Colon capsule endoscopy for colonic surveillance.

AIM: Resources used in surveillance colonoscopies are taking up an increasing proportion of colonoscopy capacity. Colon capsule endoscopy (CCE) is a promising technique for noninvasive investigation of the colon. We aimed to investigate CCE as a possible filter in colonic surveillance with the primary outcome of reducing the number of colonoscopies. METHOD: Patients scheduled for follow-up colonoscopy were subjected to a primary CCE and only supplemental conventional endoscopy if significant pathology was detected or if the CCE examination was incomplete. Significant pathology was defined as more than two small polyps, or one polyp greater than 9 mm or any polyp in patients with hereditary nonpolyposis colorectal cancer. Supplemental endoscopy was carried out to the extent needed to resect the detected polyps and investigate the parts of the colon that were not sufficiently visualized by the capsule. RESULTS: A total of 180 patients were included. Seventy-seven patients (43%) had a complete CCE with no significant findings. A complete colonoscopy was carried out in 67 patients (37%) and 36 patients (20%) underwent a sigmoidoscopy. In the 103 patients undergoing endoscopy, 59 (57%) had no adenomas detected, 33 (32%) had 'low-risk' adenomas and 11 (11%) had 'high-risk' adenomas. CONCLUSION: The introduction of CCE as filter test in colonic surveillance reduced colonoscopies by 43%, but this implies that untreated polyps are left behind and is not cost-effective. The CCE completion rate must be improved.

The PARIS score can reliably predict 7-day all-cause mortality for both acute medical and surgical patients: an international validation study.

BACKGROUND: We believe errors in the risk assessment of acutely ill patients occur because only vital signs without concurrent functional capacity are considered. We, therefore, developed the PARIS risk score based on blood pressure, age, respiratory rate, loss of independence and oxygen saturation. AIM: Validation of the PARIS score in four independent cohorts from three countries. METHODS: Retrospective cohort study of acutely ill patients admitted to hospitals in Denmark, Ireland and Uganda. Vital signs and functional capacity (registered as ability to stand or walk or get into bed unaided) was recorded upon arrival. Patients were followed up for 7 days (Denmark and Ireland) or until discharge (Uganda) and mortality recorded. The discriminatory power (ability to identify patients at increased risk) was determined using area under the receiver operating characteristics curve (AUROC) and calibration (precision) using Hosmer-Lemeshow goodness of fit test. RESULTS: Out of 14 447 patients, 327 (2.3%) died within 7 days: median age was 59 (39-75) years and 7458 (51.8%) were female. Seven-day mortality increased from 0.3% with a score of 0-26.7% with a score of 5. The score's AUROC as 0.833 [95% confidence interval (95% CI) 0.811-0.856], 0.817 (95% CI 0.792-0.842) and 0.894 (95% CI 0.813-0.974) for all patients, medical patients and surgical patients, respectively. However, except for surgical patients, calibration of the score was poor. CONCLUSION: The PARIS score can identify both high and low risk acutely admitted medical and surgical patients, but calibration was poor for medical patients.

Abrogating fibrinolysis does not improve bleeding or rFVIIa/rFVIII treatment in a non-mucosal venous injury model in haemophilic rodents.

