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Objectives: One third of infrainguinal vein bypasses may fail within the first 1.5 years. Pro- and anti-inflammatory mechanisms are thought to be involved in these graft stenoses and occlusions. In previous studies, low levels of anti-phosphorylcholine IgM (anti-PC IgM, an innate anti-inflammatory IgM) have been associated with increased cardiovascular events. In this study, the peri-operative dynamics of anti-PC IgM levels were established during leg bypass surgery, and associations assessed between anti-PC IgM levels and primary graft patency., Design and Methods: This was a prospective, observational cohort study of infrainguinal autogenous vein bypass for peripheral arterial occlusive disease involving four university affiliated hospitals. Plasma cytokine and anti-PC IgM levels were measured pre- and post-operatively. The outcome of interest was loss of primary graft patency because of occlusion or intervention for graft stenosis., Results: One hundred and forty-two consecutive patients were enrolled: mean age 66 (46-91); 91% white race and male; 72.5% critical limb ischaemia (Fontaine III or IV). Median pre-operative anti-PC IgM levels were 49 units/mL (IQR 32.3-107.7, mean 89.8 + 101 sd). During follow up of an average of 1.8 years (1 month-7.4 years), 50 (35.2%) grafts lost primary patency. Pre-operative levels of interleukin 6 or C-reactive protein did not predict graft failure. Patients with pre-operative anti-PC IgM values in the lowest quartile had a twofold increased risk of graft failure (multivariable Cox proportional hazard, p = .03, HR 2.11, 95% CI 1.09-4.07), even after accounting for the other significant factors of conduit diameter, distal anastomosis, smoking, and the severity of leg ischaemia., Conclusions: Low levels of anti-PC IgM are associated with vein bypass graft failure. This biological mediator may be a useful marker to identify patients at higher risk, and offers the potential for novel, directed therapies for vascular inflammation and its consequences., (Copyright © 2018 European Society for Vascular Surgery. Published by Elsevier B.V. All rights reserved.)

Arteriovenous fistula (AVF) remodeling is an active area of research in vascular biology given the high rates of primary failure, complications, and cost burden for the health-care system. Comorbidities such as diabetes and different types of vascular disease are known to influence AVFs outcomes. However, little is known about the effects of immunosuppression, particularly human immunodeficiency virus (HIV) infection, on AVF primary failure and patency. This retrospective study assessed the impact of HIV infection and T-cell counts on AVF outcomes. Using a retrospective cohort of 495 patients, we compared the risk of AVF primary failure and primary unassisted patency on HIV-positive and nonimmunocompromised individuals using logistic regressions and Cox proportional hazard models. Within the HIV-infected subset (n = 43), we analyzed the association between immunological parameters such as T-cell counts and primary failure. Positive predictors of primary failure were HIV infection [odds ratio (OR) = 3.09, P = 0.002] and history of a previous AVF (OR = 2.18, P = 0.003). However, there was no difference in primary unassisted patency between HIV-positive and negative individuals after excluding primary failure cases. There was no association between T-cell subset counts and AVF outcomes. Our results indicate that HIV-positive individuals have a higher risk of AVF primary failure than nonimmunocompromised patients. However, this increased susceptibility is not explained by the degree of immunosuppression., Competing Interests: None

Congenital heart disease is a leading cause of death in the newborn period, and man-made grafts currently used for reconstruction are associated with multiple complications. Tissue engineering can provide an alternative source of vascular tissue in congenital cardiac surgery. Clinical trials have been successful overall, but the most frequent complication is graft stenosis. Recent studies in animal models have indicated the important role of the recipient׳s immune response in neotissue formation, and that modulating the immune response can reduce the incidence of stenosis., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The presence of occlusion/near-occlusion of glomerular capillaries was recently added to the existing definition of glomerulitis (g). We retrospectively re-evaluated 135 renal allograft biopsies regarding g to ensure no antibody-damaged grafts were missed. Previous and revised g scores (pg and rg, respectively) were compared for clinicopathologic correlations. The g score did not change in 100 (74.1%) biopsies. Thirty-five (25.9%) biopsies were changed to a lower score. Sensitivity and specificity of pg and rg for the presence of donor-specific antibodies (DSA) were 76% vs. 58% and 70% vs. 79%, respectively. Pg score indicated graft loss with 65% sensitivity and 63% specificity, whereas rg showed 46% sensitivity and 71% specificity. Area under the curve (AUC) values in ROC analysis for DSA and graft loss were as follows: pg, 0.773; rg, 0.693; and pg, 0.635; rg, 0.577, respectively. A comparison of the two AUC values revealed a significant difference between pg and rg only for DSA (P = 0.0076). Pg and post-transplant time of biopsy independently predicted graft loss, whereas rg did not. In conclusion, revised g scores showed lesser sensitivity but higher specificity for DSA and graft loss. Recent definition of g missed antibody-mediated rejection in few cases, and it was not an independent predictor for graft loss., (© 2017 Steunstichting ESOT.)

Percutaneous coronary intervention is widely adopted to treat patients with coronary artery disease. However, restenosis remains an unsolved clinical problem after vascular interventions. The role of the systemic and local immune response in the development of restenosis is not fully understood. Hence, the aim of the current study was to investigate the role of the human immune system on subsequent neointima formation elicited by vascular injury in a humanized mouse model. Immunodeficient NOD.Cg-Prkdc scid IL2rg tm1Wjl (NSG) mice were reconstituted with human (h)PBMCs immediately after both carotid wire and femoral cuff injury were induced in order to identify how differences in the severity of injury influenced endothelial regeneration, neointima formation, and homing of human inflammatory and progenitor cells. In contrast to non-reconstituted mice, hPBMC reconstitution reduced neointima formation after femoral cuff injury whereas hPBMCs promoted neointima formation after carotid wire injury 4 weeks after induction of injury. Neointimal endothelium and smooth muscle cells in the injured arteries were of mouse origin. Our results indicate that the immune system may differentially respond to arterial injury depending on the severity of injury, which may also be influenced by the intrinsic properties of the arteries themselves, resulting in either minimal or aggravated neointima formation.

Background: Arterial cryopreserved allografts are conduit of choice for arterial bypass in case of infection. They are sometimes submitted to accelerated degeneration: stenosis, thrombosis, or aneurysmal evolution. We hypothesized that ABO group and Rhesus compatibility could play a role in arterial cryopreserved allograft complications., Methods: Patients who benefit from allograft bypass from 2006 to 2011 were retrospectively compared, regarding ABO or rhesus compatibility and irregular agglutinins., Results: Seventy-two patients were included. Regarding ABO mismatch, there was no difference in terms of death (51% vs. 43%, P = 0.820), thrombosis (25% vs. 32%, P = 0.78), rupture (9% vs. 0%, P = 0.247), stenosis (3% vs. 12%, P = 0.331), aneurysmal degeneration (16% vs. 12%, P = 1), and 5 years of secondary patency rate (34% vs. 39%, P = 0.141). There was also no difference regarding Rhesus mismatch., Conclusions: Allograft degeneration does not seem to be related to ABO mismatch. This degeneration could be related to another way of immunogenicity, particularly Human Leukocyte Antigen mismatch, and needs further exploration., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Cardiac allograft vasculopathy (CAV) is a common complication following heart transplantation (HT), resulting in diminished graft survival. The preferred strategy for preventing CAV is optimal medical management; however, for patients who develop CAV, delaying disease progression through effective medication management is equally important. A review of the literature regarding medication management of CAV was conducted via a search of the MEDLINE database. Studies were included if they were published in English, conducted in humans ≥ 18 years of age or older, and used noninvestigational medications. Immunosuppressive medications such as the antiproliferative mycophenolate, the calcineurin inhibitor tacrolimus, and the proliferation signal inhibitors sirolimus and everolimus have been shown to prevent the development of CAV. Certain cardiovascular medications, such as HMG-CoA reductase inhibitors (statins), gemfibrozil, calcium channel blockers, and angiotensin-converting enzyme inhibitors, have also demonstrated efficacy in preventing this disease process. Prevention of CAV has also been observed with prophylaxis against cytomegalovirus infection and antioxidant medications. Despite being commonly used in HT patients, neither antiplatelet agents nor glycemic control have proved effective at preventing CAV. Only sirolimus has been shown to arrest the progress of existing CAV., (© 2015 Pharmacotherapy Publications, Inc.)

