A genome-wide identification and comparative analysis of the lentil MLO genes

Revista electrónica on linePowdery mildew is a widespread fungal plant disease that can cause significant losses in many crops. Some MLO genes (Mildew resistance locus O) have proved to confer a durable resistance to powdery mildew in several species. Resistance granted by the MLO gene family members has prompted an increasing interest in characterizing these genes and implementing their use in plant breeding. Lentil (Lens culinaris Medik.) is a widely grown food legume almost exclusively consumed as dry seed with an average world production of 4.5 million tons. Powdery mildew causes severe losses on certain lentil cultivars under particular environmental conditions. Data mining of the lentil CDC Redberry draft genome allowed to identify up to 15 gene sequences with homology to known MLO genes, designated as LcMLOs. Further characterization of these gene sequences and their deduced protein sequences demonstrated conformity with key MLO protein characteristics such as the presence of transmembrane and calmodulin binding domains, as well as that of other conserved motifs. Phylogenetic and other comparative analyses revealed that LcMLO1 and LcMLO3 are the most likely gene orthologs related to powdery mildew response in other species, sharing a high similarity with other known resistance genes of dicot species, such as pea PsMLO1 and Medicago truncatula MtMLO1 and MtMLO3. Sets of primers were designed as tools to PCR amplify the genomic sequences of LcMLO1 and LcMLO3, also to screen lentil germplasm in search of resistance mutants. Primers were used to obtain the complete sequences of these two genes in all of the six wild lentil relatives. Respective to each gene, all Lens sequences shared a high similarity. Likewise, we used these primers to screen a working collection of 58 cultivated and 23 wild lentil accessions in search of length polymorphisms present in these two genes. All these data widen the insights on this gene family and can be useful for breeding programs in lentil and close related species.S

Combining electrospinning and cell sheet technology for the development of a multiscale tissue engineered ligament construct (TELC)

Ligament tissue rupture is a common sport injury. Although current treatment modalities can achieve appropriate reconstruction of the damaged ligament, they present significant drawbacks, mostly related to reduced tissue availability and pain associated with tissue harvesting. Stem cell based tissue regeneration combined with electrospun scaffolds represents a novel treatment method for torn ligaments. In this study, a low fiber density polycaprolactone (PCL) electrospun mesh and sheep mesenchymal stem cells (sMSCs) were used to develop tissue engineered ligament construct (TELC) in vitro. The assembly of the TELC was based on the spontaneous capacity of the cells to organize themselves into a cell sheet once seeded onto the electrospun mesh. The cell sheet matured over 4 weeks and strongly integrated with the low fiber density electrospun mesh which was subsequently processed into a ligament-like bundle and braided with two other bundles to develop the final construct. Live/dead assay revealed that the handling of the construct through the various phases of assembly did not cause significant difference in viability compared to the control. Mechanical evaluation demonstrated that the incorporation of the cell sheet into the braided construct resulted in significantly modifying the mechanical behavior. A stress/displacement J-curve was observed for the TELC that was similar to native ligament, whereas this particular feature was not observed in the non-cellularized specimens. The regenerative potential of the TELC was evaluated ectopically in immunocompromized rats, compared to non cellularized electrospun fiber mesh and this demonstrated that the TELC was well colonized by host cells and that a significant remodelling of the implanted construct was observed. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017

Additively manufactured biphasic construct loaded with BMP-2 for vertical bone regeneration: a pilot study in rabbit

