Bystander CD8 T-cell-mediated demyelination is interferon-gamma-dependent in a coronavirus model of multiple sclerosis

Mice infected with the coronavirus mouse hepatitis virus, strain JHM (JHM) develop a disease that shares many histological characteristics with multiple sclerosis. We previously demonstrated that JHM-infected mice that only have CD8 T cells specific for an epitope not in the virus develop demyelination on specific activation of these cells. Herein we show that this process of bystander T-cell-mediated demyelination is interferon-gamma (IFN-gamma)-dependent. The absence of IFN-gamma abrogated demyelination but did not change T-cell infiltration or expression levels of inflammatory cytokines or chemokines in the spinal cord. These results are consistent with models in which IFN-gamma contributes to CD8 T-cell-mediated demyelination by activation of macrophages/microglia, the final effector cells in the disease process.     

Phase I clinical study of LL-D49194α1 with retrospective pharmacokinetic investigations in mice and humans

Summary LL-D491941 is a new cytotoxic antibiotic selected for clinical phase I study because of its impressive pre-clinical anti-tumour activity and its low toxicity profile in experimental animals. A total of 15 patients were treated in centres in Glasgow and Amsterdam at doses ranging from 0.25 to 4 mg/m². One minor response was noted in a patient with colonic carcinoma. The study was suspended following the discovery of unexpected cardiotoxicity. As this toxicity was not consistent with the standard (EORTC) European Organisation for Research and Treatment of Cancer toxicology profile, we chose to investigate the pharmacokinetics of LL-D491941 in mice and humans in more detail to try to explain this phenomenon. A major difference in plasma protein binding was discovered between mice and patients, with a suggestion of non-linear kinetics being noted at higher doses in humans. It is likely that these differences in drug handling account for the unexpected and serious toxicity encountered in this trial.     

Modulation of immunoglobulin production and cytokine mRNA expression in peripheral blood mononuclear cells by intravenous immunoglobulin

Intravenous immunoglobulin (IVIG) has the potential to regulate Ig production, but the mechanism(s) responsible for this effect is unknown. In experiments reported here, we examined the ability of IVIG to regulate Ig production in human peripheral blood mononuclear cells (PBMCs) stimulated with pokeweed mitogen (PWM). IVIG (2–10 mg/ml) showed a potent (80–85%) inhibition of PWM-stimulated IgG, IgM, and IgA production. To determine more precisely how IVIG mediated the inhibition of Ig production, we studied Ig promoting cytokine gene expression after PWM stimulation with or without IVIG (2 and 10 mg/ml) using dot-blot techniques. RNA was isolated from PBMCs at predetermined time points and probed with cDNAs specific for human cytokines (IL-1-, IL-2, IL-2R, IL-4, IL-5, IL-6, -IFN, and TNF-). IL-6 mRNA accumulation was maximal at 4.5 hr post-PWM stimulation and was inhibited 64–75% when IVIG (10 mg/ml) was present. -IFN mRNA levels peaked at 72 hr poststimulation and were also 68–75% inhibited by IVIG. IL-2 mRNA levels peaked at 4.5 hr and were 23–46% inhibited by IVIG. The inhibitory effect of IVIG on production of these cytokines (IL-6 and -IFN) was also observed at the protein level in sonicated PBMCs after incubation with PWM and IVIG. The mRNA levels for other cytokines were not or only minimally inhibited by IVIG. Addition of IL-6, -IFN, or IL-2 partially restored Ig production in IVIG-treated PWM-stimulated cultures, suggesting that inhibition of other cytokines or another mechanism(s) independent of cytokine inhibition might also be involved, although inhibition of IL-6, -IFN, and IL-2 may be one of the critical factors in the suppression of Ig production by IVIG.     

Decreased interstitial glucose and transmural gradient in lactate during ischemia

