Allometric scaling in-vitro

About two decades ago, West and coworkers established a model which predicts that metabolic rate follows a three quarter power relationship with the mass of an organism, based on the premise that tissues are supplied nutrients through a fractal distribution network. Quarter power scaling is widely considered a universal law of biology and it is generally accepted that were in-vitro cultures to obey allometric metabolic scaling, they would have more predictive potential and could, for instance, provide a viable substitute for animals in research. This paper outlines a theoretical and computational framework for establishing quarter power scaling in three-dimensional spherical constructs in-vitro, starting where fractal distribution ends. Allometric scaling in non-vascular spherical tissue constructs was assessed using models of Michaelis Menten oxygen consumption and diffusion. The models demonstrate that physiological scaling is maintained when about 5 to 60% of the construct is exposed to oxygen concentrations less than the Michaelis Menten constant, with a significant concentration gradient in the sphere. The results have important implications for the design of downscaled in-vitro systems with physiological relevance

Experimental and Computational Methods for the Study of Cerebral Organoids: A Review

Cerebral (or brain) organoids derived from human cells have enormous potential as physiologically relevant downscaled in vitro models of the human brain. In fact, these stem cell-derived neural aggregates resemble the three-dimensional (3D) cytoarchitectural arrangement of the brain overcoming not only the unrealistic somatic flatness but also the planar neuritic outgrowth of the two-dimensional (2D) in vitro cultures. Despite the growing use of cerebral organoids in scientific research, a more critical evaluation of their reliability and reproducibility in terms of cellular diversity, mature traits, and neuronal dynamics is still required. Specifically, a quantitative framework for generating and investigating these in vitro models of the human brain is lacking. To this end, the aim of this review is to inspire new computational and technology driven ideas for methodological improvements and novel applications of brain organoids. After an overview of the organoid generation protocols described in the literature, we review the computational models employed to assess their formation, organization and resource uptake. The experimental approaches currently provided to structurally and functionally characterize brain organoid networks for studying single neuron morphology and their connections at cellular and sub-cellular resolution are also discussed. Well-established techniques based on current/voltage clamp, optogenetics, calcium imaging, and Micro-Electrode Arrays (MEAs) are proposed for monitoring intra- and extra-cellular responses underlying neuronal dynamics and functional connections. Finally, we consider critical aspects of the established procedures and the physiological limitations of these models, suggesting how a complement of engineering tools could improve the current approaches and their applications

the italian centro 3r mission and early achievements presented during the second annual meeting

Not available

Allometric Scaling and Cell Ratios in Multi-Organ in vitro Models of Human Metabolism

Intelligent in vitro models able to recapitulate the physiological interactions between tissues in the body have enormous potential as they enable detailed studies on specific two-way or higher order tissue communication. These models are the first step toward building an integrated picture of systemic metabolism and signaling in physiological or pathological conditions. However, the rational design of in vitro models of cell–cell or cell–tissue interaction is difficult as quite often cell culture experiments are driven by the device used, rather than by design considerations. Indeed, very little research has been carried out on in vitro models of metabolism connecting different cell or tissue types in a physiologically and metabolically relevant manner. Here, we analyze the physiological relationship between cells, cell metabolism, and exchange in the human body using allometric rules, downscaling them to an organ-on-a-plate device. In particular, in order to establish appropriate cell ratios in the system in a rational manner, two different allometric scaling models (cell number scaling model and metabolic and surface scaling model) are proposed and applied to a two compartment model of hepatic-vascular metabolic cross-talk. The theoretical scaling studies illustrate that the design and hence relevance of multi-organ models is principally determined by experimental constraints. Two experimentally feasible model configurations are then implemented in a multi-compartment organ-on-a-plate device. An analysis of the metabolic response of the two configurations demonstrates that their glucose and lipid balance is quite different, with only one of the two models recapitulating physiological-like homeostasis. In conclusion, not only do cross-talk and physical stimuli play an important role in in vitro models, but the numeric relationship between cells is also crucial to recreate in vitro interactions, which can be extrapolated to the in vivo reality

Experimental and Computational Methods for the Study of Cerebral Organoids: A Review

Cerebral (or brain) organoids derived from human cells have enormous potential as physiologically relevant downscaled in vitro models of the human brain. In fact, these stem cell-derived neural aggregates resemble the three-dimensional (3D) cytoarchitectural arrangement of the brain overcoming not only the unrealistic somatic flatness but also the planar neuritic outgrowth of the two-dimensional (2D) in vitro cultures. Despite the growing use of cerebral organoids in scientific research, a more critical evaluation of their reliability and reproducibility in terms of cellular diversity, mature traits, and neuronal dynamics is still required. Specifically, a quantitative framework for generating and investigating these in vitro models of the human brain is lacking. To this end, the aim of this review is to inspire new computational and technology driven ideas for methodological improvements and novel applications of brain organoids. After an overview of the organoid generation protocols described in the literature, we review the computational models employed to assess their formation, organization and resource uptake. The experimental approaches currently provided to structurally and functionally characterize brain organoid networks for studying single neuron morphology and their connections at cellular and sub-cellular resolution are also discussed. Well-established techniques based on current/voltage clamp, optogenetics, calcium imaging, and Micro-Electrode Arrays (MEAs) are proposed for monitoring intra- and extra-cellular responses underlying neuronal dynamics and functional connections. Finally, we consider critical aspects of the established procedures and the physiological limitations of these models, suggesting how a complement of engineering tools could improve the current approaches and their applications

