Over the past decade, scientists studying the development and functioning of the human body at the most fundamental level have experienced something of a renaissance, thanks to structures called organoids — tiny 3D models of organs developed from pluripotent stem cells growing in Petri dishes grow.
Organoids are derived from human pluripotent stem cells, which can be introduced into any cell type in the human body and have become an important research tool for understanding human development and disease. They have allowed scientists to move away from the simple two-dimensional growth of cells in culture and have provided important insights into the complex three-dimensional shape and function of various organs such as the lungs, brain and heart. However, growing tiny organs in a dish is a tricky process.
A University of Michigan Medical School lab led by Jason Spence, Ph.D., of the Department of Internal Medicine, has developed a new, significantly simpler method of culturing a 3D gut model that results in increased complexity and organization . The Advance, published in cell reports, describes how intestinal organoids now contain cells that make up the serous mesothelium, the protective, outermost layer of the gut. This layer is also found lining many other organ systems and is critical in producing a non-stick surface that allows for relatively friction-free movement of organs within the abdominal cavity.
Previous research into growing different types of mini-organs relied on a supportive gel called Matrigel, which forms a 3D scaffold that allowed different cell types to develop into an organoid.
“Matrigel is the gold standard for organoid culture, but it has limitations,” explained Meghan Capeling, Ph.D. Candidate in Spence’s lab and head of new research. For one, Matrigel is very expensive at around $200 for 5ml of product. Second, it’s derived from mouse tumor cells, “so if you’re considering downstream clinical applications, it wouldn’t work well because it contains unknown biological components,” Capeling said.
In a previous 2018 publication, Capeling and her colleagues found that gut organoids could be grown in a simpler, biologically inert alginate gel because they make their own supportive mesenchymal cells — cells that transform into connective and smooth tissue in the developing fetus muscle cells.
This finding led the team to question whether the cells even needed a 3D growth environment. The answer, they determined, is no. In the new work, they describe their successful generation of the human gut organoid in a simple suspension culture.
“It actually looks a bit like bubble tea,” Capeling said, “it’s just a regular tissue culture plate that’s filled with growth media.” (Growth medium is a liquid containing life-sustaining chemicals and nutrients for the growth of cells.) They compared the suspension organoids to real human tissue, as well as to organoids formed with Matrigel and alginate, and found that they looked similar at the molecular level. In fact, the suspension organoids more closely resembled actual human tissue.
The team’s next step was to see if these floating mini-guts could actually function like a developing human gut, and they attempted to use the organoids to understand how the serosa layer forms, noting that over Little to nothing is known about this process in the human development context. The team studied the chemical cues that lead to the formation of the serosa in suspension cultures, Capeling says.
“This is one of the first studies to give an idea of the specific regulators that might play a role in the proper development of gut serosa.”
Given that abnormal development of the serosa can lead to congenital defects, the team hoped to use the organoids to uncover how this layer of tissue normally forms. Using drugs that block the activity of specific proteins, Capeling and the team identified two signaling pathways, termed Wnt and Hedgehog, that are essential for the normal formation of the serosa. Although the suspension method resulted in fewer organoids overall, the method could be a game changer for researchers using human pluripotent stem cells. The team hopes that suspension culture will open up the possibility of larger organoid experiments and will be an improved system for studying human development and disease.
Additional authors include Sha Huang, Charlie Childs, Joshua H. Wu, Yu-Hwai Tsai, Angeline Wu, Neil Garg, Emily M. Holloway, Nambirajan Sundaram, Carine Bouffi, Michael Helmrath, and Jason R. Spence.