Abbey Schepers

Anti–MCP-1 Gene Therapy Inhibits Vascular Smooth Muscle Cells Proliferation and Attenuates Vein Graft Thickening Both In Vitro and In Vivo

Objective—Because late vein graft failure is caused by intimal hyperplasia (IH) and accelerated atherosclerosis, and these processes are thought to be inflammation driven, influx of monocytes is one of the first phenomena seen in IH, we would like to provide direct evidence for a role of the MCP-1 pathway in the development of vein graft disease. Methods and Results—MCP-1 expression is demonstrated in various stages of vein graft disease in a murine model in which venous interpositions are placed in the carotid arteries of hypercholesterolemic ApoE3Leiden mice and in cultured human saphenous vein (HSV) segments in which IH occurs. The functional involvement of MCP-1 in vein graft remodeling is demonstrated by blocking the MCP-1 receptor CCR-2 using 7ND-MCP-1. 7ND-MCP1 gene transfer resulted in 51% reduction in IH in the mouse model, when compared with controls. In HSV cultures neointima formation was inhibited by 53%. In addition, we demonstrate a direct inhibitory effect of 7ND-MCP-1 on the proliferation of smooth muscle cell (SMC) in HSV cultures and in SMC cell cultures. Conclusion—These data, for the first time, prove that MCP-1 has a pivotal role in vein graft thickening due to intimal hyperplasia and accelerated atherosclerosis.

Natural Killer Cells and CD4 + T-Cells Modulate Collateral Artery Development

Objective—The immune system is thought to play a crucial role in regulating collateral circulation (arteriogenesis), a vital compensatory mechanism in patients with arterial obstructive disease. Here, we studied the role of lymphocytes in a murine model of hindlimb ischemia. Methods and Results—Lymphocytes, detected with markers for NK1.1, CD3, and CD4, invaded the collateral vessel wall. Arteriogenesis was impaired in C57BL/6 mice depleted for Natural Killer (NK)-cells by anti-NK1.1 antibodies and in NK-cell–deficient transgenic mice. Arteriogenesis was, however, unaffected in J&agr;281-knockout mice that lack NK1.1+ Natural Killer T (NKT)-cells, indicating that NK-cells, rather than NKT-cells, are involved in arteriogenesis. Furthermore, arteriogenesis was impaired in C57BL/6 mice depleted for CD4+ T-lymphocytes by anti-CD4 antibodies, and in major histocompatibility complex (MHC)-class-II–deficient mice that more selectively lack mature peripheral CD4+ T-lymphocytes. This impairment was even more profound in anti-NK1.1-treated MHC-class-II–deficient mice that lack both NK- and CD4+ T-lymphocytes. Finally, collateral growth was severely reduced in BALB/c as compared with C57BL/6 mice, 2 strains with different bias in immune responsiveness. Conclusions—These data show that both NK-cells and CD4+ T-cells modulate arteriogenesis. Promoting lymphocyte activation may represent a promising method to treat ischemic disease.

