Mucosal vaccination delays or prevents prion infection via an oral route.

In recent years major outbreaks of prion disease linked to oral exposure of the prion agent have occurred in animal and human populations. These disorders are associated with a conformational change of a normal protein, PrP(C) (prion protein cellular), to a toxic and infectious form, PrP(Sc) (prion protein scrapie). None of the prionoses currently have an effective treatment. A limited number of active immunization approaches have been shown to slightly prolong the incubation period of prion infection. Active immunization in wild-type animals is hampered by auto-tolerance to PrP and potential toxicity. Here we report that mucosal vaccination with an attenuated Salmonella vaccine strain expressing the mouse PrP, is effective at overcoming tolerance to PrP and leads to a significant delay or prevention of prion disease in mice later exposed orally to the 139A scrapie strain. This mucosal vaccine induced gut anti-PrP immunoglobulin (Ig)A and systemic anti-PrP IgG. No toxicity was evident with this vaccination approach. This promising finding suggests that mucosal vaccination may be a useful method for overcoming tolerance to PrP and preventing prion infection among animal and potentially human populations at risk.

The prognostic significance of microalbuminuria in non-diabetic acute stroke patients.

BACKGROUND: Microalbuminuria (MA) is thought to be a marker of widespread vascular damage. It is associated with increased mortality in diabetes mellitus, hypertension and acute myocardial infarction. The aim of the present study was to evaluate the prognostic significance of MA in non-diabetic acute stroke patients. MATERIAL AND METHODS: We studied 52 patients (mean age 69.3 +/- 12.5 years) diagnosed with ischemic stroke confirmed by computed tomography, who were admitted to the Stroke Unit within 24 hours after the onset of symptoms. The control group consisted of 37 age- and gender-matched subjects (mean age 65.2 +/- 5.7 years), examined 3 to 18 months after ischemic stroke. We excluded patients with diabetes mellitus, positive urinalysis, proteinuria, hepatic or renal insufficiency, neoplastic disease or clinical signs of infection. The severity of the neurological deficit was assessed by the Scandinavian Stroke Scale (SSS). The albumin excretion rate was measured in daily urine collection on the second day of hospitalization, using the immunonephelometric method. The patients were followed up for three months. RESULTS: MA was found in 24 of 52 (46.1%) acute stroke patients and in 5 of 37 (13.5%) controls (p<0.05). Patients with MA scored lower on the SSS than patients without MA, both on admission and later. We found a correlation between the daily excretion of albumin and the severity of neurological deficit on admission, as expressed by the SSS score (r = -0.48, p<0.05). The 90-day mortality rate was higher in patients with MA as compared to patients without MA (45.8% vs 7.1%). Patients with MA scored lower on the Barthel Index on Day 90 (median: 65 vs 100, p<0.01). CONCLUSIONS: We found that MA can be detected in about 46% of non-diabetic patients with acute ischemic stroke. Measuring the albumin excretion rate may be a reliable predictor of increased mortality 3 months after stroke.

[The role of statins in prevention of ischemic stroke]. / Rola statyn w prewencji udaru niedokrwiennego mózgu.

Although associations between cholesterol and coronary heart disease (CHD) are well accepted, the association between cholesterol and stroke remains unclear. Epidemiological studies suggest lack of apparent correlation between cholesterol and cerebrovascular events, however meta-analyses of secondary prevention trials tested statins (HMG-CoA reductase inhibitors) efficacy in reducing cholesterol revealed a powerful statistically significant effect to reduce stroke as well as CHD (32%). Mechanism for stroke reduction can be connected with nonlipid mechanism of statins action: modifying endothelial function and inflammatory responses, plaque stabilisation and inhibition of plaque progression and thrombus formation in the intracranial and extracranial carotid arteries. Stroke events may be also reduced partially as a consequence of CHD reduction.

Cue-induced cocaine craving: neuroanatomical specificity for drug users and drug stimuli.

