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Summary. Background: Circulating endothelial cells (CECs) have been identified as markers of vascular damage in a variety of disorders, such as myocardial infarction, vasculitis, and transplantation. CD146-driven immunomagnetic isolation has gained widespread use, but the technique is hampered by the lack of a definition of CECs and the absence of a consensus for their enumeration. Aim: To evaluate several variables influencing immunomagnetic isolation of CECs, formulate a definition for CECs and propose a consensus protocol for their enumeration. Methods: We devised a protocol based on CD146-driven immunomagnetic isolation and a subsequent confirmatory step with Ulex-Europaeus-Lectin-1 staining. In a multi-center effort, we evaluated the preanalytical and analytical phases of this protocol. We evaluated the effects of storage, anticoagulation and density centrifugation, and compiled previous experience with this technique. Results: Our protocol permitted unequivocal identification of CECs with acceptable reproducibility. There was an effect of storage time in that median cell numbers declined to only 87.5% of their baseline values during 24 h of storage at 4°C. Recovery was lower with citrate than with ethylene-diamine tetra-acetic acid after 4 h of storage; density centrifugation was also associated with lower recovery. We provide a comprehensive list of technical recommendations and potential pitfalls. Finally, based on our experience with this protocol and a recent consensus workshop, we formulated a working definition for CECs. Conclusion: Our work represents an important step toward consensus regarding the CECs. Our recommendations represent the experience of three major centers and should now be scrutinized by others in the field.

Frailty, the state of increased vulnerability to physical stressors as a result of progressive and sustained degeneration in multiple physiological systems, is common in those with chronic kidney disease (CKD). In fact, the prevalence of frailty in the older adult population is reported to be 11%, whereas the prevalence of frailty has been reported to be greater than 60% in dialysis-dependent CKD patients. Frailty is independently linked with adverse clinical outcomes in all stages of CKD and has been repeatedly shown to be associated with an increased risk of mortality and hospitalization. In recent years there have been efforts to create an operationalized definition of frailty to aid its diagnosis and to categorize its severity. Two principal concepts are described, namely the Fried Phenotype Model of Physical Frailty and the Cumulative Deficit Model of Frailty. There is no agreement on which frailty assessment approach is superior, therefore, for the time being, emphasis should be placed on any efforts to identify frailty. Recognizing frailty should prompt a holistic assessment of the patient to address risk factors that may exacerbate its progression and to ensure that the patient has appropriate psychological and social support. Adequate nutritional intake is essential and individualized exercise programmes should be offered. The acknowledgement of frailty should prompt discussions that explore the future care wishes of these vulnerable patients. With further study, nephrologists may be able to use frailty assessments to inform discussions with patients about the initiation of renal replacement therapy.

Religious concerns may be an important reason why patients decline listing for a renal transplant. These issues may be equally, or even more, important when live donation is discussed. There is good reason to believe that religious concerns play a significant role much more often than clinicians and transplant teams believe. The issue is certainly further compounded by the fact that a few, if any, patients come forward with their religious concerns, not least because issue of transplantation is new to them anyway and because they meet with transplant teams whom they do not know. Health professionals, on the other hand, may wish to avoid this sensitive issue altogether or may lack knowledge on religious issues pertaining to transplantation. Some may be entirely unaware. We encountered a case in clinic that revealed our remarkable lack of knowledge in this regard. Here, we aim to provide an overview on how the different religions view transplantation and organ donation, with an emphasis on practical points for health care professionals who are involved in transplant listing, organ donation and retrieval, and transplantation itself. Knowledge of these facts may provide a background to deal with these issues professionally and appropriately and to increase transplant numbers.

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Recent research has recognised new populations of non-hematopoietic cells in the blood. One of these, circulating endothelial cells (CECs), often defined by the expression of membrane glycoprotein CD146, are rarely found in the blood in health, but raised numbers are present in a wide variety of human conditions, including inflammatory, immune, infectious, neoplastic and cardiovascular disease, and seem likely to be evidence of profound vascular insult. An additional population are endothelial progenitor cells, defined by the co-expression of endothelial and immaturity cell surface molecules and also by the ability to form colonies in vitro. Although increased numbers of CECs correlate with other markers of vascular disease, questions remain regarding the precise definition, cell biology and origin of CECs. For example, they may be damaged, necrotic or apopototic, or alive, and could possess procoagulant and/or proinflammatory properties. However, since these cells seem to be representative of in situ endothelium, their phenotype may provide useful information. Indeed, whatever their phenotype, there is growing evidence that CECs may well be a novel biomarker, the measurement of which will have utility in various clinical settings related to vascular injury. Despite this promise, progress is impeded by the diversity of methodologies used to detect these cells. Accordingly, results are sometimes inconclusive and even conflicting. Nevertheless, increased CECs predict adverse cardiovascular events in acute coronary syndromes, suggesting they may move from being simply a research index to having a role in the clinic. The objective of the present communication is to condense existing data on CECs, briefly compare them with progenitor cells, and summarise possible mechanism(s) by which they may contribute to vascular pathology.

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Renal activity and smoldering disease is difficult to assess in anti-neutrophil cytoplasmic antibody-associated vasculitis (AAV) because of renal scarring. Even repeated biopsies suffer from sampling errors in this focal disease especially in patients with chronic renal insufficiency. We applied capillary electrophoresis coupled to mass spectrometry toward urine samples from patients with active renal AAV to identify and validate urinary biomarkers that enable differential diagnosis of disease and assessment of disease activity. The data were compared with healthy individuals, patients with other renal and non-renal diseases, and patients with AAV in remission. 113 potential biomarkers were identified that differed significantly between active renal AAV and healthy individuals and patients with other chronic renal diseases. Of these, 58 could be sequenced. Sensitivity and specificity of models based on 18 sequenced biomarkers were validated using blinded urine samples of 40 patients with different renal diseases. Discrimination of AAV from other renal diseases in blinded samples was possible with 90% sensitivity and 86.7-90% specificity depending on the model. 10 patients with active AAV were followed for 6 months after initiation of treatment. Immunosuppressive therapy led to a change of the proteome toward "remission.