Practice guidelines — an emerging synthetic science

Dr. Cook is a Career Scientist of the Ontario Ministry of Health     

Cholinergic Modification of Glucose-Induced Biphasic Insulin Release in Vitro

An in vitro system for perifusion of rat pancreatic islets has been utilized to define the effects of cholinergic agents on the dynamics of insulin release. In the absence of glucose the effects of either acetylcholine or acetyl-beta-methylcholine were minimal at concentrations up to 10(-5) mM. In the presence of low glucose concentration (2.4 mM), both of the muscarinic agents produced dose-dependent biphasic insulin release. Under these conditions significant insulin release was observed over both phases at concentrations of the muscarinic agents as low as 10(-8) mM. Further, the dose response curves relating muscarinic concentration to the total amount of insulin released in each of the two phases showed lack of parallelism between the curves. Nicotinic acid in concentrations up to 10(-5) mM had no effect on insulin release in the presence of 2.4 mM glucose. When the glucose concentration was increased to 16.4 mM, the effects of the muscarinic agents were significantly less than those observed in the presence of 2.4 mM glucose. This held true whether the effect was defined as absolute increment due to the muscarinic agent or as percentage of enhancement. Atropine inhibited insulin release induced by both acetylcholine and by 16.4 mM glucose. These data indicate that cholinergic stimulation can play a significant role in modifying insulin release patterns.     

Effects of dopamine receptor blockade on cerebral blood flow response to somatosensory stimulation in the unanesthetized rat

Local cerebral blood flow (CBF) was determined in 30 cerebral structures, including four structures of the whisker-to-barrel cortex sensory pathway, by the quantitative autoradiographic [(14)C]iodoantipyrine method during unilateral vibrissal stimulation in rats administered 0.1 or 1.0 mg/kg haloperidol or its control vehicle intravenously. The low dose of haloperidol had no significant effects on resting CBF or its enhancement by vibrissal stimulation. By standard t tests, the high dose statistically significantly lowered baseline CBF in frontal and visual cortex, hippocampus, dentate gyrus, inferior olive, cerebellar cortex, and the ventral posteromedial (VPM) thalamic nucleus on the unstimulated side, and raised baseline CBF in the lateral habenula; however, these changes lost statistical significance after Bonferroni correction for multiple comparisons. Neither dose had any effects on the increases in CBF evoked by vibrissal stimulation in the principal sensory trigeminal nucleus and barrel cortex, but the higher dose statistically significantly enhanced the percent increases in CBF due to the sensory stimulation in the spinal trigeminal nucleus and VPM thalamic nucleus. These results do not support a role for direct dopaminergic vasoactive mechanisms in the increases in CBF associated with neuronal functional activation.     

Effects of antifungal interventions on the outcome of experimental infections with phenotypic switch variants of Cryptococcus neoformans

In cryptococcal infection, phenotypic switching from a smooth to a mucoid variant can occur in vivo, producing variants with enhanced virulence that are subsequently selected and affect the outcome of infection. Here, we demonstrate that antifungal treatment of the chronically infected host can promote this phenomenon. Amphotericin B treatment reduces fungal burden less effectively in mucoid variant-infected than in smooth variant-infected mice. Consequently, amphotericin B treatment resulted in a more pronounced prolongation of survival in smooth variant-infected than in mucoid variant-infected mice (20 versus 42 days; P < 0.05). Administration of anticapsular monoclonal antibody mediated better protection in smooth variant-infected than in mucoid variant-infected mice, although a protective effect was not consistently observed at all doses. Most interestingly, both antifungal drug therapy and administration of anticapsular monoclonal antibody promoted the selection of mucoid variants in smooth variant-infected mice, a phenomenon manifested by a statistically higher percentage of mucoid colonies in smooth variant-infected mice than in nontreated control mice. This finding suggests that both chemotherapeutic and immunological antifungal interventions may promote the selection of the more virulent mucoid variant, which could affect the outcome of infection in chronically infected hosts.     

