International Journal of Angiology 9:151 155 (2000) DOI: 10.1007/s005470000022 Spatial Variation of the Veno-arteriolar Reflex (VAR) and Effect of Reconstructive Surgery in Limbs with Chronic Critical Limb Ischaemia (CLI) Einar Stranden, Ph.D., 1 Kristine Jacobsen Berger, 2 Karthrine Elise Pedersen 2 1 Department of Vascular Diagnosis and Research, Aker Hospital, University of Oslo, Norway 2 Department of General Physiology, Institute of Biology, University of Oslo, Norway Abstract. Ischaemic ulcers and necrosis tend to appear first in the most distal part of the feet in patients with chronic critical limb ischaemia (CLI), indicating that derangements in local microcirculation are focally distributed. In the present study the veno-arteriolar response (VAR), as an indicator of microcirculatory regulation, was obtained in three different areas of critically ischaemic feet. Five women and seven men, mean age 75 years, with unilateral CLI were included. Two groups including young and age-matched healthy participants served as controls. Laser Doppler flux (LDF) was recorded simultaneously at the pulp of the first toe (site 1), the level of the second metatarsal body (site 2), and the anterolateral part of the ankle (site 3). LDF was recorded with the investigated limb in supine and dependent positions. Orthostatic response (OR) was calculated at all measuring sites as perfusion (LDF) in the dependent foot divided by perfusion in the supine position. There was no difference in OR between the control groups, or between sites within each group. Median OR was about 0.5, indicating that perfusion was reduced to 50% during dependency. In patients with CLI, VAR was not present in the afflicted foot and ankle, and OR at the three sites were significantly greater than in controls (p < 0.0005). Median OR at site 1 was 3.7, indicating considerably increased local perfusion when the foot was lowered. At site 2 the increase was moderate, and there was no increase present at site 3 (median OR 1.0). There was a significant trend towards normal values (OR < 0.8) after vascular reconstruction at all sites. Abolished VAR and increased local perfusion in ischaemic limbs assist in explaining why patients with ischaemic rest pain obtain relief of pain with dependency. The recovery of VAR following reconstructive surgery indicates that the microcirculatory derangement is reversible. Correspondence to: Einar Stranden, Ph.D., Department of Vascular Diagnosis and Research, Aker Hospital, University of Oslo, 0514 Oslo, Norway Introduction Chronic critical limb ischaemia (CLI) is the most advanced stage of peripheral atherosclerosis [8,25], where the perfusion of nutritive capillaries is diminished to a degree where basal requirements of tissue oxygenation are not covered. Treatment of this serious condition with high morbidity and mortality [18,29] requires optimal understanding of the underlying pathogenetic mechanisms. Patients with CLI tend to lower the painful leg to alleviate pain at rest. Under normal physiological conditions, a venoarteriolar response (VAR) leads to increased precapillary resistance when the lower limb shifts from supine to the sitting position [5,7,10]. The mechanism limits the rise in capillary pressure resulting from the vertical column of blood from the heart to the foot. Previous studies on VAR in the ischaemic lower limb revealed a disturbance of this mechanism [4,20,27,30,31,32]. An impaired VAR could cause increased intracapillary hydrostatic pressure and thereby transudation of fluid through the capillary wall leading to interstitial oedema formation. The VAR is initiated when venous transmural pressure is greater than 25 mmhg [10]. In the foot this may be achieved by different procedures, including: 1. By shifting from supine to erect or sitting position [2,32] 2. Inflating of a venous occlusion cuff at the thigh or calf to subdiastolic pressure causing venous congestion [6] 3. Exposure of a limb to subatmospheric pressure [6,27] 4. Passive lowering of a limb below heart level by flexion of the knee joint By the first method, central, as well as peripheral, local vascular reflexes may be activated, with difficulties in distinguishing between these effects. In the second method, inflation of a pneumatic cuff causes a rise in the distal venous pressure that eventually approaches cuff pressure. However, since pressure is increased most in the veins, the perfusion pressure, and hence blood flow, will decrease.
