Amniotic fluid and tissue from Adam Yanke. He was also from Rush Health Care System here with Brian. And he's been doing a lot of research in this area and a lot of review. This is a tough topic, so thank you for tackling it. So here he is. Well, I'll just start out by saying I probably did a terrible job, and everything else should be better than that. And then also, I don't know if you guys have ever read Thinking Fast and Thinking Slow with Kahneman and Tversky. And there's a saying about Tversky that's the longer it takes you to figure out you're not as smart as him, the dumber you are. And I felt a lot like that with Arnold Kaplan up here. So Dr. Moorman, thanks for buffering that a little bit. You're also extremely smart. Is that a compliment? Yeah, that came off the wrong way. Did he go to the bathroom? Is he still in the room? Good, good. This is not recorded or anything, right? And then my last statement to dig myself further in the hole is I was asked by our marketing department to do a commercial on amniotic fluid injections in a level 1 RCT that we were doing. And I was like, sure, that sounds great. And they asked me a million questions about stem cells. And I was like, that's not what it is. I was like, what are you going to put in this? That's not what I'm saying. So now if you Google Adam Yankee in stem cells, apparently that's what comes up. So I'm not proud of that. And I just want to just bring that up right now. I don't want to be called out directly either for that. OK, so amniotic products. I do have some conflicts. I've had research funding only from organogenesis. I'll tell you about a study I was involved in that's part of this talk. Otherwise, I'm not involved in any others. It's pretty simple. There's just this many that you can use. And you just have to pick one that you can get a hold of. So it's clearly not simple. This is what's kind of self-proclaimed composition and storage and processing of different products that are available. The point of this is it's meant to be a busy slide. It's hard to figure out. There's a lot of different options. That's part of what this is all about in putting science behind it. If you do go to clinicaltrials.gov, there's five studies that are actively enrolling in the US for amniotic products to be used for the treatment of knee arthritis. If you look at studies that do not involve the knee, there's over 40 more that involve other parts of the body. So a lot of this has been done with the membrane itself, either in ophthalmology or in wound care. And there's GI, renal, all different sources. So we're kind of late to the party. But there certainly could play a role. This is something that we don't have as much evidence for. But a lot of other fields have a lot of evidence for. So it could certainly play a role. It's good to understand the contents and the processing of amniotic products and really where they come from. And so something from a patient perspective is to always mention these are healthy elective C-sections with mothers that are screened. And so that's where this comes from. It's not something that comes from an amniocentesis or anything else along those lines. When we look at the contents, there's the maternal tissue. And adjacent to that is the chorion and then the amnion. Some products have amnion alone. Some have amnion and chorion. I'm not aware of any that are chorion alone. That may be the case, but I'm not aware of any. They're very loosely associated. So it is very easily to bluntly dissect off the chorion from the amnion. And so when people talk about amniotic membrane, typically they're referring to that amnion layer. That layer has multiple components to it. It does have some natural functions, which may be important for considering homologous use with the FDA. But none of these, except for synthesis of growth factors, would be relevant. And that's really not going to happen unless you have viable cells, which most products, I would just say arguably, may not have. There's some that claim that they do have. But I think that that still needs to be proven objectively separately by a third party. So the amnion is separated into the mesenchyme, the basement membrane, and then the epithelium. And these have different components within them. So there are MSCs that are specific to the mesenchyme. This layer is more important for adherence. And that's really what it does within the body. It's got different collagen types, one and three, and some proteoglycan and glycoproteins that are present as well. When you look at the epithelium, again, it has specific MSCs. Also can produce growth factors. Again, that's adjacent to the amniotic fluid itself. And so that's why that layer does that. And both of these have some chondrogenic differentiation potential, which has been done in the lab. So this, again, is a tissue engineering type, not in vivo differentiation, as was discussed. And they can also decrease immune response. And so the contents are these layers. Then there's obviously the fluid component. And then there's also other aspects that I don't get into as much with this talk, but umbilical cord and then combinations of different types of forms that we see here. This is just a study that did demonstrate the chondrogenic differentiation potential of the amniotic fluid-derived cells that are seen here. And then this is a study that was done at our institution with Brian Cole that was through a Korean umbilical cord product in combination with microfracture for cartilage restoration. So the rest of this, I'd like to talk about amniotic products specifically that come from the membrane. When you look at these, there's different types of processing. So you can have aseptic processing or terminal sterilization. But one way or another, you can't be transmitting disease. You can have fresh hypothermically stored tissue, or it can be dehydrated or lyophilized. So lyophilized is when you freeze it, usually flash freezing, depending on how much you're doing. And then it goes to sublimation, where the water goes directly to gas. And then you can have cryopreserved tissue as well. Hard to know what the actual benefit between cryopreserved and lyophilization is. You probably are going to get some different structures of the cell, whether they're lysed or not might be different, depending on the freezing protocol. And there may be some difference in protein storing as well there. When you looked at fresh tissue, this was not in orthopedics. But I believe this was an ophthalmology study. There was an infection rate that was basically unacceptably high. So it's pretty rare that we use fresh tissue in this setting. And certainly, I'm