The goal of this project is to determine how changes in kidney extracellular matrix (ECM) stiffness caused byadvanced glycation impact progression of diabetic nephropathy. Our overarching hypothesis is that advancedglycation end products (AGEs) form crosslinks that increase ECM stiffness in the kidney and alter integrinsignaling to trigger increased extracellular matrix synthesis. The newly synthesized matrix is further crosslinked,thus creating a feed forward cycle of increased stiffness and ECM synthesis. We propose that this mechanismcontributes to accumulation of ECM in the glomerulus and tubulointerstitium that is characteristic of diabetickidney disease. Our preliminary studies show that incubating renal tubules with glucose or ribose significantlyincreases the stiffness of the tubular basement membrane.
The aims of this application are to (1) Determine themechanism by which sugar exposure alters the stiffness of kidney tubular and glomerular extracellular matrixand (2) Determine the mechanism by which increased stiffness alters integrin signaling and leads to increasedECM production. The methods employed in aim 1 will include measurement of tubular basement membranestiffness in normal and sugar modified kidney tubules using our newly developed microcantilever-based stiffnessmeasurement technique. Glomerular stiffness will be measured using microscale compression testing.Mechanical characterization of the tissue will be correlated with biochemical analysis via mass spectrometry toevaluate crosslink formation.
For aim 2, we will use in vitro cell culture models that mimic the mechanicalproperties of normal and sugar modified tubular and glomerular ECM. Stiffness induced changes in cellphenotype, cell proliferation and ECM production will be evaluated in wild type and integrin ?1 and ?2-null tubularepithelial and glomerular mesangial cells. Successful completion of these aims will establish the role of AGE-mediated ECM stiffening as a potential contributor to progression of diabetic kidney disease and will informsubsequent studies in diabetic and integrin-null mouse models.
Diabetes is one of the most significant public health issues in the Unites States; and it is the most common causeof kidney failure requiring dialysis. Additional insight into the underlying causes of diabetic kidney disease isneeded in order to develop new strategies to treat and prevent the disease. This study will investigate a newpotential factor that may influence development and progression of diabetic kidney disease.
