Platelet cold agglutinins: a flow cytometric analysis.

Spontaneous EDTA-independent cold platelet agglutination is a rare phenomenon that produces pseudothrombocytopenia when blood samples are analyzed in automated cell counters. We report a case of platelet cold agglutinins and an analysis by flow cytometry. A 49 year old woman presented with abnormal vaginal bleed secondary to uterine fibroids. Platelet clumping was observed in blood samples taken in EDTA-, heparin- and citrate-containing tubes. In flow cytometric tests, patient serum agglutinated 16% of normal platelets at 22 degrees C, and 7% of platelets after incubation at 37 degrees C; in contrast, 3% and < 1% of platelets were agglutinated at 22 and 37 degrees C, respectively, after incubation with normal serum. Minimal agglutination (< 10%) was observed with patient serum at a titre of 1:5 or at temperatures > 30 degrees C. After incubation at 4 degrees C, IgM antibody and C3 were increased on the patient's platelets; no significant amount of IgM or C3 was detected on normal platelets. The specificity of the platelet cold agglutinin was determined by competitive inhibition by monoclonal anti-CD41(GPIIbIIIa). Before the addition of monoclonal antibody, patient's serum agglutinated 16% of normal platelets at 22 degrees C; after addition of anti-CD41 only 2% of the platelets were agglutinated. This blocking effect was not observed with anti-CD42. The patient's platelets functioned normally as determined by CD62 and CD63 expression in response to thrombin, normal platelet aggregation in response to collagen, ADP, and ristocetin, and a normal template bleeding time. In summary, platelet agglutination by a platelet cold agglutinin was quantitated by flow cytometry, the responsible antibody was characterized as a low titre IgM with minimal activity > 30 degrees C, and competitive binding studies supported the GPIIbIIIa complex as the binding site for the antibody. Since the antibody did not affect platelet function, we believe that these patients will not suffer complications from their platelet cold agglutinin, but it could pose a problem under circumstances such as cardiac surgery with hypothermia.     

Appendizitissonographie durch chirurgen — Erfahrungssache? Eine prospektive studie

A number of studies have shown that ultrasound has an advantage over physical examination in the diagnosis of acute appendicitis. Most of these studies were conducted by experts in the field of ultrasonography. In this study the influence of experience on the results of the sonography of actue appendicitis were evaluated. All 203 patients admitted to our unit between December 1990 and December 1992 were examined physically and sonographically by a team of surgeons consisting of one experienced sonographer and six inexperienced surgical trainees. Laparotomy was performed in 136 patients (46%). Appendicitis was demonstrated histologically in 119 cases (39.4%). Initial clinical findings were positive in 87 (28.8%). Sonography was positive in 119 patients (39.4%). The 163 patients not operated on demonstrated other pathology on ultrasound in 60 cases (19.9%). The rate of negative laparotomies amounted to 7.2% in our study. Sensitivity and specificity for the sonographic diagnosis were 92% and 95%, respectively. They were only 81% and 80% for physical examination. Overall accuracy was 92% for sonography. Sensitivity and specificity for the inexperienced surgeons were 87% and 93%, respectively, while the experienced surgeon reached values of 97% and 98%, respectively. The results of both groups are comparable with values in the literature, suggesting that ultrasound evaluation of appendicitis is not a diagnostic tool limited to a few experienced sonographers.     

Hypoxia aggravates lipopolysaccharide-induced lung injury

The animal model of inflammatory response induced by intratracheal application of lipopolysaccharide includes many typical features of acute lung injury or the acute respiratory distress syndrome. A number of experimental investigations have been performed to characterize the nature of this injury more effectively. In inflammatory conditions, hypoxia occurs frequently before and in parallel with pulmonary and non-pulmonary pathological events. This current study was designed to examine the in vivo effect of hypoxia as a potentially aggravating condition in endotoxin-induced lung injury. Lipopolysaccharide, 150 microg, was instilled intratracheally into rat lungs, and thereafter animals were exposed to either normoxia or hypoxia (10% oxygen). Lungs were collected 2, 4, 6 and 8 h later. Inflammatory response and tissue damage were evaluated by quantitative analysis of inflammatory cells and mediators, surfactant protein and vascular permeability. A significantly enhanced neutrophil recruitment was seen in lipopolysaccharide-animals exposed to hypoxia compared to lipopolysaccharide-animals under normoxia. This increased neutrophil accumulation was triggered by inflammatory mediators such as tumour necrosis factor-alpha and macrophage inflammatory protein-1beta, secreted by alveolar macrophages. Determination of vascular permeability and surfactant protein-B showed enhanced concentrations in lipopolysaccharide-lungs exposed to hypoxia, which was absent in animals previously alveolar macrophage-depleted. This study demonstrates that hypoxia aggravates lipopolysaccharide injury and therefore represents a second hit injury. The additional hypoxia-induced inflammatory reaction seems to be predominantly localized in the respiratory compartment, underlining the compartmentalized nature of the inflammatory response.     

Flavonoid glycosides from Alchemilla speciosa

The new flavonol glycosides quercetin 3- O-beta-(2'- O-alpha- L-rhamnopyranosyl)-glucopyranoside uronic acid and kaempferol 3- O-beta-(2'- O-alpha- L-rhamnopyranosyl)-glucopyranoside uronic acid were isolated from the leaves of Alchemilla speciosa Buser together with 13 known flavonol and flavone glycosides and (+)-catechin. The structures were determined by spectroscopic methods.     

