Tissue specific dysregulated protein subnetworks in type 2 diabetic bladder urothelium and detrusor muscle.

Jan. 8, 2015

Diabetes mellitus is well known to cause bladder dysfunction; however, the molecular mechanisms governing this process and the effects on individual tissue elements within the bladder are poorly understood, particularly in type 2 diabetes. A shotgun proteomics approach was applied to identify proteins differentially expressed between type 2 diabetic (TallyHo) and control (SWR/J) mice in the bladder smooth muscle and urothelium, separately. We were able to identify 1760 non-redundant proteins from the detrusor smooth muscle and 3169 non-redundant proteins from urothelium. Pathway and network analysis of significantly dysregulated proteins was conducted to investigate the molecular processes associated with diabetes. This pinpointed ERK1/2 signaling as a key regulatory node in the diabetes-induced pathophysiology for both tissue types. The detrusor muscle samples showed diabetes-induced increased tissue remodeling-type events such as Actin Cytoskeleton Signaling and Signaling by Rho Family GTPases. The diabetic urothelium samples exhibited oxidative stress responses, as seen in the suppression of protein expression for key players in the NRF2-Mediated Oxidative Stress Response pathway. These results suggest that diabetes induced elevated inflammatory responses, oxidative stress, and tissue remodeling are involved in the development of tissue specific diabetic bladder dysfunctions. Validation of signaling dysregulation as a function of diabetes was performed using Western blotting. These data illustrated changes in ERK1/2 phosphorylation as a function of diabetes, with significant decreases in diabetes-associated phosphorylation in urothelium, but the opposite effect in detrusor muscle. These data highlight the importance of understanding tissue specific effects of disease process in understanding pathophysiology in complex disease and pave the way for future studies to better understand important molecular targets in reversing bladder dysfunction.

Figure: A label-free proteomics approach was applied to monitor changes in the soluble proteomes of urothelium and detrusor muscle of diabetic mice (TallyHo) compared to their age-matched controls (SWR/J). Four biological replicates for each tissue type and each condition, control vs. diabetic, were prepared and analyzed simultaneously. The detrusor muscle and urothelium samples were separated on a reverse phase column using a 4 h gradient. Within each tissue type, the chromatograms for the diabetic and control conditions look similar to each other. However, a clear distinction is evident in comparison of the chromatographic separations of the urothelium samples with those of the detrusor muscle. The most dramatic differences between the tissue types are shown by the black boxes in the bottom chromatogram, showing broad peaks in the region of 94-118 min and a series of low intensity broad peaks at 134-180 min in the diabetic and control detrusor muscle chromatograms, compared with sharper and higher intensity peaks in those regions in the urothelium chromatograms.

Results from: Tomechko SE, Liu G, Tao M, Schlatzer D, Powell CT, Gupta S, Chance MR, Daneshgari F. Mol Cell Proteomics. 2015 Jan 8. pii: mcp.M114.041863. [Epub ahead of print]