David Severs

Brain Renin–Angiotensin SystemNovelty and Significance: Does It Exist?

Because of the presence of the blood–brain barrier, brain renin–angiotensin system activity should depend on local (pro)renin synthesis. Indeed, an intracellular form of renin has been described in the brain, but whether it displays angiotensin (Ang) I–generating activity (AGA) is unknown. Here, we quantified brain (pro)renin, before and after buffer perfusion of the brain, in wild-type mice, renin knockout mice, deoxycorticosterone acetate salt–treated mice, and Ang II–infused mice. Brain regions were homogenized and incubated with excess angiotensinogen to detect AGA, before and after prorenin activation, using a renin inhibitor to correct for nonrenin-mediated AGA. Renin-dependent AGA was readily detectable in brain regions, the highest AGA being present in brain stem (>thalamus=cerebellum=striatum=midbrain>hippocampus=cortex). Brain AGA increased marginally after prorenin activation, suggesting that brain prorenin is low. Buffer perfusion reduced AGA in all brain areas by >60%. Plasma renin (per mL) was 40× to 800× higher than brain renin (per gram). Renin was undetectable in plasma and brain of renin knockout mice. Deoxycorticosterone acetate salt and Ang II suppressed plasma renin and brain renin in parallel, without upregulating brain prorenin. Finally, Ang I was undetectable in brains of spontaneously hypertensive rats, while their brain/plasma Ang II concentration ratio decreased by 80% after Ang II type 1 receptor blockade. In conclusion, brain renin levels (per gram) correspond with the amount of renin present in 1 to 20 &mgr;L of plasma. Brain renin disappears after buffer perfusion and varies in association with plasma renin. This indicates that brain renin represents trapped plasma renin. Brain Ang II represents Ang II taken up from blood rather than locally synthesized Ang II.

Salt Sensitivity of Angiogenesis Inhibition–Induced Blood Pressure Rise: Role of Interstitial Sodium Accumulation?

In response to salt loading, Na+ and Cl− accumulate in the skin in excess of water, stimulating skin lymphangiogenesis via activation of the mononuclear phagocyte system cell-derived vascular endothelial growth factor-C–vascular endothelial growth factor type 3 receptor signaling pathway. Inhibition of this pathway results in salt-sensitive hypertension. Sunitinib is an antiangiogenic, anticancer agent that blocks all 3 vascular endothelial growth factor receptors and increases blood pressure. We explored the salt dependency of sunitinib-induced hypertension and whether impairment of skin lymphangiogenesis is an underlying mechanism. Normotensive Wistar–Kyoto rats were exposed to a normal or high salt with or without sunitinib administration. Sunitinib induced a 15 mm Hg rise in telemetrically measured blood pressure, which was aggravated by a high-salt diet (HSD), resulting in a decline of the slope of the pressure–natriuresis curve. Without affecting body weight, plasma Na+ concentration or renal function, Na+ and Cl− skin content increased by 31% and 32% with the high salt and by 49% and 50% with the HSD plus sunitinib, whereas skin water increased by 17% and 24%, respectively. Skin mononuclear phagocyte system cell density increased both during sunitinib and a HSD, but no further increment was seen when HSD and sunitinib were combined. HSD increased skin lymphangiogenesis, while sunitinib tended to decrease lymphangiogenesis, both during a normal-salt diet and HSD. We conclude that sunitinib induces hypertension that is aggravated by high salt intake and not accompanied by impaired skin lymphangiogenesis.

Mycophenolate Mofetil Attenuates DOCA-Salt Hypertension: Effects on Vascular Tone.

Inflammation is increasingly recognized as a driver of hypertension. Both genetic and pharmacological inhibition of B and T cells attenuates most forms of experimental hypertension. Accordingly, the immunosuppressive drug mycophenolate mofetil (MMF) reduces blood pressure in the deoxycorticosterone acetate (DOCA-) salt model. However, the mechanisms by which MMF prevent hypertension in the DOCA-salt model remain unclear. Recent studies indicate that immunosuppression can inhibit sodium transporter activity in the kidney, but its effect on vascular tone is not well characterized. Therefore, the aim of the present study was to analyze the vascular and renal tubular effects of MMF in the DOCA-salt model in rats (4 weeks without uninephrectomy). Co-treatment with MMF attenuated the rise in blood pressure from day 11 onward resulting in a significantly lower telemetric mean arterial pressure after 4 weeks of treatment (108 ± 7 vs. 130 ± 9 mmHg, P < 0.001 by two-way analysis of variance). MMF significantly reduced the number of CD3+ cells in kidney cortex and inner medulla, but not in outer medulla. In addition, MMF significantly reduced urinary interferon-γ excretion. Vascular tone was studied ex vivo using wire myographs. An angiotensin II type 2 (AT2) receptor antagonist blocked the effects of angiotensin II (Ang II) only in the vehicle group. Conversely, L-NAME significantly increased the Ang II response only in the MMF group. An endothelin A receptor blocker prevented vasoconstriction by endothelin-1 in the MMF but not in the vehicle group. MMF did not reduce the abundances of the kidney sodium transporters NHE3, NKCC2, NCC, or ENaC. Together, our ex vivo results suggest that DOCA-salt induces AT2 receptor-mediated vasoconstriction. MMF prevents this response and increases nitric oxide availability. These data provide insight in the antihypertensive mechanism of MMF and the role of inflammation in dysregulating vascular tone.

