Cluster Of Differentiation 14 Research Articles

CD14 blockade preserves left ventricular structure and systolic function in a murine model of ST-elevation myocardial infarction

Abstract Background Cluster of differentiation-14 (CD14) facilitates the excessive pro-inflammatory monocyte-macrophage response to myocardial injury. Both acute and chronic infiltration of pro-inflammatory macrophages are implicated in the immunological progression of ischemic heart failure. However, investigation of clinically practicable strategies of CD14 blockade are yet to be reported. Purpose This fully blinded and randomised preclinical study evaluated the efficacy of a neutralising anti-CD14 antibody given at reperfusion in a murine model of reperfused ST-elevation myocardial infarction (STEMI) with key clinical features. Methods Large anterior STEMI was induced in 48 adult mice by left anterior descending coronary artery occlusion for 1 h prior to reperfusion and randomisation to receive anti-CD14 antibody, isotype control (IgG2a) or saline weekly for four weeks (independently blinded). Outcome measures included functional assessments by 9.4T cardiac magnetic resonance (CMR) imaging and ultra-high-resolution echocardiography at 21- and 28-days post-STEMI, respectively. Results Multi-plane short axis CMR at 21-days post-STEMI revealed greater left ventricular (LV) ejection fraction in the anti-CD14 antibody-treated group compared with saline- and isotype-treated controls (30±1 vs 19±1 and 20±1 %, respectively, p<0.001; Figure 1A). CMR also revealed reduced LV dilatation with anti-CD14-treatment compared to control groups (76±5 vs 102±8 and 107±7 µl EDV in saline- and isotype-treated controls, respectively, p<0.05). Parasternal long axis echocardiography at 28 days post-STEMI confirmed that anti-CD14 antibody treatment was associated with greater LV systolic function compared to saline and isotype controls (30±1 vs 24±1 and 25±2 % EF, respectively, p<0.05; Figure 1B). Preservation of LV end-diastolic (86±4 vs 111±6 and 101±5 µl, p<0.01) and end-systolic (61±3 vs 84±5 and 77±5 µl, p<0.01) volumes was also observed with anti-CD14 treatment. Global longitudinal strain (GLS) of the LV was also significantly greater at 28 days post-STEMI with anti-CD14 treatment relative to saline and isotype controls (-8.5±0.4 vs -6.8±0.3 and -6.6±0.4 %, respectively, p<0.05). Conclusion(s) The findings of this fully blinded and randomised preclinical study provide evidence of a clinically practicable strategy of CD14 blockade initiated at reperfusion to mitigate progressive LV systolic dysfunction and dilatation post-STEMI.Figure 1.Functional outcome measures

Reducing dietary lipopolysaccharides to slow progression of cognitive impairment and Alzheimer’s disease

We hypothesize that a dietary reduction of lipopolysaccharide (LPS) could reduce risk and slow progression of cognitive impairment and Alzheimer’s disease by reducing neuroinflammation. LPS is a component of the highly inflammatory cell membrane of Gram-negative bacteria. LPS and inflammatory cytokines are elevated in Alzheimer’s disease, both in plasma and in the central nervous system. Inflammatory cytokines, triggered by LPS in food, can enter the brain via active transport and induce microglial activation and inflammatory damage, including loss of synapses and neurons. Excess LPS and inflammatory cytokines can also induce blood-brain barrier damage and increase amyloid-β production. The highest dietary sources of LPS are processed meat and dairy products, which can rapidly raise levels of plasma LPS within one to two hours. For example, there was a 65-fold increase in plasma LPS after eating a thin-crust cheese pizza. The LPS in these foods is transported via chylomicrons into the bloodstream. Smaller amounts of LPS from colonic microbiota do not account for the sharp postprandial increase of plasma LPS. Once in the bloodstream, LPS can increase inflammatory cytokines 2-fold to 68-fold within two hours. LPS increases cytokines via LPS binding protein and cluster of differentiation-14, triggering toll-like receptor-4, and activating nuclear factor kappa-B. Nuclear factor kappa-B can increase cytokine levels, including interleukin-1-beta, interleukin-6, and tumor necrosis factor-alpha. Practical implications: Even low plasma levels of LPS can reduce cognition. Decreasing brain inflammation by reducing dietary LPS can lower risk of cognitive impairment and dementia.