OBJECTIVE: Cocaine-related cues have been hypothesized to perpetuate drug abuse by inducing a craving response that prompts drug-seeking behavior. However, the mechanisms, underlying neuroanatomy, and specificity of this neuroanatomy are not yet fully understood. METHOD: To address these issues, experienced cocaine users (N=17) and comparison subjects (N=14) underwent functional magnetic resonance imaging while viewing three separate films that portrayed 1 ) individuals smoking crack cocaine, 2) outdoor nature scenes, and 3) explicit sexual content. Candidate craving sites were identified as those that showed significant activation in the cocaine users when viewing the cocaine film. These sites were then required to show significantly greater activation when contrasted with comparison subjects viewing the cocaine film (population specificity) and cocaine users viewing the nature film (content specificity). RESULTS: Brain regions that satisfied these criteria were largely left lateralized and included the frontal lobe (medial and middle frontal gyri, bilateral inferior frontal gyrus), parietal lobe (bilateral inferior parietal lobule), insula, and limbic lobe (anterior and posterior cingulate gyrus). Of the 13 regions identified as putative craving sites, just three (anterior cingulate, right inferior parietal lobule, and the caudate/lateral dorsal nucleus) showed significantly greater activation during the cocaine film than during the sex film in the cocaine users, which suggests that cocaine cues activated similar neuroanatomical substrates as naturally evocative stimuli in the cocaine users. Finally, contrary to the effects of the cocaine film, cocaine users showed a smaller response than the comparison subjects to the sex film. CONCLUSIONS: These data suggest that cocaine craving is not associated with a dedicated and unique neuroanatomical circuitry; instead, unique to the cocaine user is the ability of learned, drug-related cues to produce brain activation comparable to that seen with nondrug evocative stimuli in healthy comparison subjects.

Hepatitis C virus infection in cocaine users--a silent epidemic.

Over a recent three year period, approximately 600 individuals responded to newspaper advertisements for research studies requiring healthy, cocaine using subjects. These subjects were screened using a standard phone interview in order to eliminate individuals with known medical or psychiatric illnesses that would exclude them from ongoing neuroimaging studies of drug abuse. Individuals were specifically asked about their hepatitis and HIV status. Of these, 170 subjects passed the phone screen, having no known medical or psychiatric illness outside of cocaine abuse/dependence and were willing to be further evaluated for the studies. These subjects were brought to the Medical College of Wisconsin's General Clinical Research Center and tested for, among other measures, hepatitis B, hepatitis C, and HIV. Of these, 144 completed the examination and all testing. In this cohort of assumed healthy subjects, 47 (33%) tested positive for antibodies to the hepatitis C virus (HCV). Only 7 (5%) tested positive for the hepatitis B surface antigen and 2 (1.4%) to HIV. The demographics of this cohort are 56% African-American, 81% male, 75% never-married, 55% unemployed with a mean age of 36 years. The percentage of subjects reporting any lifetime intravenous drug use among the HCV(+) and the HCV(-) cohorts was 77% vs. 29% respectively. Some routes of HCV transmission are still unclear and may reflect lifestyle or other factors related to cocaine use outside of parenteral drug use. Since almost all HCV infections become chronic, and many progress to chronic active hepatitis, cirrhosis, and ultimately hepatocellular carcinoma, these observations suggest a significant epidemic in an unsuspecting population with little regular access to health care. These individuals also form a large pool for the continued transmission of HCV to the general population. Additional public health interventions are suggested.

Neurochemical adaptation to cocaine abuse: reduction of N-acetyl aspartate in thalamus of human cocaine abusers.

BACKGROUND: The aim of this study was to determine the existence of, and possible mechanisms for, chronic cocaine use-induced neurotoxicity in the human brain. Because in vivo magnetic resonance spectroscopy (MRS) provides a noninvasive way to detect biochemical and physiological changes in the brain, we sought to specifically determine the neurochemical adaptations in chronic cocaine-dependent subjects. METHODS: Twenty-one cocaine users and 13 non-drug-using, age-matched normal volunteers were recruited for an in vivo proton MRS study. Following screening that included physical examination, histories, and blood testing, cocaine group subjects received a spectral scan on a 1.5-T GE Signa scanner. Spectra were obtained from the left basal ganglia and/or the left thalamus from subjects in both groups using an rf bird-cage type head coil with single-voxel localization. RESULTS: The level of N-acetyl aspartate in the region of left thalamus was lower (17%) in the chronic cocaine user group but not in the region of left basal ganglia, compared with the control group. CONCLUSIONS: These results suggest that chronic cocaine use may induce abnormal neurochemical activity and a state of neuronal dysregulation and/or neurotoxicity. It will now be important to determine if these alterations are reversible during withdrawal and what the functional implications of this observation are with respect to cognitive function and drug relapse.

