Fas death receptor signalling: roles of Bid and XIAP

Fas (also called CD95 or APO-1), a member of a subgroup of the tumour necrosis factor receptor superfamily that contain an intracellular death domain, can initiate apoptosis signalling and has a critical role in the regulation of the immune system. Fas-induced apoptosis requires recruitment and activation of the initiator caspase, caspase-8 (in humans also caspase-10), within the death-inducing signalling complex. In so-called type 1 cells, proteolytic activation of effector caspases (-3 and -7) by caspase-8 suffices for efficient apoptosis induction. In so-called type 2 cells, however, killing requires amplification of the caspase cascade. This can be achieved through caspase-8-mediated proteolytic activation of the pro-apoptotic Bcl-2 homology domain (BH)3-only protein BH3-interacting domain death agonist (Bid), which then causes mitochondrial outer membrane permeabilisation. This in turn leads to mitochondrial release of apoptogenic proteins, such as cytochrome c and, pertinent for Fas death receptor (DR)-induced apoptosis, Smac/DIABLO (second mitochondria-derived activator of caspase/direct IAP binding protein with low Pi), an antagonist of X-linked inhibitor of apoptosis (XIAP), which imposes a brake on effector caspases. In this review, written in honour of Juerg Tschopp who contributed so much to research on cell death and immunology, we discuss the functions of Bid and XIAP in the control of Fas DR-induced apoptosis signalling, and we speculate on how this knowledge could be exploited to develop novel regimes for treatment of cancer.

Concomitant vascular GRP-receptor and VEGF-receptor expression in human tumors: molecular basis for dual targeting of tumoral vasculature

Gastrin-releasing peptide (GRP) and GRP receptors (GRPR) play a role in tumor angiogenesis. Recently, GRPR were found to be frequently expressed in the vasculature of a large variety of human cancers. Here, we characterize these GRPR by comparing the vascular GRPR expression and localization in a selection of human cancers with that of an established biological marker of neoangiogenesis, the vascular endothelial growth factor (VEGF) receptor. In vitro quantitative receptor autoradiography was performed in parallel for GRPR and VEGF receptors (VEGFR) in 32 human tumors of various origins, using ¹²⁵I-Tyr-bombesin and ¹²⁵I-VEGF₁₆₅ as radioligands, respectively. Moreover, VEGFR-2 was evaluated immunohistochemically. All tumors expressed GRPR and VEGFR in their vascular system. VEGFR were expressed in the endothelium in the majority of the vessels. GRPR were expressed in a subpopulation of vessels, preferably in their muscular coat. The vessels expressing GRPR were all VEGFR-positive whereas the VEGFR-expressing vessels were not all GRPR-positive. GRPR expressing vessels were found immunohistochemically to co-express VEGFR-2. Remarkably, the density of vascular GRPR was much higher than that of VEGFR. The concomitant expression of GRPR with VEGFR appears to be a frequent phenomenon in many human cancers. The GRPR, localized and expressed in extremely high density in a subgroup of vessels, may function as target for antiangiogenic tumor therapy or angiodestructive targeted radiotherapy with radiolabeled bombesin analogs alone, or preferably together with VEGFR targeted therapy.

Deletions of 11q22.3-q25 are associated with atypical lung carcinoids and poor clinical outcome

Carcinoids are slow-growing neuroendocrine tumors that, in the lung, can be subclassified as typical (TC) or atypical (AC). To identify genetic alterations that improve the prediction of prognosis, we investigated 34 carcinoid tumors of the lung (18 TCs, 15 ACs, and 1 unclassified) by using array comparative genomic hybridization (array CGH) on 3700 genomic bacterial artificial chromosome arrays (resolution ?1 Mb). When comparing ACs with TCs, the data revealed: i) a significant difference in the average number of chromosome arms altered (9.6 versus 4.2, respectively; P = 0.036), with one subgroup of five ACs having more than 15 chromosome arms altered; ii) chromosomal changes in 30% of ACs or more with additions at 9q (?1 Mb) and losses at 1p, 2q, 10q, and 11q; and iii) 11q deletions in 8 of 15 ACs versus 1 of 18 TCs (P = 0.004), which was confirmed via fluorescence in situ hybridization. The four critical regions of interest in 45% ACs or more comprised 11q14.1, 11q22.1-q22.3, 11q22.3-q23.2, and 11q24.2-q25, all telomeric of MEN1 at 11q13. Results were correlated with patient clinical data and long-term follow-up. Thus, there is a strong association of 11q22.3-q25 loss with poorer prognosis, alone or in combination with absence of 9q34.11 alterations (P = 0.0022 and P = 0.00026, respectively).

