Non-genomic activities of retinoic acid receptor alpha control actin cytoskeletal events in human platelets.

UNLABELLED: Essentials Platelets employ proteins/signaling pathways traditionally thought reserved for nuclear niche. We determined retinoic-acid-receptor alpha (RARα) expression and function in human platelets. RARα/actin-related protein-2/3 complex (Arp2/3) interact via non-genomic signaling in platelets. RARα regulates Arp2/3-mediated actin cytoskeletal dynamics and platelet spreading. SUMMARY: Background Platelets utilize proteins and pathways classically reserved for the nuclear niche. Methods We determined whether human platelets express retinoic-acid-receptor family members, traditionally thought of as nuclear transcription factors, and deciphered the function of RARα. Results We found that RARα is robustly expressed in human platelets and megakaryocytes and interacts directly with actin-related protein-2/3 complex (Arp2/3) subunit 5 (Arp2/3s5). Arp2/3s5 co-localized with RARα in situ and regulated platelet cytoskeletal processes. The RARα ligand all-trans retinoic acid (atRA) disrupted RARα-Arp2/3 interactions. When isolated human platelets were treated with atRA, rapid cytoskeletal events (e.g. platelet spreading) were inhibited. In addition, when platelets were cultured for 18 h in the presence of atRA, actin-dependent morphological changes (e.g. extended cell body formation) were similarly inhibited. Using in vitro actin branching assays, RARα and Arp2/3-regulated complex actin branch formation was demonstrated. Consistent with inhibition of cytoskeletal processes in platelets, atRA, when added to this branching assay, resulted in dysregulated actin branching. Conclusion Our findings identify a previously unknown mechanism by which RARα regulates Arp2/3-mediated actin cytoskeletal dynamics through a non-genomic signaling pathway. These findings have broad implications in both nucleated and anucleate cells, where actin cytoskeletal events regulate cell morphology, movement and division.

Protein synthesis by platelets: historical and new perspectives.

In the late 1960s, numerous investigators independently demonstrated that platelets are capable of synthesizing proteins. Studies continued at a steady pace over the next 30 years and into the 21st century. Collectively, these investigations confirmed that platelets synthesize proteins and that the pattern of protein synthesis changes in response to cellular activation. More recent studies have characterized the mechanisms by which platelets synthesize proteins and have shown that protein synthesis alters the phenotype and functions of platelets. Here, we chronologically review our increased understanding of protein synthetic responses in platelets and discuss how the field may evolve over the next decade.

Differential regulation of matrix metalloproteinase-9 by monocytes adherent to collagen and platelets.

Circulating monocytes adhere to platelets and matrix proteins at sites of vascular injury, where engagement of specific surface tethering molecules mediates outside-in signaling and synthesis of gene products by the leukocytes. Here we demonstrate that interaction of isolated human monocytes with collagen induces matrix metalloproteinase-9 (MMP-9; gelatinase B) synthesis by monocytes, a process that is greatly enhanced in the presence of platelets. MMP-9 is a potent matrix degrading enzyme implicated in atherosclerotic plaque rupture, aneurysm formation, and other vascular syndromes. Synthesis of MMP-9 by monocytes is tightly regulated and synergistically increased following adhesion to collagen and platelets. Adhesion to control matrix proteins alone did not result in MMP-9 protein production and, similarly, adhesion of monocytes to platelets activated with thrombin in suspension was not sufficient to induce MMP-9 synthesis in the absence of monocyte adhesion to collagen. Interruption of intercellular contact between platelets and monocytes dramatically inhibited MMP-9 synthesis. These observations demonstrate that discrete adhesion-dependent signaling pathways govern MMP-9 synthesis by monocytes. The synthesis of MMP-9 by monocytes may be critical in vascular syndromes and other pathological processes that are dependent on dysregulated cell-cell and cell-matrix interactions.

Integrins regulate the intracellular distribution of eukaryotic initiation factor 4E in platelets. A checkpoint for translational control.

