Circulating trimethyllysine and risk of acute myocardial infarction in patients with suspected stable coronary heart disease.

BACKGROUND: The carnitine precursor trimethyllysine (TML) is associated with progression of atherosclerosis, possibly through a relationship with trimethylamine-N-oxide (TMAO). Riboflavin is a cofactor in TMAO synthesis. We examined prospective relationships of circulating TML and TMAO with acute myocardial infarction (AMI) and potential effect modifications by riboflavin status. METHODS: By Cox modelling, risk associations were examined amongst 4098 patients (71.8% men) with suspected stable angina pectoris. Subgroup analyses were performed according to median plasma riboflavin. RESULTS: During a median follow-up of 4.9 years, 336 (8.2%) patients experienced an AMI. The age- and sex-adjusted hazard ratio (HR) (95% CI) comparing the 4th vs. 1st TML quartile was 2.19 (1.56-3.09). Multivariable adjustment for traditional cardiovascular risk factors and indices of renal function only slightly attenuated the risk estimates [HR (95% CI) 1.79 (1.23-2.59)], which were particularly strong amongst patients with riboflavin levels above the median (Pint  = 0.035). Plasma TML and TMAO were strongly correlated (rs  = 0.41; P < 0.001); however, plasma TMAO was not associated with AMI risk in adjusted analyses [HR (95% CI) 0.81 (0.58-1.14)]. No interaction between TML and TMAO was observed. CONCLUSION: Amongst patients with suspected stable angina pectoris, plasma TML, but not TMAO, independently predicted risk of AMI. Our results motivate further research on metabolic processes determining TML levels and their potential associations with cardiovascular disease. We did not adjust for multiple comparisons, and the subgroup analyses should be interpreted with caution.

Allo-HLA Cross-Reactivities of Cytomegalovirus-, Influenza-, and Varicella Zoster Virus-Specific Memory T Cells Are Shared by Different Healthy Individuals.

Virus-specific T cells can recognize allogeneic HLA (allo-HLA) through TCR cross-reactivity. The allospecificity often differs by individual (private cross-reactivity) but also can be shared by multiple individuals (public cross-reactivity); however, only a few examples of the latter have been described. Because these could facilitate alloreactivity prediction in transplantation, we aimed to identify novel public cross-reactivities of human virus-specific CD8+ T cells directed against allo-HLA by assessing their reactivity in mixed-lymphocyte reactions. Further characterization was done by studying TCR usage with primer-based DNA sequencing, cytokine production with ELISAs, and cytotoxicity with 51 chromium-release assays. We identified three novel public allo-HLA cross-reactivities of human virus-specific CD8+ T cells. CMV B35/IPS CD8+ T cells cross-reacted with HLA-B51 and/or HLA-B58/B57 (23% of tetramer-positive individuals), FLU A2/GIL (influenza IMP[58-66] HLA-A*02:01/GILGFVFTL) CD8+ T cells with HLA-B38 (90% of tetramer-positive individuals), and VZV A2/ALW (varicella zoster virus IE62[593-601] HLA-A*02:01/ALWALPHAA) CD8+ T cells with HLA-B55 (two unrelated individuals). Cross-reactivity was tested against different cell types including endothelial and epithelial cells. All cross-reactive T cells expressed a memory phenotype, emphasizing the importance for transplantation. We conclude that public allo-HLA cross-reactivity of virus-specific memory T cells is not uncommon and may create novel opportunities for alloreactivity prediction and risk estimation in transplantation.

Microbiota-dependent metabolite trimethylamine-N-oxide is associated with disease severity and survival of patients with chronic heart failure.

