Background Interpreting hematology analytes in children is challenging due to the extensive changes in hematopoiesis that accompany physiological development and lead to pronounced sex- and age-specific dynamics. Continuous percentile charts from birth to adulthood allow accurate consideration of these dynamics. However, the ethical and practical challenges unique to pediatric reference intervals have restricted the creation of such percentile charts, and limitations in current approaches to laboratory test result displays restrict their use when guiding clinical decisions. Methods We employed an improved data-driven approach to create percentile charts from laboratory data collected during patient care in 10 German centers (9,576,910 samples from 358,292 patients, 412,905–1,278,987 samples per analyte). We demonstrate visualization of hematology test results using percentile charts and z-scores (www.pedref.org/hematology) and assess the potential of percentiles and z-scores to support diagnosis of different hematological diseases. Results We created percentile charts for hemoglobin, hematocrit, red cell indices, red cell count, red cell distribution width, white cell count and platelet count in girls and boys from birth to 18 years of age. Comparison of pediatricians evaluating complex clinical scenarios using percentile charts versus conventional/tabular representations shows that percentile charts can enhance physician assessment in selected example cases. Age-specific percentiles and z-scores, compared with absolute test results, improve the identification of children with blood count abnormalities and the discrimination between different hematological diseases. Conclusions The provided reference intervals enable precise assessment of pediatric hematology test results. Representation of test results using percentiles and z-scores facilitates their interpretation and demonstrates the potential of digital approaches to improve clinical decision-making.

We investigated regulation of macrophage prostaglandin production during activation by interferon gamma (IFN-gamma) and lipopolysaccharide (LPS). An in vitro model was established using the mouse macrophage-like cell line RAW 264.7. Cells were cultivated in the presence of IFN-gamma and LPS for up to 48 h and changes in the secretion of nitric oxide (NO.) and tumor necrosis factor alpha (TNF-alpha) were observed as activation markers. Under these conditions a prompt and strong increase in PGE2 production was found in the first 8 h followed by nearly constant generation of PGE2 during the next 40 h. In contrast, the activity of prostaglandin endoperoxide synthase (PGHS), measured as PGE2 production of microsomal protein fractions, was also increased, but reached a clear maximum at 24 h. Recently a second form of PGHS was cloned (PGHS-2) and specific antibodies and mRNA probes for both isoforms are available. PGHS-2 enzyme was expressed maximally after 24 h of activation whereas PGHS-1 was not influenced. In the presence of IFN-gamma and LPS, PGHS-2 mRNA expression reached a maximum at 8 h but PGHS-1 mRNA was not induced during the whole time period. These data indicate that changes in PG synthesis following macrophage activation are due to regulation of PGHS-2 expression.

The immunosuppressive drug cyclosporin A (CsA) has considerable nephrotoxic side effects which seem to be related to its interference with the synthesis of vasoactive prostanoids. Therefore, the molecular mechanism of the effect of CsA on the synthesis of prostaglandin E2 (PGE2) was investigated in rat renal mesangial cells (RMC). CsA effectively inhibited the PGE2 synthesis induced by inflammatory cytokines such as interleukin 1 (IL-1) or tumor necrosis alpha (TNF alpha). The induction by IL-1 and the inhibition by CsA were reflected in the enzyme activity of the cyclooxygenase. The changes in activity could be correlated with the expression of the inducible cyclooxygenase isoform (COX2), which is characterized by its 4.4 kb mRNA: the expression of this enzyme was enhanced by IL-1 and suppressed by CsA on the mRNA and protein level as determined by Northern and Western blot analyses. Suppression of COX2 mRNA was also observed when the message was induced by LPS or ionophore A23187. The expression of the basal cyclooxygenase isoform (COX1), which was constitutively expressed in proliferating mesangial cells, was not affected by IL-1 or CsA. Interferon gamma, which did not induce prostaglandin synthesis or influence COX mRNA expression, augmented the expression of MHC antigens in RMC. This induction was insensitive to CsA treatment. We could thus show that the inducible cyclooxygenase isoform in mesangial cells is a molecular target for CsA providing a possible mechanism for the interference of the drug with the balance of vasoactive prostanoids.

The human monocytic cell line U937 was used as a model system to investigate the effects of glucocorticoids on monocytic differentiation. Upon incubation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) (5 x 10(-9) M) for 48 to 72 h, the immature U937 cells ceased to proliferate and became morphologically and functionally macrophage-like. Preincubation of the cells with glucocorticoids (dexamethasone and prednisolone, 10(-7) and 10(-6) M) but not progesterone (10(-6) M) had marked effects: The cells remained in suspension and developed very little cell-cell interaction. This correlated with decreased expression of the surface molecules ICAM-1 and CD18 as determined by fluorescence-activated cell sorter analysis. The TPA-induced ability of the cells to release lysozyme or to generate reactive oxygen radicals (determined as reduction of nitroblue tetrazolium) was markedly reduced. The induction of cyclooxygenase activity and thus the ability to release prostanoids was almost completely abolished. Inhibition of prostanoid synthesis was also observed when the glucocorticoids were administered 24 or 48 h after TPA. The primary step of TPA induction, the activation and translocation of protein kinase C, however, was not affected by glucocorticoids as determined by activity measurements and Western blot analysis. There was no change in the subsequent TPA-induced induction of c-fos. The down-regulation of the differentiation-related oncogenes c-myc and c-myb was the same in cells treated with TPA in the presence or absence of glucocorticoids. Furthermore, no significant effect of glucocorticoids on the TPA-induced growth arrest was observed. Glucocorticoids thus interfere with TPA-induced functions, which are typical for activated macrophages; however, they do not impair the differentiation process and concomitant growth inhibition.

Significant progress in the investigation of the regulation of prostanoid formation has recently been made by cloning a second gene coding for prostaglandin G/H synthase (PGHS; EC 1.14.99.1). In this study we examined the expression of the two PGHS isoforms during phorbol ester induced monocytic differentiation of human myeloid leukemia cells (U937). Murine and ovine PGHS‐1 probes hybridized to 2.8‐ and 5.5‐kb mRNA species, whereas the murine PGHS‐2 probe hybridized to a 5.3‐kb species. Western blot analysis using antisera to mouse PGHS‐1 and to a synthetic peptide derived from a mouse PGHS‐2‐specific region revealed a band of 70 kDa for PGHS‐1 and a doublet of about 85 kDa for PGHS‐2. Unlike PGHS‐2, which was not expressed in U937 control cells, both PGHS‐1 protein and mRNA were detected in untreated U937 cells. TPA strongly induced PGHS‐2 protein and also increased the amount of PGHS‐1 protein. Correspondingly, a marked induction of PGHS‐2 mRNA was found, but virtually no change in the expression of the PGHS‐1 2.8‐kb mRNA occurred. The induction of both PGHS isoforms turned out to be dexamethasone‐sensitive. The suppression of PGHS‐2 induction was more pronounced. These results suggest that both PGHS‐1 and to a larger extent PGHS‐2 contribute to the upregulation of prostanoid synthesis during monocytic differentiation.