S-adenosyl-L-methionine prevents 5-HT(1A) receptors up-regulation induced by acute imipramine in the frontal cortex of the rat

S-adenosyl-L-methionine (SAM) has shown efficacy in speeding the onset of the antidepressant effect of imipramine in depressed patients. This effect may be related to their interactions at the serotonin(1A) (5-HT(1A)) receptors. Acute imipramine up-regulated the frontal cortex 5-HT(1A) receptors (B(max), 51.5 +/- 8.4 fmol/mg protein) vs. saline (B(max), 27.5 +/- 5.9 fmol/mg protein), and did not show antidepressant effect. Acute SAM and imipramine+SAM did not modify frontal cortex 5-HT(1A) receptors, and showed antidepressant effects (decrease of the immobility response of 26%, P<0.01; and 47%, P<0.001) vs. saline. All the chronic treatments showed antidepressant effects and up-regulated the hippocampus 5-HT(1A) receptors. SAM prevents the 5-HT(1A) receptor up-regulation induced by acute imipramine in the frontal cortex. This mechanism may contribute to imipramine's antidepressant effect.     

Immunophenotyping of leukocytes in peripheral blood of BALB/c mice infected with mouse herpesvirus isolate 72

We characterized leukocytes in peripheral blood of BALB/c mice infected with mouse herpesvirus isolate 72 (MHV-72) representing an isolate of mouse herpesvirus strain 68 (MHV-68, species Murid herpesvirus 4, genus Rhadinovirus, subfamily Gammaherpesvirinae, family Herpesviridae) (van Regenmortel et al., 2000). In acute infection (up to day 30 post infection (p.i.)) the number of CD8+ T cells increased, reaching a maximum at day 11 p.i. This increase correlated with that of CD4+ T, activated CD 19+ B and natural killer (NK) cells. At day 30 p.i. the numbers of CD4+, CD8+, CD14+ and CD19+ cells decreased to normal values. A similar increase in the number of these cells was observed at day 730 p. i. In the course of persistent infection (after day 30 p.i.) some of the mice developed a leukemia-like syndrome characterized by an increase in the number of leukocytes and appearance of atypical, blastic immature forms of leukocytes. The latter forms of leukocytes were characteristic by an increased amount of argyrophilic proteins. These results show further similarities between MHV-72 (another isolate of MHV-68) and EBV infections and justify the use of MHV-68 or MHV-72 as an appropriate mouse model for the study of EBV infection of humans.     

Cell proliferation depends on mitochondrial Ca2+ uptake: inhibition by salicylate

Store-operated Ca²⁺ entry (SOCE) is a ubiquitous Ca²⁺ influx pathway involved in control of multiple cellular and physiological processes including cell proliferation. Recent evidence has shown that SOCE depends critically on mitochondrial sinking of entering Ca²⁺ to avoid Ca²⁺-dependent inactivation. Thus, a role of mitochondria in control of cell proliferation could be anticipated. We show here that activation of SOCE induces cytosolic high [Ca²⁺] domains that are large enough to be sensed and avidly taken up by a pool of nearby mitochondria. Prevention of mitochondrial clearance of the entering Ca²⁺ inhibited both SOCE and cell proliferation in several cell types including Jurkat and human colon cancer cells. In addition, we find that therapeutic concentrations of salicylate, the major metabolite of aspirin, depolarize partially mitochondria and inhibit mitochondrial Ca²⁺ uptake, as revealed by mitochondrial Ca²⁺ measurements with targeted aequorins. This salicylate-induced inhibition of mitochondrial Ca²⁺ sinking prevented SOCE and impaired cell growth of Jurkat and human colon cancer cells. Finally, direct blockade of SOCE by the pyrazole derivative BTP-2 was sufficient to arrest cell growth. Taken together, our results reveal that cell proliferation depends critically on mitochondrial Ca²⁺ uptake and suggest that inhibition of tumour cell proliferation by salicylate may be due to interference with mitochondrial Ca²⁺ uptake, which is essential for sustaining SOCE. This novel mechanism may contribute to explaining the reported anti-proliferative and anti-tumoral actions of aspirin and dietary salicylates.     