Recent evidence from our laboratory demonstrates that platelets synthesize numerous proteins in a signal-dependent fashion (Pabla, R., Weyrich, A. S., Dixon, D. A., Bray, P. F., McIntyre, T. M., Prescott, S. M., and Zimmerman, G. A. (1999) J. Cell Biol. 144, 175-184; Weyrich, A. S., Dixon, D. A., Pabla, R., Elstad, M. R., McIntyre, T. M., Prescott, S. M., and Zimmerman, G. A. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 5556-5561). Protein synthesis in platelets is controlled at the translational level; however, the mechanisms of regulation are not known. Here we demonstrate that translation initiation factors are redistributed to mRNA-rich areas in aggregated platelets, an event that induces protein synthesis. Interrogation of cDNA arrays revealed that platelet-derived mRNAs are primarily associated with the cytoskeletal core. In contrast, eukaryotic initiation factor 4E (eIF4E), the essential mRNA cap-binding protein that controls global translation rates, is localized in the membrane skeleton and soluble fraction of platelets, physically separated from most mRNAs. Platelet activation redistributes eIF4E to the cytoskeleton and increases interactions of eIF4E with mRNA cap structures. Redistribution of eIF4E to the mRNA-rich cytoskeleton coincides with a marked increase in protein synthesis, a process that is blocked when intracellular actin is disrupted. Additional studies demonstrated that beta(3) integrins are the primary membrane receptor that distributes eIF4E within the cell. These results imply that integrins link receptor-mediated pathways with mRNA-rich cytoskeletal domains and thereby modulate the organization of intracellular translational complexes. They also indicate that the functional status of eIF4E is regulated by its intracellular distribution.

Flow-induced DNA synthesis requires signaling to a translational control pathway.

BACKGROUND: The mTOR translational control pathway that signals to the P70/P85 S6 kinase (pp70(S6k)) is essential for mitogenesis. We have previously shown that pp70(S6k) is activated by fluid flow. We hypothesized that oscillatory fluid flow in the absence of exogenous mitogens would induce endothelial cells to synthesize DNA via activation of the mTOR pathway. For comparison, we also studied the ERK1/2 transcriptional signaling pathway. METHODS: Confluent human umbilical vein endothelial cells (HUVECs) were exposed to oscillatory flow (12 dyn/cm(2) peak shear stress; 3.3 Hz) or kept static in serum-deprived culture medium. Rapamycin or PD98059 was used to inhibit pp70(S6k) or ERK1/2 activation, respectively. RESULTS: Oscillatory flow activated both the pp70(S6k) and ERK1/2 signaling pathways. Rapamycin blocked activation of pp70(S6k) but not ERK1/2, while PD98059 blocked ERK1/2 but not pp70(S6k). DNA synthesis, as measured by [3H]thymidine uptake, increased by approximately twofold (P < 0.01) in HUVEC cultures exposed to oscillatory flow compared with those kept static. Rapamycin completely abolished the flow-induced increase in DNA synthesis while PD98059 did not. Oscillatory flow upregulated expression of cyclin-dependent kinases 1 and 4 mRNA in a temporal pattern consistent with cell cycle entry; rapamycin also inhibited these changes. CONCLUSIONS: Oscillatory flow activates both the ERK 1/2 and pp70(S6k) signaling pathways in HUVECs and induces DNA synthesis in the absence of other exogenous mitogens. Complete blockade of [3H]thymidine uptake by the mTOR pathway inhibitor rapamycin indicates that separate and distinct signaling to a translational control pathway is necessary to mediate flow-induced DNA synthesis by endothelial cells. Oscillatory flow-induced endothelial proliferation may contribute to atherogenesis.

Fluid flow activates a regulator of translation, p70/p85 S6 kinase, in human endothelial cells.