OBJECTIVES: Recent metabolomic, experimental and clinical studies have demonstrated that trimethylamine-N-oxide (TMAO), a microbiota-dependent metabolite from dietary phosphatidylcholine and carnitine, is a strong predictor of coronary artery disease (CAD). This finding suggests a link between the gut microbiota and atherosclerosis. The potential impact of TMAO in chronic heart failure (HF) is unknown. We hypothesized that TMAO levels would provide prognostic information about adverse outcomes in chronic HF. DESIGN: Prospective, observational study including 155 consecutive patients with chronic HF. In addition, 100 patients with stable CAD without HF and 33 matched healthy individuals were included as controls. Plasma levels of TMAO and its precursors choline and betaine were measured, and associations with symptoms, aetiology and transplant-free survival in the patients with HF were explored. RESULTS: Plasma levels of TMAO (P = 0.01), choline (P < 0.001) and betaine (P < 0.001) were elevated in patients with chronic HF compared to control subjects, with the highest levels in patients with New York Heart Association (NYHA) classes III and IV. Furthermore, TMAO levels were highest in individuals with ischaemic HF, followed by those with stable CAD and nonischaemic HF. TMAO, but not choline or betaine, was associated with reduced transplant-free survival: approximately 50% of patients in the upper tertile of TMAO levels died or received a heart transplant during 5.2 years of follow-up (unadjusted Cox-regression: hazard ratio 2.24, 95% confidence interval 1.28-3.92, P = 0.005). CONCLUSIONS: TMAO levels were elevated in patients with HF and associated with NYHA class, ischaemic aetiology and adverse outcomes. Future studies should focus on gut microbiota, dietary composition and intestinal dysfunction in relation to TMAO levels and clinical outcome in HF.

Fatty acid composition in chronic heart failure: low circulating levels of eicosatetraenoic acid and high levels of vaccenic acid are associated with disease severity and mortality.

OBJECTIVES: Free fatty acids (FFAs) are the major energy sources of the heart, and fatty acids (FAs) are active components of biological membranes. Data indicate that levels of FAs and their composition may influence myocardial function and inflammation. The aim of this study was to investigate whether total levels and composition of FAs and FFAs in plasma are altered in clinical heart failure (HF) and whether any alterations in these parameters are correlated with the severity of HF. SUBJECTS: Plasma from 183 patients with stable HF was compared with plasma from 44 healthy control subjects. RESULTS: Our main findings are as follows: (i) patients with HF had decreased levels of several lipid parameters and increased levels of FFAs in plasma, compared with controls, which were significantly correlated with clinical disease severity. (ii) Patients with HF also had a decreased proportion in the plasma of several n-3 polyunsaturated FAs, an increased proportion of several monounsaturated FAs, and a decreased proportion of some readily oxidized long-chain saturated FAs. (iii) These changes in FA composition were significantly associated with functional class, impaired cardiac function (i.e., decreased cardiac index and increased plasma N-terminal pro-B-type natriuretic peptide levels) and enhanced systemic inflammation (i.e., increased high-sensitivity C-reactive protein levels). (iv) Low levels of C20:4n-3 (eicosatetraenoic acid) and in particular high levels of C18:1n-7 (vaccenic acid) were significantly associated with total mortality in this HF population. CONCLUSIONS: Our data demonstrate that patients with HF are characterized by a certain FA phenotype and may support a link between disturbed FA composition and the progression of HF.

Dietary supplementation of tetradecylthioacetic acid increases feed intake but reduces body weight gain and adipose depot sizes in rats fed on high-fat diets.

AIM: The pan-peroxisome proliferator-activated receptor (PPAR) ligand and fatty acid analogue tetradecylthioacetic acid (TTA) may reduce plasma lipids and enhance hepatic lipid metabolism, as well as reduce adipose tissue sizes in rats fed on high-fat diets. This study further explores the effects of TTA on weight gain, feed intake and adipose tissue functions in rats that are fed a high-fat diet for 7 weeks. METHODS: The effects on feed intake and body weight during 7 weeks' dietary supplement with TTA ( approximately 200 mg/kg bw) were studied in male Wistar rats fed on a lard-based diet containing approximately 40% energy from fat. Adipose tissue mass, body composition and expression of relevant genes in fat depots and liver were measured at the end of the feeding. RESULTS: Despite higher feed intake during the final 2 weeks of the study, rats fed on TTA gained less body weight than lard-fed rats and had markedly decreased subcutaneous, epididymal, perirenal and mesenteric adipose depots. The effects of TTA feeding with reduced body weight gain and energy efficiency (weight gain/feed intake) started between day 10 and 13. Body contents of fat, protein and water were reduced after feeding lard plus TTA, with a stronger decrease in fat relative to protein. Plasma lipids, including Non-Esterified Fatty Acids (NEFA), were significantly reduced, whereas fatty acid beta-oxidation in liver and heart was enhanced in lard plus TTA-fed rats. Hepatic UCP3 was expressed ectopically both at protein and mRNA level (>1900-fold), whereas Ucp1 mRNA was increased approximately 30-fold in epididymal and approximately 90-fold in mesenteric fat after lard plus TTA feeding. CONCLUSION: Our data support the hypothesis that TTA feeding may increase hepatic fatty acid beta-oxidation, and thereby reduce the size of adipose tissues. The functional importance of ectopic hepatic UCP3 is unknown, but might be associated with enhanced energy expenditure and thus the reduced feed efficiency.