Essentials The efficacy of systemic antifibrinolytics for hemophilic non-mucosal bleeding is undetermined. The effect of systemically inhibiting fibrinolysis in hemophilic mice and rats was explored. Neither bleeding nor the response to factor treatment was improved after inhibiting fibrinolysis. The non-mucosal bleeding phenotype in hemophilia A appears largely unaffected by fibrinolysis. SUMMARY: Background Fibrinolysis may exacerbate bleeding in patients with hemophilia A (HA). Accordingly, antifibrinolytics have been used to help maintain hemostatic control. Although antifibrinolytic drugs have been proven to be effective in the treatment of mucosal bleeds in the oral cavity, their efficacy in non-mucosal tissues remain an open question of significant clinical interest. Objective To determine whether inhibiting fibrinolysis improves the outcome in non-mucosal hemophilic tail vein transection (TVT) bleeding models, and to determine whether a standard ex vivo clotting/fibrinolysis assay can be used as a predictive surrogate for in vivo efficacy. Methods A highly sensitive TVT model was employed in hemophilic rodents with a suppressed fibrinolytic system to examine the effect of inhibiting fibrinolysis on bleeding in non-mucosal tissue. In mice, induced and congenital hemophilia models were combined with fibrinolytic attenuation achieved either genetically or pharmacologically (tranexamic acid [TXA]). In hemophilic rats, tail bleeding was followed by whole blood rotational thromboelastometry evaluation of the same animals to gauge the predictive value of such assays. Results The beneficial effect of systemic TXA therapy observed ex vivo could not be confirmed in vivo in hemophilic rats. Furthermore, neither intravenously administered TXA nor congenital knockout of the fibrinolytic genes encoding plasminogen or tissue-type plasminogen activator markedly improved the TVT bleeding phenotype or response to factor therapy in hemophilic mice. Conclusions The findings here suggest that inhibition of fibrinolysis is not effective in limiting the TVT bleeding phenotype of HA rodents in non-mucosal tissues.

Rapid switch-off of the human myosin heavy chain IIX gene after heavy load muscle contractions is sustained for at least four days.

Long-term heavy load contractions decrease the relative amount of the myosin heavy chain (MHC) IIX isoform in human skeletal muscle, but the timing of the down-regulation in the short term is unknown. Untrained subjects performed two resistance bouts, in two consecutive days, with one leg, the other leg serving as a control (age 24±1, n=5). Muscle biopsies were obtained in both legs before, immediately after, and 24, 54, and 96 hours after exercise. Serial cryosection analysis combined immunohistochemistry and ATPase histochemistry with In Situ hybridization to identify the distribution of MHC isoforms and their corresponding transcripts, enabling identification of transitional fibers. Fibers positive solely for MHC IIX mRNA decreased in the exercised leg throughout the study period. At 96 hours post-exercise, no fibers solely expressed MHC IIX mRNA. In contrast, the number of fibers expressing MHC IIA mRNA increased throughout the study period. The percentage of fibers expressing mRNA for MHC I was unchanged in both legs at all time points. Pronounced depletion of glycogen in the MHC IIX fibers of the exercised leg verifies that the type IIX fibers were active during the heavy load contractions. Major mismatch between MHC at the mRNA and protein levels was only found in the fibers of the exercised leg. These data provide unequivocal in situ evidence of an immediate shutdown of the MHC IIX gene after resistance exercise. A further novel finding was that the silencing of the MHC IIX gene is sustained at least 4 days after removal of the stimulus.

Programming microphysiological systems for children's health protection.

Microphysiological systems (MPS) and computer simulation models that recapitulate the underlying biology and toxicology of critical developmental transitions are emerging tools for developmental effects assessment of drugs/chemicals. Opportunities and challenges exist for their application to alternative, more public health relevant and efficient chemical toxicity testing methods. This is especially pertinent to children's health research and the evaluation of complex embryological and reproductive impacts of drug/chemical exposure. Scaling these technologies to higher throughput is a key challenge and drives the need for in silico models for quantitative prediction of developmental toxicity to inform safety assessments. One example is cellular agent-based models, constructed from extant embryology, that produce data useful to simulate critical developmental transitions and thereby predict phenotypic consequences of disruption in silico. Biologically inspired MPS models built from human induced pluripotent stem (iPS)-derived cells and synthetic matrices that recapitulate organ-specific physiologies and native tissue architectures are providing exciting new research opportunities to advance the assessment of developmental toxicity and offer the possibility of deriving a full 'human on a chip' system, or a 'Homunculus.' Impact statement This 'commentary' summarizes research needs and opportunities for engineered MPS models for developmental and reproductive toxicity testing. Emerging concepts can be taken forward to a virtual tissue modeling framework for assessing chemical (and non-chemical) stressors on human development. These models will advance children's health research, both basic and translational and new ways to evaluate complex embryological and reproductive impacts of drug and chemical exposures to inform safety assessments.

Development of a tail vein transection bleeding model in fully anaesthetized haemophilia A mice - characterization of two novel FVIII molecules.

