29Animal models of bone marrow have limited spatial and temporal resolution to observe biological 30 events (intravasation and cellular egress) and are inadequate to dissect dynamic events at the niche level 31 (100 microns). Utilizing microfluidic and stem cell technology, we present a 3D in vitro model of human 32 bone marrow that contains perivascular and endosteal niches complete with dynamic, perfusable vascular 33 networks. We demonstrate that our model can perform in vivo functions including maintenance and 34 differentiation of CD34 + hematopoietic stem/progenitor cells (HSPC) for up to fourteen days, egress of 35 myeloid progenitors, and expression of markers consistent with in vivo human bone marrow. The platform 36 design enables the addition of tissue niches at a later timepoint to probe mechanisms such as tumor cell 37 migration. Overall, we present a novel organ-on-a-chip platform that is capable of recapitulating the human 38 bone marrow microenvironment to observe hematopoietic phenomena at high spatial and temporal 39 resolution. 40 41 Manuscript Template Page 2 of 34 MAIN TEXT 42 43the abluminal surface of a blood vessel comprises the perivascular niche (4, 9). 51Both the perivascular and endosteal niches contain sinusoidal blood vessels as a key feature of the bone 52 marrow. These specialized vessels are surrounded by many different cell types including mesenchymal stem cells 53 (MSC), specialized CXCL12 abundant reticular (CAR) cells, HSC, leukocytes at different stages of differentiation, 54 and adipocytes (10). Bone marrow stromal cells are known to secrete chemotactic signals such as stem cell factor 55 (SCF) (4) and CXCL-12/SDF-1 (11). Endothelial cells express specialized ligands on the luminal surface such as E-56 selectin, ICAM, and VCAM (3, 12). These chemokines and adhesion molecules cooperatively work to facilitate 57 homing of circulating HSC to bone marrow (3, 13). In vivo, osteoblasts also express VCAM and CXCL-12 in 58 addition to osteopontin, which has been implicated in maintaining HSC quiescence (7, 14). Moreover, blood vessels 59 found within the endosteal niche express higher levels of E-selectin, which play an important role in regulating HSC 60 cell cycling (15), and has been implicated as a regulator of breast cancer metastasis to bone (12, 16). We endeavored 61to create a unified model of these two bone marrow niches (perivascular and endosteal) using an in vitro using a 62 microfluidic organ-on-a-chip system. 63Advances in tissue engineering and microfluidics have created "organ-on-a-chip" technologies to generate 64 3D microphysiological mimics that utilize a broad range of human cells, thus providing specific advantages over 65 traditional 2D culture and mouse models. Moreover, microfluidic technology lends itself well to capturing the spatial 66 scale of the adjacent endosteal and perivascular bone marrow microenvironments. Although several labs have utilized 67 similar tissue engineering and microfluidic approaches to recreate aspects of the perivascular and endost...
An atomic-scale mechanism describing the role of mechanosensing in T Cell Receptor (TCR) recognition of peptides in the binding groove of the peptide-major histocompatibility complex (pMHC) may inform the design of novel TCRs for immunotherapies. Using steered molecular dynamic simulations, our study demonstrates that mutations to peptides in the binding groove of the pMHC – which are known to discretely alter the T cell response to an antigen – influence MHC conformation and thus the overall strength of the TCR-pMHC bond including duration and length under constant load. Moreover, physiochemical features of the TCR-pMHC dynamic bond strength, such as hydrogen bonds and Lennard-Jones contacts, correlate with the immunogenic response elicited by the specific peptide in the MHC groove. Thus, formation of transient TCR-pMHC bonds is a characteristic of immunogenic peptides and is mediated by stabilized interactions.x
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