Micro-mechanical fingerprints of the rat bladder change in actinic cystitis and tumor presence.

Tissue mechanics determines tissue homeostasis, disease development and progression. Bladder strongly relies on its mechanical properties to perform its physiological function, but these are poorly unveiled under normal and pathological conditions. Here we characterize the mechanical fingerprints at the micro-scale level of the three tissue layers which compose the healthy bladder wall, and identify modifications associated with the onset and progression of pathological conditions (i.e., actinic cystitis and bladder cancer). We use two indentation-based instruments (an Atomic Force Microscope and a nanoindenter) and compare the micromechanical maps with a comprehensive histological analysis. We find that the healthy bladder wall is a mechanically inhomogeneous tissue, with a gradient of increasing stiffness from the urothelium to the lamina propria, which gradually decreases when reaching the muscle outer layer. Stiffening in fibrotic tissues correlate with increased deposition of dense extracellular matrix in the lamina propria. An increase in tissue compliance is observed before the onset and invasion of the tumor. By providing high resolution micromechanical investigation of each tissue layer of the bladder, we depict the intrinsic mechanical heterogeneity of the layers of a healthy bladder as compared with the mechanical properties alterations associated with either actinic cystitis or bladder tumor.

Negative regulation of diacylglycerol kinase theta mediates adenosine-dependent hepatocyte preconditioning.

In liver ischemic preconditioning (IP), stimulation of adenosine A2a receptors (A2aR) prevents ischemia/reperfusion injury by promoting diacylglycerol-mediated activation of protein kinase C (PKC). By concerting diacylglycerol to phosphatidic acid, diacylglycerol kinases (DGKs) act as terminator of diacylglycerol signalling. This study investigates the role of DGK in the development of hepatocyte IP. DGK activity and cell viability were evaluated in isolated rat hepatocytes preconditioned by 10 min hypoxia followed by 10 min re-oxygenation or by the treatment with the A2aR agonist, CGS21680, and subsequently exposed to prolonged hypoxia. We observed that after IP or A2aR activation, a decrease in DGK activity was associated with the onset of hepatocyte tolerance to hypoxia. CGS21680-induced stimulation of A2aR specifically inhibited DGK isoform theta by activating RhoA-GTPase. Consistently, both siRNA-mediated downregulation of DGK theta and hepatocyte pretreatment with the DGK inhibitor R59949 induced cell tolerance to hypoxia. The pharmacological inhibition of DGK was associated with the diacylglycerol-dependent activation of PKC delta and epsilon and of their downstream target p38 MAPK. In conclusion, we unveil a novel signalling pathway contributing to the onset of hepatocyte preconditioning, which through RhoA-GTPase, couples A2aR to the downregulation of DGK. Such an inhibition is essential for the sustained accumulation of diacylglycerol required for triggering PKC-mediated survival signals.