Cytokines and Extracellular Matrix

Key Personnel

• Michael White
  University of Chicago

Project Description

Morphogenetic signals from the cell’s microenvironment play important roles in stem and progenitor cell differentiation and in multicellular morphogenesis, being driven by extracellular matrix (ECM) proteins and cytokines. We focus on how signaling from cytokines is modulated by the presence of and interactions with ECM proteins and how this modulates morphogenesis. This interaction leads to synergistic signaling between adhesion receptors and cytokine receptors. We develop molecular engineering approaches to create recombinant cytokine variants that bind the ECM with unusually high affinity, to enhance their translational potential. Recombinant ECM proteins can be built into biomaterials, or they can be assembled into a gel network to be used as a biomaterial matrix themselves. Our goal is to use these matrix and cytokine engineering approaches to support the multicellular morphogenesis that occurs between nephron progenitor cells, the developing microvasculature, and other stromal cells.

Publications

1. Abstract

   Kidney repair and regeneration: perspectives of the NIDDK (Re)Building a Kidney consortium

   Naved, Bilal A.; Bonventre, Joseph V.; Hubbell, Jeffrey A.; Hukriede, Neil A.; Humphreys, Benjamin D.; Kesselman, Carl; Valerius, M. Todd; McMahon, Andrew P.; Shankland, Stuart J.; Wertheim, Jason A.; White, Michael J.V.; de Caestecker, Mark P.; Drummond, Iain A. Kidney International . March 2022.

   Acute kidney injury impacts ∼13.3 million individuals and causes ∼1.7 million deaths per year globally. Numerous injury pathways contribute to acute kidney injury, including cell cycle arrest, senescence, inflammation, mitochondrial dysfunction, and endothelial injury and dysfunction, and can lead to chronic inflammation and fibrosis. However, factors enabling productive repair versus nonproductive, persistent injury states remain less understood. The (Re)Building a Kidney (RBK) consortium is a National Institute of Diabetes and Digestive and Kidney Diseases consortium focused on both endogenous kidney repair mechanisms and the generation of new kidney tissue. This short review provides an update on RBK studies of endogenous nephron repair, addressing the following questions: (i) What is productive nephron repair? (ii) What are the cellular sources and drivers of repair? and (iii) How do RBK studies promote development of therapeutics? Also, we provide a guide to RBK’s open access data hub for accessing, downloading, and further analyzing data sets.

2. Abstract

   Engineered collagen-targeting therapeutics reverse lung and kidney fibrosis in mice (Preprint)

   White, Michael JV; Raczy, Michal M; Budina, Erica; Yuba, Eiji; Solanki, Ani; Shim, Ha-Na; Zhang, Zheng Jenny; Gray, Laura T; Cao, Shijie; Alpar, Aaron T.; Hubbell, Jeffrey A. bioRxiv . 2022.

   Fibrotic diseases are involved in 45% of deaths in the United States. In particular, fibrosis of the kidney and lung are major public health concerns due to their high prevalence and lack of existing treatment options. Here, we harness the pathophysiological features of fibrotic diseases, namely leaky vasculature and aberrant extracellular matrix (ECM) protein deposition (i.e. collagen), to target an anti-fibrotic biologic and a small molecule drug to disease sites of fibrosis, thus improving their therapeutic potential in mouse models of lung and kidney fibrosis. First, we identify and validate collagen-targeting drug delivery systems that preferentially accumulate in the diseased organs: von Willebrand Factor’s A3 domain (VWF-A3) and decorin-derived collagen-binding peptide-conjugated micelles (CBP-micelles). We then engineer and recombinantly express novel candidate biologic therapies based on the anti-inflammatory cytokine IL-10: A3-IL-10 and A3-Serum Albumin-IL-10 (A3-SA-IL-10). Simultaneously, we stably encapsulate the potential anti-fibrotic water-insoluble drug, rapamycin, in CBP-micelles. We show that these novel formulations of therapeutics bind to collagen in vitro and that their efficacy in mouse models of lung and kidney fibrosis is improved, compared to free, untargeted drugs. Our results demonstrate that collagen-targeted anti-fibrotic drugs may be next generation therapies of high clinical potential.Competing Interest StatementThe authors have declared no competing interest.

3. Abstract

   Myofibroblast differentiation is governed by adhesion mechanics, and inhibition of the stress sensor Talin2 reverses lung fibrosis (Preprint)

   White, Michael JV; Ozkan, Melis; Gomez Medellin, Jorge Emiliano; Zent, Roy; Critchley, David; Hubbell, Jeffrey A. bioRxiv . 2021.

