Ingrid M. Garrelds

Vascular aging is marked by increased levels of reactive oxygen species in endothelial cells. Reactive oxygen species can, among others, be produced by dysfunctional mitochondria, contributing to acceleration of vascular aging by promoting DNA damage response and senescence. In the aged vasculature, impaired endothelial cell function causes decreased vasodilation, which may also have an impact on peripheral organs such as the kidney. The aim of this study was to investigate the effect of chronic treatment with SUL-138 (30 mg/kg/day), a novel mitochondrial protective compound, on DNA damage-prompted, accelerated endothelial aging and associated kidney dysfunction in mice. Endothelial-specific aging was induced by knock-out (KO) of DNA repair endonuclease Ercc1 in mice [endothelial-cell specific Ercc1 KO (EC-KO) mice]. We showed that impaired endothelium-dependent vasodilation and expression of DNA damage response markers in EC-KO mice were restored after the treatment with SUL-138. The underlying mechanism of improved vasodilation was an increase in endothelium-derived hyperpolarization (EDH). Endothelial-specific aging induced tubular injury, sodium wasting, and increased inflammatory markers in the kidney, which were normalized by the treatment with SUL-138. We conclude that accelerated endothelial aging adversely affects vascular function and causes kidney tubular injury. SUL-138 rescues endothelial aging, restores vasodilation by increasing EDH, and protects the kidney. Thus, preservation of mitochondrial function is a potential pharmacotherapeutic target in aging-related dysfunction provoked by the DNA damage response.

BACKGROUND: The microbiota-derived short chain fatty acid butyrate has been shown to lower blood pressure (BP) in rodent studies. Nonetheless, the net effect of butyrate on hypertension in humans remains uncovered. In this study, for the first time, we aimed to determine the effect of oral butyrate on BP in patients with hypertension. METHODS: We performed a double-blind randomized placebo-controlled trial including 23 patients with hypertension. Antihypertensive medication was discontinued for the duration of the study with a washout period of 4 weeks before starting the intervention. Participants received daily oral capsules containing either sodium butyrate or placebo with an equivalent dosage of sodium chloride for 4 weeks. The primary outcome was daytime 24-hour systolic BP. Differences between groups over time were assessed using linear mixed models (group-by-time interaction). RESULTS: Study participants (59.0±3.7 years; 56.5% female) had an average baseline office systolic BP of 143.5±14.6 mm Hg and diastolic BP of 93.0±8.3 mm Hg. Daytime 24-hour systolic and diastolic BP significantly increased over the intervention period in the butyrate compared with the placebo group, with an increase of +9.63 (95% CI, 2.02–17.20) mm Hg in daytime 24-hour systolic BP and +5.08 (95% CI, 1.34–8.78) mm Hg in diastolic BP over 4 weeks. Butyrate levels significantly increased in plasma, but not in feces, upon butyrate intake, underscoring its absorption. CONCLUSIONS: Four-week treatment with oral butyrate increased daytime systolic and diastolic BP in subjects with hypertension. Our findings implicate that butyrate does not have beneficial effects on human hypertension, which warrants caution in future butyrate intervention studies. REGISTRATION: URL: https://onderzoekmetmensen.nl/ ; Unique identifier: NL8924.

BACKGROUND: Small-interfering RNA (siRNA) targeting hepatic AGT (angiotensinogen) mRNA depletes AGT, lowering blood pressure for up to 6 months. However, certain situations may require a rapid angiotensin increase. The REVERSIR (RVR) - reverse siRNA silencing technology a potential approach to counteract siRNA effects. METHODS: Spontaneously hypertensive rats received 10 mg/kg AGT siRNA, and 3 weeks later were given AGT-RVR (1, 10, or 20 mg/kg). One week after AGT-RVR dosing, a redose of AGT siRNA assessed its post-AGT-RVR effectiveness for 2 weeks. Additionally, the impact of AGT-RVR after an equihypotensive dose of valsartan (4 mg/kg per day) was examined. RESULTS: Baseline mean arterial pressure (MAP) was 144±1 mm Hg. AGT siRNA reduced MAP by ≈16 mm Hg and AGT by >95%, while renin increased 25-fold. All AGT-RVR doses restored MAP to baseline within 4 to 7 days. Notably, 10 and 20 mg/kg restored AGT and renin to baseline, while 1 mg/kg allowed ≈50% AGT restoration, with renin remaining above baseline. A second AGT siRNA treatment, following 1 mg/kg AGT-RVR, reduced MAP to the same degree as the initial dose, while following 10 mg/kg AGT-RVR, it resulted in ≈50% of the first dose’s MAP effect at 2 weeks. The valsartan-induced MAP reduction was unaffected by AGT-RVR. CONCLUSIONS: In spontaneously hypertensive rats, angiotensinogen-RVR dose-dependently reversed AGT siRNA-induced AGT reduction, normalizing MAP. MAP normalization persisted even with 50% recovered AGT levels, likely due to upregulated renin maintaining adequate angiotensin generation. Post-AGT-RVR dosing, a second AGT siRNA dose lowered MAP again.

