Thanks for allowing me to present here. It really is an honor. So we'll go ahead and get into it. Our conflicts are listed here as well as are available online. You've heard a lot of this already so I don't need to reiterate all of it. But clearly the onset of arthritis after post-traumatic injuries or chronic degenerative arthritis involves a cascade of different cytokines that are time dependent. And there's some literature that's looked at acute inflammation for post-traumatic arthritis to try to mitigate the onset of arthritis. We looked at an impaction study where we took intact cartilage plugs with and without healthy synovium and found we were able to save viability or prevent the viability that happens when you don't have healthy synovium present in a osteochondral allograft model. When we looked at the mechanism of how that was being accomplished, we saw that there was a spike of IL-1 receptor antagonist or IRAP around day seven that was being secreted by the healthy synovium that was not present at the time of the impacted event. So when we think about treating patients that have arthritis, it's different than the post-traumatic arthritis model when it's chronic and degenerative. And just as Adam mentioned, sometimes we try things and see if they work first before we figure out why they might work. So we're trying to get two products that were, one was, both were available in the market when the study came out originally and now one is available through a study only and the other is still available. We wanted to try to figure out any potential mechanisms and are these truly anti-inflammatory treatments or how do they enact their clinical improvement. The two we looked at are bone marrow aspirate concentrate from the Iliac crest and the other is amniotic suspension allograft. Both of these had fairly limited clinical studies to support them, however, amniotic suspension allograft did have a level one RCT looking at hyaluronic acid and saline and found that it was superior to both of those treatments out to a year. There's also been murine models in the amniotic suspension allograft as well as equine in the bone marrow aspirate concentrate that show improved tissue formation. So they have some potential benefit in clinical and basic science studies. When we look at some of the contents of amniotic suspension allograft, this is data from the, it's white paper data, essentially it shows there's IRAP present and we've recently been a third party validation of this that show that there's a decent amount of IRAP in the amniotic suspension allograft. And while there's no magic bullet, we do seem to see a lot of increase in that in studies that show improvement of or reduction of inflammatory cytokines in arthritis. So our study was to look at the anti-inflammatory and disease modifying potential of BMAC and ASA in the setting of a synovium and an arthritic co-culture model. This was set up for 17 patients that were undergoing total knee arthroplasty and after the cuts were made for arthroplasty, we harvested explants of arthritic chondrocytes from those shavings and then we took synovium from those same patients. How am I doing? Good. So the, I don't know what that was. The synovium and the explants came from the same patients when we used the BMAC, it also came from those patients having total knees. So that's the population that we harvested from. We used clinically relevant concentrations when we did these co-cultures. So we used the same concentration that was used in the FDA clinical trial for ASA and we used the natural concentration that was available for the patient at the time of aspiration for BMAC. Our outcomes included ELISA of the media essentially, which would be the synovial fluid equivalent. We then looked at PCR and gene expression of the synovium and the cartilage and then we did histology with Mencken grading. This is probably the most important slide. So if we look on the left here, the boxes around the IL-1 beta levels and we saw is that the application of amniotic suspension allograft decreases IL-1 to 50% of what it is in the control group. And so that was a statistically significant improvement and the BMAC was basically no improvement whatsoever, almost the exact same numbers. When we did look at BMAC, there was a significant reduction of about 20% of IL-6. So not quite as drastic, but potentially different mechanisms here with regards to how inflammatory cytokines are responding. This data here, we're still trying to figure out how to key this in, but when we looked at gene expression in the cartilage, we saw that there was less gene expression of collagen 1A with BMAC compared to AMAC suspension allograft. However, neither of these were statistically different than the control. Conversely, when we looked at synovial gene expression, BMAC had significantly less, I'm sorry, significantly greater expression of collagen 2 compared to AMAC suspension allograft. Again, neither of these were statistically different from the control. When we looked at histology, we basically found things on both sides of the spectrum. Sometimes there was better proteoglycans standing with treatment, sometimes not. And ultimately the histology findings were not statistically significant. So this study does have limitations. We did not characterize the BMAC or AMAC suspension products that we used. We have separate studies that have done that, but it wasn't done for this. So we can't tie it to the results that we got and the relative effectiveness. The equivocal gene expression results may be due to the wrong timeframe to look at these, or it could purely be two small numbers and not be powered for that. There's also a decent amount of heterogeneity in the tissue that we were harvesting, and the relative amount of arthritis that was present was obviously variable depending on the indication for the total need. In conclusion, arthritic chondrocytes do seem to respond to BMAC and ASA by potentially reducing IL-1 data and IL-6. However, we did not see any mitigation of the overall disease process in these explants. I'd like to thank my co-authors, and in particular, ORIF and Charlie Hannon, who received a Clinician Scientist Grant from ORIF for this study, as well as support from CSU through the Translational Medicine Institute. Thank you.

anti-inflammatory effects

disease modifying effects

bone marrow aspirate concentrate

amniotic suspension allograft

total knee arthroplasty