Prevalence of Abdominal Aortic Aneurysm in Men Aged 65-74 Years in a Metropolitan Area in North-East Spain.

BACKGROUND: A declining prevalence of AAA and a shift in the distribution towards the older population have been observed during the last decade in Europe. The aim was to estimate the current screening prevalence of AAA in men aged 65-74 years in a metropolitan area in north-east Spain and to identify associated risk factors. METHODS: A cross sectional prevalence study in men registered in L'Hospitalet Primary Healthcare Services (Barcelona, Spain) was performed. There were 619 randomly selected subjects (expected prevalence of aneurysm, 5%; accuracy of estimation, ±2%; loss to follow up, 30%). Exclusion criteria were life expectancy <1 year, limited quality of life, previous diagnosis of AAA, prior aorto-femoral surgery, and non-Caucasian. The following were measured: internal diameter of the infrarenal abdominal aorta using ultrasound, cardiovascular risk factors, personal (heart disease, stroke, peripheral vascular disease) and family history (AAA), physical examination, and blood tests. We estimated the prevalence and 95% confidence interval of AAA, and used logistic regression analysis to identify risk factors for AAA. RESULTS: Among the 651 individuals included in the analysis the prevalence of aneurysm was 2.30% (95% CI, 1.30-3.77%). In the regression analysis, AAA was associated with smoking (0-10, 11-20, or >20 cigarettes/day), diagnosis of myocardial infarction, and being taller than the median (165 cm). CONCLUSIONS: The current screening prevalence of AAA among men aged 65-74 years in a metropolitan area in north-east Spain is similar to that in northern Europe. Smoking, myocardial infarction, and height were associated with the presence of AAA.

Isotopic discrimination of stable isotopes of nitrogen (δ15N) and carbon (δ13C) in a host-specific holocephalan tapeworm.

During the past decade, parasites have been considered important components of their ecosystems since they can modify food-web structures and functioning. One constraint to the inclusion of parasites in food-web models is the scarcity of available information on their feeding habits and host-parasite relationships. The stable isotope approach is suggested as a useful methodology to determine the trophic position and feeding habits of parasites. However, the isotopic approach is limited by the lack of information on the isotopic discrimination (ID) values of parasites, which is pivotal to avoiding the biased interpretation of isotopic results. In the present study we aimed to provide the first ID values of δ(15)N and δ(13)C between the gyrocotylidean tapeworm Gyrocotyle urna and its definitive host, the holocephalan Chimaera monstrosa. We also test the effect of host body size (body length and body mass) and sex of the host on the ID values. Finally, we illustrate how the trophic relationships of the fish host C. monstrosa and the tapeworm G. urna could vary relative to ID values. Similar to other studies with parasites, the ID values of the parasite-host system were negative for both isotopic values of N (Δδ(15)N = - 3.33 ± 0.63‰) and C (Δδ(13)C = - 1.32 ± 0.65‰), independent of the sex and size of the host. By comparing the specific ID obtained here with ID from other studies, we illustrate the importance of using specific ID in parasite-host systems to avoid potential errors in the interpretation of the results when surrogate values from similar systems or organisms are used.

DNA copy number profiling reveals extensive genomic loss in hereditary BRCA1 and BRCA2 ovarian carcinomas.

