Designing 3D Synthetic Matrices for Studying Interactions Between Bacterial Peptidoglycan and Macrophages in the Context of Immune Misrecognition Diseases

Abstract

The immune system is a complex network comprised of various cellular components that work together to protect the body from damage, with a key function of differentiating between pathogenic bacteria and the 39 trillion commensal bacteria that constitute our microbiome. Misrecognition of commensal bacteria has been linked to many inflammatory bowel disorders in humans yet remains not fully understood and difficult to study. Enabled by recent advances in molecular tools, fragments of the bacterial cell wall peptidoglycan have been identified that bind to pattern recognition receptors within macrophages, which activate macrophages to start a signaling cascade that turns on the immune response. Typically, these interactions are studied using two‐dimensional invasion assays on tissue culture plastic plates; however, the dimensionality and ‘stiffness’ of these traditional culture systems may not be representative of the cell‐microenvironment interactions that exist in vivo. In order to better simulate the native cellular environment, three‐dimensional (3D) cell culture systems can be used to control key biochemical and mechanical properties of the cell microenvironment. Here, we established an approach for the encapsulation and culture of human macrophages in 3D within well‐defined, biomimetic hydrogels decorated with integrin binding peptides that mimic the natural in vivo environment and linked with cell‐degradable sites for cell‐driven migration and remodeling to probe macrophage response to bacterial invasion. With this approach, we have shown invasion with bacteria whose glycans are remodeled and their engulfment through immunostaining and assessed activation of macrophages using RT‐qPCR and ELISA. These studies provide new insights into the interactions between bacteria and immune cells and this new method affords unique opportunities for examining how our immune system recognizes and misrecognizes bacteria toward a better understanding of the mechanisms of immune misrecognition diseases.Support or Funding InformationNIH CBI Training Grant‐5T32GM008550NIH U01 Common Fund program‐U01CA221230‐01