Production of a Human Growth Plate Organ-Chip Model of Skeletal Development
Lead Research Organisation:
Queen Mary University of London
Department Name: School of Engineering & Materials Scienc
Abstract
The growth plate is an area of tissue at the ends of bones in children which determines the development of the skeleton. It represents a transition between bone and cartilage and is supported by blood vessels. Due to the importance of the growth plate, there is considerable interest in studying this organ and its role in development, ageing, and disease. To do so necessitates robust, reproducible and physiologically relevant experimental 'model' systems. Currently many studies rely on animals, typically mice and rats. Since mechanical forces are known to influence skeletal development in the body, some of the animal models include techniques to control mechanical forces on the growth plate whilst the animal is still alive. These widely used animal models often have limited reproducibility and fail to represent key behaviour in humans as well as prompting ethical concerns. There is therefore an urgent need to replace some of these animal models with non-animal, human experimental models. This would improve scientific rigour and relevance to human physiology as well as reducing the use of animals in science.
Previously reported non-animal models of the growth plate fail to replicate the key features of the organ including a gradient of bone-cartilage tissue, mechanical loading and incorporation of blood vessels linking to the bone. Therefore, we will develop and validate a new human growth plate model using organ-on-a-chip technology. An organ-chip is an experimental model system in which human cells can be grown within interconnected channels through which fluid can be pumped providing the cells with the necessary nutrients to keep them alive. We will develop a growth plate organ-chip with the following key features:
- A composite tissue graded from bone to cartilage
- A channel replicating the blood vessels
- Mechanical loading of the developing bone-cartilage growth plate
- A reproducible, scalable system that can easily be adopted by other researchers
To achieve the above, we will use human adult stem cells expanded from bone marrow. These stem cells will be grown within a 3D gel material in one of the channels in an organ-chip. The cells will then be differentiated into bone and cartilage cells using natural growth factors to create the graded bone-cartilage growth plate tissue. We will also create a blood vessel channel lined with human endothelial cells that form blood vessel walls. This blood vessel channel will provide a conduit for nutrient and hormone delivery to the developing growth plate as occurs in the body. To ensure reproducibility and scalability, the organ-chip model will be created with commercially available human stem cell and endothelial cells and within the commercially available organ-chip provided by Emulate Inc. This chip consists of two channels separated by a semi-permeable membrane allowing interaction between the developing bone-cartilage growth plate tissue in one channel and the blood vessel channel. In addition, we will utilise the ability of the Emulate organ-chip to provide controlled mechanical loading to the channels replicating the mechanical environment within the body.
In this way, our vision is to create a highly reproducible and validated human vascularised growth plate organ-chip model which can be readily adopted by the scientific community reducing reliance on animal models and improving the quality of research into skeletal development in health and disease.
Previously reported non-animal models of the growth plate fail to replicate the key features of the organ including a gradient of bone-cartilage tissue, mechanical loading and incorporation of blood vessels linking to the bone. Therefore, we will develop and validate a new human growth plate model using organ-on-a-chip technology. An organ-chip is an experimental model system in which human cells can be grown within interconnected channels through which fluid can be pumped providing the cells with the necessary nutrients to keep them alive. We will develop a growth plate organ-chip with the following key features:
- A composite tissue graded from bone to cartilage
- A channel replicating the blood vessels
- Mechanical loading of the developing bone-cartilage growth plate
- A reproducible, scalable system that can easily be adopted by other researchers
To achieve the above, we will use human adult stem cells expanded from bone marrow. These stem cells will be grown within a 3D gel material in one of the channels in an organ-chip. The cells will then be differentiated into bone and cartilage cells using natural growth factors to create the graded bone-cartilage growth plate tissue. We will also create a blood vessel channel lined with human endothelial cells that form blood vessel walls. This blood vessel channel will provide a conduit for nutrient and hormone delivery to the developing growth plate as occurs in the body. To ensure reproducibility and scalability, the organ-chip model will be created with commercially available human stem cell and endothelial cells and within the commercially available organ-chip provided by Emulate Inc. This chip consists of two channels separated by a semi-permeable membrane allowing interaction between the developing bone-cartilage growth plate tissue in one channel and the blood vessel channel. In addition, we will utilise the ability of the Emulate organ-chip to provide controlled mechanical loading to the channels replicating the mechanical environment within the body.
In this way, our vision is to create a highly reproducible and validated human vascularised growth plate organ-chip model which can be readily adopted by the scientific community reducing reliance on animal models and improving the quality of research into skeletal development in health and disease.
Technical Summary
We will develop and validate a human growth plate organ-chip model with the following key characteristics:
- 3D spatial gradient of growth factors producing a cartilage-bone construct within an organ-chip
- Vascularisation of the growth plate via the developing bone to recapitulate in vivo behaviour
- Biomechanical loading to replicate the physiological growth plate maturation
- Reproducible and scalable proving access to researchers across academia and industry
A buoyancy-driven gradient in bone morphogenic protein BMP-2 will be created using gelatin methacryloyl hydrogel seeded with human mesenchymal stem cells and maintained within the Emulate organ-chip. Osteochondral differentiation media combined with the BMP-2 gradient will induce an osteochondral growth plate tissue with cartilage and bone (WP1). This will be supported by a bone vascular channel lined with human bone marrow endothelial cells providing a conduit for nutrient exchange and hormone regulation (WP2). To ensure reproducibility and scalability, the model will be created with commercially available human stem cells and endothelial cell lines (Lonza and ATCC) within the blank S1 organ-chip provided by Emulate Inc. This chip consists of two channels separated by a semi-permeable membrane allowing interaction between the developing osteochondral tissue and the vascular channel. We will provide mechanically stimulation to the 3D osteochondral tissue by way of the vacuum channels which causes deformation of the microfluidic channels (WP3).
We will optimise the protocols and characterise the model in terms of biophysical properties and spatio-temporal protein expression using CellDIVE imaging and histology. We will validate against existing in vivo data and by determining the response to oestrogen which regulates growth plate closure in humans (WP4). Full protocols and characterisation will be published with Emulate supporting the roll out of this novel and transformative non-animal technology.
- 3D spatial gradient of growth factors producing a cartilage-bone construct within an organ-chip
- Vascularisation of the growth plate via the developing bone to recapitulate in vivo behaviour
- Biomechanical loading to replicate the physiological growth plate maturation
- Reproducible and scalable proving access to researchers across academia and industry
A buoyancy-driven gradient in bone morphogenic protein BMP-2 will be created using gelatin methacryloyl hydrogel seeded with human mesenchymal stem cells and maintained within the Emulate organ-chip. Osteochondral differentiation media combined with the BMP-2 gradient will induce an osteochondral growth plate tissue with cartilage and bone (WP1). This will be supported by a bone vascular channel lined with human bone marrow endothelial cells providing a conduit for nutrient exchange and hormone regulation (WP2). To ensure reproducibility and scalability, the model will be created with commercially available human stem cells and endothelial cell lines (Lonza and ATCC) within the blank S1 organ-chip provided by Emulate Inc. This chip consists of two channels separated by a semi-permeable membrane allowing interaction between the developing osteochondral tissue and the vascular channel. We will provide mechanically stimulation to the 3D osteochondral tissue by way of the vacuum channels which causes deformation of the microfluidic channels (WP3).
We will optimise the protocols and characterise the model in terms of biophysical properties and spatio-temporal protein expression using CellDIVE imaging and histology. We will validate against existing in vivo data and by determining the response to oestrogen which regulates growth plate closure in humans (WP4). Full protocols and characterisation will be published with Emulate supporting the roll out of this novel and transformative non-animal technology.
