Marie Luise Gross

Elevated systemic TGF-β impairs aortic vasomotor function through activation of NADPH oxidase-driven superoxide production and leads to hypertension, myocardial remodeling, and increased plaque formation in apoE−/− mice

The role of circulating, systemic TGF-beta levels in endothelial function is not clear. TGF-beta(1) may cause endothelial dysfunction in apolipoprotein E-deficient (apoE(-/-)) mice via stimulation of reactive oxygen species (ROS) production by the NADPH oxidase (NOX) system and aggravate aortic and heart remodeling and hypertension. Thoracic aorta (TA) were isolated from 4-mo-old control (C57Bl/6), apoE(-/-), TGF-beta(1)-overexpressing (TGFbeta(1)), and crossbred apoE(-/-) x TGFbeta(1) mice. Endothelium-dependent relaxation was measured before and after incubation with apocynin (NOX inhibitor) or superoxide dismutase (SOD; ROS scavenger). Superoxide production within the vessel wall was determined by dihydroethidine staining under confocal microscope. In 8-mo-old mice, aortic and myocardial morphometric changes, plaque formation by en face fat staining, and blood pressure were determined. Serum TGF-beta(1) levels (ELISA) were elevated in TGFbeta(1) mice without downregulation of TGF-beta-I receptor (immunohistochemistry). In the aortic wall, superoxide production was enhanced and NO-dependent relaxation diminished in apoE(-/-) x TGFbeta(1) mice but improved significantly after apocynin or SOD. Myocardial capillary density was reduced, fibrocyte density increased, aortic wall was thicker, combined lesion area was greater, and blood pressure was higher in the apoE(-/-) x TGFbeta vs. C57Bl/6 mice. Our results demonstrate that elevated circulating TGF-beta(1) causes endothelial dysfunction through NOX activation-induced oxidative stress, accelerating atherosclerosis and hypertension in apoE(-/-) mice. These findings may provide a mechanism explaining accelerated atherosclerosis in patients with elevated plasma TGFbeta(1).

Renal toxicity mediated by glucose degradation products in a rat model of advanced renal failure

Background  In peritoneal dialysis (PD) residual renal function contributes to improved patient survival and quality of life. Glucose degradation products (GDP) generated by heat sterilization of PD fluids do not only impair the peritoneal membrane, but also appear in the systemic circulation with the potential for organ toxicity. Here we show that in a rat model of advanced renal failure, GDP affect the structure and function of the remnant kidney.

RAGE expression in the human peritoneal membrane

BACKGROUND Experimental animal models have demonstrated that the interaction of advanced glycation end-products (AGE) with their receptor RAGE is, at least in part, responsible for peritoneal damage. This study investigates the in vivo expression of RAGE in the peritoneal membrane of uraemic human patients. METHODS Peritoneal biopsies of 89 subjects (48 uraemic and 41 healthy age-matched patients) were examined. The expression of CD3, IL-6, activated NFkappaBp65, VEGF, transforming growth factor (TGF)-beta1, smooth-muscle actin (SMA), methylglyoxal (MGO) and RAGE was analysed immunohistochemically. Additionally, in 4 of the 48 uraemic patients, peritoneal biopsies were repeated after 15 months at the time of catheter removal to analyse the above parameters and the extent of NFkappaB-binding activity determined by electrophoretic mobility shift assay (EMSA) in the long-term follow-up. RESULTS In comparison to the healthy controls, uraemic patients showed a significant increase in fibrosis, angiogenesis, submesothelial thickness, MGO-derived protein adducts, RAGE, IL-6, VEGF, TGF-beta1, SMA and NFkappaBp65 in their peritonea. Four patients, followed up longitudinally from peritoneal dialysis (PD) catheter insertion to removal, demonstrated further significant increase in the above parameters, particularly in RAGE expression and NFkappaB activation. CONCLUSIONS Along with a higher expression of several indicators for inflammation, angiogenesis, fibrosis and AGE accumulation, the peritoneal membrane of the uraemic patients showed an increased submesothelial thickness and a marked induction of RAGE expression and NFkappaB-binding activity, which both further increased after PD treatment. These findings in human peritoneum support the concept of the AGE-RAGE interaction being crucial in peritoneal damage due to uraemia and PD.