Tumor necrosis factor-α plays an important role in restenosis development

Genetic factors appear to be important in the restenotic process after percutaneous coronary intervention (PCI), as well as in inflammation, a pivotal factor in restenosis. TNFα, a key regulator of inflammatory responses, may exert critical influence on the development of restenosis after PCI. The GENetic DEterminants of Restenosis (GENDER) project included 3104 patients who underwent a successful PCI. Systematic genotyping for six polymorphisms in the TNFα gene was performed. The role of TNFα in restenosis was also assessed in ApoE*3‐Leiden mice, TNFα knockout mice, and by local delivery of a TNFα biosynthesis inhibitor, thalidomide. The −238G‐1031T haplotype of the TNFα gene increased clinical and angiographic risk of restenosis (P=0.02 and P=0.002, respectively). In a mouse model of reactive stenosis, arterial TNFα mRNA was significantly time‐dependently up‐regulated. Mice lacking TNFα or treated locally with thalidomide showed a reduction in reactive stenosis (P=0.01 and P=0.005, respectively). Clinical and preclinical data indicate that TNFα plays an important role in restenosis. Therefore, TNFα genotype may be used as a risk marker for restenosis and may contribute to individual patient screening prior to PCI in clinical practice. Inhibition of TNFα may be an anti‐restenotic target strategy.—Monraats, P. S., Pires, N. M. M., Schepers, A., Agema, W. R. P., Boesten, L. S. M., deVries, M. R., Zwinderman, A. H., deMaat, M. P. M., Doevendans, P. A. F. M., deWinter, R. J., Tio, R. A., Waltenberger, J., tHart, L. M., Frants, R. R., Quax, P. H. A., van, Vlijmen, B. J. M., Havekes, L. M., van derLaarse, A., van derWall, E. E., Jukema, J. W. Tumor necrosis factor‐α plays an important role in restenosis development. FASEB J. 19, 1998–2004 (2005)

Genetic inflammatory factors predict restenosis after percutaneous coronary interventions

Background— Restenosis is a negative effect of percutaneous coronary intervention (PCI). No clinical factors are available that allow good risk stratification. However, evidence exists that genetic factors are important in the restenotic process as well as in the process of inflammation, a pivotal factor in restenosis. Association studies have identified genes that may predispose to restenosis, but confirmation by large prospective studies is lacking. Our aim was to identify polymorphisms and haplotypes in genes involved in inflammatory pathways that predispose to restenosis. Methods and Results— The GENetic DEterminants of Restenosis (GENDER) project is a multicenter prospective study, including 3104 consecutive patients after successful PCI. Forty-eight polymorphisms in 34 genes in pathways possibly involved in the inflammatory process were analyzed. The 16Gly variant of the β2-adrenergic receptor gave an increased risk of target vessel revascularization (TVR). The rare alleles of the CD14 gene (−260T/T), colony-stimulating factor 2 gene (117Thr/Thr), and eotaxin gene (−1328A/A) were associated with decreased risk of TVR. However, through the use of multiple testing corrections with permutation analysis, the probability of finding 4 significant markers by chance was 12%. Conclusions— Polymorphisms in 4 genes considered involved in the inflammatory reaction showed an association with TVR after PCI. Our results may contribute to the unraveling of the restenotic process. Given the explorative nature of this analysis, our results need to be replicated in other studies.

Inhibition of Complement Component C3 Reduces Vein Graft Atherosclerosis in Apolipoprotein E3–Leiden Transgenic Mice

Background— Venous bypass grafts may fail because of development of intimal hyperplasia and accelerated atherosclerosis. Inflammation plays a major role in these processes. Complement is an important part of the immune system and participates in the regulation of inflammation. The exact role of complement in the process of accelerated atherosclerosis of vein grafts has not yet been explored, however. Methods and Results— To assess the role of complement in the development of vein graft atherosclerosis, a mouse model, in which a venous interposition was placed in the common carotid artery, was used. In this model, vein graft thickening appeared within 4 weeks. The expression of complement components was studied with the use of immunohistochemistry on sections of the thickened vein graft. C1q, C3, C9, and the regulatory proteins CD59 and complement receptor-related gene y could be detected in the lesions 4 weeks after surgery. Quantitative mRNA analysis for C1q, C3, CD59, and complement receptor-related gene y revealed expression of these molecules in the thickened vein graft, whereas C9 did not show local mRNA expression. Furthermore, interference with C3 activation with complement receptor-related gene y–Ig was associated with reduced vein graft thickening, reduced C3 and C9 deposition, and reduced inflammation as assessed by analysis of influx of inflammatory cells, such as leukocytes, T cells, and monocytes. In addition, changes in apoptosis and proliferation were observed. When C3 was inhibited by cobra venom factor, a similar reduction in vein graft thickening was observed. Conclusions— The complement cascade is involved in vein graft thickening and may be a target for therapy in vein graft failure disease.