Tetradecylthioacetic acid attenuates dyslipidaemia in male patients with type 2 diabetes mellitus, possibly by dual PPAR-alpha/delta activation and increased mitochondrial fatty acid oxidation.

AIM: We previously demonstrated that a modified fatty acid, tetradecylthioacetic acid (TTA), improves transport and utilization of lipids and increases mitochondrial fatty acid oxidation in animal and cell studies. We conducted an exploratory study of safety and effects of this novel drug in patients with type 2 diabetes mellitus and investigated the mechanism of action in human cell lines. METHODS: Sixteen male patients with type 2 diabetes mellitus received 1 g TTA daily for 28 days in an open-labelled study, with measurement of parameters of lipid metabolism, glucose metabolism and safety (ClinicalTrials.gov NCT00605787). The mechanism of action was further investigated in a human liver cell line (HepG2) and in cultured human skeletal muscle cells (myotubes). RESULTS: Mean LDL cholesterol level declined from 4.2 to 3.7 mmol/l (p < 0.001), accompanied by increased levels of the HDL apolipoproteins A1 and A2, and a decline in LDL/HDL ratio from 4.00 to 3.66 (p = 0.008). Total fatty acid levels declined, especially the fraction of the polyunsaturated n-3 fatty acids docosahexaenoic acid (-13%, p = 0.002) and eicosapentaenoic acid (-10%, p = 0.07). Glucose metabolism was not altered and the drug was well tolerated. In cultured liver cells, TTA acted as a pan-PPAR agonist with predominant PPAR-alpha and PPAR-delta activation at low TTA concentrations. In myotubes, TTA and a PPAR-delta agonist, but not the PPAR-alpha or PPAR-gamma agonists, increased the fatty acid oxidation. CONCLUSIONS: We demonstrate for the first time that TTA attenuates dyslipidaemia in patients with type 2 diabetes mellitus. These effects may occur through mechanisms involving PPAR-alpha and PPAR-delta activation, resulting in increased mitochondrial fatty acid oxidation.

Increased survival by feeding tetradecylthioacetic acid during a natural outbreak of heart and skeletal muscle inflammation in S0 Atlantic salmon, Salmo salar L.

We have previously documented increased survival by feeding tetradecylthioacetic acid (TTA) during a natural outbreak of infectious pancreatic necrosis in post-smolt S1 Atlantic salmon. The aim of the present study was to test the effects of dietary TTA in S0 smolt at a location where fish often experience natural outbreaks of heart and skeletal muscle inflammation (HSMI) during their first spring at sea. The experimental groups were fed a diet supplemented with 0.25% TTA for a 6-week period prior to a natural outbreak of HSMI in May 2007. Relative percent survival for the groups fed TTA was 45% compared with control diets, reducing mortality from 4.7% to 2.5%. Expression of genes related to lipid oxidation was higher in cardiac ventricles from salmon fed TTA compared with controls. In addition, salmon fed TTA had periodically reduced levels of plasma urea, and increased cardiosomatic index and growth. Reduced mortality and increased growth after administration of TTA may be related to a combination of anti-inflammatory effects, and an altered metabolic balance with better protein conservation because of increased lipid degradation.

Effect of fish oil supplementation and restricted feeding on body fat distribution and blood lipid profile in a rabbit model of castration-induced obesity.

This study investigates the effect of fish oil supplementation and restricted feeding on body fat distribution and blood lipid profile in experimentally induced obesity in rabbits. The trial was carried out with 30 male rabbits, divided into 5 groups of 6 animals each (NC - non-castrated, non-treated, full-diet fed; C100 - castrated, non-treated, full-diet fed; FO100 - castrated, treated with fish oil, full-diet fed; C50 - castrated, non-treated, 50% restricted fed; FO50 - castrated, treated with fish oil, 50% restricted fed). At the end of the experiment, plasma lipids measurement and quantification of fat distribution was performed. The results of this study indicate that fish oil supplementation reduces obesity-associated abnormalities in lipid profile (high-density lipoprotein cholesterol to low-density lipoprotein cholesterol ratio and non-esterified fatty acids) and in body fat distribution in full-diet fed rabbits. Restricted feeding (C50) alone and the combination of restricted feeding and fish oil supplementation (FO50) in particular, has a detrimental effect on the lipid profile despite the marked reduction in intra-abdominal fat.

