INTRODUCTION

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CIRCUMFERENTIAL WALL STRESS and wall shear stress are considered to be important driving forces during development of the arterial wall (10, 27). They also modulate arterial wall structure in the adult. Elevation of transmural pressure, as in chronic hypertension, leads in many cases to an increase in arterial wall mass (8). The consequence of this hypertrophy on lumen diameter depends on the anatomic location of the vessel and on the etiology of the hypertension (5, 12). That different processes influence arterial wall mass and arterial lumen diameter may help in understanding the inward or outward nature of arterial remodeling (22). With respect to the regulation of arterial structural lumen diameter, several studies (14, 36) have shown that chronic elevation of blood flow leads to a widening of large arteries. Less is known about the effects of chronic blood flow reduction. They have been investigated in large arteries without changes of transmural pressure (3, 17) and also in poststenotic arteries that experience reductions in both flow and transmural pressure (7). Nonetheless, few studies have investigated the long-term effects of blood flow alterations on the structure of small resistance-sized muscular arteries (33, 34). These may be of particular interest from both the pathophysiological and mechanistic point of view. Small collateral vessels can play an important role in tissue perfusion after large artery occlusion (4, 26). Unlike large arteries, the density of smooth muscle cells is particularly high in small muscular arteries. This component of the arterial wall may be more susceptible to structural modulation than the extracellular matrix components such as collagen and elastin.

In the present study, we evaluated structural changes in rat mesenteric small arteries exposed for 4 wk to either an increase or decrease in blood flow. To this end we 1) ligated the connections to the arterial anastomosis for every other first-order side branch of the superior mesenteric artery that run parallel to the gut; 2) determined the acute (30 min) and chronic (4 wk) effects of these ligations on the blood flow through the trunks of the ligated and intermittent trees; 3) isolated first-order vessels that had been exposed to elevated or reduced flow for 4 wk; and 4) analyzed structural, mechanical, and contractile properties by organ chamber and histological techniques. For comparison, we also investigated vessels from 6-wk-old rats and 10-wk-old sham-operated (Sham) rats, representing the control conditions at initiation and termination, respectively, of the experimental intervention.

	MATERIAL AND METHODS

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Experimental groups. Six-week-old Wistar-Kyoto rats (Central Animal Facilities, Universiteit Maastricht, The Netherlands) that weighed 136 ± 8 g were randomly divided into three groups of seven animals each. In the first group, surgery was performed to modify blood flow in the mesenteric arteries (Fig. 1). In the second group, a similar surgical intervention was performed but without flow modification (Sham). The experimental and Sham groups were used 4 wk later. The third group, which was not subjected to surgery, was used for in vitro experimentation at 6 wk of age. The experimental protocols were approved by the Ethical Committee for Animal Welfare of the Universiteit Maastricht.

View larger version (23K): [in this window] [in a new window]   Fig. 1.   Schematic representation of mesenteric circulation of rat and location of blood flow measurements that were performed before and after ligation of arterial connections to peri-ileal anastomoses. This intervention resulted in a reduction in blood flow in stems of obstructed arterial trees [low-flow (LF) vessels] and an elevation in blood flow in nonligated vessels [high-flow (HF) vessels].

Blood flow measurements and arterial ligations. The animals were anesthetized with pentobarbital sodium (60 mg/kg ip), and a medial laparotomy was performed. Body temperature was maintained at 37.5°C by a thermostatically controlled heating platform. A section of the ileum was extracted and spread over a gauze swab that had been dampened with a sterile physiological salt solution (PSS). A segment of a first-order mesenteric artery side branch was gently freed of fat and connective tissue under a dissection microscope. With the use of a micromanipulator, a transit-time ultrasonic flow probe (0.5-mm V-series, Transonic Systems, Ithaca, NY) was placed around the artery (Fig. 1). Flow was determined with a T106 flowmeter (Transonic Systems) linked to a microcomputer. A zero-flow reading was obtained by transiently clamping the artery under investigation. Then the mean flow was determined over a 15-min period. Flow was measured again after ligation of the second-order arterial and venous branches near the gut (Fig. 1). These ligations were performed with 6-0 silk surgical thread and were applied on every other first-order mesenteric artery side branch (Fig. 1). When all ligations had been realized, blood flow was also measured in a parallel nonligated mesenteric arterial branch (Fig. 1). Similar surgery but without the ligations and similar flow measurements were performed in the Sham group.