Cellular phenotype is determined not only by genetic transcription but also by subsequent translation of mRNA into protein. Extracellular signals trigger intracellular pathways that distinctly activate translation. The 70/85-kDa S6 kinase (pp70(S6k)) is a central enzyme in the signal-dependent control of translation, but its regulation in endothelial cells is largely unknown. Here we show that fluid flow (in the absence of an exogenous mitogen) as well as humoral agonists activate endothelial pp70(S6k). Rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR), and wortmannin, a phosphatidylinositol 3-kinase inhibitor, blocked flow-induced pp70(S6k) activation; FK-506, a rapamycin analog with minimal mTOR inhibitory activity, and PD-98059, an inhibitor of the flow-sensitive mitogen-activated protein kinase pathway, had no effect. Synthesis of Bcl-3, a protein whose translation is controlled by an mTOR-dependent pathway, was induced by flow and inhibited by rapamycin and wortmannin. Transcriptional blockade did not abolish the flow-induced upregulation of Bcl-3. Fluid forces may therefore modify endothelial phenotype by specifically regulating translation of certain mRNA transcripts into protein.

Endovascular-assisted versus conventional in situ saphenous vein bypass grafting: cumulative patency, limb salvage, and cost results in a 39-month multicenter study.

OBJECTIVE: In this retrospective multicenter study, the results of a minimally invasive method of endovascular-assisted in situ bypass grafting (EISB) versus "open" conventional in situ bypass grafting (CISB) were evaluated with a comparison of primary and secondary patency, limb salvage, and cost. METHODS: Enrolled in this study were 273 patients: 117 underwent CISB (42 femoropopliteal, 75 femorocrural) and 156 underwent EISB (41 femoropopliteal, 115 femorocrural). EISB was performed with an angioscopic Side Branch Occlusion system and an angioscopically guided valvulotome. All the patients underwent follow-up examination with serial color-flow ultrasound scanning. RESULTS: Both groups had similar comorbid risk factors for diabetes mellitus, coronary artery heart disease, hypertension, and cigarette smoking. The primary patency rates (CISB, 78.2% +/- 5% [SE]; EISB, 70.5% +/- 5%; P =.156), the secondary patency rates (CISB, 84.1% +/- 4%; EISB, 82.9% +/- 5%; P =.26), and the limb salvage rates (CISB, 85.8%; EISB, 88.4%; P =.127) were statistically similar, with a follow-up period that extended to 39 months (mean, 16.6 months; range, 1 to 40 months). In veins that were less than 2.5 to 3.0 mm in diameter, the EISB grafts fared poorly, with an increased incidence of early (12-month) graft thromboses (CISB, 10 grafts, 8.5%; EISB, 24 grafts, 15.3%). However, wound complications (CISB, 23%; EISB, 4%; P =.003), mean hospital length of stay (CISB, 6.5 days +/- 4.83; EISB, 3.2 days +/- 3.19; P =.001), and mean hospital charges (CISB, $25,349 +/- $19,476; EISB, $18,096 +/- $14,573; P =.001) were all significantly reduced in the EISB group. CONCLUSION: The CISB and EISB midterm primary and secondary patency and limb salvage rates were statistically similar. In smaller veins (< 2.5 to 3.0 mm in diameter), however, EISB is not appropriate because overly aggressive instrumentation may cause intimal trauma, with resultant early graft failure. With the avoidance of a long leg incision in the EISB group, wound complications and hospital length of stay were significantly reduced, which lowered hospital charges and justified the additional cost of the endovascular instruments. When in situ bypass grafting is contemplated, EISB in appropriate patients is a safe, minimally invasive, and cost-effective alternative to CISB.

An assessment of the current applicability of the EVT endovascular graft for treatment of patients with an infrarenal abdominal aortic aneurysm.

