So let's go to the second part of this problem. You know how to get these embryonic stem cells. Now the goal is how to differentiate them into different tissue. And one way to see embryonic differentiation is to imagine these as a series of binary choices. And they don't have to be binary, but that's the easiest way to see these. And at every step of the way, the cells have to choose a path to become something at the expense of the possibility of becoming something else. So when you made it all the way here, at every corner, you made a decision. And you chose one out of all the possibilities that took you closer to this place, but by definition, further to any other place that you could have gone on the other side. So in this case, let's say the cell, it's been told be one of the three germ layers. And after that said, okay, I cannot be gut. I can be skin or brain, because they come from the same embryonic origin. Okay, after that I'm going to decide not to be skin, I'm going to decide, or I've been told, to be brain. So I don't have the ability to be skin anymore, but I have the ability to become every part of your brain or the spinal cord. And after that, they will have another decision that would say, out of all the central nervous system, you are going to be part of the spinal cord, okay? You lost the ability now to be endoderm, to be gut, you lost the ability to be skin, then you lost the ability to be brain. Now you only retain the ability to be spinal cord. Within the spinal cord, there are many neurons or many cell types. And then you are being told, we want particular cell in the spinal cord. So in the way of deciding or being told what to do, the cell became closer to what it's going to be at the expense of not being able to do other tissues. That is called a differentiation path commitment. You will hear these words over and over again. So you can simplify this as a cell that can be everything, transition into a series of steps, becoming more and more and more differentiated, becoming closer to the terminal product as development proceeds. The cell is being usually told what it's going to become. If we would know that for everything, that means we could make every tissue that we want, and that is the challenge. So these cells, during development, they do it all the time. And usually we like to draw them as a terminal fate, in this case, as a neuron. That it's the there and live forever. By the way, you got your neurons and now very few parts of the brain are regenerating neurons. Living is about killing the neurons, [LAUGH] in your bran, I guess. Some cells retain one ability, and this is the terminal fate has two properties. One, it's able to regenerate itself. And the other one, it's able to produce some differentiated cells. How do we call these type of cells? Stem cells, the pluripotent will be how many cells they can make. They can pluripotent and multipotent, but the stem cell is nothing but a cell that can regenerate itself and make something else. So that's the definition of a stem cell, it's functional. There is no universal stem cell marker. There's no universal stem cell thing. If you give me a cell, I cannot tell you if it's a stem cell or not until I see it doing these two things, which makes research difficult. Yeah, which makes isolating stem cells from organs particularly difficult. Yeah, so because a stem cell can do two things, make more copies of itself, and given the right cues, differentiate into at least one other cell type, there are two types of stem cells. There are stem cells that we were talking earlier, which is embryonic stem cells, which are these particular stage in development we freeze the lab, this type. But we are full of adult stem cells that are in charge of the regular turnover and regeneration of tissues. And some animals are great at that. Planaria, I think Zara talk about planaria first. You can chop a planaria any way you want, it has an amazing quality, the stem cells are fantastic, you get a whole animal. And that's called asexual reproduction. Asexual, all one word, sorry for my pronunciation, reproduction. That's a form of reproduction. Where you take an animal, it splits in two, and it generates two. So one hot area of research is understanding how these animals do it. Because maybe we can trick ourselves to do the same. So adult stem cells, they live in us, we have them now. And they are in charge of regenerating our skin, our blood cells, the hair of many of you, the guts all the time. But they're also dangerous in a way, because if you remember, a stem cell can do two things, it can make more of itself or differentiate. When the balance between these two goes wrong and it's skewed towards making more of itself, this sounds like what? >> Cancer. >> Cancer. >> It's called mutation [INAUDIBLE] >> A mutation is a change in the DNA sequence. Not all mutations cause cancer. Cancer can be caused by a mutation. The great, great majority of the mutation are lethal for the cell or don't do anything. There are some mutations that sadly break this balance and allow the cells to proliferate more, and that's not good. So we go back to these, we have adult stem cells. And adult stem cells, this is the only mention that I'm going to make to them. They're extremely important. We already see what they can be great at. They know how to do something. That's a great thing. They already come from school, they learn how to make a certain tissue. So stem cells in our skin know how to make more skin. So that's why skin transplant works. And that's why hair follicle transplant works. Because they know how to do hair. And so if you put them there, they keep doing hair. If you can extend those types of therapies for other tissues that will be phenomenal. So if you think about, going back now, linking the