Rifaximin alters gut microbiota profile, but does not affect systemic inflammation - a randomized controlled trial in common variable immunodeficiency.

Common variable immunodeficiency (CVID) patients have reduced gut microbial diversity compared to healthy controls. The reduced diversity is associated with gut leakage, increased systemic inflammation and ten "key" bacteria that capture the gut dysbiosis (dysbiosis index) in CVID. Rifaximin is a broad-spectrum non-absorbable antibiotic known to reduce gut leakage (lipopolysaccharides, LPS) in liver disease. In this study, we explored as a 'proof of concept' that altering gut microbial composition could reduce systemic inflammation, using CVID as a disease model. Forty adult CVID patients were randomized, (1:1) to twice-daily oral rifaximin 550 mg versus no treatment for 2 weeks in an open-label, single-centre study. Primary endpoints were reduction in plasma/serum levels of soluble (s) CD14, sCD25, sCD163, neopterin, CRP, TNF, LPS and selected cytokines measured at 0, 2 and 8 weeks. Secondary endpoint was changes in intra-individual bacterial diversity in stool samples. Rifaximin-use did not significantly change any of the inflammation or gut leakage markers, but decreased gut microbial diversity compared with no treatment (p = 0.002). Importantly, the gut bacteria in the CVID dysbiosis index were not changed by rifaximin. The results suggest that modulating gut microbiota by rifaximin is not the chosen intervention to affect systemic inflammation, at least not in CVID.

[Late complications following renal transplantation]. / Late complicaties na niertransplantatie.

The number of renal transplant recipients is increasing steadily. Physicians from all specialties are ever more likely to encounter this vulnerable group of patients. They constitute a susceptible group because of increased mortality and morbidity. Half of the renal transplants are lost due to chronic transplant failure. The primary cause of chronic transplant failure is chronic allograft nephropathy. Other causes of transplant failure are calcineurin inhibitor toxicity, recurrence of the original renal disease such as glomerulonephritis and diabetes mellitus, stenosis of the renal artery in the transplant, and urological complications. The other half of the renal transplants are lost due to the death of the recipient. The primary cause of death is cardiovascular disease due to former chronic renal, hypertension and dyslipidemia following the use of immunosuppressants. In addition malignancies, infections and bone abnormalities do occur more frequently as compared to the normal populations. Alertness is warranted following kidney transplantation by both the patients themselves as well as all the treating specialists. Careful periodical monitoring for life is required because of the risk of the abovementioned complications.

Ultrasound measurement of anatomical parameters of the upper airway in adults. / Medición ecográfica de parámetros anatómicos de la vía aérea superior en adultos.

BACKGROUND: Most of the works on ultrasound airway anatomy are limited to a morphological description. A study was conducted in order to provide an objective normal range of measurements. MATERIALS AND METHODS: Observational study to describe the ultrasound characteristics of the upper airway in adults without clinical difficult airway criteria, compared to cadaver dissection anatomical models. RESULTS: The study included 45 volunteers (27 men and 18 women), and 3fresh cadavers. The quality of the examination was very good/good in 93% of the cases. MEASUREMENTS: tracheal diameter (1.3±0.3cm), vocal cord (1.6±0.5cm), cricothyroid membrane (0.94±0.32cm), cricotracheal membrane (0.3±0.09cm), thickness of the muscles in the floor of the mouth (MFM) (1.5±0.26cm), sub-mandibular subcutaneous fat plus MFM (2.11±0.34cm), hyoid-mandible distance (5.35±0.69cm), palate-floor of the mouth distance (4.92±0.5cm), palate-anterior border of the mandible (5.51±0.7cm), and palate-pharynx angle (114±14). Observed differences: Males had a larger tracheal diameter than females (M: 1.4±0.3 vs. F: 1.2±0.2cm, p=0.014). Subject height showed a significant correlation with the tracheal diameter (R: 0.501, p<0.001), as well as the length of the vocal cord (R: 0.363, p=0.016), the thickness of MFM (R=0.299, p=0.046) as well as the hyoid-mandible (R: 0.556; p<0.001) and palate-mandible distances (R: 0.362; p=0.015). CONCLUSIONS: Ultrasound allows the anatomy of upper airway to be evaluated, as well as to calculate distances between the anatomical structures. The study defines these distances in adult volunteers without clinical difficult airway criteria.

