The role of Periostin in regulating the biomechanical properties of cushion tissue

Abstract only availableDuring embryonic heart development the atrio-ventricular (AV) cushions swell and fuse to form the valves and septa of the adult heart. Initially, the cushions appear as swellings on the interior wall of the AV canal and eventually fuse to form the septum and valvular leaflets. The morphogenetic event that the cushions undergo during the fusion process is, in part, driven by the cohesive energy of the tissue, which can be described by the tissue's surface tension. It has been shown earlier that many properties of embryonic tissues can be interpreted by using the analogy that they behave as liquids and it is this analogy that gives rise to apparent tissue surface tension. Periostin is hypothesized to affect cushion tissue surface tension, through its possible binding of the extracellular matrix of the tissue. In this study virus containing the sense strand of Periostin DNA is introduced into hanging drops containing living explants of AV cushion tissue. Overnight the tissue explants rounded up to form spheroids allowing their surface tension to be measured and compared to the surface tension of AV cushion tissue explants exposed to a LacZ promoter control virus. The surface tension was determined using a specifically designed apparatus that measures the viscoelastic response of spherical explants due to a compressive force. It was expected that the increased production of Periostin in the cushion explants due to exposure to the virus will result in an increased surface tension compared to that of explants exposed to the control virus. The preliminary results of the experiment have displayed no significant difference of surface tension between the control virus and the Periostin virus. Since earlier research has shown a significant difference in the rate of fusion of cushions exposed to Periostin DNA virus and those exposed to the control virus, and because fusion time is characterized by the ratio of the surface tension and the viscosity of the tissue, we believe that Periostin may be affecting the viscosity of the tissue explants instead of the surface tension.NSF-REU Program in Biosystems Modeling and Analysi

A potential mouse model for the erosive vitreoretinopathy of Wagner disease

Patients with the very rare eye pathology Wagner disease (OMIM #143200) present with an abnormal (empty) vitreous, retinal detachment and altered electroretinogram (ERG). The disease is progressive and can eventually lead to blindness. No therapy can be offered to date. The genetic basis is the presence of mutations in the VCAN gene, encoding the large extracellular matrix molecule versican, which is a component of the vitreous. All identified mutations map to the canonical splice sites flanking exon 8, resulting in low number of aberrant splice products and a severe increase in two (V2, V3) of the four naturally occurring splice variants. The pathomechanism of Wagner's disease is poorly understood and a mouse model may afford further insight. The hdf -/- mice, named for their initial phenotype of heart defects, carry a null allele for Vcan that leads to embryonic lethality when homozygous, but heterozygote animals are viable. Here we investigated a possible eye phenotype in the heterozygous animals. While the overall morphology of retina and ciliary body appears to be normal, older (17 months) mutant animals show a decrease in ERG signaling profiles affecting the a-, b- and c-waves. This aspect of altered ERG profile demonstrates similarities to the human disease manifestation and underlines the suitability of heterozygous hdf+/- mice as a model for Wagner disease

Fibulin-1 is required for morphogenesis of neural crest-derived structures

AbstractHere we report that mouse embryos homozygous for a gene trap insertion in the fibulin-1 (Fbln1) gene are deficient in Fbln1 and exhibit cardiac ventricular wall thinning and ventricular septal defects with double outlet right ventricle or overriding aorta. Fbln1 nulls also display anomalies of aortic arch arteries, hypoplasia of the thymus and thyroid, underdeveloped skull bones, malformations of cranial nerves and hemorrhagic blood vessels in the head and neck. The spectrum of malformations is consistent with Fbln1 influencing neural crest cell (NCC)-dependent development of these tissues. This is supported by evidence that Fbln1 expression is associated with streams of cranial NCCs migrating adjacent to rhombomeres 2–7 and that Fbln1-deficient embryos display patterning anomalies of NCCs forming cranial nerves IX and X, which derive from rhombomeres 6 and 7. Additionally, Fbln1-deficient embryos show increased apoptosis in areas populated by NCCs derived from rhombomeres 4, 6 and 7. Based on these findings, it is concluded that Fbln1 is required for the directed migration and survival of cranial NCCs contributing to the development of pharyngeal glands, craniofacial skeleton, cranial nerves, aortic arch arteries, cardiac outflow tract and cephalic blood vessels

The ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin motifs) family

The ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin motifs) enzymes are secreted, multi-domain matrix-associated zinc metalloendopeptidases that have diverse roles in tissue morphogenesis and patho-physiological remodeling, in inflammation and in vascular biology. The human family includes 19 members that can be sub-grouped on the basis of their known substrates, namely the aggrecanases or proteoglycanases (ADAMTS1, 4, 5, 8, 9, 15 and 20), the procollagen N-propeptidases (ADAMTS2, 3 and 14), the cartilage oligomeric matrix protein-cleaving enzymes (ADAMTS7 and 12), the von-Willebrand Factor proteinase (ADAMTS13) and a group of orphan enzymes (ADAMTS6, 10, 16, 17, 18 and 19). Control of the structure and function of the extracellular matrix (ECM) is a central theme of the biology of the ADAMTS, as exemplified by the actions of the procollagen-N-propeptidases in collagen fibril assembly and of the aggrecanases in the cleavage or modification of ECM proteoglycans. Defects in certain family members give rise to inherited genetic disorders, while the aberrant expression or function of others is associated with arthritis, cancer and cardiovascular disease. In particular, ADAMTS4 and 5 have emerged as therapeutic targets in arthritis. Multiple ADAMTSs from different sub-groupings exert either positive or negative effects on tumorigenesis and metastasis, with both metalloproteinase-dependent and -independent actions known to occur. The basic ADAMTS structure comprises a metalloproteinase catalytic domain and a carboxy-terminal ancillary domain, the latter determining substrate specificity and the localization of the protease and its interaction partners; ancillary domains probably also have independent biological functions. Focusing primarily on the aggrecanases and proteoglycanases, this review provides a perspective on the evolution of the ADAMTS family, their links with developmental and disease mechanisms, and key questions for the future

Altered versican cleavage in ADAMTS5 deficient mice : a novel etiology of myxomatous valve disease

AbstractIn fetal valve maturation the mechanisms by which the relatively homogeneous proteoglycan-rich extracellular matrix (ECM) of endocardial cushions is replaced by a specialized and stratified ECM found in mature valves are not understood. Therefore, we reasoned that uncovering proteases critical for ‘remodeling’ the proteoglycan rich (extracellular matrix) ECM may elucidate novel mechanisms of valve development. We have determined that mice deficient in ADAMTS5, (A Disintegrin-like And Metalloprotease domain with ThromboSpondin-type 1 motifs) which we demonstrated is expressed predominantly by valvular endocardium during cardiac valve maturation, exhibited enlarged valves. ADAMTS5 deficient valves displayed a reduction in cleavage of its substrate versican, a critical cardiac proteoglycan. In vivo reduction of versican, in Adamts5−/− mice, achieved through Vcan heterozygosity, substantially rescued the valve anomalies. An increase in BMP2 immunolocalization, Sox9 expression and mesenchymal cell proliferation were observed in Adamts5−/− valve mesenchyme and correlated with expansion of the spongiosa (proteoglycan-rich) region in Adamts5−/− valve cusps. Furthermore, these data suggest that ECM remodeling via ADAMTS5 is required for endocardial to mesenchymal signaling in late fetal valve development. Although adult Adamts5−/− mice are viable they do not recover from developmental valve anomalies and have myxomatous cardiac valves with 100% penetrance. Since the accumulation of proteoglycans is a hallmark of myxomatous valve disease, based on these data we hypothesize that a lack of versican cleavage during fetal valve development may be a potential etiology of adult myxomatous valve disease

