Sex differences and role of lysyl oxidase-like 2 in angiotensin II-induced hypertension in mice.

Hypertension, a disease with known sexual dimorphism, accelerates aging-associated arterial stiffening, partly because of the activation of matrix remodeling caused by increased biomechanical load. In this study, we tested the effect of biological sex and the role of the matrix remodeling enzyme lysyl oxidase-like 2 (LOXL2) in hypertension-induced arterial stiffening. Hypertension was induced by angiotensin II (ANG II) infusion via osmotic minipumps in 12- to 14-wk-old male and female mice. Blood pressure and pulse wave velocity (PWV) were measured noninvasively. Wire myography and uniaxial tensile testing were used to test aortic vasoreactivity and mechanical properties. Aortic wall composition was examined by histology and Western blotting. Uniaxial stretch of cultured cells was used to evaluate the effect of biomechanical strain. LOXL2's catalytic function was examined using knockout and inhibition. ANG II infusion-induced hypertension in both genotypes and sexes. Wild-type (WT) males exhibited arterial stiffening in vivo and ex vivo. Aortic remodeling with increased wall thickness, intralamellar distance, higher LOXL2, and collagen I and IV content was noted in WT males. Female mice did not exhibit increased PWV despite the onset of hypertension. LOXL2 depletion improved vascular reactivity and mechanics in hypertensive males. LOXL2 depletion improved aortic mechanics but worsened hypercontractility in females. Hypertensive cyclic strain contributed to LOXL2 upregulation in the cell-derived matrix in vascular smooth muscle cells (VSMCs) but not endothelial cells. LOXL2's catalytic function facilitated VSMC alignment in response to biomechanical strain. In conclusion, in males, arterial stiffening in hypertension is driven both by VSMC response and matrix remodeling. Females are protected from PWV elevation in hypertension. LOXL2 depletion is protective in males with improved mechanical and functional aortic properties. VSMCs are the primary source of LOXL2 in the aorta, and hypertension increases LOXL2 processing and shifts to collagen I accumulation. Overall, LOXL2 depletion offers protection in young hypertensive males and females.NEW & NOTEWORTHY We examined the effect of sex on the evolution of angiotensin II (ANG II)-induced hypertension and the role of lysyl oxidase-like 2 (LOXL2), an enzyme that catalyzes matrix cross linking. While ANG II led to hypertension and worsening vascular reactivity in both sexes, aortic remodeling and stiffening occurred only in males. LOXL2 depletion improved outcomes in males but not females. Thus males and females exhibit a distinct etiology of hypertension and LOXL2 is an effective target in males.

An evolutionarily conserved olfactory receptor is required for sex differences in blood pressure.

Sex differences in blood pressure are well-established, with premenopausal women having lower blood pressure than men by ~10 millimeters of mercury; however, the underlying mechanisms are not fully understood. We report here that sex differences in blood pressure are absent in olfactory receptor 558 knockout (KO) mice. Olfr558 localizes to renin-positive cells in the kidney and to vascular smooth muscle cells. Female KOs exhibit increased blood pressure and increased pulse wave velocity. In contrast, male KO mice have decreased renin expression and activity, altered vascular reactivity, and decreased diastolic pressure. A rare OR51E1 (human ortholog) missense variant has a statistically significant sex interaction effect with diastolic blood pressure, increasing diastolic blood pressure in women but decreasing it in men. In summary, our findings demonstrate an evolutionarily conserved role for OLFR558/OR51E1 to mediate sex differences in blood pressure.

Optimization of resting tension for wire myography in male rat pulmonary arteries.

Wire myography to test vasomotor functions of blood vessels ex-vivo are well-established for the systemic circulation, however, there is no consensus on protocols for pulmonary arteries. We created a standardized wire myography protocol for healthy rat PAs and validated this in a pulmonary hypertension (PH) model. Vessels stretched to higher initial tensions (5.0, 7.5 and 10.0 mN) exhibited a uniform response to phenylephrine, a larger dynamic range, and lower EC50 values. The endothelium-mediated relaxation showed that moderate tensions (7.5 and 10.0 mN) produced robust responses with higher maximum relaxation and lower EC50 values. For endothelium independent responses, the higher initial tension groups had lower and more consistent EC50 values than the lower initial tension groups. Pulmonary arteries from rats with PH were more responsive to vasoactive drugs when subjected to a higher initial tension. Notably, vessels in the PH group subjected to 15.0 mN exhibited high dynamic ranges in contractile and relaxation responses without tearing. Lastly, we observed attenuated cholinergic responses in these vessels-consistent with endothelial dysfunction in PH. Therefore, a moderate initial tension of 7.5-10.0 mN is optimal for healthy rat pulmonary arteries and a higher initial tension of 15.0 mN is optimal for pulmonary arteries from animals with PH.