BACKGROUND: Few studies have attempted to characterise genomic changes occurring in hereditary epithelial ovarian carcinomas (EOCs) and inconsistent results have been obtained. Given the relevance of DNA copy number alterations in ovarian oncogenesis and growing clinical implications of the BRCA-gene status, we aimed to characterise the genomic profiles of hereditary and sporadic ovarian tumours. METHODS: High-resolution array Comparative Genomic Hybridisation profiling of 53 familial (21 BRCA1, 6 BRCA2 and 26 non-BRCA1/2) and 15 sporadic tumours in combination with supervised and unsupervised analysis was used to define common and/or specific copy number features. RESULTS: Unsupervised hierarchical clustering did not stratify tumours according to their familial or sporadic condition or to their BRCA1/2 mutation status. Common recurrent changes, spanning genes potentially fundamental for ovarian carcinogenesis, regardless of BRCA mutations, and several candidate subtype-specific events were defined. Despite similarities, greater contribution of losses was revealed to be a hallmark of BRCA1 and BRCA2 tumours. CONCLUSION: Somatic alterations occurring in the development of familial EOCs do not differ substantially from the ones occurring in sporadic carcinomas. However, some specific features like extensive genomic loss observed in BRCA1/2 tumours may be of clinical relevance helping to identify BRCA-related patients likely to respond to PARP inhibitors.

Juvenile Huntington's disease: a case report and literature review.

Huntington's disease is the most frequent neurodegenerative disease with a prevalence of fewer than 10 cases per 10,000 inhabitants; the juvenile form is responsible for less than 10% of all cases. Huntington's disease belongs to the group known as "triad syndromes," which evolve with cognitive, motor and neuropsychiatric manifestations. Around 30% of patients debut with behavioral symptoms, which are a major challenge for management by patients, families, and caregivers. Huntington's disease (HD) is reviewed and a case of juvenile onset is reported in this article. The characteristics of juvenile-onset Huntington's disease (HD) differ from those of adult-onset HD, as chorea does not occur, although bradykinesia, dystonia, and signs of cerebellar disorder, such as rigidity, are present, frequently in association with convulsive episodes and psychotic manifestations.

Early graft dysfunction after liver transplant: Comparison of different diagnostic criteria in a single-center prospective cohort.

OBJECTIVE: Comparison of different diagnostic criteria for early liver allograft dysfunction (EAD) and their capability to predict mortality. DESIGN: Single-center, prospective, cohort study. SETTINGS: ICU in a Regional Hospital with a liver transplant program since 1997. PATIENTS: 253 consecutive patients admitted to our ICU immediately after liver transplantation between 2009 and 2015. VARIABLES OF INTEREST: Differences in the incidence of EAD and its relation with ICU, Hospital and 2-year mortality depending on the definition applied using as comparator the UNOS (United Network for Organ Sharing) primary non-function criterion. RESULTS: The incidence of early liver allograft dysfunction according to UNOS was 13.8%, to Makowka 6.3%, to Ardite 10.7%, to Nanashima 20.6%, to Dhillon 30.8% and to MEAF 13.4%. Kappa test did not show a good correlation among these criteria. EAD was related with ICU mortality for all diagnostic criteria except Dhillon but only UNOS, Makowka and MEAF were associated with 2-year mortality. Hospital mortality was poorly predicted by all criteria except for the MEAF score. CONCLUSION: We found a poor agreement between different criteria analyzed for the diagnosis of EAD. In our population, the MEAF score showed the best relationship with short- and long-term mortality.

C-reactive protein at ICU admission as a marker of early graft dysfunction after liver transplant. A prospective, single-center cohort study.

OBJECTIVE: To explore the behavior of C-reactive protein (CRP) after orthotopic liver transplantation (OLT) during the first postoperative days, and its usefulness as a marker of severe early allograft dysfunction (EAD). DESIGN: A prospective, single-center cohort study was carried out. SETTING: The Intensive Care Unit (ICU) of a regional hospital with a liver transplant program since 1997. PATIENTS: The study comprised a total of 183 patients admitted to our ICU immediately after liver transplantation between 2009 and 2015. VARIABLES OF INTEREST: C-reactive protein levels upon ICU admission and after 24 and 48h, severe EAD and hospital mortality. RESULTS: The CRP levels after OLT were: upon ICU admission 57.5 (51.6-63.3)mg/L, after 24h 80.1 (72.9-87.3)mg/L and after 48h 69.9 (62.5-77.4)mg/L. Severe EAD patients (14.2%) had higher mortality (23.1 vs 2.5; OR 11.48: 2.98-44.19) and lower CRP upon ICU admission (39.3 [29.8-48.7]mg/L) than the patients without EAD (0.5 [53.9-67.0]; p<0.05] - the best cut-off point being 68mg/L (sensitivity 92.3%; specificity 40.1%; Youden index 0.33). Lower CRP upon ICU admission was correlated to higher mortality (24.5 [9.2-39.7] vs 59.4 [53.4-65.4]; p<0.01, AUC 0.79 [0.65-0.92]). CONCLUSION: Liver transplant is a strong inflammatory stimulus accompanied by high levels of C-reactive protein. A blunted rise in CRP on the first postoperative day after OLT may be a marker of poor allograft function and is related to hospital mortality.