Spatial confined enzyme-linked immunoassay for ultrasensitive biomarker detection using a microfluidic magnetic delivery disc

Herein, a spatial confined enzyme-linked immunoassay (ELIA) is developed for ultrasensitive biomarker detection using a microfluidic magnetic delivery (μMD) disc. In μMD-ELIA, magnetic immunocomplexes are delivered into microchannel and trapping/preconcentration in the microchamber of μMD disc for fluorescence detection. The microchamber offers a confined space both for effective magnetic preconcentration and accelerated concentration of enzyme products. After μMD disc optimization, the assay performances of μMD-ELIA have been evaluated by model targets of human monocyte differentiation antigen cluster of differentiation14 (CD14) and interleukin (IL)–8. The limits of detection (LOD) of μMD-ELIA for CD14 and IL-8 are 0.17 pg/mL and 0.012 pg/mL, respectively, which corresponded to an over 300-fold LOD sensitivity enhancement compared to their standard ELISA results. In human serum analysis, good accuracy is obtained on both CD14 and IL-8 when compared to the results of ELISA kits, and satisfied recoveries of 81.3–112 % with RSDs of 1.0–9.7 % for IL-8 were obtained. Moreover, the μMD-ELIA is than applied to IL-6 concentration variation monitor in rat plasma during the progression of inflammatory reaction successfully, which further confirmed that the μMD-ELIA has enormous potential in disease early diagnosis and curative effect monitoring.

Dioleoylphosphatidylglycerol Inhibits Heat Shock Protein B4 (HSPB4)-Induced Inflammatory Pathways In Vitro.

Our previous work shows that dioleoylphosphatidylglycerol (DOPG) accelerates corneal epithelial healing in vitro and in vivo by unknown mechanisms. Prior data demonstrate that DOPG inhibits toll-like receptor (TLR) activation and inflammation induced by microbial components (pathogen-associated molecular patterns, PAMPs) and by endogenous molecules upregulated in psoriatic skin, which act as danger-associated molecular patterns (DAMPs) to activate TLRs and promote inflammation. In the injured cornea, sterile inflammation can result from the release of the DAMP molecule, heat shock protein B4 (HSPB4), to contribute to delayed wound healing. Here, we show in vitro that DOPG inhibits TLR2 activation induced in response to HSPB4, as well as DAMPs that are elevated in diabetes, a disease that also slows corneal wound healing. Further, we show that the co-receptor, cluster of differentiation-14 (CD14), is necessary for PAMP/DAMP-induced activation of TLR2, as well as of TLR4. Finally, we simulated the high-glucose environment of diabetes to show that elevated glucose levels enhance TLR4 activation by a DAMP known to be upregulated in diabetes. Together, our results demonstrate the anti-inflammatory actions of DOPG and support further investigation into its development as a possible therapy for corneal injury, especially in diabetic patients at high risk of vision-threatening complications.

Evaluation of miRNA-7, miRNA-10 and miRNA-21 as diagnostic non-invasive biomarkers of hepatocellular carcinoma.