Copper (Cu)-bearing stainless steel with release of Cu2+ ions is a novel material for coronary stents that could reduce the in-stent restenosis after the stent implantation. The inflammation has been recently recognized as an important factor to smooth muscle cells proliferation, thrombosis, and hence the restenosis post-angioplasty. The objective of this study is to further investigate the effect and relevant mechanism of Cu-bearing stainless steel (316L-Cu SS) on the inflammation reaction after stent implantation. The results demonstrated that, compared with commercial coronary stent material (316L SS), 316L-Cu SS could inhibit the inflammation caused by endothelial dysfunction through blockading the inflammatory factors (TNF-α, IL-1β, 6, 8), which would then reduce the recruitment and infiltration of leukocytes, rather than have direct effect on leukocytes. This finding further explained the reduction effect of 316L-Cu SS on in-stent restenosis from a novel view.

Objective: To investigated whether CBS3830, a highly selectively inhibitor of p38MAPK, could ameliorate inflammation and intimal hyperplasia in arterialized vein grafts (AVGs)., Methods: Sixty male Sprague-Dawley rats underwent a reversed right jugular vein to common carotid artery interposition graft and were randomly treatment with vehicle (control) or single-dose (3 mg/kg, preoperative) or double-dose (3 mg/kg, preoperative and 4 d postoperative) CBS3830. Twenty rats underwent sham operation. The levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) were determined by ELISA. Vein grafts were analyzed by intimal/medial morphometry, proliferating cell nuclear antigen (PCNA) expression, and p38MAPK phosphorylation., Results: TNF-α, IL-1β, and IL-6 gradually increased then slowly decreased in AVG rats. However, at 4 d and 7 d, TNF-α levels decreased by 37.5% and 29.5% (p = 0.003, 0.05, respectively) in the single-dose CBS3830 group, and by 37.6% and 32.5%, respectively (both p = 0.003) in the double-dose group compared with those of control. IL-1β levels significantly reduced at 4 d and 14 d in both dosage groups. IL-6 levels significantly reduced at 7 d in both groups. Intima and medial thickening were significantly reduced in both dosage treated groups at 7, 14, and 28 d (all p = 0.000) compared to the controls. Further study showed CBS3830 inhibited p38MAPK phosphorylation and decreased PCNA expression., Conclusions: CBS3830 significantly decreases inflammation and intimal hyperplasia in AVGs.

Background: Heparin, the standard perioperative anticoagulant for the prevention of graft vessel thrombosis in patients undergoing liver transplantation (LT), binds to the chemokine platelet factor 4 (PF4). Antibodies that are formed against the resulting PF4/heparin complexes can induce heparin-induced thrombocytopenia. LT is a clinical situation that allows the study of T-cell dependency of immune responses because T-cell function is largely suppressed pharmacologically in these patients to prevent graft rejection., Objectives: To investigate the immune response against PF4/heparin complexes in patients undergoing LT., Patients and Methods: In this prospective cohort study, 38 consecutive patients undergoing LT were systematically screened for anti-PF4/heparin antibodies (enzyme immunoassay and heparin-induced platelet aggregation assay), platelet count, liver function, and engraftment., Results: At baseline, 5 (13%) of 38 patients tested positive for anti-PF4/heparin IgG (non-platelet-activating) antibodies. By day 20, an additional 5 (15%) of 33 patients seroconverted for immunoglobulin G (two platelet-activating) antibodies. No patient developed clinical heparin-induced thrombocytopenia. Two of six patients with graft function failure had anti-PF4/heparin IgG antibodies at the time of graft function failure. Graft liver biopsy samples from these patients showed thrombotic occlusions of the microcirculation., Conclusions: Anti-PF4/heparin IgG antibodies are generated despite strong pharmacologic suppression of T cells, indicating that T cells likely have a limited role in the immune response to PF4/heparin complexes in humans., (© 2014 International Society on Thrombosis and Haemostasis.)

Background: Non-HLA antibodies (Abs) targeting vascular receptors are considered to have an influence on renal transplant injury. Anti-endothelin-1 type A receptor (anti-ETAR) antibodies were associated with cellular and antibody-mediated rejection and early onset of vasculopathy in heart transplant patients but their role in renal transplantation remains unclear. The aim of our study was to assess the incidence and importance of anti-ETAR antibodies and their impact on renal transplant during the first year observation., Methods: We evaluated the presence of anti-ETAR antibodies in 116 consecutive renal transplant recipients in pre- and post-transplant screening (before and in 1st, 3rd, 6th, 12th month after transplantation). Additionally, we assessed the presence of anti-HLA antibodies. Anti-ETAR antibodies were assayed by ELISA. The diagnosis of acute rejection was based on the Banff criteria., Results: Anti-ETAR antibodies were observed in 55 (47.4%) of the analyzed recipients before transplantation. The function of renal transplant was significantly worse in the anti-ETAR(+) group compared to the anti-ETAR(-) group during the first post-transplant year. One month after transplantation the serum creatinine in anti-ETAR (+) patients (pts) was 1.86±0.8mg/dl and 1.51±0.5 in anti-ETAR(-) pts (p=0.009). Twelve months after transplantation the difference between the groups was still observed 1.70±0.7 vs. 1.40±0.4 (p=0.04). Biopsy proven acute rejection was recognized in 8/55 (14.5%) in ETAR(+) and 9/61 (14.8%) in ETAR(-) patients but cases with mild to severe intimal arteritis (v1-v3) were more often observed in patients with the presence of anti-ETAR Abs 4/55 (7.2%) comparing with 1/61 (1.6%) in anti-ETAR(-) patients. The anti-ETAR antibody levels varied at different measurement intervals during the one-year follow-up., Conclusions: The presence of anti-ETAR antibodies is associated with a worse renal transplant function during the first 12months after transplantation. Including anti-ETAR antibodies in the diagnostics of renal transplant recipient immune status should be considered to provide comprehensive assessment of humoral alloimmunity., (© 2013. Published by Elsevier B.V. All rights reserved.)

The article deals with an analysis of the literature data concerning immunological mechanisms of the formation of restenoses after damage of the arterial wall, considering the participants of the early and late phases of inflammatory response initiated by endothelial damage. Also given is characteristics of the process of formation on the neointima, followed by description of the role of intercellular adhesion molecules in initiation and maintaining of the processes of acute and chronic inflammation.

Background: All humans have natural, protective antibodies directed against phosphorylcholine (PC) epitopes, a common inflammatory danger signal appearing at sites of cell injury, oxidative stress, and on bacterial capsules. In large human cohorts, low levels of anti-PC IgM were associated with a significantly increased risk of stroke or myocardial infarction. However, it is not known if these antibodies protect against the premature closure of arterial reconstructions., Methods: A prospective, observational study of patients undergoing elective, infrainguinal, autogenous vein bypasses for atherosclerotic occlusive disease of the legs was conducted. Clinical data were recorded prospectively, and preoperative levels of anti-PC IgM measured with the CVDefine kit from Athera Biotechnologies (Solna, Sweden). The principal clinical end point was the loss of primary patency (loss of graft flow, or any intervention for stenosis). Patients were followed regularly by duplex ultrasound at 1, 3, 6, 12, 18 months, and yearly thereafter., Results: Fifty-six patients were studied, for an average of 1.3 years. Indications for surgery were claudication (33.9%), ischemic rest pain (17.9%), and ischemia with ulceration or gangrene (48.2%). Seventeen (30.4%) patients experienced loss of primary patency (10 graft occlusions, seven surgical or endovascular revisions of graft stenoses). Kaplan-Meier survival analysis showed that the quartile of patients with the lowest anti-PC IgM levels had significantly worse primary graft patency (log-rank test, P = .0085). Uni- and multivariate Cox proportional hazards analysis revealed that the preoperative anti-PC IgM level was an important predictor of graft failure. Patients with IgM values in the lowest quartile had a 3.6-fold increased risk of graft failure (95% confidence interval: 1.1-12.1), even after accounting for other significant clinical or technical factors such as indication for surgery, site of distal anastomosis, or vein graft diameter., Conclusions: A naturally occurring IgM antibody directed against the proinflammatory epitope PC may be protective against vein graft stenosis and failure, through anti-inflammatory mechanisms. Measurement of this antibody may be a useful prognostic indicator, although larger studies of more diverse populations will be needed to confirm these results. The biological actions of anti-PC IgM suggest it may be useful in developing immunotherapies to improve bypass longevity., (Published by Mosby, Inc.)