Complement factor C5a as mast cell activator mediates vascular remodelling in vein graft disease

AIMSnFailure of vein graft conduits due to vein graft thickening, accelerated atherosclerosis, and subsequent plaque rupture is applicable to 50% of all vein grafts within 10 years. New potential therapeutic targets to treat vein graft disease may be found in components of the innate immune system, such as mast cells and complement factors, which are known to be involved in atherosclerosis and plaque destabilization. Interestingly, mast cells can be activated by complement factor C5a and, therefore, a direct role for C5a-mediated mast cell activation in vein graft disease is anticipated. We hypothesize that C5a-mediated mast cell activation is involved in the development and destabilization of vein graft lesions.nnnMETHODS AND RESULTSnMast cells accumulated in time in murine vein graft lesions, and C5a and C5a-receptor (CD88) expression was up-regulated during vein graft disease in apolipoprotein E-deficient mice. Mast cell activation with dinitrophenyl resulted in a profound increase in vein graft thickening and in the number of plaque disruptions. C5a application enhanced vein graft lesion formation, while treatment with a C5a-receptor antagonist resulted in decreased vein graft disease. C5a most likely exerts its function via mast cell activation since the mast cell inhibitor cromolyn totally blocked C5a-enhanced vein graft disease.nnnCONCLUSIONnThese data provide evidence that complement factor C5a-induced mast cell activation is highly involved in vein graft disease, which identifies new targets to prevent vein graft disease.

Intraoperative guidance in parathyroid surgery using near-infrared fluorescence imaging and low-dose Methylene Blue

BACKGROUNDnIdentification of diseased and normal parathyroid glands during parathyroid surgery can be challenging. The aim of this study was to assess whether near-infrared (NIR) fluorescence imaging using administration of a low-dose Methylene Blue (MB) at the start of the operation could provide optical guidance during parathyroid surgery and assist in the detection of parathyroid adenomas.nnnMETHODSnPatients diagnosed with primary hyperparathyroidism planned for parathyroidectomy were included. Patients received 0.5 mg/kg MB intravenously directly after start of anesthesia. During the operation, NIR fluorescence imaging was performed to identify parathyroid adenomas. Imaging results were compared with a previous published feasibility study in which 12 patients received MB after intraoperative identification of the adenoma.nnnRESULTSnA total of 13 patients were included in the current study. In 10 of 12 patients with a histologically proven adenoma, the adenoma was fluorescent. Mean signal to background ratio was 3.1 ± 2.8. Mean diameter of the resected lesions was 17 ± 9 mm (range 5-28 mm). Adenomas could be identified up to 145 minutes after administration, which was the longest timespan until resection. Interestingly, in 3 patients, a total of 6 normal parathyroid glands (median diameter 2.5 mm) with a signal to background ratio of 1.8 ± 0.4 were identified using NIR fluorescence imaging.nnnCONCLUSIONnEarly administration of low-dose MB provided guidance during parathyroidectomy by identifying both parathyroid adenomas and normal parathyroid glands. In patients in whom difficult identification of the parathyroid adenoma is expected or when normal glands have to be identified, the administration of MB may improve surgical outcome.

C1-esterase inhibitor protects against early vein graft remodeling under arterial blood pressure.

OBJECTIVESnArterial pressure induced vein graft injury can result in endothelial loss, accelerated atherosclerosis and vein graft failure. Inflammation, including complement activation, is assumed to play a pivotal role herein. Here, we analyzed the effects of C1-esterase inhibitor (C1inh) on early vein graft remodeling.nnnMETHODSnHuman saphenous vein graft segments (n=8) were perfused in vitro with autologous blood either supplemented or not with purified human C1inh at arterial pressure for 6h. The vein segments and perfusion blood were analyzed for cell damage and complement activation. In addition, the effect of purified C1inh on vein graft remodeling was analyzed in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery.nnnRESULTSnApplication of C1inh in the in vitro perfusion model resulted in significantly higher blood levels and significantly more depositions of C1inh in the vein wall. This coincided with a significant reduction in endothelial loss and deposition of C3d and C4d in the vein wall, especially in the circular layer, compared to vein segments perfused without supplemented C1inh. Administration of purified C1inh significantly inhibited vein graft intimal thickening in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery.nnnCONCLUSIONnC1inh significantly protects against early vein graft remodeling, including loss of endothelium and intimal thickening. These data suggest that it may be worth considering its use in patients undergoing coronary artery bypass grafting.

