Polyamine deprivation provokes an antalgic effect

It is well established that inhibition of putrescine formation using D,L-2-(difluoromethyl)ornithine and feeding a polyamine-deficient diet together with non-absorbable antibiotics (neomycin and metronidazole), prevent almost completely the growth of tumors in rats. A similar regimen given to patients with prostate cancer not only reduced the titer of prostate specific antigen in serum, but surprisingly provoked at the same time an antalgic effect. This observation led us to study the potentiation effect of polyamine deprivation on pain threshold in healthy rats. Animals were fed for 2 weeks with an artificial diet of known polyamine content, in combination with antibiotics and 2-(difluoromethyl)ornithine, and were then submitted to pain stimuli using two models, the Randall-Selitto test and the Tail-Flick test. Polyamine deprivation produced in these models an increase in the latency of the response, even under conditions which did not produce significant changes of the polyamine concentrations in blood and brain. From these observations, we may conclude that the polyamines play a role in the perception of nociceptive stimuli under physiological conditions.     

Mediastinal masses: diagnostic approach

In most clinical situations the modern radiological approach to a mediastinal mass consists of performing a CT scan following the chest radiograph. Magnetic resonance imaging is indicated when CT findings are equivocal and as the first-line method in particular situations such as suspected involvement of the posterior mediastinum. In both techniques, tissular components of the mass assessed by density or signal intensity analysis, together with the precise location, are the leading edge of the radiological diagnosis. This review deals mainly with the differential diagnosis of primary neoplasms according to CT and MRI findings.     

Evidence that different mechanisms underlie smooth muscle relaxation to nitric oxide and nitric oxide donors in the rabbit isolated carotid artery

1. The endothelium-dependent relaxants acetylcholine (ACh; 0.03-10 microM) and A23187 (0.03-10 microM), and nitric oxide (NO), applied either as authentic NO (0.01-10 microM) or as the NO donors 3-morpholino-sydnonimine (SIN-1; 0.1-10 microM) and S-nitroso-N-acetylpenicillamine (SNAP; 0.1-10 microM), each evoked concentration-dependent relaxation in phenylephrine stimulated (1-3 microM; mean contraction and depolarization, 45.8+/-5.3 mV and 31.5+/-3.3 mN; n=10) segments of rabbit isolated carotid artery. In each case, relaxation closely correlated with repolarization of the smooth muscle membrane potential and stimulated a maximal reversal of around 95% and 98% of the phenylephrine-induced depolarization and contraction, respectively. 2. In tissues stimulated with 30 mM KCl rather than phenylephrine, smooth muscle hyperpolarization and relaxation to ACh, A23187, authentic NO and the NO donors were dissociated. Whereas the hyperpolarization was reduced by 75-80% to around a total of 10 mV, relaxation was only inhibited by 35% (n=4-7 in each case; P<0.01). The responses which persisted to ACh and A23187 in the presence of 30 mM KCl were abolished by either the NO synthase inhibitor L-NG-nitroarginine methyl ester (L-NAME; 100 microM) or the inhibitor of soluble guanylyl cyclase 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 microM; 10 min; n=4 in each case; P<0.01). 3. Exposure to ODQ significantly attenuated both repolarization and relaxation to ACh, A23187 and authentic NO, reducing the maximum changes in both membrane potential and tension to each relaxant to around 60% of control values (n=4 in each case; P<0.01). In contrast, ODQ almost completely inhibited repolarization and relaxation to SIN-1 and SNAP, reducing the maximum responses to around 8% in each case (n=3-5; P<0.01). 4. The potassium channel blockers glibenclamide (10 microM), iberiotoxin (100 nM) and apamin (50 nM), alone or in combination, had no significant effect on relaxation to ACh, A23187, authentic NO, or the NO donors SIN-1 and SNAP (n=4 in each case; P>0.05). Charybdotoxin (ChTX; 50 nM) almost abolished repolarization to ACh (n=4; P<0.01) and inhibited the maximum relaxation to ACh, A23187 and authentic NO each by 30% (n=4-8; P<0.01). Application of ODQ (10 microM; 10 min) abolished the ChTX-insensitive responses to ACh, A23187 and authentic NO (n=4 in each case; P<0.01 5. When the concentration of phenylephrine was reduced (to 0.3-0.5 microM) to ensure the level of smooth muscle contraction was the same as in the absence of potassium channel blocker, ChTX had no effect on the subsequent relaxation to SIN-1 (n=4; P>0.05). However, in the presence of tone induced by 1-3 microM phenylephrine (51.2+/-3.3 mN; n=4), ChTX significantly reduced relaxation to SIN-1 by nearly 50% (maximum relaxation 53.2+/-6.3%, n=4; P<0.01). 6. These data indicate that NO-evoked relaxation of the rabbit isolated carotid artery can be mediated by three distinct mechanisms: (a) a cyclic GMP-dependent, voltage-independent pathway, (b) cyclic GMP-mediated smooth muscle repolarization and (c) cyclic GMP-independent, ChTX-sensitive smooth muscle repolarization. Relaxation and repolarization to both authentic and endothelium-derived NO in this large conduit artery appear to be mediated by parallel cyclic GMP-dependent and -independent pathways. In contrast, relaxation to the NO-donors SIN-1 and SNAP appears to be mediated entirely via cyclic GMP-dependent mechanisms.     