152 Methods three and four do not seem to have these limitations. In the present study method four was chosen because it mimics the situation that is common with CLI and pain at bed rest. The VAR was measured by shifting the afflicted limb position while the patients were kept supine to eliminate or minimize involvement of central orthostatic regulatory mechanisms. Because ischaemic ulcers and necrosis tend to appear first in the most distal part of the feet in patients with CLI, there is reason to assume that derangements in local microcirculation are focally distributed. The main aim of the present study was therefore to establish any variations in VAR in different areas of the critically ischaemic foot, and the effect of vascular reconstructive surgery on this distribution. To investigate whether any derangement could be ascribed to age, two control groups were included: healthy young subjects and healthy age-matched subjects. Material and Methods The study comprised one group of patients with CLI and two control groups, one group with healthy young subjects (C1) and one group with healthy age-matched subjects (C2). Control group C1 included ten healthy participants (five women and five men) with a mean age of 26 years ±4 (SD), and group C2 included ten participants (six women and four men) with a mean age of 69 years ±12. All control subjects had normal foot pulses and blood pressures. None had symptoms, or earlier history of cardiovascular disease or diabetes mellitus. Twelve patients (five women and seven men with a mean age of 75 years ±14) with unilateral CLI were included. CLI was defined according to the Second European Consensus Document on CLI [25] by either of the following two criteria: Persistently recurring ischemic rest pain requiring opiate analgesia for more than two weeks, or ulceration or gangrene of the foot or toes; either of these clinical entities combined with an ankle systolic pressure (ASP) 50 mm Hg or toe systolic pressure 30 mmhg. Mean ankle brachial index (ABI) in the limbs with CLI was 0.21 ± 0.23 (SD) with a mean ASP of 31 ± 18 mmhg. In four limbs with CLI ultrasound, Doppler signals could not be detected, and distal ankle systolic pressure was defined as 15 mm Hg. Following vascular surgery (7 10 days postoperatively), the mean ABI had increased to 79.5 ± 18.6 mmhg. Patients with an amputated contralateral limb, clinical signs of congestive heart failure, diabetes mellitus or manifest venous insufficiency were excluded from the study. No patients had signs of local infection in the critically ischaemic foot. The patients were investigated three times: Prior to vascular reconstructive surgery for CLI, 4 7 days, and 60 90 days following surgery. The Regional Ethical Committee of Southern Norway accepted the study. Laser Doppler Fluxmetry Stranden et al.: Spatial Variation of the Veno-arteriolar Reflex in CLI Fig. 1. Schematic illustration of the experimental set-up with the laser Doppler fluxmeter probes attached to the limbs with CLI: The pulp of the first toe (Site 1), at the level of the second metatarsal body (Site 2), and at the anterolateral part of ankle (Site 3). A fourth probe was attached to the first toe of the contralateral limb as reference. M is a water filled, openended manometer line connected to a pressure monitor for recording variation in the transmural pressure. Two laser Doppler (LD) fluxmeters (Periflux PF 4001 and 4002, Perimed AB, Järfälla, Sweden), with two channels each, were used simultaneously to evaluate the microcirculation at four different areas of the skin. The depth of measurement using PF 4001 and 4002 is roughly 0.5 1 mm, but varies considerably depending on blood content of the tissue, degree of oxygenation and skin pigmentation [22]. The laser Doppler flux (LDF) values, expressed as perfusion unit (PU), were saved by Perisoft acquisition and analysis software program (Perimed AB) in a computer for offline analysis. The laser probes were applied by means of a probe holder and double adhesive tape at the pulp of the first toe (site 1), the level of the second metatarsal body (site 2), and the anterolateral part of ankle (site 3). A fourth probe was attached to the first toe of the contralateral limb as reference (Figure 1). Measurements were performed after a 10 15 minute period of rest, with the subject lying down with both limbs in the horizontal position. The room temperature was kept between 23 25 C. First, skin perfusion was continuously recorded for three minutes in horisontal position. Mean PU during this period represented the LDF value for the supine position. The limb was then passively lowered by flexion in the knee joint with the three laser probes still attached. Three minutes of recording represented the LDF values for the dependent limb. Carefully, the limb was placed in horizontal position again to repeat the same measurements. This procedure was repeated three times for each patient (Fig. 1). Variation in transmural pressure The VAR is activated when venous transmural pressure is greater than 25 mmhg [10]. To record changes in this pressure, a water filled, open-ended manometer line connected to a pressure monitor (Pressure Monitor, Stranden) was attached to the skin at the dorsum of the foot (Fig. 1). This device recorded vertical movements of the foot during the measurements. Alterations in pressure indicated variations in the vertical position of the foot, which in turn represented changes of the transmural pressure in the foot veins. Calculation of the orthostatic response (OR) OR is calculated as the quotient between skin perfusion at dependency (PU d ) and in the horizontal position (PU h ): OR PU d /PU h. OR <0.8 indicates arteriolar constriction in the measured skin area, which normally occurs when the limb is lowered. In our study the finding of an OR 0.8 was defined as an abolished or disturbed VAR. Statistics The Kruskal-Wallis test was used to test for statistical significance within three or more groups. For comparison between study sites or between groups, Wilcoxon signed rank sum test (two-tailed) for paired observations and Mann-Whitney test (two-tailed) for unpaired data were used. GraphPad Prism 3.0 was used for analysis and presentation of data (GraphPad Software, Inc., San Diego, CA, USA). Differences were considered significant when p-values were less than 0.05. The values are given as mean ± standard deviation (SD) or medians and range in parenthesis. In the box plots, results are given as median value, 25% and 75% percentiles, and total range.