The Combination of ACE Inhibition plus Sympathetic Denervation Is Superior to ACE Inhibitor Monotherapy in the Rat Renal Ablation Model

Background: The blood pressure-independent renoprotective actions of the blockade of the renin-angiotensin and the sympathetic nervous system are well documented, but monotherapies fail to completely abrogate progression. We investigated whether combined inhibition of the two systems provides additive renoprotection. Methods: After subtotal nephrectomy (SNX) or sham operation, rats underwent resection of dorsal roots, i.e. rhizotomy or sham rhizotomy. Subsequently, they received tap water or quinapril in drinking water for 16 weeks (n = 18/group). Albuminuria, blood pressure and kidneys were assessed (morphometry, immunohistochemistry). Results: At the end of the study telemetric blood pressure in SNX was 118 ± 16 mm Hg, in SNX + rhizotomy 110 ± 10 mm Hg, in SNX + quinapril 103 ± 9 mm Hg and in SNX + quinapril + rhizotomy 95 ± 7 mm Hg. Albuminuria in the respective groups was 169 ± 75, 86 ± 45, 15 ± 23 and 5 ± 4 mg/24 h. The glomerulosclerosis index was 1.40 ± 0.6, 0.80 ± 0.23, 0.37 ± 0.16 and 0.31 ± 0.15 (p < 0.001). Only combined intervention caused significant reduction of the glomerular volume and podocyte hypertrophy. The lowest indices for nitrotyrosine, NOS-1 (nNOS), TGF-β and interstitial collagen were seen with combined interventions (p < 0.05). Conclusion: In angiotensin-converting enzyme inhibitor-treated SNX animals, abrogation of sympathetic overactivity provides additional renoprotection and less nitro-oxidative stress of podocytes than single interventions. The added benefits were partially blood pressure independent.

Selective phosphodiesterase-5 inhibition reduces neointimal hyperplasia in rat carotid arteries after surgical endarterectomy

OBJECTIVE Long-term results of surgical vessel reconstruction are compromised by restenosis caused by neointimal hyperplasia. Recent studies suggest that reduced cyclic guanosine monophosphate signaling is associated with neointima formation. In a rat model of endarterectomy, we investigated the effect of pharmacologic inhibition of cyclic guanosine monophosphate degradation on neointima formation by using the selective phosphodiesterase-5 inhibitor vardenafil. METHODS Carotid endarterectomy was performed in male Sprague-Dawley rats by means of incision of the right common carotid artery with removal of intima. Four groups were studied: unoperated control rats (n = 4), sham-operated rats (n = 9), control rats with endarterectomy (n = 9), or endarterectomized rats treated with vardenafil (10 mg/kg/day) postoperatively (n = 9). After 3 weeks, vessel compartment areas were measured by means of conventional microscopy with hematoxylin and eosin staining. Immunohistochemical analysis was performed to confirm neointima formation and the local cyclic guanosine monophosphate content. Plasma levels of cyclic guanosine monophosphate were determined by means of enzyme immunoassay. Students t test was used for statistical evaluation. RESULTS Immunohistochemical analysis demonstrated intensive staining for transforming growth factor beta1 and alpha-smooth muscle actin in the control neointima. Vardenafil significantly reduced the stenosis grade (24.64% +/- 7.46% vs 54.12% +/- 10.30% in the control group, P < .05) and expression of transforming growth factor beta1, as well as alpha-smooth muscle actin, in the neointima. The immunohistochemical score for cyclic guanosine monophosphate was higher in the treated neointima (4.80 +/- 0.76 vs 2.84 +/- 0.40 in the control group, P < .05), and increased plasma cyclic guanosine monophosphate levels were found by means of enzyme immunoassay as well (84.65 +/- 12.77 pmol/mL vs 43.50 +/- 3.30 pmol/mL in the control group, P < .05). CONCLUSIONS Treatment with vardenafil can be considered a new possibility to prevent neointimal hyperplasia after endarterectomy.

Increased renoprotection with ACE inhibitor plus aldosterone antagonist as compared to monotherapies—the effect on podocytes

BACKGROUND Blockade of the renin-angiotensin-aldosterone system (RAAS) does not completely prevent progression of renal disease. Mineralocorticoid receptor blockade provides additional renoprotection over ACE-inhibition monotherapy. We examined the mechanisms underlying superior renoprotection in the subtotal nephrectomy (SNX) model. METHODS Sprague-Dawley rats were randomized into six groups: (1) sham-op, (2) SNX without treatment, (3) SNX + quinapril (Q), (4) SNX + spironolactone (S), (5) SNX + combination therapy (Q+S), (6) SNX + combination hydrochlorothiazide + reserpin + hydralazine (HRH). Albuminuria and blood pressure were monitored, and kidneys were examined by morphometric and molecular methods. RESULTS In SNX rats, albumin excretion was significantly higher than in sham-op rats. Blood pressure reduction was not significantly different between the treatment groups. All therapies (S, Q, Q+S and HRH) reduced albuminuria; the values were lowest in animals treated with Q+S. The volume density of glomerular matrix and the number of mesangial cells were significantly increased in SNX and were lowest in SNX treated with Q+S. The number of podocytes was reduced in SNX, but was normalized in SNX treated with Q+S. Glomerular volumes and podocyte volumes were significantly higher in SNX than in sham-op. Both volumes were reduced by all interventions, but almost normalized by treatment with Q+S. Expression of collagen IV, TGF-beta(1) and desmin was increased after SNX and significantly reduced by treatment with Q and Q+S. CONCLUSIONS In subtotally nephrectomized rats, mineralocorticoid blockade provided additional renoprotection over and above ACE inhibition. Such benefit was paralleled by major changes in podocyte number and morphology and was not blood pressure dependent.

