Subsequent Intimal Hyperplasia Research Articles

α-Iso-cubebene attenuates neointima formation by inhibiting HMGB1-induced monocyte to macrophage differentiation via suppressing ROS production

α-Iso-cubebene (ICB) is a dibenzocyclooctadiene lignin contained in Schisandra chinensis, a medicinal herb used to improve cardiovascular symptoms. To investigate the mechanisms involved, the effects of ICB on cellular production of reactive oxygen species (ROS) was determined using cultured human THP-1 cells. When THP-1 cells were stimulated with HMGB1, cellular concentration of ROS was increased in dose- and time-dependent manners. These increases were significantly attenuated in cells pretreated with NADPH oxidase inhibitors, diphenyleneiodonium chloride and apocynin, but not by other inhibitors related to ROS generation in monocytes. The expression of constitutively expressed NADPH oxidase (NOX) subunits including NOX1, NOX2, NOX4 and NOX5 was not affected by HMGB1, but HMGB1-induced ROS production was exclusively attenuated in NOX2-deficient cells using siRNA, suggesting an enhanced NOX2 complex assembly. When cells were stimulated with HMGB1, p47phox phosphorylation at ser345, ser359 and ser370 was increased in dose- and time-dependent manners, which were significantly attenuated in ICB (3–10 μg/mL)-pretreated cells. In addition, HMGB1-induced monocyte–macrophage differentiation (MMD) in bone marrow-derived cells isolated from mice were significantly attenuated in cells treated with apocynin and ICB. Also, macrophage infiltration and intimal hyperplasia in the wire-injured femoral artery were significantly attenuated in ICB-treated mice compared to wild-type control mice. The results of this study show that ICB inhibits HMGB1-induced MMD by suppressing ROS production in monocytes, thus suggest that ICB has therapeutic potential for vascular inflammation with subsequent intimal hyperplasia related to vascular injury.

VOLUMETRIC ASSESSMENT OF INTRACORONARY PATHOLOGICAL FORMATIONS USING OCT IMAGING IN KAWASAKI DISEASE

Kawasaki disease (KD) can lead to coronary artery aneurysms with subsequent intimal hyperplasia, neovascularization, fibrosis, calcification, and macrophage accumulation. Intracoronary Optical Coherence Tomography (OCT) was found promising for characterization of coronary artery tissue layers in KD patients. Cross-sectional OCT images of coronary arteries play a crucial role to estimate the thickness variations of arterial wall layers, but the visualization is limited on a 2D plane. Automatic volumetric assessment of pathological formations of each tissue would be of high interest to quantify pathological formations. Hence, the goal of this study is to perform automatic registration of 2D OCT cross-sections for volumetric analysis of pathological formations in KD patients. In this study, 26 OCT cross-sectional pullbacks were obtained from different pediatric patients with KD using ILUMIEN ® OCT system. Each pullback sequence consists of ∼100 images per patient. We designed a model to compound 2D images of coronary artery tissues using a Convolutional Neural Network (CNN). CNNs were found to be very effective techniques to extract automatically specific and discriminative attributes of various tissues from OCT images. A fully-automated tissue classification model is used to characterize intima, media, neovascularization, fibrosis, and calcification using CCN and deep learning algorithms. Similarity between deep features extracted from different coronary artery tissues is used for registration. Paired t-test is applied between every two consequent frames of each pullback to validate tissue alignment before and after applying CNN-based registration. Alpha is set to 0.05 and significant difference was observed if the p-value was less than 0.05. The 3D models (figure) show a good alignment of intracoronary tissue layers and pathological formations with very low p-values (-log10 (p-value) = 34.35 ± 26.41 and significant difference of 0.03 ± 0.01 for all patient’s pullback). We applied our technique to create 3D reconstruction and analyze volume variations of coronary artery layers and pathological formations caused by KD on coronary artery tissues. By doing so, we can create an automated diagnostic framework providing clinicians with an operator-independent diagnosis of histological coronary lesions. From a visual 3D rendering of qualitative findings, we can generate a quantitative diagnostic output. This study contributes to estimate volumetric variations of various pathological formations caused by KD on coronary artery tissue layers. A visual 3D rendering of qualitative findings, together with a quantitative diagnostic report can greatly enhance patient care and management.