The rats were caged in pairs and had free access to rat chow (Hope Farms, Woerden, The Netherlands) and tap water until their use 4 wk later. At this time, the rats were again anesthetized with pentobarbital sodium. The flow through the ligated and nonligated mesenteric arteries and through the first-order mesenteric artery side branches of Sham animals was determined, again with an acutely applied Transonic flow probe.

Systemic blood pressure. Three days before in vitro experimentation (i.e., 25 days after the first surgery), the rats were anesthetized with pentobarbital sodium (60 mg/kg ip), and an arterial catheter was introduced into the right femoral artery, positioned above the iliac bifurcation, and exteriorized at the neck. Two days later, the catheter was connected to a pressure transducer (CP-01, Century Technology, Inglewood, CA), and the analog signal was digitized. For each conscious animal, the value of the mean blood pressure was determined from the mean of 30-s readings obtained over 15 min.

Pressure myograph experiments. After the flow measurements at 4 wk after surgery were made, first-order mesenteric artery segments were isolated. For each animal in the experimental group, two segments were dissected, one corresponding to an artery with low flow (LF) and the other corresponding to an artery with high flow (HF). For the control and Sham groups, only one arterial segment was taken from each rat. All preparations were mounted in an arteriograph system (Living System Instrumentation, Burlington, VT), in which wall thickness and internal diameter could be continuously monitored while intraluminal pressure and flow were controlled (11). Segments were cannulated at both ends with a glass micropipette (internal diameter at tip 200 µm), tied to the pipettes with 11-0 surgical suture, and filled with filtered 1% albumin-PSS. The vessels were kept in a bath (7 ml) filled with warmed (37°C) and oxygenated (5% CO₂ in O₂) PSS. In the absence of flow, the pressure was increased to 50 mmHg. The pressure-servo system that controlled the intraluminal pressure was placed in the manual mode (i.e., without automatic control of pressure) to check for leaks in the cannulated preparation. Leaks were detected by a drop in pressure. In the absence of leaks, the pressure-servo system was set back to the automatic mode. The distance between the pipettes was set during pressurization at 150 mmHg and was adjusted by positioning the movable cannula to prevent warping, axial compression, or stretch of the vessel. Thereafter, the preparation was pressurized at 100 mmHg in the absence of flow and was allowed to equilibrate for 1 h.

The arteriograph system was placed on the stage of an inverted microscope (Nikon TMS) equipped with a black-and-white video camera (Stemmer). An electronic system (Living System Instrumentation) analyzed the signal obtained from the video image and continuously determined the internal diameter and wall thickness of the vessel. These parameters were recorded together with the intraluminal pressure with a chart recorder.

Experimental protocols. After the equilibration period, constrictions were studied by recording the decrease in the internal diameter in response to a high K⁺ solution (125 mM). After the preparations were washed with PSS until complete relaxation, a concentration-response curve was constructed by the cumulative addition of norepinephrine (NE; 0.01-10 µM) in the presence of cocaine (1 µM). Then the preparation was washed until a complete dilation was observed.

Spontaneous tone was assessed by recording the difference in arterial diameter before and after the addition of sodium nitroprusside (100 µM) to the bath solution. Thereafter, the bath solution was replaced by Ca²⁺-free PSS + 0.3 mM ethylene glycol-bis(-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), and stepwise increases in intraluminal pressure from 10 to 160 mmHg were applied, with an incremental pressure of 10 mmHg. Internal diameter and wall thickness of the preparation were measured at each pressure step. The intraluminal pressure was maintained for 1 min at each step.

Calculation of parameters. From parameters measured with the arteriograph system, other vascular parameters were calculated (11). Wall cross-sectional area (CSA) was defined as CSA = (D²_e  D²_i)/4, where D_e is the external diameter (in mm) and D_i the internal diameter (in mm). Wall tension was calculated as T = PD_i/2, where T is tension (in N/m) and P is intraluminal pressure (in N/m²). Circumferential wall stress (WS) was calculated with the formula WS = T/h, where h is wall thickness (in m). Wall shear rate (WSR) was calculated with the formula WSR = 4/(r³), where is blood flow (in ml/s) and r is the vessel radius (in cm).