[Immunological defense to viral infections: focus on virus-specific T cells]. / Immunologische afweer tegen virale infecties: virusspecifieke T-cellen in beeld.

T cells play a central role in the control of and the protection against viral infections. The T cell population consists ofa diversity ofvirus-specific memory T cells. The characteristics of these T cells seem to depend largely on the type of virus for which they are specific. T cells directed against latent viruses, such as cytomegalovirus, are cytotoxic cells. T cells directed against viruses that after the initial infection are completely removed by the immune system, such as the influenza virus, are non-cytotoxic cells. The development of new immunological techniques, such as the detection of virus-specific cells with HLA-peptide tetrameric complexes, enables the characterization of the properties of virus-specific T cells in the blood and organs.

A predictive test for difficult intubation in laryngeal microsurgery. Validation study. / Test predictivo de intubación difícil en microcirugía de laringe. Estudio de validación.

BACKGROUND: In 2003 a simple test to detect difficult intubation (DI) in patients underwent laryngeal microsurgery was described. The present study was conducted to validate the test. METHODS: Patients with laryngeal disease scheduled for laryngeal microsurgery were included. The variables evaluated were eleven parameters: Mallampati Score ≥III, thyromental distance <6.5cm, mouth opening <40mm, limitation of the temporomandibular joint, pathological dentition, arched paladet, maxillary deficiency, neck mobility <90°, body mass index, symptoms of laryngeal dysfunction, and tumour of supraglottic region. A rapid examination of the airway by an expert anaesthesiologist was carried out just before induction. DI was defined by a Cormack laryngeal view gradeiii-iv and/or if auxiliary equipment was required to achieve orotracheal intubation. A comparison of ROC curves was conducted to estimate the best predicting cut-off, as well as differences between the index described in 2003 and the rapid assessment by an expert anaesthesiologist. RESULTS: One hundred and five patients were included in the study. The incidence of DI according to the predefined criteria was 45% (47 patients). The score at the cut-off of 5 provides a sensitivity of 89% and a specificity of 55%. The fast-evaluation by an expert anaesthesiologist obtained similar sensitivity (89%) and better specificity (81%). Area under curve (AUC) was 0.87, similar to a fast evaluation (AUC: 0.85; P=.71). CONCLUSION: The index was validated at a cut-off of ≥5, being a useful and reliable test to predict DI in patients with laryngeal diseases and it is a useful tool for anaesthesiologists with conventional training.

Ultrasound-guided genicular nerve block for pain control after total knee replacement: Preliminary case series and technical note. / Bloqueo ecoguiado de los nervios geniculados en el manejo analgésico de la artroplastia de rodilla: descripción de la técnica y resultados clínicos preliminares.

INTRODUCTION: Total knee arthroplasty (TKA) is an operation with moderate to severe postoperative pain. The Fast-Track models employ local infiltration techniques with anaesthetics at high volumes (100-150ml). We proposed a genicular nerve block with low volume of local anaesthetic. The aim of our study is to evaluate the periarticular distribution of these blocks in a fresh cadaver model and to describe the technique in a preliminary group of patients submitted to TKA. MATERIALS AND METHODS: In the anatomical phase, 4 genicular nerves (superior medial, superior lateral, inferior medial and inferior lateral) were blocked with 4ml of local anaesthetic with iodinated contrast and methylene blue in each (16ml in total). It was performed on a fresh cadaver and the distribution of the injected medium was evaluated by means of a CT-scan and coronal anatomical sections on both knees. The clinical phase included 12 patients scheduled for TKA. Ultrasound-guided block of the 4 genicular nerves was performed preoperatively and their clinical efficacy evaluated by assessing pain after the reversal of the spinal block and at 12h after the block. Pain was measured using the numerical scale and the need for rescue analgesia was evaluated. RESULTS: A wide periarticular distribution of contrast was observed by CT-scan, which was later evaluated in the coronal sections. The distribution followed the joint capsule without entering the joint, both in the femur and in the tibia. The pain after the reversal of the subarachnoid block was 2±1, requiring rescue analgesia in 42% of the patients. At 12h, the pain according to the numerical scale was 4±1, 33% required rescue analgesia. CONCLUSION: The administration of 4ml of local anaesthetic at the level of the 4 genicular nerves of the knee produces a wide periarticular distribution. Our preliminary data in a series of 12 patients undergoing TKA seems to be clinically effective. Nevertheless, extensive case series and comparative studies with local infiltration techniques with anaesthetics are needed to support these encouraging results.

