Technical note: Simultaneous carotenoid and vitamin analysis of milk from total mixed ration-fed cows optimized for xanthophyll detection

Concentrations of retinol, α-tocopherol, and major carotenoids in dairy products are often determined simultaneously by liquid chromatography. These compounds have different polarity and solubility; thus, extracting them simultaneously can be difficult and inefficient. In milks with low carotenoid concentrations, the xanthophylls lutein and zeaxanthin may not be completely resolved using common extraction techniques. A simplified method was developed to optimize extraction efficiency and the limit of detection and limit of quantification (LoQ) of lutein and zeaxanthin in bovine milk without decreasing sensitivity to other vitamins or carotenoids. The developed method evaluates lutein, zeaxanthin, β-carotene, retinol, and α-tocopherol simultaneously by ultra-high performance liquid chromatography–photodiode array detection. Common saponification temperatures (40–60°C) and concentrations of KOH in water (10–50% KOH wt/vol) were evaluated. Multiple solvents were evaluated for optimal xanthophyll extraction (diethyl ether, dichloromethane, hexane, and tetrahydrofuran) following saponification. The limit of detection and LoQ were defined as 3:1 and 10:1 signal-to-noise ratio, respectively. All experiments were performed in triplicate. The optimal saponification procedure was a concentration of 25% KOH at either 40 or 50°C. Saponified extracts solubilized in solutions containing diethyl ether had greater concentrations of lutein- than hexane- or tetrahydrofuran-based solutions, with peak areas above LoQ values. The solution containing diethyl ether solubilized similar concentrations of retinol, α-tocopherol, and β-carotene when compared with other solutions. The proposed optimized method allows for the simultaneous determination of carotenoids from milk with increased lutein and zeaxanthin sensitivity without sacrificing recovery of retinol, α-tocopherol, and β-carotene.     

P2Y11 Agonism Prevents Hypoxia/Reoxygenation- and Angiotensin II-Induced Vascular Dysfunction and Intimal Hyperplasia Development

Vascular dysfunction in cardiovascular diseases includes vasomotor response impairments, endothelial cells (ECs) activation, and smooth muscle cells (SMCs) proliferation and migration to the intima. This results in intimal hyperplasia and vessel failure. We previously reported that activation of the P2Y11 receptor (P2Y11R) in human dendritic cells, cardiofibroblasts and cardiomyocytes was protective against hypoxia/reoxygenation (HR) lesions. In this study, we investigated the role of P2Y11R signaling in vascular dysfunction. P2Y11R activity was modulated using its pharmacological agonist NF546 and antagonist NF340. Rat aortic rings were exposed to angiotensin II (AngII) and evaluated for their vasomotor response. The P2Y11R agonist NF546 reduced AngII-induced vascular dysfunction by promoting EC-dependent vasorelaxation, through an increased nitric oxide (NO) bioavailability and reduced AngII-induced H₂O₂ release; these effects were prevented by the use of the P2Y11R antagonist NF340. Human vascular SMCs and ECs were subjected to AngII or H/R simulation in vitro. P2Y11R agonist modulated vasoactive factors in human ECs, that is, endothelial nitric oxide synthase (eNOS) and endothelin-1, reduced SMC proliferation and prevented the switch towards a synthetic phenotype. H/R and AngII increased ECs secretome-induced SMC proliferation, an effect prevented by P2Y11R activation. Thus, our data suggest that P2Y11R activation may protect blood vessels from HR-/AngII-induced injury and reduce vascular dysfunctions. These results open the way for new vasculoprotective interventions.     

