Vascular growth factors play an important role in maintaining the structure and integrity of the glomerular filtration barrier. In healthy adult glomeruli, the proendothelial survival factors vascular endothelial growth factor-A (VEGF-A) and angiopoietin-1 are constitutively expressed in glomerular podocyte epithelia. We demonstrate that this milieu of vascular growth factors is altered in streptozotocin-induced type 1 diabetic mice, with decreased angiopoietin-1 levels, VEGF-A upregulation, decreased soluble VEGF receptor-1 (VEGFR1), and increased VEGFR2 phosphorylation. This was accompanied by marked albuminuria, nephromegaly, hyperfiltration, glomerular ultrastructural alterations, and aberrant angiogenesis. We subsequently hypothesized that restoration of angiopoietin-1 expression within glomeruli might ameliorate manifestations of early diabetic glomerulopathy. Podocyte-specific inducible repletion of angiopoietin-1 in diabetic mice caused a 70% reduction of albuminuria and prevented diabetes-induced glomerular endothelial cell proliferation; hyperfiltration and renal morphology were unchanged. Furthermore, angiopoietin-1 repletion in diabetic mice increased Tie-2 phosphorylation, elevated soluble VEGFR1, and was paralleled by a decrease in VEGFR2 phosphorylation and increased endothelial nitric oxide synthase Ser 1177 phosphorylation. Diabetes-induced nephrin phosphorylation was also reduced in mice with angiopoietin-1 repletion. In conclusion, targeted angiopoietin-1 therapy shows promise as a renoprotective tool in the early stages of diabetic kidney disease.

Polycystic kidney diseases (PKD) are genetic disorders characterized by progressive epithelial cyst growth leading to destruction of normally functioning renal tissue. Current therapies have focused on the cyst epithelium, and little is known about how the blood and lymphatic microvasculature modulates cystogenesis. Hypomorphic Pkd1 nl/nl mice were examined, showing that cystogenesis was associated with a disorganized pericystic network of vessels expressing platelet/endothelial cell adhesion molecule 1 and vascular endothelial growth factor receptor 3 (VEGFR3). The major ligand for VEGFR3 is VEGFC, and there were lower levels of Vegfc mRNA within the kidneys during the early stages of cystogenesis in 7-day-old Pkd1 nl/nl mice. Seven-day-old mice were treated with exogenous VEGFC for 2 weeks on the premise that this would remodel both the VEGFR3 + pericystic vascular network and larger renal lymphatics that may also affect the severity of PKD. Treatment with VEGFC enhanced VEGFR3 phosphorylation in the kidney, normalized the pattern of the pericystic network of vessels, and widened the large lymphatics in Pkd1 nl/nl mice.These effects were associated with significant reductions in cystic disease, BUN and serum creatinine levels. Furthermore, VEGFC administration reduced M2 macrophage pericystic infiltrate, which has been implicated in the progression of PKD. VEGFC administration also improved cystic disease in Cys1 cpk/cpk mice, a model of autosomal recessive PKD, leading to a modest but significant increase in lifespan. Overall, this study highlights VEGFC as a potential new treatment for some aspects of PKD, with the possibility for synergy with current epithelially targeted approaches.

A wide variety of nanomaterials are currently being developed for use in the detection and treatment of human diseases. However, there is no systematic way to measure and predict the action of such materials in biological contexts. Lipid-encapsulated nanoparticles (NPs) are a class of nanomaterials that includes the liposomes, the most widely used and clinically proven type of NPs. Liposomes can, however, activate the complement system, an important branch of innate immunity, resulting in undesirable consequences. Here, we describe the complement response to lipid-encapsulated NPs that are functionalized on the surface with various lipid-anchored gadolinium chelates. We developed a quantitative approach to examine the interaction of NPs with the complement system using in vitro assays and correlating these results with those obtained in an in vivo mouse model. Our results indicate that surface functionalization of NPs with certain chemical structures elicits swift complement activation that is initiated by a natural IgM antibody and propagated via the classical pathway. The intensity of the response is dependent on the chemical structures of the lipid-anchored chelates and not zeta potential effects alone. Moreover, the extent of complement activation may be tempered by complement inhibiting regulatory proteins that bind to the surface of NPs. These findings represent a step forward in the understanding of the interactions between nanomaterials and the host innate immune response and provide the basis for a systematic structure-activity relationship study to establish guidelines that are critical to the future development of biocompatible nanotherapeutics.

Glomerular disease often features altered histologic patterns of extracellular matrix (ECM). Despite this, the potential complexities of the glomerular ECM in both health and disease are poorly understood. To explore whether genetic background and sex determine glomerular ECM composition, we investigated two mouse strains, FVB and B6, using RNA microarrays of isolated glomeruli combined with proteomic glomerular ECM analyses. These studies, undertaken in healthy young adult animals, revealed unique strain-and sexdependent glomerular ECM signatures, which correlated with variations in levels of albuminuria and known predisposition to progressive nephropathy. Among the variation, we observed changes in netrin 4, fibroblast growth factor 2, tenascin C, collagen 1, meprin 1-a, and meprin 1-b. Differences in protein abundance were validated by quantitative immunohistochemistry and Western blot analysis, and the collective differences were not explained by mutations in known ECM or glomerular disease genes. Within the distinct signatures, we discovered a core set of structural ECM proteins that form multiple protein-protein interactions and are conserved from mouse to man. Furthermore, we found striking ultrastructural changes in glomerular basement membranes in FVB mice. Pathway analysis of merged transcriptomic and proteomic datasets identified potential ECM regulatory pathways involving inhibition of matrix metalloproteases, liver X receptor/retinoid X receptor, nuclear factor erythroid 2-related factor 2, notch, and cyclin-dependent kinase 5. These pathways may therefore alter ECM and confer susceptibility to disease.

Nanoparticles offer new options for medical diagnosis and therapeutics with their capacity to specifically target cells and tissues with imaging agents and/or drug payloads. The unique physical aspects of nanoparticles present new challenges for this promising technology. Studies indicate that nanoparticles often elicit moderate to severe complement activation. Using human in vitro assays that corroborated the mouse in vivo results we previously presented mechanistic studies that define the pathway and key components involved in modulating complement interactions with several gadolinium-functionalized perfluorocarbon nanoparticles (PFOB). Here we employ a modified in vitro hemolysis-based assay developed in conjunction with the mouse in vivo model to broaden our analysis to include PFOBs of varying size, charge and surface chemistry and examine the variations in nanoparticle-mediated complement activity between individuals. This approach may provide the tools for an in-depth structure-activity relationship study that will guide the eventual development of biocompatible nanoparticles.