A novel system to diagnose cutaneous adverse drug reactions employing the cellscan--comparison with histamine releasing test and Inf-gamma Releasing Test

Background: There are several mechanisms to describe allergic drug reactions yet the methods to diagnose them are limited. Objective: To compare several conventional clinical and laboratory methods to diagnose skin reactions to drugs to a new method of diagnosing drug reactions by the CellScan system. Methods: The study entailed 21 patients who were diagnosed as suffering from drug eruptions, and 105 healthy controls with no history of drug allergy. The drugs were classified into two groups according to suspicion of causing drug allergy: high and low. Most of the patients were on more than one drug, leading to 41 patient-drug interactions (assays). Histamine releasing test (HRT), interferon (INF)-γ releasing test and CellScan examination were performed on lymphocytes of the patients and controls. Results: The HRT was interpreted as positive in 9 out of 18 (50%) patients and in 13 out of 35 (37%) assays. Based on the INF-γ releasing test, positive results were observed in 16 out of 21 (76%) patients and in 24 out of 41 (59%) assays. In the CellScan test (CST), positive results were observed in 17 out of 21 (81%) patients and in 29 out of 41 (71%) assays. The rate of identifying the drug for eruption in the high suspicion level drugs was 9 out of 22 (41%) assays in the HRT, 20 out of 24 (83%) assays in the INF-γ releasing test, and 21 out of 24 (87%) studies with the CellScan method. The rate of determining of the drug that caused the eruption in the low suspicion level drugs was 4 out of 13 (31%) in the HRT, 4 out of 17 (24%) assays in the INF-γ releasing test, and 8 out of 17 (47%) analyses in the CST. When examined in the CellScan, 99 out of 105 (94%) controls were interpreted as negative. Conclusion: This preliminary study indicates that the CellScan seems to be an easy and promising method for the detection of drugs responsible for adverse skin reactions. In contrast to the HRT and to the Interferon-γ secretion test, the CellScan method is characterized by its ability to track and monitor the reaction of individual cells. By measuring the kinetic parameters of selected cells before and after adding the suspected drug, we were able to identify the culprit drug. The CellScan method had the highest sensitivity, and the interferon-γ secretion test had the highest specificity for detection of the culprit drug. In contrast, the analysis of 105 normal control sera disclosed a high specificity of 94% for the CellScan method.     

Induction of biologically active antineutrophil cytoplasmic antibodies by immunization with human apoptotic polymorphonuclear leukocytes

Translocation of intracellular components to the cell surface during the priming or apoptosis of polymorphonuclear leukocytes (PMN) is an important mechanism for interaction of antineutrophil cytoplasmic antibodies (ANCA) with these antigens. To test the capacity of apoptotic PMN to trigger production of ANCA, six groups of mice were immunized with either live or apoptotic lymphocytes, or with live, apoptotic, formalin-fixed, or lysed PMN. Mice immunized with both live and apoptotic neutrophils developed high titers of antibodies which gave a granular cytoplasmic immunofluorescent pattern. These antibodies were specific for lactoferrin and myeloperoxidase. Following a second intravenous infusion of apoptotic PMNs, mice developed anti-PR3 antibodies. Vasculitis lesions were not found in mice which developed ANCA. The ANCA-containing IgG fraction induced superoxide production by human PMNs. These results support the hypothesis that neutrophil-specific antigens presented on the cell membranes of apoptotic PMN may induce ANCA in the proper conditions.     

Inhibition of ras by farnesylthiosalicylate significantly reduces the levels of autoantibodies in two animal models of the antiphospholipid syndrome

BACKGROUND: Stimulation and proliferation of lymphocytes require activation of Ras. S-farnesylthiosalicylic acid (FTS) is a synthetic substance that detaches Ras from the inner cell membrane and induces its rapid degradation. Antiphospholipid antibodies (aPL) are a heterogeneous group of antibodies detected in patients with antiphospholipid syndrome (APS), which is associated with thrombosis, pregnancy losses, and thrombocytopenia. OBJECTIVE: To examine the effect of FTS treatment on aPL levels in a genetic autoimmune model (the MRL/lpr mice) and in an induced model of APS. METHODS: Female Balb/C mice immunized once with beta2-glycoprotein I (beta2-GPI) in complete Freund's adjuvant (CFA) and female MRL/lpr mice were treated intraperitoneally with either FTS (5 mg/Kg/day) or saline 3-5 times a week. aPL and anti-beta2-GPI antibodies were measured by ELISA. RESULTS: FTS treatment 3 times a week resulted in significant decreases of aPL and anti-beta2-GPI antibodies in both animal models. In contrast, more frequent treatment (5 times a week) had no significant effect on autantibody levels in both animal models. We further compared 2 protocols in the induced APS model, one for alternate day treatment and the other for daily treatment on the first 3 days each week, and found a decrease in autoantibody levels only in the alternate day protocol. CONCLUSIONS: Inhibition of Ras activation by FTS is effective in decreasing autoantibody levels in models of APS. The differential modulation of immune function by alternate day compared to daily treatment may provide better understanding of the role of Ras activation in this system.     

