Plasma P-selectin is increased in thrombotic consumptive platelet disorders

P-selectin is a 140-kD protein found in the alpha-granules of platelets and the Weibel-Palade bodies of endothelial cells that on cell activation is expressed on the cell surface and also secreted into the plasma. The secreted form of P-selectin, like plasma P-selectin, differed from platelet membrane P-selectin in that its molecular mass was approximately 3 kD lower under reducing conditions. Both the secreted and plasma forms of P-selectin contained cytoplasmic sequence as determined by Western blot analysis with an affinity-purified rabbit anti-P-selectin cytoplasmic peptide antibody. We have measured plasma P- selectin and beta-thromboglobulin (beta TG) concurrently in (1) patients with consumptive thrombotic disorders, including disseminated intravascular coagulation (DIC), heparin-induced thrombocytopenia (HIT), and thrombotic thrombocytopenic purpura (TTP)/haemolytic uremic syndrome (HUS); (2) patients with idiopathic thrombocytopenic purpura (ITP); and (3) healthy controls. Patients with DIC, HIT, and TTP/HUS, but not ITP, had significantly elevated plasma P-selectin and beta TG levels when compared with their age-matched healthy controls. The increased plasma P-selectin and beta TG in patients with thrombotic disorders were likely to be the result of in vivo platelet and endothelial cell damage or activation. We also found that avoidance of veno-occlusion and other tedious measures customarily taken during blood collection and sample preparation to prevent in vitro platelet activation did not affect plasma P-selectin assay results. In addition, plasma P-selectin levels were not influenced by the presence of renal failure or heparin administration. These results indicate that plasma P- selectin may be a useful new marker for thrombotic diseases.     

Protein expression changes in a cell system of beta-cell maturation reflect an acquired sensitivity to IL-1β

Aim/hypothesis Type 1 diabetes mellitus (T1DM) is caused by specific destruction of the pancreatic beta cells in the islets of Langerhans. Increased sensitivity to cytokines, in particular to interleukin-1 (IL-1) seems to be an acquired trait during beta-cell maturation. In response to cytokines both protective and deleterious mechanisms are induced in beta cells, and when the deleterious prevail, T1DM develops. The aims of this study were to identify perturbation in protein patterns (PiPP) associated with beta-cell maturation, and compare these changes to previous analyses of IL-1 exposed rat islets. For this purpose, proteome analyses were carried out using a cell-line, which matures from a glucagon-producing pre-beta-cell phenotype (NHI-glu) to an insulin-producing beta-cell phenotype (NHI-ins). We have previously shown that this maturation is accompanied by acquired sensitivity to the toxic effects of IL-1.Methods 2D-gel electrophoresis was used to separate the proteins and MALDI-MS and database searches were performed to identify the proteins.Results During beta-cell maturation 135 protein spots out of 2239 detectable changed expression levels. Of these, 74 were down-regulated, 44 up-regulated, 16 were suppressed and 1 was expressed de novo. Using MALDI-MS, positive identification was obtained for 93 out of the 135 protein-spots revealing 97 different proteins. Of these, 22 proteins were in common with changes identified in previous proteome analysis of perturbation in protein pattern in IL-1 exposed rat islets. Several of the proteins were present in more than one spot suggesting post-translational modification.Conclusion/interpretation Several proteins and protein modifications were identified that could be critically involved in beta-cell maturation, insulin-gene expression and the acquired IL-1 sensitivity.Abbreviations T1DM Type 1 diabetes mellitus - PiPP perturbation in protein pattern - NO nitric oxide - iNOS inducible nitric oxide synthase - 2D-GE 2 dimensional gel electrophoresis - IEF isoelectric focusing - NEPHGE non-equilibrium pH-gradient electrophoresis - WF Wistar Furth - BB Bio Breeding - PDX-1 pancreatic duodenum homeobox 1 - ASS argininosuccinate synthetase - HSP heat shock protein - Picot PKC-interacting cousin of thioredoxin - JNK Jun N-terminal kinase - VDAC voltage-dependent anion channel - GST glutathione-S-transferase - Mw molecular weight - pI isoelectric point     

Influence of clinical status on the association between plasma total and unbound bilirubin and death or adverse neurodevelopmental outcomes in extremely low birth weight infants

Objectives: To assess the influence of clinical status on the association between total plasma bilirubin and unbound bilirubin on death or adverse neurodevelopmental outcomes at 18–22 months corrected age in extremely low birth weight infants. Method: Total plasma bilirubin and unbound bilirubin were measured in 1101 extremely low birth weight infants at 5 ± 1 days of age. Clinical criteria were used to classify infants as clinically stable or unstable. Survivors were examined at 18–22 months corrected age by certified examiners. Outcome variables were death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death prior to follow‐up. For all outcomes, the interaction between bilirubin variables and clinical status was assessed in logistic regression analyses adjusted for multiple risk factors. Results: Regardless of clinical status, an increasing level of unbound bilirubin was associated with higher rates of death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss and death before follow‐up. Total plasma bilirubin values were directly associated with death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death before follow‐up in unstable infants, but not in stable infants. An inverse association between total plasma bilirubin and death or cerebral palsy was found in stable infants. Conclusions: In extremely low birth weight infants, clinical status at 5 days of age affects the association between total plasma bilirubin and death or adverse neurodevelopmental outcomes at 18–22 months of corrected age. An increasing level of UB is associated a higher risk of death or adverse neurodevelopmental outcomes regardless of clinical status. Increasing levels of total plasma bilirubin are directly associated with increasing risk of death or adverse neurodevelopmental outcomes in unstable, but not in stable infants.     

Human papillomavirus type 16 (HPV 16) E7 and major histocompatibility complex (MHC) class I and II expression in human keratinocytes in culture

Archives of Virology - The low expression of major histocompatibility complex (MHC) class I antigens on human papillomavirus (HPV)-infected cervical carcinoma cells may be responsible for an...     

In situ hybridization with nonisotopic probes using different detection systems.

In order to evaluate a sensitive nonisotopic in situ hybridization method for routine work in pathology laboratories, we compared seven different detection systems, using digoxigenin- and biotin-labeled probes. The sensitivity of these methods was tested on four cases of cervical condyloma all known to be positive for HPV 6. Four of these methods gave satisfactory results without any background staining. The single biotin method and the single digoxigenin method were equally sensitive, while the two triple biotin methods, using mouse anti-biotin/anti-mouse IgG/alkaline phosphatase mouse anti-alkaline phosphatase or mouse anti-biotin/alkaline phosphatase anti-mouse IgG/alkaline phosphatase mouse anti-alkaline phosphatase as the detection systems, tremendously improved the sensitivity. The enhanced sensitivity of the nonisotopic in situ hybridization method make it useful in investigation of pathologic tissues.