OBJECTIVE: To determine the percentage of elective abdominal aortic aneurysms (AAAs)/aortoiliac aneurysms that currently can be repaired with endovascular grafts (EVGs), the reasons for rejection of EVGs, and the future role of EVG in the treatment of AAA. METHODS: From January 1997 to May 1998, patients at three hospitals (a university hospital, a university-affiliated teaching hospital, and a Veterans Administration hospital with university faculty and residents) were evaluated for EVGs as part of a national clinical trial with grafts manufactured by Endovascular Technologies (EVT, Menlo Park, Calif). All patients at two hospitals and patients treated by the participating surgeons at the third hospital were screened for EVG. Patients with AAAs that were ruptured, symptomatic, or involved renal or mesenteric arteries and patients who declined treatment were excluded from the study. Evaluation included clinical examination, computed tomography scan, and selective arteriography. The decision to proceed with EVG was made by the vascular surgeon, with input and concurrence of medical personnel from a company with extensive experience in endograft repair. The main outcome measures were the determination of the percentage of elective AAAs currently being treated with an EVG and the reasons for exclusion of patients from EVG placement. RESULTS: A total of 162 patients underwent elective treatment of an AAA, 22 (14%) with an EVG (14 bifurcated, eight tube) and 140 (86%) with traditional resection. Indications for not proceeding with an EVG included insufficient proximal cuff in 29 patients (21%), distal common iliac aneurysm or insufficient distal iliac neck in 29 patients (21%), proximal neck too large for an EVG in 24 patients (17%), symptomatic iliac stenosis in 23 patients (16%), iliac stenosis precluding introducer passage in 17 patients (12%), patient preference in 11 patients (8%), and calcification, kink, or extensive thrombus involving the proximal neck precluding safe graft attachment in seven patients (5%). Of the 22 patients treated with an EVG, three were converted to open resection, because of iliac stenosis in two patients and premature stent deployment in one patient (initial technical success rate, 86%). CONCLUSION: Based on currently available technology, 80% of patients were not candidates for an EVG because of proximal calcification, short aortic or distal cuff, coexisting distal iliac aneurysm, and stenotic iliac disease. Even with the use of adjunctive procedures, most patients still require open repair. Significant changes in design will be necessary to apply these devices to most patients with an AAA.

PDGFbeta receptor blockade inhibits intimal hyperplasia in the baboon.

BACKGROUND: We have evaluated the use of a mouse/human chimeric anti-platelet-derived growth factor-beta receptor antibody in combination with heparin to inhibit intimal hyperplasia in the saphenous artery of the baboon after balloon angioplasty. METHODS AND RESULTS: The study evaluated lesion development in sequential injuries made 28 days apart. Each animal received control treatment after the first injury and antibody/heparin therapy after the second injury to the contralateral artery. The antibody was administered by bolus intravenous injections (10 mg/kg) on study days 1, 4, 8, 15, and 22 and heparin coadministered by continuous intravenous infusion at a dose of 0.13 mg/kg per hour. Morphometric analysis of tissue sections showed a 53% decrease in intimal area after antibody/heparin treatment (P=0.005), corresponding to a 40% decrease in the intima-to-media ratio (P=0.005). Smooth muscle cell proliferation in the injured wall, measured at both 4 and 29 days after balloon injury, were similar in the control and antibody/heparin-treated animals. CONCLUSIONS: These data suggest that platelet-derived growth factor plays a key role in the development of intimal lesions at sites of acute vascular injury in the nonhuman primate.

Is smooth muscle growth in primate arteries regulated by endothelial nitric oxide synthase?

PURPOSE: We investigated whether control of constitutive endothelial cell nitric oxide synthase (cNOS) and nitric oxide (NO) by changes in shear stress might be important for the regulation of smooth muscle cell (SMC) growth and vascular diameter. METHODS: Bilateral femoral arteriovenous fistulas were placed in baboons to increase the blood flow in the external iliac arteries. At 2 months, the fistula was ligated on one side to restore normal flow (flow switch). RESULTS: In response to flow switch and a decrease in shear stress, iliac artery lumenal area decreased and SMC proliferation was induced. A decline in NO production, cNOS messenger RNA (mRNA), and protein were associated with these biological effects. In a subset of animals with iliac arteries under high flow, infusion of N(omega)-nitro-L-arginine, an inhibitor of cNOS, did not induce proliferation. CONCLUSION: Shear stress can regulate cNOS, vasoconstriction, and SMC proliferation. A decrease in nitric oxide may be necessary, but is not sufficient to induce SMC proliferation in response to a decrease in blood flow.

