acase@tulane.edu / Increased activity of the intrarenal renin-angiotensin system (RAS), in which proximal tubular angiotensinogen (AGT) is a key factor, has been implicated in the progression of diabetic nephropathy. AGT expression is upregulated in renal proximal tubular cells (PTC) by high glucose due to elevated reactive oxygen species (ROS) generation. Angiotensin II (Ang II) also promotes ROS generation. Glucose reabsorption in PTC occurs mainly through sodium-glucose co-transporter 2 (SGLT2). This study was performed to demonstrate that SGLT2 mediates AGT augmentation in PTC under hyperglycemic conditions. Furthermore, the enhancing effect of Ang II was investigated. Established mouse PTC were treated with 5 (normal), 15, or 25 mM D-glucose or D-mannitol (osmotic control). Pyruvate was used to investigate the role of glycolysis on AGT regulation. Glycolytic activity was quantified using a Seahorse metabolic analyzer. Tempol, an antioxidant, was used to determine the role of ROS. SGLT2 expression was silenced using shRNA. PTC were treated with high glucose and 10-10-10-7 M Ang II or an Ang II receptor blocker. AGT protein levels were increased by 15 (4.4 ± 0.2-fold over control) and 25 mM (4.6 ± 0.2-fold) glucose. AGT mRNA was also augmented (31.1 ± 3.5-fold) by 25 mM glucose, but not mannitol. AGT expression was stimulated by pyruvate (10.7 ± 1.0-fold over control), and exposure to 10, 15, or 25 mM glucose increased glycolytic activity (3.10 ± 0.28-fold, 2.74 ± 0.20-fold, and 2.75 ± 0.34-fold, respectively), suggesting that enhanced glycolysis stimulates AGT expression. ROS accumulation increased (3.03 ± 0.29-fold) in 25 mM glucose over control. Tempol attenuated glucose-induced AGT augmentation by 77%, suggesting that ROS generation contributes to AGT upregulation. SGLT2 knockdown prevented AGT augmentation in 15 mM glucose, indicating that SGLT2 plays a key role mediating AGT upregulation by high glucose. Ang II receptor blockade did not alter AGT levels, and Ang II did not enhance AGT expression in normal or high glucose. Similarly, SGLT2 expression was unchanged by glucose or Ang II. These results indicate that SGLT2 contributes to AGT upregulation in PTC by high glucose, which helps to explain the mechanisms causing intrarenal RAS activation and consequent diabetic nephropathy. / 1 / Michael W. Cypress

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

Thesis (Ph. D.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 164-201). Also available in print.

The role of the hypothalamic paraventricular nucleus (PVN) in cardiovascular and neuroendocrine regulation has been well documented. Much remains unknown however about the integration of synaptic signals within this nucleus and the neuronal subtypes and chemical messengers governing these processes. Angiotensin II (ANG) has been demonstrated to act as a neurotransmitter in PVN where it exerts considerable influence on neuronal excitability. The studies within this thesis were undertaken to delineate the actions of ANG on the membrane properties of PVN neurons and its effect on synaptic transmission within this nucleus. We report that ANG activates a nitric oxide mediated negative feedback loop in the PVN. The magnitude of the depolarizing response to ANG appears to be dependant on this GABAergic inhibitory input demonstrating there exists within PVN an intrinsic negative feedback loop which modulates neuronal excitability in response to peptidergic excitation. We also demonstrate that the depolarizing response to ANG in magnocellular neurons is in part dependent upon increases in glutamatergic excitatory synaptic input. These data in combination highlight the multiple levels of synaptic integration controlling the output of magnocellular neurons in PVN. PVN also contains significant populations of neurosecretory parvocellular neurons which exercise considerable influence over the adenohypophysis and therefore neuroendocrine regulation. ANG caused an AT1 receptor mediated depolarization of these neurosecretory neurons. Voltage-clamp analysis revealed that ANG activated a non-selective cationic current and reduced a sustained potassium current characteristic of IK. These studies identify multiple post-synaptic modulatory sites through which ANG can influence the excitability of neurosecretory parvocellular PVN neurons. The findings in this thesis provide the framework for a cellular model of action of ANG within PVN to regulate the activity of this nucleus not only through direct cellular mediated ion channel interactions but also through modulation of synaptic input within the magnocellular system of PVN. / Thesis (Ph.D, Physiology) -- Queen's University, 2008-01-18 14:10:22.319

Thesis (Ph. D.)--University of Missouri--Columbia, 1996. / "December 1996" Typescript. Vita. Includes bibliographical references (l. 121-128). Also available on the Internet.

Abstract not available

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2006. / Härtill 4 uppsatser.

Abstract not available

Long-term stimuli of many systems leads to decreased cellular responsiveness, or desensitization. We characterized the desensitization of angiotensin II (Ang 11)-mediated inositol phospholipid (IP) hydrolysis in cultured human aortic smooth muscle cells (HASMC). Although it has been suggested that the desensitization induced by long-term Mg II exposure may result partially from down-regulation of Ang II receptor, this is not sufficient to explain fully desensitization in many systems. Post-receptor desensitization of IP hydrolysis may also result from phosphorylation or changes in protein levels of the effector enzyme, PLC-β. We identified the major PLC-β isoenzymes expressed by HASMC as PLC-β1 and PLC-β3. Ang II pretreatment reduced IP accumulation induced by Ang II (1μM) in a time-dependent manner. Phorbol ester-12-myristrate-13-acetate (PMA), a protein kinase C (PKC) activator, also reduced Ang II-stimulated IP accumulation. These results suggest that PKC activation may negatively regulate Ang II-stimulated IP signaling in HASMC, similar to rat cells. In addition, PKC also reduced IP accumulation stimulated by A1F₄⁻, directly activating the G protein. It suggests that the majority of PKC-induced desensitization of Ang II-stimulated IP signaling occurs downstream of the Ang II receptor in HASMC. However, both PLC-β1 and PLC-β3, expected candidates for PKC phosphorylation, were phosphorylated independently of PKC activation or inhibition, indicating that PKC might not be involved in direct phosphorylation of PLC-β1 and PLC-β3. Furthermore, PLC-β1, but not PLC-β3, was highly phosphorylated under basal conditions, suggesting that PLC-β1 and PLC-β3 may play different roles in IP signaling in HASMC. / Graduation date: 2003

Thesis (Ph.D.)--Michigan State University. Dept. of Pharmacology and Toxicology, 2008. / Title from PDF t.p. (viewed on Mar. 30, 2009) Includes bibliographical references (p.177-201). Also issued in print.