[Middle cerebral artery ischemic stroke presentation of miliar tuberculosis]. / Infarto de la arteria cerebral media como forma de presentación de la tuberculosis miliar.

Presence of central nervous system by extrapulmonary tuberculosis is an infrequent disease specially among non HIV infected patients, and it is associated with poor prognosis and high mortality rates. We report a case with a middle cerebral artery ischemic strocke as a first symptom of miliar tuberculosis.

Differential roles of Akt, Rac, and Ral in R-Ras-mediated cellular transformation, adhesion, and survival.

Multiple biological functions have been ascribed to the Ras-related G protein R-Ras. These include the ability to transform NIH 3T3 fibroblasts, the promotion of cell adhesion, and the regulation of apoptotic responses in hematopoietic cells. To investigate the signaling mechanisms responsible for these biological phenotypes, we compared three R-Ras effector loop mutants (S61, G63, and C66) for their relative biological and biochemical properties. While the S61 mutant retained the ability to cause transformation, both the G63 and the C66 mutants were defective in this biological activity. On the other hand, while both the S61 and the C66 mutants failed to promote cell adhesion and survival in 32D cells, the G63 mutant retained the ability to induce these biological activities. Thus, the ability of R-Ras to transform cells could be dissociated from its propensity to promote cell adhesion and survival. Although the transformation-competent S61 mutant bound preferentially to c-Raf, it only weakly stimulated the mitogen-activated protein kinase (MAPK) activity, and a dominant negative mutant of MEK did not significantly perturb R-Ras oncogenicity. Instead, a dominant negative mutant of phosphatidylinositol 3-kinase (PI3-K) drastically inhibited the oncogenic potential of R-Ras. Interestingly, the ability of the G63 mutant to induce cell adhesion and survival was closely associated with the PI3-K-dependent signaling cascades. To further delineate R-Ras downstream signaling events, we observed that while a dominant negative mutant of Akt/protein kinase inhibited the ability of R-Ras to promote cell survival, both dominant negative mutants of Rac and Ral suppressed cell adhesion stimulated by R-Ras. Thus, the biological actions of R-Ras are mediated by multiple effectors, with PI3-K-dependent signaling cascades being critical to its functions.

[Infected cholangiocarcinoma]. / Colangiocarcinoma infectado.

Cholangiocarcinomas are malignancies of the biliary duct system. They are encountered in 3 geographic regions: intrahepatic, proximal extrahepatic, and distal extrahepatic. The etiology of most bile duct cancers remains undetermined but some risk factors, like gallstone, have been suggested to play a role by inducing malignant transformation. The prognosis and clinical manifestations depend on the anatomical location and clinical presentation may be confuses or by means of complications like sepsis. We present a case of cholangiocarcinoma which made debut with cholestasis and sepsis in a cholecystectomiced patient, who had a long standing lithiasic cholecystitis.

Synergism between two growth regulatory pathways: cooperative transformation of NIH3T3 cells by G alpha 12 and c-raf-1.