Revision of reversed infrainguinal bypass grafts without preoperative arteriography.

PURPOSE: To determine whether graft revision on the basis of a duplex scan alone without an arteriogram is effective in identifying graft stenosis and allowing for repair to preserve bypass graft patency. METHODS: From 1994 to 1997, all patients in whom infrainguinal grafts were placed at a University-affiliated teaching hospital were entered into a prospective protocol using duplex scanning to detect stenotic lesions. Studies were performed after the operation, at 1 month, at 3 months, and every 3 months thereafter. All grafts were composed of reversed autogenous vein and were placed subcutaneously to allow for easier monitoring and correction. Patients who had failing grafts underwent operative correction without preoperative arteriography. RESULTS: During this interval, 48 lesions in 31 grafts were repaired. The indication for repair was a velocity ratio greater than 2.5 in all patients and greater than 3.0 for 43 lesions. Forty-four lesions had a peak systolic velocity greater than 250 cm/sec. Twenty-nine lesions reduced the distal graft velocity to less than 45 cm/sec. Sixteen lesions involved the proximal anastomosis, 26 the body of the graft, three the distal anastomosis, two involved inflow arteries, and one affected the outflow vessel. Repair included patch angioplasty for 39 lesions, resection with interposition graft for five, a proximal jump graft for three, and a distal extension graft for one. The severity and location of the stenosis was confirmed at operation in all cases. Twenty-eight of the 31 patients (90%) are currently alive, and follow-up on these patients has ranged from 5 to 36 months (mean, 19 months). Twenty-nine of the 31 grafts (94%) remained patent, with a 92% patency rate by life table analysis at 3 years. Follow-up duplex scans found improvement in the ankle-brachial index (mean increase, 0.33) and distal graft velocity (mean increase, 43 cm/sec) in all patients. After repair, 27 patients had a distal graft velocity greater than 45 cm/sec and no patient had a velocity ratio greater than 1.5. Complications included wound infection in two patients and bleeding that required reoperation in one. All symptomatic patients had clinical improvement, and none required early reexploration for residual stenosis. CONCLUSIONS: Graft repair may be safely performed on the basis of duplex scanning alone with preservation of bypass patency and correction of hemodynamic deterioration. Duplex scanning can detect inflow or outflow disease in addition to intrinsic graft stenoses and can identify sequential lesions, eliminating the need for, expense of, and risk of arteriography.

Acute reductions in blood flow and shear stress induce platelet-derived growth factor-A expression in baboon prosthetic grafts.

Abrupt reductions in fluid shear stress induce subendothelial smooth muscle cells (SMCs) to proliferate in experimental prosthetic grafts. Platelet-derived growth factor (PDGF), an important SMC mitogen, is expressed by cultured endothelial cells and modulated by shear stress. We hypothesized that this growth factor would be modulated by changes in shear stress in vivo. Bilateral aortoiliac prosthetic grafts were implanted into five baboons. High flow was generated by construction of femoral arteriovenous fistulas on both sides. Two months later, one of the fistulas was ligated, reducing shear stress in the upstream graft by 78 +/- 6%. Four days after fistula ligation, all grafts were removed and analyzed. As previously reported, SMC proliferation in low-flow grafts exceeded that in high-flow grafts, although the neointimal area was similar. mRNA levels for PDGF-A were significantly increased in low-flow grafts compared with high-flow grafts. In situ hybridization and immunohistochemical studies localized the increased PDGF-A mRNA and protein to the luminal endothelium and subjacent SMCs. Abrupt reductions in blood flow and fluid shear stress may induce accelerated neointimal thickening by a PDGF-A-mediated mechanism, since endothelial expression of this gene is temporally and anatomically associated with neointimal SMC proliferation.

Pharmacologic intervention to prevent graft failure.