Transient receptor potential (TRP) channels are pivotal in modulating vascular functions. In fact, topical application of cinnamaldehyde or capsaicin (TRPA1 and TRPV1 channel agonists, respectively) induces “local” changes in blood flow by releasing vasodilator neuropeptides. We investigated TRP channels’ contributions and the pharmacological mechanisms driving vasodilation in human isolated dermal arteries. Ex vivo studies assessed the vascular function of artery segments and analyzed the effects of different compounds. Concentration–response curves to cinnamaldehyde, pregnenolone sulfate (PregS, TRPM3 agonist), and capsaicin were constructed to evaluate the effect of the antagonists HC030031 (TRPA1); isosakuranetin (TRPM3); and capsazepine (TRPV1). Additionally, the antagonists/inhibitors olcegepant (CGRP receptor); L-NAME (nitric oxide synthase); indomethacin (cyclooxygenase); TRAM-34 plus apamin (K⁺ channels); and MK-801 (NMDA receptors, only for PregS) were used. Moreover, CGRP release was assessed in the organ bath fluid post-agonist-exposure. In dermal arteries, cinnamaldehyde- and capsaicin-induced relaxation remained unchanged after the aforementioned antagonists, while PregS-induced relaxation was significantly inhibited by isosakuranetin, L-NAME and MK-801. Furthermore, there was a significant increase in CGRP levels post-agonist-exposure. In our experimental model, TRPA1 and TRPV1 channels seem not to be involved in cinnamaldehyde- or capsaicin-induced relaxation, respectively, whereas TRPM3 channels contribute to PregS-induced relaxation, possibly via CGRP-independent mechanisms.

Background: A single dose of small interfering RNA (siRNA) targeting liver angiotensinogen eliminates hepatic angiotensinogen and lowers blood pressure. Angiotensinogen elimination raises concerns for clinical application because an angiotensin rise is needed to maintain perfusion pressure during hypovolemia. Here, we investigated whether conventional vasopressors can raise arterial pressure after angiotensinogen depletion. Methods and Results: Spontaneously hypertensive rats on a low‐salt diet were treated with siRNA (10 mg/kg fortnightly) for 4 weeks, supplemented during the final 2 weeks with fludrocortisone (6 mg/kg per day), the α‐adrenergic agonist midodrine (4 mg/kg per day), or a high‐salt diet (all groups n=6–7). Pressor responsiveness to angiotensin II and norepinephrine was assessed before and after siRNA administration. Blood pressure was measured via radiotelemetry. Depletion of liver angiotensinogen by siRNA lowered plasma angiotensinogen concentrations by 99.2±0.1% and mean arterial pressure by 19 mm Hg. siRNA‐mediated blood pressure lowering was rapidly reversed by intravenous angiotensin II or norepinephrine, or gradually reversed by fludrocortisone or high salt intake. Midodrine had no effect. Unexpectedly, fludrocortisone partially restored plasma angiotensinogen concentrations in siRNA‐treated rats, and nearly abolished plasma renin concentrations. To investigate whether this angiotensinogen originated from nonhepatic sources, fludrocortisone was administered to mice lacking hepatic angiotensinogen. Fludrocortisone did not increase angiotensinogen in these mice, implying that the rise in angiotensinogen in the siRNA‐treated rats must have depended on the liver, most likely reflecting diminished cleavage by renin. Conclusions: Intact pressor responsiveness to conventional vasopressors provides pharmacological means to regulate the blood pressure–lowering effect of angiotensinogen siRNA and may support future therapeutic implementation of siRNA.

Changes in the renin–angiotensin system, known for its critical role in the regulation of blood pressure and sodium homeostasis, may contribute to aging and age-related diseases. While the renin–angiotensin system is suppressed during aging, little is known about its regulation and activity within tissues. However, this knowledge is required to successively treat or prevent renal disease in the elderly. Ercc1 is involved in important DNA repair pathways, and when mutated causes accelerated aging phenotypes in humans and mice. In this study, we hypothesized that unrepaired DNA damage contributes to accelerated kidney failure. We tested the use of the renin-activatable near-infrared fluorescent probe ReninSense680™ in progeroid Ercc1^d/− mice and compared renin activity levels in vivo to wild-type mice. First, we validated the specificity of the probe by detecting increased intrarenal activity after losartan treatment and the virtual absence of fluorescence in renin knock-out mice. Second, age-related kidney pathology, tubular anisokaryosis, glomerulosclerosis and increased apoptosis were confirmed in the kidneys of 24-week-old Ercc1^d/− mice, while initial renal development was normal. Next, we examined the in vivo renin activity in these Ercc1^d/− mice. Interestingly, increased intrarenal renin activity was detected by ReninSense in Ercc1^d/− compared to WT mice, while their plasma renin concentrations were lower. Hence, this study demonstrates that intrarenal RAS activity does not necessarily run in parallel with circulating renin in the aging mouse. In addition, our study supports the use of this probe for longitudinal imaging of altered RAS signaling in aging.