Micro-mechanical viscoelastic properties of crosslinked hydrogels using the nano-epsilon dot method

Engineering materials that recapitulate pathophysiological mechanical properties of native tissues in vitro is of interest for the development of biomimetic organ models. To date, the majority of studies have focused on designing hydrogels for cell cultures which mimic native tissue stiffness or quasi-static elastic moduli through a variety of crosslinking strategies, while their viscoelastic (time-dependent) behavior has been largely ignored. To provide a more complete description of the biomechanical environment felt by cells, we focused on characterizing the micro-mechanical viscoelastic properties of crosslinked hydrogels at typical cell length scales. In particular, gelatin hydrogels crosslinked with different glutaraldehyde (GTA) concentrations were analyzed via nano-indentation tests using the nano-epsilon dot method. The experimental data were fitted to a Maxwell Standard Linear Solid model, showing that increasing GTA concentration results in increased instantaneous and equilibrium elastic moduli and in a higher characteristic relaxation time. Therefore, not only do gelatin hydrogels become stiffer with increasing crosslinker concentration (as reported in the literature), but there is also a concomitant change in their viscoelastic behavior towards a more elastic one. As the degree of crosslinking alters both the elastic and viscous behavior of hydrogels, caution should be taken when attributing cell response merely to substrate stiffness, as the two effects cannot be decoupled

Inauguration of the Centro 3R for the promotion of 3Rs principles in teaching and research

The first European interuniversity center dedicated to promoting 3Rs principles in teaching and research was inaugurated in Pisa, Italy on March 14, 2018. The Centro 3R1 was spearheaded by the Universities of Pisa and Genova. Membership is open to all Italian universities and agreements for twinning across Europe and other countries are being pursued

clarifying mid brain organoids application of the clarity protocol to unperfusable samples

The aim of this study was to apply a workflow, integrating delipidation methods and advanced 3D imaging techniques for mapping of the global neuronal organization of brain organoids. These are self-organizing constructs in vitro generated from human pluripotent stem cells encased in a Matrigel shell, which resemble downscaled structural and functional features of human brains. In particular, we focused on midbrain organoids, widely considered a promising tool for studying dopaminergic neuron degeneration in Parkinson's Disease. The evaluation of the microanatomical alterations at a patient-level will potentially guide future research of this neuropathy, providing meaningful human specific data in line with the European Directives and the 3Rs principles

Improving African healthcare through open source biomedical engineering

The lack of accessible quality healthcare is one of the biggest problems in Africa and other developing countries. This is not only due to the unavailability of resources, but also to the absence of a structured formative process for the design and management of healthcare facilities. Crucial to the effective and efficient exploitation of healthcare facilities and biomedical technology is the support of Biomedical engineers, who form the link between technology and medical practice. Indeed Biomedical engineers, together with nurses and doctors, form the pillars of healthcare systems in the developed world. In this paper, the Open Source for BioMedical Engineering (OS4BME) project and its kick off summer school are presented. The OS4BME project aims at developing a new generation of biomedical engineers, able to exploit emerging technologies generated by the recent "Makers" revolution. During the one week summer school, students from various sub-Saharan countries were introduced to these new design, development and sharing paradigms. Students worked together to identify new simple biomedical devices, which could help in daily clinical practice in their countries. A cheap and easy-to-use neonatal monitoring device was chosen as a Crowd design project. The OS4BME Baby Monitor was designed and assembled by the students during the one week summer school, demonstrating the creative potential of the new generation of biomedical engineers empowered with the paradigms of crowdsourcing and rapid prototyping

The Kahawa Declaration: a manifesto for the democratization of medical technology

Most medical technology is employed and accepted passively by patients and doctors who have little or no influence in its design or usability. Patients are not involved in the development of medical technology, which is undertaken behind closed doors and whose global impact is hindered by proprietary know-how and by costs. This has so far impeded equitable healthcare as most of the world does not have access to the technology or healthcare coverage. Understanding the relevance of international partnerships for achieving the Sustainable Development Goals, feeling specially committed to the promotion of the Goal on “Good Health and Well-Being”, and convinced about the role that open-source biomedical engineering approaches may play in the future of medical technology, we commit ourselves, through the Kahawa Declaration, to enlighten the transformation of the biomedical engineering field, towards the democratization of medical technology as a key for achieving universal equitable health care. This paper presents the content of the Kahawa Declaration, which was signed in Nairobi in December 2017