   Fibrosis is involved in 45% of deaths in the United States, and no treatment exists to reverse progression of the disease. In order to find novel targets for fibrosis therapeutics, we developed a model for the differentiation of monocytes to myofibroblasts that allowed us to screen for proteins involved in myofibroblast differentiation. We assessed these screening results for proteins to target for novel fibrosis therapeutics. Here we test whether inhibition of a novel protein target generated by our model, talin2, can prevent and even reverse myofibroblast differentiation. We find that knockdown of talin2 de-differentiates myofibroblasts, altering myofibroblast morphology, α-smooth muscle actin and collagen content, and the secretome. Talin2 inhibition reverses bleomycin-induced lung fibrosis in mice. Talin2 inhibition could be a novel treatment for reversing lung fibrosis.One Sentence Summary Silencing the spring protein Talin2 reverses myofibroblast differentiation and reverses existing fibrosis.Competing Interest StatementMJVW and JH are authors on a patent regarding this work(α-SMA)alpha-smooth muscle actin(ALT)Alanine Transferase(AST)Aspartate Transferase(BAL)broncheo-alveolar lavaged(BUN)blood urea nitrogen(FCS)fetal calf serum(FAs)focal adhesions(FBs)Fibrillar adhesions(IHC)Immunohistochemistry(MRC-5)Human fibroblasts(kilopascals)kPa(MCP1)macrophage-chemotactic protein-1(NIH-3T3)mouse fibroblasts(PBMC)peripheral blood mononuclear cells(PBS)phosphate buffered saline(SFM)serum-free media(siRNA)Silencing RNA(TGFβ)Transforming growth factor β(Tln2)Talin2(TNFα)Tumor necrosis factor α(UUO)Unilateral Ureteral Obstruction

4. Abstract

   Blocking antibodies against integrin-α3, integrin-αM, and integrin-αMβ2 de-differentiate myofibroblasts and reverse lung and kidney fibroses in a mouse model (Preprint)

   White, Michael JV; Ozkan, Melis; Rączy, Michal M; Gomez-Medellin, Jorge Emiliano; Koss, Kyle M; Alpar, Aaron T.; Naved, Bilal; Wertheim, Jason; Hubbell, Jeffrey A. bioRxiv . 2021.

   Fibrosis is involved in 45% of deaths in the United States, and no treatment exists to reverse the progression of the disease. Myofibroblasts are key to the progression and maintenance of fibrosis. We investigated features of cell adhesion necessary for monocytes to differentiate into myofibroblasts, seeking to identify pathways key to myofibroblast differentiation. Blocking antibodies against integrins α3, αM, and αMβ2 de-differentiate myofibroblasts in vitro, lower the pro-fibrotic secretome of myofibroblasts, and reverse fibrosis in vivo. Blocking key integrins may be an effective therapeutic for the treatment and reversal of fibrosis. Teaser: Blocking integrin-α3, integrin-αM, and integrin-αMβ2 reverses myofibroblast differentiation and reverses existing fibrosis. Competing Interest Statement: MJVW and JH are authors on a patent regarding this submission Abbreviations: (α-SMA) alpha-smooth muscle actin (CBP-α-αMβ2) CBP-functionalized neutralizing antibodies against αMβ2 (CBP-α-αM) CBP-functionalized neutralizing antibodies against αM (CBP-α-α3) CBP-functionalized neutralizing antibodies against α3 (CBP)(LRELHLNNNC) collagen-binding peptide (FAs) focal adhesions (FCS) fetal calf serum (FBs) Fibrillar adhesions (CY7-α-α3) Fluorescently labeled Cy7-neutralizing antibodies against α3 (CY7-α-αMβ2) Fluorescently labeled Cy7-neutralizing antibodies against αMβ2 (CY7-α-αM) Fluorescently labeled Cy7-neutralizing antibodies against αM (antibody which stabilizes the association of integrin α2β1) Gi14 (MRC-5) Human fibroblasts (called MAC1) heterodimer αMβ2 (IL12p40) IL-12 subunit p40 (α3) Integrin-α3 (αM) Integrin-αM (β2)(CD18) Integrin-β2 (MCP1) macrophage-chemotactic protein-1 (NIH-3T3) mouse fibroblasts (α-αMβ2) neutralizing antibodies against αMβ2 (α-αM) neutralizing antibodies against αM (α-α3) neutralizing antibodies against α3 (PBMC) peripheral blood mononuclear cells (PBS) phosphate buffered saline (SFM) serum-free media (TGFβ) Transforming growth factor β (TNFα) Tumor necrosis factor α (UUO) Unilateral ureteral obstruction

5. Abstract

   (Re)Building a Kidney.

   Oxburgh, L; Carroll, TJ; Cleaver, O; Gossett, DR; Hoshizaki, DK; Hubbell, JA; Humphreys, BD; Jain, S; Jensen, J; Kaplan, DL; Kesselman, C; Ketchum, CJ; Little, MH; McMahon, AP; Shankland, SJ; Spence, JR; Valerius, MT; Wertheim, JA; Wessely, O; Zheng, Y; Drummond, IA. J Am Soc Nephrol . 28(5):1370–1378. May 2017.

   (Re)Building a Kidney is a National Institute of Diabetes and Digestive and Kidney Diseases-led consortium to optimize approaches for the isolation, expansion, and differentiation of appropriate kidney cell types and the integration of these cells into complex structures that replicate human kidney function. The ultimate goals of the consortium are two-fold: to develop and implement strategies for in vitro engineering of replacement kidney tissue, and to devise strategies to stimulate regeneration of nephrons in situ to restore failing kidney function. Projects within the consortium will answer fundamental questions regarding human gene expression in the developing kidney, essential signaling crosstalk between distinct cell types of the developing kidney, how to derive the many cell types of the kidney through directed differentiation of human pluripotent stem cells, which bioengineering or scaffolding strategies have the most potential for kidney tissue formation, and basic parameters of the regenerative response to injury. As these projects progress, the consortium will incorporate systematic investigations in physiologic function of in vitro and in vivo differentiated kidney tissue, strategies for engraftment in experimental animals, and development of therapeutic approaches to activate innate reparative responses.