Detection of circulating human platelet fragments by using monoclonal antibodies against platelet glycoprotein IIb-IIIa complex

A radioimmunoassay (RIA) for the detection of platelets and platelet fragments was developed. A sandwich of two monoclonal antibodies directed against the platelet-specific glycoprotein complex IIb-IIIa (GP IIb-IIIa) was used in this assay. A discontinuous 7.5-20% (v/v) albumin gradient was applied to separate platelets and their fragments of various sizes. In platelet suspensions fractionated in this way, we observed that particles smaller than normal platelets still carried the GP IIb-IIIa antigens. This procedure enabled us to detect platelet-derived particles in platelet-rich plasma from thrombocytopenic patients.     

Severe Electrolyte Disturbances in Lethal Subclavian Catheter Dialysis

Abstract: Syncope, irreversible shock, and respiratory and circulatory arrest developed in a 54-year-old diabetic man chronically dialyzed with a subclavian catheter (SC) minutes after the end of a dialysis session. Resuscitation attempts remained unsuccessful. During the resuscitation attempt, a blood analysis showed severe hyponatremia, acidosis, and hypochloremia. Respiratory and cardiac arrest developed during dialysis in a 64-year-old woman on chronic SC dialysis. Resuscitation was unsuccessful; chloremia levels were 79 mEq/L, and calcemia levels were 20 mg%. Both patients were dialyzed with a standard dialysate solution. The reasons for the electrolyte disturbances could not be explained technically. The autopsy showed myocardial perforation by the SC and accumulation in the pericardium of the fluids administered during the resuscitation attempt (e.g., glucose 5%, bicarbonate, Ca gluconate, human albumines), thus explaining the erroneous electrolyte results. The reason for the perforation was a too-rigid central femoral vein catheter, erroneously labeled a subclavian catheter by the supplying firm. Because of a syndrome of progressive vena subcla-via and vena cava sclerosis with insufficient arterial phase flow and venous-phase bleeding around the puncture site during single-needle dialysis, the original SC had to be replaced by a longer one with the tip located in the atrium (this SC was actually a femoral catheter). Analysis of the fluid aspirated through the SC can determine the diagnosis in sudden death of SC dialysis patients.     

Alterations in the cortical thymic epithelium of rats after in vivo exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): an (immuno)histological study.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces thymic atrophy in rats. The present study was initiated to provide (immuno)histological data on the mechanism of action. Juvenile male Wistar rats were orally intubated once with 50 or 150 micrograms/kg TCDD. They were euthanized 4 or 10 days thereafter, or were allowed to stay alive until Day 20 or 26. Growth retardation occurred rapidly in all TCDD-treated animals. Lethality was demonstrated within 20-21 days after administration. At Days 4 and 10 after intubation, thymic atrophy was shown by reduction of thymic weight and cortex/medulla ratio. Staining patterns for T-cell markers in the atrophic thymuses coincided with the reduction of cortical areas. There was no evidence indicating that the effects were indirectly caused by stress. TCDD-induced thymic atrophy persisted until Day 26 after administration. Immunohistochemical analysis revealed prominent changes in the cortical thymic epithelium at the 150-micrograms/kg dose level. First, in the cortex epithelial cell aggregates were observed both at Day 4 and at Day 10 after administration. Apparently, the architecture of the epithelium had changed in these animals. Second, at 10 days after administration epithelial cells were found with the simultaneous expression of markers that in the normal uninvoluted thymus only occur either in the subcapsular/medullary area or in the cortex. This phenotype points to an unusual stage of differentiation. We conclude that TCDD exposure affects the cortical epithelium of the rat thymus at a high dose level. Apparently, it disturbs the epithelial network and interfers with the differentiation of epithelial cells.     

Keratin and vimentin distribution patterns in the epithelial structures of the cannie anal region

The intermediate filament labeling pattern of the epithelial structures of the canine anal region was studied with different polypeptide specific keratin monoclonal antibodies (MoAbs) and with a monoclonal and polyclonal vimentin antibody. The epithelial structures in this region could be discriminated and characterized by differences in their keratin staining pattern. The basal cells in the different epithelial structures showed a similar staining pattern characterized by reactivity with MoAbs staining keratins 5, 8, 14, and 17. Columnar epithelial cells showed a completely different phenotype mostly characterized by reactivity with MoAbs staining keratins 7, 5, 8, 18, and 19. A restricted number of differentiated perianal gland cells showed perinuclear vimentin staining. Myoepithelial cells did not stain for vimentin, but, as other basal cells, were positive for MoAbs staining keratins 5, 8, 14, and 17.© Willey-Liss, Inc.     

The gap between legal rules and practice in advertising non-registered pharmaceutical products : A new method of analysis

The market of non-registered pharmaceutical products is growing fast in number and overall costs, not only in the Netherlands, but also in other European countries. These products often give the impression that the consumer may expect 'an effect as from a drug'. Legally, there is a clear distinction between 'drugs' and 'commodities' in the Netherlands; the question is whether legislation and practice concur. In an investigation we analysed texts of advertisements for non-registered pharmaceutical products published in a popular magazine. A method was developed, based on the legal definition of a drug and jurisprudence, to determine in a qualitative and quantitative way the application of medicinal claims. It transpired that in 65% of the analysed advertisements explicit or implicit claims were made. These products should therefore be subject to drugs legislation. Thus, in the Netherlands there is a gap between legislation and practice in advertising non-registered pharmaceutical products.     

