Dual ifgMosaic: A Versatile Method for Multispectral and Combinatorial Mosaic Gene-Function Analysis

Improved methods for manipulating and analyzing gene function have provided a better understanding of how genes work during organ development and disease. Inducible functional genetic mosaics can be extraordinarily useful in the study of biological systems; however, this experimental approach is still rarely used in vertebrates. This is mainly due to technical difficulties in the assembly of large DNA constructs carrying multiple genes and regulatory elements and their targeting to the genome. In addition, mosaic phenotypic analysis, unlike classical single gene-function analysis, requires clear labeling and detection of multiple cell clones in the same tissue. Here, we describe several methods for the rapid generation of transgenic or gene-targeted mice and embryonic stem (ES) cell lines containing all the necessary elements for inducible, fluorescent, and functional genetic mosaic (ifgMosaic) analysis. This technology enables the interrogation of multiple and combinatorial gene function with high temporal and cellular resolution.This work was supported by grants to the PI R.B. from the Spanish Ministry of Economy, Industry and Competitiveness (SAF2013-44329-P, SAF2013-42359-ERC, and RYC-2013-13209) and European Research Council (ERC-2014-StG - 638028). S.P.-Q., M.F.-C., and I.G.-G. were supported by PhD fellowships from Fundacion La Caixa (CX-SO-2013-02, CX\_E-2015-01, and CX-SO-16-1, respectively). W.L. by a FP7-PEOPLE-2012-COFUND GA600396 postdoctoral contract. We thank Simon Bartlett for English editing, Ralf H. Adams for sharing the Cdh5(PAC)-CreERT2 mice, Jose Luis de La Pompa for comments throughout the project and for sharing the Tie2-Cre mice, Gonzalo Gancedo for the help with the mouse colony, Valeria Caiolfa for the help with the microscopy, and all the members of the CNIC gene targeting, transgenesis, cellomics, and microscopy units. The CNIC is supported by MEIC/MINECO and the Pro CNIC Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505).S

Glycosaminoglycan and Proteoglycan Biotherapeutics in Articular Cartilage Protection and Repair Strategies: Novel Approaches to Visco?supplementation in Orthobiologics

The aim of this study is to review developments in glycosaminoglycan and proteoglycan research relevant to cartilage repair biology and in particular the treatment of osteoarthritis (OA). Glycosaminoglycans decorate a diverse range of extracellular matrix and cell associated proteoglycans conveying structural organization and physico‐chemical properties to tissues. They play key roles mediating cellular interactions with bioactive growth factors, cytokines, and morphogenetic proteins, and structural fibrillar collagens, cell interactive and extracellular matrix proteoglycans, and glycoproteins which define tissue function. Proteoglycan degradation detrimentally affects tissue functional properties. Therapeutic strategies have been developed to counter these degenerative changes. Neo‐proteoglycans prepared from chondroitin sulfate or hyaluronan and hyaluronan or collagen‐binding peptides emulate the interactive, water imbibing, weight bearing, and surface lubricative properties of native proteoglycans. Many neo‐proteoglycans outperform native proteoglycans in terms of water imbibition, matrix stabilization, and resistance to proteolytic degradation. The biospecificity of recombinant proteoglycans however, provides precise attachment to native target molecules. Visco‐supplements augmented with growth factors/therapeutic cells, hyaluronan, and lubricin (orthobiologicals) have the capacity to lubricate and protect cartilage, control inflammation, and promote cartilage repair and regeneration of early cartilage lesions and may represent a more effective therapeutic approach to the treatment of mild to moderate OA and deserve further study

Coronary vessel formation in development and disease: mechanisms and insights for therapy

The formation of new blood vessels after myocardial infarction (MI) is essential for the survival of existing and regenerated cardiac tissue. However, the extent of endogenous revascularization after MI is insufficient, and MI can often result in ventricular remodelling, progression to heart failure and premature death. The neutral results of numerous clinical trials that have evaluated the efficacy of angiogenic therapy to revascularize the infarcted heart reflect our poor understanding of the processes required to form a functional coronary vasculature. In this Review, we describe the latest advances in our understanding of the processes involved in coronary vessel formation, with mechanistic insights taken from developmental studies. Coronary vessels originate from multiple cellular sources during development and form through a number of distinct and carefully orchestrated processes. The ectopic reactivation of developmental programmes has been proposed as a new paradigm for regenerative medicine, therefore, a complete understanding of these processes is crucial. Furthermore, knowledge of how these processes differ between the embryonic and adult heart, and how they might be more closely recapitulated after injury are critical for our understanding of regenerative biology, and might facilitate the identification of tractable molecular targets to therapeutically promote neovascularization and regeneration of the infarcted heart