Sex Differences and Role of Lysyl Oxidase Like 2 (LOXL2) in Angiotensin II-Induced Hypertension in Mice.

BACKGROUND: Hypertension, a disease with known sexual dimorphism, accelerates aging associated arterial stiffening, in part due to the activation of matrix remodeling caused by increased biomechanical load. In this study, we tested the effect of biological sex and the role of the matrix remodeling enzyme lysyl oxidase like 2 (LOXL2) in hypertension induced arterial stiffening. METHODS: Angiotensin II (Ang II) was delivered using osmotic pumps in Loxl2+/- and WT male and female mice. Blood pressure and pulse wave velocity (PWV) were measured noninvasively to assess hypertension and aortic stiffness. Wire myography and uniaxial tensile testing were used to test aortic vasoreactivity and mechanical properties. Aortic wall composition was examined by histology and Western blotting. The effect of biomechanical strain on LOXL2 expression and secretion by vascular smooth muscle (VSMC) and endothelial cells (EC) was evaluated by uniaxial cyclic stretching of cultured cells. The role of LOXL2 catalytic function on VSMC alignment in response to mechanical loading was determined with LOXL2 inhibition and knockout. RESULTS: Ang II infusion induced hypertension in WT and Loxl2+/- mice of both sexes and increased PWV in WT males but not in Loxl2+/- males, WT females, or Loxl2+/- females. LOXL2 depletion protected males from Ang II mediated potentiation of vasoconstriction but worsened in females and improved aortic mechanical properties in both sexes. Histological analysis showed increased aortic wall thickness in hypertensive WT males but not females and increased intralamellar distance in both sexes, that was ameliorated in Loxl2+/- mice. Western blotting revealed increased collagen I, decreased collagen IV, and increased LOXL2 accumulation and processing in hypertensive mice. Hypertensive cyclic strain contributed to LOXL2 upregulation in the cell-derived matrix in VSMCs but not ECs. LOXL2 catalytic function facilitated VSMC alignment in response to biomechanical strain. CONCLUSIONS: In males, arterial stiffening in hypertension is driven both by VSMC response and matrix remodeling. Females exhibit a delayed onset of Ang II-induced hypertension with minimal ECM remodeling but with VSMC dysfunction. LOXL2 depletion ameliorates arterial stiffening and preserves functional contractility and aortic structure in male hypertensive mice. LOXL2 depletion improves aortic mechanics but worsens aortic contractility in hypertensive females. VSMCs are the primary source of LOXL2 in the aorta and hypertension increases LOXL2 processing and shifts to collagen I accumulation. Overall, LOXL2 depletion offers protection in young hypertensive males and females.

Neonatal exposure to hypoxia induces early arterial stiffening via activation of lysyl oxidases.

Hypoxia in the neonatal period is associated with early manifestations of adverse cardiovascular health in adulthood including higher risk of hypertension and atherosclerosis. We hypothesize that this occurs due to activation of lysyl oxidases (LOXs) and the remodeling of the large conduit vessels, leading to early arterial stiffening. Newborn C57Bl/6 mice were exposed to hypoxia (FiO2  = 11.5%) from postnatal day 1 (P1) to postnatal day 11 (P11), followed by resumption of normoxia. Controls were maintained in normoxia. Using in vivo (pulse wave velocity; PWV) and ex vivo (tensile testing) arterial stiffness indexes, we determined that mice exposed to neonatal hypoxia had significantly higher arterial stiffness compared with normoxia controls by young adulthood (P60), and it increased further by P120. Echocardiography performed at P60 showed that mice exposed to hypoxia displayed a compensated dilated cardiomyopathy. Western blotting revelated that neonatal hypoxia accelerated age-related increase in LOXL2 protein expression in the aorta and elevated LOXL2 expression in the PA at P11 with a delayed decay toward normoxic controls. In the heart and lung, gene and protein expression of LOX/LOXL2 were upregulated at P11, with a delayed decay when compared to normoxic controls. Neonatal hypoxia results in a significant increase in arterial stiffness in early adulthood due to aberrant LOX/LOXL2 expression. This suggests an acceleration in the mechanical decline of the cardiovascular system, that contributes to increased risk of hypertension in young adults exposed to neonatal hypoxia that may increase susceptibility to further insults.