Heart Fields: Spatial Polarity and Temporal Dynamics

In chick and mouse, heart fields undergo dynamic morphological spatiotemporal changes during heart tube formation. Here, the dynamic change in spatial polarity of such fields is discussed and a new perspective on the heart fields is proposed. The heart progenitor cells delaminate through the primitive streak and migrate in a semicircular trajectory craniolaterally forming the bilateral heart fields as part of the splanchnic mesoderm. They switch their polarity from anteroposterior to mediolateral. The anterior intestinal portal posterior descent inverts the newly formed heart field mediolateral polarity into lateromedial by 125° bending. The heart fields revert back to their original anteroposterior polarity and fuse at the midline forming a semi heart tube by completing their half circle movement. Several names and roles were assigned to different portions of the heart fields: posterior versus anterior, first versus second, and primary versus secondary heart field. The posterior and anterior heart fields define basically physical fields that form the inflow–outflow axis of the heart tube. The first and second heart fields are, in contrast, temporal fields of differentiating cardiomyocytes expressing myosin light chain 2a and undifferentiated and proliferating precardiac mesoderm expressing Isl1 gene, respectively. The two markers present a complementary pattern and are expressed transiently in all myocardial lineages. Thus, Isl1 is not restricted to a portion of the heart field or one of the two heart lineages as has been often assumed. Anat Rec, 297:175–182, 2014. © 2013 Wiley Periodicals, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/102627/1/ar22831.pd

ADAMTS metalloproteases generate active versican fragments that regulate interdigital web regression

SummaryWe show that combinatorial mouse alleles for the secreted metalloproteases Adamts5, Adamts20 (bt), and Adamts9 result in fully penetrant soft-tissue syndactyly. Interdigital webs in Adamts5−/−;bt/bt mice had reduced apoptosis and decreased cleavage of the proteoglycan versican; however, the BMP-FGF axis, which regulates interdigital apoptosis was unaffected. BMP4 induced apoptosis, but without concomitant versican proteolysis. Haploinsufficiency of either Vcan or Fbln1, a cofactor for versican processing by ADAMTS5, led to highly penetrant syndactyly in bt mice, suggesting that cleaved versican was essential for web regression. The local application of an aminoterminal versican fragment corresponding to ADAMTS-processed versican, induced cell death in Adamts5−/−;bt/bt webs. Thus, ADAMTS proteases cooperatively maintain versican proteolysis above a required threshold to create a permissive environment for apoptosis. The data highlight the developmental significance of proteolytic action on the ECM, not only as a clearance mechanism, but also as a means to generate bioactive versican fragments

Alteration of chondroitin sulfate composition on proteoglycan produced by knock-in mouse embryonic fibroblasts whose versican lacks the A subdomain

Versican/proteoglycan-mesenchymal (PG-M) is a large chondroitin sulfate (CS) proteoglycan of the extracellular matrix (ECM) that is constitutively expressed in adult tissues such as dermis and blood vessels. It serves as a structural macromolecule of the ECM, while in embryonic tissue it is transiently expressed at high levels and regulates cell adhesion, migration, proliferation, and differentiation. Knock-in mouse embryonic (Cspg2Δ3/Δ3) fibroblasts whose versican lack the A subdomain of the G1 domain exhibit low proliferation rates and acquire senescence. It was suspected that chondroitin sulfate on versican core protein would be altered when the A subdomain was disrupted, so fibroblasts were made from homozygous Cspg2Δ3/Δ3 mouse embryos to investigate the hypothesis. Analysis of the resulting versican deposition demonstrated that the total versican deposited in the Cspg2Δ3/Δ3 fibroblasts culture was approximately 50% of that of the wild type (WT), while the versican deposited in the ECM of Cspg2Δ3/Δ3 fibroblasts culture was 35% of that of the WT, demonstrating the lower capacity of mutant (Cspg2Δ3/Δ3) versican deposited in the ECM. The analysis of CS expression in the Cspg2Δ3/Δ3 fibroblasts culture compared with wild-type fibroblasts showed that the composition of the non-sulfate chondroitin sulfate isomer on the versican core protein increased in the cell layer but decreased in the culture medium. Interestingly, chondroitin sulfate E isomer was found in the culture medium. The amount of CS in the Cspg2Δ3/Δ3 cell layer of fibroblasts with mutant versican was dramatically decreased, contrasted to the amount in the culture medium, which increased. It was concluded that the disruption of the A subdomain of the versican molecule leads to lowering of the amount of versican deposited in the ECM and the alteration of the composition and content of CS on the versican molecule

Gene Knock-Outs of Inositol 1,4,5-Trisphosphate Receptors Types 1 and 2 Result in Perturbation of Cardiogenesis

Rs in cardiogenesis remain unclear.Rs mimicked the phenotype of the AV valve defect that result from the inhibition of calcineurin, and it could be rescued by constitutively active calcineurin.Rs in cardiogenesis in part through the regulation of calcineurin-NFAT signaling