Glomerulonephritis is most commonly an autoimmune-induced inflammatory event wherein infiltration of the kidney by macrophages results in loss of renal function due to fibrosis. The long-term goal of our laboratory is to identify the molecular switches that govern macrophage adhesion and thus regulate the inflammatory response. We previously identified tyrosine phosphorylation of 23 integrin adhesion receptors as one such switch and have demonstrated that 23 phosphorylation is necessary for macrophage adhesion to some ligands. Blockade of 23 phosphorylation results in loss of adhesion due to a failure of the actin cytoskeleton to organize into stress fibers. In this application we present evidence that this unique mechanism is required for macrophage entry into renal tissues. We hypothesize that macrophage passage through renal basement membrane is mediated by an interaction between the proteoglycan agrin and phosphorylated 23 integrins. In this application we will assess the role of 23 integrins and their phosphorylation in migration to renal tissues during experimental glomerulonephritis. We will determine whether agrin in renal basement membrane regulates macrophage infiltration by in vitro transmigration studies. We will characterize the production of macrophage podosomal adhesions and pseudopods that are required for migration into three-dimensional tissues by TIRF microscopy of living macrophages. We will biochemically determine the signaling from phosphorylated 23 that controls actin cytoskeletal reorganization in the macrophage pseudopod and podosome. Identification of mechanisms used during organ-specific macrophage immune trafficking can provide novel targets for therapeutic intervention and complement global immune-suppressive therapies.

Public Health Relevance

r laboratory has identified a new mechanism of white blood cell adhesion to tissues. This research proposal will determine whether a particular adhesion molecule called 23 integrin, found on a subset of white blood cells, contributes to glomerulonephritis (inflammation of the kidneys). Understanding the mechanism of glomerulonephritis can provide new targets for drugs to reduce the progression of this disease.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
Project #
Application #
Study Section
Intercellular Interactions (ICI)
Program Officer
Flessner, Michael Francis
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Upstate Medical University
Anatomy/Cell Biology
Schools of Medicine
United States
Zip Code
Miller, Matthew R; Miller, Eric W; Blystone, Scott D (2017) Non-canonical activity of the podosomal formin FMNL1? supports immune cell migration. J Cell Sci 130:1730-1739
Miller, Matthew R; Blystone, Scott D (2015) Human Macrophages Utilize the Podosome Formin FMNL1 for Adhesion and Migration. Cellbio (Irvine, Calif) 4:1-11
Miller, Matthew R; Blystone, Scott D (2015) Reliable and inexpensive expression of large, tagged, exogenous proteins in murine bone marrow-derived macrophages using a second generation lentiviral system. J Biol Methods 2:e23
Krainer, Elisabeth C; Ouderkirk, Jessica L; Miller, Eric W et al. (2013) The multiplicity of human formins: Expression patterns in cells and tissues. Cytoskeleton (Hoboken) 70:424-38
Yagi, Mayumi; Murray, Jacqueline; Strand, Kurt et al. (2012) Heparin modulates the conformation and signaling of platelet integrin ?IIb?3. Thromb Res 129:743-9
Yamodo, Innocent H; Blystone, Scott D (2012) Calcium Integrin Binding Protein Associates with Integrins ?V?3 and ?IIb?3 Independent of ?3 Activation Motifs. Cellbio (Irvine, Calif) 1:30-37
Gianni, Tatiana; Cerretani, Arianna; Dubois, Rebecca et al. (2010) Herpes simplex virus glycoproteins H/L bind to cells independently of {alpha}V{beta}3 integrin and inhibit virus entry, and their constitutive expression restricts infection. J Virol 84:4013-25
Mersich, Akos T; Miller, Matthew R; Chkourko, Halina et al. (2010) The formin FRL1 (FMNL1) is an essential component of macrophage podosomes. Cytoskeleton (Hoboken) 67:573-85
Gao, Chunlei; Blystone, Scott D (2009) A Pyk2-Vav1 complex is recruited to beta3-adhesion sites to initiate Rho activation. Biochem J 420:49-56
Christophi, George P; Panos, Michael; Hudson, Chad A et al. (2009) Macrophages of multiple sclerosis patients display deficient SHP-1 expression and enhanced inflammatory phenotype. Lab Invest 89:742-59