not aware of any products on the market using the orthopedic setting that are fresh. So most of the ones that you see will be cryopreserved or lyophilized. And this does affect the actual basement membrane. I don't think anybody would really be using these for structural support. But it does decrease the structural aspects of the amniotic membrane when they are lyophilized or cryopreserved. So these are the theoretical benefits. And again, some of this has been shown with reducing scar formation, improving ulcer healing. And I just think that we have to see how this plays a role in more complex scenarios in the knees, such as osteoarthritis, and affecting that entire environment. And so as we're using these, you want to say, what's in there? And you don't want to have this response, which I don't know if anybody's seen UHF, which is nothing. So you want to know what's in there and what you're using. And you don't want to just be applying this just because it's something new and interesting. So you really should be doing targeted approaches. So what can be in there? Well, there can be growth factors, cytokines. There's the extracellular matrix components that we talked about. And there's plus-minus cell viability that you could be delivering to the tissue of interest. The different ways to apply it currently are to use it as a physical barrier, as an injectable, or a possible scaffold for reinforcing tissue regeneration. When we look at some of the basic science here, this was a study looking at a meniscectomy model in rats, where we take rats that are meniscectomized. They're predisposed to developing arthritis, essentially, or cartilage defects. We can see that, what happens here, with injection of saline. And in this slide, they show decreased saphenin O staining, but also gross examination demonstrated increased arthritis progression. And then with the injection of a micronized amnion-chorion combination, it did seem to mitigate that progression of those abnormalities. And so that's some just good basic science work to say that there may be some role here in preventing arthritis, or possibly treating arthritis. Similar study looking at a rat model of osteoarthritis using two different doses. I think that's the point I want to bring home with this study, is that there was a dose response. So with the lower dose, they did not see a difference. And with the higher dose, there is a difference. This is just like Dr. Dragoo was saying. It matters the content of what we're putting in, probably also the relative concentration compared to the tissue that you're treating as well. So one concentration within the joint may not work at the same concentration in the tendons. And so this really is going to become very highly specialized. And with having so many different options, it makes it very difficult to figure out. This is level four evidence. And so again, this is meant to be busy on purpose. This is a study looking at 40 patients treated for all sorts of different diagnoses. This doesn't help us with clinical treatment. It's good to show that it's been safe and didn't have any side effects, but that's about all you can take from that. Similarly, Jack Farr did a study on an injectable for amniotic membrane that is cryopreserved for knee arthritis. And again, this showed some clinical improvements, but really was good as a safety study, just showing that there was no adverse events with this. This expanded into a study that we were involved in as well that was a randomized control trial looking at ASA, which is the amniotic suspension allograft compared to saline and hyaluronic acid. That was a single injection hyaluronic acid formulation. This is the first ASA for, I'm sorry, the first RCT for amniotic products in the knee for arthritis. And what this showed is that at three months, it was superior to HA, and at six months, it was superior to HA and saline. So that's a pretty good long-term outcome for an injection-type study. We also had a placebo response of the saline. Again, that probably is dilutional, or it's just the placebo response of being enrolled in a study and potentially having a treatment group that's amniotic membrane. And then finally, this is the study that was mentioned. So for plantar fasciitis, this is the only other RCT I'm aware of in orthopedics for an amniotic product. And they did show significant improvement in VAS compared to saline. The problem is this is at three months, so it's just hard to know if that's really a long enough time point or not, and is that really a lasting effect or not. And so something else that's gonna be coming up in the future is a possible barrier to utilization of these products. And so this is important to know. When they're in sheets, that's considered minimally manipulated, and as long as you're using them as a barrier, that's what the amniotic membrane was used for natively, and so that's theoretically okay. If it's something that is lyophilized, or ground up, or minced, that is more than minimally manipulated. My understanding is that this is gonna be in place for people still to use for approximately three years, and after that point, products will require a BLA for that to occur. And I do know, I know of at least one company that's pursuing that for knee arthritis as an injectable, so people are definitely trying to make that possible. And this is self-serving, but this is just a little bit of a summary for what we were talking about earlier for the different products that are available and how to kind of approach this. And then in conclusion, I just think we need to be thoughtful. As Dr. Dragoo said, you wanna give all these things a good name. There's a lot of science here, and there's a lot of knowledge in this room, and the more that we can use these things thoughtfully and not as a sales tactic, the better off we're all gonna be. And the reality of how this tends to go in my experience is you do something like an RCT, you develop evidence first, then you take, I hope that's not gonna happen this whole time. That's annoying. Then you find out if there's a clinical effect. If there is a clinical effect, then you look to see what the contents of what you injected actually was. As long as it was safe, we frequently don't care that much about the details of the content, unfortunately. Once we know the content, we can start to do targeted therapies based on pathology and the different tissues that we're trying to go after, and then you go back to more RCTs based on that theory. Thank you very much, and thanks for inviting me. Thank you.

amniotic products

orthopedics

evidence-based practice

processing methods

positive outcomes