Primary biliary cirrhosis

Primary biliary cirrhosis (PBC) is a chronic, progressive cholestatic liver disease, which is invariably fatal. Circumstantial and indirect evidence suggests that autoimmune mechanisms have a role in the pathogenesis of PBC. Antimitochondrial antibodies (AMA) are highly sensitive and specific markers that can predict the development of the disease in a healthy individual. Long-term administration of ursodeoxycholic acid (UDCA), a naturally occurring bile acid, safely slows the progression of PBC, delays the need for liver transplantation, and postpones death. An effort should be made to identify the patients with PBC in the asymptomatic stage by the presence of AMA and to conduct a clinical trial in order to assess the benefit of long-term administration of UDCA on the prevention of the overt disease in these individuals.     

Aspirin-Interleukin-3 Interrelationships in Patients With Anti-Phospholipid Syndrome

PROBLEM: Previously we reported on the generation of experimental anti-phospholipid syndrome (APS) in mice. These models were employed to evaluate the efficacy of various novel therapeutic modalities including interleukin-3 (IL-3) and low dose aspirin. The efficacy of the latter was found to be interrelated. Low dose aspirin is capable of inhibiting the activity of the enzyme cyclooxygenase which is responsible for the metabolism of arachidonic acid towards the production of prostaglandins. This shifts the metabolism of arachidonic acid to the other pathway and leads to an overproduction of leukotrienes. The leukotrienes act as stimulators of IL-3 production, a positive cytokine in pregnancy which enhances placental and fetal development. In the current study we evaluated the IL-3 levels in pregnant women with APS and expanded our knowledge on the interrelationships between aspirin, arachidonic acid metabolites and IL-3 in the human system. METHODS: IL-3 levels were recorded in the serum of pregnant women with APS and compared to a control pregnant group. In addition peripheral blood mononuclear cells from healthy subjects were incubated with different concentrations of aspirin or with arachidonic acid metabolites (Leukotriene B4, C4 or PGE₂), and IL-3 production in the culture fluids was evaluated. RESULTS: Serum level of IL-3 in pregnant patients with primary APS, APS secondary to SLE and SLE was lower in comparison to the control group. The in vitro studies revealed that only low dose aspirin (10 mg/μl) stimulated IL-3 production while higher doses of the drug failed to induce the cytokine generation. Leukotriene B4 and C4 were stimulatory whereas PGE₂ acted as inhibitor of IL-3 production. CONCLUSIONS: The serum level of IL-3 is decreased to pregnant women with primary or secondary APS. Low dose aspirin is capable of stimulating EL-3 production in vitro most probably through an elevation of leukotriene production, which may explain its beneficial activity in preventing the manifestations of APS.     

Spinal cord injury in the setting of traumatic thoracolumbar fracture is not reliably associated with increased risk of associated intra-abdominal injury following blunt trauma: An analysis of a National Trauma Registry database

Purpose: There is a common opinion that spinal fractures usually reflect the substantial impact of injuries and therefore may be used as a marker of significant associated injuries, specifically for intraabdominal injury (IAI). The impact of concomitant spinal cord injury (SCI) with the risk of associated IAI has not been well clarified. The aim of this study was to evaluate the incidence and severity of IAIs in patients suffering from spinal fractures with or without SCI. Methods: A retrospective cohort study using the Israeli National Trauma Registry was conducted. Patients with thoracic, lumbar and thoracolumbar fractures resulting from blunt mechanisms of injury from January 1, 1997 to December 31, 2018 were examined, comparing the incidence, severity and mortality of IAIs in patients with or without SCI. The collected variables included age, gender, mechanism of injury, incidence and severity of the concomitant IAIs and pelvic fractures, abbreviated injury scale, injury severity score, and mortality. Statistical analysis was performed using GraphPad InStat ® Version 3.10, with Chi-square test for independence and two sided Fisher’s exact probability test. Results: Review of the Israeli National Trauma Database revealed a total of 16,878 patients with spinal fractures. Combined thoracic and lumbar fractures were observed in 1272 patients (7.5%), isolated thoracic fractures in 4967 patients (29.4%) and isolated lumbar fractures in 10,639 patients (63.0%). The incidence of concomitant SCI was found in 4.95% (63/1272), 7.65% (380/4967) and 2.50% (266/10639) of these patients, respectively. The overall mortality was 2.5%, proving higher among isolated thoracic fracture patient than among isolated lumbar fracture counterparts (11.3% vs. 4.6%, p < 0.001). Isolated thoracic fractures with SCI were significantly more likely to die than non-SCI counterparts (8.2% vs. 3.1%, p < 0.001). There were no differences in the incidence of IAIs between patients with or without SCI following thoracolumbar fractures overall or in isolated thoracic fractures; although isolated lumbar fractures patients with SCI were more likely to have renal (3.4% vs. 1.6%, p = 0.02) or bowel injuries (2.3% vs. 1.0%, p = 0.04) than the non-SCI counterparts. Conclusion: SCI in the setting of thoracolumbar fracture does not appear to be a marker for associated IAI. However, in a subset of isolated lumbar fractures, SCI patient is associated with increased risks for renal and bowel injury.     