Histology and morphometry. After contractile reactivity was recorded, the preparations were removed from the arteriograph and fixed without intraluminal pressure and longitudinal stretch. They were placed overnight in phosphate-buffered formaldehyde (4%), stored in ethanol, and embedded in paraffin.

Cross sections (4 µm) were stained with Lawson's solution (Boom, Meppel, The Netherlands) to visualize internal and external elastic laminae. Video images were generated from the cross section with a Zeiss axioscope (Zeiss) and a standard charge-coupled device camera (Sony). Internal and external media circumferences, demarcated by the internal and external elastic laminae and external circumference adventitia, were measured (Sigma Scan, Jandel Scientific, Corte Madera, CA). From these values, media and adventitia cross-sectional areas, internal radius, and media thickness were calculated for each section (2).

Additional cross sections were stained with eosin and hematoxylin, and the number of nuclear profiles in the medial and adventitial parts of the sections were counted separately at ×400 magnification by two independent investigators.

Biochemical analysis. The DNA content of first-order mesenteric side branches was determined with the fluorometric assay of Labarca and Paigen (16) with calf thymus DNA as internal standard. The method exploits the enhancement by DNA of the fluorescence of bisbenzimidazole (Hoechst 33258). The quantity of DNA (in ng) determined in the preparations was divided by the length of the vessel segments (in mm).

Solutions and drugs. The composition of the PSS was (in mM) 119 NaCl, 4.7 KCl, 2.5 CaCl₂, 1.2 MgSO₄, 25 NaHCO₃, 1.2 KH₂PO₄, and 5.5 glucose. In the high K⁺ solution, all NaCl was replaced by an equimolar amount of KCl. In Ca²⁺-free PSS, CaCl₂ was omitted from the normal PSS and 0.3 mM EGTA was added. The drugs used were pentobarbital sodium (Sanofi, Bordeaux, France), Hoechst 33258 (Biochem, Bruges, Belgium), and norepinephrine, cocaine, sodium nitroprusside, EGTA, calf thymus DNA, and bovine serum albumin (Sigma Chemical; St. Louis, MO). All drugs were dissolved in distilled water.

Data analysis. In all experiments, n equals the number of rats. Results are shown as means ± SE. Statistical significance of differences was evaluated with two-way analysis of variance followed by post hoc comparisons with Fisher's least significant difference test. A P value of <0.05 was considered significant.

	RESULTS

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Blood flow. In anesthetized 6-wk-old rats, blood flow in first-order mesenteric artery branches was on the order of 0.3 ml/min and did not differ among experimental groups (Table 1). After ligation of the connections to the mesenteric arterial anastomosis, a drastic reduction in blood flow (close to 90%) was observed in the upstream arteries. In the feeding vessel of the nonligated arterial trees, a marked increase in blood flow (close to 80%) was recorded (Table 1). Four weeks later, all rats were still alive, and flow changes were still present in the obstructed arteries and the unobstructed parallel vessels (Table 1). These vessels will be referred to as LF and HF arteries, respectively. The late changes in blood flow were comparable to those noted immediately after the ligations (Table 1). In Sham control rats, blood flow values had not changed during the 4-wk period.

Mean intra-aortic blood pressure, measured in awake conditions at the end of the experimental period, did not differ significantly between experimental (125 ± 4 mmHg; n = 7) and Sham animals (120 ± 2 mmHg; n = 7). Also, body weight did not differ among groups (268 ± 10 vs. 255 ± 11 g for experimental and Sham groups, respectively).

Vessel structure. Figure 2 illustrates that overall vessel wall structure was well preserved after 4 wk of modified blood flow. For instance, in neither HF nor LF vessels was disruption of the internal elastic lamina or a neointimal layer observed. Table 2 summarizes quantitative changes in vessel wall structure. No statistically significant differences were observed between 6-wk-old rats and 10-wk-old Sham control rats. In HF compared with Sham vessels, lumen diameter and media cross-sectional area were significantly increased. In LF compared with either Sham or HF vessels, lumen diameter, media and adventitia cross-sectional areas, and media thickness were significantly reduced.

View larger version (42K): [in this window] [in a new window]   Fig. 2.   Typical examples of histological cross sections (Lawson staining) of 1st-order mesenteric artery side branches from a 6-wk-old control (top) and a 10-wk-old sham-operated (Sham) rat (bottom middle) and vessels exposed for 4 wk to HF (bottom left) or LF (bottom right).