Impact of Complications After Surgery for Colorectal Liver Metastasis on Patient Survival

BACKGROUNDnIn some patients with colorectal liver metastases it is not clear whether liver resection or isolated liver perfusion is the best treatment option. For instance, the risk of complications after surgery may be so substantial and affect subsequent survival. Aim of the present study is to compare complication occurrence and its effect on survival after liver resection and perfusion.nnnMETHODSnPatient records of all 225 patients with colorectal liver metastases treated with liver resection (n = 121) or liver perfusion (n = 104) in the period 1997-2006 were reviewed for complications during the initial hospitalisation until 30 d after discharge, and to assess patient survival until the last hospital visit. Median duration of follow-up was 38 mo for overall survival and 22 mo for survival after surgery.nnnRESULTSnComplications occurred less often in patients undergoing resection than perfusion (29.8% versus 49.0%, X(2) = 8.77, P < 0.01). Postoperative mortality rates were similar in both groups (4.1% and 4.8%, respectively). As expected, long term survival after liver surgery was better in the resection group: at 3 y, 60% of patients survived in the resection group, compared with 22% after liver perfusion (log rank X(2) = 35.29 P < 0.001). However, liver resection patients with postoperative complications, had similar survival as perfusion patients without complications (log rank X(2) = 2.45, p = 0.12). This remained after adjustment for differences between the patient groups at time of surgery.nnnCONCLUSIONnLiver resection has superior long-term survival, but survival is significantly reduced by the occurrence of post-surgical complications. When complications occur after liver resection, survival is comparable to patients who underwent uncomplicated liver perfusion.

Characteristics of contralateral carcinomas in patients with differentiated thyroid cancer larger than 1 cm

PurposeTraditionally, total thyroidectomy has been advocated for patients with tumors larger than 1xa0cm. However, according to the ATA and NCCN guidelines (2015, USA), patients with tumors up to 4xa0cm are now eligible for lobectomy. A rationale for adhering to total thyroidectomy might be the presence of contralateral carcinomas. The purpose of this study was to describe the characteristics of contralateral carcinomas in patients with differentiated thyroid cancer (DTC) larger than 1xa0cm.MethodsA retrospective study was performed including patients from 17 centers in 5 countries. Adults diagnosed with DTC stage T1b-T3 N0-1a M0 who all underwent a total thyroidectomy were included. The primary endpoint was the presence of a contralateral carcinoma.ResultsA total of 1313 patients were included, of whom 426 (32xa0%) had a contralateral carcinoma. The contralateral carcinomas consisted of 288 (67xa0%) papillary thyroid carcinomas (PTC), 124 (30xa0%) follicular variant of a papillary thyroid carcinoma (FvPTC), 5 (1xa0%) follicular thyroid carcinomas (FTC), and 3 (1xa0%) Hürthle cell carcinomas (HTC). Ipsilateral multifocality was strongly associated with the presence of contralateral carcinomas (OR 2.62). Of all contralateral carcinomas, 82xa0% were ≤10xa0mm and of those 99xa0% were PTC or FvPTC. Even if the primary tumor was a FTC or HTC, the contralateral carcinoma was (Fv)PTC in 92xa0% of cases.ConclusionsThis international multicenter study performed on patients with DTC larger than 1xa0cm shows that contralateral carcinomas occur in one third of patients and, independently of primary tumor subtype, predominantly consist of microPTC.