(D-150) Toward Measuring the Influence of Perivascular Tissue on Pressure-Diameter Relationships of Murine Common Carotid and Femoral Arteries

Introduction::

Perivascular adipose tissue (PVAT) is known to be capable of affecting biological properties of blood vessels such as arteries. Not only do these surrounding tissues provide external support to blood vessels, but PVAT also acts as an endocrine organ, supports inflammation during atherosclerosis and vascular remodeling and has vasodilatory and contractile effects [1]. To date little is known about the extent to which PVAT influences the mechanical properties of blood vessels while they are maintained inside the body. Recent findings from our lab have indicated that transitional elastic arteries, such as the common carotid and femoral arteries, increase their diameter in response to increasing pressures to a lesser extent when surrounded by PVAT (in situ) than in traditional ex vivo testing with PVAT removed. In this study, we sought to quantify the difference in the average pressure-diameter relationship between common carotid arteries and femoral arteries in both in situ and ex vivo conditions. We hypothesized despite geometric differences between carotid and femoral arteries, both would exhibit greater outer diameters (OD) during ex vivo testing, relative to in situ testing. Common carotid arteries supply blood to the brain and are often subject to atherosclerosis and treatments against strokes. Femoral arteries, on the other hand, are prone to peripheral arterial disease which prevents blood flow to the lower limbs. A better knowledge of PVAT's influence on the mechanical properties of these vessels can allow for improvements in the treatment of these and other vascular conditions.

Materials and Methods::

In situ mechanical testing. 8-12 weeks-old male wild-type mice were gently dissected to reveal the femoral arteries. A 25G needle attached to a hydrostatic pressure system was then introduced in the left ventricle of the heart to allow for Hank's buffered saline solution to flow through the vasculature. To minimize fluid leakage, the needle was clamped in-place in the ventricle and the inferior vena cava was ligated with single strand of 9/0 nylon suture. The exposed femorals were then imaged under an Optical Coherence Tomography (OCT) microscope (Thorlabs, Inc.) to visualize vessel cross-sections (Figure 1A,B) at increasing intraluminal pressures (using a pressure controller; Fluigent). A similar technique was used when imaging the right and left common carotids, however the descending thoracic aorta was ligated. OCT images were taken from 20 to 140mmHg with a 20mmHg step and ImageJ was used to determine the vessels’ OD based on a known pixel size from the OCT acquisition.

 

Ex vivo mechanical testing. After in situ testing, the common carotid and femoral arteries were dissected and cleaned of any excess perivascular tissues. Vessels were then ligated to a metal cannula (30G for femorals; 27G for carotids) at each end using 6/0 silk suture. When present, branches were tied off with 9/0 nylon sutures. Cannulated vessels were then mounted on a computer-controlled biaxial mechanical testing system which allows for the measurement of OD and axial force in response to pressurization and axial stretch [2]. For paired comparisons with in situ data, pressure-diameter curves were retrieved.

Results, Conclusions, and Discussions::

A paired T-test between the ODs measured in situ and ex vivo at set pressure values revealed a significant difference between them in both the common carotids and femorals at every pressure value (stars; Figure 1C,D). To compare the pressure-diameter relationship between carotids and femorals, we ran unpaired T-tests between the differences in measured ODs (e.g., ex vivo OD - in situ OD) at each pressure, testing the hypothesis that carotids have a larger difference in diameter, compared to femorals. However, we found significant differences in pressure-diameter relationships only at 20mmHg and 40mmHg (pound signs; Figure 1C,D), suggesting similar differences in measured diameters at higher pressures. In Figure 1C,D, the in-situ curves for both carotid and femoral arteries follow a linear trend while the ex vivo curves exhibit characteristic non-linear responses. A Pearson Correlation test for both in situ and ex vivo curves with respect to pressure showed a significant linear correlation between in situ curves, but not ex vivo curves, at the 0.1% significance level.

 

Using OCT to test vessel mechanical properties in situ is a novel approach to investigating blood vessel behavior inside organisms. The linear in situ vs. nonlinear ex vivo response of both carotids and femorals is critical as it suggests that these vessels behave differently with PVAT present. The decrease in compliance observed in arteries surrounded by PVAT confirms that PVAT contributes to attenuating the mechanical action of elastin and smooth muscle in regulating the arterial inner diameter in response to controlled pressurization. This decrease was insignificantly different between carotids and femorals, implying that the PVAT surrounding them similarly influences their degree of dilation (at least for pressures above 40mmHg). Although not significant, the greater difference between the in situ and ex vivo measurements in carotids compared to femorals may be due to differences in perivascular tissue structure/composition.

Furthering this study with a more diverse pool of arteries in different disease states (e.g., hypertension and, atherosclerosis, etc.) could provide findings with the potential to improve the constitutive modeling of arteries, and inform (non)surgical treatments of vascular disease in an effort to better cardiovascular health.

Acknowledgements (Optional): : We would like to acknowledge the support of the American Heart Association (AHA) and the WashU Cardiovascular Summer Research (CardS) program. This work was supported in part by the National Institutes of Health (R00 HL146951 to MRB).

References (Optional): :

1. Brown, NK., et al. Arteriosclerosis, Thrombosis, and Vascular Biology, 2014.
2. Ferruzzi, J., et al.. Annals of Biomedical Engineering, 2013.