Prevalence and significance of CYP2C19*2 and CYP2C19*17 alleles in a New Zealand acute coronary syndrome population.

BACKGROUND: High on-treatment platelet reactivity has been associated with poor outcomes following acute coronary syndromes (ACS). Both the loss of function CYP2C19*2 allele and the gain of function CYP2C19*17 allele along with a range of clinical characteristics have been associated with variation in the response to clopidogrel. AIM: The study aims to examine the frequency of CYP2C19 variants and understand the factors associated with on-treatment platelet reactivity in a New Zealand ACS population. METHODS: We prospectively enrolled 312 ACS patients. We collected clinical characteristics and measured on-treatment platelet reactivity using two validated point-of-care assays, VerifyNow and Multiplate. DNA was extracted and CYP2C19*2 and *17 alleles were identified using real-time polymerase chain reaction. RESULTS: CYP2C19*2 or CYP2C19*17 alleles were observed in 101 (32%) and 106 (34%) of patients, respectively, with significant differences in distribution by ethnicity. In Maori and Pacific Island patients, 47% (confidence interval (CI) 31-63%) had CYP2C19*2 and 11% (CI 4-19%) CYP2C19*17 compared with 26% (CI 19-32%) and 41% (CI 32-49%) in white people. Carriage of CYP2C19*2 alleles was associated with higher levels of platelet reactivity measured by either assay, but we observed no relationship between platelet reactivity and CYP2C19*17. In multivariate analysis diabetes, clopidogrel dose and CYP2C19*2 status were all significant independent predictors of platelet reactivity. CONCLUSIONS: Both CYP2C19*2 and *17 were common in a New Zealand ACS population, with CYP2C19*2 observed in almost half the Maori and Pacific Island patients. CYP2C19*2, diabetes and clopidogrel dose were independent contributors to on-treatment platelet reactivity.

Differential patterns of liver proteins in experimental murine hepatosplenic schistosomiasis.

Schistosoma mansoni eggs produced by adult worms in the mesenteric vasculature become trapped in the liver, where they induce granulomatous lesions and strong immune responses. Infected individuals suffer from intestinal schistosomiasis (INT) in 90% of cases, whereas the remaining 10% present with severe hepatosplenic schistosomiasis (HS). The CBA/J mouse model mimics human disease, with 20% of infected mice developing hypersplenomegaly syndrome (HSS) that resembles HS and 80% developing moderate splenomegaly syndrome (MSS) similar to INT. We studied differential patterns of protein expression in livers of 20-week-infected CBA/J mice with MSS or HSS to understand the molecular changes that underlie these two disease forms. Using differential in-gel electrophoresis to identify differentially expressed protein spots, we found 80 protein spots significantly changed with infection and 35 changes specific to severe disease. In particular, the abundances of prohibitin 2, transferrin isoforms, and major urinary protein isoforms were significantly altered in HSS mice. Furthermore, annexin 5, glutathione S-transferase pi class, and S. mansoni phosphoenolpyruvate carboxykinase expression levels changed significantly with schistosome infection. Additionally, levels of major urinary protein decreased and levels of transferrin increased significantly in the sera of HSS mice compared to levels in sera of MSS or control mice, and these differences correlated to the degree of splenomegaly. These findings indicate that the liver protein abundances differ between MSS and HSS mice and may be used for the development of diagnostic markers for the early detection of hepatosplenic schistosomiasis.

Prasugrel inhibits platelet-enhanced pro-inflammatory CD4+ T cell responses in humans.