Intestinal cell cycle regulations. Interactions of cyclin D1, Cdk4, and p21Cip1

OBJECTIVE: The p21Cip1 protein is a potent stoichiometric inhibitor of cyclin-dependent kinase activity, and p21Cip1 mRNA expression is localized to the nonproliferative compartment of the intestinal villus, suggesting an in vivo growth-inhibitory role in the gut. The authors determined whether nontransformed rat intestinal epithelial cells (IECs) underwent reversible cell cycle arrest by contact inhibition, and determined whether increases in the relative amount of p21 associated with cyclin D/Cdk4 protein complexes were associated with cell growth arrest. METHODS: Density arrest was achieved by prolonged culture IEC-6 in confluent conditions (5 or more days). Release from density arrest was achieved by detaching the cells from the culture plate and reseeding them at a 1:4 ratio. The DNA synthesis was estimated by [3H]-thymidine incorporation and expressed as mean plus or minus standard error of the mean (n = 4). Cyclin D1, Cdk4, and p21 mRNA and protein levels were determined by standard Northern and Western blot analyses, respectively. Cyclin D1, Cdk4, and p21 protein complex formation was analyzed by immunoprecipitating the complexes from cell lysates with an antibody to one of the constituents, followed by SDS polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis of the precipitated complexes using antibodies to the other proteins. The kinase activity of the immunoprecipitated Cdk4 was determined using recombinant Rb as substrate. RESULTS: The IEC-6[3H]-thymidine incorporation was decreased 7.5-fold from day 1 confluence to day 7 of confluence. Twenty-four hours after release from density arrest, there was a 43-fold increase in [3H]-thymidine incorporation. Cyclin D1 and Cdk4 mRNA levels remained relatively constant during contact inhibition, whereas immunoblotting showed that the levels of cyclin D1 and Cdk4 proteins decreased by 70.9% and 68.7%, respectively, comparing day 3 with day 9 during density arrest. The levels of cyclin D1 increased 5.8-fold and Cdk4 increased by 4.4-fold by 24 hours after reseeding the day 9 density-arrested cultures, coincident with the increase in DNA synthesis. The amount of p21 associated with the cyclin D1 and Cdk4 complex in the density-arrested cells was 170% of that observed in the reseeded, proliferating cells. More important, the p21::Cdk4 ratio was 6.4-fold higher in the density-arrested (quiescent) cells as compared with rapidly proliferating cells by 24 hours after release from growth arrest. Recovery of Cdk4-dependent kinase activity occurred by 4 hours after release from growth arrest, coincident with decreased binding of p21 to the complex. CONCLUSIONS: Intestinal epithelial cells in culture can undergo density-dependent growth arrest. This process involves downregulation of cyclin D1 and Cdk4 at the level of protein expression, whereas the mRNA levels remain relatively unchanged. Further, during contact inhibition, there is more p21 associated with cyclin D1/Cdk4, which further contributes to the inhibition of the kinase complex. The authors also have shown that the process of contact inhibition is reversible, which may explain partly the ability of the intestinal epithelium to increase proliferative activity in response to injury.     

Mental chronometry with simple linear regression

Typically, mental chronometry is performed by means of introducing an independent variable postulated to affect selectively some stage of a presumed multistage process. However, the effect could be a global one that spreads proportionally over all stages of the process. Currently, there is no method to test this possibility although simple linear regression might serve the purpose. In the present study, the regression approach was tested with tasks (memory scanning and mental rotation) that involved a selective effect and with a task (word superiority effect) that involved a global effect, by the dominant theories. The results indicate (1) the manipulation of the size of a memory set or of angular disparity affects the intercept of the regression function that relates the times for memory scanning with different set sizes or for mental rotation with different angular disparities and (2) the manipulation of context affects the slope of the regression function that relates the times for detecting a target character under word and nonword conditions. These ratify the regression approach as a useful method for doing mental chronometry.     

The distribution of IL-13 receptor alpha1 expression on B cells, T cells and monocytes and its regulation by IL-13 and IL-4

To study the expression of IL-13 receptor alpha1 (IL-13Ralpha1), specific monoclonal antibodies (mAb) were generated. Surface expression of the IL-13Ralpha1 on B cells, monocytes and T cells was assessed by flow cytometry using these specific mAb. Among tonsillar B cells, the expression was the highest on the IgD+ CD38- B cell subpopulation which is believed to represent naive B cells. Expression was also detectable on a large fraction of the IgD-CD38- B cells but not on CD38+ B cells. Activation under conditions which promote B cell Ig class switching up-regulated the expression of the receptor. However, the same stimuli had an opposite effect for IL-13Ralpha1 expression levels on monocytes. While IL-13Ralpha1 mRNA was clearly detectable in T cell preparations, no surface expression was detected. However, permeabilization of the T cells showed a clear intracellular expression of the receptor. A soluble form of the receptor was immunoprecipitated from the supernatant of activated peripheral T cells, suggesting that T cell IL-13Ralpha1 might have functions unrelated to the capacity to form a type II IL-4/IL-13R with IL-4Ralpha.     