Factor Structure of the Neck Disability Index

BACKGROUND CONTEXT: The Neck Disability Index frequently is used to measure outcomes of the neck. The statistical rigor of the Neck Disability Index has been assessed with conflicting outcomes. To date, Confirmatory Factor Analysis of the Neck Disability Index has not been reported for a suitably large population study. Because the Neck Disability Index is not a condition-specific measure of neck function, initial Confirmatory Factor Analysis should consider problematic neck patients as a homogenous group. PURPOSE: We sought to analyze the factor structure of the Neck Disability Index through Confirmatory Factor Analysis in a symptomatic, homogeneous, neck population, with respect to pooled populations and gender subgroups. STUDY DESIGN: This was a secondary analysis of pooled data. PATIENT SAMPLE: A total of 1,278 symptomatic neck patients (67.5% female, median age 41 years), 803 nonspecific and 475 with whiplash-associated disorder. OUTCOME MEASURES: The Neck Disability Index was used to measure outcomes. METHODS: We analyzed pooled baseline data from six independent studies of patients with neck problems who completed Neck Disability Index questionnaires at baseline. The Confirmatory Factor Analysis was considered in three scenarios: the full sample and separate sexes. Models were compared empirically for best fit. RESULTS: Two-factor models have good psychometric properties across both the pooled and sex subgroups. However, according to these analyses, the one-factor solution is preferable from both a statistical perspective and parsimony. The two-factor model was close to significant for the male subgroup (p<.07) where questions separated into constructs of mental function (pain, reading headaches and concentration) and physical function (personal care, lifting, work, driving, sleep, and recreation). CONCLUSIONS: The Neck Disability Index demonstrated a one-factor structure when analyzed by Confirmatory Factor Analysis in a pooled, homogenous sample of neck problem patients. However, a two-factor model did approach significance for male subjects where questions separated into constructs of mental and physical function. Further investigations in different conditions, subgroup and sex-specific populations are warranted.

Chapter VI: Follow-up after revascularisation

Structured follow-up after revascularisation for chronic critical limb ischaemia (CLI) aims at sustained treatment success and continued best patient care. Thereby, efforts need to address three fundamental domains: (A) best medical therapy, both to protect the arterial reconstruction locally and to reduce atherosclerotic burden systemically; (B) surveillance of the arterial reconstruction; and (C) timely initiation of repeat interventions. As most CLI patients are elderly and frail, sustained resolution of CLI and preserved ambulatory capacity may decide over independent living and overall prognosis. Despite this importance, previous guidelines have largely ignored follow-up after CLI; arguably because of a striking lack of evidence and because of a widespread assumption that, in the context of CLI, efficacy of initial revascularisation will determine prognosis during the short remaining life expectancy. This chapter of the current CLI guidelines aims to challenge this disposition and to recommend evidentially best clinical practice by critically appraising available evidence in all of the above domains, including antiplatelet and antithrombotic therapy, clinical surveillance, use of duplex ultrasound, and indications for and preferred type of repeat interventions for failing and failed reconstructions. However, as corresponding studies are rarely performed among CLI patients specifically, evidence has to be consulted that derives from expanded patient populations. Therefore, most recommendations are based on extrapolations or subgroup analyses, which leads to an almost systematic degradation of their strength. Endovascular reconstruction and surgical bypass are considered separately, as are specific contexts such as diabetes or renal failure; and critical issues are highlighted throughout to inform future studies.