Background: Anti-PF4/heparin antibodies are frequently generated after coronary artery bypass grafting (CABG) surgery, with platelet-activating IgG implicated in heparin-induced thrombocytopenia (HIT). It is controversial whether non-platelet-activating antibodies are associated with thrombosis., Objectives: To determine in post-CABG patients whether thromboprophylaxis using fondaparinux vs. unfractionated heparin (UFH) reduces the frequency of anti-PF4/heparin antibodies, and whether anti-PF4/heparin antibodies are associated with early graft occlusion., Methods/patients: In a pre-planned secondary analysis of a randomized control trial (RCT) comparing fondaparinux vs. UFH thromboprophylaxis post-CABG, we determined the frequency of anti-PF4/heparin antibody formation by solid-phase enzyme-immunoassay (EIA) and of platelet-activating antibodies by serotonin-release assay (SRA); the SRA and fluid-phase EIA were used to assess fondaparinux cross-reactivity. We also examined whether anti-PF4/heparin antibodies were associated with early arterial or venous graft occlusion (6-week CT angiography)., Results: We found no significant difference in the frequency of antibody formation between patients who received fondaparinux vs. UFH (65.3% vs. 46.0%; P = 0.069), and no significant fondaparinux cross-reactivity. Venous graft occlusion(s) occurred in 6/26 patients who formed 'strong' IgG antibodies (≥ 1.0 optical density [OD] units and ≥ 2× baseline) vs. 3/66 who did not (P = 0.0139). In both unadjusted and adjusted analyses, strong postoperative (but not pre-operative) anti-PF4/heparin IgG responses were associated with a markedly increased risk of early venous (but not arterial) graft occlusion (adjusted OR, 9.25 [95% CI, 1.73, 49.43]; P = 0.0093); notably, none of the three SRA-positive patients developed a venous graft occlusion., Conclusions: Fondaparinux vs. UFH thromboprophylaxis postCABG does not reduce anti-PF4/heparin antibody formation. Non-platelet-activating anti-PF4/heparin IgG antibodies generated post operatively are associated with early venous graft occlusion., (© 2012 International Society on Thrombosis and Haemostasis.)

Synthetic arteriovenous (AV) grafts, placed between an artery and vein, are used for hemodialysis but often fail due to stenosis, typically at the vein-graft anastomosis. This study recorded T lymphocyte and macrophage accumulation at the vein-graft anastomosis, suggesting a role for inflammation in stenosis development. Epoxyeicosatrienoic acids (EETs), products of cytochrome P-450 epoxidation of arachidonic acid, have vasculoprotective and anti-inflammatory effects including inhibition of platelet activation, cell migration, and adhesion. EETs are hydrolyzed by soluble epoxide hydrolase (sEH) to less active diols. The effects of a specific inhibitor of sEH (sEHI) on cytokine release from human monocytes and mouse bone marrow-derived macrophages (BMMΦ) from wild-type (WT) and sEH knockout (KO) animals were investigated. Expression of sEH protein increased over time at the anastomosis as evaluated by immunohistochemistry. Pre-exposure of adherent human monocytes to sEHI (5 μM) significantly inhibited lipopolysaccharide-induced release of monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor-α and enhanced the EET-to-diol ratio. Release of MCP-1 from WT BMMΦ was significantly inhibited but release from sEH KO BMMΦ was not attenuated indicating the specificity of the sEHI. In contrast, sEHI did not inhibit the release of macrophage inflammatory protein-1 or interleukin-6. Nuclear translocation of NF-κB, as assessed by immunocytochemical staining, was not decreased with sEHI in monocytes, but the phosphorylation of JNK was completely abrogated, suggesting this pathway is the target of sEHI effects in monocytes. These results suggest that sEHI may be useful for inhibition of inflammation and subsequently stenosis in AV grafts.

Objective: Infrainguinal autogenous vein grafts are especially prone to narrowing and failure, and both inflammatory and thrombotic pathways are implicated. Platelets and monocytes are the key thrombo-inflammatory cells that arrive first at sites of vascular injury. These cells have potent interactions that recruit and activate one another, propagating thrombotic and inflammatory responses within the vessel wall. We therefore hypothesized that elevated levels of platelet-monocyte aggregates (PMA) might be associated with stenosis, and could possibly discriminate between patients with or without vein graft stenosis., Methods: Thirty-six vascular surgery patients were studied, in a stable quiescent period after infrainguinal autogenous vein graft bypasses for occlusive disease. Eighteen patients had hemodynamically significant graft stenoses confirmed by imaging, and 18 were free from stenosis. The level of PMA in whole blood was quantified after blood draw using two-color flow cytometry. Three measurements were made per sample: the basal, in-vivo level of aggregates (baseline PMA); the predisposition to spontaneously generate PMA (spontaneous PMA); and PMA generation by the addition of exogenous thrombin receptor-activating peptide (stimulated PMA). The baseline, in-vivo level of PMA was estimated by immediate flow analysis. The predisposition to spontaneously generate PMA was measured after in vitro incubation. Responsiveness to thrombin stimulation of the blood was quantified by the in vitro dose response to an exogenous thrombin receptor-activating peptide (sfllrn)., Results: Baseline PMA levels were similar in patients with vein graft stenosis vs nonstenosis (14.8% ± 3.2 vs 10.1% ± 1.5, respectively, mean ± SEM). However, patients with stenosis showed higher spontaneous PMA levels (58.5% ± 4.5 vs 28.3% ± 4.3; P < .001) and higher stimulated PMA levels (P < .001; analysis of variance). Covariables of smoking, diabetes, statin, or antithrombotic therapy could not account for these differences., Conclusions: Platelet-monocyte reactivity may play a role in the development of vein graft stenoses. Those with/without stenosis differed primarily in their threshold, or predisposition to form aggregates (spontaneous PMA), while their basal circulating levels of PMA (baseline PMA) were similar. These measurements may unmask pathologic differences in thrombo-inflammatory responsiveness that are not apparent in basal measurements. Understanding the causes and mechanisms leading to abnormal platelet-monocyte responses may improve approaches to predicting or preventing vein graft stenosis., (Copyright © 2011 Society for Vascular Surgery. All rights reserved.)

Background: It is well established that primed/memory T cells play a critical role in heart transplant rejection. This contributes to the challenges faced in the transplant clinic because current treatments that are efficient in controlling naïve T cell alloresponses have limited efficacy on primed T cell responders., Methods: Fully MHC-mismatched heart transplantation was performed from BALB/c to C57BL/6 mice presensitized with BALB/c splenocytes 14 days pretransplantation. A combination therapy comprising CD70-, CD154-, and CD8-specific antibodies (Abs) was administered at day 0 and 4 posttransplantation with rapamycin on days 0 to 4., Results: The Ab combination therapy extended heart transplant survival in presensitized recipients from median survival time 8 days (MST) to MST 78 days. A decrease in the number of splenic interferon-gamma-secreting cells measured by ELISpot assay was seen in the treated group compared with the untreated controls. However, graft-infiltrating CD8+ and CD4+ T cells persisted despite treatment and the number of intragraft CD4+ T cells increased at day 30 posttransplantation. When an additional "rescue therapy" comprising the same Abs was readministered at days 30, 60, and 90 posttransplantation, T cell infiltration was reduced and indefinite graft survival was observed. Furthermore, rescue therapy resulted in gradual decrease in titer and, by day 90 posttransplantation, the complete loss of the preexisting, donor-specific Abs., Conclusion: We conclude that our Ab combination therapy extends allograft survival in presensitized recipients. When combined with intermittent Ab-mediated rescue therapy, this results in indefinite allograft survival and a loss of the preexisting, donor-specific Abs from the circulation.