Salt Sensitivity of Angiogenesis Inhibition–Induced Blood Pressure RiseNovelty and Significance: Role of Interstitial Sodium Accumulation?

In response to salt loading, Na+ and Cl− accumulate in the skin in excess of water, stimulating skin lymphangiogenesis via activation of the mononuclear phagocyte system cell-derived vascular endothelial growth factor-C–vascular endothelial growth factor type 3 receptor signaling pathway. Inhibition of this pathway results in salt-sensitive hypertension. Sunitinib is an antiangiogenic, anticancer agent that blocks all 3 vascular endothelial growth factor receptors and increases blood pressure. We explored the salt dependency of sunitinib-induced hypertension and whether impairment of skin lymphangiogenesis is an underlying mechanism. Normotensive Wistar–Kyoto rats were exposed to a normal or high salt with or without sunitinib administration. Sunitinib induced a 15 mm Hg rise in telemetrically measured blood pressure, which was aggravated by a high-salt diet (HSD), resulting in a decline of the slope of the pressure–natriuresis curve. Without affecting body weight, plasma Na+ concentration or renal function, Na+ and Cl− skin content increased by 31% and 32% with the high salt and by 49% and 50% with the HSD plus sunitinib, whereas skin water increased by 17% and 24%, respectively. Skin mononuclear phagocyte system cell density increased both during sunitinib and a HSD, but no further increment was seen when HSD and sunitinib were combined. HSD increased skin lymphangiogenesis, while sunitinib tended to decrease lymphangiogenesis, both during a normal-salt diet and HSD. We conclude that sunitinib induces hypertension that is aggravated by high salt intake and not accompanied by impaired skin lymphangiogenesis.

Salt Sensitivity of Angiogenesis Inhibition–Induced Blood Pressure RiseNovelty and Significance

In response to salt loading, Na+ and Cl− accumulate in the skin in excess of water, stimulating skin lymphangiogenesis via activation of the mononuclear phagocyte system cell-derived vascular endothelial growth factor-C–vascular endothelial growth factor type 3 receptor signaling pathway. Inhibition of this pathway results in salt-sensitive hypertension. Sunitinib is an antiangiogenic, anticancer agent that blocks all 3 vascular endothelial growth factor receptors and increases blood pressure. We explored the salt dependency of sunitinib-induced hypertension and whether impairment of skin lymphangiogenesis is an underlying mechanism. Normotensive Wistar–Kyoto rats were exposed to a normal or high salt with or without sunitinib administration. Sunitinib induced a 15 mm Hg rise in telemetrically measured blood pressure, which was aggravated by a high-salt diet (HSD), resulting in a decline of the slope of the pressure–natriuresis curve. Without affecting body weight, plasma Na+ concentration or renal function, Na+ and Cl− skin content increased by 31% and 32% with the high salt and by 49% and 50% with the HSD plus sunitinib, whereas skin water increased by 17% and 24%, respectively. Skin mononuclear phagocyte system cell density increased both during sunitinib and a HSD, but no further increment was seen when HSD and sunitinib were combined. HSD increased skin lymphangiogenesis, while sunitinib tended to decrease lymphangiogenesis, both during a normal-salt diet and HSD. We conclude that sunitinib induces hypertension that is aggravated by high salt intake and not accompanied by impaired skin lymphangiogenesis.

Salt Sensitivity of Angiogenesis Inhibition–Induced Blood Pressure Rise

In response to salt loading, Na+ and Cl− accumulate in the skin in excess of water, stimulating skin lymphangiogenesis via activation of the mononuclear phagocyte system cell-derived vascular endothelial growth factor-C–vascular endothelial growth factor type 3 receptor signaling pathway. Inhibition of this pathway results in salt-sensitive hypertension. Sunitinib is an antiangiogenic, anticancer agent that blocks all 3 vascular endothelial growth factor receptors and increases blood pressure. We explored the salt dependency of sunitinib-induced hypertension and whether impairment of skin lymphangiogenesis is an underlying mechanism. Normotensive Wistar–Kyoto rats were exposed to a normal or high salt with or without sunitinib administration. Sunitinib induced a 15 mm Hg rise in telemetrically measured blood pressure, which was aggravated by a high-salt diet (HSD), resulting in a decline of the slope of the pressure–natriuresis curve. Without affecting body weight, plasma Na+ concentration or renal function, Na+ and Cl− skin content increased by 31% and 32% with the high salt and by 49% and 50% with the HSD plus sunitinib, whereas skin water increased by 17% and 24%, respectively. Skin mononuclear phagocyte system cell density increased both during sunitinib and a HSD, but no further increment was seen when HSD and sunitinib were combined. HSD increased skin lymphangiogenesis, while sunitinib tended to decrease lymphangiogenesis, both during a normal-salt diet and HSD. We conclude that sunitinib induces hypertension that is aggravated by high salt intake and not accompanied by impaired skin lymphangiogenesis.

