Linkage analysis by genotyping of sibling populations : a genetic map for the potato cyst nematode constructed using a "pseudo-F2" mapping strategy

A mapping strategy is described for the construction of a linkage map of a non-inbred species in which individual offspring genotypes are not amenable to marker analysis. After one extra generation of random mating, the segregating progeny was propagated, and bulked populations of offspring were analyzed. Although the resulting population structure is different from that of commonly used mapping populations, we show that the maximum likelihood formula for a normal F2 is applicable for the estimation of recombination. This "pseudo-F2" mapping strategy, in combination with the development of an AFLP assay for single cysts, facilitated the construction of a linkage map for the potato cyst nematode Globodera rostochiensis. Using 12 pre-selected AFLP primer combinations, a total of 66 segregating markers were identified, 62 of which were mapped to nine linkage groups. These 62 AFLP markers are randomly distributed and cover about 65% of the genome. An estimate of the physical size of the Globodera genome was obtained from comparisons of the number of AFLP fragments obtained with the values for Caenorhabditis elegans. The methodology presented here resulted in the first genomic map for a cyst nematode. The low value of the kilobase/centimorgan (kb/cM) ratio for the Globodera genome will facilitate map-based cloning of genes that mediate the interaction between the nematode and its host plan

An online catalogue of AFLP markers covering the potato genome

An AFLP marker catalogue is presented for gene mapping within cultivated potato. The catalogue is comprised of AFLP fingerprint images of 733 chromosome-specific AFLP markers which are mapped relative to 220 RFLP loci, isozyme loci, morphological characteristics and disease resistance traits. Use of the catalogue is based on identification of common AFLP markers which are visually recognized on autoradiogram images as co-migrating bands in fingerprints generated from different genotypes. Images of AFLP fingerprints combined with detailed information on the genomic location of all AFLP markers are available at URL: http://www.spg.wau.nl/pv/aflp/catalog.htm. It is demonstrated that the comparison of autoradiogram images and subsequent identification of common AFLP markers solely are efficient means for alignment of linkage groups and mapping target genes

Imaging Collagen in Intact Viable Healthy and Atherosclerotic Arteries Using Fluorescently Labeled CNA35 and Two-Photon Laser Scanning Microscopy

We evaluated CNA35 as a collagen marker in healthy and atherosclerotic arteries of mice after both ex vivo and in vivo administration and as a molecular imaging agent for the detection of atherosclerosis. CNA35 conjugated with fluorescent Oregon Green 488 (CNA35/OG488) was administered ex vivo to mounted viable muscular (uterine), elastic (carotid), and atherosclerotic (carotid) arteries and fresh arterial rings. Two-photon microscopy was used for imaging. CNA35/OG488 labeling in healthy elastic arteries was compared with collagen type I, III, and IV antibody labeling in histologic sections. For in vivo labeling experiments, CNA35/OG488 was injected intravenously in C57BL6/J and apolipoprotein E −/− mice. Ex vivo CNA35/OG488 strongly labeled collagen in the tunica adventitia, media, and intima of muscular arteries. In healthy elastic arteries, tunica adventitia was strongly labeled, but labeling in tunica media and intima was prevented by endothelium and elastic laminae. Histology confirmed the affinity of CNA35 for type I, III, and IV collagen in arteries. Strong CNA35/OG488 labeling was found in atherosclerotic plaques. In vivo applied CNA35/OG488 minimally labeled the tunica intima of healthy carotid arteries. Atherosclerotic plaques in apolipoprotein E −/− mice exhibited large uptake. CNA35/OG488 imaging in organs revealed endothelium as a limiting barrier for in vivo uptake. CNA35/OG488 is a good molecular imaging agent for atherosclerosis