Stranden et al.: Spatial Variation of the Veno-arteriolar Reflex in CLI 153 Fig. 2. Orthostatic response obtained by lowering the foot in the two control groups, C1 (young subjects) and C2 (old subjects). Three areas were recorded: The pulp of the first toe (Site 1), at the level of the second metatarsal body (Site 2) and anterolateral part of the ankle (Site 3). OR is calculated as the quotient between skin perfusion at dependency (PU d ) and in the horizontal position (PU h ). The box plots indicate median value, 25% and 75% percentiles and total range. K-WT: Kruskal-Wallis test. Results Figure 2 summarizes calculations of the orthostatic response in the two control groups for the three measuring sites of the investigated extremity. Except for three measurements (5%), all OR were less than 1.0. In 53 out of 60 sites OR was below 0.8, indicating activated VAR. The median OR for both groups and sites 1 3 was approximately 0.5, indicating a reduction in perfusion of 50% during dependency. No trend between the different sites for any of the groups (Kruskal-Wallis test), or between the groups (Mann- Whitney s test) was found. Hence, the study did not indicate any spatial variation of the VAR in normal feet, or any age-dependent variation. Accordingly, the groups C1 and C2 were pooled for comparison with the patient group. The calculated OR for the first toe of the contralateral limb (site 4, supine) was 0.98 (0.85 1.18) and 0.99 (0.76 1.16) for groups C1 and C2, respectively. Lowering of a limb consequently did not cause any generalized variation in perfusion, as judged from the contralateral foot. Figure 3 shows the distribution of OR in limbs with CLI at the different sites as compared with the pooled control groups. Except for one patient at site 2 and two patients at site 3, OR was greater than 0.8 in all patients, indicating no VAR. For all three sites, OR in the patient group was significantly higher than in controls, p < 0.0005. For site 1 the median OR in patients was 3.7, meaning that perfusion in the first toe increased 3.7 times when lowering the leg. A statistically significant reduction in OR in proximal direction from the toe was found in the patients, 3.7 (1.54 47.00) (site 1), 1.1 (0.60 2.00) (site 2), 1.0 (0.67 2.28) (site 3), p < 0.0001 (Kruskal-Wallis test). There was, however, no difference between site 2 and site 3 (p 0.32). In the patient group, the calculated OR for the first toe of the contralateral limb (site 4, supine) was 1.13 (0.58 1.33). Fig. 3. Comparison of the orthostatic response between the pooled control groups (C) and patients with CLI before vascular reconstructive surgery (P). The values are plotted in a logarithmic scale. For explanation see figure 2. Figure 4 illustrates the variation in OR for the 3 sites following vascular reconstructive surgery, demonstrating a trend towards normal values (p < 0.05, Kruskal-Wallis test). For sites 2 and 3 the median OR got below 0.8 in the first postoperative period (4 7 days). Normalization at site 1 was not achieved until the second postoperative control (60 90 days). The increase in venous transmural pressure at the foot level by the present method is dependent on the length of the leg from the knee joint to the foot. For young controls the increase was 40 (36 48) mmhg, older controls 39.0 (36 45) mmhg and patients 35 (26 42) mmhg. For all subjects studied, the increase was more than 25 mmhg. Discussion Changing the limbs with CLI from horizontal to dependent position resulted in an increased skin perfusion at all measuring sites of that limb. The same manoeuvre in the control groups elicited a reduction of skin perfusion. These findings indicate that lowering of the limb in subjects with normal circulation induced arteriolar constriction, whereas this mechanism was abolished in the ischemic foot. Postural induced anteriolar constriction in the skin is a well-known physiological phenomenon [5]. The VAR is regarded as a homeostatic response of the blood circulation to changes in the position of the lower limb. Probably this is one of the factors limiting oedema formation in the dependent lower limb [7,12]. Lowering of the limb leads to an increased venous transmural pressure and venous distension [9,10,27]. This in turn supposedly evokes a sympathetic axon reflex from the veins to the arterioles causing arteriolar constriction [9,10]. Patients with CLI relieve their pain at rest by lowering the ischemic foot. This movement increases blood perfusion because of an abolished VAR [12,24,27,30]. Increased transmural pressure may furthermore distend the vessels
