Transmural pressure and axial loading interactively regulate arterial remodeling ex vivo

Abstract

Physiological axial strains range between 40 and 60% in arteries, resulting in stresses comparable to those due to normal blood pressure or flow. To investigate the contribution of axial strain to arterial remodeling and function, porcine carotid arteries were cultured for 9 days at physiological and reduced axial stretch ratios in the presence of normotensive and hypertensive transmural pressures by ex vivo perfusion techniques. Consistent with previous in vivo studies, vessels cultured with physiological levels of axial strain and exposed to hypertensive pressure had greater mass, wall area, and outer diameter relative to those cultured at the same axial stretch ratio and normotensive pressure. Reducing the amount of axial strain resulted in mass loss and decreased cell proliferation. Culture in a hypertensive pressure environment at reduced axial strain produced arteries with greater contractility in response to norepinephrine. Arteries cultured at reduced axial strain with the matrix metalloproteinase inhibitor GM6001 maintained their masses over culture, indicating a possible mechanism for this model of axial stretch-dependent remodeling. Although not historically considered one of the primary stimuli for remodeling, multiple linear regression analysis revealed that axial strain had an impact similar to or greater than transmural pressure on various remodeling indexes (i.e., outer diameter, wall area, and wet mass), suggesting that axial strain is a primary mediator of vascular remodeling.

Fig. 1.

Calculations of stresses due to axial strain for arteries cultured at normotensive (100 mmHg) transmural pressure and an axial stretch ratio of 1.0, 1.3, or 1.5 by use of measured axial force at the stretch ratio at which the artery was cultured. Included for comparison are calculated circumferential stresses for arteries cultured with normotensive or hypertensive transmual pressure. n = 3 for each axial stretch ratio; τ is a physiologically representative mean shear stress magnitude of 1.5 Pa; n = 5 and 6 for normotensive and hypertensive pressures, respectively.

Fig. 2.

Normalized outer diameters over a 9-day perfusion period. A: there was an increase in outer diameter with respect to time for the hypertensive group at physiological axial strain. This response was abolished at reduced axial strain in B and C. *P < 0.05 within the hypertensive (200 mmHg) series compared with t = 0. **P < 0.05 within the normotensive (100 mmHg) series compared with t = 0. #P < 0.05 between the normotensive and hypertensive series. ##P < 0.01 between the normotensive and hypertensive series. In C, P ≤ 0.11 for all time points in the normotensive series with respect to t = 0 except day 6, when P = 0.16. When comparing the final values for all 6 groups by Duncan's test, the normalized outer diameter for the hypertensive λ = 1.5 group was greater than all other groups (P < 0.001) whereas the other groups were not significanlty different from one another.

Fig. 3.

A: percent change in wall area over the 9-day culture period. Wall structure measurements of histological sections from segments pressure-fixed at 100 mmHg were used to calculate wall areas for arteries perfused under hypertensive (solid bars) and normotensive (open bars) pressures and various axial stretch ratios. B: percent change in wet mass in arteries after 9-day culture in the ex vivo perfusion system. The hypertensive pressure/λ = 1.5 group was significantly greater (*P < 0.05) than all groups, with no statistical differences among the remaining groups.

Fig. 4.

A: cell proliferation effectively became nonexistent as axial stretch ratio during ex vivo culture was reduced. Images were captured with a ×10 objective lens. B: a mitotic index was developed for each experimental group [hypertensive (solid bars) and normotensive (open bars) pressures]. Axial strain had a much greater effect than transmural pressure on proliferation; n = 5 for all groups except hypertensive/λ = 1.3 (n = 4) and hypertensive/λ = 1.5 (n = 6). BrdU, 5-bromo-2-deoxyuridine. *P < 0.05 compared with the corresponding pressure group at λ = 1.0. #P < 0.05 compared with the corresponding pressure group at λ = 1.3. C: arteries with the matrix metalloproteinase (MMP) inhibitor GM6001 added to the culture medium experienced less cell proliferation than the control group (labeled Steady Flow); n = 5 for the steady flow and pulsatile groups, n = 4 for the steady flow w/GM6001 group.

Fig. 5.

Contractile responses to norepinephrine for arteries cultured under either normotensive (A/open bars; n = 5 for each level of axial stretch) or hypertensive [B/solid bars; λ = 1.5 (n = 6); λ = 1.3 (n = 5); λ = 1.0 (n = 4)] pressure at varying axial stretch ratios along with contractile responses of fresh arteries under different levels of axial stretch (C/shaded bars; n = 3 for each level of axial stretch). *Significantly different compared with mean for λ = 1.5 (P < 0.05).