INTRODUCTION: The tail tip bleeding model and the tail vein transection survival model in mice are important tools for assessment of in vivo effect in haemostasis research. While the tail vein transection model exhibits the best sensitivity to pharmacological intervention it uses death or near-death as endpoint which is fully avoided in the tail tip bleeding model. AIM: The aim of this study was to develop a new tail bleeding model maintaining the sensitivity of the previous survival model but avoiding death/near-death as endpoint. METHODS: Combining the two existing tail bleeding models we developed an optimized version of the survival model with full anaesthetic coverage and short duration of experiments. Using this model, we characterized the effect of turoctocog alfa, a B-domain truncated FVIII molecule (NovoEight(®) ), as well as the prolonged half-life version of the same molecule (turoctocog alfa pegol, N8-GP). RESULTS: Data showed that the model was sensitive to clinically relevant doses of both turoctocog alfa as well as N8-GP when dosed for 'on demand' treatment. The model also correctly identified a longer duration of effect for N8-GP compared with turoctocog alfa. Moreover, the model allowed the use of mice of both genders and was reproducible over time. CONCLUSION: The optimized tail vein transection bleeding model is sensitive to standard as well as half-life prolonged FVIII molecules and should be a valuable alternative to both the tail tip bleeding model, enhancing sensitivity to pharmacological intervention, as well as to the previously used tail vein transection survival model, avoiding death or near-death as endpoint.

Expression of tissue factor in canine mammary tumours and correlation with grade, stage and markers of haemostasis and inflammation.

Tissue factor (TF) expression in human cancers has been associated with a procoagulant state and facilitation of metastasis. This study was conducted in order to evaluate if TF was expressed in canine mammary tumours. Forty epithelial mammary tumours from 28 dogs were included. TF expression of the tumours was evaluated by immunohistochemistry using a polyclonal antibody against recombinant canine TF. In addition, thromboelastography, haemostatic and inflammatory parameters were evaluated in the patients. TF was recognized in 44% of benign and 58% of malignant tumours. TF localized to the cytoplasmic membrane of neoplastic luminal epithelial cells and/or diffusely in the cytoplasm. No association was found between TF expression and stage or grade of disease. A significant association between TF expression and antithrombin and plasminogen was found, and extensive TF expression was seen in a lymph node metastasis classified as anaplastic mammary carcinoma from a dog with concomitant disseminated intravascular coagulation (DIC).

Physical activity enhances metabolic fitness independently of cardiorespiratory fitness in marathon runners.

High levels of cardiovascular fitness (CRF) and physical activity (PA) are associated with decreased mortality and risk to develop metabolic diseases. The independent contributions of CRF and PA to metabolic disease risk factors are unknown. We tested the hypothesis that runners who run consistently >50 km/wk and/or >2 marathons/yr for the last 5 years have superior metabolic fitness compared to matched sedentary subjects (CRF, age, gender, and BMI). Case-control recruitment of 31 pairs of runner-sedentary subjects identified 10 matched pairs with similar VO2max (mL/min/kg) (similar-VO2max). The similar-VO2max group was compared with a group of age, gender, and BMI matched pairs who had the largest difference in VO2max (different-VO2max). Primary outcomes that defined metabolic fitness including insulin response to an oral glucose tolerance test, fasting lipids, and fasting insulin were superior in runners versus sedentary controls despite similar VO2max. Furthermore, performance (velocity at VO2max, running economy), improved exercise metabolism (lactate threshold), and skeletal muscle levels of mitochondrial proteins were superior in runners versus sedentary controls with similar VO2max. In conclusion subjects with a high amount of PA have more positive metabolic health parameters independent of CRF. PA is thus a good marker against metabolic diseases.

Consequences of crown shortening canine teeth in Greenland sled dogs.

