The existence of a cardiac renin-angiotensin system, independent of the circulating renin-angiotensin system, is still controversial. We compared the tissue levels of reninangiotensin system components in the heart with the levels in blood plasma in healthy pigs and 30 hours after nephrectomy. Angiotensin I (Ang I)-generating activity of cardiac tissue was identified as renin by its inhibition with a specific active site-directed renin inhibitor. We took precautions to prevent the ex vivo generation and breakdown of cardiac angiotensins and made appropriate corrections for any losses of intact Ang I and II during extraction and assay. Tissue levels of renin (n=ll) and Ang I (n=7) and II (n=7) in the left and right atria were higher than in the corresponding ventricles (P< .05). Cardiac renin and Ang I levels (expressed per gram wet weight) were similar to the plasma levels, and Ang II in cardiac tissue was higher than in plasma (P<.05). The presence of these renin-angiotensin system components in cardiac tissue therefore cannot be accounted for by trapped plasma or simple diffusion from plasma into the interstitial fluid. Angiotensinogen levels (n=ll) in cardiac tissue were 10% to 25% of the A ngiotensin I (Ang I) is produced in the circulating / \ blood by the action of renin from the kidney on -Z A . angiotensinogen produced by the liver. Ang I is converted to Ang II, a potent vasoconstrictor, by angiotensin-converting enzyme (ACE) located on the luminal surface of the vascular endothelium. It is now well established that Ang I and II are not only produced in the blood compartment but also locally in tissues. Recent evidence suggests that complete local reninangiotensin systems (RAS) are present in a number of organs, for instance, kidney, adrenal gland, and ovary. 13In such local RAS, the production of Ang I and II is thought to depend on in situ synthesized renin rather than plasma-derived renin.A local cardiac RAS has also been postulated. 4 ' 5 However, direct evidence for Ang I and II production in the heart by in situ synthesized renin is still lacking. Renin mRNA levels in the heart are usually low and can be detected only by polymerase chain reaction. 68 Early studies showed Ang I-generating activity in left ventric-

Duchenne muscular dystrophy (DMD) is a severe progressive muscular disorder caused by reading frame disrupting mutations in the DMD gene, preventing the synthesis of functional dystrophin. As dystrophin provides muscle fiber stability during contractions, dystrophin negative fibers are prone to exercise-induced damage. Upon exhaustion of the regenerative capacity, fibers will be replaced by fibrotic and fat tissue resulting in a progressive loss of function eventually leading to death in the early thirties. With several promising approaches for the treatment of DMD aiming at dystrophin restoration in clinical trials, there is an increasing need to determine more precisely which dystrophin levels are sufficient to restore muscle fiber integrity, protect against muscle damage and improve muscle function.To address this we generated a new mouse model (mdx-Xist Δhs) with varying, low dystrophin levels (3–47%, mean 22.7%, stdev 12.1, n = 24) due to skewed X-inactivation. Longitudinal sections revealed that within individual fibers, some nuclei did and some did not express dystrophin, resulting in a random, mosaic pattern of dystrophin expression within fibers. Mdx-Xist Δhs, mdx and wild type females underwent a 12 week functional test regime consisting of different tests to assess muscle function at base line, or after chronic treadmill running exercise. Overall, mdx-Xist Δhs mice with 3–14% dystrophin outperformed mdx mice in the functional tests. Improved histopathology was observed in mice with 15–29% dystrophin and these levels also resulted in normalized expression of pro-inflammatory biomarker genes, while for other parameters >30% of dystrophin was needed. Chronic exercise clearly worsened pathology, which needed dystrophin levels >20% for protection. Based on these findings, we conclude that while even dystrophin levels below 15% can improve pathology and performance, levels of >20% are needed to fully protect muscle fibers from exercise-induced damage.