While platelets have well characterized effects on monocytes, the effect of platelet activation on CD4+ T-cell differentiation and cytokine production is not clear. To examine the effects of platelet T-cell interactions on T-cell phenotype, and whether these interactions were altered by prasugrel, we conducted a randomized, double-blind, placebo-controlled crossover study in healthy subjects. At baseline the addition of platelets to CD4+ T-cells resulted in an increase in the release of pro-inflammatory cytokine IFN-γ (192% increase in IFN-γ levels, p = 0.01) and pro-inflammatory CD4+ phenotypes, (38% and 58% increase in Th1 and Th17 phenotypic markers respectively, p = 0.01) but no change in Tregs. Prasugrel abolished the effects of platelets on CD4+ T-cells with similar levels of pro-inflammatory cytokines and cell numbers to T-cells stimulated. Antiplatelet therapy may provide therapeutic benefit both from direct platelet inhibition and also through indirect effects on immune response development.

Expression of Trypanosoma cruzi surface antigen FL-160 is controlled by elements in the 3' untranslated, the 3' intergenic, and the coding regions.

The FL-160 surface antigen gene family of T. cruzi consists of hundreds of members of 160 kDa glycoproteins expressed in trypomastigotes, but not in epimastigotes. Steady-state levels of FL-160 mRNA were 80 to 100-fold higher in trypomastigotes than in epimastigotes, yet transcription rates were equivalent between the lifecycle stages. Luciferase reporter constructs demonstrated that the 3' untranslated region (UTR) and intergenic region (IR) following the coding sequence of FL-160 was sufficient to generate 8-fold higher luciferase expression in trypomastigotes compared with epimastigotes. Transfection of 3' UTR/IR deletion constructs revealed cis-acting elements which conferred a trypomastigote-specific expression pattern similar to that of FL-160. Parasites treated with translation and transcription inhibitors, cyclohexamide and Actinomycin D, respectively, displayed a stage-specific pattern of FL-160 mRNA degradation. Epimastigotes, but not trypomastigotes, treated with the inhibitors accumulated a 1.4 Kb FL-160 cleavage product. The cleavage site mapped to a 31 base poly-purine tract in the FL-160 coding region. The first 526 aa of FL-160, containing the 31 base poly-purine tract and several smaller tracts, were fused to green fluorescent protein (GFP) and expressed from the T. cruzi tubulin locus. Stable transformants expressed 4-fold more FL-160:GFP fusion mRNA and 12-fold more fusion protein in the trypomastigote stage than in the epimastigote stage suggesting post-transcriptional and translational control elements. These data reveal at least two distinct control mechanisms for trypomastigote-specific expression of FL-160 surface glycoproteins, one involving the 3' UTR/IR and one involving the coding region of FL-160.

Expression of mammalian cytokines by Trypanosoma cruzi indicates unique signal sequence requirements and processing.

A vector based upon the calmodulin-ubiquitin 2.65 locus of Trypanosoma cruzi has enabled the expression and secretion of the murine cytokines interleukin-2 (IL-2) and gamma-interferon (gamma-IFN) by transfected T. cruzi. The T. cruzi-derived cytokines were bioactive and produced by both epimastigotes and mammalian forms. The native coding sequence of IL-2 was sufficient to cause secretion of the protein, but the gamma-IFN signal sequence had to be replaced by the IL-2 signal sequence (IL-2/gamma-IFN) to allow efficient secretion of gamma-IFN. The amino acid sequences at the N-termini of the secreted T. cruzi-derived cytokines were different from the expected murine secreted protein. The secreted IL-2 was cleaved six amino acids downstream from the murine signal sequence cleavage site, and the hybrid IL-2/gamma-IFN molecule was cleaved three amino acids downstream from the predicted signal cleavage site in the IL-2/gamma-IFN molecule. These apparent differences in signal peptide sequence requirements and cleavage sites most likely indicate that the signal sequence processing in trypanosomes is distinct from that of higher eukaryotes.

The initiation and function of Th2 responses during infection with Schistosoma mansoni.

