Back in 2005, Drs. Irina and Michael Conboy showed that joining the circulatory systems of young and old mice together in a procedure called parabiosis could rejuvenate aged tissues and reverse some aspects of aging in old mice.
Following this discovery, many researchers concluded that there must be something special in young blood that was able to spur rejuvenation in aged animals, and various companies have been trying to find out what. Indeed, we recently reported that researchers were apparently successful in halving the epigenetic age of old rats by treating them with Elixir, a proprietary mix of pro-youthful factors normally found in young blood.
However, a question still remains: was the rejuvenation the result of there being something beneficial in the young blood, or is it more a case of dilution of the harmful factors present in old blood?
Today, we want to spotlight a new study by Drs. Irina and Michael Conboy, which again lends more weight to the idea that the rejuvenation is most likely due to a dilution of pro-aging factors in old blood rather than there being any special sauce in young blood [1].
During the study, the research team discovered that by replacing half of the blood plasma in old mice with a saline and albumin mixture, the albumin replacing the lost protein that was removed when the original old blood plasma was taken, they could achieve a similar or even greater rejuvenation effect in brain, liver, and muscle tissues as joining two mice together through parabiosis or giving old mice young blood.
We had the opportunity to interview Drs. Irina and Michael Conboy about this new discovery and to see if we could get to the bottom of the mystery surrounding aged blood rejuvenation.
Steve: This recent paper builds on the 2015 paper of TGF beta, but it goes even further back to the days when you guys had a lab next door to Amy Wagers and Tony Wyss-Coray and you all shared the techniques, including the parabiosis technique.
Irina: Yes. Actually, I would like also to thank you, Elena, and the whole organization for highlighting our work and giving us an opportunity to speak in interviews.
Steve: You are very welcome. So, is this dilution? Is it what you put in that’s more important, is it what you take out, or is it both? I personally think that the evidence strongly suggests that it’s more what you take out, but that doesn’t necessarily mean that there isn’t good stuff in young blood.
Irina: Since our 2005 heterochronic parabiosis paper, many people jumped into this boat of young blood, thinking that the reason for rejuvenation is that there are less young factors in an old animal and we provided them. Meanwhile, all our work even leading to that paper suggested the opposite outcome: that there are excessive factors in old blood that are actually good proteins; for example, TGF beta. You cannot live without TGF beta. But, when people age, the levels of this protein become elevated, and they start doing counterproductive things for tissue repair, induce inflammation, increase fibrosis, and prevent proliferation of tissue stem cells. That was our point of view for the past 15 years, and every single paper that we published since was putting forward the general idea that it is not the young blood, it is the old blood that needs thought and attention.
Eventually, we decided to set up an unambiguous experimental system in which we can resolve the lack of young versus excess of old blood factors paradigm. As you mentioned, there might be a combination of things: something declines with age, and something else increases with age, but it does not mean that rejuvenation can operate by adding age-diminished molecules to old mammals with their age-elevated dominant inhibitors of tissue health and repair. Once we developed a small-circuit blood exchange, which is a well-controlled system, it’s not like parabiosis, now we were able to answer this question directly: If you dilute old blood, will there be rejuvenation? If you dilute young blood, will there be aging? Because if you hypothesize that, with age, you have diminishing positive factors in blood, if you dilute young blood, you will make mammals older because you have diminished their young factors. That was the premise of the paper, and we did not know what to expect.
We started out with a clean slate; let’s see what happens. What happened is that it seems that rejuvenation in all these previous experiments, such as parabiosis, exchanging old animals with young blood, and so forth, all of this works through blood dilution, through normalizing age-elevated circulating proteins. If you dilute the blood of young animals, they do not become older. It is not really that you have some decline with aging, it is that you have excess with aging. That was the major fork in the road where most people went in the direction of adding young blood fractions or factors (with lackluster clinical success). We went in the opposite direction from the same fork in the road. It is paradigm shifting, because if you cannot rejuvenate old mammals by young blood or fractions or proteins, then it’s better to stop doing that and move into something more productive that has a direct route to clinical translation, because therapeutic plasma exchange is already FDA approved.
Steve: Tony Wyss-Coray’s lab at Alkahest is about to do a human trial for a blood scrubber. It’s a medical device that goes in-line with dialysis, and it’s designed to remove B2M from the blood. That’s the one that Saul Villeda identified a while ago. Are they heading in the right direction?