Background: Vein graft disease is a chronic inflammatory disease and limits the long-term clinical outcome of coronary revascularization. Because calcitonin gene-related peptide (CGRP) inhibits macrophage infiltration and inflammatory mediators, we hypothesized that transfected CGRP gene would inhibit macrophage infiltration and expression of inflammatory mediators in vein graft disease., Methods: Autologous rabbit jugular vein grafts were incubated ex vivo in a solution of mosaic adeno-associated virus vectors containing CGRP gene (AAV2/1.CGRP) or Escherichia coli B-galactosidase gene (LacZ) or a saline solution and then interposed in the carotid artery. Expression of CGRP gene was identified by reverse transcription-polymerase chain reaction, and E. coli LacZ gene expression was identified by X-gal staining. Intima to media ratios were evaluated at postoperative 4 weeks. Macrophages were identified with CD68 antibody by immunocytochemistry. Inflammatory mediators were measured with real-time polymerase chain reaction., Results: The CGRP and LacZ gene expression were positive at postoperative 4 weeks. The intima to media ratio was significantly inhibited in the AAV2/1.CGRP group. Macrophage infiltration and expression of inflammatory mediators including monocyte chemoattractant protein-1, tumor necrosis factor-alpha, inducible nitric oxide synthase, and matrix metalloproteinase-9 were also significantly inhibited in the AAV2/1.CGRP group., Conclusions: Transfection of AAV2/1.CGRP inhibited inflammatory mediator expression, macrophage infiltration, and neointimal hyperplasia in experimental vein graft disease., (Copyright 2010 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Using a clinically relevant, fully disparate, allogeneic aortic transplant mouse model of allograft vasculopathy, we have demonstrated that neointimal proliferation is dependent on CD8(+) T cell effector pathways in the presence of therapeutic doses of calcineurin inhibitor (CNI) immunosuppression. CD4(+) T cell pathways are ablated by CNI immunosuppression. In the current study, we examined the relationship between CD8(+) T cell activities, medial SMC loss and neointimal hyperplasia. We demonstrate that at 5-6wk post transplantation in a wild type/wild type transplant CD8(+) T cell infiltration, CD8(+) CTL effector cell mediator expression and medial SMC loss all occur within aortic interposition grafts in the face of CNI immunosuppression. Both IFN-gamma and CTL mediated effector function is required for SMC loss and lesion formation under these conditions. Using strain combinations and reconstitution models, we provide data that blockade of the perforin/granzyme pathway does not prevent lesion formation but that blockade of the Fas/FasL pathway of cytotoxicity dramatically reduces SMC loss and prevents neointimal lesion formation. Both of these blockade strategies are in the face of an active IFN-gamma pathway. These data suggest a cooperative role between Fas/FasL and IFN-gamma mediated effector functions in medial SMC loss and neointimal lesion formation., (Copyright 2009 Elsevier B.V. All rights reserved.)

Objective: The aim of this 12 months observational study was to investigate risk factors of major adverse coronary events, such as death or Q wave myocardial infarction due to stent thrombosis or in stent restenosis., Material and Methods: One hundred fifty four patients with ST segment elevation acute coronary syndrome were treated with percutaneous coronary intervention (PCI) and with implantation of metal stent. TIMI and CADILLAC scores were used for evaluation of initial risk. Blood levels of cytokines and sP selectin were measured on day 1 before PCI and on day 10 of hospitalization., Results: We proved that CADILLAC score was applicable for evaluation of prognosis in patients with acute coronary syndrome and ST segment elevation treated with coronary stenting. High levels of tumor necrosis factor during first 24 hours of acute coronary syndrome and interleikin 8 on day 10 after PCI were found to be risk factors of major adverse coronary events during subsequent 12 months. High sP selectin level on day 10 predicted stent thrombosis during long term follow up.

Percutaneous stent implantation has revolutionized the clinical treatment of occluded arteries. Nevertheless, there is still a large unmet need to prevent re-occlusion after implantation. Consequently, a niche exists for a cost-effective pre-clinical method of evaluating novel interventional devices in human models. Therefore, the development of a coronary model artery offers tremendous potential for the treatment of endothelial cell dysfunction and restenosis. As a first step, we employ tissue-engineering principles to examine the effect of stent deployment upon endothelial cells in a tubular in vitro system capable of replicating the coronary artery biomechanical environment. In particular, the cellular and molecular changes pertaining to inflammation, proliferation, and death were assessed after stent deployment. Real-time quantitative PCR demonstrated increased expression of genes encoding for E-Selectin, ICAM-1, and VCAM-1; markers associated with an inflammatory response in vivo. Further, an increase in the pro-apoptotic protein Bax was paralleled with a decrease in the anti-apoptotic protein Bcl-2; however, apoptotic morphology was not observed. Interestingly, transcription of c-fos increased, whereas Ki67 levels fell over the same period. One hypothesis is that these results are in response to the altered local hemodynamic environment induced by stent deployment. Most significantly, this study highlights the potential of a biomimetic hemodynamic bioreactor combined with a gene expression analysis to evaluate, with greater specificity, the performance and interaction of stents with the endothelial layer in a controlled environment.

Purpose: Time changes in plasma concentrations of six different cytokines were investigated to evaluate the inflammatory response to renal artery stent placement., Materials and Methods: A total of 22 patients (17 men; mean age, 66 years +/- 13) with ostial renal artery stenosis and poorly controlled hypertension treated with stent placement were studied. Blood samples were collected at baseline and at 24 hours and 6 months after the intervention. Plasma concentrations of (i) tumor necrosis factor-alpha, (ii) interleukin-6 (IL-6), (iii) monocyte chemoattractant protein-1, (iv) intercellular adhesion molecule-1, (v) vascular cell adhesion molecule-1, and (vi) regulated upon activatin normal T-cell expressed presumed secreted were measured. Restenosis diagnosed with imaging follow-up at 6 months was recorded. Plasma concentrations of the aforementioned cytokines were compared between patients with and without restenosis., Results: IL-6 concentration increased significantly 24 hours after stent placement (8.3 pg/mL +/- 1.24 vs. 2.76 pg/mL +/- 1.27 at baseline) and returned to baseline levels (2.6 pg/mL +/- 1.77) at 6-month follow-up (P < .0001). No significant changes occurred in the concentrations of any other cytokines at the three time points. Baseline and 6-month concentrations of IL-6 were significantly higher in patients with restenosis than in those without restenosis (8.13 pg/mL +/- 4 vs 0.75 pg/mL +/- 0.47 [P < .005] and 9.55 pg/mL +/- 6.5 vs 0.42 pg/mL +/- 0.35 [P < .02], respectively)., Conclusions: Renal artery angioplasty with stent placement induces an inflammatory response, as evidenced by increased IL-6 production. Additionally, IL-6 seems to identify patients prone to develop restenosis; therefore, it might be used as an early predictor of restenosis after renal angioplasty with stent placement. However, larger studies are required to confirm IL-6 as a potential predictor of restenosis.

Present study was conducted to evaluate the association of IgG anticardiolipin antibodies with instent restenosis in patients having undergone percutaneous intervention with bare metal or drug eluting stents. Coronary artery disease patients with stent placement at least 6 months prior were screened for eligibility. 26 satisfied the inclusion/exclusion criteria. 10 patients with symptoms of restenosis, confirmed on check angiography served as cases and 16 without symptoms of restenosis served as control. Unpaired t- test was applied to ascertain the significance of any difference between control and study groups. Antibody levels were estimated on ELISA reader. The mean (+/- SD) anticardiolipin antibodies levels in cases and controls were 11.8 +/- 5.1 GPL/U/ml and 14.3 +/- 10.2 GPL/U/ml, respectively. The difference was not statistically significant (P > 0.05). In conclusion, we did not observe any significant correlation between the level of IgG aCL and instent restenosis.