Generation of Splenic Follicular Structure and B Cell Movement in Tumor Necrosis Factor–deficient Mice

Secondary lymphoid tissue organogenesis requires tumor necrosis factor (TNF) and lymphotoxin α (LTα). The role of TNF in B cell positioning and formation of follicular structure was studied by comparing the location of newly produced naive recirculating and antigen-stimulated B cells in TNF^−/− and TNF/LTα^−/− mice. By creating radiation bone marrow chimeras from wild-type and TNF^−/− mice, formation of normal splenic B cell follicles was shown to depend on TNF production by radiation-sensitive cells of hemopoietic origin. Reciprocal adoptive transfers of mature B cells between wild-type and knockout mice indicated that normal follicular tropism of recirculating naive B cells occurs independently of TNF derived from the recipient spleen. Moreover, soluble TNF receptor–IgG fusion protein administered in vivo failed to prevent B cell localization to the follicle or the germinal center reaction. Normal T zone tropism was observed when antigen-stimulated B cells were transferred into TNF^−/− recipients, but not into TNF/LTα^−/− recipients. This result appeared to account for the defect in isotype switching observed in intact TNF/LTα^−/− mice because TNF/LTα^−/− B cells, when stimulated in vitro, switched isotypes normally. Thus, TNF is necessary for creating the permissive environment for B cell movement and function, but is not itself responsible for these processes.     

Benzalkonium interference with test methods for potassium and sodium

Six automated instruments that measure sodium and potassium were tested for interference from two compounds used in catheters. Tridodecylmethylammonium heparin did not interfere with any of the methods. However, benzalkonium heparin falsely increased sodium measurement with the Kodak Ektachem, and falsely increased potassium measurements with three instruments (Beckman Astra, Baxter Paramax, and the Instrumentation Laboratory Monarch) in which ion-selective electrodes measure potassium in diluted serum. Three instruments in which ion-selective electrodes measure serum directly--Du Pont Dimension, Abbott Spectrum, and Kodak Ektachem--experienced no interference with potassium measurements. Interference of benzalkonium with potassium measurements may result from its interaction with the electrode membranes, which is accentuated in diluted serum.     

New strategies for clinical trials in patients with sepsis and septic shock

OBJECTIVE: The difficulty in identifying new treatment modalities that significantly reduce the mortality and morbidity rates associated with sepsis has highlighted the need to reevaluate the approach to clinical trial design. The United Kingdom Medical Research Council convened an International Working Party to address these issues. DATA SOURCES: The subject areas that were to be the focus of discussion were identified by the co-chairs, and group leaders were nominated. Preconference reading material was circulated to group members. STUDY SELECTION AND DATA EXTRACTION: Small-group discussion fed into an iterative process of feedback from plenary sessions, followed by the formulation of recommendations. Finally, each working group prepared a summary of its recommendations and these are reported herein. DATA SYNTHESIS: There were five key recommendations. First, investigators should no longer rely solely on the American College of Chest Physicians/Society of Critical Care Medicine definitions of sepsis or sepsis syndrome as the basis of trial entry. Entry criteria should be based on three principles: a) All patients should have infection; b) there should be evidence of a pathologic process that represents a biologically plausible target for the proposed intervention, for example, an abnormal circulating level of a biological marker pertinent to the study drug; and c) patients should fall into an appropriate category of severity (usually severe sepsis). Second, investigators should use a scoring system for organ dysfunctions that has been validated and that can be incorporated into all sepsis studies; agreement on the use of a single system would simplify comparisons between studies. Third, the primary outcome measure generally should be mortality rates, but under appropriate circumstances major morbidities could be considered as primary end points. Regardless of choice of the duration to primary end point, patients should be followed for > or =90 days. Fourth, sample size needs to be based on a realistic assessment of achievable effect size based on knowledge of the at-risk population. Fifth, subgroups should be few in number and should be defined a priori on the basis of variables present before randomization. CONCLUSIONS: Important changes in several aspects of trial design may improve the quality of clinical studies in sepsis and maximize the chance of identifying effective therapeutic agents.