Evaluation of the determinants of flow-mediated radial artery vasodilatation in humans

The relative importance of the early peak response during hyperaemia and of the duration of the hyperaemic phase (t1/2: blood flow velocity half time and AUCt1/2: area under the curve of flow velocity at t1/2) in the magnitude of the flow-dependent vasodilatation of the radial artery was determined in humans. Radial artery diameter was measured continuously in 18 healthy volunteers using an echo-tracking system coupled to a Doppler device for the measurement of the radial blood flow. In 9 subjects, arterial parameters were measured at baseline and during 3 hyperaemic tests performed after 2, 5 or 10 minutes of ischaemia. Reproducibility of the measured parameters was studied in 9 other subjects. Radial artery diameter, AUCt1/2 and t1/2 increased proportionally with the duration of ischaemia. In contrast, the peak flow response was already maximal after 5 minutes of ischaemia. The regression analysis showed that the best fit model after stepwise analysis only included t1/2 (r = 0.85, p < 0.001). There was no correlation between the peak flow values and the duration of hyperaemia (r = 0.29, p = 0.14). These results demonstrate that conduit arteries postischaemic flow-dependent vasodilatation in humans is both determined by the peak value and by the duration of the hyperaemic phase and suggest that these two components must be considered when comparing this index of NO release between different groups of subjects.     

New treatment for ileal pouch-anal or coloanal anastomotic stenosis

Persistent anastomotic stricture following ileal pouch-anal or coloanal anastomoses can be treated by transanal resection using a stapler or a more complex procedure, such as transanal pouch advancement with neoanastomosis. We propose an easier and faster technique, which does not require any particular device. Its long-term functional results are satisfactory in most patients.     

CD4+ T cell responses in mice lacking MHC class II molecules specifically on B cells

The role of B lymphocytes in initiating and maintaining a CD4+ T cell response has been examined using a variety of strategies, but remains controversial because of weaknesses inherent to each of the approaches. Here, we address this issue by measuring CD4+ T cell priming both in mutant mice devoid of B cells and in chimeric animals lacking major histocompatibility complex class II molecules specifically on B cells. We find that peptide and some protein antigens do not require B cells expressing class II molecules, nor B cells themselves, to efficiently prime. This could be demonstrated by the usual lymph node proliferation assay, a rather indirect in vitro measure of priming, and by a direct ex vivo assay of population expansion and activation marker expression. Interestingly, one protein antigen, conalbumin, could not prime in the absence of B cells, but could in the presence of B cells devoid of class II molecules. This finding constrains the possible mechanisms whereby B lymphocytes contribute to the initiation of a CD4+ T cell response, arguing against the importance of surface immunoglobulin-mediated antigen presentation by B cells.     

The influence of positive selection on RAG expression in thymocytes

Many human cancers present deletions of the short arm of chromosome 17, which includes the TP53 locus. We detected a new polymorphism in intron 2 of the TP53 gene using PCR-SSCP and used this polymorphic site as a marker to detect loss of heterozygosity in 135 human tumors (73 soft tissue sarcomas, and 48 colorectal and 14 bladder carcinomas). Heterozygosity for this site was 41.5% in this study group and tumor-specific loss of alleles occurred in 43% of informative cases. Allelic losses were more frequently detected at this site than at that in which restriction fragment length polymorphism (RFLP) is located, as detected by the pHp53B probe. It is concluded that this novel approach has several advantages, including detection of a high incidence of informative cases and minimal tissue requirements.     

Arthrotic sciatica caused by radicular compression of osteophytic origin in the lateral recess. Apropos of 18 cases

The authors report 18 cases of arthrosic sciatica due to toot compression in the lateral recess by posterior corporeal and/or posterior apophyseal osteophytosis. The authors study the clinical and radiological characteristics that may indicate the diagnosis and discuss the different mechanisms by which vertebral arthrosis can lead to radicular compression. When surgery is necessary because of the persistent nature of the sciatica, a broader approach should be undertaken than that required for excision of the disc, in order to explore fully the roots, the multiplicity of possible compression sites being one of the essential characteristics of these cases of arthrosic radiculopathy. Study of the literature and of the series of sciatica patients operated upon by the authors shows that although discal hernia is far from being the most frequent cause of common sciatica, arthrosic compression is a cause that cannot be ignored, especially in aged subjects.