Immunosuppressive treatments are available to suppress acute cardiac rejection; however, no viable treatment exists for long-term cardiac graft failure. Moreover, extended use of calcineurin inhibitor immunosuppressants, the mainstay of the current therapeutic for cardiac transplantation, leads to significant associated pathologies such as nephrotoxicity and increased risk of cardiac disease. For the last ten years alternatives to calcineurin inhibitors, or adjuvant therapies designed to complement their activities, have been explored. In tandem with this development, there has been considerable interest in Traditional Chinese Medicines (TCM) as sources for novel therapeutics. Our study examines the ability of the TCM Cordyceps sinensis to reduce acute and chronic rejection associated with cardiac transplantation. The objectives of this study were to first determine if oral delivery of the extract could reduce acute rejection in a rat heterotopic heart model of transplantation. The second objective was to determine, in vitro, if a sterile, aqueous extract of C. sinensis could decrease CD8+ T cell activity. The third objective was to determine if oral delivery of the extract could ablate allograft vasculopathy in a mouse abdominal aortic transplant model. We found that oral delivery of the extract demonstrated a reduction in acute rejection when used in conjunction with a sub-therapeutic dose of Cyclosporine. Further, we found, using a mixed lymphocyte reaction, that the extract was able to significantly reduce CD8+ T cell activity. Finally, we demonstrate that oral delivery of the extract, used with a therapeutic dose of Cyclosporine to suppress acute rejection, ablates allograft vasculopathy.

We tested if Quilty (endocardial infiltration of lymphocytes) in routinely processed endomyocardial biopsy is associated with poor outcome after heart transplantation (HTx). Biopsies (n=9829) harvested within the first post-transplant year from 938 patients (778 men, mean age 49 years) were evaluated for Quilty and acute cellular rejection (according to the International Society for Heart and Lung Transplantation, ISHLT, classification). Transplant vasculopathy was evaluated by coronary angiography, and severe stenosis was found in 19% of patients. Survival was tested by Kaplan-Meier and Cox regression analyses for all-cause mortality and major cardiac events (lethal acute cellular rejection, graft loss or myocardial infarction). We found 1840 (19%) Quilty-positive biopsies in 487 Quilty-positive patients (52%). Quilty was more prevalent in women (p=0.038) and younger men (p=0.001), and was correlated with ISHLT grade 1R (OR 1.45, 95% CI 1.36-1.55; p<0.001) and ISHLT grade 2R (OR 2.48, 95% CI 2.21-3.41; p<0.001). Quilty in any biopsy was associated with a higher all-cause mortality (log rank p=0.045) due to a higher risk for major cardiac event (p=0.0001). Multivariate regression analysis showed Quilty (RR 1.69, 95%CI 1.05-2.73) and transplant vasculopathy (RR 2.78, 95%CI 1.68-4.61) as risk factors for major cardiac events and treated hyperlipidemia as lowering the risk for major cardiac events (RR 0.47, 95%CI 0.28-0.77). Quilty is associated with graft loss and poor outcome post HTx. Index biopsy during the first post-transplant year is a useful tool to identify patients at risk and is recommended during routine post-transplant management.

Background: The aim was to assess the results of a decellularized bovine ureter graft (SynerGraft) for complex venous access., Methods: Bovine ureter conduits were implanted in patients with a failed fistula or access graft in whom native vessels were unsuitable as conduits. Graft histories were obtained from all patients who had undergone this procedure at one institution. Failed grafts were explanted and subjected to histological examination. A sample of fresh bovine ureter was immunostained for galactose (alpha1 --> 3) galactose (alpha-Gal)., Results: Nine patients with a median age of 46 (range 25-70) years underwent complex venous access surgery between August 2004 and November 2006 using a SynerGraft. Graft types included loop superficial femoral artery to stump of long saphenous vein (four patients), loop brachial artery to vein (two), brachial artery to axillary vein (two) and left axillary artery to innominate vein (one). Three grafts developed aneurysmal dilatation and two thrombosed. Histological assessment of the explanted bovine ureters revealed acute and chronic transmural inflammation. Immunostaining of fresh bovine ureter suggested residual cells and the xenoantigen alpha-Gal., Conclusion: Graft failure with aneurysmal dilatation and thrombosis in complex arteriovenous conduits using bovine ureter may be due to residual xenoantigens., (2008 British Journal of Surgery Society Ltd. Published by John Wiley & Sons, Ltd.)

The monoclonal antibody (mAb) to the platelet glycoprotein (GP) IIb/IIIa receptor has potent antiplatelet and antithrombotic characteristics shown to reduce thrombus-related major complications after coronary angioplasty. This mAb can be incorporated in drug-eluting stents capable of releasing single or multiple bioactive agents into the bloodstream and surrounding tissues. Stents eluting the monoclonal mouse anti-human platelet glycoprotein IIb/IIIa antibody SZ-262 were tested for their effectiveness in improving the blood compatibility and the antithrombotic characteristics by immunofluorescence and scanning electron microscopy (SEM). The SEM results convincingly demonstrated that the surface of the mAb eluting-stents was completely free of platelet uptake without any sign of cellular debris or proteinaceous deposits, compared with controls. The deformation index of platelets on the L-polylactic acid (L-PLA) coated stents were higher than bare Nitinol intravascular stents, as shown by SEM images. Monoclonal antibody to the platelet GP IIb/IIIa receptor, when eluting from L-PLA polymer-coated stents, effectively inhibits platelet aggregation in the stent microenvironment, thus demonstrating a potential capacity of reducing thrombosis, improving blood flow and arterial patency rates, and inhibiting cyclic blood flow variations. These results highlight the possibility of such monoclonal antibody-eluting stents to reduce or possibly eliminate thrombosis and in-stent restenosis., (Copyright 2007 Wiley Periodicals, Inc.)

The development of coronary stent has revolutionized the field of interventional cardiology by reducing the incidence of restenosis after balloon angioplasty. However, the stent has still associated with a serious complication, namely, in-stent restenosis. Although, restenosis following coronary stenting has long been attributed to neointimal proliferation, thrombosis, and negative remodeling, the inflammation may be a trigger for those vascular reactions following coronary stenting. Both experimental and clinical studies have demonstrated a marked activation of local and systemic inflammatory response following stent implantation, suggesting that inflammation may play an important role in determining in-stent restenosis via neointimal proliferation. The key role of inflammation in vascular healing and in-stent retsenosis has also been increasingly well understood. Recently, drug-eluting stents (DESs) have been shown to decrease in-stent restenosis in a large number of clinical studies. In addition to their anti-proliferative activity, DESs have been considered to possess an anti-inflammatory property, especially for sirolimus-eluting stent compared with bare metal stent. Moreover, the benefit of the anti-inflammatory therapy during the peri-procedural period and long-term follow-up by means of drug administration is also dependent on the inflammatory status during percutaneous coronary intervention. Measurement of cytokine and acute phase proteins, such as C-reactive protein, therefore, may be important to identify high-risk subjects and develop specific treatment tailored to the individual patients with stent restenosis. Thus, therapeutic approach should be further directed toward increasing local resistance to proliferative inflammatory stimuli by means of anti-proliferative, locally delivered drugs and reducing the magnitude and persistence of systemic inflammation.

Hepatitis C virus causes various extrahepatic immunologic abnormalities. Vascular access thrombosis (VAT) is a major cause of morbidity in chronic haemodialysis (HD) patients. Immunoglobulin-G anticardiolipin antibody (IgG-ACA) is strongly associated with venous and arterial thrombosis in patients with normal renal function. Previous investigations have reported the association of raised IgG-ACA titre recurrent with VAT in HD patient, and also few equivalent studies were reported the same in Taiwan. This study attempted to determine whether raised IgG-ACA titres are associated with increased risk of recurrent VAT in HD patients with chronic hepatitis C. This study enrolled 98 chronic hepatitis C patients undergoing HD. IgG-ACA titre and hepatitis C marker were measured for all subjects. Raised IgG-ACA titres were present in 29.6% (29/98) of patients. In both groups (raised and normal IgG-ACA), the type of shunt did not differ (p = 0.416). There was strong association between raised IgG-ACA titre and recurrent VAT (p = 0.0004). In predicting for more or one episodes of VAT using multiple logistic regression, synthetic graft (p < 0.0001), raised IgG-ACA titre (p = 0.039), presence of hepatitis B (p = 0.004) and haemodialysis duration (p = 0.039) were significant factors. The prevalence of raised IgG-ACA titres was 39.6% among chronic hepatitis C with HD patients. There was strong association between raised IgG-ACA titre and recurrent VAT, and this finding may be the consequence of pathogenetic role of raised IgG-ACA titres on the development of VAT status in HD patients with chronic hepatitis C.