differentiation with the stem cell. You can think of a stem cell, of an embryonic stem cell, of something that sets at the base of this tree, of this decision making tree. So what used to happen in the embryo, and we touched upon that already, is well, now we have the stem cells that are at the base. So we know that the stem cells can do it. Our job is to find how we tell the stem cells how to do it. And that is why we are kind of struggling right now. So let me tell the source of inspiration of how to do that. And I'll show you, just for fun, also because I happen to, for a class, I did this experiment and I was blown away. So look at this, look at what will happen. So this is Spemann and Mangold, remember we talked about gender discrepancies. He won the Nobel prize, she did not. So these are two embryos. There are many cells in it, this is are a single cell. This is an eyebrow to get an idea of the scale. So the researcher is going to cut a piece of this embryo, and put it into that one. Nothing will be removed from this one. It will add one piece into the other one. So here is the donor piece if you want, this is the host of the embryo, and small incision. It's pluged there, and this will allow it to develop. What happened? Two heads. Yeah? So, what are the alternatives? What could happen here? Anybody has a clue or a, what could have been when we take a piece of one embryo, and we put it into another one without removing anything, and now this animal has two heads. So one way to think about these is, the piece you transfer from one embryo to the other one, was destined to be the head in the other embryo. Now, that would be a logical conclusion. So now this thing has two pieces that were already programmed to be head, so now it has two heads. That's not what's happening. No. If that would have been the case, the new head would be made with cells and material coming from the embryo that you took the piece, correct? What happens is this little piece of embryo when you put it into the other one, it's able to convince other cells that were there that we're not going to be the head, to become a head. So, that was fabulous because people now start thinking wait a minute, therefore there is a piece of the embryo that is able to tell other cells what to do. How would you track the cells where they come from? At the time, they used eggs that will produce frogs of two colors. >> [INAUDIBLE]. >> Now we can actually put them a little tag, a little genetic label, and they will emit light of a green color, for example, and we just follow the green color. Yeah? So, the marvelous discovery here was that there were these cells that were able to organize structures. There were these cells that were called organizers. And these organizers will send signals to the other cells to become something. Now this has to start ticking now. There are cells that send a signal, To other cells and force these cells to differentiate into a particular tissue. And for that, Spemann won the Nobel Prize. It was a race to identify what was the signal. This is it, the graft was sending some signals that were received from some cells, and those cells were forced to become head, anterior. So, people went bananas trying to figure out what were the signals, many approaches, one of them is finding mutations. Remember I told you there are these animals that's raised in the lab, this model organisms, to find mutations in genes that will cause animals to lose the head. And that's how many of these signals were identified, for animals that when crossed and when homozygous, two copies of that gene being delivered, no head. So basically, but this is an important point. The graft was able to send a signal for the host cells to become something. That's exactly what we want to do. We have embryonic stem cells that can become everything. That's you, you want to send a signal for the stem cells to become the organ that you want them to be. And that's the game, period. That's it, yeah? So if you learn all the signals, now extrapolate this into many decisions and may signals. If we learn all the signals required of each step for the cell in these binary decisions, we can provide these signals to the differentiated cells in the dish, and guide them through all this path, yeah? So I'll go quickly through this. So, one of the things we do, and the details are not important, if you think about the central nervous system, it comes from a tube. Some point in development it was just a simple tube. And you can see this tube has a series of coordinates. A one coordinate, this axis, which is the dorsal ventral, and one coordinate on this axis which is anterior, posterior. Or would hear me saying because that's a more precise definition, but it's anterior to posterior. And depending on where the cell is in this tube, it will produce different types of neurons. And because you as a taxpayer funded research for a very long time, we know a lot about the signals that control this pattern of the central nervous system. This neural tube, and the details are irrelevant, just get the concept please. The neural tube that looks like this at the beginning, well, a little bit later, the receives all these signals from the surrounding tissue. And these signals they are soluble, they are molecules that we can add to a liquid medium, where the cells grow. Are able to tell the cells in this tube where they are in both orientations in the anterior, posterior, and the ventral and dorsal. So one way to get the cells to become what you want, is to modulate in these signals at the right time and at the right concentration and with the right goodies. To navigate this two-dimensional space, and get the different cell neuronal types. So, a particular concentration, let's say of this molecule called retinoic acid and another molecule