The alternatively spliced domains EIIIB and EIIIA of human fibronectin affect cell adhesion and spreading

Fibronectin has a complex pattern of alternative splicing at the pre-mRNA level leading to the expression of different isoforms. The alternatively spliced domains EIIIB and EIIIA are known to be prominently expressed during development and wound healing. While the other spliced domain (CS-segment) is known to promote cell adhesion in a cell type specific manner, the biological functions of the spliced domains EIIIB and EIIIA are not well understood. In the present study, we have prepared expression proteins of specific domains of human fibronectin using a prokaryotic expression system and used the purified fragments to test their ability to support adhesion and spreading of cultured cells. Fragments from type-III domains #7 to #12 were prepared in various combinations to include or exclude the spliced domains EIIIB and EIIIA. The results indicate that cultured NIL fibroblasts adhere to many of the fragments tested. However, the cell adhesion and spreading are enhanced, especially at lower concentrations, to fragments including the domain EIIIB. The inclusion of domain EIIIA led to a decrease in the adhesion of cells and those that adhered did not spread well. When tested in a centrifugal cell adhesion assay, fragments including domain EIIIB resisted the detaching forces and stayed adhered. Fragments that included domain EIIIA were unable to resist the detaching centrifugal forces to the same extent as the fragments that included domain EIIIB alone. These results suggest that the spliced domain EIIIB may be serving important biological functions in enhancing cell adhesion and spreading. This is likely to be mediated by conformational effects because domain EIIIB alone neither exhibited any adhesive activity nor competed in inhibiting adhesion to fragments #7-10

Expression of Extracellular Matrix Proteins in Human Periodontal Ligament Cells During Mineralization In Vitro

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/142249/1/jper0320.pd

Current concepts in periodontal bioengineering

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73112/1/j.1601-6343.2005.00352.x.pd

Exploring hypotheses of the actions of TGF-beta 1 in epidermal wound healing using a 3D computational multiscale model of the human epidermis

In vivo and in vitro studies give a paradoxical picture of the actions of the key regulatory factor TGF-beta 1 in epidermal wound healing with it stimulating migration of keratinocytes but also inhibiting their proliferation. To try to reconcile these into an easily visualized 3D model of wound healing amenable for experimentation by cell biologists, a multiscale model of the formation of a 3D skin epithelium was established with TGF-beta 1 literature-derived rule sets and equations embedded within it. At the cellular level, an agent-based bottom-up model that focuses on individual interacting units ( keratinocytes) was used. This was based on literature-derived rules governing keratinocyte behavior and keratinocyte/ECM interactions. The selection of these rule sets is described in detail in this paper. The agent-based model was then linked with a subcellular model of TGF-beta 1 production and its action on keratinocytes simulated with a complex pathway simulator. This multiscale model can be run at a cellular level only or at a combined cellular/subcellular level. It was then initially challenged ( by wounding) to investigate the behavior of keratinocytes in wound healing at the cellular level. To investigate the possible actions of TGF-beta 1, several hypotheses were then explored by deliberately manipulating some of these rule sets at subcellular levels. This exercise readily eliminated some hypotheses and identified a sequence of spatial-temporal actions of TGF-beta 1 for normal successful wound healing in an easy-to-follow 3D model. We suggest this multiscale model offers a valuable, easy-to-visualize aid to our understanding of the actions of this key regulator in wound healing, and provides a model that can now be used to explore pathologies of wound healing

Development of a Three Dimensional Multiscale Computational Model of the Human Epidermis

Transforming Growth Factor (TGF-β1) is a member of the TGF-beta superfamily ligand-receptor network. and plays a crucial role in tissue regeneration. The extensive in vitro and in vivo experimental literature describing its actions nevertheless describe an apparent paradox in that during re-epithelialisation it acts as proliferation inhibitor for keratinocytes. The majority of biological models focus on certain aspects of TGF-β1 behaviour and no one model provides a comprehensive story of this regulatory factor's action. Accordingly our aim was to develop a computational model to act as a complementary approach to improve our understanding of TGF-β1. In our previous study, an agent-based model of keratinocyte colony formation in 2D culture was developed. In this study this model was extensively developed into a three dimensional multiscale model of the human epidermis which is comprised of three interacting and integrated layers: (1) an agent-based model which captures the biological rules governing the cells in the human epidermis at the cellular level and includes the rules for injury induced emergent behaviours, (2) a COmplex PAthway SImulator (COPASI) model which simulates the expression and signalling of TGF-β1 at the sub-cellular level and (3) a mechanical layer embodied by a numerical physical solver responsible for resolving the forces exerted between cells at the multi-cellular level. The integrated model was initially validated by using it to grow a piece of virtual epidermis in 3D and comparing the in virtuo simulations of keratinocyte behaviour and of TGF-β1 signalling with the extensive research literature describing this key regulatory protein. This research reinforces the idea that computational modelling can be an effective additional tool to aid our understanding of complex systems. In the accompanying paper the model is used to explore hypotheses of the functions of TGF-β1 at the cellular and subcellular level on different keratinocyte populations during epidermal wound healing

The role of tenascin-C in tissue injury and tumorigenesis

The extracellular matrix molecule tenascin-C is highly expressed during embryonic development, tissue repair and in pathological situations such as chronic inflammation and cancer. Tenascin-C interacts with several other extracellular matrix molecules and cell-surface receptors, thus affecting tissue architecture, tissue resilience and cell responses. Tenascin-C modulates cell migration, proliferation and cellular signaling through induction of pro-inflammatory cytokines and oncogenic signaling molecules amongst other mechanisms. Given the causal role of inflammation in cancer progression, common mechanisms might be controlled by tenascin-C during both events. Drugs targeting the expression or function of tenascin-C or the tenascin-C protein itself are currently being developed and some drugs have already reached advanced clinical trials. This generates hope that increased knowledge about tenascin-C will further improve management of diseases with high tenascin-C expression such as chronic inflammation, heart failure, artheriosclerosis and cancer