Biphasic Change of Tau (τ) in Mice as Arterial Load Acutely Increased with Phenylephrine Injection

<div><p>Background</p><p>Diastolic dysfunction is the hemodynamic hallmark of hypertensive heart disease. Tau (τ) has been used to describe left ventricle relaxation. The relationship between τ and afterload has been controversial. Our goal was to demonstrate this relationship in mice, because genetically-modified mouse models have been used extensively for studies in cardiovascular diseases.</p> <p>Methods</p><p>Increased arterial load was produced by phenylephrine administration (50 µg/kg iv) (n = 10). A series of pressure-volume loops was recorded with a Millar conductance catheter <i>in vivo</i> as the left ventricle pressure reached the maximum. The arterial load was expressed as Ea (effective arterial elastance). Tau values were computed using three mathematical methods: τ_Weiss, τ_Glantz, and τ_Logistic.</p> <p>Results</p><p>A correlation plot between τ and Ea showed a biphasic relationship a flat phase I and an inclined phase II. The existence of an inflection point was proved mathematically with biphasic linear regression. Pressure-volume area (PVA), a parameter linearly related to myocardial O₂ consumption (MVO₂), was found to be directly proportional to Ea. The plot of τ versus PVA was also biphasic.</p> <p>Conclusion</p><p>We concluded that a small increase of the arterial load by phenylephrine increased PVA (index of MVO₂) but had little effect on τ. However, after an inflection point, further increase of arterial load and PVA resulted in the linear increase of τ.</p> </div

Arterial load was acutely increased by phenylephrine intravenous administration.

<p>A: Real time recording of series of pressure-volume loops during phenylephrine injection by Millar conductance catheter. The mean systolic peak pressure (B) and the mean arterial load (Ea) (C) of 10 mice were calculated before phenylephrine injection (Pre-PE) and at the maximum effects of phenylephrine (PE). PE = phenylephrine, Ea (effective arterial elastance) = Pes/SV (end systolic pressure/stroke volume).</p

The relationship between τ, Ea, LV peak systolic pressure, and pressure-volume area (PVA).

<p>Plots of the three τ computations versus Ea (A), peak systolic pressure (B), and PVA (C) with phenylephrine injection. (D) Combined plots of τ () and PVA versus Ea (PVA = 282.63×Ea +1178.5, R² = 0.9858).</p

Changes in heart rate, dP/dt max, and dP/dt min with phenylephrine injection.

<p>(A): Heart rate before phenylephrine injection (baseline), at Inflection Point (IP), at middle point of phase II (MP), and at the maximum effects of phenylephrine (Peak). (B): dP/dt max and dP/dt min before phenylephrine injection (baseline), at inflection point (IP), and at the maximum effects of phenylephrine (Peak). a, aa: p<0.05 or p<0.01 versus baseline; b: p<0.05 versus IP. IP: inflection point; MP: middle point.</p

The Ea (A), PVA (B), and systolic peak pressure (C) at inflection point.

<p>The Ea, PVA, and systolic peak pressure were significantly greater at inflection point than those at baseline. PE: phenylephrine; IP: inflection point. **: p<0.01, ***: p<0.001, self-paired t-test.</p

Eberson raw data

Each sheet contains Figures 1-5

Effect of Lysyl Oxidase Inhibition on Angiotensin II-Induced Arterial Hypertension, Remodeling, and Stiffness

<div><p>It is well accepted that angiotensin II (Ang II) induces altered vascular stiffness through responses including both structural and material remodeling. Concurrent with remodeling is the induction of the enzyme lysyl oxidase (LOX) through which ECM proteins are cross-linked. The study objective was to determine the effect of LOX mediated cross-linking on vascular mechanical properties. Three-month old mice were chronically treated with Ang II with or without the LOX blocker, β -aminopropionitrile (BAPN), for 14 days. Pulse wave velocity (PWV) from Doppler measurements of the aortic flow wave was used to quantify in vivo vascular stiffness in terms of an effective Young’s modulus. The increase in effective Young’s modulus with Ang II administration was abolished with the addition of BAPN, suggesting that the material properties are a major controlling element in vascular stiffness. BAPN inhibited the Ang II induced collagen cross-link formation by 2-fold and PWV by 44% (P<0.05). Consistent with this observation, morphometric analysis showed that BAPN did not affect the Ang II mediated increase in medial thickness but significantly reduced the adventitial thickness. Since the hypertensive state contributes to the measured in vivo PWV stiffness, we removed the Ang II infusion pumps on Day 14 and achieved normal arterial blood pressures. With pump removal we observed a decrease of the PWV in the Ang II group to 25% above that of the control values (P=0.002), with a complete return to control values in the Ang II plus BAPN group. In conclusion, we have shown that the increase in vascular stiffness with 14 day Ang II administration results from a combination of hypertension-induced wall strain, adventitial wall thickening and Ang II mediated LOX ECM cross-linking, which is a major material source of vascular stiffening, and that the increased PWV was significantly inhibited with co-administration of BAPN.</p></div

Effect of Ang II and BAPN on LOX, collagen, and cross-linking in aortas.

<p>The effect of LOX is not confined to the crosslinking of collagen and elastin and therefore we examined the effect of LOX on gene expression of other key ECM proteins and enzymes. <b>A</b>, Lysyl-oxidase enzymatic activity measured from the lower thoracic aorta represented by the production of H₂O₂ and detected by Amplex red oxidation. <b>B</b>, The total collagen content using hydroxyproline concentrations assuming collagen contains 13.5% hydroxyproline. <b>C</b>, Cross-linked collagen, using cyanogen bromide as digestion. <b>D</b>, Percent of collagen cross-linking. n = 6. *P<0.05 compared to control, ^ƗP<0.05 compared with BAPN group, ^#P<0.05 compared with Ang II group at their respective day.</p

Cessation of Ang II infusion.

<p>Hypertension contributes to vascular stiffness together with structural and material remodeling. Therefore the Alzet pumps were removed to determine the contribution of each to the total vascular stiffness. <b>A</b>, The Alzet pumps were removed on Day 14 and the blood pressure returned to control values by day + 4. <b>B</b>, The pulse wave velocity measured with and without Ang II infusion on days 14 and + 4 days after pump removal. n = 6. *P<0.05 compared to control, ^ƗP<0.05 compared with Ang II group, ^#P<0.05 compared with Ang II + BAPN + 4 group.</p

Vascular morphology.

<p>Aortic histomorphometry was performed with the elastin specific stain Verhoeff’s Van Gieson (VVG) and the collagen specific stain picrosirius red (PSR). <b>A</b>, Representative histological sections are presented according to treatment. <b>B</b>, The medial thickness and adventitial thickness were analyzed to describe the aortic morphological changes in response to the treatments. <b>C</b>, The aortic structural dimensions as measured with VVG and PSR stains. <b>D</b>, The systolic luminal diameter were measured with the M-mode ECHO, collagen content was quantified with PSR histological staining, and elastin content was measured with VVG stain. n = 6. *P<0.05 compared to control, ^ƗP<0.05 compared with BAPN group.</p