Irina: They published a Nature Medicine paper saying that infusion of small volumes of young plasma into old mice is rejuvenative, and then they spent millions trying to apply this to people in three years of clinical trials, which unfortunately did not show much improvement in AD, if I’m not mistaken. Removing age-elevated molecules, might be a similar thing that therapeutic plasma exchange accomplishes in an already FDA-approved way. There was, in fact, a recent promising clinical trial with TPE that slowed progression of AD, and Dr. Dobri Kiprov was the clinical director of that trial. Now, with respect to specific molecules, Beta-2 microglobulin is an invariant chain of MHC class 1, which is present on most cells in your body and presents viral antigens, so, if a cell is infected by a virus, the immune system can recognize this. It is not really a conventional systemic factor; and the levels of B2M are a reflection of inflammation. If you have chronic inflammation, tissue will have more of this molecule on the surface of more cells. Some of that will be shed into circulation. So, a high B2M level is a marker for inflammation. Interestingly, we did not find that B2M increases with age in the bloodstream, but it becomes more prevalent locally in some parts of old, inflamed tissues; we published on this in our 2016 heterochronic blood exchange paper in Nature Communications.
Steve: It’s at least partly regulated by the NF-kB complex, which is part of inflammation.
Irina: Well if you think what this molecule is, B2M is a very well-known protein that is present on cell surfaces in combination with other proteins and has a defined function of presenting cells’ own peptides. In some diseases where inflammation is predominant, you might have so much MHC that the part that sheds into the bloodstream is now elevated. Will you reduce inflammation if you scrub that molecule from the bloodstream? Not likely. Inflammation will persist locally in tissues through elevated gene expression for this molecule and many other proinflammatory proteins, being regulated by NF-kB, TGF beta, interferon gamma, IL-6, etc. So, each cell will still produce more and more MHC if there is an inflammatory environment.
On top of that, the reason that we decided to publish our paper is that if you can reposition an already FDA-approved procedure to treat the disease, that repositioning will be much faster and less costly than the development of anything new. Therapeutic plasma exchange can accomplish the same thing as specific targeting of systemic Beta-2 microglobulin, plus, as we publish, it can simultaneously normalize a number of age-elevated products, not just B2M. Commercialization of TPE as a repositioned therapy will be much faster and more effective, as compared to additional intricate specific devices or instruments. We are starting a company, which is called IMYu, focusing on the application of therapeutic plasma exchange and defined pharmacology for preventing, attenuating and reversing diseases of later years; applying designer approaches for combating a number of inflammatory fibrotic, degenerative and metabolic pathologies.
Steve: I was wondering whether restoring blood to a younger state would help reduce or prevent fibrotic tissue from forming. It seems it might?
Irina: We saw that it does. In the muscle, for example, there is less fibrosis, and we showed in the liver there was less fibrosis. So we showed therapeutic effects on fibrotic diseases.
Steve: With GDF-11. We still really don’t know what’s going on with that, even now.
Irina: GDF-11 is a member of the TGF beta family, and many members of that family promote blood vessel formation. So through that property, which never was disputed and was discovered before, there should be some improvement and tissue repair, but on the minus side, you also will promote growth of any tumor or precancerous mass. And GDF-11 has a very strong association with a number of human cancers, for example, colon cancer. GDF-11 typically is so minute that you cannot even find it in plasma. If you had a lot of it, you would promote blood vessel formation, but it also might promote cancers; and GDF-11 is not unique; you can use VEGF, which also improves tissue repair and promotes cancers as well.
Steve: The one big question that people usually ask is, oh, okay, about muscle, bone and other tissues, but they always say, but what about the brain? You’ve got the blood-brain barrier, and the brain is weird, it doesn’t work like the rest of the body. What was the evidence for rejuvenation in the brain?
In our paper, we looked at the hippocampal neurogenesis, which is the formation of new neurons in the area of the brain that is responsible for learning and memory. We saw an eightfold increase in all the animals after neutral blood exchange, which is much higher than anything else that was observed either by heterochronic parabiosis or reported by young blood infusion. In old mice transfused with 50% young blood, we did not see an improvement in neurogenesis, which, again, puts a question mark on potency of young blood in old mammals. Does young blood by itself work as a medicine or not? If you dilute old blood by 50%, in neutral age blood exchange – with albumin-supplemented saline, there is quickly better neurogenesis. In our work in progress, we also see a reduction of neuro-inflammation by neutral blood exchange and improvement in animal cognition. We decided to have another dedicated study on those two parameters. That work is collaborative with professor Yi Zuo from UC Santa Cruz.