Through experiments in which pro-inflammatory mediators have been neutralized in schistosomeinfected WT and IL-4 -/- mice we have been investigating the nature of the immune response that is required to allow survival during the period of acute disease that accompanies the onset of egg production by the parasitic worms. The developing picture is that of an early pro-inflammatory type-1 like response (characterized by the production of IFN-gamma and TNF-alpha and NO) which in the short term is able to control the deleterious effects on the liver associated with the arrival of the eggs, followed rapidly by a potent egg-induced Th2 response which simultaneously subsumes the role of protecting the liver and downregulates the production of the inflammatory mediators, thereby averting the serious consequence associated with the continued high level production of NO. Current research is directed towards: 1) understanding how pro-inflammatory mediators are protective during schistosomiasis, and 2) elucidating the underlying mechanisms through which the Th response is biased in a Th2 direction following exposure to schistosome eggs.

Muscle-saving lateral axillary thoracotomy.

Seventy-five consecutive thoracotomies through a lateral axillary thoracotomy incision are reviewed. The limited approach is a muscle-splitting incision with preservation of the Latissimus Dorsi and Pectoralis Major muscles and splitting of the Serratus Anterior muscle. Detailed description of the operative technique is given, and a review of morbidity and mortality is included. We conclude that the lateral axillary incision is a good alternative to the standard postero-lateral approach, as it provides excellent visibility and allows for all pulmonary surgical procedures, with minimal postoperative discomfort.

[The treatment of gastroesophageal reflux using the Kaunitz technic]. / De behandeling van gastro-oesofagiale reflux bij middel van de Kaunitz techniek.

The Kaunitz' procedure in the surgical treatment of gastroesophageal reflux associated with a hiatal hernia. In patients with pathological gastroesophageal reflux associated with a hiatal hernia, simple repositioning of the lower esophageal sphincter (L.E.S.) without fundoplication can restore the competence of the L.E.S., provided there is adequate mobilisation of the esophagus and fixation to strong tissues. The procedure proposed by Kaunitz consists of a left thoracotomy, with fixation of the gastroesophageal junction to the diaphragm. In the period May 74-May 83 we performed the procedure on 85 patients. There was no operative mortality and a low early complication rate. All but one patient showed a good repositioning on the postoperative barium swallow and improved dramatically, with complete cessation of reflux symptoms. Seventy-eight patients (92,8%) are followed up regularly by means of a standard questionnaire, barium swallow and esophagoscopy. At this stage of the study, with follow-up periods ranging from 6 months to 9 years, 65 patients (83,3%) show an excellent result, six (7,7%) are greatly improved, with disappearance of the preoperatively existing esophagitis, but keep some mild residual reflux symptoms. In one patient repositioning was impossible. Six recurrences were noted, four of them in patients operated during the early period of our experience with the Kaunitz technique. This simple and effective procedure is a valuable alternative in the surgical treatment of pathological gastroesophageal reflux.

Technical note: the use of arterialized arm veins in infrainguinal bypass.

In order to augment the vein donation area of arm veins for infrainguinal bypass, we performed an arteriovenous AV fistula prior to the revascularization procedure. This technique was used in 5 patients. The diameter enlargement of the vein obtained by this technique enabled us to construct completely autologous grafts with gratifying results.

[Surgery of acute cholecystitis in patients 80 years of age or older]. / Chirurgie de la cholécystite aiguë au-delà de 80 ans.

In 17 Belgian surgical centers, 324 operations have been performed for acute cholecystitis in patients 80 years of age or older. In this group of patients: 1. The incidence of acute cholecystitis is hardly higher in women than in men. 2. The patients were operated within 24 hours of admission in 38% of cases. The main bile duct was surgically explored in 1/3 of patients. Cholecystostomy or simple drainage were used in 1/10 of cases only. 3. Operative mortality was 19%. It was higher for patients operated as emergencies than for delayed operations and higher also when the cholecystectomy was associated with an exploration of the bile duct. The ideal treatment for such patients should thus be a cholecystectomy with an endoscopic sphincterotomy.

[Surgery in the octogenarian (introduction). Results of a national survey]. / Chirurgie de l'octogénaire (introduction). Résultats d'une enquête nationale.

