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Circulating cell-free DNA (cfDNA) has been investigated as a screening tool for many diseases. To avoid expensive and time-consuming DNA isolation, direct quantification PCR assays can be established. However, rigorous validation is required to provide reliable data in the clinical and non-clinical context. Considering International Organization for Standardization, as well as bioanalytical method validation guidelines we provide a comprehensive procedure to validate assays for cfDNA quantification from unpurified blood plasma. A 90 and 222 bp assay was validated to study the kinetics of cfDNA after exercise in patients with systemic lupus erythematosus. The assays showed ultra-low limit of quantification (LOQ) with 0.47 and 0.69 ng/ml, repeatability ≤ 11.6% (95% CI: 8.1–20.3), and intermediate precision ≤ 12.1% (95% CI: 9.2-17.7). Incurred sample reanalysis confirmed the precision of the procedure. The additional consideration of pre-analytical factors shows that centrifugation speed and temperature do not change cfDNA concentrations. In SLE patients cfDNA increases ∼2 fold after all out walking exercise, normalizing after 60 min of rest. The established assays allow reliable and cost-efficient quantification of cfDNA in minute amounts of plasma in the clinical setting and can be used as a standard to control pre-analytical factors including cfDNA losses during purification.

Immunophenotypic distinction between neoplastic and reactive T-cell clones can be challenging, as peripheral T-cell lymphomas (PTCLs) lack an immunophenotypic marker of clonality. Systematic screening of 10,510 cases analyzed by immunophenotyping at our institution between 2006 and 2012 resulted in 49 cases with aberrant T-cell populations of unclear significance. Review of patient charts allowed us to assign these cases to three categories. In 21 cases, PTCL could later be confirmed by complementary diagnostics (PTCL group). In 20 cases, follow-up confirmed the reactive nature of the aberrant T-cells (non-PTCL group). Eight cases remained of unclear significance. Neither the population size nor the number of aberrant markers differed significantly between the PTCL and non-PTCL groups. Only loss of CD7 was found significantly more often in patients with PTCL than in patients with non-PTCL (p = 0.037). Our data show that aberrant T-cell populations need to be interpreted in the clinicopathological context, as reactive and neoplastic phenotypes largely overlap.

Recent findings support the hypothesis that the CD34+-cell population in bone marrow and peripheral blood contains hematopoietic and endothelial progenitor and stem cells. In this study, we report that human AC133+ cells from granulocyte colony-stimulating factor–mobilized peripheral blood have the capacity to differentiate into endothelial cells (ECs). When cultured in the presence of vascular endothelial growth factor (VEGF) and the novel cytokine stem cell growth factor (SCGF), AC133+ progenitors generate both adherent and proliferating nonadherent cells. Phenotypic analysis of the cells within the adherent population reveals that the majority display endothelial features, including the expression of KDR, Tie-2, Ulexeuropaeus agglutinin-1, and von Willebrand factor. Electron microscopic studies of these cells show structures compatible with Weibel-Palade bodies that are found exclusively in vascular endothelium. AC133-derived nonadherent cells give rise to both hematopoietic and endothelial colonies in semisolid medium. On transfer to fresh liquid culture with VEGF and SCGF, nonadherent cells again produce an adherent and a nonadherent population. In mice with severe combined immunodeficiency, AC133-derived cells form new blood vessels in vivo when injected subcutaneously together with A549 lung cancer cells. These data indicate that the AC133+-cell population consists of progenitor and stem cells not only with hematopoietic potential but also with the capacity to differentiate into ECs. Whether these hematopoietic and endothelial progenitors develop from a common precursor, the hemangioblast will be studied at the single-cell level.

Wir berichten uber eine 60jahrige Patientin, welche nach haufigen Zeckenstichereignissen histologisch typische juxta-artikulare fibroide Knoten (JFK) und rotlich-livide Hautschwellungen im Sinne einer Acrodermatitis chronica atrophicans (ACA) zeigte. Es erfolgte eine systemische Antibiose mit Ceftriaxon (Rocephin) 2 g/die parenteral uber 21 Tage. Unerwunschte Nebenwirkungen traten nicht auf.