Lower extremity vascular grafts, either vein or synthetic, fail for diverse reasons. Technical defects or poor surgical judgment doom a graft beyond any benefit pharmacotherapy can offer. Graft failure due to spontaneous thrombosis particularly affects prosthetic conduits, and use of antiplatelet agents (dextran, ASA) or anticoagulants (heparin, warfarin) is probably useful in this setting. An effective way to inhibit vein graft or anastomotic intimal hyperplasia remains elusive. Perhaps the most permanent and longstanding influence on lower extremity graft survival can be made through risk factor intervention aimed at arresting the progression of atherosclerosis. Aggressive treatment of hyperlipidemia, hypertension, smoking, and other known risk factors should be routinely and aggressively pursued in patients with lower extremity grafts, either autogenous or prosthetic. Lower extremity graft patency is optimally ensured by technically adept insertion of a proper autologous conduit in a well-selected patient. Pharmacotherapy may have a significant adjunctive role in the maintenance of graft patency, especially in high-risk settings such as limb salvage with associated poor outflow, a marginal vein graft, or the obligatory use of prosthetic material.

Short-stay carotid endarterectomy is safe and cost-effective.

BACKGROUND: Carotid endarterectomy (CEA) is conventionally performed following a contrast arteriogram, under general anesthesia, and with postoperative admission to an intensive care unit (ICU). We investigated whether any of these traditional adjuncts to CEA is necessary. PATIENTS AND METHODS: Eighteen consecutive patients had CEA performed according to a protocol of duplex scanning only, operation under regional anesthesia, and admission to the ICU only in cases of a proven need for services unique to the ICU (group I). Utilization of preoperative arteriography, admission to the ICU, postoperative complications, total hospital length of stay, and hospital charges were calculated for this group and results were compared with a group of 178 patients undergoing conventional CEA (arteriography, general anesthesia, routine ICU admission) during the same period (group II). RESULTS: In group I, 1 patient (6%) underwent preoperative arteriography and 4 patients (22%) were admitted to the ICU after CEA. Most group II patients (114 of 178, or 64%) underwent preoperative arteriography and virtually all (175 of 178, or 98%) were admitted to the ICU. Compared with group II, the average hospital length of stay for group I was significantly shorter (1.3 +/- 0.1 versus 3.1 +/- 0.3 days, P = 0.03) and hospital charges were significantly reduced ($5,861 +/- 229 versus $11,140 +/- 729, P = 0.02). CONCLUSIONS: This pilot study suggests that CEA can be safely performed without routine preoperative carotid arteriography; that routine ICU admission is unnecessary for the majority of cases; and that elimination of routine arteriography and ICU admission can reduce hospital charges for CEA by nearly one half.

Regional expression of the platelet-derived growth factor and its receptors in a primate graft model of vessel wall assembly.

Healing baboon polytetrafluoroethylene grafts express PDGF mRNA in the neointima. Perfusates of graft segments also contain PDGF-like mitogenic activity. To extend these findings, we studied the expression and regional distribution of the PDGF protein isoforms and their receptors in this prosthetic graft model. By immunohistochemistry, as well as ELISA and Western blot analysis of tissue extracts, both PDGF-A and PDGF-B were identified in macrophages within the interstices of the synthetic material. In contrast, the neointima contained predominantly PDGF-A localized to the endothelial surface and the immediate subjacent smooth muscle cell layers. Tissue extracts of neointima and graft material were mitogenic for baboon aortic smooth muscle cells in culture; nearly all of this proliferative activity was blocked by a neutralizing anti-PDGF antibody. PDGF receptor beta-subunit mRNA and protein were easily detectable in the neointima and graft material. PDGF receptor alpha-subunit mRNA was also observed in the graft matrix and at lower levels in the neointima. This pattern of ligand and receptor expression further implicates locally produced PDGF as a regulator of neointimal smooth muscle cell growth in this model. The coexpression of ligand and receptor in the macrophage-rich matrix also suggests that PDGF may participate in the foreign body response.

Accuracy of duplex scanning for measurement of arterial volume flow.

