The pattern of cardiovascular alterations induced by infusion of platelet-activating factor in rabbit is modified by pretreatment with H1-H2 receptor antagonists but not by cyclooxygenase inhibition

The intravenous infusion of platelet activating factor (PAF) (0.8 g/kg b.w.) induced ECG and hemodynamic alterations characterized by the following sequential three phases. Phase I (15 sec) consisted of a transient bradycardia with reduction in left ventricular pressure (LVPs), mean arterial pressure (MAP) and cardiac output (CO). Phase II developed within 30 sec and consisted of a rise in cardiac frequency, increase in LVPs, MAP and total peripheral resistances (TPR), which were associated with a decrease in CO. Finally, phase III, that occurred about 90 sec after PAF infusion, was characterized by marked ECG changes (ST segment depression and conduction arrhythmias), a decrease in LVPs and MAP, as well as a rise in TPR and in right atrial pressure (RAP). All these alterations were reversible within 30–60 min. Pretreatment with promethazine and cimetidine, as H₁ and H₂ histamine receptor antagonists, markedly prevented the development of phase II, namely the rise in cardiac frequency, LVPs, MAP and TPR, but did not significantly modify phase I and III. In contrast, pretreatment with indomethacin, an inhibitor of cyclooxygenase, moderatively attenuated, but did not abolish, the three phases of cardiovascular changes induced by PAF infusion.     

Role of double-stranded RNA and Npro of classical swine fever virus in the activation of monocyte-derived dendritic cells

Classical swine fever virus (CSFV) is a noncytopathogenic (ncp) positive-sense RNA virus that replicates in myeloid cells including macrophages and dendritic cells (DC). The virus does not induce type I interferon (IFN-alpha/beta), which in macrophages has been related to the presence of the viral Npro gene. In the present work, the role of viral double-stranded (ds)RNA and Npro in the virus-host cell interaction has been analyzed. Higher levels of detectable dsRNA were produced by a genetically engineered cytopathogenic (cp) CSFV compared with ncp CSFV, and cp CSFV induced IFN-alpha/beta in PK-15 cells. With DC, there was only a small difference in the levels of dsRNA between the cp and ncp viruses, and no IFN-alpha/beta was produced. However, the cp virus induced a higher degree of DC maturation, in terms of CD80/86 and MHC II expression. Npro deletion mutants induced an increase in DC maturation and IFN-alpha/beta production-for both ncp and cp viruses-despite reduced replication efficiency in the DC. Deletion of Npro did not influence dsRNA levels, indicating that the interference was downstream of dsRNA turnover regulation. In conclusion, the capacity of CSFV to replicate in myeloid DC, and prevent IFN-alpha/beta induction and DC maturation, requires both regulated dsRNA levels and the presence of viral Npro.     

Influences of hormone replacement therapy on postmenopausal women's health perceptions

OBJECTIVE: To assess the beliefs of climacteric women regarding their health, menopause, and hormone replacement therapy (HRT). DESIGN: Medical students asked to interview 526 healthy women, ranging from 40 to 64 years of age, between January and February of 2002. Of that number, 26 (4.9%) declined to participate in the interview. Thus, 500 women were interviewed about their beliefs and perceptions regarding their quality of life and health risks, as well as their opinions on menopause and HRT. RESULTS: The mean age of the sample was 53.3 +/- 6.2 years; 83.4% were postmenopausal, and 18.8% were HRT users. Of the women interviewed, 38.6% believed that their health was good. Although 78.8% thought that cancer is the main cause of death, 64% of them considered themselves to be at high risk for cardiovascular disease and osteoporosis. Most (64%) believed that menopause deteriorates the quality of life and that it increases cardiovascular risk (52.4%) and osteoporosis (72.0%). The HRT users perceived that they had better health status (48.9% v 36.2%, P < 0.02) and smaller cardiovascular risk (54.3% v 66.3%, P < 0.04) than did the nonusers; however, they ignored the preventive effect of estrogens in osteoporosis. CONCLUSIONS: Women believe that menopause deteriorates their health. The HRT users perceived themselves to be healthier and to have a smaller risk for cardiovascular disease.     

Vasopressin-induced taurine efflux from rat pituicytes: a potential negative feedback for hormone secretion

Previous work on the whole neurohypophysis has shown that hypotonic conditions increase release of taurine from neurohypophysial astrocytes (pituicytes). The present work confirms that taurine is present in cultured pituicytes, and that its specific release increases in response to a hypotonic shock. We next show that vasopressin (VP) and oxytocin (OT) also specifically release taurine from pituicytes. With an EC₅₀ of ∼2 n m , VP is much more potent than OT, and the effects of both hormones are blocked by SR 49059, a V_1a receptor antagonist. This pharmacological profile matches the one for VP- and OT-evoked calcium signals in pituicytes, consistent with the fact that VP-induced taurine efflux is blocked by BAPTA-AM. However, BAPTA-AM also blocks the taurine efflux induced by a 270 mosmol l⁻¹ challenge, which per se does not evoke any calcium signal, suggesting a permissive role for calcium in this case. Nevertheless, the fact that structurally unrelated calcium-mobilizing agents and ionomycin are able to induce taurine efflux suggests that calcium may also play a signalling role in this event. It is widely accepted that in hypotonic conditions taurine exits cells through anionic channels. Antagonism by the chloride channel inhibitors 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) and 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) suggests the same pathway for VP-induced taurine efflux, which is also blocked in hypertonic conditions (330 mosmol l⁻¹). Moreover, it is likely that the osmosensitivity of the taurine channel is up-regulated by calcium. These results, together with our in situ experiments showing stimulation of taurine release by endogenous VP, strengthen the concept of a glial control of neurohormone output.     