Summary The purpose of this investigation was to assess the effects of ischemia and reperfusion on the transmural levels of glucose and lactate in the interstitium in 11 open-chest swine. Microdialysis probes were used to estimate changes in interstitial metabolities across the ventricular wall. Probes were placed in the subepicardium and the subendocardium of the left anterior descending (LAD) coronary artery perfusion bed and in the midmyocardium of the circumflex (CFX) perfusion bed. The LAD coronary artery was cannulated and perfused with blood from the femoral artery through an extracorporal perfusion circuit. Ischemia was induced in the LAD perfusion bed by reducing the flow of the LAD perfusion pump by 60% for 50 min, and was followed by 30 min of reperfusion. Regional myocardial blood flow was assessed with fluorescent microspheres. Ischemia resulted in a transmural gradient in blood flow, with the most severe reduction in flow occurring in the subendocardium (p<0.05). We found a significant reduction in interstitial glucose in both the LAD subepicardium (1.26±0.24 mM) (p=0.0009) and subendocardium (0.89±0.21 mM) (p=0.0001) during ischemia compared to the aerobic (non-ischemic) period (1.97±0.25 mM, 2.03±0.29 mM for the subepicardium and subendocardium, respectively). This coincided with a significant reduction in glucose delivery (LAD pump flow^* arterial glucose) to the LAD perfusion bed during ischemia (54.5±8.5 mol/min) compared to aerobic values (182.1±25.3 mol/min) (p<0.05). Interstitial lactate levels were significantly increased during ischemia in the LAD subendocardium (3.39±0.46 mM) compared to the aerobic values (1.73±0.46 mM) (p<0.0029). A transmural gradient in interstitial lactate levels was observed during ischemia: this gradient was not seen during the aerobic period and was negated upon reperfusion. In conclusion, ischemia resulted in a decrease in interstitial glucose in both the LAD subepicardium and subendocardium, and an increase in interstitial lactate in the LAD subendocardium. Further, a transmural gradient in interstitial lactate levels was observed during ischemia, with the highest lactate values appearing in the subendocardium.     

Induction of gene expression of amyloid precursor protein (APP) in activated human lymphoblastoid cells and lymphocytes

To understand the possible role of amyloid precursor protein (APP) in human lymphocytes, and the regulation of APP gene expression in this cell type, we determined levels of cellular APP protein and of mRNA in human T-cell-derived Jurkat cells that were treated with lectin, phorbol ester, and calcium ionophore. We also related these levels to cell aggregation and adhesion. Cell-cell aggregation and cell-plastic adhesion were observed over a 24-h period after incubating cells for 2 h with phytohemagglutinin or phorbol myristate acetate. Cells treated with a calcium ionophore showed no aggregation or adhesion. Western blots indicated no obvious alteration in the level of cellular APP with different treatments. Northern blots showed a significant transient increase of APP mRNA after incubation with the calcium ionophore, whereas phorbol ester treatment showed a slight increase of APP mRNA. We analyzed the level of APP mRNA in human peripheral T cells which had been separated from peripheral lymphocytes. The level increased transiently by up to threefold after treatment with calcium ionophore plus phorbol esters. These data suggest that cell-cell aggregation and cell-matrix adhesion by human lymphocytes are not associated with an increased level of cellular APP protein or of mRNA.     

Treatment of systemic and renal-limited vasculitic disorders with pooled human intravenous immune globulin

Idiopathic crescentic glomerulonephritis is characterized by an absence of immunohistological evidence of immune deposits, often with evidence of segmental glomerular necrosis. Such pauciimmune crescentic glomerulonephritis is the most common renal manifestation seen in patients with Wegener's granulomatosis, polyarteritis nodosa, and glomerulonephritis associated with other systemic vasculitic disorders (i.e., Churg-Strauss syndrome). Recently, the idiopathic crescentic glomerulonephritides, either in renal-limited form or in association with other systemic vasculitic disorders, were found to have in common a serologic marker, antineutrophil cytoplasmic autoantibodies. These cytoplasmic and perinuclear antineutrophil cytoplasmic autoantibodies are specific for constituents of neutrophil primary granules and monocyte lysosomes. As serologic markers for vasculitic disorders, they are also felt to be directly involved in the pathogenesis of necrotizing vascular injury.In vitro, both perinuclear and cytoplasmic antineutrophil cytoplasmic autoantibodies are capable of causing cytokineprimed neutrophils to undergo degranulation and respiratory burst, releasing toxic oxygen species and lytic enzymes. Antiidiotype antibodies which inhibit antineutrophil cytoplasmic autoantibodiesin vitro, in a V region-dependent manner, are found in pooled humanγ-globulin preparations. Intravenous immune globulin infusionsin vivo have produced dramatic improvements in the necrotizing vascular injury produced by antineutrophil cytoplasmic autoantibodies, and a rapid reduction in these autoantibody levels is seen post-intravenous immune globulin infusion in most patients. The proposed mechanisms of action of intravenous immune globulin in vasculitic disorders include Fc-dependent mechanisms, and F(ab′)₂-dependent mechanisms are likely important. Intravenous immune globulin infusions appear to have an important place in the management of the necrotizing vascular injury. Blinded, randomized, placebo-controlled trials will be necessary to establish definitively intravenous immune globulin as a therapeutic option in vasculitic disorders.     