Four new HLA class I alleles in Cauacasoids

Four new HLA classical class I alleles in the three loci are described in Caucasian individuals. A*3012 was first suspected by an abnormal serologic pattern that would be explained by the single amino acid substitution at the A30-specific Ser17. B*270505 differs from B*270502 in a silent substitution at an up to now constant position in the B locus. B*3541 encodes for a new Cys at position 118 that has not been encountered in neither human nor primate alleles. Cw*0716 seems to be originated by a large-scale interallelic recombination event between Cw*0701/*0706/*0718 and Cw*020202, giving rise to a new antigen-binding cleft conformation.     

Functional and pharmacological properties of GABArho1delta51 receptors

Gamma-aminobutyrate is the main inhibitory neurotransmitter in the vertebrate brain, and the gamma-aminobutyric acid (GABA) receptor subunit GABArho1delta51 is an alternatively spliced form of the GABArho1 receptor that was recently isolated from human retina cDNA libraries. The rho1delta51 receptor subunit lacks 17 amino acids in the extracellular N-terminal domain and, when expressed in Xenopus oocytes, forms functional homomeric GABA receptors. Unexpectedly, even after a such a big deletion, the fundamental properties of the deleted variant receptors are very similar to those of the complete GABArho1 receptors. For example, both types of receptors are bicuculline resistant, desensitize very little, and are negatively modulated by Zn2+ and positively modulated by La3+. In spite of such similarities, the GABArho1delta51 receptors are more sensitive to GABA, to the specific GABA(C) antagonist (1,2,5,6-tetrahydropyridine-4-yl)methylphosphinic acid and to Zn2+, than the complete GABArho1 receptors. The GABArho1delta51 receptors extend the variety of inhibitory receptors in the retina. Their functional significance still remains to be determined.     

Synergistic effect of azithromycin on the phagocytic killing ofStaphylococcus aureus by human polymorphonuclear leukocytes

Many macrolides have been shown to affect the interaction between bacteria and various immune defense mechanisms, such as chemotaxis, accumulation, and bioactivity within phagocytic cells. The interaction of azithromycin with human polymorphonuclear leukocytes (PMNs) was studied in vitro and compared with the interactions between other macrolides and PMNs. The opsonophagocytic killing ofStaphylococcus aureus was synergistically enhanced by azithromycin at concentrations below and above the minimal inhibitory concentration, with a reduction of up to 2.82 log₁₀ cfu/ml with 2 mg/ml of azithromycin. Other macrolides were effective only at subinhibitory concentrations. The beneficial azithromycin-leukocyte interaction may explain azithromycin's efficacy against intracellular pathogens.     

Phase I clinical evaluation of a neutralizing monoclonal antibody against epidermal growth factor receptor in advanced brain tumor patients: preliminary study

High levels of growth factors and their receptors have been demonstrated in human tumors. Gliomas and meningiomas are characterized by overexpression of epidermal growth factor receptor (EGF-R). Ior egf/r3, is a neutralizing murine monoclonal antibody (MAb) against EGF-R, and was generated at the Cuban Institute of Oncology. The antibody recognizes EGF-R with high affinity, inhibiting tyrosine kinase activation. A clinical trial was conducted in brain tumor patients to evaluate toxicity, immunogenicity, and clinical benefit of escalating doses of the antibody. Nine patients with histologically confirmed gliomas or meningiomas, who had active or recurrent disease after receiving conventional treatment, received four intravenous doses of ior egf/r3. Total dosages ranged from 160 to 480 mg. As inclusion criteria, radioimmunoscintigraphy with the same MAb labeled with 99mTechnetium (99mTc) was performed. Immune response against the murine antibody was also evaluated. After four doses of ior egf/r3 MAb, no significant toxicity was found, except in one patient who developed a grade 4 allergic adverse event. This reaction was probably related with previous sensitization to the same MAb and the development of human anti-mouse antibodies (HAMA) response. Despite no major objective antitumor responses, eight patients had stable disease on the 6-month evaluation, and two patients remain alive after four years of MAb therapy.     

