Abstract 22: Formyl Peptide Receptor-1 Activation Is Crucial For The Cause Of Spontaneous Hypertension In Dahl Salt Sensitive Rats

Mitochondria evolved from bacteria and use N-formylated peptides (NFPs) to synthetize protein. Bacterial and mitochondrial NFPs activate formyl peptide receptor 1 (FPR-1) and lead to vascular injury. We previously observed that Dahl Salt Sensitive rats (S) fed a low-salt (LS, 0.3% NaCl) diet presented spontaneous hypertension, vascular dysfunction, and overexpression of FPR-1 in arteries when compared to Dahl Salt Resistant (R) rats. High salt (HS, 2% NaCl) diet worsened these phenotypes in S rats. Interestingly, HS diet induced leaky gut and amoxicillin (AMO) treatment decreased BP in S-HS. Due to the dual sources of NFPs (microbiota and host mitochondria), we hypothesized that cell death-derived mitochondria and/or leaky gut-derived bacterial NFPs lead to FPR-1 activation, vascular injury and elevated BP in S rats independent of HS diet. For this, we used flow cytometry to measure cell necrosis and early and late apoptosis in kidney, bone marrow-derived macrophages and mesenteric resistance arteries (MRA) from male S and R rats (8-week old) on a LS diet. Zonulin, a biomarker for leaky gut, was measured in plasma. In another group, rats were treated with FPR-1 antagonist [Cyclosporin H (CsH), 0.3 mg/kg/day, osmotic mini-pump, 14 days], vehicle (VEH) or received water with AMO (5 mg/kg/day) for 21 days to deplete bacteria. BP was measured by telemetry and vascular function and structure were assessed in MRA. S rats presented increased kidney cell necrosis (R: 3.8±0.3 vs. S: 5.3±0.5* %). CsH decreased spontaneous elevation of BP [Diastolic: R+VEH: 77±2.7 vs. R+CsH: 81±1.2 vs. S+VEH: 126±3.0* vs. S+CsH:115±2.7 # ] and vascular hypercontractility [KCl (120mM): R+VEH: 9.4±1 vs. R+CsH: 10.2±0.4; S+VEH: 15.5±0.9* vs. S+CsH:11.7±0.8 # mN; Phenylephrine (10μM): R+VEH: 9.3±1 vs. R+CsH: 9.7±1; S+VEH: 14.5±1*vs. S+CsH: 11.4±0.6 # mN) in S-LS rats. AMO did not change vascular contraction or BP. Leaky gut was not observed in Dahl S-LS diet. In conclusion, FPR-1 can serve as a causative agent for the spontaneous elevation of BP and kidney-derived mitochondria, but not gut-derived microbiota, are the main source for NFPs.

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Abstract P217: Mitochondrial N-Formyl Peptides Elicit Changes in Endothelial Cell Cytoskeleton via Formyl Peptide Receptor Activation

Hypertension ◽

10.1161/hyp.68.suppl_1.p217 ◽

2016 ◽

Vol 68 (suppl_1) ◽

Author(s):

Camilla F Wenceslau ◽

Cameron G McCarthy ◽

R.Clinton Webb

Keyword(s):

Endothelial Cells ◽

Endothelial Cell ◽

Vascular Dysfunction ◽

Receptor Activation ◽

Vascular Leakage ◽

Formyl Peptide Receptor ◽

Experimental Hypertension ◽

Peptide Receptor ◽

Formyl Peptide ◽

Formyl Peptides

One major pathophysiological characteristic of cardiovascular disease, including hypertension, is vascular dysfunction. Recently, we demonstrated that mitochondrial damage-associated molecular patterns are elevated in the circulation of SHR. Mitochondria carry hallmarks of their bacterial ancestry and one of these hallmarks is that this organelle still uses an N-formyl-methionyl-tRNA as an initiator of protein synthesis. We observed that mitochondrial N-formyl peptides (F-MIT) infusion into rats induces inflammation and vascular dysfunction, including vascular leakage, via formyl peptide receptor (FPR) activation. However, neutrophil depletion did not change this response. Therefore, we hypothesize that F-MIT via FPR activation elicits changes directly to cytoskeleton-regulating proteins in vascular cells, which may lead to increased vascular permeability. To test this hypothesis we used vascular smooth muscle cells (VSMC) and endothelial cells harvested from aortas of Sprague-Dawley rats (n=5) and human donors (n=5), respectively. Cells were divided into three groups for Western blot analysis of cytoskeleton-regulating proteins. The cells were incubated for 20 minutes in medium with either vehicle (non-formylated peptide), F-MIT (10 μM), or F-MIT after a 5-minute pre-incubation with FPR1 and 2 antagonists (Cyclosporine H, CsH, 1 μM and WRW4, 10 μM). In endothelial cells, the treatment with F-MIT increased the protein expression of RhoA/ROCK (Rho: 1.8 fold vs. Veh; ROCK: 1.4 fold vs. Veh, p<0.05), cell division control protein 42 (CDC42) (2.0 fold vs. Veh, p<0.05) and phospho-myosin light chain (MLC) Thr/Ser19 (1.5 fold vs. Veh, p<0.05). These changes were all abolished in the presence of FPR antagonists. On the other hand, F-MIT decreased expression of phospho-MLC (0.6 fold vs. Veh, p<0.05) and CDC42 (0.5 fold vs. Veh, p<0.05) and did not change RhoA/ROCK expression in VSMC. In conclusion, F-MIT, via FPR activation, elicits direct changes in endothelial cell and VSMC cytoskeleton-regulating proteins. This interaction can lead to endothelial contraction, increased vascular leakage and attenuated barrier function as observed in clinical and experimental hypertension.

