Project

Discipline

Steroids; Kidney ; VEGF; Aldosterone; VEGF receptors; CYP11B1; Diabetic nephropathy; Proliferation

Lead
LeadBisher wurde angenommen, dass ein bestimmtes Steroidhormon der Nebenniere (Aldosteron) ausschliesslich über das Renin-Angiotensin II System reguliert wird. Ich konnte zeigen, dass ein gefässstimulierender Faktor (VEGF) ebenfalls Aldosteron zu stimulieren vermag. Da bei einigen Nierenerkrankungen erhöhte Werte von Aldosteron und VEGF im Blut gefunden werden, wird diese Studie deren Zusammenspiel und die Auswirkungen auf die Pathogenese bei Nierenerkrankungen untersuchen.
Lay summary
Ziele des Forschungsprojekts Aldosteron ist ein Hormon, welches Natrium und Wasser im Körper zurückhält. Bislang wurde angenommen, dass es im Wesentlichen durch Renin und Angiotensin II stimuliert wird. Es erhöht Blutdruck, schädigt Herz, Gefässe und Nieren. Neben den Nebennieren bilden u.a. das Herz, das Gehirn, die Lunge, die Leber und die Nieren selbst Aldosteron. Unklar ist, ob auch Gefässe Aldosteron bilden können. Entgegen einiger Literatur konnte ich aber zeigen, dass Aldosteron weder in Gefässen, noch in der Niere selber gebildet werden kann. Bei vielen Nierenerkrankungen werden erhöhte Mengen eines gefässstimulierenden Faktors (VEGF) gefunden. Ich konnte zeigen, dass VEGF unabhängig von Renin die Aldosteronproduktion in Nebennierenzellen stimulieren kann. Das Ziel dieses Projekts war es, zu evaluieren, welche Rolle die Interaktion von VEGF und Aldosteron bei ausgewählten Nierenerkrankungen („diabetische Nephropathie“ und „Mesangioproliferative Glomerulonephritis“) spielt. In diesem Projekt konnte gezeigt werden, dass Diabetiker über ein anderes hormonelles Ausscheidungsprofil verfügen als Nicht-Diabetiker und dass verschiedene Hormone Nierenprobleme bei Diabetikern fördern können. Ein erhöhter Blutzuckerspiegel, wie er bei Diabetikern vorkommt, hat einen direkten negativen Effekt auf die Nierengefässe. Dies ist ebenfalls eine neue Erkenntnis dieser Studie.  Kontext und Bedeutung des Projekts Resultate aus dieser Studie sollten zu einem besseren Verständnis der Pathogenese von Nierenerkrankungen führen und zielgerichtete Therapiemöglichkeiten aufzeigen. Key words aldosterone, VEGF(vascular endothelial growth factor), VEGF receptors, mineralocorticoid receptor,  diabetic nephropathy, steroids, proliferation