Total DNA content did not differ significantly between the mesenteric arteries of 6-wk-old rats and those of Sham rats (Fig. 3). In HF vessels, the DNA content was approximately twice as large as in Sham vessels (Fig. 3). The DNA content was, however, not modified in vessels that had been chronically exposed to reduced blood flow (Fig. 3). The number of nuclear profiles in both the medial and adventitial parts of the 4-µm-thick cross sections was significantly larger in HF than in Sham vessels (Table 2). In LF vessels, on the other hand, the number of nuclear profiles was not modified in the media and significantly increased in the adventitia compared with Sham vessels (Table 2). Nuclear density, assessed as the number of nuclear profiles per unit surface area, was significantly elevated in the media and adventitia of LF compared with Sham vessels.

View larger version (10K): [in this window] [in a new window]   Fig. 3.   Total DNA content (ng/mm vessel length) in mesenteric arteries from 6-wk-old control and 10-wk-old sham-operated rats and vessels exposed for 4 wk to HF or LF. Values are means ± SE. *** Significant difference from Sham rats, P < 0.001.

Pressure-diameter relationship. Figure 4 depicts the relationship between internal diameter and intraluminal pressure observed in isolated cannulated arteries incubated in Ca²⁺-free solution. Between 10 and 160 mmHg, the internal diameter did not differ between mesenteric arteries of 6-wk-old rats and those of Sham rats. Over this range of pressures, the diameters were significantly greater in HF and significantly smaller in LF than in Sham vessels.

View larger version (19K): [in this window] [in a new window]   Fig. 4.   Diameter-pressure relationships in isolated cannulated mesenteric arteries maintained at fixed axial length and incubated in Ca2+-free solution. , Vessels from intact 6-wk-old rats; , vessels from Sham rats; , vessels exposed for 4 wk to HF; , vessels from the same rats exposed for 4 wk to LF. Values are means ± SE; n = 6-7 rats.

Circumferential wall stress calculated at an in vitro intraluminal pressure of 100 mmHg did not differ significantly among HF (122 ± 8 N/m²), LF (110 ± 5 N/m²), and Sham vessels (125 ± 6 N/m²) and vessels from 6-wk-old intact rats (120 ± 4 N/m²). This was the case within a broad range of pressures (e.g., 60-160 mmHg) (data not shown). To estimate wall shear rate, the blood flow measured in vivo was combined with the internal diameters observed in vitro at 100 mmHg. This estimate did not differ significantly among HF (665 ± 96 s¹), LF (590 ± 106 s¹), and Sham vessels (692 ± 54 s¹) and vessels from 6-wk-old intact rats (745 ± 60 s¹).

Contractile reactivity. In HF, LF, and Sham vessels equilibrated at 100 mmHg in the presence of 2.5 mM Ca²⁺, the administration of 100 µM sodium nitroprusside and the removal of extracellular Ca²⁺ failed to induce significant dilatation (data not shown). This indicates the absence of basal tone in all three experimental groups.

All HF and Sham vessels constricted in response to a high K⁺ solution (125 mM) and to NE (10 µM,). Twenty-five percent of the LF vessels failed to respond to these vasoconstrictor stimuli and were not included in this study. Constrictor responses to K⁺ and NE did not differ between vessels from intact 6-wk-old rats and Sham control rats (Table 3, Fig. 5). The percent diameter reduction induced by K⁺ was significantly larger, but the accompanying reduction in media stress was comparable in HF compared with Sham vessels (Table 3). Constrictor responses to K⁺ were markedly smaller in LF than in Sham vessels. In HF vessels, the maximal response to NE was increased vs. control vessels when expressed as percent diameter reduction but not when expressed as change in media stress (Table 3, Fig. 5). In LF vessels, the maximal constrictor responses to NE were significantly reduced in both respects.

View larger version (16K): [in this window] [in a new window]   Fig. 5.   Cumulative concentration-response curves for vasoconstrictor effect of norepinephrine (NE) in isolated cannulated mesenteric arteries maintained at fixed axial length in presence of 2.5 mM Ca2+. , Vessels from intact 6-wk-old rats; , vessels from Sham rats; , vessels exposed for 4 wk to HF; , vessels from the same rats exposed for 4 wk to LF. Values are means ± SE; n = 6-7 rats.

