Selective gamma-ketoaldehyde scavengers protect Nav1.5 from oxidant-induced inactivation.

The cardiac sodium channel (SCN5A, Na(V)1.5) is a key determinant of electrical impulse conduction in cardiac tissue. Acute myocardial infarction leads to diminished sodium channel availability, both because of decreased channel expression and because of greater inactivation of channels already present. Myocardial infarction leads to significant increases in reactive oxygen species and their downstream effectors including lipoxidation products. The effects of reactive oxygen species on Na(V)1.5 function in whole hearts can be modeled in cultured myocytes, where oxidants shift the availability curve of I(Na) to hyperpolarized potentials, decreasing cardiac sodium current at the normal activation threshold. We recently examined potential mediators of the oxidant-induced inactivation and found that one specific lipoxidation product, the isoketals, recapitulated the effects of oxidant on sodium currents. Isoketals are highly reactive gamma-ketoaldehydes formed by the peroxidation of arachidonic acid that covalently modify the lysine residues of proteins. We now confirm that exposure to oxidants induces lipoxidative modification of Na(V)1.5 and that the selective isoketal scavengers block voltage-dependent changes in sodium current by the oxidant tert-butylhydroperoxide, both in cells heterologously expressing Na(V)1.5 and in a mouse cardiac myocyte cell line (HL-1). Thus, inhibition of this lipoxidative modification pathway is sufficient to protect the sodium channel from oxidant induced inactivation and suggests the potential use of isoketal scavengers as novel therapeutics to prevent arrhythmogenesis during myocardial infarction.

Novel SCN5A mutation leading either to isolated cardiac conduction defect or Brugada syndrome in a large French family.

BACKGROUND: The SCN5A gene encoding the human cardiac sodium channel alpha subunit plays a key role in cardiac electrophysiology. Mutations in SCN5A lead to a large spectrum of phenotypes, including long-QT syndrome, Brugada syndrome, and isolated progressive cardiac conduction defect (Lenègre disease). METHODS AND RESULTS: In the present study, we report the identification of a novel single SCN5A missense mutation causing either Brugada syndrome or an isolated cardiac conduction defect in the same family. A G-to-T mutation at position 4372 was identified by direct sequencing and was predicted to change a glycine for an arginine (G1406R) between the DIII-S5 and DIII-S6 domain of the sodium channel protein. Among 45 family members, 13 were carrying the G1406R SCN5A mutation. Four individuals from 2 family collateral branches showed typical Brugada phenotypes, including ST-segment elevation in the right precordial leads and right bundle branch block. One symptomatic patient with the Brugada phenotype required implantation of a cardioverter-defibrillator. Seven individuals from 3 other family collateral branches had isolated cardiac conduction defects but no Brugada phenotype. Three flecainide test were negative. One patient with an isolated cardiac conduction defect had an episode of syncope and required pacemaker implantation. An expression study of the G1406R-mutated SCN5A showed no detectable Na(+) current but normal protein trafficking. CONCLUSIONS: We conclude that the same mutation in the SCN5A gene can lead either to Brugada syndrome or to an isolated cardiac conduction defect. Our findings suggest that modifier gene(s) may influence the phenotypic consequences of a SCN5A mutation.

Gastrointestinal prokinetic drugs have different affinity for the human cardiac human ether-à-gogo K(+) channel.

Agonists of the serotonin 5-hydroxytryptamine 4 (5-HT4) receptor are widely used to activate motility in the gastrointestinal tract. Among these, cisapride was recently withdrawn from the U.S. market because of its proarrhythmic effects. Cisapride is a potent blocker of human ether-à-gogo (HERG) K(+) channels and prolongs the cardiac action potential in a reverse use dependence manner. We compared the effects of four different 5-HT4 receptor agonists (cisapride, prucalopride, renzapride and mosapride) on cloned HERG channels with the objective to evaluate and compare their proarrhythmic potential. K(+) currents from HERG-transfected COS-7 cells were recorded under physiological conditions using the whole cell configuration of the patch-clamp technique. Short (500 ms) depolarizing prepulses were used and following deactivating HERG currents were measured. Cisapride inhibited the HERG channels in a concentration-dependent manner with an IC(50) of 2.4 10(-7) M. The IC(50) value for prucalopride to block HERG (5.7 10(-6) M) was 20-fold higher than that of cisapride. Renzapride was slightly more potent than prucalopride (IC(50) = 1.8 10(-6) M). Mosapride produced no significant effects on the recombinant HERG current. The voltage dependence of HERG block was also investigated. The block mediated by cisapride or renzapride was voltage-dependent whereas that produced by prucalopride was not. We conclude that the rank order of potency of 5-HT4 agonists to block HERG is cisapride > renzapride > prucalopride > mosapride. We also conclude that 5-HT4 agonists devoid of side effects on the HERG current such as mosapride can be found as a safe alternative to cisapride.