Presentation of the results of a national, multicentric and retrospective study regarding the surgery of the octogenarian. The series counts 7,407 patients accepted for surgery, and which can be divided into 4,581 women (61.8%) and 2,826 men (38.2%) of an average age of 84.4 years old (extremities 80-101 years old). The global mortality of this population of elderly (whether operated on or not) amounts to 16.7%. The postoperative mortality of the 4,177 operated patients is 18.2% with different rates according to the surgical disciplines: 34.7% for surgery of the colon, 23.2% for bone surgery, 22.4% for vascular surgery, 15.3% for biliary surgery, 8.7% for urologic surgery and 5.6% for cardiac surgery.

[Implantation of a cardiac pacemaker in the octogenarian]. / Implantation de stimulateur cardiaque chez l'octogénaire.

Thirteen centres, specialized in pacemakers and studying the implantation, brought together their material. Consequently, 785 patients living with a pacemaker (PM) were grouped. The average age at the time of the first implantation is 83,5 years old. The mortality of 7.5% as well as the morbidity of 16% lie very low. Nevertheless, their causes are discussed and compared to those resulting from other types of surgical pathologies on the octogenarian. It seems that the implantation of a PM, even on a person of 80 or older, is an efficient therapeutic gesture, benign and capable of normalizing the chances and quality of life on a long term basis.

[Arterial thrombosis of the lower limb in patients over 80 years of age. Belgian multicentric experience concerning 802 patients]. / Arteriële thrombose van het onderste lidmaat boven de 80 jaar. Belgische multicentrische ervaring betreffende 802 patiënten.

This report concerns a joint study of fourteen centres about the treatment of arterial occlusion of the low extremity. Eight hundred and two patients older than 80 years have been studied during a period from 1 to 27 years. The operative mortality was 25.2%. There was no significant difference in mortality between emergency cases and those patients who were operated under elective conditions. Leg amputations are followed by a significant higher mortality than more conservative surgery such as arterial by-pass, sympathectomy or embolectomy. The most frequent cause of death was from cardiopulmonar origin (57%). The postoperative morbidity of cardiac, pulmonary, urinary or infectious origin was frequent (50%). Surgical complications in the true sence of the word are quite rare and their frequency is limited to 7%. The conclusion of this study is that conservative surgery such as reascularisation or sympathectomy is, whenever possible, to be preferred over amputation not only because of their lower mortality (13 to 19%) but also since they permit better revalidation of these elderly patients.

Suboptimal response to clopidogrel and the effect of prasugrel in acute coronary syndromes.

BACKGROUND: High on clopidogrel platelet reactivity (HPR) has been associated with adverse outcomes following acute coronary syndromes (ACS). This study investigated the rate of HPR in a New Zealand ACS population and examined the effectiveness of prasugrel in reducing platelet reactivity in those with HPR. METHODS: In this prospective cohort study, 250 patients with ACS were pretreated with aspirin and clopidogrel and residual platelet reactivity was measured using whole blood multiple electrode platelet aggregometry. Twenty-seven of the patients with HPR were treated with prasugrel at the discretion of their physician, and platelet reactivity retested. RESULTS: Ninety-five patients (38%) had HPR. Maori and Pacific Island patients had a higher rate of HPR compared to Europeans (57% versus 35.9%, p=0.013). Additionally, patients with diabetes were also found to have higher rate of HPR compared to non-diabetics (50% versus 34.8%, p=0.045). Patients treated with a low dose clopidogrel regimen had significantly higher rates of HPR (45.4%) compared to those treated with intermediate (25.4%) or high dose regimens (26.8%, p=0.009). All of the 27 patients with HPR who were subsequently treated with prasugrel (60 mg) had a significant decrease in platelet reactivity (660 AU min (565-770) before versus 230 AU min (110-345) after, p<0.001), and was reduced to below the HPR cutoff in 24 (88.9%) of the patients. CONCLUSIONS: Ethnicity, diabetes and clopidogrel dose contributed to HPR. The use of prasugrel in those with HPR resulted in a consistent and marked reduction in platelet reactivity.