The alpha-subunit of the heterotrimeric G-protein, G12, has been shown to induce cellular transformation when overexpressed or oncogenically activated in rodent fibroblasts. To investigate the interaction between Galpha12 transforming pathway and the Ras-Raf-MAPK pathway, we examined the ability of mitogenic signaling molecules in cooperating with Galpha12 in transforming NIH3T3 fibroblasts. We observed a striking cooperative effect on focus-forming ability when Galpha12 and c-raf-1 cDNAs were co-transfected into NIH3T3 cells. NIH3T3 cells coexpressing both Galpha12 and c-raf-1 resulted in the constitutive activation of the mitogenic-activated protein kinase (MAPK). In addition, the levels of GTP-bound Ras were elevated in Galpha12 transformed NIH3T3 cells. Our results provide a model for studying the effects of simultaneous activation of two distinct growth regulatory pathways in cellular transformation.

Oncogenic activation of human R-ras by point mutations analogous to those of prototype H-ras oncogenes.

R-ras, K-rev-1/rap and TC21, are more closely related to prototype H-ras than any other known members of the ras superfamily. We recently isolated a mutationally activated TC21 oncogene from a human ovarian carcinoma cell line. Based upon these observations, we sought to re-examine the transforming potential of R-ras, which was reported earlier to lack transforming capacity. Mutations were introduced into the R-ras gene at codons 38 or 87, analogous to positions 12 and 61, respectively, responsible for H-ras oncogene activation. While both mutations resulted in acquisition of R-ras transforming capacity for NIH3T3 cells the position 61 was shown to be more active. Transfectants expressing either R-ras mutant formed colonies in soft agar and were tumorigenic in vivo. As has been reported for H-ras, R-ras cooperated with c-raf-1 in inducing transformation of NIH3T3 cells. These results imply interactions in R-ras and c-raf-1 signaling pathways. We observed R-ras transcripts of 4.6 and 1.2 kb ubiquitously expressed in each of a variety of tissues examined. All these findings raise the possibility that R-ras, like prototype ras genes, may be mutationally activated as an oncogene in some human malignancies.

Cooperative transformation of NIH3T3 cells by G alpha12 and Rac1.

The heterotrimeric G-protein, G alpha12, together with the closely-related G alpha13, are members of the G12 class of alpha-subunits important in mediating the signaling from seven transmembrane domain-spanning receptors. Recent evidence implicating both G alpha12 and G alpha13 in the activation of signaling pathways involving members of the RHO gene family led us to examine the role of Rac1, RhoA and Cdc42Hs in the transforming properties of G alpha12. Asparagine 17 (Asn 17) dominant inhibitory mutants of Rac1, and to a lesser extent RhoA, block focus forming ability of the GTPase-deficient mutant of G alpha12 (G alpha12 Leu 229) in NIH3T3 cells. In turn, wild-type G alpha12 cooperates well with Rac1 Val 12 but not with RhoA Leu 63 mutant in transforming NIH3T3 cells. Interestingly, the morphology of foci induced by G alpha12 and RhoA mutants are strikingly similar and is distinct from those displayed by Rac1 Val 12 mutant. The fact that G alpha12's ability to induce mitogenesis in NIH3T3 cells is not significantly perturbed by C3 ribosyltransferase suggested that RhoA does not play a major role in G alpha12-induced mitogenic events. Activated mutant of Rac1 has previously been demonstrated to stimulate the activity of the stress-induced c-Jun N-terminal kinase/stress-activated protein kinases (JNK/SAPKs). Transient co-transfection of Rac1 Val 12 mutant with the wild-type G alpha12 in COS7 cells leads to the further activation of an exogenously expressed hemagglutinin(HA)-tagged JNK. Furthermore, the cooperation between G alpha12 and Rac1 in cellular transformation is correlated with their ability to stimulate transcription from c-fos serum response element (SRE).

A novel insertional mutation in the TC21 gene activates its transforming activity in a human leiomyosarcoma cell line.