Inhibition of gastric emptying and intestinal transit by amphetamine through a mechanism involving an increased secretion of CCK in male rats

1. The effect of amphetamine on gastrointestinal (GI) transit and the plasma levels of cholecystokinin (CCK) were studied in male rats. 2. Gastric emptying was inhibited both acutely and chronically by the administration of amphetamine. GI transit was decreased by the acute administration of amphetamine but not affected by the chronic administration of amphetamine. 3. Plasma CCK levels were increased dose-dependently by amphetamine. 4. Proglumide, a CCK receptor antagonist, prevented amphetamine-induced inhibition of gastric emptying and the decrease in GI transit in male rats. 5. The selective CCK(A) receptor antagonist, lorglumide, dose-dependently attenuated the amphetamine-induced inhibition of gastric emptying in male rats. In contrast, the selective CCK(B) receptor antagonist, PD 135,158, did not reverse the effect of amphetamine on gastric emptying. 6. Both lorglumide and PD 135,158 reversed the inhibitory effect of amphetamine on GI transit in male rats. 7. These results suggest that amphetamine-induced inhibition of gastric emptying and intestinal transit is due in part to a mechanism associated with the hypersecretion of endogenous CCK.     

Long-term follow-up of patients with Crohn's disease treated with azathioprine or 6-mercaptopurine

BACKGROUND: Crohn's ulcerative gastrointestinal disease is presently managed through a variety of medical interventions, including-according to severity of illness-anti-inflammatory, immunosuppressive, and corticosteroid agents; and with remedial surgery to correct anatomical abnormalities caused by disease processes. The immunosuppressant azathioprine (or its metabolite, 6-mercaptopurine) is considered an efficient maintenance therapy for Crohn's, but there is always concern about bone-marrow suppression, liver damage, and other adverse effects. For how long persons with this disease should be given these drugs has not been determined. METHODS: Patients who were treated with azathioprine or 6-mercaptopurine for more than 6 months, and who were in prolonged clinical remission (> 6 months without steroids) were followed. The time-to-relapse was analysed in those on treatment, in those who stopped treatment for reasons other than a relapse, and in the whole sample, taking into account that they could be treated with the drugs or not, as a function of time. The influence of concomitant variables on time-to-relapse rate was examined using the Cox proportional hazard model. FINDINGS: In the 157 patients who continued to take the therapy, cumulative probabilities of relapse at 1 and 5 years were 11% and 32% respectively. Female gender, younger age, and a time for achieving remission more than 6 months were associated with a higher risk of relapse. In 42 patients who stopped therapy, probabilities of relapse at 1 and 5 years were 38% and 75%, respectively. Male gender, younger age and duration of remission less than 4 years were associated with a higher risk of relapse. After 4 years of remission on these drugs, the risk of relapse appeared to be similar, whether the therapy was maintained or stopped. INTERPRETATION: Taking into account the potential risks of long-term immunosuppressive therapy, the usefulness of maintaining azathioprine or 6-mercaptopurine in patients who have been in remission for more than 4 years is questionable.     

Influence of cigarette smoking on rate of reopening of the infarct-related coronary artery after myocardial infarction: a multivariate analysis

OBJECTIVES: This study sought to determine whether the reopening of the infarct-related vessel is related to clinical characteristics or cardiovascular risk factors, or both. BACKGROUND: In acute myocardial infarction, thrombolytic therapy reduces mortality by restoring the patency of the infarct-related vessel. However, despite the use of thrombolytic agents, the infarct-related vessel remains occluded in up to 40% of patients. METHODS: We studied 295 consecutive patients with an acute myocardial infarction who underwent coronary angiography within 15 days (mean [+/- SD] 6.7 +/- 3.2 days) of the onset of symptoms. Infarct-related artery patency was defined by Thrombolysis in Myocardial Infarction trial flow grade > or = 2. Four cardiovascular risk factors--smoking, hypertension, hypercholesterolemia and diabetes mellitus--and eight different variables-age, gender, in-hospital death, history of previous myocardial infarction, location of current myocardial infarction, use of thrombolytic agents, time interval between onset of symptoms, thrombolytic therapy and coronary angiography--were recorded in all patients. RESULTS: Thrombolysis in current smokers and anterior infard location on admission were the three independent factors highly correlated with the patency of the infarct-related vessel (odds ratios 3.2, 3.0 and 1.9, respectively). In smokers, thrombolytic therapy was associated with a higher reopening rate of the infard vessel, from 35% to 77% (p < 0.001). Nonsmokers did not benefit from thrombolytic therapy, regardless of infarct location. CONCLUSIONS: These observational data, if replicated, suggest that in patients with acute myocardial infarction, thrombolytic therapy may be most effective in current smokers, whereas nonsmokers and ex-smokers may require other management strategies, such as emergency percutaneous transluminal coronary angioplasty.