The ontogeny of New Zealand groper (Polyprion oxygeneios) lymphoid organs and IgM.

This study investigates the ontogeny of New Zealand groper (Polyprion oxygeneios) immune system, a new species for aquaculture in the Southern Pacific Ocean. In the eggs, both lysozyme and IgM were detected. Egg IgM was found at 1.07-1.56 µg/g wet weight and consisted of monomers compared to the polymerized IgM found in adult serum. In larvae, the head-kidney (HK) was first observed at 6 dph, followed by the spleen at 16 dph, and thymus at 20 dph, and within these organs IgM(+) cells were first detected in the HK (12 dph), then the spleen (32 dph) and finally in the thymus and the gastrointestinal tract (45 dph). Low levels of Igµ heavy chain transcripts were detected at 2 and 3 dph and they increased at 9 dph. Igµ expression further increased from day 45 onwards. In juveniles (115 dph), the HK and blood showed similar percentages of IgM(+) cells as the adult groper. These results highlight the important maturation steps that occur during the development of the immune system in the marine teleost P. oxygeneios.

Chronic exposure to schistosome eggs reduces serum cholesterol but has no effect on atherosclerotic lesion development.

Previous studies have shown that people infected with schistosomiasis have lower levels of serum cholesterol than uninfected controls. To better understand the impact of this parasitic infection on serum cholesterol levels and on atherosclerotic lesion development induced by hypercholesterolemia, apolipoprotein E (ApoE)-deficient mice were chronically exposed to the eggs of Schistosoma mansoni over a period of 16 weeks. Total serum cholesterol and low-density lipoprotein (LDL) were reduced in egg-exposed ApoE-deficient mice fed a diet high in cholesterol compared to unexposed controls. However, exposure to eggs had no effect on atherosclerotic lesion size or progression in ApoE-deficient mice. Macrophages isolated from egg-exposed mice had an enhanced ability to take up LDL but not acetylated LDL (acLDL). This study suggests that schistosome eggs alone may alter serum lipid profiles through enhancing LDL uptake by macrophages, but these changes do not ultimately affect atherosclerotic lesion development.

The absence of IL-6 does not affect Th2 cell development in vivo, but does lead to impaired proliferation, IL-2 receptor expression, and B cell responses.

The role of IL-6 in Th2 cell differentiation and response development after the injection of eggs from Schistosoma mansoni was investigated using wild-type (WT) and IL-6-/- mice. IL-6 was induced in the lymph nodes (LN) of WT mice within 24 h of egg injection, and IL-4 production by WT LN cells and CD4 T cells isolated 24 h after egg injection and stimulated in vitro was observed. In the absence of IL-6, this early production of IL-4 by LN cells and purified CD4 T cells was not abolished; although the level of IL-4 produced by IL-6-/- LN cells was similar to WT, IL-4 production by purified IL-6-/- CD4 T cells was reduced compared with WT. Despite the difference in CD4 T cell production of IL-4, the development of egg-specific Th2 cells 7 days after egg injection was not affected by the absence of IL-6. Nevertheless, Ab production was impaired and the in vitro proliferative response of whole LN cell populations, CD4 and CD8 T cells, and B cells from IL-6-/- mice was poor compared with WT. The proliferative defect in the IL-6-/- cells correlated with decreased IL-2R expression, and addition of exogenous IL-6 enhanced IL-2R expression as well as proliferation of LN cells from IL-6-/- mice. These studies demonstrate that Th2 differentiation and response development in vivo is not dependent on IL-6, but that Th-dependent and independent B cell responses are. Our results also emphasize the importance of IL-6 for lymphoproliferation, possibly through induction of IL-2R expression.