Vertical bone augmentation of the jaws is required when the height of bone is insufficient at the site of dental implant placement. In this proof of concept study, we investigated the potential of a biphasic polycaprolactone construct combined with a hyaluronic acid based hydrogel loaded with recombinant human bone morphogenetic growth factor-2 (BMP-2) for vertical bone regeneration. The biphasic scaffold consisted of an outer shell manufactured by fused deposition modelling, mimicking native cortical bone and providing mechanical and space maintenance properties essential for bone formation. Within this shell, a 90% porous melt electrospun microfibrous mesh mimicking the architecture of cancellous bone was incorporated in order to facilitate hydrogel loading and subsequent osteogenesis and angiogenesis. The in vitro performances of the biphasic construct demonstrated that BMP-2 was released in a sustained manner over several weeks and that cell viability was maintained in the hydrogel over 21 days. qRT-PCR demonstrated the upregulation of bone markers such as osteopontin, osteocalcin and collagen 1A1 at day 3 and 14 in the constructs loaded with BMP2. In vivo assessment of the biphasic scaffold was performed using a dose of 30 μg of BMP-2 in a rabbit calvarial vertical bone augmentation model. The histology and micro-CT analysis of the elevated space demonstrated that the hydrogel and the presence of BMP-2 enabled bone formation. However, this was limited to the immediate vicinity of the calvarial bone. The amount of newly formed bone was relatively small which was likely due to poor vascularisation of the extraskeletal space. The utilisation of this biomimetic biphasic construct with excellent space maintenance properties can be of interest in dentistry although the in vivo model requires refinement to demonstrated appropriate efficacy

Titania nanopores with dual micro-/nano-topography for selective cellular bioactivity

This letter describes a simple surface modification strategy based on a single-step electrochemical anodization towards generating dual micro- and nano-rough horizontally-aligned TiO nanopores on the surface of clinically utilized micro-grooved titanium implants. Primary macrophages, osteoblasts and fibroblasts were cultured on the nano-engineered implants, and it was demonstrated that the modified surfaces selectively reduced the proliferation of macrophages (immunomodulation), while augmenting the activity of osteoblasts (osseo-integration) and fibroblasts (soft-tissue integration). Additionally, the mechanically robust nanopores also stimulated osteoblast and fibroblast adhesion, attachment and alignment along the direction of the pores/grooves, while macrophages remained oval-shaped and sparsely distributed. This study for the first time reports the use of cost-effectively prepared nano-engineered titanium surface via anodization, with aligned multi-scale micro/nano features for selective cellular bioactivity, without the use of any therapeutics

Chitin and chitosan in selected biomedical applications

Chitin (CT), the well-known natural biopolymer and chitosan (CS) (bio-based or "artificial polymer") are non-toxic, biodegradable and biocompatible in nature. The advantages of these biomaterials are such that, they can be easily processed into different forms such as membranes, sponges, gels, scaffolds, microparticles, nanoparticles and nanofibers for a variety of biomedical applications such as drug delivery, gene therapy, tissue engineering and wound healing. Present review focuses on the diverse applications of CT and CS membranes and scaffolds for drug delivery, tissue engineering and targeted regenerative medicine. The chitinous scaffolds of marine sponges' origin are discussed here for the first time. These CT based scaffolds obtained from Porifera possess remarkable and unique properties such as hydration, interconnected channels and diverse structural architecture. This review will provide a brief overview of CT and CS membranes and scaffolds toward different kinds of delivery applications such as anticancer drug delivery, osteogenic drug delivery, and growth factor delivery, because of their inimitable release behavior, degradation profile, mucoadhesive nature, etc. The review also provides an overview of the key features of CT and CS membranes and scaffolds such as their biodegradability, cytocompatibility and mechanical properties toward applications in tissue engineering and wound healing

Surface modification of 3D printed polycaprolactone constructs via a solvent treatment: impact on physical and osteogenic properties

One promising strategy to reconstruct bone defects relies on 3D printed porous structures. In spite of several studies having been carried out to fabricate controlled, interconnected porous constructs, the control over surface features at, or below, the microscopic scale remains elusive for 3D polymeric scaffolds. In this study, we developed and refined a methodology which can be applied to homogeneously and reproducibly modify the surface of polymeric 3D printed scaffolds. We have demonstrated that the combination of a polymer solvent and the utilization of ultrasound was essential for achieving appropriate surface modification without damaging the structural integrity of the construct. The modification created on the scaffold profoundly affected the macroscopic and microscopic properties of the scaffold with an increased roughness, greater surface area, and reduced hydrophobicity. Furthermore, to assess the performance of such materials in bone tissue engineering, human mesenchymal stem cells (hMSC) were cultured in vitro on the scaffolds for up to 7 days. Our results demonstrate a stronger commitment toward early osteogenic differentiation of hMSC. Finally, we demonstrated that the increased in the specific surface area of the scaffold did not necessarily correlate with improved adsorption of protein and that other factors, such as surface chemistry and hydrophilicity, may also play a major role