Cathepsin C gene: First compound heterozygous patient with Papillon-Lefèvre syndrome and a novel symptomless mutation

Papillon-Lefèvre syndrome (PLS) has recently been shown to be caused by mutations in the cathepsin C gene resulting in periodontal disease and palmoplantar keratosis. Thirteen different homozygous mutations have been characterised in PLS patients of different ethnic origin. In the present paper, a PLS patient is described who carries two novel mutations (706G>T and 872G>A) in the paternal and maternal chromosomes, respectively. This is the first compound patient described so far. In addition, a novel symptomless mutation (458C>T) in the cathepsin C gene is described in three homozygous individuals. Thus, not all mutations should be considered as a cause of disease, whether case studies or general population screening is performed. Another already described mutation that provoked the Haim-Munk syndrome (HMS) in Indian Jews has also been found to give rise to PLS in a Spanish family from Madrid. On the other hand, PLS patients are ameliorated by retinoids, which indicates that retinoids may be used as therapeutic agents in this immune system deficiency.     

Looking for ferns: optimization of digestion pretreatment in fluorescence in situ hybridization (FISH) technique on paraffin-embedded tissues.

Fluorescence in situ hybridization (FISH) is a useful cytogenetic technique for the detection of chromosome aberrations. However, applying this technique routinely on paraffin-embedded tissue is hampered by technical problems. The efficiency of hybridization is influenced by formalin fixation time, and this may vary considerably between specimens. We present a simple method for improving hybridization by microscopically monitoring the time of enzymatic digestion. To establish optimal digestion time, enzymatic digestion was stopped at 3-minute intervals for biopsies and 10-minute intervals for autopsies in 24 paraffin-embedded samples. At every stop, tissue morphology was examined under light microscopy to determine if observed changes could be correlated with subsequent FISH results. The appearance of fernlike formations was found to mark the optimal digestion time that produced the strongest hybridization signals. Using this method of digestion time control, an additional 41 cases were evaluated for FISH with various types of probe. Monitoring under the microscope could be more spaced if the morphology did not change after the first visual control and could be adapted to the type of sample (in general, endoscopic samples, total digestion time of about 10 min; routine biopsies, 15 to 30 min; autopsy samples, 20 to 40 min). In every case, the appearance of the fernlike pattern correlated with proper hybridization signal. Monitoring digestion time for the appearance of fernlike structures is a useful method for improving reproducibility of FISH technique on paraffin-embedded samples. It is particularly useful when dealing with samples under heterogeneous fixation conditions (consultations, autopsies, etc.), because it eliminates the need for repetition.     

The effect of oral hormone replacement therapy on lipoprotein profile, resistance of LDL to oxidation and LDL particle size

Objectives: To disclose if oral estradiol (E₂), alone or in combination with natural progesterone (P) or medroxyprogesterone acetate (MPA), may modify the oxidizability of low density lipoprotein (LDL), and if the effect is achieved at physiological dosages. LDL oxidizability was assessed by the resistance to oxidation by copper and by the particle size profile, since small particles have increased oxidation susceptibility. Methods: Thirty-three women received two consecutive, two-month length doses of 1 and 2 mg/day of oral E₂. They were then randomly assigned to a fourteen-day treatment of 2 mg/day E₂ plus either 300 mg/day P or 5 mg/day MPA. A parallel group of experiments was performed on a pool of baseline plasma, where hormones were added at the desired concentration. Lipoprotein levels, resistance of LDL to oxidation, and LDL particle diameter, were measured at baseline and after each treatment. Results: Estradiol reduced LDL levels and increased high density lipoprotein (HDL) and triglycerides. P abolished these changes, whereas MPA only reversed the increase of HDL. Estradiol protected LDL from oxidation in a dose-dependent manner, although only at pharmacological concentrations (1 μM or higher). Both P and MPA were inert at either physiological or pharmacological concentrations. The size of the LDL particles remained unaffected except under MPA, in which it was reduced. Conclusions: Estradiol has a protective effect against LDL oxidation, although only at pharmacological dosages. P and MPA did not limit the E₂ action. The size of the LDL particles remained unaltered after each E₂ dose, but MPA, and not P, was associated with a diminution.