called. It's applied on embryonic stem cells, where embryonic stem cells receive in a timely and concentration and dose controlled manner of the signals, to guide them all the way to make motor neurons. The neurons that die in Lou Gehrig's disease patients or kids with SMA for example. And this is a picture generated in our lab, where this is a stem cell and a few days later, having all the goodies, now you get these neurons. They are the same cells that die in an ALS patient, so now we can study ALS in a dish. Now we can generate different type of motor neurons out of stem cells. And ALS is characterized Lou Gehrig's disease by progressive paralysis, and that is because these neurons that innervate the muscles die, yeah? And once you see an ALS patients or a Parkinsonian patients, or something like that The plane is on the ground, the cells have died, and they're dying, that's why the patient has a problem. It's very hard to reconstruct what happened during that time. So what we can do now, is to have these cells in the lab, and see. And in an analogy, we see the plane in the air, and we're going to study what went wrong. How it goes wrong, and how we can do to remediate that, so it's like a flight simulator. So if we find why they die and if we find out ways to keep them alive, then that will be great and then we'll try that. And one thing that is doing is I'm telling you that ALS patients get paralyzed and they cannot move. But how does Steve Hawking, the physicist, drive the wheelchair and write, does anybody know? >> His eyes. >> Eyes, because the few neurons that innervate the eyes and makes the eyes move don't die. Wouldn't it be fantastic to know why these neuron don't die? >> Mm-hm. >> Okay, that's what is doing, because now we can generate from stem cells changing these queues. When, with a genetic trick, we can make the neurons that die in adolescence and neurons that don't die in adolescence. So for the first time ever, now we have two planes, one plane that crashes, and the other plane that crashes later, or doesn't crash. So now we're studying the differences between these two planes and trying to understand what goes wrong in one, and why the other one is able to weather that much better than the other. Yeah, so we think that by using the stem cell as a model system, as a disease in a dish, it's called, we will learn a lot about the pathology. And it's a faster and cheaper way to try things that otherwise would be crazy to try either in patients or even in mouse models. So that's an important application of embryonic stem cell technology. It's not all about generating the organ to transplant, it's way more about understanding and modeling diseases, and then learn. So we'll speed up now, and go into another next part. So this is basically the protocol, you have an embryonic stem cell, you differentiate it into the affected cell type. You learn something about the deceased, what drugs could be useful for the deceased, and then you give the drug to the patient. Or the other way, you make the cells and then you implant them back, but is it possible for us to have embryonic stem cells now? Can I have a stem cell that is the genotype of that patient? No, and you've heard about IPS and so on, so one way to get around this trick is to change something about the idea of clinic. And this is something that we don't do anymore, but it will be important for you to get the concept. Sperm, egg, half of genetic material, they get together. So now I change the sperm, and instead of having the sperm, I have a cell from the adult body. This cell has the complete genetic material, I cannot mix it with one half, because that will be one and a half, that's not viable. So, as you saw the trick of the mitochondrial replacement therapies, we can play a similar trick. We can remove the nucleus, which is complete genetic material, we can remove half of this egg, combine, and now we have an egg with a complete genetic material. With one difference, this is not half and half, this is all coming from that guy. If we allow this to develop and have a blastocyst, now we can have two outcomes. A clone, if we allow this to develop, or generate a patient specific embryonic stem cells. Horrible idea, you can go to jail if you do this. It's egg donors by the millions to get somebody a clone, this is illegal, on top of that you go to jail. So the powerful reprogramming technology that you learned last time, produces the same effect. It's able to recreate this inner cell mass without the need of doing this cloning procedure. So reprogramming, take something back to an inner cell mass-like state. And now that, without the need of egg donors or anything, we can differentiate in the lab exactly in the same way or a similar way that we did with embryonic stem cells. Given the conditions that that reprogrammed cell line, as was saying, is genetically normal and so on, and so all the caveats you learned. Yeah, so now we can close the loop because now we can have a patient, reprogramming, and now we can do exactly what we learned for embryonic stem cells. And just an anecdote here, the first time that this loop was closed was exactly with motor neurons. The protocol you saw where an ALS patient got reprogrammed and they made motor neurons from an ALS patient. So the first time this loop was closed was making the cells you learn and you're going to see today. So the elephant in the room is how to generate these cells that we can use for something. And I tell you that the organs are made of multiple cells that are even more difficult to make. You also know about chimera, the chimera is a fictional animal, not so much anymore, that is composed by parts of different animals. And you saw one, you saw an embryo that received cells from a different embryo, and so that was a chimeric