are challenging to analyze using proteomic assays, primarily due to extensive crosslinking and glycosylation of the majority of high Mr ECM proteins. In this study, we implemented an LC-MS/MS-based strategy to characterize the rat vocal fold mucosa proteome. Our sample preparation protocol successfully solubilized both proteins and certain high Mr glycoconjugates and resulted in the identification of hundreds of mucosal proteins. A straightforward approach to the treatment of protein identifications attributed to single peptide hits allowed the retention of potentially important low abundance identifications (validated by a cross-sample match and de novo interpretation of relevant spectra) while still eliminating potentially spurious identifications (global single peptide hits with no cross-sample match). The resulting vocal fold mucosa proteome was characterized by a wide range of cellular and extracellular proteins spanning 12 functional categories

Loss-of-function mutations in the X-linked biglycan gene cause a severe syndromic form of thoracic aortic aneurysms and dissections.

Thoracic aortic aneurysm and dissection (TAAD) is typically inherited in an autosomal dominant manner, but rare X-linked families have been described. So far, the only known X-linked gene is FLNA, which is associated with the periventricular nodular heterotopia type of Ehlers-Danlos syndrome. However, mutations in this gene explain only a small number of X-linked TAAD families. We performed targeted resequencing of 368 candidate genes in a cohort of 11 molecularly unexplained Marfan probands. Subsequently, Sanger sequencing of BGN in 360 male and 155 female molecularly unexplained TAAD probands was performed. We found five individuals with loss-of-function mutations in BGN encoding the small leucine-rich proteoglycan biglycan. The clinical phenotype is characterized by early-onset aortic aneurysm and dissection. Other recurrent findings include hypertelorism, pectus deformity, joint hypermobility, contractures, and mild skeletal dysplasia. Fluorescent staining revealed an increase in TGF-β signaling, evidenced by an increase in nuclear pSMAD2 in the aortic wall. Our results are in line with those of prior reports demonstrating that Bgn-deficient male BALB/cA mice die from aortic rupture. In conclusion, BGN gene defects in humans cause an X-linked syndromic form of severe TAAD that is associated with preservation of elastic fibers and increased TGF-β signaling.Genet Med 19 4, 386-395

Knock-Down of Cathepsin D Affects the Retinal Pigment Epithelium, Impairs Swim-Bladder Ontogenesis and Causes Premature Death in Zebrafish

The lysosomal aspartic protease Cathepsin D (CD) is ubiquitously expressed in eukaryotic organisms. CD activity is essential to accomplish the acid-dependent extensive or partial proteolysis of protein substrates within endosomal and lysosomal compartments therein delivered via endocytosis, phagocytosis or autophagocytosis. CD may also act at physiological pH on small-size substrates in the cytosol and in the extracellular milieu. Mouse and fruit fly CD knock-out models have highlighted the multi-pathophysiological roles of CD in tissue homeostasis and organ development. Here we report the first phenotypic description of the lack of CD expression during zebrafish (Danio rerio) development obtained by morpholino-mediated knock-down of CD mRNA. Since the un-fertilized eggs were shown to be supplied with maternal CD mRNA, only a morpholino targeting a sequence containing the starting ATG codon was effective. The main phenotypic alterations produced by CD knock-down in zebrafish were: 1. abnormal development of the eye and of retinal pigment epithelium; 2. absence of the swim-bladder; 3. skin hyper-pigmentation; 4. reduced growth and premature death. Rescue experiments confirmed the involvement of CD in the developmental processes leading to these phenotypic alterations. Our findings add to the list of CD functions in organ development and patho-physiology in vertebrates