BLTR1 in Monocytes Emerges as a Therapeutic Target For Vascular Inflammation With a Subsequent Intimal Hyperplasia in a Murine Wire-Injured Femoral Artery

Given the importance of high-mobility group box 1 (HMGB1) and 5-lipoxygenase (5-LO) signaling in vascular inflammation, we investigated the role of leukotriene signaling in monocytes on monocyte-to-macrophage differentiation (MMD) induced by HMGB1, and on vascular inflammation and subsequent intimal hyperplasia in a mouse model of wire-injured femoral artery. In cultured primary bone marrow-derived cells (BMDCs) stimulated with HMGB1, the number of cells with macrophage-like morphology was markedly increased in association with an increased expression of CD11b/Mac-1, which were attenuated in cells pre-treated with Zileuton, a 5-LO inhibitor as well as in 5-LO-deficient BMDCs. Of various leukotriene receptor inhibitors examined, which included leukotriene B4 receptors (BLTRs) and cysteinyl leukotriene receptors (cysLTRs), the BLTR1 inhibitor (U75302) exclusively suppressed MMD induction by HMGB1. The importance of BLTR1 in HMGB1-induced MMD was also observed in BMDCs isolated from BLTR1-deficient mice and BMDCs transfected with BLTR1 siRNA. Although leukotriene B4 (LTB4) had minimal direct effects on MMD in control and 5-LO-deficient BMDCs, MMD attenuation by HMGB1 in 5-LO-deficient BMDCs was significantly reversed by exogenous LTB4, but not in BLTR1-deficient BMDCs, suggesting that LTB4/BLTR1-mediated priming of monocytes is a prerequisite of HMGB1-induced MMD. In vivo, both macrophage infiltration and intimal hyperplasia in our wire-injured femoral artery were markedly attenuated in BLTR1-deficient mice as compared with wild-type controls, but these effects were reversed in BLTR1-deficient mice transplanted with monocytes from control mice. These results suggest that BLTR1 in monocytes is a pivotal player in MMD with subsequent macrophage infiltration into neointima, leading to vascular remodeling after vascular injury.

Abstract 238: Sub-Endothelial Matrix Targeted Liposomal Nanoparticles For Vascular Therapeutics

Objectives: Vascular intervention results in intimal denudation, exposure of the sub-endothelial matrix, and subsequent intimal hyperplasia (IH), under the control of numerous remodeling mechanisms. Reduction of IH-induced restenosis may be achieved by manipulation of these remodeling pathways through targeted molecular inhibition. Spatially-controlled nanoparticles designed to co-localize to exposed sub-endothelial matrices could provide an optimal delivery system for targeted vascular therapeutics. To his end, we aimed to develop the framework for a surface-modified liposomal drug delivery platform designed to preferentially bind collagen type IV. Methods: Non-targeted control liposomes (NTL) were formed with bulk DOPC-PEG, 30% cholesterol, and 0.1 mol% Rhodamine-DOPE. DSPE-PEG-DBCO lipids were conjugated to peptides previously shown to bind collagen IV via copper-free click chemistry and inserted at 5 mol% to form collagen-targeting liposomes (CTL), either at hydration (PreCTL) or by post-insertion via micellar transfer (PostCTL). Peptide conjugation was confirmed by MALDI-TOF, and liposomes were characterized by DLS and electrophoretic mobility. Liposome binding was assayed on collagen IV matrices dried at 3ug/cm 2 and quantified by fluorescence at 0-24hr static 37°C incubation. Results: All liposome formulations exhibited a narrow size distribution (~100nm) and neutral-low positive charge. CTLs demonstrated a significant increase in binding vs. NTLs (Fig1). Conclusions: CTLs demonstrated significant affinity for collagen IV binding in a static environment compared with NTLs. Future studies aim to optimize the binding capacity of CTLs via further lipid modifications and under flow conditions mimicking vessel wall hemodynamics. Considering the efficacy demonstrated here, CTLs show promise as the framework for a spatially-controlled drug delivery platform for future application in targeted vascular therapeutics.