Nitric oxide mediates the mitogenic effects of insulin and vascular endothelial growth factor but not of leptin in endothelial cells.

The regulation of vascular wall homeostasis by nitric oxide (NO) generated by endothelium is being intensively studied. In the present paper, the involvement of NO in the vascular endothelial growth factor (VEGF), insulin or leptin-stimulated proliferation of human endothelial cells (HUVEC) was measured by [3H]thymidine or bromodeoxyuridine incorporation. VEGF and insulin, but not leptin, increased NO generation in HUVEC, as detected with ISO-NO electrode. Proliferation of HUVEC induced by leptin was not changed or was higher in the presence of N(omega)-nitro-L-arginine methyl ester (L-NAME) a nitric oxide synthase (NOS) inhibitor. In contrast, L-NAME blunted the proproliferative effect of VEGF and insulin. Furthermore, we demonstrated that, in human arterial smooth muscle cells (hASMC) transfected with endothelial NOS (eNOS) gene, the generation of biologically active VEGF protein was NO-dependent. Inhibition of NO generation by L-NAME decreased the synthesis of VEGF protein and attenuated HUVEC proliferation induced by conditioned media from transfected hASMC. Endothelium-derived NO seems to participate in VEGF and insulin, but not leptin, mitogenic activity. Additionally, the small amounts of NO released from endothelial cells, as mimicked by eNOS transfection into hASMC, may activate generation of VEGF in sub-endothelial smooth muscle cells, leading to increased synthesis of VEGF protein necessary for turnover and restitution of endothelial cells.

Determination of drug-induced changes in functional MRI signal using a pharmacokinetic model.

As the applications of functional magnetic resonance imaging (fMRI) expand, there is a need for the development of new strategies for data extraction and analysis that do not require the presentation of stimuli in a repeated on/off pattern. A description and evaluation of a method and computer algorithm for the detection and analysis of brain activation patterns following acute drug administration using fMRI are presented. A waveform analysis protocol (WAP) input function has been developed that is based upon the single-dose pharmacokinetics of a drug of interest. As a result of this analysis, regional brain activation can be characterized by its localization and intensity of activation, onset of action, time to peak effect, and duration of action. A global statistical test for significant drug effects based upon the probability of a voxel being activated by a saline vehicle injection is applied to grouped data on a voxel by voxel basis. Representative data are presented using nicotine as a prototypical agent. Using this method, statistically significant drug-induced brain activation has been identified in several key cortical and subcortical brain regions.

Nicotine-induced limbic cortical activation in the human brain: a functional MRI study.

OBJECTIVE: Nicotine is a highly addictive substance, and cigarette smoking is a major cause of premature death among humans. Little is known about the neuropharmacology and sites of action of nicotine in the human brain. Such knowledge might help in the development of new behavioral and pharmacological therapies to aid in treating nicotine dependence and to improve smoking cessation success rates. METHOD: Functional magnetic resonance imaging, a real-time imaging technique, was used to determine the acute CNS effects of intravenous nicotine in 16 active cigarette smokers. An injection of saline followed by injections of three doses of nicotine (0.75, 1.50, and 2.25 mg/70 kg of weight) were each administered intravenously over 1-minute periods in an ascending, cumulative-dosing paradigm while whole brain gradient-echo, echo-planar images were acquired every 6 seconds during consecutive 20-minute trials. RESULTS: Nicotine induced a dose-dependent increase in several behavioral parameters, including feelings of "rush" and "high" and drug liking. Nicotine also induced a dose-dependent increase in neuronal activity in a distributed system of brain regions, including the nucleus accumbens, amygdala, cingulate, and frontal lobes. Activation in these structures is consistent with nicotine's behavior-arousing and behavior-reinforcing properties in humans. CONCLUSIONS: The identified brain regions have been previously shown to participate in the reinforcing, mood-elevating, and cognitive properties of other abused drugs such as cocaine, amphetamine, and opiates, suggesting that nicotine acts similarly in the human brain to produce its reinforcing and dependence properties.