3D ultrasound estimation of the effective volume for popliteal block at the level of division. / Estimación del volumen de anestésico local en contacto con el nervio ciático en el bloqueo poplíteo.

Local anaesthetic injection between the tibial and commmon peroneal nerves within connective tissue sheath results in a predictable diffusion and allows for a reduction in the volume needed to achieve a consistent sciatic popliteal block. Using 3D ultrasound volumetric acquisition, we quantified the visible volume in contact with the nerve along a 5cm segment. METHODS: We included 20 consecutive patients scheduled for bunion surgery. Ultrasound guided popliteal block was performed using a posterior, out of plane approach at the level of división of the sciatic nerve. Thirty ml of mepivacaine 1.5% and levobupivacaine 0.5% were slowly injected while assessing the injection pressure and the diffusion of the local anaesthetic. Volumetric acquisition was performed before and after the block to quantify the the volume of the sciatic nerve and the volume of the surrounding hypoechoic halo contained inside the connective tissue in a 5cm segment. RESULTS: All blocks were successful within 20min after the injection. The total estimated volume contained inside the common connective tissue sheath was 6.8±2.6cm3. Of this, the volume of the halo sorrounding the nerve was 4.4±1.7cm3 and the volume inside the sciatic nerve was 2.4±1.7cm3. CONCLUSIONS: The volume of local anaesthetic in close contact with the sciatic nerve can be estimated by volumetric acquisition. Our results suggest that the effective volume of local anaesthetic needed for a successful sciatic popliteal block could be reduced to less than 7ml.

Cost-effectiveness of post-operative cell salvage in total knee arthroplasty. Should we continue to recommend its use today? / Análisis de coste-eficacia del recuperador postoperatorio de sangre de drenajes en cirugía protésica primaria de rodilla. ¿Debemos seguir recomendando su empleo en la actualidad?

OBJECTIVES: Total knee arthroplasty (TKA) has a high transfusion rate. In our protocol, the use of postoperative cell salvage is indicated in patients with contraindications to tranexamic acid (TA). An analysis was performed on the effect of post-operative cell salvage (POCS) regarding transfusion rate and costs in patients undergoing TKA. MATERIAL AND METHODS: A prospective analysis was conducted on 518 patients, of whom 434 received TA, and 84 were contraindicated. The red cell mass, blood volume, and the percentage of lost blood volume were calculated. Incidents associated with the use of post-operative re-perfusion of drained blood and the rate of transfusion were recorded. An analysis was performed on the costs associated with allogeneic transfusion prevention methods. RESULTS: A POCS drain was not inserted in 10 out of the 84 patients not candidates for TA. In the 74 in which it was placed, 158±72ml of red cell mass was reinfused. The allogeneic transfusion rate was 36%, and was 52% in those with no drain inserted. Relative risk of transfusion using POCS was 0.69 (0.41 to 1.16) with an absolute risk reduction of 16% (-8 to 40%). The number needed to treat to avoid allogeneic transfusion was 7. The direct costs to avoid allogeneic transfusion were €1,610. No complications associated with blood re-infusion were observed. CONCLUSIONS: The use of POCS would be required in 7 patients after TKA to avoid one allogeneic transfusion with a cost over 10 times that of a transfusion of red cell concentrates.

Altered gut microbiota profile in common variable immunodeficiency associates with levels of lipopolysaccharide and markers of systemic immune activation.