animal. Do not mix with a hybrid, a mule is a hybrid, yeah? Mule has half genetic material from a horse and half from a donkey. This is a chimera, it contains cells that you can trace back, as she was saying, genetically to two different individuals. So if you mix some cells, that's called a trace chimera, and you are used to seeing them, human cancer's been studying mouse. They take a few cells from a human cancer and they propagate them under the skin of a mouse, and they test drugs on them and so on. That's a mouse and human chimera, and nobody's horrified by that, there's a tumor of a human growing under the skin of a mouse and that's normal. Now let's twist that a little bit, you know that embryonic stem cells or IPS cells are equivalent to the inner cell muscle of the blastocyst. When it's well done, they are actually equivalent, they are interchangeable, if they are interchangeable, then we can do this. We can have a mouse blastocyst and we can add some mouse embryonic stem cells in there. And the mouse embryonic stem cells will be integrated there, and because they are of the similar thing, they will grow and develop a mouse. But also it will be possible to, instead of this mouse, embryonic stem cells that I dropped there and will make a mouse that contains, in this case they were blue. So most of the mouse will be white, like the embryo was before, and we'll have some cells that are blue. These embryonic stem cells could be from a rat. They are the same stage, they're equivalent. So now this mouse, these blue patches, instead of being mouse, they're going to be rat, and that's okay. Now we'll have rat skin in some places, but let's keep thinking about that. You will have some of the sperm or some of the eggs will be mouse, but some will be rat because the cells that are generating the gametes come from these blue cells. And if we keep pushing the parts of this brain of that mouse are going to be rat. So at certain if at particular behavior is controlled by three neurons, and these three neurons happened to be rat, for that particular behavior, this guy all though it looks like a mouse is going to behave like a rat. Now there's going to be a percentage of blue cells over white. If there is 1%, this is going to be a 1% chimera. If I keep adding blue cells, this mouse is going to become, blue, blue, blue, blue, blue, blue or rat, rat, rat, rat, rat, rat, rat, okay? Are we okay with that? Do we understand the concept? Replace the mouse with a pig and replace the stem cells with human stem cells. Everything I told you about the rat and the mouse it's translated into a pig that will be great. It will contain some organ tissue. It will contain also, Gametes generating from the pig cells, but gametes generated from the human cells. Moreover, it will contain parts of the brain that are human. If it's 1%, we can talk about it. But you have to consider what happens when that percentage goes increasing. And at some point, you're going to look at that pig and that pig is going to look back at you- >> [LAUGH] >> And you have to decide if there is something in there, And I think quote [LAUGH] that I used last time is, college or grill. >> [LAUGH] >> At some point, you go from butchering to murder. And we know both sides of the coin exist, because we have no problem butchering pigs for human consumption. It's a huge problem to kill a person. It's a huge problem to kill a person that contains a heart valve from a pig. That's still murder. But we already move away and that person that contains a heart valve from a pig, it's some percentage pig. And that's murder. So now let's increase that percentage. We know that at some point it's okay, because 100% pig is okay to kill, and that's called butchering. So these two things have to meet at some point. And at some point, we're going to need to decide that it's not enough. Is it a part of the brain, it's not a part of the brain? How do we talk? The answer is [LAUGH] we don't know, and that's what's come. So where is the technology coming? So the question is, if you grow an organ inside a pig, it will never be 100% human. Well, the proof of principle has been done. In the rat and mouse experiment, the host mouse had a mutation that couldn't grow a pancreas. When they put the rat cells, the pancreas was only made by the rat cells that were not mutant for that gene. You can think about engineering that pig that would never make a heart. When you sprinkle in your human stem cells, the stem cell, the heart will be made only from human. The other issues, then we can solve that part. How do we restrict the human cells so they don't contribute to the brain? How do we restrict the human cells so they don't contribute to the gametes? I could tell you, look, I'm infertile. I cannot produce sperm. I want to have a child. You're giving him a heart and making him survive. I want to have kids. Why can't I have sperm made like he has a heart being made from the pig? It becomes completely arbitrary on that case. By the same token, somebody can say look I'm Parkinsonian. I need the neurons that are in the brain. You are giving him a heart. Why can't you give me my neurons to survive? So it's a tough call, and we have to be ready to start addressing it, because there were already grants submitted to NIH to do this study. So to conclude on something that we might see is 99.9% of the time you're going to see this diagram that I show. And this is what we are worth. What some people are trying to introduce is the pig into this diagram. Either to study, to make it an organ and transplant it back or a very useful tool. Have a human organ inside of an animal model that we can study, we can treat. We can see what happens with the genetic mutation or this drug. >> Okay. >> [APPLAUSE] >> Let's give you a round of applause. >> [APPLAUSE]