	DISCUSSION

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In rat mesenteric small arteries that had been chronically exposed to altered blood flow, both the wall mass and lumen diameter were modified. Elevated blood flow resulted in outward hypertrophic remodeling and reduced blood flow in inward hypotrophic remodeling (32). These seem to involve different cellular changes.

The superior mesenteric artery supplies the arterial blood flow to most of the small intestine. In the rat, the third-order side branches of this vessel run parallel to the gut and interconnect with the ~20 first-order branches. Thanks to these anastomoses, most of the outflow of every other first-order mesenteric artery side branch could be obstructed without noticeable necrotizing injury to the ileum as has previously been reported (32, 33). Although flow was drastically reduced in the obstructed vessels, blood flow increased acutely to approximately the same extent in the parallel vessels that had not been obstructed. Four weeks after this intervention, the ligated vessels were still present and perfused, possibly due to the small side branches of these vessels that perfuse the mesentery. Blood flow was still drastically reduced in the ligated vessels, and blood flow was still markedly enhanced in the nonligated parallel vessels. In 10-wk-old rats that had been subjected at 6 wk of age to a sham procedure, blood flow did not differ significantly between first-order mesenteric artery side branches. Furthermore, wall structure, lumen diameter, and vasoconstrictor responses did not differ between vessels of 6-wk-old rats and those of 10-wk-old Sham rats. Collectively, these observations indicate that persistent reductions and elevations in blood flow could be induced in parallel mesenteric small arteries of the same rats and that these changes were superimposed on stable vessel wall structure and reactivity.

Measurements of lumen diameter and wall thickness in isolated cannulated arteries and morphometric measurements of immersion-fixed preparations were used to evaluate the structure of the vessels. These approaches indicate that both wall mass and lumen diameter were increased in the HF arteries and that both wall mass and lumen diameter were reduced in the LF vessels. Pressure-diameter curves indicate that the alteration of lumen diameter was statistically significant over a broad range of intraluminal pressures. These lumen diameter changes are primarily structural in nature because, for all experimental groups, removal of extracellular Ca²⁺ and a high concentration of sodium nitroprusside failed to modify lumen diameter. When evaluated at identical pressure (e.g., 100 mmHg), circumferential wall stress in HF and LF vessels did not differ from control values. Collectively, these observations indicate that 4 wk of elevated blood flow resulted in outward hypertrophic remodeling, that 4 wk of reduced blood flow resulted in inward hypotrophic remodeling, and that both sets of structural alterations led to a restoration of circumferential wall stress. The latter is in line with earlier findings in the circulation of the cremaster muscle (34, 35) and mesentery of the rat (32, 33) as far as adaptation to elevated flow is concerned. It is tempting to speculate that wall shear stress was also normalized by the structural responses to altered blood flow (14, 17, 36). This cannot be directly concluded from our observations but is supported by calculated wall shear rate based on flows and diameters measured in vivo and in vitro, respectively.

The increase in wall mass noted in the HF vessels was accompanied by a substantial increase in the DNA content of these vessels (+100%) and by a significant increase in the number of nuclear profiles in cross sections of the media and adventitia (+45 and +120%, respectively). The reduction in wall mass seen in the LF vessels was not accompanied by a reduction in DNA content, and a comparable number of nuclear profiles could be observed in the media of these vessels. Nonetheless, a marked increase in the number of nuclei was noted in the adventitia (+100%). The role of the adventitia in the overall structure of the blood vessel wall is largely unknown. Yet, because we observed different changes in adventitial and medial cells in LF vessels, future mechanistic analyses will require techniques with a high degree of spatial resolution rather than biochemical studies of the entire vessel segments. Because two independent techniques, aimed at changes in cell number, yielded conflicting results, analysis of the changes in cell number, cell volume, and ploidy in the present experimental setting will require more sophisticated techniques. That different cellular changes are involved in the structural responses to flow elevation and flow reduction is also indicated by the observed alterations in contractile reactivity. In the HF vessels, the absolute and relative reductions in lumen diameter evoked by either high K⁺ or a high concentration of NE were increased, but the accompanying reduction in media stress did not differ from that in vessels from Sham animals. So, in this condition, new contractile elements (e.g., cells) seem to have been added both in series and in parallel to the preexisting ones (23). In the LF vessels, the vasoconstrictor responses were markedly reduced, not only when assessed as an absolute reduction in diameter but also in terms of alteration in media stress. So, in the LF vessels, hyporeactivity to vasoconstrictor stimuli seemed to involve a reduction in both mass and contractility of the arterial smooth muscle.