Steve: There’s a population of specialized stem cells in the hypothalamus, and some researchers think that, as they die, their ability to regulate hormones fails. Is that possibly what starts the cascade off?
Irina: I don’t think so. There are so many things that can start the cascade. For example, thymic involution, after which our immune system is not the same. Then, there’s a physical decline in the numbers of memory T cells and B cells because they have no telomerase, so once their telomeres shorten, they die. Overall damage to tissues everywhere and an increase of senescent cells, there are so many different directions. I don’t think that the only one is hypothalamus.
Steve: Changes to the microbiome as well.
Irina: Exactly, and that’s why I don’t think a silver bullet will be able to resolve diseases of aging. You know, therapeutic plasma exchange or neutral blood exchange are excellent in this regard because they molecularly recalibrate numerous factors simultaneously. They restore the health of numerous organs because multiple age-elevated determinants change to younger levels.
Steve: It’s a relatively low-hanging fruit by the look of it. Some people have asked if that technology would be easily scalable or cost-effective for a global audience. Wouldn’t it be, because it’s already approved?
Irina: It’s already approved. Dr. Kiprov was doing it in that clinic for 25 years. Anecdotally, people were telling him that they had more energy after this procedure was done. They feel younger, they feel better, and they do not succumb to viral illnesses that much. So, again, it’s just anecdotal evidence, but it certainly is worthy of attention in the current pandemic situation. People who underwent repetitive TPE did not get sick with the flu. This was back in 2017, 2018 before COVID, and they were from the age group 65 to 70, and 60% of flu hospitalizations were in this age group. None of the people who were in Dr. Kiprov’s care, even though they did not get the flu shot, got sick. So, better immunity is predicted as well.
Steve: They do transfusions a lot as well; there’s anecdotal evidence that you hear from time to time about people giving blood, and it makes them feel better as well.
Irina: Absolutely, yeah. I noticed in some recent publications when authors want to prove that young blood works, they have to just pump old animals with huge volumes of young blood. In actuality, de facto diluting the old blood. Even then, the data is highly variable and not as strong as if you take out some factors and replace them with a saline supplemented with albumin.
Steve: So we’re not saying it’s dilution, but it’s dilution.
Irina: Exactly.
Steve: That all feeds into the idea of inflammaging, as they call it, effectively all of the sources of inflammation that are creating this inflammaging are a major reason why we age. And if you remove that inflammation, things start working again, like NAD+, for example, that’s actively destroyed by CD38, which is present in the SASP. So, if you remove it now, levels go back up, and that’s why people may feel that they’re more energetic.
Irina: Another point is that the so-called young factors, of course, don’t disappear from our bodies. Where did they go? They’re still there. They’re suppressed by the age-elevated proteins. Once you remove that suppression, now, all of a sudden, you also have all of the young, systemic tissue determinants. They were not expressed appropriately but now they are, so you don’t need to add them back.
Steve: A lot of people are downstream of the problem. NAD levels drop, we’ll try and restore it with nicotinamide riboside, NMN, and other things like that. They’re all trying to compensate, and it does seem to have benefits, but obviously, it’s not a great long-term fix.
Irina: This is somewhat metaphorically similar to the application of insulin. You can not just keep exchanging blood and have good effects. In the same way, if somebody is diabetic, they need insulin to live, but if you keep injecting insulin, they’re going to die. This is a somewhat invasive procedure where your whole blood goes through a machine and the cells are returned to you and the plasma is diluted. It is not completely benign, and you cannot just keep doing it every day. So, to know how long the effects last and to know it for each individual is an additional know-how part that we found out from a couple of years of research and are studying further. I’m sure there will be competition, but I just want to warn everybody, do not go somewhere and get your blood replaced with saline. Wait until there are reputable clinical trials and people know a little bit more.
Steve: So, no popping over the border for a quick oil change, not advised at this point, although I suppose the principle is similar to changing the oil in the car.
Michael: Or like a fish tank. You get the poor goldfish in murky water and then you change the water to clean it up, and that perks up the fish.
Irina: As a principle, it’s better and safer than infusions of blood or bodily fluids, and it is more effective than any single factor, but we still need to know how to apply the procedure in a way that is effective for you. Because we’re not aging in the same way. We all age slightly differently.