Non-invasive testing of acquired long QT syndrome: evidence for multiple arrhythmogenic substrates.

BACKGROUND: Although well-defined clinically and electrocardiographically, Acquired Long QT Syndrome (LQTS) remains elusive from a pathophysiologic point of view. An increasingly accepted hypothesis is that it represents an attenuated form of Congenital Long QT Syndrome. To test this hypothesis further, we investigated patients with Acquired LQTS, using various investigations that are known to give information in patients with Congenital LQTS. METHODS: All the investigations were performed in patients with a history of Acquired Long QT Syndrome, defined by marked transient QT lengthening (QT>600 ms) and/or torsades de pointes. Measurement of the QT interval dispersion, the interlead difference for the QT interval on a 12-lead ECG, was performed in 18 patients and compared with 18 controls, matched for age and sex. To assess sympathetic myocardial innervation, I-123 Meta-iodobenzylguanidine (I-123-MIBG) scintigraphy was performed in 12 patients, together with Thallium scintigraphy, to rule out abnormal myocardial perfusion. Time-frequency analysis of a high-resolution ECG using a wavelet technique, was made for nine patients and compared with 38 healthy controls. Finally, genetic studies were performed prospectively in 16 consecutive patients, to look for HERG, KCNE1, KCNE2 and KCNQ1 mutations. The functional profile of a mutated HERG protein was performed using the patch-clamp technique. RESULTS: Compared with the control group, a significant increase in QT dispersion was observed in the patients with a history of Acquired LQTS (55+/-15 vs. 33+/-9 ms, P<0.001). In another group of patients with Acquired LQTS, 123 I-MIBG tomoscintigraphy demonstrated a decrease in the sympathetic myocardial innervation. Time--frequency analysis using wavelet transform, demonstrated an abnormal frequency content within the QRS complexes, in the patients with Acquired LQTS, similar to that found in Congenital LQTS patients. Molecular screening in 16 consecutive patients, identified one patient with a missense mutation on HERG, one of the LQTS genes. Expression of the mutated HERG protein led to altered K(+) channel function. CONCLUSION: Our results suggest that Acquired and Congenital Long QT Syndromes have some common features. They allow the mechanism of the clinical heterogeneity, found in both syndromes, to be understood. Further multi-facet approaches are needed to decipher the complex interplay between the main determinants of these arrhythmogenic diseases.

AKAP proteins anchor cAMP-dependent protein kinase to KvLQT1/IsK channel complex.

In cardiac myocytes, the slow component of the delayed rectifier K(+) current (I(Ks)) is regulated by cAMP. Elevated cAMP increases I(Ks) amplitude, slows its deactivation kinetics, and shifts its activation curve. At the molecular level, I(Ks) channels are composed of KvLQT1/IsK complexes. In a variety of mammalian heterologous expression systems maintained at physiological temperature, we explored cAMP regulation of recombinant KvLQT1/IsK complexes. In these systems, KvLQT1/IsK complexes were totally insensitive to cAMP regulation. cAMP regulation was not restored by coexpression with the dominant negative isoform of KvLQT1 or with the cystic fibrosis transmembrane regulator. In contrast, coexpression of the neuronal A kinase anchoring protein (AKAP)79, a fragment of a cardiac AKAP (mAKAP), or cardiac AKAP15/18 restored cAMP regulation of KvLQT1/IsK complexes inasmuch as cAMP stimulation increased the I(Ks) amplitude, increased its deactivation time constant, and negatively shifted its activation curve. However, in cells expressing an AKAP, the effects of cAMP stimulation on the I(Ks) amplitude remained modest compared with those previously reported in cardiac myocytes. The effects of cAMP stimulation were fully prevented by including the Ht31 peptide (a global disruptor of protein kinase A anchoring) in the intracellular medium. We concluded that cAMP regulation of I(Ks) requires protein kinase A anchoring by AKAPs, which therefore participate with the channel protein complex underlying I(Ks).

