Chicken Muscle-Derived ACE2-Upregulating Peptide VVHPKESF Reduces Blood Pressure Associated with the ACE2/Ang (1-7)/MasR Axis in Spontaneously Hypertensive Rats

H. Fan, N. Shang, S. T. Davidge, J. Wu, Chicken Muscle-Derived ACE2-Upregulating Peptide VVHPKESF Reduces Blood Pressure Associated with the ACE2/Ang (1-7)/MasR Axis in Spontaneously Hypertensive Rats. Mol. Nutr. Food Res. 2024, 68, 2300524. https://doi.org/10.1002/mnfr.202300524

About

Hypertension is a major contributor to global morbidity and mortality. While synthetic pharmaceuticals remain the primary therapy for hypertension treatment, prolonged pharmacological interventions often have drawbacks. Consequently, lifestyle changes such as increasing physical activity and adopting the Dietary Approaches to Stop Hypertension diet are recommended. Bioactive peptides represent a burgeoning category of natural dietary compounds that show promise in managing hypertension. Chicken muscle proteome is a great source of bioactive peptides. Our previous work reported a chicken (laying hen) muscle hydrolysate demonstrating anti-inflammatory effects in vascular cells. These findings indicate the potential of peptides VRP, LKY, VRY, and V-F in reducing blood pressure (BP) in vivo and prompt an exploration into the mechanisms underpinning their antihypertensive effects. The present study aims to contribute valuable insights into the role of these peptides in hypertension management.

Approach

Peptides (VRP, LKY, VRY, and V-F with purity > 98%) were added to the cultured media to study ACE2 upregulation in rat aortic smooth muscle A7r5 cells at the concentration of 50 µm for 24 h, and perform the Caco-2 transport experiment at 5 mm initial concentration in the apical chamber. Animals (SHRs, 12–14 weeks old) were randomly assigned into five groups (n = 6): the untreated group and four peptides (VRP, LKY, VRY, and V-F) (15 mg kg−1 BW per day). Peptides were dissolved in 20 mL of 10% (in ddH2O v/v) Ensure (Abbott Nutrition, QC, Canada) and orally administered to animals from day 1 for 18 days; the untreated group was given 10% Ensure only. BP was recorded for a continuous 24 h (10 s of every 1 min) every 3 days until day 18 (BP on day 0 was recorded as the baseline). Systolic blood pressure (SBP), diastolic blood pressure (DBP), mean arterial pressure (MAP), and heart rate (HR, bpm) were recorded. At the end of the experiment, animals were sacrificed by cardiac blood collection under anesthesia. Blood and tissues were collected and stored at −80 °C for further analysis. Plasma was collected and plasma ACE activity was measured using an ACE assay kit. Concentrations of Ang II (CSB-E04494r), Ang (1-7) (CSB-E14241r), and ACE2 (CSB-E14308r) were evaluated by enzyme-linked immunosorbent assay (ELISA) kits. Circulating cytokines were quantified using rat cytokine strips.

Analysis of Results

SBP of the untreated SHRs was (173.7 ± 9.0) mmHg on day 0 and steadily increased to (179.5 ± 11.9) mmHg on day 18; a similar trend was observed for DBP and MAP. Only peptide V-F (15 mg kg−1 body weight [BW]) reduced BP over the treatment period, with SBP being reduced from (171.2 ± 9.6) mmHg on day 0 to (159.9 ± 15.2) mmHg on day 18. No significant change in HR was observed after peptide treatment. Thus, only the group of V-F was used for subsequent plasma and tissue analyses. Treatment with peptide V-F lowered the circulating level of Ang II (p = 0.011) while increased Ang (1-7) and ACE2 levels; no change in the circulating ACE activity was detected. Treatment with peptide V-F reduced circulating inflammatory cytokines or chemokines including tumor necrosis factor alpha (TNFα), monocyte chemoattractant protein-1 (MCP-1), and interleukins (IL-1α and IL-15). Although peptides VRP, LKY, and VRY showed high in vitro ACE-inhibitory activity and antioxidant activity, no change in BP reduction was observed in SHRs administered with these peptides.

Application

SHRs are the most widely used animal model for studying human essential hypertension due to their similar pathogenesis. SHRs develop persistent high BP in their early adulthood (9–15 weeks) till up to >180 mmHg (16–28 weeks). In summary, only peptide V-F reduced BP in SHRs among the four chicken-derived peptides; its activity involved the activation of the ACE2-Ang (1-7)-MasR axis and amelioration of vascular inflammation and oxidative stress. The lack of BP reduction in SHRs administered with LKY and VRY was likely due to their poor GI stability. Our findings further pinpointed the importance of bioavailability of peptides in dictating their in vivo efficacies; however, being unable to fully address the failure of peptides VRP, LKY, and VRY is another limitation of the present study. The presence of antihypertensive peptides in chicken muscle supports their potential uses as antihypertensive functional food ingredients and nutraceuticals.

Abstract

Scope

This study aims to investigate the antihypertensive effect of four chicken muscle-derived angiotensin (Ang)-converting enzymes (ACE)-regulating peptides: Val-Arg-Pro (VRP, ACE inhibition), Leu-Lys-Tyr and Val-Arg-Tyr (LKY and VRY, ACE inhibition and ACE2 upregulation), and Val-Val-His-Pro-Lys-Glu-Ser-Phe (VVHPKESF [V-F], ACE2 upregulation) in spontaneously hypertensive rats.

Methods and results

Rats (12–14 weeks old) are grouped: 1) untreated, 2) VRP, 3) LKY, 4) VRY, and 5) V-F. Blood pressure (BP) is monitored using implantable telemetry technology. Over 18-day oral administration of 15 mg kg⁻¹ body weight (BW) per day, only peptide V-F significantly (p < 0.05) reduces BP, decreases circulating Ang II, and increases ACE2 and Ang (1-7) levels, and enhances aortic expressions of ACE2 and Mas receptor (MasR). Peptide V-F also attenuates vascular inflammation (TNFα, MCP-1, IL-1α, IL-15, and cyclooxygenase 2 [COX2]) and vascular oxidative stress (nitrotyrosine). The gastrointestinal (GI)-degraded fragment of peptide V-F, Val-Val-His-Pro-Lys (VVHPK), is also an ACE2-upregulating peptide. Peptides VRP, LKY, and VRY do not reduce BP, possibly due to low bioavailability or other unknown reasons.

Conclusions

Peptide V-F is the first ACE2-upregulating peptide, purified and fractionated from food proteins based on in vitro ACE2 upregulation, that reduces BP associated with the activation of ACE2/Ang (1-7)/MasR axis; the N-terminal moiety VVHPK may be responsible for the antihypertensive effect of V-F.

KEYWORDS: bioactive peptides; blood pressure; chicken; inflammation; oxidative stress; spontaneously hypertensive rat