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A2926 - Decoupling of Biochemical and Biophysical Contributions to Fibrocyte Accumulation, Density and Aspect Ratio in the Scleroderma Lung Microenvironment
Author Block: H. Sun1, J. Winkler2, M. Minasyan1, H. Pan3, D. Omkar1, A. Gonzalez1, A. Pellowe1, J. Meng4, X. Peng5, E. Herzog6; 1Internal Medicine, Yale University, New Haven, CT, United States, 2Pulmonary, Critical Care and Sleep Medicine, Yale University, new haven, CT, United States, 3Yale Univeristy, New Haven, CT, United States, 4Department of Respirology, Xiangya Hospital, Central South University, Changsha, China, 5Yale University, New Haven, CT, United States, 6Pulmonary Medicine, Yale University School of Medicine, New Haven, CT, United States.
Decoupling of Biochemical and Biophysical Contributions to Fibrocyte Accumulation, Density and Aspect Ratio in the Scleroderma Lung Microenvironment
Huanxing Sun, Julia Winkler, Maksym Minasyan, Hongyi Pan, Omkar Desai, Anjelica Gonzalez, Amanda Pellowe, Jie Meng, Xueyan Peng, Erica L. Herzog
Yale University School of Medicine/Department of Internal Medicine/Section of Pulmonary/Erica Herzog Lab, 300 Cedar Street, New Haven, CT/06520, USA
Supported by Scleroderma Foundation (HS) and NIH awards HL109233 and HL125250 (ELH).
Rationale:
Fibrocytes are collagen-producing leukocytes that accumulate in patients with scleroderma (systemic scleroderma, SSc) associated interstitial lung disease (SSc-ILD) via unknown mechanisms. Our prior work showed that fibrocytes accumulate in an ex vivo mimetic of SSc-ILD generated by the decellularization of explanted human lungs. However, the biophysical and biochemical factors accounting for this observation remain unclear. In this study, we used peripheral blood mononuclear cells (PBMCs), which contain the population of fibrocytes, and a novel bioengineering based culture platform of “tunable” hydrogels conjugated with purified human lung ECM to perform previously impossible decoupling of the contribution of ECM proteins and tissue stiffness to the accumulation of fibrocytes.
Materials and methods:
PBMCs obtained from subjects with SSc-ILD were seeded onto soft (1kPa) or stiff (20kPa) hydrogels to which ECM derived from control lung scaffold (CLS) or SSc-ILD lung scaffold (SLS) had been conjugated. PBMCs were cultured for up to 14 days and CD45+Pro-ColIa1+ cells meeting criteria for fibrocytes were evaluated along with cellular density, morphology, and aspect ratio.
Results:
Relative to cells grown on soft hydrogels conjugated to CLS, there was increased accumulation of fibrocytes seen in PBMCs grown in soft hydrogel conjugated with SLS. The cells appeared clustered into foci with aspect ratio changed. . In contrast, cells grown on stiff hydrogels conjugated to CLS showed a numerical increase in fibrocytes with an elongated, spindle shaped morphology and an increase in aspect ratio. Cellular density was uniform. Finally, the stiff hydrogels conjugated to SLS showed the most collagen expression, an intermediate aspect ratio, and marked organization of fibrocytes into large foci.
Conclusion:
Biophysical and biochemical properties of the SSc-ILD lung ECM regulate production by fibrocytes or fibrocytes-like cells. Stiffness appears to regulate aspect ratio whereas matrix composition appears to regulate cell density and clustering into foci. Application of this novel method to further studies of SSc-ILD, as well as other forms of lung remodeling and cell populations, may provide important new insights into cell:matrix interactions in the human lung.