Transplant coronary artery disease is the pre-eminent cause of late cardiac allograft failure. It is primarily characterized by a concentric intimal hyperplasia, which we designate transplant intimal hyperplasia (TIH). Although the pathogenesis of TIH is predominately immune driven, the specific role of alloantibodies in the disease process remains undefined. In this study we investigated the contribution of alloantibodies to the development of TIH in a murine model. Orthotopic, carotid artery transplantation was performed between B10A(2R) (H-2(h2)) donor mice and B-cell deficient muMT(-/-) knockout or wild-type C57BL/6 (H-2(b)) recipients in the absence of immunosuppression. Grafts were harvested at 35 days and subjected to planimetry and immunohistochemistry. Alloantibodies were detectable in wild-type recipients within 7 days of transplantation and recipients developed marked TIH at 35 days. Allografts harvested from B-cell deficient recipient mice also developed TIH, which was comparable in severity with wild-type recipients. However, whereas allografts from wild-type recipients showed marked intimal smooth muscle cell (SMC) proliferation, the neointima in B-cell deficient recipients lacked SMCs. Post-transplantation administration of anti-donor serum to muMT(-/-) recipients restored neointimal SMC population but did not influence the severity of TIH. Significant neointimal formation occurs in the absence of alloantibodies but lacks a SMC component. Therefore, SMC migration and proliferation is antibody dependent.

Background: Saphenous veins are often used for coronary artery bypass grafting (CABG), but loss of patency is a problem. The surgical procedure may contribute to graft injury. Our aim was to study the impact of surgical handling of saphenous veins on graft inflammation and vascular function., Methods: Biopsy samples of saphenous veins were taken from 9 patients undergoing elective CABG at the start of vein harvesting (open technique) and after the last proximal anastomosis was sutured. Messenger RNA was extracted and amplified with semiquantitative reverse transcription polymerase chain reaction. Gene expression of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta), leukocyte adhesion molecules (E-selectin, intercellular adhesion molecule-1), and vasoactive substances (endothelin-1, inducible and endothelial nitric oxide synthase) was investigated. Translocation of nuclear factor-kappaB (NFkappaB) was evaluated with electrophoretic mobility shift assay. Immunostaining for von Willebrand factor was performed to evaluate loss of endothelium, and in vitro vein reactivity to phenylephrine and endothelin-1 was studied., Results: Gene expression of cytokines and leukocyte adhesion molecules increased after graft harvesting and storage, whereas vasoactive substances did not change. Nuclear translocation of NFkappaB occurred after surgical handling, concurrent with partial loss of endothelium and impaired contractile function., Conclusions: Standard surgical handling of vein grafts induces NFkappaB-driven inflammation in the vessel wall and impairs vascular function. This may potentially contribute to both early and late graft occlusion.

Objective: Chronic vascular rejection, the main cause of allograft failure, is characterized by the destruction of smooth muscle cells (SMCs) in the media concomitantly with the proliferation of SMCs in the adjacent neointima. We hypothesized that alloantibodies might be responsible for these 2 opposite but coordinated events., Methods and Results: We used the rat aortic interposition model of chronic vascular rejection. During the rejection process, a neointima composed of proliferating SMCs from the recipient developed, whereas the SMCs in the media, all of donor origin, underwent apoptosis. Alloantibody deposition was detected only in the media. Using in vitro cultures experiments, we observed that alloantibody binding to donor SMCs exerts (1) a rapid upregulation of the transcription of growth factors genes, followed by (2) the induction of apoptosis after 24 hours. The transient production of growth factors by donor SMCs in response to the binding of alloantibodies induced the proliferation of recipient SMCs in culture supernatant transfer experiments. Additional data suggest that among the repertoire of alloantibodies, those directed against major histocompatibility complex I might carry the remodeling effect., Conclusions: Our data suggest that during chronic vascular rejection, alloantibody binding to donor medial SMCs is a crucial event that links neointimal and medial remodeling.

In order to prove the feasibility of preparation of the drug-incorporated stent by immersing stent wires in the monoclonal antibody (mAb) solution, fluorescence stain and image analysis were used to evaluate the L-PLA-coated stent. Absorption was measured using a radioisotope technique after preparing the mAb-incorporated stent, and the absorption curve was determined from the absorption data. In an in vitro perfusion circuit, the antibody was eluted from the stent matrices, and the related influence factors were evaluated based on the release data.

Coronary artery disease, leading to myocardial infarction and ischaemia, affects millions of persons and is one of the leading causes of morbidity and mortality worldwide. Invasive techniques such as coronary artery bypass grafting are used to alleviate the sequelae of arterial occlusion. Unfortunately, restenosis or occlusion of the grafted conduit occurs over a time frame of months to years with a gradual reduction in patency, especially in vein grafts. The events leading to intimal hyperplasia (IH) formation involve numerous cellular and molecular components. Various cellular elements of the vessel wall are involved as are leucocyte-endothelial interactions that trigger the coagulation cascade leading to localized thrombus formation. Subsequent phenotypic modification of the medial smooth muscle cells and their intimal migration is the basis of the lesion formation that is thought to be propagated by an immune-mediated reaction. Despite intense scrutiny, the pathophysiology of IH remains an enigma. Although several growth factors, cytokines and numerous other biomolecules have been implicated and their relationship to prohyperplasia pathways such as the phosphatidyl-inositol 3-kinase (PI3K)-Akt pathway has been established, many pieces of the puzzle are still missing. An in-depth understanding of early vein graft adaptation and progression is necessary to improve the long-term prognosis and develop more effective therapeutic measures. In this review, we have critically evaluated and summarized the literature to elucidate and interlink the numerous established and emerging factors that play a key role in the development of IH leading to vein graft restenosis.

Most patients subjected to coronary artery bypass grafting develop graft occlusions in some time after successful coronary surgery. Causes of graft occlusions comprise hyperplasia of intima occurring as a response to release of cytokines and angiotensin II. Angiotensin II is formed in human body with participation of angiotensin converting enzyme (ACE) or chymase. We found high concentrations of ACE in some smooth muscle cells in hyperplasia zone. Proliferation of smooth muscle cells, their hypertrophy and increased synthesis of intercellular space proteins facilitate graft occlusion. Moreover ACE was found in cytoplasm of macrophages infiltrating graft's intima. Thus ACE can be an important contributor to the process of coronary artery bypass graft obstruction.

Graft coronary arteriosclerosis (GCA) is the leading cause of long-term mortality after heart transplantation (HTx). The goal of this study was to demonstrate that inhibition of immunemediated injury by cyclosporine (CsA) protects the allograft from GCA. ACI-to-Lewis rat allografts were disparate in major and nonmajor histocompatibility loci. Isografts (Lewis-Lewis) were controls. Treatment groups received either olive oil or CsA at 2.5, 5, 10, or 20 mg/kg/day for 3 months. Histology (elastin) and immunohistochemistry using monoclonal antibodies to CD4, CD8, CD45R, RT1B, CD11b/c, CD25, and alpha-actin was performed to examine the epicardial and intramyocardial coronary arteries. Computerized image morphometry was utilized to measure intimal and medial thickness and area. Rats receiving olive oil or CsA at 2.5 mg/kg/day had severe rejection and no graft survival. CsA at 5 mg/kg/day resulted in less severe rejection with significant intimal and medial proliferation (P < 0.001). CsA at 10-20 mg/kg/day paralleled Lewis-Lewis isograft outcomes and inhibited arteriosclerotic vascular changes in the allograft (P < 0.001). Perivascular T-helper cells and macrophages were a characteristic finding with low-dose CsA but rare with higher CsA doses. In this new model of accelerated GCA in rats, immune-mediated antigen-dependent vasculopathy as a result of inadequate immunosuppresion is fundamental in the development of GCA, which appeared equally in epicardial arteries and intramyocardial arterioles. CsA prevents GCA in a dose-dependent fashion in the rat allograft.