OBJECTIVES: To evaluate the consequences of crown shortening, focusing on the prevalence of pulp exposure and periapical pathology in Greenland sled dogs that had had their canine crowns shortened at an early age. METHODS: Five cadaver heads and 54 sled dogs underwent an oral examination for dental fractures and pulp exposure of canines. All canines were radiographed and evaluated for periapical pathology. RESULTS: The prevalence of canine pulp exposure in 12 (5 heads and 7 dogs) crown shortened dogs was 91 · 7%, and 21 · 3% in 47 not-crown shortened dogs. A significant (P < 0 · 001) risk of pulp exposure of the canines in the crown shortened group compared to the not-crown shortened group was seen with a relative risk of 4 · 3 on a dog basis and a relative risk of 12 · 2 on a tooth basis. In dogs with pulp exposure of canines (n = 51) the prevalence of periapical pathology was 82 · 4%, but only 0 · 8% in dogs without pulp exposure (n = 133) resulting in a significant (relative risk, 109 · 5; P < 0 · 001) risk of periapical pathology in teeth with pulp exposure compared to teeth without pulp exposure. CLINICAL SIGNIFICANCE: The high risk of periapical pathology observed in teeth with pulp exposure confirms that these teeth should not be neglected in affected dogs.

Immediate and long-term consequences of vascular toxicity during zebrafish development.

Proper formation of the vascular system is necessary for embryogenesis, and chemical disruption of vascular development may be a key event driving developmental toxicity. In order to test the effect of environmental chemicals on this critical process, we evaluated a quantitative assay in transgenic zebrafish using angiogenesis inhibitors that target VEGFR2 (PTK787) or EGFR (AG1478). Both PTK787 and AG1478 exposure impaired intersegmental vessel (ISV) sprouting, while AG1478 also produced caudal and pectoral fin defects at concentrations below those necessary to blunt ISV morphogenesis. The functional consequences of vessel toxicity during early development included decreased body length and survival in juvenile cohorts developmentally exposed to inhibitor concentrations sufficient to completely block ISV sprouting angiogenesis. These data show that concentration-dependent disruption of the presumed targets for these inhibitors produce adverse outcomes at advanced life stages.

Release of targeted p53 from the mitochondrion as an early signal during mitochondrial dysfunction.

Increased accumulation of p53 tumor suppressor protein is an early response to low-level stressors. To investigate the fate of mitochondrial-sequestered p53, mouse embryonic fibroblast cells (MEFs) on a p53-deficient genetic background were transfected with p53-EGFP fusion protein led by a sense (m53-EGFP) or antisense (c53-EGFP) mitochondrial import signal. Rotenone exposure (100nM, 1h) triggered the translocation of m53-EGFP from the mitochondrion to the nucleus, thus shifting the transfected cells from a mitochondrial p53 to a nuclear p53 state. Antibodies for p53 serine phosphorylation or lysine acetylation indicated a different post-translational status of recombinant p53 in the nucleus and mitochondrion, respectively. These data suggest that cycling of p53 through the mitochondria may establish a direct pathway for p53 signaling from the mitochondria to the nucleus during mitochondrial dysfunction. PK11195, a pharmacological ligand of mitochondrial TSPO (formerly known as the peripheral-type benzodiazepine receptor), partially suppressed the release of mitochondria-sequestered p53. These findings support the notion that p53 function mediates a direct signaling pathway from the mitochondria to nucleus during mitochondrial dysfunction.

eHealth: individualisation of infliximab treatment and disease course via a self-managed web-based solution in Crohn's disease.

BACKGROUND: Infliximab (IFX) maintenance therapy for Crohn's disease (CD) is administered every 8 weeks, but inter-patient variation in optimal treatment intervals may exist. AIM: To assess, in a prospective pilot study, the efficacy, safety and quality of life (QoL) of IFX maintenance treatment scheduled through web-based self-monitoring of disease activity. METHODS: Twenty-seven CD patients in IFX maintenance therapy were enrolled and received a standardised disease education and web-training. Using the http://www.cd.constant-care.dk concept, patients recorded their disease activity and faecal calprotectin weekly. From this, the inflammatory burden (IB) score was calculated, placing patients in the green, yellow or red zones of a 'traffic light' system. If placed in the yellow or red zones, the computer directed these patients to consult their physician for IFX infusion. RESULTS: Seventeen patients (63%) completed 52 weeks of follow-up, 6 (22%) completed 26 weeks and 4 (15%) were excluded due to loss of response, patient decision or non-adherence. In total, 121 IFX infusions were given with a median interval of 9 (range: 4­18) weeks. Only 10% of infusions were given at 8-week intervals, whereas 39% were administered with shorter and 50% with longer intervals respectively. The mean IB and the QoL remained stable during the web-treatment. One mild infusion reaction and one case of folliculitis were observed, while three patients underwent surgery. CONCLUSIONS: The program http://www.cd.constant-care.dk appears to be a practical and safe concept for the individualised scheduling of maintenance treatment with IFX in patients with Crohn's disease. Larger studies are awaited to confirm this preliminary outcome.