Determination of nitrite/nitrate in human biological material by the simple Griess reaction.

Since a number of pathological processes such as septic shock, inflammation, graft rejection, diabetes, etc. are associated with a release of nitric oxide (NO), rapid and accurate methods of monitoring of NO concentration are of interest. Various methods for measurement of nitrite and nitrate (NO2-, NO3- ) -- the stable metabolites of NO -- are commonly used for this purpose. In this paper we have shown that the proper Griess procedure for nitrite determination significantly increases the sensitivity of this method. This procedure, supplemented with deproteinization and reduction of nitrates to nitrites in the presence of NADPH-sensitive reductase, can be successfully applied for measurement of NOx levels in human body fluids (serum, urine and CSF). Deproteinization of samples with methanol/diethylether is required and does not influence the sensitivity of detection of NO metabolites. The recovery of the method is 88%+/-6% (n = 30). The NOx concentrations measured by this procedure ranged from 25.0 to 39.0 micromol/l in blood, 4.6 to 14.6 micromol/l in CSF and 0.37 to 2.52 mmol/l (adjusted to creatinine concentration) in urine. The coefficient of variation for this method was between 1.3-2.2%. This method can also be recommended for measurement of NOx produced by cells in tissue cell culture.

Apo E isoforms, insulin output and plasma lipid levels in essential hypertension.

BACKGROUND: The association between apo E isoforms and insulin output during the oral glucose test (OGTT) in 60 non-diabetic, non-obese patients with essential hypertension and in control subjects (non-obese, non-diabetic normotensive subjects) was estimated. METHODS: According to low or high insulin output during OGTT, the subjects were divided into the following groups: normotensive subjects with low (NLI) and high (NHI) and hypertensive subjects with low (HLI) and high (HHI) insulin output. RESULTS: The apo E 4/2 phenotype was detected in 32% of hypertensive subjects but not in control subjects. The frequency of apo E 3/2 phenotype in hypertensive subjects was 5% and in normotensive subjects 15%. An increased frequency of phenotype apo E 4/3 was noticed both in HHI (46%) and in NHI (50%) compared with HLI (22%) and NLI (17%) groups. CONCLUSION: The results suggest that the determination of phenotypes apo E and insulin output may contribute to an early detection of individuals at high risk of hypertension development.

An inhibitory effect of camonagrel-a new thromboxane synthase inhibitor, on P-selectin-mediated platelet/PMN adhesion.

P-selectin (PADGEM protein, GMP-140 or CD 62) is a glycoprotein of platelet a-granules and endothelial Weibel-Palade bodies that, by mediating cellular adhesion, initiates recruitment of leukocytes and lymphocytes into injured tissue. Both of the endothelial antiplatelet autacoids prostacyclin (PGI(2)) and nitric oxide (NO) have been demonstrated to inhibit P-selectin expression. Prostaglandin endoperoxides PGG(2)/PGH(2) that are generated by activated platelets have been demonstrated to be used by endothelium for generation of prostacyclin. In an experimental model in vitro that resembles vessel wall/platelet/PMN interaction in vivo, we found that aspirin (100 µM), a COX inhibitor, but not L-NMMA (100 µM) and a NO-synthase inhibitor, reversed the inhibitory effect of arterial wall on P-selectin mediated platelet/PMN adhesion. The anti-adhesive potency of vessel wall reversed by aspirin was dose-dependently restored by camonagrel (3-100 µM), a new TXA(2) synthase inhibitor. We conclude that selective TXA(2)-synthase inhibitors may inhibit P-selectin mediated platelet/PMN adhesion by augmenting formation of prostacyclin by vessel walls.