Determinants of blood flow and ultrafiltration in continuous arteriovenous haemodiafiltration: theoretical predictions and laboratory and clinical observations

In continuous arteriovenous haemofiltration (CAVH) or haemodiafiltration (CAVHD), it is important to obtain an adequate blood flow through the haemofilter to minimise the risk of excessive haemoconcentration and clotting. In this study we determined the resistance to blood flow of the extracorporeal device as well as the hydraulic permeability of the filter membrane is intensive care patients treated with CAVHD. Data were obtained for CAVH catheters and Scribner shunts and for a polyacrylonitrile (AN-69) plate filter, an AN-69 capillary filter and a polysulphone (PS) capillary filter. In accordance with recent literature we also predicted the resistance to flow by using Poiseuille's law and a formula for the estimation of blood viscosity. Although with all three filters an adequate blood flow was usually obtained, the resistance to blood flow was 2-3 times greater than the predicted value. With continued use of the filter, resistance to blood flow remained largely unchanged. When, in the laboratory, the AN-69 capillary filter was perfused with saline and with a viscous sucrose solution, the resistance to flow was only 1.4 time the predicted value, a difference that might result from small deviations of the capillary diameter. When perfused with blood, the resistance was 2.6 times greater than the predicted value. This was largely explained by gross underestimation of blood viscosity in these patients. By combining laboratory data on filter resistance during saline perfusion and a more accurate estimation of blood viscosity, a reasonably accurate prediction of blood flow rate would be feasible. In the clinic the hydraulic permeability of the filters decreased with time.(ABSTRACT TRUNCATED AT 250 WORDS)     

Platelet-Associated IgG in Thrombocytopenia: A Comparison of Two Techniques

The results obtained in the analysis of 130 thrombocytopenic patients with a radioimmunoassay (RIA) for platelet-associated IgG (PA-IgG) and the platelet suspension immunofluorescence test (PIFT) were compared. The RIA was positive in 33 of 41 (82.9%) patients with idiopathic thrombocytopenia (ITP) and in 51 of 79 (64.4%) patients with secondary thrombocytopenia (STP). The PIFT was positive in 37 of the 41 (90.2%) ITP patients and in 57 of the 79 (72.2%) STP patients. Sensitivity and specificity for the diagnosis of ITP of both tests were comparable: 82.9 and 40.9% for the PA-IgG(RIA) and 90.2 and 36.7% for the PIFT. A significant positive correlation was observed between the mean amount of PA-IgG measured and the height of PIFT scores with anti-IgG. Of 38 discrepancies between PA-IgG(RIA) and PIFT with anti-IgG, 15 were due to borderline results, 17 were associated with abnormal platelet-size distribution and 20 were associated with occurrence of IgM antibodies. These results suggest influences of platelet fragments and/or aggregates on accurate measurement of PA-IgG. Both fragments and aggregates escape from accurate platelet counting, while their contribution to the total IgG content remains. Therefore, a falsely elevated PA-IgG (RIA) may be measured.     

Differential expression of cell cycle and apoptosis related proteins in colorectal mucosa, primary colon tumours, and liver metastases

AIMS: Tumour cell growth results from a disturbance in the balance between the rate of proliferation and cell death. In this study, proteins involved in the regulation of cell cycle arrest and apoptosis were studied as possible factors responsible for uncontrolled cell growth in colorectal cancer. METHODS: The expression of proteins involved in these processes was investigated in 48 metastases from patients with colorectal cancer and compared with eight normal colon mucosa samples and 14 primary tumours. Both primary tumours and metastases were obtained from eight patients. The expression of thymidylate synthase (TS), p53, retinoblastoma protein (Rb), Fas receptor, Fas ligand, bcl-2, mcl-1, bax, and bcl-x was measured using immunohistochemistry. Proliferation was determined by Ki67 staining, whereas apoptosis was assessed by M30 immunostaining, which recognises cleaved cytokeratin 18. RESULTS: In the limited number of cases in which paired comparisons were possible, the expression of TS and Ki67 was significantly higher in metastases than in the matched primary tumour samples (p = 0.014 and 0.016, respectively), whereas Rb expression was lower in metastases than in primary tumours (p = 0.024). Fas receptor expression was high in normal mucosa but absent in primary tumours and metastases, whereas the opposite was seen for p53. The expression of bax, mcl-1, and bcl-x in normal mucosa was more apical than that seen in malignant cells, where a more diffuse expression pattern was seen (p < 0.04). Apoptosis was more abundant in primary tumours than in metastases. CONCLUSIONS: These results demonstrate that proliferation and apoptosis are disturbed during colorectal cancer progression, and this is accompanied by loss of Rb and Fas expression, the accumulation of p53 and TS, and changes in the expression patterns of bax, mcl-1, and bcl-xl.