Reduced number of Langerhans cells in oesophageal mucosa from AIDS patients.

AIMS: The oesophageal mucosa is a frequent target of opportunistic infections in human immunodeficiency virus (HIV) infection. Langerhans cells (LC) are known as a target and reservoir of HIV in the skin. The aim of this study was to characterize oesophageal LC in HIV-infected patients. METHODS AND RESULTS: Thirty oesophageal biopsies were obtained from 29 patients (median age 35.5), all in stage IV of the HIV Center of Disease Control Classification. We performed histological assessment of the oesophageal mucosa and immunohistochemical detection of oesophageal LC using an anti-CD1a antibody, followed by morphometric analysis. Biopsies from 17 noninfected patients were studied using the same procedure. LC in oesophageal mucosa of the HIV positive patients showed a significantly and dramatically decreased number (LC(N) median = 5.85/mm2) and surface/epithelial surface (LC (S) ratio = 0.09) when compared with HIV-negative controls (LC(N) median = 29.7/mm2, LC(S) ratio = 1.83) with P = 0.003 for LC(N) and P < 0.0001 for LC(S). CONCLUSION: These data suggest that oesophageal LC are, like their epidermal counterparts, a preferential target for HIV infection. Their alterations may provide a clue to the pathogenesis of the decreased local oesophageal immunity and to the occurrence of opportunistic infections.

A diffuse T lymphocytic gastrointestinal mucosal infiltration associated with Sjögren's syndrome resulting in a watery diarrhea syndrome and responsive to immunosuppressive therapy.

We report the case of a 45-yr-old white man, investigated for chronic diarrhea, malabsorption and weight loss associated with sicca syndrome. Endoscopic and x-ray examinations showed normal macroscopic mucosa in gastrointestinal tract (GIT). Immunohistochemistry showed diffuse polyclonal T cell lymphocytes infiltrating either epithelium and lamina propria in GIT. There was no villous atrophy in the jejunum and ileum. Corticosteroids, azathioprine, and cyclosporine failed to improve symptoms. Monthly intravenous cyclophosphamide administered over 1 yr, stopped the diarrhea and weight loss. The patient is free of symptoms up to a 5-yr follow-up.

[Paraneoplastic intestinal pseudo-obstruction revealing small cell lung carcinoma: "the anti-Hu syndrome"]. / Pseudo-obstruction intestinale paranéoplasique révélatrice d'un carcinome pulmonaire à petites cellules: le syndrome anti-Hu.

A 67-year-old woman was admitted for intestinal pseudoobstruction associated with peripheral sensitive neuropathy. Jejunal biopsies performed during laparotomy, showed axonal degeneration and lympho-plasmocytic infiltration in myenteric plexus. High titer of seric anti-Hu antibodies suggested a paraneoplastic syndrome. Thoracic CT scan showed mediastinal lymph nodes. Their histological examination confirmed the diagnosis of metastatic small-cell lung carcinoma. Her condition gradually deteriorated despite supportive parenteral nutrition, chemotherapy, steroids and intravenous immunoglobulins. She died 12 months after the onset of symptoms.

Ticlopidine induced colitis: a histopathological study including apoptosis.

AIMS: To describe ticlopidine related microscopic colitis and to assess the occurrence of apoptosis in the colon epithelium. METHODS: A series of colorectal biopsy samples from nine patients with ticlopidine related chronic diarrhoea were analysed. Biopsies were also taken from five of these patients between two and four months after ticlopidine withdrawal. The number of apoptotic cells in the crypts/mm2 (apoptotic index) was calculated using in situ labelling by terminal deoxyribonucleotidyl transferase (TdT) mediated dUTP-biotin nick end labelling (TUNEL). All specimens were matched to normal colorectal specimens from a control group of comparable age and sex distribution. RESULTS: Histological examination of the colon biopsy specimens taken from all nine patients with ticlopidine related chronic diarrhoea showed characteristic features of microscopic colitis. The histology returned to normal when ticlopidine was withdrawn. Apoptotic cells were rarely found in controls, and the mean apoptotic index was 0.53. The apoptotic index was significantly higher (16.53) in ticlopidine related colitis, but decreased dramatically to control value when ticlopidine was withdrawn. CONCLUSION: Microscopic colitis can be induced by ticlopidine and is accompanied by an increase in epithelial apoptosis. Hence, increased apoptosis might be related to drug injury or might be part of microscopic colitis.

