Towards More Predictive, Physiological and Animal-free In Vitro Models: Advances in Cell and Tissue Culture 2020 Conference Proceedings.

Experimental systems that faithfully replicate human physiology at cellular, tissue and organ level are crucial to the development of efficacious and safe therapies with high success rates and low cost. The development of such systems is challenging and requires skills, expertise and inputs from a diverse range of experts, such as biologists, physicists, engineers, clinicians and regulatory bodies. Kirkstall Limited, a biotechnology company based in York, UK, organised the annual conference, Advances in Cell and Tissue Culture (ACTC), which brought together people having a variety of expertise and interests, to present and discuss the latest developments in the field of cell and tissue culture and in vitro modelling. The conference has also been influential in engaging animal welfare organisations in the promotion of research, collaborative projects and funding opportunities. This report describes the proceedings of the latest ACTC conference, which was held virtually on 30th September and 1st October 2020, and included sessions on in vitro models in the following areas: advanced skin and respiratory models, neurological disease, cancer research, advanced models including 3-D, fluid flow and co-cultures, diabetes and other age-related disorders, and animal-free research. The roundtable session on the second day was very interactive and drew huge interest, with intriguing discussion taking place among all participants on the theme of replacement of animal models of disease.

Microdomain heterogeneity in 3D affects the mechanics of neonatal cardiac myocyte contraction.

Cardiac muscle cells are known to adapt to their physical surroundings, optimizing intracellular organization and contractile function for a given culture environment. A previously developed in vitro model system has shown that the inclusion of discrete microscale domains (or microrods) in three dimensions (3D) can alter long-term growth responses of neonatal ventricular myocytes. The aim of this work was to understand how cellular contact with such a domain affects various mechanical changes involved in cardiac muscle cell remodeling. Myocytes were maintained in 3D gels over 5 days in the presence or absence of 100-µm-long microrods, and the effect of this local heterogeneity on cell behavior was analyzed via several imaging techniques. Microrod abutment resulted in approximately twofold increases in the maximum displacement of spontaneously beating myocytes, as based on confocal microscopy scans of the gel xy-plane or the myocyte long axis. In addition, microrods caused significant increases in the proportion of aligned myofibrils (≤20° deviation from long axis) in fixed myocytes. Microrod-related differences in axial contraction could be abrogated by long-term interruption of certain signals of the RhoA-/Rho-associated kinase (ROCK) or protein kinase C (PKC) pathway. Furthermore, microrod-induced increases in myocyte size and protein content were prevented by ROCK inhibition. In all, the data suggest that microdomain heterogeneity in 3D appears to promote the development of axially aligned contractile machinery in muscle cells, an observation that may have relevance to a number of cardiac tissue engineering interventions.

Digital image correlation of bone sequential microscopic observations.

A method of image correlation is presented to study sequential microscopic observations of human Haversian cortical bone. Imaging biological tissues is sometimes challenging owing to their complex microstructures in particular when microcracks appear. Bone microfractures can be studied in micro compression tests where the progressive growth of small cracks is imaged by light microscopy. The two-dimensional displacement field on the sample surface is then tracked by various digital image correlation methods based on cross-correlation formulation. Because of the potential high number of sequential observations, the method calculates the displacements at given growth steps obtained either by direct comparison of the studied step and the undeformed initial state, called 'direct correlation', or by iterative comparisons of successive pairs of observations, called 'gradual correlation'. In the gradual procedure, two cases are studied, referred to as 'invariant gradual correlation' and 'varying gradual correlation', when the correlation domain is transferred till the last observation or reinitialised for each image pairs. As bone is highly heterogeneous, two types of correlation procedures are considered with or without domain partition (WDP or WODP) delimiting material and strong discontinuities. The precision of the methods is specifically evaluated for experimental observations.