Massive pulmonary embolism: surgical embolectomy versus thrombolytic therapy--should surgical indications be revisited?

The treatment of massive pulmonary embolism (PE) is a matter of debate. We present our institutional experience of patients suffering from massive PE with the aim of comparing the early results, the outcome and quality of life (QoL) between patients primarily assigned to either pulmonary surgical embolectomy (SE) or thrombolytic therapy (TL). A subgroup of patients (TS) with failed responses to TL requiring SE was separately analysed.

Multiplicity of data in trial reports and the reliability of meta-analyses: empirical study

Objectives To examine the extent of multiplicity of data in trial reports and to assess the impact of multiplicity on meta-analysis results. Design Empirical study on a cohort of Cochrane systematic reviews. Data sources All Cochrane systematic reviews published from issue 3 in 2006 to issue 2 in 2007 that presented a result as a standardised mean difference (SMD). We retrieved trial reports contributing to the first SMD result in each review, and downloaded review protocols. We used these SMDs to identify a specific outcome for each meta-analysis from its protocol. Review methods Reviews were eligible if SMD results were based on two to ten randomised trials and if protocols described the outcome. We excluded reviews if they only presented results of subgroup analyses. Based on review protocols and index outcomes, two observers independently extracted the data necessary to calculate SMDs from the original trial reports for any intervention group, time point, or outcome measure compatible with the protocol. From the extracted data, we used Monte Carlo simulations to calculate all possible SMDs for every meta-analysis. Results We identified 19 eligible meta-analyses (including 83 trials). Published review protocols often lacked information about which data to choose. Twenty-four (29%) trials reported data for multiple intervention groups, 30 (36%) reported data for multiple time points, and 29 (35%) reported the index outcome measured on multiple scales. In 18 meta-analyses, we found multiplicity of data in at least one trial report; the median difference between the smallest and largest SMD results within a meta-analysis was 0.40 standard deviation units (range 0.04 to 0.91). Conclusions Multiplicity of data can affect the findings of systematic reviews and meta-analyses. To reduce the risk of bias, reviews and meta-analyses should comply with prespecified protocols that clearly identify time points, intervention groups, and scales of interest.

Association between circulating cytokine levels, diabetes and insulin resistance in a population-based sample (CoLaus study)

OBJECTIVE: The associations between inflammation, diabetes and insulin resistance remain controversial. Hence, we assessed the associations between diabetes, insulin resistance (using HOMA-IR) and metabolic syndrome with the inflammatory markers high sensitivity C-reactive protein (hs-CRP), interleukin-1beta (IL-1β), interleukin-6 (IL-6) and tumour necrosis factor-α (TNF-α). DESIGN: CROSS-SECTIONAL STUDY: PARTICIPANTS: 2884 MEN AND 3201 WOMEN AGED 35 TO 75: METHODS: CRP was assessed by immunoassay and cytokines by multiplexed flow cytometric assay. In a subgroup of 532 participants an oral glucose tolerance test was performed to screen for impaired glucose tolerance (IGT). RESULTS: IL-6, TNF-α and hs-CRP were significantly and positively correlated with fasting plasma glucose, insulin and HOMA-IR. Participants with diabetes had higher IL-6, TNF-α and hs-CRP levels than participants without diabetes; this difference persisted for hs-CRP after multivariate adjustment. Participants with metabolic syndrome had increased IL-6, TNF-α and hs-CRP levels; these differences persisted after multivariate adjustment. Participants in the highest quartile of HOMA-IR had increased IL-6, TNF-α and hs-CRP levels; these differences persisted for TNF-α and hs-CRP after multivariate adjustment. No association was found between IL-1β levels and all diabetes and insulin resistance markers studied. Finally, participants with IGT had higher hs-CRP levels than participants with a normal OGTT, but this difference disappeared after controlling for body mass index (BMI). CONCLUSION: subjects with diabetes, metabolic syndrome and increased insulin resistance present with increased levels of IL6, TNF-α and hs-CRP, while no association was found with IL-1β. The increased inflammatory state of subjects with IGT is partially explained by increased BMI. © 2012 Blackwell Publishing Ltd.