Safety and tolerability of oral daily and intermittent ibandronate are not influenced by age

OBJECTIVE: The risk of osteoporosis increases exponentially with age. Elderly patients, who are often frail, have declining functional status and take multiple medications, and require osteoporosis therapies that are not only effective, but also very well tolerated. Ibandronate is a potent nitrogen-containing bisphosphonate that can be given intermittently with extended between-dose intervals. Oral daily and intermittent ibandronate (interval between doses > 2 mo) was found to significantly reduce the risk of new morphometric vertebral fractures by 62% and 50%, respectively, compared with calcium and vitamin D supplementation alone. We investigated the effect of age on the safety profile of oral daily and intermittent ibandronate, with particular emphasis on the upper gastrointestinal (GI) safety profile of ibandronate. METHODS: A predefined subgroup analysis examined the tolerability of oral ibandronate in women aged < 70 and > or = 70 years. RESULTS: The incidence of adverse events in patients aged > or = 70 years receiving oral daily and intermittent ibandronate was similar and comparable to placebo. The incidence of upper GI adverse events, including dyspepsia and esophagitis, was also similar between the 2 treatment groups and placebo. CONCLUSION: Older patients (> or = 70 yrs) receiving oral daily and intermittent ibandronate are at no greater risk of adverse events than older patients receiving placebo. Older patients were at no greater risk of upper GI adverse events than younger patients or patients receiving placebo. As a result of the good efficacy and tolerability observed in this trial, a once-monthly oral regimen of ibandronate is in late-stage clinical development.     

Steroidogenesis and extracellular cAMP accumulation in adrenal tumor cell cultures.

ACTH stimulated steroidogenesis and cAMP (adenosine 3',5'-monophosphate) accumulation in an adrenocortical mouse tumor cell line (clone Y1) with Kd values which differed by more than one order of magnitude (5.2 X 10(-11) M and 7 X 10(-10) M, respectively). All of the cAMP formed in response to added ACTH appeared extracellularly in 5- or 30-min incubations. ACTH, at 5 and 10 muU/ml, stimulated steroidogenesis to 25% and 40% of maximum activity; and increased the extracellular accumulation of cAMP 1.4-fold and 2.3-fold, respectively. The effects of ACTH appeared to be via an action on intracellular ATP, specific for cAMP and dependent on an ACTH-sensitive adenylate cyclase system. These observations indicate that ACTH increases cAMP accumulation in Y1 cells at virtually all steroidogenic concentrations and suggest that cAMP is an essential component of ACTH-stimulated steroidogenesis.     

Allogeneic or autologous bone marrow transplantation (BMT) for non-Hodgkin's lymphoma (NHL): results of a provincial strategy. Ontario BMT Network, Canada

In 1986, the bone marrow transplant centers in Ontario agreed to a strategy for the treatment of patients with NHL. Suitable patients would undergo autotransplant but be referred for allotransplant if they had persistent marrow involvement or an inadequate marrow/stem cell harvest. Data of all patients were recorded in a database. We reviewed this database to compare these transplant modalities with respect to overall survival, rate of relapse and treatment-related mortality. Between January 1986 and August 1997, 429 patients underwent BMT for NHL - 385 autotransplants and 44 allotransplants. Sixty-eight percent of patients received their transplant for aggressive NHL, while the others had indolent lymphoma. Three-year actuarial survival did not differ between allogeneic and autologous BMT: 71% vs 62%, respectively (P = 0.5330 by log-rank testing). Three-year actuarial rate of relapse was lower after allotransplant than autotransplant: 6% vs 41%, respectively (P = 0.0006 by log-rank testing). Treatment-related mortality was higher after allotransplant than autotransplant: 23% vs 6%, respectively (P = 0.001 by chi2 analysis). For further comparison, autotransplant patients were randomly matched 2:1 with the allotransplant patients for age +/- 5 years, disease status at BMT, disease histology, and year of BMT. In the matched comparison, survival did not differ (relative risk of death after allotransplant: 0.711 (95% CI: 0.309-1.637)). Relapse rate was significantly lower in the allotransplant group (relative risk of relapse for allotransplant: 0.190 (95% CI: 0.043-0.834)) and treatment-related mortality was not significantly different (relative risk for allotransplant: 1.425 (95% CI: 0.527-3.851)). In conclusion, a review of a provincial strategy for treatment of NHL, shows that survival is not different after allogeneic or autologous BMT, but the rate of relapse is lower after allotransplant. These data support continuing the current provincial strategy.     

Small-molecule XIAP inhibitors derepress downstream effector caspases and induce apoptosis of acute myeloid leukemia cells

We tested the effects of small-molecule XIAP antagonists based on a polyphenylurea pharmacophore on cultured acute myelogenous leukemia (AML) cell lines and primary patient samples. X-linked inhibitor of apoptosis protein (XIAP) antagonist N-[(5R)-6-[(anilinocarbonyl)amino]-5-((anilinocarbonyl){[(2R)-1-(4-cyclohexylbutyl)pyrrolidin-2-yl]methyl}amino)hexyl]-N-methyl-N'-phenylurea (1396-12), but not a structurally related control compound, induced apoptosis of primary leukemia samples with a lethal dose (LD50) of less than 10 microM in 16 of 27 (60%) samples. In contrast, XIAP antagonist 1396-12 was not lethal to the normal hematopoietic cells in short-term cytotoxicity assays. Response of primary AML specimens to XIAP inhibitor correlated with XIAP protein levels, with higher levels of XIAP associated with sensitivity. The XIAP antagonist 1396-12 induced activation of downstream caspases 3 and 7 prior to the activation of upstream caspase 8 and caspase 9. Apoptosis induction was also independent of B-cell lymphoma protein-2 (Bcl-2) or caspase 8, indicative of a downstream effect on apoptotic pathways. Thus, polyphenylurea-based XIAP antagonsists directly induce apoptosis of leukemia cells and AML patient samples at low micromolar concentrations through a mechanism of action distinct from conventional chemotherapeutic agents.