Analysis of CD44 expression in serous and mucinous borderline tumours of the ovary: comparison with cystadenomas and overt carcinomas.

AIMS: To determine the diagnostic and prognostic value of the immunohistochemical analysis of CD44 variants in benign borderline and malignant tumours of the ovary. METHODS AND RESULTS: The reactivity of tumour cells with three monoclonal antibodies, respectively, directed to all CD44 variants, CD44-v3 isoform and CD44-v6 isoform, was assessed by using an indirect immunoperoxidase technique applied to formalin-fixed, paraffin-embedded samples of 36 cases of borderline, as compared to 20 cases of benign tumours and 20 cases of carcinomas. CD44 variants were detected in 97% of borderline tumours, as compared to 60% of benign tumours and 100% of carcinomas. CD44-v3 was detected in 25% of borderline tumours, as compared to 0% of benign tumours (P = 0.003) and 55% of carcinomas (P = 0.065). The expression of CD44-v6 was detected in 28% of borderline tumours, as compared to 20% of benign tumours and 30% of carcinomas. In borderline tumours, as in carcinomas, CD44-v6, but not CD44-v3, expression was correlated with an increased proliferative index and with a higher incidence of p53 expression. CONCLUSION: Borderline tumours of the ovary present frequent quantitative and qualitative alterations in the pattern of expression of CD44 proteins. However, these alterations are unlikely to represent useful diagnostic or prognostic markers.

In situ Rantes and interferon-gamma gene expression in pediatric small bowel Crohn's disease.

BACKGROUND: Rantes (regulated upon activation, normal T cell expressed and secreted) is a chemotactic cytokine for memory T lymphocytes, monocytes, and eosinophils. The cytokine interferon-gamma (IFN-gamma) plays a key in the immune response. Their distributions and possible roles in the selective accumulation of inflammatory cells in Crohn's disease (CD) were examined by determining the expression of Rantes and IFN-gamma genes in patients with CD using in situ hybridization (ISH) on frozen and paraffin-embedded tissue sections. METHODS: Intestinal and mesenteric lymph node samples from 9 children who had undergone ileal resection for CD were examined for the presence of epithelioid-giant cell granulomas (EGCG) and Rantes and IFN-gamma messenger RNA (mRNA). Normal pediatric intestine (n = 5) and lymph nodes (n = 2) served as controls. RESULTS: Many cells in all CD specimens in the epithelial compartment, lamina propria, and the EGCG gave positive signal with the Rantes antisense probe. Labelled cells were identified on paraffin sections as lymphocytes, macrophages, and epithelioid cells. There were Rantes-positive cells in the control intestinal tissues, but many Rantes-positive cells in control lymph nodes that showed follicular hyperplasia. IFN-gamma-positive cells were present in all CD ileal and lymph node specimens, predominantly in close contact with EGCC. No positive signal was obtained with the Rantes and IFN-gamma sense control probes. CONCLUSIONS: These findings suggest that Rantes and IFN-gamma contribute to the selective accumulation of macrophages and memory T helper lymphocytes inside the granulomas and inflammatory infiltrates that are characteristic of CD.

Expression of cadherins in benign, borderline, and malignant ovarian epithelial tumors: a clinicopathologic study of 60 cases.

We have analyzed the expression of E- and N-cadherins in benign, borderline, and maligant ovarian tumors, and we have correlated the pattern of cadherin expression with the standard clinicopathological parameters. An immunohistochemical technique has been applied to formalin-fixed, paraffin-embedded samples of 20 benign cystic tumors, 20 borderline tumors, and 20 cancers. Expression of E- and N-cadherins immunostaining were compared with the histological type, degree of histological differentiation, International Federation of Gynecology and Obstetrics (FIGO) stage, presence of ascites, occurrence of recurrence, and survival. E-cadherin was homogeneosuly expressed in benign tumors but was heterogeneously expressed or undetectable in most borderline and malignant tumors. In contrast, N-cadherin was detected in most benign and borderline tumors but was absent or heterogeneous in most carcinomas. The difference of expression of E-cadherin and N-cadherin between the three groups of ovarian tumors was statistically significant (respectively, P = .03 and P < .001). In ovarian carcinoma, patients with negative E-cadherin staining present a significantly shorter survival. No correlation was found between cadherin expression and clinicopathological parameters in borderline tumors. Our results suggest that alterations in E-cadherin and N-cadherin expressions are differentially involved in ovarian carcinogenesis and may have diagnostic and prognostic values.