Common variable immunodeficiency (CVID) is the most common symptomatic primary immunodeficiency characterized by low immunoglobulin (Ig)G and IgA, and/or IgM. In addition to bacterial infections, a large subgroup has noninfectious inflammatory and autoimmune complications. We performed 16S ribosomal RNA-based profiling of stool samples in 44 CVID patients, 45 patients with inflammatory bowel disease (disease controls), and 263 healthy controls. We measured plasma lipopolysaccharide (LPS) and markers of immune cell activation (i.e., soluble (s) CD14 and sCD25) in an expanded cohort of 104 patients with CVID and in 30 healthy controls. We found a large shift in the microbiota of CVID patients characterized by a reduced within-individual bacterial diversity (alpha diversity, P<0.001) without obvious associations to antibiotics use. Plasma levels of both LPS (P=0.001) and sCD25 (P<0.0001) were elevated in CVID, correlating negatively with alpha diversity and positively with a dysbiosis index calculated from the taxonomic profile. Low alpha diversity and high dysbiosis index, LPS, and immune markers were most pronounced in the subgroup with inflammatory and autoimmune complications. Low level of IgA was associated with decreased alpha diversity, but not independently from sCD25 and LPS. Our findings suggest a link between immunodeficiency, systemic immune activation, LPS, and altered gut microbiota.

Purification and characterization of a long-chain acyl-CoA hydrolase from rat liver microsomes.

A long-chain acyl-CoA hydrolase from rat liver microsomes has been purified by solvent extraction and gel chromatography to homogeneity as judged by polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate. The enzyme was a monomer of molecular weight 59 000. In a sucrose gradient it sedimented at 4.3 S. The isoelectric point, pI was 6.9, and the Stokes radius was approx. 31 A. The enzyme hydrolyzed long-chain fatty acyl-CoA (C7--C18) with maximum activity for palmitoyl-CoA. Bovine serum albumin activation of the enzyme was related to the ratio acyl-CoA/bovine serum albumin, and at high ratios, acyl-CoA inhibited the enzyme activity. Disregarding the substrate inhibition, an apparent Km of 65 nmol/mg protein or 1-10(-7) M and a V of 750 nmol/mg protein per min were calculated. The enzyme was inhibited by p-hydroxymercuribenzoate and N-ethylmaleimide. Reactivation by means of dithiothreitol was not complete.

Correlation between the cellular level of long-chain acyl-CoA, peroxisomal beta-oxidation, and palmitoyl-CoA hydrolase activity in rat liver. Are the two enzyme systems regulated by a substrate-induced mechanism?

Data obtained in earlier studies with rats fed diets containing high doses of peroxisome proliferators (niadenate, tiadenol, clofibrate, or nitotinic acid) are used to look for a quantitative relationship between peroxisomal beta-oxidation, palmitoyl-CoA hydrolase, palmitoyl-CoA synthetase and carnitine palmitoyltransferase activities, and the cellular concentration of their substrate and reaction products. The order of the hyperlipidemic drugs with regard to their effect on CoA derivatives and enzyme activities was niadenate greater than tiadenol greater than clofibrate greater than nicotinic acid. Linear regression analysis of long-chain acyl-CoA content versus palmitoyl-CoA hydrolase and peroxisomal beta-oxidation activity showed highly significant linear correlations both in the total liver homogenate and in the peroxisome-enriched fractions. A dose-response curve of tiadenol showed that carnitine palmitoyltransferase and palmitoyl-CoA synthetase activities and the ratio of long-chain acyl-CoA to free CoASH in total homogenate rose at low doses before detectable changes occurred in the peroxisomal beta-oxidation and palmitoyl-CoA hydrolase activity. A plot of this ratio parallelled the palmitoyl-CoA synthetase activity. The specific activity of microsomally localized carnitine palmitoyl-transferase was low and unchanged up to a dose where no enhanced peroxisomal beta-oxidation was observed, but over this dose the activity increased considerably so that the specific of the enzyme in the mitochondrial and microsomal fractions became comparable. The mitochondrial palmitoyl-CoA synthetase activity decreased gradually. The correlations may be interpreted as reflecting a common regulation mechanism for palmitoyl-CoA hydrolase and peroxisomal beta-oxidation enzymes, i.e., the cellular level of long-chain acyl-CoA acting as the metabolic message for peroxisomal proliferation resulting in induction of peroxisomal beta-oxidation and palmitoyl-CoA hydrolase activity. The findings are discussed with regard to their possible consequences for mitochondrial fatty acid oxidation and the conversion of long-chain acyl-L-carnitine to acyl-CoA derivatives.