Direct link to Lay Summary

Last update: 11.07.2016

Active participation

Role of Renin-independent Extra-adrenal Aldosterone Production in Kidney Disease Aldosterone is a major steroid hormone maintaining plasma volume and blood pressure. Besides its action on renal tubules to promote Na+ and water retention, other less beneficial effects such as enhanced tissue fibrosis have been observed in numerous organs including the kidney. The main production site of aldosterone are the adrenal glands which can be stimulated by the renin angiotensin system (RAS). Aldosterone production has also been demonstrated in a variety of other organs including brain, heart, peripheral vessels, lung, liver and kidneys. In renal inflammatory disease, enhanced aldosterone expression has been observed in mesangial cells. Effects of aldosterone on these cells are so far controversial. Aldosterone is an important player in pregnancy. In normal pregnant women serum aldosterone levels increase throughout pregnancy and go back to normal levels after delivery. In preeclampsia (PE), a life-threatening disease in pregnancy characterized by arterial hypertension and proteinuria, aldosterone levels are inappropriately low. The current pathomechanism consist of disturbed angiogenic signaling and endothelial integrity. sFlt-1, a splice variant of the vasuclar endothelial growth factor receptor 1 (VEGFR1) is increased in preeclamptic pregnancies and effectively traps VEGF. As a consequence, there is a decreased VEGF availability for the vessel maintenance in preeclamptic pregnancies which results in glomerular endotheliosis and podocyte injury. My recent research field focussed on the VEGF-aldosterone problematic in pregnancy. I discovered that angiogenic (vascular endothelial growth factor (VEGF)) and anti-angiogenic (soluble fms-like tyrosine kinase (sFlt-1)) factors play a crucial role in the stimulation of adrenal aldosterone production and that this stimulation is renin-independent. We and others could show that reduced VEGF signaling leads to morphological changes in the adrenal gland of pregnant rats. The zone glomerulosa of rats over-expressing sFlt-1 showed reduced capillary density and capillary fenestrations. These changes go together with a reduced Kdr (VEGF-receptor 2) expression. I could further show that VEGF directly stimulates adrenal aldosterone production. VEGF is able to increase the mRNA expression of the aldosterone synthase (CYP11B2) as well as its protein expression directly. Increased Aldosterone levels measured by ELISA and functional assays showing increased CYP11B2 activity after VEGF stimulation confirm these results. Co-culture experiments with primary human umbilical vein endothelial cells (HUVECs) and adrenal cells (H295R) showed also an increase in aldosterone production. While testing aldosterone synthesis in my negative control, HUVECs, I could surprisingly detect a prominent aldosterone production in these cells. Repetitive independent experiments with HUVECs showed a significant aldosterone production on protein level. Unfortunately I could not detect any mRNA of CYP11B2 so far. But the activity of CYP11B2 can be stimulated to almost 40% with corticosterone, a precursor for aldosterone. The HUVEC model might be extended to other vascular beds such as to glomerular endothelial cells. I am convinced that besides the already known extra-adrenal aldosterone production sites, other organs or organ systems are able to produce relevant amounts of aldosterone as well. Clinically, it has been observed that directly blocking the aldosterone receptor (MR) has beneficial effects on organ preservation other than just inhibiting the RAS. This leads to the assumption that other mechanisms than the RAS favor aldosterone production. This supports our findings where VEGF stimulated adrenal aldosterone production renin-independently.While high aldosterone availability appears to be beneficial in pregnancy, the opposite seems to be true in renal disease. Since VEGF expression is enhanced in many renal diseases this provides the potential for an unfavorable interplay between aldosterone and VEGF. Placental growth factor (PlGF) is mainly produced in the placenta and is a VEGF homologue. I and others could show that there is a strong positive correlation of PlGF and aldosterone in pregnant women. In contrast to VEGF, PlGF is unable to promote adrenal aldosterone production by itself (manuscript in preparation). Aldosterone has been described to promote PlGF production in peripheral vessels via a mineralocorticoid responsive element in the PlGF gene. This regulation has been described in an atherosclerotic animal model due to aldosterone excess.Given these findings, extra-adrenal renal aldosterone production might be regulated via the RAS, via VEGF itself, via PlGF enhancing a given VEGF availability such as via an interaction with the glomerular endothelial layer or via yet unrecognized factors. Furthermore, substrate availability for aldosterone production might be crucial since HUVECs produce aldosterone only in the presence of Corticosterone but not Progesterone or Deoxycorticosterone. My own data show that VEGF does not regualte adrenal aldosterone production over the steroidogenic acute regulatory protein (StAR) such as ANG.II (RAS) does. I hypothesize, first, that an extraadrenal renal aldosterone production takes place in renal disease; second, that renal glomerular aldosterone production depends on its VEGF availability and is regulated independent of the RAS; and third, that pro-angiogenic factors such as PlGF or the endothelium itself might stimulate excessive renal aldosterone production. The aim of the study is to assess and compare the aldosterone production in HUVECs with systemic human endothelial cells, human glomerular endothelial and mesangial cells by measuring CYP11B2 mRNA, CYP11B2 protein, CYP11B2 activity, localization of CYP11B2 in mitochondria and substrate specificities in response to stimulation with either Ang II or VEGF. Next, I will assess specifically the expression of VEGF, PlGF and their receptors by aldosterone in these cells either alone or in co-culture conditions to identify potential forward feedback loops. Finally, I will correlate the expression of CYP11B2, VEGF, PlGF and VEGF receptors (VEGFR1, Kdr, VEGFR3, Neuropillin-1 and 2) by immunohistochemistry and laser capture microdissection analyses of kidney biopsies from patients with inflammatory glomerular diseases such as mesangioproliferative glomerulonephritis but also diabetic nephropathy, using unaffected regions of kidneys removed for malignant tumors as controls. The current investigation will open a new field in the role of aldosterone and VEGF in kidney diseases such as diabetic nephropathy, mesangioproliferative glomerulonephritis or premature arteriosclerosis of the kidney.