Keeping cells alive
One of the big challenges with making and sustaining organs in the lab is plumbing- getting nutrients in, and waste out. This is the job of the vascular system, that carries blood around the body and is vital to keeping tissues alive. Mark Skylarr-Scott and his colleagues at the Wyss Institute for Biologically Inspired Engineering at Harvard University have recently come up with a new technique for 3D printing large, vascularized human organ building blocks. The technique lays down tracks for blood vessels between clumps of cells to help keep tissues nourished. Using this method, they’ve managed to keep clumps of heart cells alive for weeks, which would previously have been almost impossible. Mark took Katie Haylor through the process...
Mark - Step one would be prepare approximately one cup of stem cells. In a lab that involves using something called a bio-reactor to generate vast numbers of stem cells in three dimensions. We then need to instruct those cells to become heart cells. So we put the correct ingredients into this bio-reactor so that they are able to develop into a heart cell.
Mark - So stem cells, and then we provide chemicals that make the stem cells think that they're supposed to develop into beating heart cells. You know, this is a fairly standard process up to this point. And now if we have hundreds of millions of heart cells, a heart isn't a hundred million cells floating around in a bio-reactor. It's actually a solid organ that beats and you know, pumps out blood. We now need to think of a way to compile these together into a tissue.
Mark - So then step two, so we now have hundreds of millions of cells. At the moment, little pieces of tissue. So we actually, they're not single cells, they are in little clumps, about half a millimeter across. If we now take these little clumps and we put them in a centrifuge, we are able to sort of push them all together. We spin them down, they form, a little pellet of cells. Cool it on ice, so it's now at zero degrees.
Mark - We then come with a 3D printer and inject gelatin - solid at room temperature and liquid when it's 37 degrees - in three dimensions into this group of cellular aggregates. Now, if this material were liquid, the gelatin would just sink and I wouldn't be able to create a 3D structure. If the material, if my cells were too solid, I would essentially be carving it like a turkey as I come in with a nozzle and 3D printer and inject gelatin in 3D. And this would also break the tissue. But because these cells are halfway between a liquid and a solid, I'm actually able to come in and lay material, this gelatin material in three dimensions, and it will hold in place so that when I now raise the temperature, my cells all stick together. So now my tissue is become solid-like, and the gelatin that I printed melts and becomes liquid-like. If I now flush that gelatin out, I'm left with space. I'm left with channels that I can now connect a pump to those channels and actually keep the tissue alive, keep it perfused and viable.
Katie - So it's a bit like a Goldilocks porridge situation. And how does this compare to how a full scale heart would be vascularized?
Mark - This is very different in terms of the process of how we develop, but this is obviously because the goal of creating an organ for transplantation, you can't wait, you know, 20 years for an adult heart to develop, we need to be able to manufacture it quickly. So the process is very different.
Mark - In terms of the architecture, we similarly have blood vessels in our body and in our organ that start very large. The aorta comes from the heart and then it splits and it splits and you're down to what's called arterioles, little arteries. And then the arterioles become capillaries. And then the capillaries rejoin to form veins and then larger veins. So this hierarchical arrangement of blood vessels we're able to reproduce, with a 3D printer - not necessarily at the resolution of capillaries, but certainly in terms of having these branched hierarchical networks.
Mark - This is actually important for transplantation. If a surgeon wants to be able to connect tissue to the patient, they don't want to have a hundred different tubes that they need to suture to connect to be able to feed that tissue. They want a single inlet that will then split, you know, and feed the full volume of the tissue and then a single outlet that they can plumb into.
Mark - I'd say the advantage of our method here is these organoids that are being developed, they really leverage biology's natural ability to make complex structures on their own. The instructions for generating the sort of patterns that you see in organs, of course, it's in our DNA. Organoid protocols, they take advantage of that to form these tissues that can exhibit amazingly complicated architectures that self assemble. They develop on their own for free essentially. So because at the smallest scale we have all of these structures already in place in the organoids, if we can now compile hundreds of thousands of these organoids together into a larger tissue that we can keep alive, we hope that through 3D printing, we get the large scale structures and vasculature necessary to keep it alive, but through developmental biology and the fact that the stem cell derived organoids that have all the microstructure already present, that we get the small scale structure in place as well.