The Relation between the Level of Serum Tumor Necrosis Factor – Alpha and Hemodialysis Adequacy in Diabetic and Non Diabetic Patients on Maintenance Hemodialysis

1.1.Background: Hemodialysis is still the most common renal replacement therapy (RRT) modality in end stage renal disease patients (ESRD), the first problem to be faced when choosing hemodialysis for patients with ESRD is the vascular access, dia­lysis delivery should be adequate not only to improve quality of life but also to prolong sur­vival, quality of life adjusted for life expectancy defined kt/v of 1.3 as the optimal cost-effective dialysis, An ideal access delivers a flow rate to the dialyzer adequate for the dialysis prescription, has a long use-life, and has a low rate of complications (eg, infection, stenosis, thrombosis, aneurysm, and limb ischemia). Of available accesses, the surgically created fistula comes closest to fulfilling these criteria, working fistula must have all the following characteristics; blood flow adequate to support dialysis which usually equates to blood flow greater than 600 ml/min, a diameter greater than 0.6 cm, with a location accessible for cannulation and a depth of approximately 0.6 cm (ideally between 0.5 and 1cm) from the skin surface. In hemodialysis patients with an arteriovenous fistula (AVF), access failure is primarily due to fistula stenosis, which predisposes to thrombosis and subsequent access loss. The risk for access failure differs individually, Fistula stenosis is histologically characterized by endothelial cell injury and intimal hyperplasia induced by factors like TNF-α, which could induce proliferation of vascular smooth muscles leading to subsequent intimal hyperplasia. Resulting in fistula stenosis and subsequent access failure. TNF-alpha influences the risk for hemodialysis access failure in diabetic ESRD patients there is advanced calcified atherosclerosis which leads to frequently inadequate arterial inflow and eventually also to venous run-off problems. So ESRD patients with diabetes have worse access survival rates and hemodialysis adequacy. 1.2.Methods:The study was conducted to 60 ESRD patients divided to group I (30 Diabetic ESRD patients on HD) and group II (30 Non - diabetic ESRD patients on HD). We estimate serum TNF-alpha in all patients. Assess AVF by Doppler U/S and estimate hemodialysis adequacy by using single pool Kt/v. 1.3.Results: We found that serum TNF-alpha level is significantly elevated in diabetic group, Kt/v is significantly decreased in diabetic group, AVF vein diameter was statistically significantly decreased in diabetic group, we also found that TNF-alpha was statically positively significant with duration of dialysis, FBG and lastly we found that TNF-alpha may affect hemodialysis adequacy adversely particularly in diabetic ESRD patients on HD 1.4.Conclusion:TNF-alpha is significantly elevated in diabetic ESRD patients on HD, vein diameter was significantly decreased in diabetic ESRD patients on HD.TNF-alpha is significantly positive correlated with duration of dialysis and duration of arteriovenous fistula so TNF -alpha may affect hemodialysis adequacy adversely particularly in diabetic patients on HD.

Cycling related common femoral artery disease: an unusual case in an otherwise healthy male

A cyclist reported gradual onset, bilateral leg claudication without any atherosclerotic risk factors and with a normal vascular examination. Duplex ultrasound and magnetic resonance angiography revealed bilateral focal stenoses due to coralline plaques of the common femoral arteries (CFAs) but normal proximal and distal vasculature. A surgical endarterectomy with vein patch angioplasty resolved all symptoms. Cyclist's iliac syndrome (whereby mechanical and anatomical factors in competitive cyclists and runners lead to arterial kinking and subsequent intimal hyperplasia, endofibrosis and, ultimately, stenoses of the external iliac arteries) is well documented. This is the first report to our knowledge of a similar process isolated to the CFA.

Preliminary Study of the Effects of Smectite Granules (WoundStat) on Vascular Repair and Wound Healing in a Swine Survival Model

WoundStat (WS) (TraumaCure, Bethesda, MD) is a topical hemostatic agent that effectively stops severe hemorrhage in animal models. To the best of our knowledge, no survival study has been conducted to ensure long-term product safety. We evaluated vascular patency and tissue responses to WS in a swine femoral artery injury model with survival up to 5 weeks. Anesthetized swine received a standardized femoral artery injury with free hemorrhage for 45 seconds followed by WS application. One hour after application, the WS was removed, the wound copiously irrigated, and the artery repaired using a vein patch. Six groups of three animals received WS and were killed either immediately after surgery or at weekly intervals up to 5 weeks. Three control animals were treated with gauze packing and direct pressure followed by identical vascular repair and survival for 1 week. At the time of killing, angiograms were performed, and tissue was collected for histopathology. Hemostasis was complete in all WS animals. All animals survived the procedure, and there were no clinically evident postoperative complications. Vascular repairs were angiographically patent in 15 of 18 animals (83%) receiving WS. Histopathologic examination of WS animals revealed severe diffuse fibrogranulomatous inflammation, early endothelial degeneration with subsequent intimal hyperplasia, moderate myocyte necrosis, and fibrogranulomatous nerve entrapment with axonal degeneration. Although an effective hemostatic agent, WS use was associated with a substantial local inflammatory response and neurovascular changes up to 5 weeks postinjury.