TC21 is the fourth member of the ras gene family to exhibit oncogenic activation in human tumor cells. To assess the prevalence of activated TC21 oncogenes in human tumors, we have developed sensitive single-strand conformational polymorphism (SSCP) conditions and immunological reagents for the detection of both single base alterations and/or overt overexpression in a wide spectrum of human tumor cell lines and surgical samples. In an initial examination of 33 human tumor specimens, we observed a novel nine basepair three amino acids insertion at TC21 codon 24 in one human uterine leiomyosarcoma cell line, SK-UT-1. This mutant allele when transfected into NIH3T3 cells, displayed high transforming activity comparable to that of the Leu72 oncogenic mutant identified by expression cDNA cloning from a human ovarian carcinoma cell line. Comparing the level of GTP-binding by the mutant and normal TC21 products revealed that this novel lesion increases the GTP-bound form of the TC21 molecule. These findings imply that the mechanism by which mutations activate the oncogenic properties of this ras-related molecule is analogous to that of previously known ras family members.

Isoform-specific insertion near the Grb2-binding domain modulates the intrinsic guanine nucleotide exchange activity of hSos1.

Two human hSos1 isoforms (Isf I and Isf II; Rojas et al., Oncogene 12, 2291-2300, 1996) defined by the presence of a distinct 15 amino acid stretch in one of them, were compared biologically and biochemically using representative NIH3T3 transfectants overexpressing either one. We showed that hSos1-Isf II is significantly more effective than hSos1-Isf I to induce proliferation or malignant transformation of rodent fibroblasts when transfected alone or in conjunction with normal H-Ras (Gly12). The hSos1-Isf II-Ras cotransfectants consistently exhibited higher saturation density, lower cell-doubling times, increased focus-forming activity and higher ability to grow on semisolid medium and at low serum concentration than their hSos1-Isf I-Ras counterparts. Furthermore, the ratio of GTP/GDP bound to cellular p21ras was consistently higher in the hSos1-Isf II-transfected clones, both under basal and stimulated conditions. However, no significant differences were detected in vivo between Isf I- and Isf II-transfected clones regarding the amount, stability and subcellular localization of Sos1-Grb2 complex, or the level of hSos1 phosphorylation upon cellular stimulation. Interestingly, direct Ras guanine nucleotide exchange activity assays in cellular lysates showed that Isf II transfectants consistently exhibited about threefold higher activity than Isf I transfectants under basal, unstimulated conditions. Microinjection into Xenopus oocytes of purified peptides corresponding to the C-terminal region of both isoforms (encompassing the 15 amino acid insertion area and the first Grb2-binding motif) showed that only the Isf II peptide, but not its corresponding Isf I peptide, was able to induce measurable rates of meiotic maturation, and synergyzed with insulin, but not progesterone, in induction of GVBD. Our results suggest that the increased biological potency displayed by hSos1-Isf II is due to higher intrinsic guanine nucleotide exchange activity conferred upon this isoform by the 15 a.a. insertion located in proximity to its Grb2 binding region.

Phase I clinical investigation of amonafide.

Amonafide (benzisoquinolinedione, NSC 308847) is a new synthetic imide antineoplastic agent with DNA intercalative properties that has been evaluated in a phase I clinical trial. The drug was administered as a single intravenous (IV) infusion over 30 to 120 minutes repeated every 28 days. Ninety-five courses of therapy at doses ranging from 18 to 1,104 mg/m2 were administered to 38 patients with refractory solid tumors. Granulocytopenia was dose limiting. Leukopenia was seen in 13 of 31 courses at doses of 690 mg/m2 or greater. Life-threatening granulocytopenia (less than or equal to 250 microliters) was noted in 1/6 patients treated at 800 mg/m2, 1/8 patients treated at 918 mg/m2, and 2/5 patients treated at 1,104 mg/m2. No definite relationship between myelotoxicity and prior treatment status was noted. Rate-of-infusion dependent, nonhematologic toxicities included diaphoresis, flushing, dizziness, and tinnitus, all of which were ameliorated by increasing the duration of drug infusion to 120 minutes. In addition, nausea and vomiting (grades 1 and 2) were seen in 29/56 courses at doses greater than or equal to 519 mg/m2, but were easily controlled by phenothiazine antiemetics. Amonafide plasma and urine concentrations were determined by high-pressure liquid chromatography (HPLC). Plasma concentrations declined biexponetially with a terminal harmonic mean terminal half-life (t 1/2) of 5.5 h. The mean apparent volume of distribution at steady-state and total body clearance were 532 L/m2 and 84 L/h/m2, respectively. Less than 5% of the total dose of amonafide was excreted unchanged in the urine. Antitumor activity has been noted in one patient with non-small-cell lung cancer (one complete response exceeding 29 months duration) and in one patient with prostatic cancer (complete pain relief and improvement in bone scan for 9 months). The recommended dose for phase II trials with this schedule of amonafide is 918 mg/m2 with dose escalation to amonafide is 918 mg/m2 with dose escalation to myelotoxicity.