Objective: Interferon-gamma (IFN-gamma), a cytokine produced primarily by T cells and by activated macrophages, plays a central role in the pathogenesis of graft arterial disease (GAD). This study investigated whether T cells can induce GAD in the absence of humoral alloresponses and whether activated macrophages or other host cell types can substitute as sources of IFN-gamma in GAD., Methods: Wild-type (WT), IFN-gamma-/-, or recombination-activating-gene-1-/- (RAG-1-/-; lacking mature T and B cells) mice received MHC II-disparate hearts. The grafts were harvested 8 weeks post-transplant and histological and immunohistochemical analyses, RNase protection assay (RPA), and flow cytometry were used to evaluate GAD lesions, infiltrating cell populations, and IFN-gamma expression by infiltrating cells., Results: Moderate-to-severe GAD developed in WT recipient allografts, associated with abundant IFN-gamma expression by both infiltrating T cells and macrophages. No GAD developed in IFN-gamma-/- or in RAG-1-/- hosts, nor was any IFN-gamma expression evident. RAG-1-/- hosts receiving naïve WT or IFN-gamma-/- T cells (10(7)) after heart transplantation demonstrated no mature B cells but showed persistence of transferred T cells up to 8 weeks post-transplant. In the complete absence of B cells and alloantibody, transfer of WT T cells into RAG-1-/- recipients yielded GAD, with associated IFN-gamma expression by the transferred T cells and the host macrophages. Transfer of IFN-gamma-/- T cells induced neither GAD nor host macrophage IFN-gamma expression., Conclusions: T cells, even in the absence of B cells, suffice to induce GAD, and T cell-derived IFN-gamma plays a critical role in GAD pathogenesis., (Copyright 2004 European Society of Cardiology)

Mononuclear leukocytes infiltrate the artery wall via integrin-mediated mechanisms and play an integral role in intimal formation after stenting. We sought to determine if acute antagonism of the alpha4 subunit of very late antigen-4 is sufficient for the late attenuation of stent intimal area (IA). Twenty-four hypercholesterolemic rabbits underwent iliac artery balloon injury, followed 2 weeks later by stent implantation, and the animals were randomized to receive an anti-alpha4 antibody (HP1/2) or a nonspecific isotypic control immunoglobulin (1E6) intravenously 1 h before stenting. Compared with controls, HP1/2-treated rabbits showed 50%, 51%, and 44% reductions in the percentage on intimal cells that were macrophages on days 3, 7, and 28 after stenting and a 59% reduction in intimal proliferation on day 3. Although stent IA was reduced by 63% and 48% in the antibody-treated group compared with the control group on days 3 and 7, this difference was not present on day 28. These data highlight the need for sustained, anti-inflammatory therapies for the prevention of stent intimal formation.

Objective: C-type natriuretic peptide (CNP), which is produced by vascular endothelial cells, exhibits anti-proliferative and anti-inflammatory effects. Cytotoxic T-lymphocytes may be involved in vein graft disease. Attenuation of vein graft disease necessitates a remodelling of the arterialized vein towards a more contractile phenotype which is characterized, among other factors, by the calponin amount. We investigated the effects of perivascularly applied CNP in a mouse model of vein graft disease., Methods: C57BL6J mice underwent interposition of the inferior vena cava from isogenic donor mice into the common carotid artery using a previously described cuff technique. In the treatment group, 10(-6)mol/l of CNP were applied locally in pluronic gel. The control group did not receive local treatment. Grafts were harvested at 1, 2, 4, and 8 weeks and underwent morphometric analysis as well as immunohistochemical analysis., Results: In grafted veins without treatment (controls) median intimal thickness was 10 (6-29), 12 (8-40)microm, was 47 (12-58), and 79 (62-146)microm after 1, 2, 4 and 8 weeks, respectively. In the treatment groups, which received 10(-6)mol/l of CNP, the intimal thickness was 5 (3-6), 6 (4-15), 32 (5-54), and 43 (39-70)microm after 1, 2, 4 and 8 weeks, respectively. This reduction of intimal thickness was significant at 1, 2 and 8 weeks. Immunohistochemically, the reduction of intimal thickness was associated with a decreased infiltration of CD-8 positive cells and an increased amount of calponin in the CNP-treated grafts., Conclusion: We conclude that perivascular application of CNP inhibits neointimal hyperplasia of vein grafts in a mouse model. These results suggest that CNP may have a therapeutic potential for the prevention of vein graft disease.

Cardiac allograft vasculopathy (CAV) continues to be a major cause of late graft failure after cardiac transplantation. We have demonstrated that Rho-kinase, an effector of the small GTPase Rho, plays an important role in the pathogenesis of arteriosclerosis. In this study, we examined whether the Rho-kinase-mediated pathway is also involved in the pathogenesis of CAV using a specific Rho-kinase inhibitor and a dominant-negative Rho-kinase. Hearts from AKR mice were heterotopically transplanted to C3H/He (allograft) or AKR mice (isograft), and the effects of long-term oral treatment with fasudil, which is metabolized to a specific Rho-kinase inhibitor hydroxyfasudil, on CAV were examined at 2 and 4 weeks after the transplantation. Coronary remodeling in the allografts characterized by intimal thickening and perivascular fibrosis was dose-dependently suppressed in the fasudil group compared with the control group (P<0.01, n=9 to 10). The inhibitory effects of hydroxyfasudil were mimicked by in vivo gene transfer of dominant-negative Rho-kinase (P<0.05, n=4). Among the proinflammatory cytokines examined, those of macrophage migration inhibitory factor, interferon-gamma, and transforming growth factor-beta1 were upregulated in the control group and were dose-dependently inhibited in the fasudil group (P<0.01, n=5). Vascular inflammation in the allografts, as evidenced by accumulation of inflammatory cells (macrophages and T cells), was also significantly inhibited in the fasudil group (P<0.05, n=5 to 10). These results indicate that long-term treatment with fasudil suppresses CAV in mice, suggesting that Rho-kinase is an important therapeutic target for the prevention of CAV.

Background: Innate immunity is of major importance in vascular repair. The present study evaluated whether systemic and transient depletion of monocytes and macrophages with liposome-encapsulated bisphosphonates inhibits experimental in-stent neointimal formation., Methods and Results: Rabbits fed on a hypercholesterolemic diet underwent bilateral iliac artery balloon denudation and stent deployment. Liposomal alendronate (3 or 6 mg/kg) was given concurrently with stenting. Monocyte counts were reduced by >90% 24 to 48 hours after a single injection of liposomal alendronate, returning to basal levels at 6 days. This treatment significantly reduced intimal area at 28 days, from 3.88+/-0.93 to 2.08+/-0.58 and 2.16+/-0.62 mm2. Lumen area was increased from 2.87+/-0.44 to 3.57+/-0.65 and 3.45+/-0.58 mm2, and arterial stenosis was reduced from 58+/-11% to 37+/-8% and 38+/-7% in controls, rabbits treated with 3 mg/kg, and rabbits treated with 6 mg/kg, respectively (mean+/-SD, n=8 rabbits/group, P<0.01 for all 3 parameters). No drug-related adverse effects were observed. Reduction in neointimal formation was associated with reduced arterial macrophage infiltration and proliferation at 6 days and with an equal reduction in intimal macrophage and smooth muscle cell content at 28 days after injury. Conversely, drug regimens ineffective in reducing monocyte levels did not inhibit neointimal formation., Conclusions: Systemic transient depletion of monocytes and macrophages, by a single liposomal bisphosphonates injection concurrent with injury, reduces in-stent neointimal formation and arterial stenosis in hypercholesterolemic rabbits.