[Hyperplastic polyps arising in islands of gastric mucosa in the duodenum]. / Polypes hyperplasiques développés sur des îlots de muqueuse gastrique dans le duodénum.

Hyperplastic polyps originating from islands of gastric mucosa can be observed in the duodenum. We report 3 cases of duodenal hyperplastic polyps on antral or fully developed fundic mucosa. They macroscopically presented as small confluent polyps or as large villous polyps. Their microscopic features were the same as those observed in hyperplastic gastric polyps. This type of polyp must be added to the list of tumor like lesions of the duodenum that may endoscopically present as polyps.

[Are lymphocytic and collagenous colitis two forms of a single disease? Arguments taken from a biopsy quantitative study]. / La colite lymphocytaire et la colite collagène sont-elles une seule et même maladie? Arguments tirés d'une étude quantitative sur biopsie.

Collagenous colitis and lymphocytic colitis are defined by a clinicopathologic syndrome with chronic watery diarrhea, microscopic lesions of colonic biopsies, and normal barium enema and colonoscopy. A histopathological study was performed on multiple colorectal biopsies to compare 12 cases of collagenous colitis (defined by a subepithelial collagen thicker than 10 microns) and 7 cases of lymphocytic colitis (defined by a number of intraepithelial lymphocytes more than 20 per 100 epithelial cells at least in one biopsied site). The study included a semiquantitative analysis of inflammatory infiltrate in the lamina propria, crypts distortion and epithelial detachment. The number of intraepithelial lymphocytes per 100 epithelial cells was determined in surface epithelium and crypts. The subepithelial collagen thickening was studied by computerised morphometry. The intraepithelial lymphocytes, villous atrophy and thickness of the subepithelial collagen were also determined in gastric and duodenal biopsies. In collagenous colitis, the subepithelial collagenous thickness ranged from 10 to 40 microns in the colon (median 20.99 microns). In 4 cases of collagenous colitis, no thickening of the collagen plate was seen in the rectum. We found constant epithelial detachment and mucosal distortion. In lymphocytic colitis, the thickness of the subepithelial collagen ranged from 6 to 10 microns in 4 cases and was less than 6 microns in 3 cases (median 6.24 microns). The median number of intraepithelial lymphocytes in surface epithelium was 22.35 (range 18.2 to 40) in lymphocytic colitis versus 12.22 (range 4.6 to 24.4) in collagenous colitis. In conclusion, we observed an overlap of both the collagenous plate thickness and the number of intraepithelial lymphocytes in collagenous colitis and lymphocytic colitis. This result favours a unified histogenesis for these two entities.

Increased expression of CD44v6 in endocrine pancreatic tumours but not in midgut carcinoid tumours.

Aims/background-To analyse the different isoforms of CD44 in various types of endocrine pancreatic and gut carcinoid tumours and to investigate the relation between their expression and tumour dissemination. This study was prompted by the recent observation that inappropriate splicing of the CD44 gene was correlated with tumour progression and metastasis formation in a number of human cancers.Methods-Expression of CD44 isoforms was studied in 38 endocrine pancreatic tumours and gut neuroendocrine tumours using antibodies directed against products of exons v3, v4-v5, v6, v7-v8 as well as against the standard CD44 molecule (CD44H). CD44 gene expression was also analysed by reverse transcription PCR (RT-PCR) in nine endocrine and seven carcinoid tumours.Results-All gastrinomas except one (nine of 10) and about half of the other endocrine pancreatic tumours (seven of 15) expressed CD44v6. Most (10/11) midgut carcinoid tumours were CD44v6 negative, with no detectable immunostaining. CD44v3, CD44v4-v5 and CD44v7-v8 were not expressed in any of these tumours. CD44 mRNA analysis illustrated a complex splice pattern and expression of large CD44 isoforms in CD44v6 positive endocrine tumours, whereas the standard form only was detected in midgut carcinoid tumours. No correlation between CD44 variant expression and tumour metastasis was observed.Conclusions-CD44 variants encoding exon v6 are preferentially expressed both in gastrinomas and in most pancreatic endocrine tumours. In contrast to other tumours, the expression of CD44v6 in pancreatic neuroendocrine tumours does not seem to be correlated with tumour dissemination.