Porphyrin-Photosensitized Processes: Their Applications in the Prevention of Arterial Restenosis

Photodynamic therapy (PDT) is based on the use of a photozensitising compound which is accumulated by rapidly proliferating cells. Subsequent irradiation with light wavelengths specifically absorbed by the photosensitiser promotes the generation of reactive short-lived oxygen species which cause an irreversible and selective damage. Endovascular interventions to correct obstructive arterial disease have been developed worldwide with excellent short term results. However, long term patency is still limited by the onset of restenosis, due to subsequent intimal hyperplasia (IH). IH is characterized by proliferation and migration of smooth muscle cells (SMC) and extracellular matrix production. Targeting of SMC by photozensitisers can be efficiently achieved by taking advantage of the receptors for low density lipoproteins (LDL) expressed by such cells. Thus, preference is given to hydrophobic compounds which readily partition in the lipid matrix of LDL. We developed a liposomal formulation of a highly hydrophobic photozensitising agent, Zn(II)-phthalocyanine (ZnPc). The liposome-delivered ZnPc was readily taken up by cultured SMC cells and preferentially localized in the Golgi apparatus. Red light irradiation of incubated SMC induced cell death. Extension of these investigations to an in vivo rabbit model showed that ZnPc mainly accumulated in the media layer, where PDT induces the main damage through cellular depletion due to apoptosis of SMC, changes in the extracellular matrix with generation of a barrier to cellular migration, and acceleration of re-endothelization. Initial clinical applications showed that PDT safely and effectively prevents restenosis after angioplasty up to a 6 month follow-up.

Vein graft arterialization causes differential activation of mitogen-activated protein kinases

Objective Vascular injury results in activation of the mitogen-activated protein kinases—extracellular-signal regulated kinases, c-jun N-terminal kinase, and p38 MAPK—which have been implicated in cell proliferation, migration, and apoptosis. The goal of this study was to characterize mitogen-activated protein kinase activation in arterialized vein grafts. Methods Carotid artery bypass using reversed external jugular vein was performed in 29 dogs. Vein grafts were harvested after 30 minutes and 3, 8, and 24 hours, and 4, 7, 14, and 28 days. Contralateral external jugular vein and external jugular vein interposition vein-to-vein grafts were used as controls. Vein graft extracts were analyzed for extracellular-signal regulated kinases, c-jun N-terminal kinase, and p38 MAPK activation. Proliferating cell nuclear antigen expression was investigated as a parameter of cell proliferation. Apoptosis was assessed by terminal deoxynucleotidyl transferase–mediated 2′-deoxyuridine 5′-triphosphate nick end labeling staining and intimal hyperplasia by morphometric examination of tissue sections. Results Significant intimal hyperplasia was observed at 28 days. Over the time points studied, vein graft arterialization resulted in bimodal activation of both extracellular-signal regulated kinase and p38 MAPK (30 minutes through 3 hours; 4 days) but did not induce activation of c-jun N-terminal kinase. Proliferating cell nuclear antigen expression increased from days 1 through 28, and apoptosis increased between 8 and 24 hours. Conclusion Vein graft arterialization induces bimodal activation of extracellular-signal regulated kinase and p38 MAPK; however, in contrast with what is described in arterial injury, it does not induce c-jun N-terminal kinase activation. These results provide the first comprehensive characterization of the mitogen-activated protein kinase signaling pathways activated in vein graft arterialization and identify mitogen-activated protein kinases as potential mediators of vein graft remodeling and subsequent intimal hyperplasia.

Stent design related neointimal tissue proliferation in human coronary arteries; an intravascular ultrasound study.