Zebrafish developmental screening of the ToxCast™ Phase I chemical library.

Zebrafish (Danio rerio) is an emerging toxicity screening model for both human health and ecology. As part of the Computational Toxicology Research Program of the U.S. EPA, the toxicity of the 309 ToxCast™ Phase I chemicals was assessed using a zebrafish screen for developmental toxicity. All exposures were by immersion from 6-8 h post fertilization (hpf) to 5 days post fertilization (dpf); nominal concentration range of 1 nM-80 µM. On 6 dpf larvae were assessed for death and overt structural defects. Results revealed that the majority (62%) of chemicals were toxic to the developing zebrafish; both toxicity incidence and potency was correlated with chemical class and hydrophobicity (logP); and inter-and intra-plate replicates showed good agreement. The zebrafish embryo screen, by providing an integrated model of the developing vertebrate, compliments the ToxCast assay portfolio and has the potential to provide information relative to overt and organismal toxicity.

Identifying developmental toxicity pathways for a subset of ToxCast chemicals using human embryonic stem cells and metabolomics.

Metabolomics analysis was performed on the supernatant of human embryonic stem (hES) cell cultures exposed to a blinded subset of 11 chemicals selected from the chemical library of EPA's ToxCast™ chemical screening and prioritization research project. Metabolites from hES cultures were evaluated for known and novel signatures that may be indicative of developmental toxicity. Significant fold changes in endogenous metabolites were detected for 83 putatively annotated mass features in response to the subset of ToxCast chemicals. The annotations were mapped to specific human metabolic pathways. This revealed strong effects on pathways for nicotinate and nicotinamide metabolism, pantothenate and CoA biosynthesis, glutathione metabolism, and arginine and proline metabolism pathways. Predictivity for adverse outcomes in mammalian prenatal developmental toxicity studies used ToxRefDB and other sources of information, including Stemina Biomarker Discovery's predictive DevTox® model trained on 23 pharmaceutical agents of known developmental toxicity and differing potency. The model initially predicted developmental toxicity from the blinded ToxCast compounds in concordance with animal data with 73% accuracy. Retraining the model with data from the unblinded test compounds at one concentration level increased the predictive accuracy for the remaining concentrations to 83%. These preliminary results on a 11-chemical subset of the ToxCast chemical library indicate that metabolomics analysis of the hES secretome provides information valuable for predictive modeling and mechanistic understanding of mammalian developmental toxicity.

Differential programming of p53-deficient embryonic cells during rotenone block.

Mitochondrial dysfunction has been implicated in chemical toxicities. The present study used an in vitro model to investigate the differential expression of metabolic pathways during cellular stress in p53-efficient embryonic fibroblasts compared to p53-deficient cells. These cell lines differed with respect to NADH/NAD(+) balance. This ratio constitutes a driving force for NAD- and NADH-dependent reactions and is inversed upon exposure to Rotenone (complex I inhibitor). Rotenone perturbed the structure of the elongated fibrillar tubulin network and decreased mRNA expression of tubulin genes both suggesting reprogramming and reorganization of the cytoskeleton in both cell lines. These changes were reflected in the abundance of specific mRNA and microRNA (miRNA) species as determined from genome-based analysis. Changes in mRNA and miRNA expression profiles reflected differences in energy utilizing pathways, consistent with the notion that the p53 pathway influences the cellular response to mitochondrial dysfunction and that at least some control may be embedded within specific mRNA/miRNA networks in embryonic cells.

