Immunoengineering

National Institute Of Biomedical Imaging And Bioengineering, Bethesda

Investigators

Linked publications, trials & patents

Abstract

Finalizing SARS-CoV-2 Research We are finalizing the publication of our last works on COVID19 research which have all completed experimentation and are being prepared for submission to scientific journals. Please refer to prior report regarding COVID19 research projects. Identification of novel mediators of injury recovery and materials response Early in response to injury and biomaterial implantation, pro-fibrotic materials (polyethylene; PE) induced the recruitment of a strong neutrophil population in comparison to pro-regenerative materials (decellularized extracellular matrix; ECM) recruit a large proportion of eosinophils, commonly associated with allergy and asthma but previously described as important mediators of tissue development in the liver and muscle after cardiotoxin injection. When shifting focus to dendritic cells which have been largely ignored (with more emphasis on macrophages), we found that when an injury was treated with ECM biomaterials there was a preferential recruitment of CD103+XCR1+ type-1 conventional dendritic cells (cDC1s) that had atypical surface marker expression including an absence of CD8 and presence of CD301b, CD206, along with low levels of CD11b (Lokwani, Josyula, Ngo et al Accepted in Principle 2023). To determine if these trauma-associated dendritic cells (tDCs) were in fact cDC1s, we evaluated their presence in a Batf3-/- model (a common depletion model for cDC1s) and found ablation of these dendritic cells in the absence of BATF3, suggesting that they are cross-presentation capable cDC1-like cells. Pathologically, the Batf3-/- mice displayed increased intramuscular adipose and fibrosis along with stochastically positioned necrotic and calcified muscle fibers. This was mirrored with gene expression patterns where we saw increases in Col1a1 and Adipoq associated with fibrosis and white fat, and Acvr1, which is associated with a genetic disease in humans characterized by severe post-traumatic soft tissue ossification (FOP). Interestingly, we also saw increases in the expression of several genes associated with muscle regeneration including Gli1, suggesting a larger regulatory role of BATF3-dependent cells in post-trauma tissue development. As these cDC1s can communicate with CD8+ T cells, we investigated adaptive immune responses to trauma. Previous literature has associated a non-canonical CD8 regulatory cell in regulating autoimmunity, and thus our interest was piqued as these cells potentially playing an immunoregulatory role early in the response to wounding. We found a number of CD4+FoxP3+ regulatory T cells as well as CD4+ and CD8+ HELIOS+ induced Tregs (iTregs). We are continuing our exploration of these CD8 iTregs in trauma. We have multiple follow-up projects to the above referenced research that we will be continuing throughout FY24. Large volume imaging of biomaterial implantation in a mouse model In collaboration with AIM we developed a method to clear tissues and image biomaterial implants made of decellularized extracellular matrix with light sheet microscopy. Using a graded methanol series followed by DCM and DBE clearing, we generated thick (1cm), optically clear quadriceps muscles and peritoneal cavities to visualize the large volumetric structure around these biomaterial implants with autofluorescence alone. We could then reconstruct the tissue in 3D, and apply machine learning methods to categorize the tissue types based on the autofluorescence emission spectra. We were able to repeatably and accurately map muscle, ECM scaffold, interstitial space, and to a lesser extent blood vessels. The role of implant location in immune response to scaffolds Biomaterial scaffolds are used in a variety of tissue contexts. Decellularized extracellular matrix (ECM) scaffolds are used for abdominal wall repair during hernia reconstruction, dural repair after neurosurgery, breast filling after lumpectomy, diabetic foot ulcer treatment, skin injury reconstruction, and have been tested for the treatment of volumetric muscle loss (VML) and more significant tissue defects. When comparing intraperitoneal implant, versus subcutaneous implant, versus VML model, we found similarities and differences based on tissue location (DeStefano et al bioRxiv). Histologically, we say that there was a strong cellular infiltration into biomaterial implants in the subcutaneous space which was increased in the presence of an injury. With intraperitoneal implants, an increased cellularity was observed within the peritoneal cavity but infiltration in the material itself was decreased and limited to clusters around the periphery of the implant. In the VML we saw a strong preference for an M2-like macrophage polarization with enrichment of CD206 and CD301b expression on MHCII- macrophages. This supports prior work where the more M2-polarized macrophages were not antigen presenting (MHCII-) whereas those that expressed MHCII (suggesting communication with T cells) did not express as strong levels of these scavenger receptors. This peaked by 21 days post-injury with greater cell infiltration when compared to the IP and SQ models. In the IP model, there were two strong differences including a lower proportion of Ly6Chi macrophages which are associated with peripheral blood recruitment and correlates with previous findings of robust tissue-resident macrophage population in the peritoneal cavity. These implants also recruited a very strong proportion of CD19+ B Cells which was in stark contrast to SQ and VML which did not recruit large numbers of B Cells. These B cells also expressed MHCII, suggesting that they could behave as antigen presenting cells. Continuing research on human immune response to traumatic injury We evaluated a panel of 59 biomarkers in the plasma of 1000 trauma patients alongside 50 healthy adults. Several interesting trends emerged, both confirming prior work and identifying new immune patterns of human traumatic injury. We found an upregulation of IL-10 which is known to increase in trauma patients, this had a correlation with trauma severity where those that ended up dying from their injuries were >70% positive for IL-10 whereas healthy controls had all but one sample return as undetected (2% positive). We are currently evaluating these markers in our mouse model of volumetric muscle loss to determine the potential correlates with human immunity and when during the course of injury recovery, we may see these similarities or differences occur. We hope for this work to be submitted for publication in FY24. We have established an MTA with the UMD Biorepository for a prospective study on discarded IRB-exempt human specimens. We have received our first sample and will continue to collect these at an estimated pace of 1 2 samples per month. We expect the first publication from this study to occur sometime in 2025 2026 pending sufficient sample collection