Does Borderline Kidney Allograft Rejection Always Require Treatment

Background. Borderline rejection (Bord-R) is a frequent diagnosis in renal transplantation, and there is increasing evidence that regulatory T lymphocytes are involved in its pathogenesis. Current histopathologic practice does not differentiate between graft-protecting and -damaging T lymphocytes, and patients with Bord-R routinely receive rejection treatment. We analyzed Treg-associated forkhead box P3 (Foxp3) gene expression in Bord-R and more severe forms of acute rejection episodes (ARE). Methods. Foxp3 transcripts were measured in 520 serial peripheral blood samples from 177 kidney graft recipients obtained during the first 20 days posttransplantation. Results. The highest Foxp3 transcripts were observed in patients with Bord-R or without rejection and the lowest in patients with ARE. Patients with Bord-R on posttransplant days 5 to 7 showed an increased Foxp3 transcript level of 156%, which increased to 302% by posttransplant days 14 to 16. In contrast, patients with ARE demonstrated significantly lower Foxp3 gene expression than that observed in Bord-R, nonrejectors, or acute tubular necrosis patients (P=0.001, P<0.001, and P=0.005, respectively, on days 11–13). Acute tubular necrosis patients demonstrated intermediately high Foxp3 gene expression. Conclusions. Our data indicate that increased Treg activity in peripheral blood is a frequent feature of Bord-R. This finding questions the appropriateness of rejection treatment in all patients with the histopathologic diagnosis “Bord-R”.

Renal failure and ACE inhibition: how much is too much?

Die Dosisantwortbeziehung zwischen pharmakologischer Blockade des Renin-Angiotensinsystems (RAS) (ACE-Hemmer oder Angiotensin-Rezeptorblocker) und Angiotensinkonzentration in der Zirkulation sowie Blutdrucksenkung ist gut dokumentiert. Hingegen ist im Moment unklar, welche Dosis von ACE-Hemmern rsp Angiotensinrezeptorblockern für die Nephroprotektion maximalen Nutzen bringt. Die Beurteilung dieser Situation wird erschwert durch die Tatsache, dass die Hemmung des RAS zu einem Filtratabfall (und damit akut zu reduzierter Nierenleistung) führt und nicht nebenwirkungsfrei ist (Natriumverlust, Anämie, Hyperkaliämie). Tierexperimentelle Befunde und neuere klinische Beobachtungen belegen, dass zur maximalen Reduktion der Proteinurie (als Surogatmarker der Nierenschädigung) und zur optimalen Nephroprotektion (verzögerter renaler Filtratverlust) höhere Dosen von ACE-Hemmern rsp. Angiotensin-Rezeptorblockern notwendig sind als zur maximalen Blutdruckhemmung. Details der optimalen Behandlungsstrategie (Dosissteigerung der Monosubstanzen rsp. Kombination von ACE-Hemmern und Angiotensin-Rezeptorblockern) sind gegenwärtig Gegenstand klinischer Untersuchungen. The dose-response relationship between pharmacological blockade of the renin-angiotensin system (RAS) and angiotensin II concentration in the circulation, on the one hand, and decrease of blood pressure, on the other hand, has been well established. In contrast it is currently unclear which dose of ACE inhibitors and/or angiotensin receptor blockers is optimal for nephroprotection. Clinical studies are rendered quite complex by an early decrease of glomerular filtration after RAS blockade and by side effects at higher doses such as renal sodium loss, hyperkalemia, anemia, etc. Animal experiments and recent clinical studies suggest that the doses of ACE inhibitors or angiotensin receptor blockers required for maximal reduction of proteinuria (as a surrogate marker) and for optimal nephroprotection (retardation of the loss of glomerular filtration) exceed those required for maximal lowering of blood pressure. Ongoing studies try to define the relative merits of high dose monotherapy (ACE inhibitors or angiotensin receptor blockers) versus a combination therapy of the two.