Pharmacokinetics, pharmacodynamics and safety of recombinant canine FVIIa in a study dosing one haemophilia A and one haemostatically normal dog.

Recombinant human FVIIa (rhFVIIa) corrects the coagulopathy in hemophilia A and B as well as FVII deficiency. This is also the case in dogs until canine anti-human FVIIa antibodies develop (~2 weeks). Recombinant canine factor VIIa (rcFVIIa), successfully over-expressed by gene transfer in haemophilia dogs, has provided long-term haemostasis (>2 years). However, pharmacokinetics (PK), pharmacodynamics (PD) and safety of rcFVIIa after pharmacological administration have not been reported. We therefore wanted to explore the safety, PK and PD of rcFVIIa in dogs. A pilot study was set up to evaluate the safety as well as PK and PD of rcFVIIa after a single intravenous dose of 270 µg kg(-1) to one HA and one haemostatically normal dog and to directly compare rcFVIIa with rhFVIIa in these two dogs. Single doses of rcFVIIa and rhFVIIa were well tolerated. No adverse events were observed. Pharmacokinetic characteristics including half-life (FVIIa activity: 1.2-1.8 h; FVIIa antigen 2.8-3.7 h) and clearance were comparable for rcFVIIa and rhFVIIa. Kaolin-activated thromboelastography approached normal in the HA dog with the improvement being most pronounced after rcFVIIa. This study provided the first evidence that administering rcFVIIa intravenously is feasible, safe, well tolerated and efficacious in correcting the haemophilic coagulopathy in canine HA and that rcFVIIa exhibits pharmacokinetic characteristics comparable to rhFVIIa in haemophilic and haemostatically competent dogs. This strengthens the hypothesis that rcFVIIa can be administered to dogs to mimic the administration of rhFVIIa to humans.

Characterization of canine coagulation factor VII and its complex formation with tissue factor: canine-human cross-species compatibility.

BACKGROUND: Canine models have been good predictors of efficacy of hemophilia treatments, including recombinant human coagulation factor (F)VIIa (hFVIIa). However, canine FVIIa and tissue factor (TF) have remained incompletely characterized. OBJECTIVE: To explore canine-human cross-species FVIIa-TF compatibility in order to strengthen the predictive value of canine models in research on FVIIa and TF. METHODS: Canine FVIIa (cFVIIa) and canine TF((1-217)) [cTF((1-217))] were produced by recombinant techniques, and canine-human cross-species FVIIa-TF interactions were characterized in vitro. RESULTS: Recombinant cFVIIa and soluble cTF((1-217)) were produced and purified to homogeneity. hFVIIa and cFVIIa bound with comparably high affinities to cTF((1-217)) (K(D)=6.0±0.7 nm and K(D)=6.0±0.3 nm, respectively) and to cell surface-expressed cTF (K(D)=8.4±0.4 nm and K(D)=7.2±1.2 nm, for (125) I-labeled hFVIIa and cFVII, respectively). In contrast, cFVIIa bound to human TF (hTF) with decreased affinity, both in solution and on cell surfaces. The decreased binding resulted in reduced activity of cFVIIa in functional assays with hTF((1-209)) . In direct comparison, cFVIIa was more active than hFVIIa, both in the absence and the presence of cognate TF. CONCLUSION: The present finding that hFVIIa binds to cTF essentially as it does to hTF substantiates the hypothesis that human FVIIa-TF biology can be reliably recapitulated in canine models on administration of hFVIIa to dogs.

Effects on cycle control and bodyweight of the combined contraceptive ring, NuvaRing, versus an oral contraceptive containing 30 microg ethinyl estradiol and 3 mg drospirenone.