High-dose chemotherapy with autologous hematopoietic stem-cell support for breast cancer in North America.

PURPOSE: To identify trends in high-dose therapy with autologous hematopoietic stem-cell support (autotransplants) for breast cancer (1989 to 1995). PATIENTS AND METHODS: Analysis of patients who received autotransplants and were reported to the Autologous Blood and Marrow Transplant Registry. Between January 1, 1989 and June 30, 1995, 19,291 autotransplants were reviewed; 5,886 were for breast cancer. Main outcomes were progression-free survival (PFS) and survival. RESULTS: Between 1989 and 1995, autotransplants for breast cancer increased sixfold. After 1992, breast cancer was the most common indication for autotransplant. Significant trends included increasing use for locally advanced rather than metastatic disease (P < .00001) and use of blood-derived rather than marrow-derived stem cells (P < .00001). One-hundred-day mortality decreased from 22% to 5% (P < .0001). Three-year PFS probabilities were 65% (95% confidence intervals [Cls], 59 to 71) for stage 2 disease, and 60% (95% Cl, 53 to 67) for stage 3 disease. In metastatic breast cancer, 3-year probabilities of PFS were 7% (95% Cl, 4 to 10) for women with no response to conventional dose chemotherapy; 13% (95% Cl, 9 to 17) for those with partial response; and 32% (95% Cl, 27 to 37) for those with complete response. Eleven percent of women with stage 2/3 disease and less than 1% of those with stage 4 disease participated in national cooperative group randomized trials. CONCLUSION: Autotransplants increasingly are used to treat breast cancer. One-hundred-day mortality has decreased substantially. Three-year survival is better in women with earlier stage disease and in those who respond to pretransplant chemotherapy.

[Pericardiocentesis in a critical care unit]. / Pericardiocentesis en una Unidad de Cuidados Intensivos.

OBJECTIVE: Study patients diagnosed of pericardial effusion and pericardiocentesis was made. We search the etiology, complications and the cnic's mortality. METHOD: 69 patients of Clinic Hospital of Zaragoza admitted in the critical care unit. RESULTS: We present 69 patients with 20-87 years old, 21 women and 48 men. The most frequent ethiology was neoplasic, 19 patients(27.5%), in most cases lung and breast cancer; 17 (24.6%) yatrogenic and 14 (20.3%) idiopathic. Diagnosis was made previously at ICU in 63.8% (44 patients). Pericardiocentesis was made in the 12 hours after admission in 40 cases (58%), and was superior than 500 cc in 37(53.6%). We registered pericardiocentesis complications in 7 (10.3%).16 patients dead (23%) most of them with pericardial effusion by mechanic cause. CONCLUSION: Was the pericardial effusion etiology has changed last years, the diagnosis is simply and minimum complications in evacuation, because the means and actual monitorization allow to make pericardiocentesis in optimum conditions.

The effect of bilayer order and fluidity on detergent-induced liposome fusion.