Effects of Echinaforce® treatment on ex vivo-stimulated blood cells

The herb Echinacea purpurea, also called purple coneflower, is regarded as an immune modulator. This study examined changes in cytokine production in blood samples from 30 volunteers before and during 8-day oral administration with an ethanolic extract of fresh Echinacea purpurea (Echinaforce(®)). Daily blood samples were ex vivo stimulated by LPS/SEB or Zymosan and analysed for a series of cytokines and haematological and metabolic parameters. Treatment reduced the proinflammatory mediators TNF-α and IL-1β by up to 24% (p<0.05) and increased anti-inflammatory IL-10 levels by 13% (p<0.05) in comparison to baseline. This demonstrated a substantial overall anti-inflammatory effect of Echinaforce(®) for the whole group (n=28). Chemokines MCP-1 and IL-8 were upregulated by 15% in samples from subjects treated with Echinaforce(®) (p<0.05). An analysis of a subgroup of volunteers who showed low pre-treatment levels of the cytokines MCP-1, IL-8, IL-10 or IFN-γ (n=8) showed significant stimulation of these factors upon Echinaforce(®) treatment (30-49% increases; p<0.05), whereas the levels in subjects with higher pre-treatment levels remained unaffected. We chose the term "adapted immune-modulation" to describe this observation. Volunteers who reported high stress levels (n=7) and more than 2 colds per year experienced a significant transient increase in IFN-γ upon Echinaforce(®) treatment (>50%). Subjects with low cortisol levels (n=11) showed significant down-regulation of the acute-phase proteins IL1-β, IL-6, IL-12 and TNF-α by Echinaforce(®) (range, 13-25%), while subjects with higher cortisol levels showed no such down-regulation. This is the first ex vivo study to demonstrate adapted immune-modulation by an Echinacea preparation. While Echinaforce(®) did not affect leukocyte counts, we speculate that the underlying therapeutic mechanism is based on differential multi-level modulation of the responses of the different types of leukocytes. Echinaforce(®) thus regulates the production of chemokines and cytokines according to current immune status, such as responsiveness to exogenous stimuli, susceptibility to viral infection and exposure to stress.

Replicative phenotyping adds value to genotypic resistance testing in heavily pre-treated HIV-infected individuals--the Swiss HIV Cohort Study

Background Replicative phenotypic HIV resistance testing (rPRT) uses recombinant infectious virus to measure viral replication in the presence of antiretroviral drugs. Due to its high sensitivity of detection of viral minorities and its dissecting power for complex viral resistance patterns and mixed virus populations rPRT might help to improve HIV resistance diagnostics, particularly for patients with multiple drug failures. The aim was to investigate whether the addition of rPRT to genotypic resistance testing (GRT) compared to GRT alone is beneficial for obtaining a virological response in heavily pre-treated HIV-infected patients. Methods Patients with resistance tests between 2002 and 2006 were followed within the Swiss HIV Cohort Study (SHCS). We assessed patients' virological success after their antiretroviral therapy was switched following resistance testing. Multilevel logistic regression models with SHCS centre as a random effect were used to investigate the association between the type of resistance test and virological response (HIV-1 RNA <50 copies/mL or ≥1.5log reduction). Results Of 1158 individuals with resistance tests 221 with GRT+rPRT and 937 with GRT were eligible for analysis. Overall virological response rates were 85.1% for GRT+rPRT and 81.4% for GRT. In the subgroup of patients with >2 previous failures, the odds ratio (OR) for virological response of GRT+rPRT compared to GRT was 1.45 (95% CI 1.00-2.09). Multivariate analyses indicate a significant improvement with GRT+rPRT compared to GRT alone (OR 1.68, 95% CI 1.31-2.15). Conclusions In heavily pre-treated patients rPRT-based resistance information adds benefit, contributing to a higher rate of treatment success.