Histological restenosis models in animals have indicated that stent design has a significant impact on vessel trauma during stent implantation and on the amount of subsequent neointimal tissue proliferation. The impact of different stent designs on intimal hyperplasia in human atherosclerotic coronary arteries has not been determined. Angiographic and intravascular ultrasound studies were performed at the 6 month follow-up in 131 consecutive native coronary lesions of 131 patients treated with 50 Multi-Link stents, 40 InFlow stents and 41 Palmaz-Schatz stents. Lumen and stent cross-sectional areas (CSA) were measured at 1 mm axial increments. Mean intimal hyperplasia cross-sectional area (stent CSA-lumen CSA) and mean intimal hyperplasia thickness were calculated. Intravascular ultrasound demonstrated different levels of intimal hyperplasia proliferation for the three stents. Mean intimal hyperplasia thickness was 0.16+/-0.08 mm for Multi-Link stents, 0.26+/-0.19 mm for Palmaz-Schatz stents and 0.39+/-0.14 mm for Inflow stents (P<0.001). Multivariate analysis proved that stent type was the only independent predictor of intimal hyperplasia thickness at follow-up (P<0.001). Coronary stent design has a significant impact on subsequent intimal hyperplasia after implantation into atherosclerotic human coronary arteries. The corrugated ring design of the Multi-Link stent proved to result in less tissue proliferation at 6-month follow-up than the tubular slotted design of Palmaz-Schatz and InFlow stents.

Effect of preintervention plaque burden on subsequent intimal hyperplasia in stented coronary artery lesions

We sought to determine if axial and circumferential distribution of plaque before stenting determines the axial and circumferential distribution of subsequent intimal hyperplasia (IH). We studied 22 patients with a single Palmaz-Schatz stent implanted in a native coronary artery, who underwent intravascular ultrasound (IVUS) imaging before intervention, after stenting, and at 6-month follow-up. For each lesion, 7 locations were analyzed: proximal and distal reference, proximal and distal edge of the stent, proximal and distal location within the body of the stent, and the articulation. Pre- and postintervention and follow-up image slices were precisely aligned and analyzed for pre- and postintervention plaque area and follow-up IH area and thickness. The location of maximal IH area was at or adjacent to the location of maximal preintervention plaque in 17 of 22 of the patients (77%). Similiarly, the circumferential distribution of IH at follow-up paralleled the eccentricity pattern of the native plaque burden in 69% (24 of 35 slices). Using multivariant analysis, the strongest predictor of IH was preintervention plaque area (p = 0.001). IH accumulates axially and circumferentially preferentially at the site of maximal preintervention plaque.

Effects of inflation pressure of balloon catheter on vascular injuries and subsequent development of intimai hyperplasia in rabbit aorta

Balloon catheter de-endothelialization is the most popular means of arterial injury in experimental animals and has been used as the model system to investigate atherogenesis and restenosis after percutaneous transluminal coronary angioplasty (PTCA). The aim of this study was to examine the relationship between balloon inflation pressure and vascular damage and also subsequent intimai hyperplasia. Retrograde pullback balloon injury of rabbit aortas was made at three different balloon pressures (1.5, 1.75, and 2.0 atm). The medial injuries, such as necrosis of smooth muscle cells and disruption of elastic lamina, were occasionally found in the injured segment of the aorta by balloon catheter at 1.75 atm and more frequently at 2.0 atm. No prominent medial injury was observed in the aortic segment to balloon catheter injury at 1.5 atm; Intimal hyperplasia developed in each animal and increased with time, 2, 4, and 8 weeks after injury. The intimal hyperplasia followed by balloon injury at 1.75 and 2.0 atm was more prominent than that at 1.5 atm, however, the development of the intimal hyperplasia was not parallel to the degree of inflation pressure. On the other hand, decrease of DNA content of the media and reduction of norepinephrine-induced vasoconstriction were observed in a pressure-dependent manner after balloon injury. These findings indicate that intimal hyperplasia is not proportionally correlated to the severity of the vascular injury. The control, of inflation pressure is very important in order to examine vascular injuries, subsequent intimal hyperplasia and vasomotor responses in animal models of balloon catheter injury.

Smooth muscle cell proliferation in the arterial intima after stretch injury: relationship between the severity of stretching and intima hyperplasia in New Zealand white rabbits.

We examined histologically whether the severity of arterial stretch injury is related to the degree of subsequent intimal hyperplasia. In six male New Zealand White rabbits, the common carotid artery was hyperextended with a 3F Fogarty balloon catheter. Two weeks later, no proliferative change was evident in the intima in the most hyperextended portion. Paradoxically, however, intimal hyperplasia due to smooth muscle cell proliferation was observed in the moderately extended portions. The intimal hyperplasia appeared to be exacerbated where the arterial stretching was more severe. It is concluded that the severity of arterial stretch injury is closely related to the intimal proliferation of smooth muscle cells.

Directional coronary atherectomy: How much lumen enlargement is optimal?