Despite advances in immunosuppressive drugs, coronary artery transplant vasculopathy (CATV) is the major cause of graft failure that limits long-term survival of cardiac transplantation. The pathogenesis of CATV involves a chronic immune response of the recipient to the donor vasculature in which activated recipient immune cells damage the endothelium and produce cytokines, resulting in vascular smooth muscle cell (VSMC) activation. Activated VSMC migrate from the media into the lumen, proliferate, and elaborate cytokines and matrix proteins, resulting in loss of lumen diameter and vascular contractility. Because of its extensive nature, interventions which are successful in patients with conventional coronary artery disease are often not applicable to the majority of patients with CATV. Although intended for immune suppression, many immunosuppressive agents owe at least part of their efficacy to their anti proliferative effects on VSMC, including rapamycin, mycophenolic acid, cyclosporin, calcium channel blockers, and HMG CoA reductase inhibitors. Because activation of VSMC is responsible for most of the obliterative arterial intimal thickening present in solid organ allografts, the induction of expression of a selected set of genes may reflect the status of acceptance of the vasculature by the recipient, and the activation, migration, and proliferation of VSMC represent potential points for therapeutic intervention. The risk of infection and malignancy associated with immunosuppressive therapy further promote the need to identify a molecular target which directly modulates the VSMC response to injury. This review will summarize the anti proliferative effects that immunosuppressive drugs have on VSMC proliferation. We will also describe efforts to define the genes which regulate the vascular response to allograft injury, and describe how some of these proteins may represent targets to reduce VSMC proliferation and attenuate CATV.

Background: Monocyte chemoattractant protein-1 (MCP-1) is associated with the development of graft coronary artery disease (GCAD) following cardiac transplantation. This study assessed whether technetium 99m ((99m)Tc)-labeled MCP-1 binds its receptors in chronic cardiac transplants and thereby provides a potential modality to assess GCAD., Methods: Allogeneic (PVG-->ACI, n = 9) and syngeneic (ACI-->ACI, n = 9) rat heterotopic heart transplants were performed. Allograft recipients were treated with 7.5 mg/kg per day of Cyclosporin A for 10 days until tolerance was achieved. After 90 days, animals were injected intravenously with (99m)Tc-MCP-1 and killed after 1 hour. Radioactivity of heart tissues was measured and standardized to uptake in the overall blood pool. Two-dimensional (99m)Tc-MCP-1 uptake (autoradiographs) was imaged by exposing 50-microm sections on a phosphoimager overnight. ED-1 staining of monocyte/macrophages was performed on serial sections. Additional sections were stained with elastin von Gieson and hematoxylin. Hearts were scored for luminal narrowing and intima/media ratio (I/M) with computerized image analysis., Results: Allografts exhibited significantly more luminal narrowing (22.5 +/- 10.7% vs 2.6 +/- 4.6, p = 0.0005) and higher I/M (0.173 +/- 0.151 vs 0.015 +/- 0.029, p = 0.0088) than isografts. The ratio of (99m)Tc-MCP-1 uptake in allografts (1.04 +/- 0.4) was greater than that of isograft controls (0.72 +/- 0.11, p = 0.03). Pixel counts of autoradiographs and ED-1-stained sections demonstrated a modest correlation between the two (R(2) = 0.50). No significant differences were seen in acute rejection scores., Conclusion: (99m)Tc-MCP-1 uptake was higher in allografts vs isografts and was consistent with a greater degree of GCAD. These data demonstrating increased radiopharmaceutical uptake in hearts with GCAD provide a foundation for the development of a potentially non-invasive imaging assay of this disease process in heart transplantation.

Background: Emerging data indicate that the inflammatory response after mechanical arterial injury correlates with the severity of neointimal hyperplasia in animal models and postangioplasty restenosis in humans. The present study was designed to examine whether a nonspecific stimulation of the innate immune system, induced in close temporal proximity to the vascular injury, would modulate the results of the procedure. Methods and Results- Rabbits subjected to iliac artery balloon injury (balloon denudation with or without stent deployment) were injected twice with a bacterial lipopolysaccharide (LPS) (500 ng/rabbit) before and after surgery. The dose was chosen to be sufficient to induce systemic inflammation but not septic shock. A systemic marker of inflammation (serum interleukin-1beta levels measured by ELISA) and monocytic stimulation (CD14 levels on monocytes measured by flow cytometry) were increased after LPS administration. Arterial macrophage infiltration at 7 days after injury was 1.7+/-1.2% of total cells in controls and 4.2+/-1.8% in LPS-treated rabbits (n=4, P<0.05). Morphometric analysis of the injured arteries 4 weeks after injury revealed significantly increased luminal stenosis (38+/-4.2% versus 23+/-2.6, mean+/- SEM; n=8, P<0.05) and neointima-to-media ratio (1.26+/-0.21 versus 0.66+/- 0.09, P<0.05) in LPS-treated animals compared with controls. This effect was abolished by anti-CD14 Ab administration. Serum interleukin-1beta levels and monocyte CD14 expression were significantly increased in correlation with the severity of intimal hyperplasia. LPS treatment increased neointimal area after stenting from 0.57+/-0.07 to 0.77+/- 0.1 mm(2) and stenosis from 9+/-1% to 13+/-1.7% (n=5, P< 0.05)., Conclusions: Nonspecific systemic stimulation of the innate immune system concurrently with arterial vascular injury facilitates neointimal formation, and conditions associated with increased inflammation may increase restenosis.

In this study, we investigated the role of T and B lymphocytes in neointimal hyperplasia after endothelial denudation. Catheter-induced endothelial denudation of wild-type mice resulted in rapid infiltration of lymphocytes to the site of injury. Mice defective in recombination-activating gene 2 (RAG2(-/-)) showed increased neointimal formation 14 days after vascular injury in comparison to their wild-type immune-competent littermates. Immunohistochemical studies revealed the preponderance of smooth muscle cells and a significantly higher number of proliferating cells in the neointima of the RAG2(-/-) mice. The neointima size and the number of proliferating smooth muscle cells in the injured vessel of RAG2(-/-) mice were similar to those observed in the injured arteries of apolipoprotein E (apoE)-deficient (apoE(-/-)) mice. Interestingly, mice with double apoE and RAG2 gene mutations (apoE(-/-) RAG2(-/-)) displayed similar neointimal characteristics as mice with a single gene defect, suggesting a similar mechanism for apoE and lymphocyte protection against injury-induced neointimal formation. The protective role of lymphocytes against neointimal formation after vascular injury directly contrasts to their reported role in the promotion of atherosclerosis, which was observed in both apoE(+/+) and apoE(-/-) mice. Thus, these results support the hypothesis of different etiology between hyperlipidemia-induced atherosclerosis and injury-induced vascular occlusion.

Objectives: We sought to investigate whether statin therapy affects the association between preprocedural C-reactive protein (CRP) levels and the risk for recurrent coronary events in patients undergoing coronary stent implantation., Background: Low-grade inflammation as detected by elevated CRP levels predicts the risk of recurrent coronary events. The effect of inflammation on coronary risk may be attenuated by statin therapy., Methods: We investigated a potential interrelation among statin therapy, serum evidence of inflammation, and the risk for recurrent coronary events in 388 consecutive patients undergoing coronary stent implantation. Patients were grouped according to the median CRP level (0.6 mg/dl) and to the presence of statin therapy., Results: A primary combined end point event occurred significantly more frequently in patients with elevated CRP levels without statin therapy (RR [relative risk] 2.37, 95% CI [confidence interval] [1.3 to 4.2]). Importantly, in the presence of statin therapy, the RR for recurrent events was significantly reduced in the patients with elevated CRP levels (RR 1.27 [0.7 to 2.1]) to about the same degree as in patients with CRP levels below 0.6 mg/dl and who did not receive statin therapy (RR 1.1 [0.8 to 1.3])., Conclusions: Statin therapy significantly attenuates the increased risk for major adverse cardiac events in patients with elevated CRP levels undergoing coronary stent implantation, suggesting that statin therapy interferes with the detrimental effects of inflammation on accelerated atherosclerotic disease progression following coronary stenting.