Agent based modelling helps in understanding the rules by which fibroblasts support keratinocyte colony formation

Background: Autologous keratincoytes are routinely expanded using irradiated mouse fibroblasts and bovine serum for clinical use. With growing concerns about the safety of these xenobiotic materials, it is desirable to culture keratinocytes in media without animal derived products. An improved understanding of epithelial/mesenchymal interactions could assist in this. Methodology/Principal Findings: A keratincyte/fibroblast o-culture model was developed by extending an agent-based keratinocyte colony formation model to include the response of keratinocytes to both fibroblasts and serum. The model was validated by comparison of the in virtuo and in vitro multicellular behaviour of keratinocytes and fibroblasts in single and co-culture in Greens medium. To test the robustness of the model, several properties of the fibroblasts were changed to investigate their influence on the multicellular morphogenesis of keratinocyes and fibroblasts. The model was then used to generate hypotheses to explore the interactions of both proliferative and growth arrested fibroblasts with keratinocytes. The key predictions arising from the model which were confirmed by in vitro experiments were that 1) the ratio of fibroblasts to keratinocytes would critically influence keratinocyte colony expansion, 2) this ratio needed to be optimum at the beginning of the co-culture, 3) proliferative fibroblasts would be more effective than irradiated cells in expanding keratinocytes and 4) in the presence of an adequate number of fibroblasts, keratinocyte expansion would be independent of serum. Conclusions: A closely associated computational and biological approach is a powerful tool for understanding complex biological systems such as the interactions between keratinocytes and fibroblasts. The key outcome of this study is the finding that the early addition of a critical ratio of proliferative fibroblasts can give rapid keratinocyte expansion without the use of irradiated mouse fibroblasts and bovine serum

The dynamics of neural fields on bounded domains: an interface approach for Dirichlet boundary conditions

Continuum neural field equations model the large scale spatio-temporal dynamics of interacting neurons on a cortical surface. They have been extensively studied, both analytically and numerically, on bounded as well as unbounded domains. Neural field models do not require the specification of boundary conditions. Relatively little attention has been paid to the imposition of neural activity on the boundary, or to its role in inducing patterned states. Here we redress this imbalance by studying neural field models of Amari type (posed on one- and two-dimensional bounded domains) with Dirichlet boundary conditions. The Amari model has a Heaviside nonlinearity that allows for a description of localised solutions of the neural field with an interface dynamics. We show how to generalise this reduced but exact description by deriving a normal velocity rule for an interface that encapsulates boundary effects. The linear stability analysis of localised states in the interface dynamics is used to understand how spatially extended patterns may develop in the absence and presence of boundary conditions. Theoretical results for pattern formation are shown to be in excellent agreement with simulations of the full neural field model. Furthermore, a numerical scheme for the interface dynamics is introduced and used to probe the way in which a Dirichlet boundary condition can limit the growth of labyrinthine structures

Co-culture of primary rat hepatocytes with rat liver epithelial cells enhances interleukin-6-induced acute-phase protein response

Three different primary rat hepatocyte culture methods were compared for their ability to allow the secretion of fibrinogen and albumin under basal and IL-6-stimulated conditions. These culture methods comprised the co-culture of hepatocytes with rat liver epithelial cells (CC-RLEC), a collagen type I sandwich culture (SW) and a conventional primary hepatocyte monolayer culture (ML). Basal albumin secretion was most stable over time in SW. Fibrinogen secretion was induced by IL-6 in all cell culture models. Compared with ML, CC-RLEC showed an almost three-fold higher fibrinogen secretion under both control and IL-6-stimulated conditions. Induction of fibrinogen release by IL-6 was lowest in SW. Albumin secretion was decreased after IL-6 stimulation in both ML and CC-RLEC. Thus, cells growing under the various primary hepatocyte cell culture techniques react differently to IL-6 stimulation with regard to acute-phase protein secretion. CC-RLEC is the preferred method for studying cytokine-mediated induction of acute-phase proteins, because of the pronounced stimulation of fibrinogen secretion upon IL-6 exposure under these conditions

Crystal structure of tarocystatin–papain complex: implications for the inhibition property of group-2 phytocystatins

Tarocystatin (CeCPI) from taro (Colocasia esculenta cv. Kaohsiung no. 1), a group-2 phytocystatin, shares a conserved N-terminal cystatin domain (NtD) with other phytocystatins but contains a C-terminal cystatin-like extension (CtE). The structure of the tarocystatin–papain complex and the domain interaction between NtD and CtE in tarocystatin have not been determined. We resolved the crystal structure of the phytocystatin–papain complex at resolution 2.03 Å. Surprisingly, the structure of the NtD–papain complex in a stoichiometry of 1:1 could be built, with no CtE observed. Only two remnant residues of CtE could be built in the structure of the CtE–papain complex. Therefore, CtE is easily digested by papain. To further characterize the interaction between NtD and CtE, three segments of tarocystatin, including the full-length (FL), NtD and CtE, were used to analyze the domain–domain interaction and the inhibition ability. The results from glutaraldehyde cross-linking and yeast two-hybrid assay indicated the existence of an intrinsic flexibility in the region linking NtD and CtE for most tarocystatin molecules. In the inhibition activity assay, the glutathione-S-transferase (GST)-fused FL showed the highest inhibition ability without residual peptidase activity, and GST-NtD and FL showed almost the same inhibition ability, which was higher than with NtD alone. On the basis of the structures, the linker flexibility and inhibition activity of tarocystatins, we propose that the overhangs from the cystatin domain may enhance the inhibition ability of the cystatin domain against papain

From gut dysbiosis to altered brain function and mental illness: mechanisms and pathways

The human body hosts an enormous abundance and diversity of microbes, which perform a range of essential and beneficial functions. Our appreciation of the importance of these microbial communities to many aspects of human physiology has grown dramatically in recent years. We know, for example, that animals raised in a germ-free environment exhibit substantially altered immune and metabolic function, while the disruption of commensal microbiota in humans is associated with the development of a growing number of diseases. Evidence is now emerging that, through interactions with the gut-brain axis, the bidirectional communication system between the central nervous system and the gastrointestinal tract, the gut microbiome can also influence neural development, cognition and behaviour, with recent evidence that changes in behaviour alter gut microbiota composition, while modifications of the microbiome can induce depressive-like behaviours. Although an association between enteropathy and certain psychiatric conditions has long been recognized, it now appears that gut microbes represent direct mediators of psychopathology. Here, we examine roles of gut microbiome in shaping brain development and neurological function, and the mechanisms by which it can contribute to mental illness. Further, we discuss how the insight provided by this new and exciting field of research can inform care and provide a basis for the design of novel, microbiota-targeted, therapies.GB Rogers, DJ Keating, RL Young, M-L Wong, J Licinio, and S Wesseling