Mitochondrial preprotein translocase of trypanosomatids has a bacterial origin

Mitochondria are found in all eukaryotic cells and derive from a bacterial endosymbiont [1, 2]. The evolution of a protein import system was a prerequisite for the conversion of the endosymbiont into a true organelle. Tom40, the essential component of the protein translocase of the outer membrane, is conserved in mitochondria of almost all eukaryotes but lacks bacterial orthologs [3-6]. It serves as the gateway through which all mitochondrial proteins are imported. The parasitic protozoa Trypanosoma brucei and its relatives do not have a Tom40-like protein, which raises the question of how proteins are imported by their mitochondria [7, 8]. Using a combination of bioinformatics and in vivo and in vitro studies, we have discovered that T. brucei likely employs a different import channel, termed ATOM (archaic translocase of the outer mitochondria! membrane). ATOM mediates the import of nuclear-encoded proteins into mitochondria and is essential for viability of trypanosomes. It is not related to Tom40 but is instead an ortholog of a subgroup of the 0mp85 protein superfamily that is involved in membrane translocation and insertion of bacterial outer membrane proteins [9]. This suggests that the protein import channel in trypanosomes is a relic of an archaic protein transport system that was operational in the ancestor of all eukaryotes.

Proposal for a 10-high-power-fields scoring method for the assessment of tumor budding in colorectal cancer

Although tumor budding is linked to adverse prognosis in colorectal cancer, it remains largely unreported in daily diagnostic work due to the absence of a standardized scoring method. Our aim was to assess the inter-observer agreement of a novel 10-high-power-fields method for assessment of tumor budding at the invasive front and to confirm the prognostic value of tumor budding in our setting of colorectal cancers. Whole tissue sections of 215 colorectal cancers with full clinico-pathological and follow-up information were stained with cytokeratin AE1/AE3 antibody. Presence of buds was scored across 10-high-power fields at the invasive front by two pathologists and two additional observers were asked to score 50 cases of tumor budding randomly selected from the larger cohort. The measurements were correlated to the patient and tumor characteristics. Inter-observer agreement and correlation between observers' scores were excellent (P<0.0001; intraclass correlation coefficient=0.96). A test subgroup of 65 patients (30%) was used to define a valid cutoff score for high-grade tumor budding and the remaining 70% of the patients were entered into the analysis. High-grade budding was defined as an average of ≥10 buds across 10-high-power fields. High-grade budding was associated with a higher tumor grade (P<0.0001), higher TNM stage (P=0.0003), vascular invasion (P<0.0001), infiltrating tumor border configuration (P<0.0001) and reduced survival (P<0.0001). Multivariate analysis confirmed its independent prognostic effect (P=0.007) when adjusting for TNM stage and adjuvant therapy. Using 10-high-power fields for evaluating tumor budding has independent prognostic value and shows excellent inter-observer agreement. Like the BRE and Gleason scores in breast and prostate cancers, respectively, tumor budding could be a basis for a prognostic score in colorectal cancer.

Infrapopliteal lesion morphology in patients with critical limb ischemia: implications for the development of anti-restenosis technologies

Purpose : To angiographically evaluate infrapopliteal arterial lesion morphology in a consecutive series of patients presenting with critical limb ischemia (CLI) and undergoing infrapopliteal angioplasty. Methods : A prospective analysis was undertaken of a consecutive series of CLI patients undergoing endovascular therapy in a tertiary referral center in the year 2011. Morphological assessment of baseline angiograms obtained prior to revascularization included lesion length, assessment of calcification using a semi-quantitative scoring system, and reference vessel diameter (RVD) measurement. Delta RVDs were assessed subtracting distal RVDs from proximal RVDs. A total of 197 infrapopliteal lesions in 105 CLI patients (n=106 limbs) were assessed. Of these, 136 lesions were treated by endovascular means. Results : The average length of treated lesions was 87.1±43.8 mm in stenoses and 124.0±78.3 mm in chronic occlusions (p<0.001). Mean RVD proximal to the lesions was 1.88 mm whereas it was 1.66 mm distal to the lesions (p≤0.03). Mean arterial calcification was 1.15. Conclusion : This prospective angiographic series underlines the complex nature and extensive longitudinal involvement of infrapopliteal lesions in CLI patients. These findings should be taken into consideration for anti-restenosis concepts in this challenging subgroup of peripheral artery disease patients.