A comparison of murine and human immunoproteomes of Helicobacter pylori validates the preclinical murine infection model for antigen screening

Preclinical mouse infection models are widely used for Helicobacter vaccine development, but how well such models mimic important aspects of human infections is unknown. A comparison of Helicobacter pylori immunoproteomes of infected mice with previously reported patient data reveals a high agreement in the antigens recognized, suggesting that H. pylori in vivo protein composition and recognition by the host immune system are comparable in mice and humans. Murine Helicobacter models may thus be valid to screen antigens for human vaccination.     

Presence of autoantibodies to apolipoprotein A-1 in patients with acute coronary syndrome further links autoimmunity to cardiovascular disease

Anti-apolipoprotein A-1 (Apo A-1) autoantibodies were described in autoimmune disorders such as systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS) and might be involved in the genesis of arterial and venous thrombotic events. To investigate the presence of these autoantibodies in patients with acute coronary syndrome (ACS) without other features of autoimmunity, we set up an enzyme-linked immunosorbent assay (ELISA) for anti-Apo A-1 antibodies. We used it to investigate their prevalence in ACS as compared to SLE and APS and correlated them to plasma Apo A-1 and serum amyloid A protein (SAA) concentrations. The prevalence of anti-Apo A-1 autoantibodies in the healthy control group was 1% (1/92), but was significantly higher in other groups: 21% (11/53) in ACS group (P=0.001), 13% (12/92) in SLE and/or APS group (P=0.005). Multiple linear regression revealed a significant correlation between plasma Apo A-1 (r=-0.72, P=0.013), plasma SAA concentration (r=0.76, P=0.0066) and anti-Apo A-1 IgG titre in ACS patients. The presence of anti-Apo A-1 autoantibodies in patients with ACS highlights an additional link between autoimmunity, inflammation and atherosclerosis.     

Molecular interaction of connexin 30.3 and connexin 31 suggests a dominant-negative mechanism associated with erythrokeratodermia variabilis

Connexins are homologous four-transmembrane-domain proteins and major components of gap junctions. We recently identified mutations in either GJB3 or GJB4 genes, encoding respectively connexin 31 (Cx31) or 30.3 (Cx30.3), as causally involved in erythrokeratodermia variabilis (EKV), a mostly autosomal dominant disorder of keratinization. Despite slight differences, phenotypes of EKV Mendes Da Costa (Cx31) and EKV Cram-Mevorah (Cx30.3) show major clinical overlap and both Cx30.3 and Cx31 are expressed in the upper epidermal layers. These similarities suggested to us that Cx30.3 and Cx31 may interact at a molecular level. Indeed, expression of wild-type Cx30.3 in HeLa cell resulted only in minor amounts of protein addressed to the plasma membrane. Mutant Cx30.3 was hardly detectable and disturbed intercellular coupling. In sharp contrast, co-expression of both wild-type proteins led to a gigantic increase of stabilized heteromeric gap junctions. Furthermore, co-expressed wild-type Cx30.3 and Cx31 coprecipitate, which demonstrates a physical interaction. Inhibitor experiments revealed that this interaction begins in the endoplasmic reticulum. These results not only provide new insights into epidermal connexin synthesis and polymerization, but also allow a novel molecular explanation for the similarity of EKV phenotypes.     

Dysferlin mutations in LGMD2B, Miyoshi myopathy, and atypical dysferlinopathies

DYSF encoding dysferlin is mutated in Miyoshi myopathy and Limb-Girdle Muscular Dystrophy type 2B, the two main phenotypes recognized in dysferlinopathies. Dysferlin deficiency in muscle is the most relevant feature for the diagnosis of dysferlinopathy and prompts the search for mutations in DYSF. DYSF, located on chromosome 2p13, contains 55 coding exons and spans 150 kb of genomic DNA. We performed a genomic analysis of the DYSF coding sequence in 34 unrelated patients from various ethnic origins. All patients showed an absence or drastic decrease of dysferlin expression in muscle. A primary screening of DYSF using SSCP or dHPLC of PCR products of each of 55 exons of the gene was followed by sequencing whenever a sequence variation was detected. All together, 54 sequence variations were identified in DYSF, 50 of which predicting either a truncated protein or one amino-acid substitution and most of them (34 out of 54) being novel. In 23 patients, we identified two pathogenic mutations, while only one was identified in 11 patients. These mutations were widely spread in the coding sequence of the gene without any mutational "hotspot."     

Formalin-fixed and paraffin-embedded nodal non-Hodgkin's lymphomas demonstrate the same chromosome changes as those found in frozen samples: a comparative study using interphase fluorescence in situ hybridization.

Cytogenetic studies in lymphomas classically require fresh or frozen tissue, whereas in many instances only paraffin-embedded biopsies are available. We applied an interphase FISH assay on nuclei extracted from thick paraffin sections to determine accuracy of molecular cytogenetics in such samples. Twenty-three lymphoma samples and 4 reactive lymph nodes were tested with various commercially available DNA probes, and hybridization patterns were compared with those obtained on frozen nuclei counterparts. Successful hybridization with all probes tested was observed for 23/27 (85%) paraffin-embedded tissues and for all (100%) frozen samples, and cut-off levels defining positivity were superimposable for both situations. Chromosome changes were detected in the same way, without any false-positive or false-negative cases. Hybridization signals observed on dewaxed samples were either those classically expected to define the relevant chromosome change or were atypical: all atypical changes could be demonstrated also into nuclei from the frozen counterpart. Moreover, all typical and atypical chromosome changes observed on frozen nuclei were also detected in paraffin-embedded tissues. Our study shows that our interphase FISH assay performed on paraffin-embedded samples is a valuable alternate to conventional methods to ascertain diagnosis of lymphomas as to include patients into therapeutic trials.