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Increase in soluble protein oligomers triggers the innate immune system promoting inflammation and vascular dysfunction in the pathogenesis of sepsis

Clinical Science ◽

10.1042/cs20180368 ◽

2018 ◽

Vol 132 (13) ◽

pp. 1433-1438

Author(s):

Amel Komic ◽

Patricia Martinez-Quinones ◽

Cameron G. McCarthy ◽

R. Clinton Webb ◽

Camilla F. Wenceslau

Keyword(s):

Soluble Protein ◽

Molecular Mechanisms ◽

Vascular Dysfunction ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Formyl Peptide Receptor ◽

Misfolded Proteins ◽

Resistance Arteries ◽

Oligomeric Protein ◽

Formyl Peptide

Sepsis is a profoundly morbid and life-threatening condition, and an increasingly alarming burden on modern healthcare economies. Patients with septic shock exhibit persistent hypotension despite adequate volume resuscitation requiring pharmacological vasoconstrictors, but the molecular mechanisms of this phenomenon remain unclear. The accumulation of misfolded proteins is linked to numerous diseases, and it has been observed that soluble oligomeric protein intermediates are the primary cytotoxic species in these conditions. Oligomeric protein assemblies have been shown to bind and activate a variety of pattern recognition receptors (PRRs) including formyl peptide receptor (FPR). While inhibition of endoplasmic reticulum (ER) stress and stabilization of protein homeostasis have been promising lines of inquiry regarding sepsis therapy, little attention has been given to the potential effects that the accumulation of misfolded proteins may have in driving sepsis pathogenesis. Here we propose that in sepsis, there is an accumulation of toxic misfolded proteins in the form of soluble protein oligomers (SPOs) that contribute to the inflammation and vascular dysfunction observed in sepsis via the activation of one or more PRRs including FPR. Our laboratory has shown increased levels of SPOs in the heart and intrarenal arteries of septic mice. We have also observed that exposure of resistance arteries and vascular smooth muscle cells to SPOs is associated with increased mitogen-activated protein kinase (MAPK) signaling including phosphorylated extracellular signal-regulated kinase (p-ERK) and p-P38 MAPK pathways, and that this response is abolished with the knockout of FPR. This hypothesis has promising clinical implications as it proposes a novel mechanism that can be exploited as a therapeutic target in sepsis.

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FPR-1 (Formyl Peptide Receptor-1) Activation Promotes Spontaneous, Premature Hypertension in Dahl Salt-Sensitive Rats

Hypertension ◽

10.1161/hypertensionaha.120.16237 ◽

2021 ◽

Author(s):

Jonnelle M. Edwards ◽

Shaunak Roy ◽

Sarah L. Galla ◽

Jeremy C. Tomcho ◽

Nicole R. Bearss ◽

...

Keyword(s):

Blood Pressure ◽

Cell Death ◽

Protein Translation ◽

Synergistic Action ◽

Formyl Peptide Receptor ◽

Leaky Gut ◽

Salt Diet ◽

Peptide Receptor ◽

Low Salt ◽

Formyl Peptide

Cell death has long been a characteristic phenotype of organ damage in hypertension, and recently, leaky gut has been revealed as a novel hypertensive phenotype. However, despite the increase in bacterial and damaged mitochondrial products in the circulation of hypertensive patients and animals, the mechanistic contribution of these two phenomena to hypertension pathophysiology is unknown. Mitochondria and bacteria both start protein translation with an N-formyl methionine residue and thus are the only sources of NFPs (N-formyl peptides), which activate the FPR-1 (formyl peptide receptor-1). We hypothesized that the synergistic action of bacterial and mitochondrial NFPs would cause the spontaneous elevation of blood pressure and vascular remodeling in male Dahl salt-sensitive rats via FPR-1. We observed that mitochondria-derived peptides originating from cell death in the kidneys are responsible for FPR-1–induced vascular hypercontractility and remodeling and premature elevation of BP in Dahl salt-sensitive rats fed a low-salt diet. However, a high-salt diet leads to gut barrier disruption and, subsequently, a synergistic action of mitochondria, and bacteria-derived leaky gut NFPs lead to a severe and established hypertension. Administration of an FPR-1 antagonist lowered blood pressure in Dahl salt-sensitive rats on a low-salt diet but amoxicillin administration did not. These results reveal for the first time that cell death can be a cause of hypertensive pathophysiology, whereas leaky gut is a consequence.

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