BACKGROUND: The objective of this study was to compare cycle control, cycle-related characteristics and bodyweight effects of NuvaRing with those of a combined oral contraceptive (COC) containing 30 microg of ethinyl estradiol and 3 mg of drospirenone. METHODS: A randomized, multicentre, open-label trial in which 983 women were treated (intent-to-treat population) with NuvaRing or the COC for 13 cycles. RESULTS: Breakthrough bleeding or spotting during cycles 2-13 was in general less frequent with NuvaRing than that with the COC (4.7-10.4%) and showed a statistically significant odds ratio of 0.61 (95% confidence interval: 0.46, 0.80) with longitudinal analysis. Intended bleeding was significantly better for all cycles with NuvaRing (55.2-68.5%) than that with the COC (35.6-56.6%) (P < 0.01). Changes from baseline in mean bodyweight and body composition parameters were relatively small for both groups with no notable between-group differences. CONCLUSION: NuvaRing was associated with better cycle control than the COC, and there was no clinically relevant difference between the two groups in bodyweight.

Gingival epithelial cells heterozygous for Toll-like receptor 4 polymorphisms Asp299Gly and Thr399ile are hypo-responsive to Porphyromonas gingivalis.

The Toll-like receptor (TLR)4 is the major sensor for bacterial lipopolysaccharide and its two common co-segregating polymorphisms, Asp299Gly and Thr399Ile, which occur at a frequency of between 6 and 10%, have been associated with infectious diseases, LPS hypo-responsiveness and cardiovascular disease. Porphyromonas gingivalis is a Gram-negative bacterium implicated in chronic periodontitis and is a known TLR4 and TLR2 agonist. We obtained two gingival epithelial cell primary cultures from subjects heterozygous for the TLR4 polymorphism Asp299Gly and compared response characteristics with similar cells from patients (four) with the wild-type TLR4 genes. Cytokine responses and transcriptome profiles of gingival epithelial cell primary culture cells to TNFalpha challenge were similar for all primary epithelial cell cultures. P. gingivalis challenge, however, gave markedly different responses for Asp299Gly heterozygous and wild-type epithelial cell cultures. The epithelial cells heterozygous for the TLR4 polymorphism Asp299Gly were functionally hypo-responsive, evidenced by differences in BD-2 mRNA expression, mRNA response profile by microarray analysis and by pro-inflammatory and chemokine cytokines at the protein and mRNA level. These findings emphasize variance in human epithelial cell TLRs, linked with Asp299Gly carriage, which results in a hypo-responsive epithelial cell phenotype less susceptible to Gram-negative diseases and associated systemic conditions.

Predictive value of soluble haemoglobin scavenger receptor CD163 serum levels for survival in verified tuberculosis patients.

Pre-treatment serum levels of sCD163 were measured in a cohort of 236 suspected tuberculosis (TB) cases from Guinea-Bissau, with a median follow-up period of 3.3 years (range 0-6.4 years). In 113 cases, the diagnosis of TB was verified by positive sputum microscopy and/or culture. Among the verified TB cases, a decreased survival rate was found in 27 patients with sCD163 levels above the upper reference limit (3.95 microg/mL). The difference in survival was significant during TB treatment (log rank, p<0.02) and after long-term follow-up (log rank, p<0.001). The decrease in survival rate during TB treatment remained significant in a multivariate Cox model controlling for human immunodeficiency virus (HIV) status, age and gender, with a mortality increase of 1.19 (95% CI, 1.04-1.36) per microg of sCD163, and a hazard ratio (HR) for sCD163 levels above the upper reference limit of 4.18 (95% CI, 1.06-16.4). The difference was not significant after excluding patients with concomitant HIV-1 and HIV-2 infection in Kaplan-Meier analyses (log rank, p 0.11). In contrast, the difference in survival remained significant in Kaplan-Meier analyses after long-term follow-up, even after excluding patients with concomitant HIV-1 and HIV-2 infection (log rank, p 0.002). In the Cox model, the mortality increase per microg of sCD163 was 1.27 (95% CI, 1.14-1.40), with an HR for elevated sCD163 levels of 2.85 (95% CI, 1.44-5.63). The HRs for concomitant HIV-1 and HIV-2 infection were 6.92 (95% CI, 3.28-14.58) and 2.48 (95% CI, 1.09-5.67), respectively. Thus, sCD163 levels appeared to be an independent predictor of survival in verified TB patients.