To study the metabolism and production of angiotensin I, highly purified monoiodinated [ synthesized mainly by the kidneys, acts on renin substrate (angiotensinogen) to form the decapeptide angiotensin I (Ang I), which is then converted to the octapeptide angiotensin II (Ang II) by angiotensin converting enzyme (ACE). All these components of the renin-angiotensin system are pres-

No abstract

The genetic defect of mdx mice resembles that of Duchenne muscular dystrophy, although their functional performance and life expectancy is nearly normal. By contrast, mice lacking utrophin and dystrophin (mdx/utrn -/-) are severely affected and die prematurely. Mice with one utrophin allele (mdx/utrn +/-) are more severely affected than mdx mice, but outlive mdx/utrn -/- mice. We subjected mdx/utrn +/+, +/-, -/- and wild type males to a 12week functional test regime of four different functional tests. Mdx/utrn +/+ and +/- mice completed the regime, while mdx/utrn -/- mice died prematurely. Mdx/utrn +/- mice performed significantly worse compared to mdx/utrn +/+ mice in functional tests. Creatine kinase levels, percentage of fibrotic/necrotic tissue, morphology of neuromuscular synapses and expression of biomarker genes were comparable, whereas mdx/utrn +/- and -/- mice had increased levels of regenerating fibers. This makes mdx/utrn +/- mice valuable for testing the benefit of potential therapies on muscle function parameters.

To estimate the contribution of angiotensin (ANG) I and II production at tissue sites to the circulating levels, ANG I and II and their radiolabeled counterparts were measured in arterial and venous plasma across various vascular beds during constant infusion of 125I-ANG I into the left cardiac ventricle of anesthetized pigs. In the combined systemic vascular beds, ANG I production was closely correlated with plasma renin activity (PRA) and ANG II production was greater than in the lungs. In the lungs virtually no ANG I but 31% of ANG II in venous plasma was derived from de novo production, which could be fully accounted for by conversion of circulating ANG I. In myocardium, head, skin, skeletal muscle, and kidney, respectively, 40, 58, 55, 67, and 94% of venous ANG I, and 32, 49, 40, 59, and 85% of venous ANG II were derived from de novo production. In these extrapulmonary beds part of de novo produced ANG I and II appeared not to be generated, respectively, by PRA and by conversion of circulating ANG I. These results indicate that production of ANG I at tissue sites contributes to its circulating level and that some circulating ANG II may not be derived from circulating ANG I.

To study regional metabolism and production of angiotensin II, we measured steady-state plasma levels of 12S I-angiotensin I and II and endogenous angiotensin I and II in the aorta and the antecubital, femoral, renal, and hepatic veins during systemic infusion of 125 I-angiotensin I or II. Extraction of arterially delivered angiotensin II ranged from 30-50% in the limbs to 80-100% in the renal and hepatomesenteric vascular beds both in essential hypertension (n=13) and in unilateral renal artery stenosis (n=7). Across the limbs, 20-30% of arterially delivered angiotensin I was converted to angiotensin II in both groups, and there was no arteriovenous gradient in endogenous angiotensin II. No conversion of arterially delivered angiotensin I was detected across the renal and hepatomesenteric beds, and there was net extraction of angiotensin II from the systemic circulation by these beds. Although regional production of angiotensin I at tissue sites made a significant contribution to its level in the veins, little of this locally produced angiotensin I reached the regional veins in the form of angiotensin II, even in the kidney with artery stenosis, where the venous levels of locally produced angiotensin I were particularly high. These results provide no evidence for a source of circulating angiotensin II other than blood-borne angiotensin I and illustrate the high degree of compartmentalization of angiotensin I and II production. sin system, circulating renin acts on circulating .Z \ -angiotensinogen to form the decapeptide angiotensin I (Ang I). During the passage of blood through the lungs, Ang I is converted to the octapeptide angiotensin II (Ang II) by angiotensin converting enzyme (ACE) that is bound to the luminal surface of the pulmonary vascular endothelium. Ang II is then transported by the bloodstream to peripheral target sites to exert its physiological actions.This view of the renin-angiotensin system, however, is an oversimplification. Observations in animals and humans have clearly demonstrated that considerable conversion of plasma Ang I occurs in organs other than the lungs,'-6 but particularly in humans, it is not known to what extent extrapulmonary conversion of Ang I to Ang II contributes to the circulating levels of Ang II.Moreover, the concept of the renin-angiotensin system as a circulating endocrine system is now being challenged.7 -10 Animal studies, in which pharmacological doses of Ang II were systemically infused, have

Our findings provide evidence for M-6-P receptor-dependent endocytosis of (pro)renin and proteolytic prorenin activation by vascular endothelial cells.