Surfactants induce fusion (or increase in size) of sonicated liposomes. This phenomenon is enhanced by cholesterol and inhibited by the intrinsic polypeptide gramicidin A. By comparison with previous physical studies we conclude that liposome 'fusion' is facilitated when both fluidity and static order of the bilayer are high.

Defining the role of novel high-dose chemotherapy regimens for the treatment of high-risk breast cancer.

We have explored several novel high-dose combinations in an attempt to increase antitumor activity while decreasing treatment-related toxicity. From October 1989 through June 1997, we performed phase I/II dose-escalation trials exploring novel high-dose regimens including ifosfamide/carboplatin/etoposide, mitoxantrone/thiotepa, and paclitaxel (Taxol; Bristol-Myers Squibb Company, Princeton, NJ)/mitoxantrone/thiotepa. We have also evaluated busulfan/cyclophosphamide and cyclophosphamide/thiotepa/carboplatin in phase II trials. Three hundred ninety-three patients have been treated in these trials and followed for a minimum of 3 months. Event-free survival (including relapses and treatment-related mortality; +/-SE) at 3 years by stage and chemosensitivity is as follows: stage II, four to nine positive nodes (n=16), 52%+/-17%; stage II, greater than nine nodes (n=30), 46%+/-11%; stage III (n=59), 50%+/-8%; inflammatory stage III (n=15), 27%+/-17%; stage IV, anthracycline responsive (n=69), 19%+/-5%; stage IV, anthracycline refractory but responsive to salvage therapy with ifosfamide, carboplatin, and etoposide or paclitaxel (n=53), 12%+/-6%; stage IV, refractory (n=128), 5%+/-2%; and stage IV, not evaluable for response (n=23), 10%+/-8%. Treatment-related mortality was 4% for both phase I and II studies involving stage II breast cancer patients, 5% for stage III breast cancer, 15% for inflammatory breast cancer, and 18% for all stage IV breast cancers, responsive and refractory. We conclude that high-dose therapy for the treatment of high-risk early stage breast cancer or metastatic breast cancer results in durable remissions. Chemosensitivity to induction regimens remains the most important prognostic indicator, although long-term survival has been seen even in patients with highly refractory disease. Further studies are necessary to define optimal high-dose strategies based on stage and chemosensitivity of disease.

Prevention of glutamate neurotoxicity in cultured neurons by 3,4-dihydro-6-hydroxy-7-methoxy-2,2-dimethyl-1(2H)-benzopyran (CR-6), a scavenger of nitric oxide.

Glutamate neurotoxicity in cerebellar neurons in culture is mediated by excessive production of nitric oxide (NO). We anticipated that 3,4-dihydro-6-hydroxy-7-methoxy-2,2-dimethyl-1(2H)-benzopyran (CR-6) could act as a scavenger of NO since it contains a position (C-5) highly activated towards nitration reaction. The aim of this work was to assess whether CR-6 acts as an NO scavenger and prevents glutamate neurotoxicity in cultures of cerebellar neurons. It was shown that CR-6 reduced, in a dose-dependent manner, glutamate-induced formation of cGMP (EC50 approximately 15 microM) and prevented glutamate neurotoxicity. The protection was approximately 50% at 3-10 microM and nearly complete at 100 microM. CR-6 did not prevent glutamate-induced activation of NO synthase, but interfered with the glutamate-NO-cGMP pathway at a later step. CR-6 reduced the formation of cGMP induced by S-nitroso-N-acetylpenicillamine (SNAP), an NO-generating agent, indicating that CR-6 acts as a scavenger of NO in cultured neurons. This was further supported by experiments showing that in neurons treated with CR-6 and glutamate, the 5-nitro derivative of CR-6 was formed, as determined by GC-MS analyses. Moreover, in vitro incubation of CR-6 with SNAP also produced the 5-nitroderivative, thus confirming that CR-6 directly reacts with NO. The results reported indicate that CR-6 acts as an NO scavenger in neurons and prevents glutamate neurotoxicity.