Habituation and transfer during sleep in cats.

In order to test the concept of "sleep learning", cats were exposed to habituation treatment of auditory stimuli during different stages of the sleep-waking cycle. While it was possible to demonstrate that habituation to an auditory stimulus can take place in paradoxical sleep (PS) and slow-wave sleep (SWS) as well as during periods of wakefulness, it was not always possible to demonstrate a transfer of habituation from the training period to other periods. A complete transfer of habituation occurred between the two sleep periods (PS and SWS); a partial transfer of habituation occurred between the waking period and both of the two sleep stages (PS and SWS), but only a minimal transfer of habituation was found between any one of the sleep stages and the waking period. When the cats were able to transfer the habituation, they also were able to discriminate between the habituating tone and a novel tone. The findings that only minimal transfer of habituation could occur between each of the sleep stages and the waking period do not lend support to the concept of "sleep learning".     

Delta sleep response to metyrapone in post-traumatic stress disorder.

Metyrapone blocks cortisol synthesis, which results in the stimulation of hypothalamic cortiocotropin-releasing factor (CRF) and a reduction in delta sleep. We examined the effect of metyrapone administration on endocrine and sleep measures in male subjects with and without chronic PTSD. We hypothesized that metyrapone would result in a decrease in delta sleep and that the magnitude of this decrease would be correlated with the endocrine response. Finally, we utilized the delta sleep response to metyrapone as an indirect measure of hypothalamic CRF activity and hypothesized that PTSD subjects would have decreased delta sleep at baseline and a greater decrease in delta sleep induced by metyrapone. Three nights of polysomnography were obtained in 24 male subjects with combat-related PTSD and 18 male combat-exposed normal controls. On day 3, metyrapone was administered during normal waking hours until habitual sleep onset preceding night 3. Endocrine responses to metyrapone were measured in plasma obtained the morning following sleep recordings, the day before and after administration. Repeated measures ANOVAs were conducted to compare the endocrine and sleep response to metyrapone in PTSD and controls. PTSD subjects had significantly less delta sleep as indexed by stages 3 and 4, and total delta integrated amplitude prior to metyrapone administration. There were no differences in premetyrapone cortisol or ACTH levels in PTSD vs controls. PTSD subjects had a significantly decreased ACTH response to metyrapone compared to controls. Metyrapone caused an increase in awakenings and a marked decrease in quantitative measures of delta sleep that was significantly greater in controls compared to PTSD. The decline in delta sleep was significantly associated with the magnitude of increase in both 11-deoxycortisol and ACTH. The results suggest that the delta sleep response to metyrapone is a measure of the brain response to increases in hypothalamic CRF. These data also suggest that the ACTH and sleep EEG response to hypothalamic CRF is decreased in PTSD.     

Impact of severity of illness bias and control group misclassification bias in case-control studies of antimicrobial-resistant organisms.

BACKGROUND: Case-control studies often analyze risk factors for antibiotic resistance. Recently published articles have illustrated that randomly selected control-patients may be preferable to those with the susceptible phenotype of the organism. A possible methodologic problem with randomly selected control-patients is potential bias due to control group misclassification. This occurs if some control-patients did not have clinical cultures performed and thus might have been unidentified case-patients. If this bias exists, these studies might be expected to report lower odds ratios (ORs) because control-patients would be more like case-patients. OBJECTIVE: To analyze potential biases that might arise due to control group misclassification and potentially larger selection biases that may be introduced if control-patients are required to have at least one clinical culture. PATIENTS: One hundred twenty case-patients, 770 control-patients in group 1, and 510 control-patients in group 2. METHODS: Two case-control studies. Case-patients had clinical cultures positive for imipenem-resistant Pseudomonas aeruginosa. The first group of control-patients were random. The second group of control-patients were identical to those in group 1 except being required to have at least one clinical culture. RESULTS: Univariate analyses showed higher ORs for case-patients versus control-patients in group 1 (imipenem [OR, 12.5], piperacillin-tazobactam [OR, 3.7], and vancomycin [OR, 4.7]) as compared with case-patients versus control-patients in group 2 (imipenem [OR, 8.0], piperacillin-tazobactam [OR, 2.5], and vancomycin [OR, 3.0]). CONCLUSION: Requiring control-patients to have at least one clinical culture introduces a selection bias likely because it eliminates patients with less severe illness.