PS 16-13 ON THE ORIGIN OF BRAIN RENIN

Objective: A renin-angiotensin system (RAS) in the brain is believed to contribute to blood pressure regulation. Given the presence of the blood-brain barrier, brain RAS activity most likely depends on synthesis of (pro)renin in the brain. In support of this concept, an intracellular, non-secreted form of renin has been described in the brain, depending on an alternative transcript of the renin gene. In the present study, we set out to quantify brain (pro)renin, both before and after buffer perfusion of the brain, in normal mice, renin knockout (KO) mice, DOCA-salt-treated mice (which have been reported to display brain RAS activation), and angiotensin II-infused mice. Design and method: Brain nuclei were homogenized and incubated with angiotensinogen to detect AGA, both before and after acid activation of prorenin, with or without the renin inhibitor aliskiren to correct for non-renin-mediated AGA. Results: Renin-dependent (i.e., aliskiren-inhibitable) AGA was readily detectable in brain nuclei, the highest AGA being present in brainstem (>thalamus = cerebellum = striatum = midbrain > hippocampus = cortex). Brain AGA increased non-significantly after prorenin activation, suggesting that brain prorenin levels are low or absent. Buffer perfusion reduced AGA in all brain areas by > 60%. Plasma renin (expressed per mL plasma) was 40–800x higher than brain renin (expressed per g tissue). Plasma prorenin levels were lower than plasma renin levels. AGA was undetectable in plasma and brain of renin KO mice. DOCA-salt and angiotensin II suppressed plasma renin, and parallel decreases were observed for brain renin. Conclusions: Brain renin levels (per g tissue) correspond with the amount of renin present in 1–20 microliter blood plasma. Brain renin disappears after buffer perfusion, and varies in association with plasma renin. This indicates that renin detected in brain nuclei represents plasma renin and/or locally activated plasma prorenin. DOCA-salt exposure does not selectively increase brain renin expression.

6B.05: SALT-SENSITIVITY OF ANGIOGENESIS INHIBITION-INDUCED BLOOD PRESSURE (BP) RISE: ROLE OF INTERSTITIAL SODIUM ACCUMULATION?

Objective: Angiogenesis inhibition with the VEGF-inhibitor sunitinib, an established anti-cancer therapy, induces hypertension and proteinuria. Exposed to osmotic stress, the Mononuclear-phagocyte-system cells produces VEGF-C and exert homeostatic regulatory activity by promoting lymphatic Na+ drainage; interference with this process resulted in salt-sensitive hypertension. Therefore, we hypothesized that sunitinib via blockade of the VEGF pathway leads to Na+ accumulation in the skin and salt-sensitive hypertension. Design and method: In male WKY rats, mean arterial pressure (MAP) was monitored telemetrically during oral treatment with sunitinib (7 mg/kg.day, n = 4–8) or vehicle (n = 4–8) after a normal salt diet (NSD: 0.5–1.0% NaCl and tap water) or a high salt diet (HSD: 8% NaCl and saline water) for 2 weeks. After 8 days of sunitinib or vehicle administration, 24-h urine was collected. After sacrificing, blood was collected for biochemical measurements and skin for Na+ concentration ([Na+]) using dry-ashing. Results: MAP during NSD was 101 ± 0.9 mmHg. HSD increased MAP by 27 ± 3 mmHg (P < 0.05 vs. NSD). Sunitinib increased MAP by 16 ± 1 mmHg during NSD (P < 0.05 vs. NSD alone) and by 23 ± 4 mmHg during HSD (P < 0.05 vs. HSD alone). Although body weight, serum [Na+] and plasma [cystatin-C] did not change in response to HSD and/or sunitinib, skin [Na+] increased from 89 ± 1 (NSD) to 92 ± 3 (HSD), and 97 ± 3 mmol/L (HSD+sunitinib), respectively (P < 0.03 for linear trend). Plasma endothelin-1 (ET-1) increased from 0.4 ± 0.09 (NSD) to 0.8 ± 0.05 pg/mL during HSD, and remained elevated with sunitinib. Skin [Na+] correlated both with MAP (r = 0.76, P < 0.01) and plasma ET-1 (r = 0.53, P < 0.05). Compared to NSD, proteinuria and endothelinuria increased during HSD, rising further (P < 0.05) with sunitinib. Conclusions: Angiogenesis inhibition-induced hypertension is salt-sensitive. The parallel increases in BP and skin [Na+], in the face of unaltered serum [Na+] and body weight, support the existence of a Na+-buffering compartment in the skin that may contribute to the salt-dependent volume and BP homeostasis during VEGF inhibition. Our data indicate that ET-1 may play a causal role in this phenomenon.