Until recently, it has not been clear how much of the effect of directional coronary atherectomy is due to tissue removal per se, and whether the long-term results of the procedure are helped or harmed when the operator attempts to obtain the “near zero percen” residual stenosis of which this technique is capable. This article summarizes the findings of a series of studies that have addressed these important questions and proposes a prescription for the optimal performance of directional atherectomy. Analysis of retrieved tissue weights compared with measured increases in luminal volume shows that about half of the improvement seen after directional atherectomy results from mechanical dilation. Because this “facilitated” dilation appears to take place within the bases of the trenches created by atherectomy cuts (rather than being randomly distributed in fractures throughout the plaque substance), a larger and smoother lumen is possible compared with that seen after conventional balloon dilation. Although the recovery of deep vessel wall components (media and even adventitia) is common, it generally does not cause either acute complications (i.e., perforation) or increase the probability of subsequent restenosis. Rather, reduction in the probability of late restenosis appears to be most directly related to the ability of directional atherectomy to provide the largest acute luminal diameter safely possible, thus providing better tolerance of subsequent intimal hyperplasia before hemodynamically significant renarrowing results at the treatment site.

Novel approach to the analysis of restenosis after the use of three new coronary devices

Restenosis after coronary intervention has remained a vexing problem despite the introduction of nearly 24 newer coronary interventional devices. To more clearly evaluate the potential impact of three such new devices on restenosis, coronary lumen diameters were measured before, immediately after and at 6 months after intervention, and restenosis was analyzed using continuous geometric techniques.Lumen diameters were measured before and immediately after intervention in 223 coronary vessels treated with one of three new devices: a single Palmaz-Schatz stent (n = 87), directional atherectomy (n = 125) and laser balloon angioplasty (n = 11); 184 (83%) of the patients underwent follow-up angiography 6 months after treatment. The immediate increase in lumen diameter produced by the intervention (acute gain) and the subsequent reduction in lumen diameter between the time of intervention to 6 month follow-up study (late loss) were examined. For each of the three interventions, the restenosis rate at follow-up study was analyzed using a traditional dichotomous definition (≥50% diameter stenosis), as well as a novel graphic technique.Although the apparent restenosis rates differed significantly among the three interventions (19% for stents, 31% for atherectomy and 50% for laser balloon angioplasty; p = 0.02), late loss among the three interventions was equivalent (average 1 mm; p = 0.91). There were, however, marked differences in the acute gain achieved by the three interventions: 2.6 mm for stents, 2.2 mm for atherectomy and 2 mm for laser balloon angioplasty; p < 0.001). It was these differences in acute gain rather than late loss that explained the observed differences in restenosis rate.By using a continuous rather than a dichotomous view of restenosis and resolving long-term device results in the components of acute gain and late loss, analysis of this patient set provides unique insights into the potential role of these new devices in reducing restenosis. The absolute magnitude of late loss in lumen diameter (reflecting local intimal hyperplasia) was equivalent among the three devices and was greater than the late loss previously reported for conventional balloon angioplasty. Therefore, the ability of a newer device to reduce restenosis may be related more to its ability to provide the greatest acute gain in lumen diameter, rather than the ability to reduce subsequent intimal hyperplasia.

Prevention of stenosis after vascular reconstruction: Pharmacologic control of intimal hyperplasia — A review

Prevention of stenosis after vascular reconstruction: Pharmacologic control of intimal hyperplasia — A review

VEIN ALLOGRAFTS FOR ARTERIAL REPLACEMENT IN RATS AN IMMUNOLOGICAL AND HISTOLOGICAL STUDY

Vein allografts were studied in the rat using the major histocompatibility complex-incompatible DA and Lewis inbred strains. Allografts were performed in the Lewis to DA and DA to Lewis combinations, with Lewis to Lewis isografts serving as controls. Vein grafts were performed by interposing a 1 cm length of fresh iliolumbar vein into a defect of the iliac artery, using microsurgical techniques. The grafts were under observation for 18 weeks for (1) patency, (2) gross structural changes, (3) histological changes, and (4) antibody responses. No immunosuppression was used. All grafts remained patient throughout the period of observation, although aneurysm formation was noted in some allografts toward the end of the observation period. Histologically, allografts and isografts were indistinguishable. In the first 2 weeks, they showed patchy areas of necrosis in the vein walls, with subsequent intimal hyperplasia and medial fibrosis. In both strain combinations a lymphocytotoxin response was induced in most animals, the response being particularly strong in the DA to Lewis combination. All cytotoxic activity could be absorbed out using red blood cells, suggesting that the specificity of the antibody was mainly or entirely directed against SD antigens.