154 Stranden et al.: Spatial Variation of the Veno-arteriolar Reflex in CLI Fig. 4. Variation in orthostatic response at the three sites of the critically ischemic foot before (Preop.), and after vascular reconstructive surgery. Postop. 1 is 4 7 days and Postop. 2 is 60 90 days, respectively postoperatively. The values are plotted in logarithmic scales. For explanation see figure 2. (arterioles, capillaries and venules), there by reducing vascular resistance [19]. Both arteriolar dilatation and elevation of venous pressure due to lowering the afflicted limb [21], increases capillary pressure, which tends to enhance transudation of fluid through the capillary wall. These may be important causative factors for the development of ischaemic oedema frequently present in patients with CLI [14,15,26]. The finding of no difference in OR between the sites in controls indicates that the VAR induces similar vascular responses despite difference in vascular anatomy. In the pulp of the toe the distribution of arteriovenous anastomoses (AVA) is abundant. The occurrence of AVA is less or absent in sites 2 and 3. Because of AVAs, the local perfusion in the toe is greater than in other skin areas [17], as it consists of both nutritive and thermoregulatory components. Since the responses in the three sites are equal, this means that the VAR affects both vascular beds similarly. There was no difference in OR between young and old controls. The lack of correspondence between OR and age has previously been established [4] using 133 Xenon clearance technique. Since present or reduced VAR has been found in patients with peripheral arterial occlusive disease (PAOD) without CLI [2,27], and absent in legs with CLI, we may assume that the abolished VAR is due to local ischaemia and not age per se. Median OR at the pulp of first toe of CLI limbs was more than three times greater than at the level of the second metatarsal body and at the ankle, while there was no difference in OR between the latter two sites. In the control limbs OR did not differ between the respective levels. The value of the OR was greatest toward the distal parts of the foot where ischaemia usually is most pronounced. In supine position these areas tend to have a lower perfusion pressure because of the vertical position relative to the heart. The difference in pressure between the toe and ankle may be as large as 10 mmhg, and at very low pressures, critical closure of the capillaries at the toe level may be induced, while capillaries at lower parts (e.g. ankle) remain open. When the foot is lowered, increased transmural pressure dilates the capillaries and local vascular resistance is reduced. Hence, because of very low perfusion in supine position, the increase in perfusion (and in OR) at the toe level during dependency may be considerably larger than in proximal areas. The cause of disturbance of VAR in the limbs with CLI is still unclear. The disturbance could either be due to diminished contractile properties of the smooth muscle cells in the arteriolar wall, by disturbances in nervous reflex mechanisms, or both [10,11,23]. The returning of VAR after successful arterial reconstructive surgery indicates that the disturbance is not due to an irreversible damage. Furthermore, a temporarily impaired precapillary sphincter function could be explained by the effect of local hypoxia and/or an inflammatory process and subsequent formation of vasoactive metabolites like oxygen-free radical [16,28]. In the present study we did not compensate for biological zero, which is the amount laser Doppler signal remaining after proximal suprasystolic cuff occlusion [3,13,32]. Prolonged cuff occlusion in these patients was very painful, and was therefore regarded unethical to perform since the effect of biological zero compensation on the ratio calculation is small. At OR values 1.0 there would have been no change, at OR >1.0, the compensated value would have been shifted upwards, and downwards at OR <1.0. Compensation hence would increase the difference between OR of patients with CLI and the controls. In conclusion, the present study demonstrated that VAR is absent in the three measured sites of the limbs with CLI. There were regional differences in these ischaemic feet and
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