Steve: There was a study out this year that identified three distinct types of ways that you age, and, based on your metrics, they gave you an ageotype, and it determines how and what kind of way you’re aging the fastest.
Irina: I don’t rely on those, to tell the truth, because I hear from others who have been told that they are seven years younger by these metrics than their chronological age, that they are surprised, as they feel quite old regardless of this “rosy” prognostication.
Steve: I feel like that. I was 45 the day that your paper was released, and I have mornings like that where everything hurts. Hopefully reducing systemic inflammation would mean fewer days like that.
Irina: Yeah, absolutely.
Steve: It is not just a question of how long can we live, but how well can we live. I think a lot of people are more focused on the healthspan aspect. I mean, another 10, 20 extra years, I’m not going to say no, but I want those 10 or 20 years to be healthy.
Irina: That’s why I don’t ever do a longevity study of how long a mouse lives. In many of those, you have an old mouse that is very sick and sits in the corner of the cage and shivers, but that is considered as a positive outcome because it did not die yet. We don’t do this type of work. We do rejuvenation work that starts with an old animal, and you analyze if you made it younger.
Steve: Using other examples of animals in nature, negligibly senescent animals in particular, we see that one of the reasons why they live so long is their ability to regenerate and repair. That’s probably why they live so long compared to us, although we don’t do badly. Then you’ve got super agers like the mole rat, the clam that lives right at the bottom of the ocean.
Irina: I don’t try to focus on either, because squirrels live as long as mole rats, and they’re a specialized case. All the squirrels that you see running around you outside, they live 20 to 30 years, while a rat, which looks exactly like a squirrel, lives only to three. So, there are many examples of enhanced longevity that we do not understand.
Steve: As far as I can tell, biological immortality, negligible senescence, or very long longevity tends to occur in extremely stable environments, such as at the bottom of the ocean where things don’t change much. If the environment doesn’t change, then there’s no environmental pressure or evolutionary pressure to change.
Irina: Clams are just so different from people that one interesting thing might be, again, a specialized case that explains how they live so long and cannot be applied to more complex systems like mammals. All of that, of course, deserves consideration and research. We should look into multiple directions, no doubt.
Steve: Comparative biology is interesting in itself. You never know what you might find out. I think you’re closing in on the situation, and it’d be interesting to see this dilution thing finally put to rest. It’s been going on a long time.
Irina: I agree.
Steve: And the actual practicality of it isn’t too bad. I recall from the interview in January that you mentioned that, because there’s so many feedback loops in the system, the reversed or younger blood state tends to be or could potentially be quite persistent for a while, so it’s not like we’re going to need to do it every week.
Irina: Yes, exactly, but you need to figure out when and how to do it. In conventional therapy plasma exchange, they remove almost all of your blood plasma, and a designer procedure could be applied where it is less harsh and has stronger positive effects. There’s still repositioning, so you should not try to use it right away. For example, if you have a fibrotic or inflammatory disease, you need to discuss with your doctor whether or not TPE is good for you. But to say that now everybody can just go and get rejuvenated, it’s a few steps removed from that; we still need to do the clinical trial. In the clinical trial, we will know so much more, then we can really prescribe the best procedure for each individual or group of people that have similar diseases.
Steve: That’s the key, because the FDA is definitely not going to approve anything for age reversal, but they will obviously go with things like disease modification. You could pick fibrotic diseases and say, “We’re going to go for this target, this is the endpoint.” That’s how you get it in.
Irina: Yes, that’s our business plan. To go for fibrotic diseases, this is our target and endpoint. For inflammatory diseases, this is a different target and endpoint. For type 2 diabetes or associated metabolic diseases, this is the target and endpoint. Once we have success for all those in clinical trials, then we feel strongly that the FDA will approve the modification or designer TPE for each group.
Steve: In its current state, it’s already approved anyway. This is the next-generation version of it. If you can demonstrate the disease reversal or modification, then that would be approved. Of course, once it’s approved, then it has lots of potential off-label use, and it goes from there.
Irina: They could compete with us, but by that time, we will have nextgen devices, products and services that are more effective and safe, and, of course, we are known names in the field.
Steve: The quantification systems and devices are getting more sophisticated now as well. Medical wearables are becoming extremely popular now with just the general public, things like that. It’s quite plausible that, in order to find out when you need to go in for your next oil change, you could have a little sensor or something implanted.