High-voltage-activated calcium channel messenger RNA expression in the 140-3 neuroblastoma-glioma cell line

Expression of calcium channel alpha1 subunits in oocytes or cell lines has proven to be a powerful method in the analysis of structure-function relations, but these experimental systems are of limited value in the examination of neuron-specific functions such as transmitter release. Cell lines derived from neurons are often capable of these functions, but their intrinsic calcium channel alpha1 subunits are complicating factors in experimental design. We have examined the biophysical and molecular properties of calcium channels in a little studied neuroblastoma-glioma hybrid cell line, 140-3, a close relative of the NG108-15 cell line, to test whether this cell line might serve a role as an expression system for neural mechanisms. This cell was selected as it contains an intact transmitter release mechanism yet secretes little in response to depolarization. Patch-clamp recording revealed only a prominent low-threshold, rapidly inactivating calcium current with a single-channel conductance of approximately 7 pS that was identified as T type. A search for calcium channel alpha1 subunit messenger RNA in the 140-3 cells with three different tests only revealed alpha1C, whereas alpha1A-alpha1C were present in the parent NG108-15 line. We made a particular effort to search for alpha1E, since this subunit has been associated with a low-voltage-activated current. Our findings suggest that, since the principal nerve terminal-associated calcium channels (alpha1A, alpha1B, alpha1E) are absent in the 140-3 cell, this cell line may prove a particularly useful model for the analysis of the role of high-voltage-activated calcium channels in complex functions of neuronal cells.     

The use of delayed computerized tomography in the evaluation of blunt abdominal trauma: a preliminary report

This purpose of this study was to analyze the use of abdominal computed tomography (CT) imaging in patients with possible blunt abdominal trauma. A retrospective analysis of all trauma patients over a 1-year period (1993-1994) was conducted, with prospective study protocol in 52 patients using serial abdominal exam and hematocrits (Hcts) instead of abdominal CT for evaluation of blunt abdominal trauma. Urgent abdominal CT was used as the initial diagnostic test for evaluation of blunt abdominal trauma in 813 patients over this 1-year period. CT was obtained in 379 (46.6%) of these patients who arrived hemodynamically stable (admission systolic blood pressure > or = 90), had a Glasgow Coma Scale > 13, and had admission Hct > or = 35 because of distracting injuries, possible traumatic brain injury, or alcohol/drug use, which might render the abdominal physical exam unreliable. Only 47 CT scans (12.4%) were positive, and three patients (0.8%) required laparotomy. In an effort to more efficiently use abdominal CT, we performed a prospective study in 52 patients with possible blunt abdominal trauma, admission systolic blood pressure > or = 90, Hct > or = 35, Glasgow Coma Scale > 13, and a normal abdominal exam on admission. These patients were followed with serial abdominal examinations and Hcts every 6 hours for 24 hours, and delayed CT, when applicable. CT was obtained in seven patients (13.5%) for evaluation of fall in Hct or abnormal abdominal examination; all were negative for abdominal injury. A protocol using serial abdominal exams, Hcts, and delayed abdominal CT imaging may be useful in select patients to decrease the high number of negative routine abdominal CTs that are obtained in the evaluation of blunt abdominal trauma.     

Transcutaneous cranial electrical stimulation (TCES): a review 1998

The Transcutaneous Cranial Electrical Stimulation (TCES) technique appeared at the beginning of the 1960s and is aimed to act at the level of the central nervous system. The current, composed of high frequency pulses interrupted with a repetitive low frequency, is delivered through three electrodes (a negative electrode placed between the eyebrows while two positive electrodes are located in the retro-mastoid region). Due to the characteristics of the current delivered, shortcomings encountered with previous electrical stimulation techniques are avoided. The main property of TCES is to potentiate some drug effects, especially opiates and neuroleptics, during anesthetic clinical procedures. This potentiation effect permits drastic reduction of pharmacological anesthetic agent and reduces post-operative complications. Animal studies performed with TCES demonstrated that this stimulation releases 5-hydroxy-indol-acetic acid and enkephalins. Despite numerous clinical and animal studies performed with this technique for several decades, TCES mechanisms are not completely elucidated but results obtained without undesirable effect are encouraging signs to continue investigations of this particular technique.