The structural and functional changes that we noted in rat mesenteric small arteries exposed to altered blood flow (HF vessels: a 25% increase in lumen diameter, a 35% increase in media cross-sectional area, and a 100% increase in DNA content; LF vessels: a 40% reduction in lumen diameter, a 30% reduction in media cross-sectional area, and a 50% reduction in contractility) are large compared with earlier investigations (14, 17, 36) of flow-induced remodeling in large conduit arteries. Considerable diameter increases have previously been described for collateral vessels in the rabbit hindlimb after femoral arterial occlusion (19) and in the rat mesentery after ligation of multiple mesenteric artery side branches (32, 33). This could be due to the muscular nature of the small arteries and may be helpful for future analyses of the mechanisms underlying flow-induced structural changes. Most arterial vasomotor (15, 28) and structural responses (3, 17) to blood flow elevation have been observed to be endothelium dependent. Endothelium-derived mitogens (25) and macrophages recruited through enhanced endothelial expression of adhesion molecules (29) have been proposed to participate in flow-induced arterial remodeling. Another possibility, which is not mutually exclusive from the former, takes into account that vasomotor responses to changes in flow affect the mechanical factors within the vessel wall. First-order mesenteric artery side branches exhibit basal vasomotor tone (24) and experience a transmural pressure that is slightly but significantly smaller than aortic pressure (31). Reduction and elevation in tone will be accompanied by an increase and a decrease, respectively, in transmural pressure in the distal part of these vessels. In view of the size of the vessels, these changes in pressure may be anticipated to be rather small. On acute local elevation in blood flow, endothelium-derived mediators such as NO and prostaglandins induce vasodilatation (15). This entails an elevation in circumferential wall stress as lumen diameter increases and wall thickness decreases. This alteration, indirectly brought about by the endothelium, has been considered to be an important direct (21) or indirect (1) hypertrophic stimulus for the arterial wall. We propose that this mechanical factor overrules the potential antimitogenic effects of the acute endothelium-derived vasodilators NO and prostaglandins (6, 9, 30). In line with this, a recent investigation (13) of arteriovenous shunts in patients revealed a positive correlation between wall shear rate and mitogenicity of vascular smooth muscle cells. Conversely, on acute local reduction in blood flow, vasomotor tone is increased as a result of a reduced supply of endothelium-derived relaxing factors and possibly increased endothelial release of the potent vasoconstrictor and mitogenic substance endothelin (20). Circumferential wall stress is thereby reduced, and in the long run, atrophy of the arterial wall may develop. Also in this case, changes in wall stress are suggested to override the mitogenic effect of reduced and increased supply of NO and endothelin, respectively. The accompanying hyporeactivity could result from prolonged excessive vasoconstriction (2). Alternatively, this may be a consequence of long-term exposure to endothelin. In deoxycorticosterone acetate salt hypertensive rats, which exhibit elevated tissue and circulating levels of endothelin (18), mesenteric small artery contractile reactivity is reduced, and this can be prevented by chronic treatment with the endothelin-receptor antagonist bosentan (18).

We are not the first who evaluated structural changes in rat mesenteric small arteries in response to chronic changes in blood flow (32, 33). Still, our study revealed aspects that were not addressed before. These include 1) the possibility that structural adaptation to reduced flow evolves through mechanisms other than those to elevated flow, 2) changes in the adventitia, and 3) changes in contractile reactivity accompanying changes in arterial diameter.

In summary, we observed that after chronic exposure to elevated and reduced blood flow, circumferential wall stress was normalized by outward hypertrophic remodeling and inward hypotrophic remodeling, respectively. The former may involve an increase in arterial cell number, the latter a reduction in arterial smooth muscle cell volume and contractility. Wall mass changes in response to chronic changes in blood flow contrast with suspected effects on arterial smooth muscle cell growth of endothelium-derived vasoactive agents implicated in the acute control of wall shear stress.