Incidence and treatment results of Endurant endograft occlusion

Objective: The Endurant endograft (Medtronic Inc, Minneapolis, Minn) is a new-generation device specifically developed to perform well in complex abdominal aortic aneurysm anatomy. Previous reports on the 1- and 2-year results of endovascular aneurysm repair (EVAR) with the Endurant endograft showed excellent outcome, including prevention of migration and type I endoleaks, but occurrence and outcome of post-EVAR occlusion have not been determined in a large multicenter patient cohort with midterm follow-up, which is the objective of this study. Methods: Data of consecutive patients treated with the Endurant from December 2007 to April 2012 in three Dutch tertiary vascular referral hospitals were prospectively gathered and retrospectively analyzed. Follow-up consisted of regular office visits, computed tomography angiography at 1 and 12 months after EVAR, and subsequently, duplex ultrasound imaging or computed tomography angiography at regular intervals. Patients with ruptured aneurysms or with earlier abdominal aortic surgery were excluded. The incidence and clinical outcome of endograft occlusions were analyzed. An expert review board assessed all cases in the search for possible causes of occlusion. Results: Included were 496 patients (87.7% male), who were a median age of 74 years (range, 68-78 years). Median follow-up was 1.7 years (range, 0-4.6 years). Twenty graft occlusions (4.0%) occurred during follow-up. Median time between primary EVAR and detection of the occlusion was 1 month, with 55% occurring ≤60 postoperative days and 90% ≤1 year. No association was found between occlusion and sex (P =.28), age (P =.96), or use of an aortouniiliac device (P =.66). Technical error was the considered cause of the occlusion in 12 patients (60%). The estimated freedom from occlusion was 98.4% at 30 days, 95.7% at 1 year, and 95.3% at 3 years. Presenting symptoms of occlusion were acute limb ischemia in 50%. Treatment was surgical (75%) or percutaneous (25%). Successful revascularization was achieved in 17 of 20 patients, but reocclusions occurred in five, resulting in a transfemoral amputation in one patient. Occlusion-related mortality was 0.6% (3 of 496). Conclusions: At a median follow-up of 1.7 years, Endurant endograft occlusion occurred in 4.0% of 496 patients. Most occlusions occurred ≤2 months after EVAR, and rarely after 1 year. A technical justification for occlusion could be found for 60% of patients. A more liberal intraoperative and early postoperative (re)intervention strategy may reduce the occlusion rates and improve outcome

Nε-(Carboxymethyl)lysine-Receptor for Advanced Glycation End Product Axis Is a Key Modulator of Obesity-Induced Dysregulation of Adipokine Expression and Insulin Resistance

OBJECTIVE: Dysregulation of inflammatory adipokines by the adipose an important role in obesity-associated insulin resistance. Pathways this dysregulation remain largely unknown. We hypothesized that the advanced glycation end products (RAGEs) and the ligand Nepsilon-(carboxymethyl)lysine (CML) are increased in adipose tissue moreover, that activation of the CML-RAGE axis plays an important role obesity-associated inflammation and insulin resistance. APPROACH AND this study, we observed a strong CML accumulation and increased RAGE in adipose tissue in obesity. We confirmed in cultured human that adipogenesis is associated with increased levels of CML and RAGE Moreover, CML induced a dysregulation of inflammatory adipokines in via a RAGE-dependent pathway. To test the role of RAGE in obesity- inflammation further, we constructed an obese mouse model that is RAGE (ie, RAGE-/-/LeptrDb-/- mice). RAGE-/-/LeptrDb-/- mice displayed an inflammatory profile and glucose homeostasis when compared with RAGE+/+/LeptrDb-/- mice. In addition, CML was trapped in adipose tissue RAGE+/+/LeptrDb-/- mice but not in RAGE-/-/LeptrDb-/-. RAGE-mediated adipose tissue provides a mechanism underlying CML accumulation in and explaining decreased CML plasma levels in obese subjects. Decreased plasma levels in obese individuals were strongly associated with insulin resistance. CONCLUSIONS: RAGE-mediated CML accumulation in adipose activation of the CML-RAGE axis are an important mechanism involved in dysregulation of adipokines in obesity, thereby contributing to the of obesity-associated insulin resistance