**2. Materials, methods**

#### **2.1 Arteries**

Abdominal aortas from freshly slaughtered young white pigs were obtained from an abbattoir conforming to European animal welfare standards (EC Regulation 1099/2009) and were transported at 4°C Ringer's lactate solution. The average length of the test segment was 120 mm (SD 5 mm), the internal diameter of the proximal end 18.1 mm (SD+/− 0.54) and that of the distal end 13.8 mm (SD+/−2.7).

In each artery adherent loose areolar-lymphatic tissue was removed whist preserving the adventitia and all side branches were ligated at their origins with 2/0 silk whilst keeping the artery moist and cool in Ringers-lactate solution. The segments of artery used extended from the coeliac trunk to the aortic bifurcation.

For studies on the stress strain relationship across the intima in pulsatile flow conditions a fluid-filled 0.9 mm polythene saline-filled catheter was introduced through a side branch and secured by a ligature. The tip of the catheter was positioned near the midpoint of the test artery, this point being marked on the exterior wall of the artery using Gentian Violet to facilitate the subsequent positioning of the ultrasound probe. During each test run this catheter was perfused at 0.1 ml/hr. with saline using a syringe pump. Once the catheter was in position a thin deformable latex membrane (Préservatif classique, PHR Lab, Boulogne Billancourt, Fr) was introduced along the full length of the arterial lumen. The membrane overlapped the ends and was smoothed against the internal wall of the artery by filling the lumen with saline. This membrane was intended to take the role of a deformable FI. **Figure 1** shows the appearance of the lined artery and the movement of the FI in relation to the wall on B-mode ultrasound when the artery is subjected to pulsatile flow under test conditions. The presence of this lining membrane also had the advantage of preventing minor leaks. The specimen, consisting of the artery and the FI, was then mounted in the test bath illustrated in **Figure 2**, being secured over the tube at either end by sliding it over two O- rings positioned on the mounting tube and holding it between the rings with a double turn of a 4 mm Silastic sling.

The artery was rotated until the catheter tip lay inferiorly, directly opposite the ultrasound probe. The tubes upon which the artery was mounted were then slid far enough apart for the artery to be lie in a relaxed straight line, not under tension. The bath was filled with oxygenated Ringer's lactate at 37°C, up to but not over the top of the uppermost wall of the artery. Contact jelly was applied and the ultrasound probe set up over the tip of the catheter so as to give a view of the artery an the membrane in transverse section. The probe was held in a clamp in contact with the superior wall of the artery but not pressing sufficiently to deform it.

**Figure 1.**

*A. The movement in B-mode ultrasound of the posterior wall of the artery and the false intima (FI) subjected to a pulse pressure of 60 mmHg and a peak pressure wave of an intermediate level of sharpness. B. The thin latex false intima lining the opened test artery; the perfused catheter is placed between the FI and the inner arterial wall to measure the differential pressure between this space and the adjacent arterial lumen.*

#### **Figure 2.**

*The artery in its test bath. The stress imposed on the FI throughout the pressure cycle is observed by recording the differential pressure between the perfused catheter and the adjacent lumen, whilst the strain response of the FI to the passage of the pulse wave is observed using M-mode ultrasound.*

#### **2.2 Perfusion apparatus**

The arteries were perfused with isotonic saline or sucrose/saline solutions with sucrose concentration adjusted to provide a range of viscosities using the apparatus shown in **Figure 3**. The perfusate was circulated from a 1 L reservoir by a centrifugal pump, forming a primary circuit by which an adjustable head of pressure could be obtained through adjustment of the return circuit valve. This circuit was tapped through a side branch to form a secondary circuit by means of which the test artery was perfused. This circuit circulated fluid at the established head of pressure through the test artery then back to the reservoir via a barostatic valve and flow meter. Pulsatile flow was imposed on the fluid by a 60 ml glass syringe pump which was operated by means of a series of cams of different profiles turned by a geared and governed slow speed electric motor. These cams were designed to reduplicate a variety of human aortic waveforms [18]. Eight cams were manufactured to deliver stroke volumes between 8 and 32mls.

*Role of Arterial Pressure, Wall Stiffness, Pulse Pressure and Waveform in Arterial Wall… DOI: http://dx.doi.org/10.5772/intechopen.100048*

Arterial pressure and pulse pressure wave profile were recorded via side tap taken off the upstream side of the inflow tube enabling differential pressure to be measured between the arterial lumen and the water bath adjacent to the artery at the same level (the arterial pressure). CTS was measured by recording the differential pressure between the perfused catheter in the sub-membrane space and a static port in the adjacent arterial lumen. These two differential pressures were measured using Honeywell 24PCBPFAD transducers calibrated using a mercury manometer, and recorded simultaneously using a dual channel Thornton 464 (Waltham, Ma) chart recorder.

The deformation of the arterial wall in response to pressure change and the displacement of the FI during pulsatile flow was observed during different phases of the pressure cycle using a linear 7.5Mhz ultrasound probe, by means of which the transverse section artery, its wall and the FI could be visualised using split B and M-mode ultrasound, **Figure 1**. Care was taken to ensure that the artery itself and the catheters and tubes connected to the differential pressure transducers were free of bubbles.