This study examined the accuracy of duplex ultrasound measurements of volume flow in a baboon model. Volume flow (Vf) through the external iliac artery was calculated from measurements of blood velocity averaged over several cardiac cycles (time-averaged velocity [TAV]) and vessel cross-sectional area (A) measured from the B-mode image: Vf = TAV x A. Fourteen anesthetized baboons were studied with a duplex scanner with a 7 MHz imaging transducer and 5 MHz pulsed Doppler. B-mode ultrasound measurements of external iliac artery diameters (2.5 +/- 0.2 mm) were used for calculation of cross-sectional area. Timed blood collections obtained through a cannula inserted into the common femoral artery and TAV measurements were obtained simultaneously during 6 to 15-second intervals. These measurements were repeated three to five times per animal with different flow rates each time. Flow rates ranged from 56 to 280 ml/min (170 +/- 54 ml/min). Average velocity was 55 +/- 17 cm/sec. There was no significant difference between the two methods of volume flow measurement (Student t test). Linear regression analysis revealed a high degree of correlation (r = 0.90, slope 0.95, and p = 0.0001). The absolute percentage error was 13% +/- 8%. Volume flow measured by duplex scanning correlates highly with timed blood collections. This method has potential application for the evaluation of diseased arteries and bypass grafts whose rates of flow and waveform patterns are similar to those of this experiment.

Increased blood flow inhibits neointimal hyperplasia in endothelialized vascular grafts.

Intimal hyperplasia is a primary cause of failure after vascular reconstruction and may be affected by blood flow. We have studied the effects of increased blood flow on intimal hyperplasia in porous polytetrafluoroethylene grafts implanted in baboons. These grafts develop an endothelial lining by 2 weeks and neointimal thickening due to proliferation of underlying smooth muscle cells by 1 month. Creation of a distal arteriovenous fistula increased flow (from 230 +/- 35 to 785 +/- 101 ml/min, p less than 0.001) and mean shear (from 26 +/- 4 to 78 +/- 10 dynes/cm2, p less than 0.001) without causing a drop in pressure across the grafts. Fistula flow did not alter the pattern of endothelial coverage but did cause a marked reduction in the cross-sectional area of the neointima (from 2.60 +/- 0.52 to 0.42 +/- 0.07 mm2 at 3 months, p less than 0.01). Detailed morphometric analysis revealed an equivalent percentage decrease in smooth muscle cells and matrix content, suggesting that the primary effect of increased flow was to reduce smooth muscle cell number without affecting the amount of matrix produced by individual cells. The neointima remained sensitive to changes in flow at late times; ligation of the fistula after 2 months resulted in a rapid increase in neointimal thickness (from 0.60 +/- 0.03 mm2 after 2 months of fistula flow to 3.88 +/- 0.55 mm2 1 month after ligation of fistula, p less than 0.01). These results support the hypothesis that changes in blood flow affect the structure of diseased as well as normal vessels.

Shear stress regulates smooth muscle proliferation and neointimal thickening in porous polytetrafluoroethylene grafts.

High shear stress appears to decrease wall thickening in diseased arteries and vascular grafts. To determine if increased shear stress diminishes smooth muscle (SMC) proliferation, we studied the effect of increased blood flow on neointimal thickening in porous polytetrafluoroethylene grafts implanted in baboons. An aorto-aortic 5-mm graft was placed in tandem with a pair of aorto-iliac 5-mm grafts, so that the proximal graft supplied all flow to both distal grafts. At 12 weeks, calculated luminal shear stress in proximal grafts was twice that in distal grafts (24 +/- 8 versus 11 +/- 5 dynes/cm2; p less than 0.05). All grafts were completely endothelialized. The neointimal cross-sectional area in proximal grafts was about half as large as in distal grafts (3.36 +/- 1.61 versus 5.93 +/- 0.61 mm2; p less than 0.05). Proximal grafts also had significantly less SMC proliferation (0.14 +/- 0.05% versus 0.24 +/- 0.10%; p less than 0.05) and SMC volume (6.1 +/- 4.0 versus 12.4 +/- 2.6 mm3/cm graft; p less than 0.01) when compared with distal grafts. We conclude that the elevation in shear stress in the proximal graft, which remained within the physiological range, inhibits SMC proliferation and neointimal thickening in these grafts.