Prevalence of vitamin B12 depletion and deficiency in Liechtenstein

OBJECTIVE: Data about vitamin B12 (B12) deficiency in the general population are scarce. The present study was performed to determine the prevalence of B12 deficiency in the general population of the Principality of Liechtenstein, as well as to identify sub-populations potentially at high risk. DESIGN: Retrospective study. SETTING: Ambulatory setting, population of the Principality of Liechtenstein. SUBJECTS: Seven thousand four hundred and twenty-four patients seeking medical attention whose serum samples were referred for routine work-up in an ambulatory setting were consecutively enrolled. Serum total B12 was determined in all patients in this cohort. In addition, for a subgroup of 1328 patients, serum holotranscobalamin was also measured. Prevalence of B12 deficiency was calculated. Further, multivariate logistical regression models were applied to identify covariates independently associated with B12 deficiency and depletion. RESULTS: Nearly 8 % of the general population was suffering from either B12 depletion or deficiency. The ratio between B12 depletion and deficiency was 2:1 for all age ranges. Pathological changes were detected predominantly in older people. Female gender was a significant predictor of B12 depletion. In the cohort, nearly 40 % exhibited either depletion or deficiency of B12. CONCLUSIONS: B12 depletion and deficiency are common in Liechtenstein, a Central European country. The measurement of biochemical markers represents a cost-efficient and valid assessment of the B12 state. When a deficiency of B12 is diagnosed at an early stage, many cases can be treated or prevented, with beneficial effects on individual outcomes and subsequent potential reductions in health-care costs.

Common obesity risk alleles in childhood attention-deficit/hyperactivity disorder

Children with attention-deficit/hyperactivity disorder (ADHD) have a higher rate of obesity than children without ADHD. Obesity risk alleles may overlap with those relevant for ADHD. We examined whether risk alleles for an increased body mass index (BMI) are associated with ADHD and related quantitative traits (inattention and hyperactivity/impulsivity). We screened 32 obesity risk alleles of single nucleotide polymorphisms (SNPs) in a genome-wide association study (GWAS) for ADHD based on 495 patients and 1,300 population-based controls and performed in silico analyses of the SNPs in an ADHD meta-analysis comprising 2,064 trios, 896 independent cases, and 2,455 controls. In the German sample rs206936 in the NUDT3 gene (nudix; nucleoside diphosphate linked moiety X-type motif 3) was associated with ADHD risk (OR: 1.39; P = 3.4 × 10(-4) ; Pcorr  = 0.01). In the meta-analysis data we found rs6497416 in the intronic region of the GPRC5B gene (G protein-coupled receptor, family C, group 5, member B; P = 7.2 × 10(-4) ; Pcorr  = 0.02) as a risk allele for ADHD. GPRC5B belongs to the metabotropic glutamate receptor family, which has been implicated in the etiology of ADHD. In the German sample rs206936 (NUDT3) and rs10938397 in the glucosamine-6-phosphate deaminase 2 gene (GNPDA2) were associated with inattention, whereas markers in the mitogen-activated protein kinase 5 gene (MAP2K5) and in the cell adhesion molecule 2 gene (CADM2) were associated with hyperactivity. In the meta-analysis data, MAP2K5 was associated with inattention, GPRC5B with hyperactivity/impulsivity and inattention and CADM2 with hyperactivity/impulsivity. Our results justify further research on the elucidation of the common genetic background of ADHD and obesity.