Irina: That’s exactly one of the future directions, a wearable analytical device, the direction which is illustrated in our recent paper in Lab on Chip. So yes, you can envision, not now but let’s say in seven – ten years, that you have a wearable device on your wrist, which tells you that, hey, your TGF beta is a little bit too high. This is the schedule for you to come and have the procedure.
Steve: It could be a bit like the little service light coming on your car dash. We talked about how, in the future, a filtering system could be reduced in size eventually to potentially be a wearable device?
Irina: I don’t think so. I don’t think that our filter system could be reduced to that. Maybe at some point, not soon.
Steve: Yeah, right now, you’d have to walk around with it on a trolley, which is a little bit inconvenient.
Irina: It’s a little inconvenient, but for somebody who has a degenerative pathology, it’s a better alternative than progressive health decline.
Elena: From what age do you think this kind of therapy might be useful for people? I’m asking because some of the rejuvenation potential of this therapy, so it seems will be very interesting for women who would like to extend their period of fertility, and they wonder also, as women naturally lose some blood every month, at least some of them do, does it make any sense to sort of replenish the blood with this kind of therapy?
Irina: Those are very interesting questions. We have a project that addresses effects on reproductive function, both in males and females, and there is at least anecdotal evidence of improvement. We believe that our approach will be helpful for extending reproductive health, but we have to try it first in experimental animals to answer this question. This specific age might differ from person to person, so that’s, again, personalized rejuvenation; this is our mission and know-how as well. With respect to the monthly blood loss, this is an amazing idea; I have never thought of it. Never really came to mind, but I have known that, for example, women are healthier than men for some time with respect to cardiovascular disease. I wonder how much of that comes from this effect of dilution of blood? I don’t know.
Steve: I would say estrogen also could have quite an effect on longevity. There was a study back in 2009 that showed how estrogen actually interacts with TERT and activates telomerase.
Irina: What if we start this treatment before the loss of estrogen or loss of testosterone, can we maintain those positive hormones for decades longer? Something to be tested.
Steve: Yes and they’ve already got implanted devices that can help regulate hormones as well.
Irina: The thing is, and this is again the major paradigm shift of our paper, it suggests that hormonal therapy was unsuccessful because of the aged combination of multiple factors, molecules and cells, that dominantly inhibit anything positive that you introduce. A metaphor is that if you have a cluster of rotten bananas, and you throw a healthy banana, a nice, good banana, into that cluster, is that going to help? You are not going to rejuvenate the cluster. The other bananas will rot, and your banana will rot as well. That applies not just to hormone replacement but also to cell transplantation, when people try to put young, healthy stem cells or tissues into old mammals. There is a need to reduce the inhibitory components of the aged environment, and there is a potential you don’t need to add anything because hormones do not disappear from our body. They simply are not expressed. They might be re-expressed again, once you remove their inhibitors.
Steve: Well, it deals with the central theme, which is inflammation. Inflammation interrupts signaling and normal function, makes the immune system not work properly, reduces NAD, it does all those things.
Irina: This plethora of things which go wrong with aging is dealt with at once. Kind of like how antibiotics are dealing with bacterial infections, as a class.
Steve: Somebody once said to me, if you consider the hallmarks of aging and its distinct categories, they are like a wagon wheel. Imagine each spoke as a hallmark, and at the center of the wheel that joins all the spokes together is inflammation, because that’s how all of these hallmarks are linked.
Irina: I don’t know, I’m not a fan of such simplicity. I prefer to go from the other direction where you have a hypothesis, and you find a way to test that hypothesis. And then, science takes care of you because it tells you this is how it is instead of trying to drag science into a particular direction. Take care of science and science will take care of you.
Literature
[1] Conboy, I. M., Conboy, M. J., Kiprov, D., Kato, C., Etienne, J., Liu, C., … & Mehdipour, M. (2020). Rejuvenation of three germ layers tissues by exchanging old blood plasma with saline-albumin. Aging, 12(10), 8790-8819.
[2] Calado, R. T., Yewdell, W. T., Wilkerson, K. L., Regal, J. A., Kajigaya, S., Stratakis, C. A., & Young, N. S. (2009). Sex hormones, acting on the TERT gene, increase telomerase activity in human primary hematopoietic cells. Blood, The Journal of the American Society of Hematology, 114(11), 2236-2243.