Liver cancer (hepatocellular carcinoma - HCC) remains a serious health challenge; it is the fourth leading cause of death worldwide. Egypt ranks fifteenth worldwide and the third in Africa in terms of HCC burden. The present study aimed to assess some microRNAs (miRNAs) including miRNA-7, miRNA-10, and miRNA-21, serum markers such as cluster of differentiation-14 (CD-14) and transforming growth factor b1 (TGF-b1), and other biochemical parameters as non-invasive tools for HCC diagnosis. The study included 100 participants divided into five groups: group I (20 normal subjects as a healthy group), group II (20 participants with chronic HCV infection but non-cirrhotic), group III (20 volunteers with chronic HCV infection and compensated cirrhosis), group IV (20 patients with chronic HCV infection and decompensated cirrhosis), and group V (20 participants with HCC). Levels of miR-7, miR-10, and miR-21 were evaluated using qRT-PCR. Serum ALT, AST, total bilirubin, total protein, albumin, PT, INR, and platelet count were determined. FIB-4 and APRI test levels were also calculated. CD-14 and TGF-β1 serum levels were estimated using enzyme-linked immunosorbent assay (ELISA) kits. The expression levels of miR-21 followed by miR-10 showed high sensitivity and specificity in predicting HCC. Serum CD-14 and TGF-b1 levels were significantly increased in all patient groups. From the study, it is concluded that the expression level of miR-21 has the highest sensitivity and specificity, followed by miR-10, which has high sensitivity and low specificity as non-invasive markers for HCC detection, while miR-7 exhibits high sensitivity and reasonable specificity in fibrosis detection.

Ameliorative effects of supranutritional selenium on TLR-4-NF-kB-TNF-α-mediated hepatic oxidative injury and inflammation in goats fed high concentrate diet.

We examined whether surplus dietary selenium (Se) supply could alleviate high concentrate (HC) diet-induced hepatic oxidative stress (OS) and inflammation. Eighteen young goats were distributed into three groups; were fed low (LC, concentrate: forage; 35: 65), high concentrate (HC, 65: 35), or Se-supplemented HC (HCSe, 65: 35 +0.5mg Se kg-1 diet) diets for 10 weeks. Short chain fatty acids, OS markers and immunoinflammatory genes expressions were assessed through gas chromatograph, kits, and RT-qPCR, respectively. Compared with LC, HC diet increased (p< .05) colonic and serum lipopolysaccharide (LPS) levels and induced hepatic oxidative injury by increasing (p< .05) malondialdehyde (MDA) levels and decreasing (p< .05) activities of glutathione peroxidase, superoxide dismutase, and catalase. HC diet altered hepatic mRNA expressions of toll-like receptor-4 (TLR-4), cluster of differentiation-14 (CD-14), tumor necrosis factor-α (TNF-α), TNF receptor-associated factor-6 (TRAF-6), nuclear factor kappa B (NF-κB), interleukin-1β (IL-1β), IL-10, IL-13, LPS-binding protein (LBP), serum amyloid A (SAA), α-acid glycoprotein (AGP), and albumin (ALB). Conversely, extra-Se supply lowered LPS and attenuated antioxidant status and inflammation in liver. In conclusion, HC diet induced oxidative lesions and TLR-4 pathway-mediated inflammation, whereas supranutritional Se alleviated oxidative and inflammatory lesions through TLR-4 pathway regulation in goat liver.

Sexual Dimorphic Role of CD14 (Cluster of Differentiation 14) in Salt-Sensitive Hypertension and Renal Injury.

Genomic sequence and gene expression association studies in animals and humans have identified genes that may be integral in the pathogenesis of various diseases. CD14 (cluster of differentiation 14)-a cell surface protein involved in innate immune system activation-is one such gene associated with cardiovascular and hypertensive disease. We previously showed that this gene is upregulated in renal macrophages of Dahl salt-sensitive animals fed a high-salt diet; here we test the hypothesis that CD14 contributes to the elevated pressure and renal injury observed in salt-sensitive hypertension. Using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9), we created a targeted mutation in the CD14 gene on the Dahl SS (SS/JrHSDMcwi) background and validated the absence of CD14 peptides via mass spectrometry. Radiotelemetry was used to monitor blood pressure in wild-type and CD14-/- animals challenged with high salt and identified infiltrating renal immune cells via flow cytometry. Germline knockout of CD14 exacerbated salt-sensitive hypertension and renal injury in female animals but not males. CD14-/- females demonstrated increased infiltrating macrophages but no difference in infiltrating lymphocytes. Transplant of CD14+/+ or CD14-/- bone marrow was used to isolate the effects of CD14 knockout to hematopoietic cells and confirmed that the differential phenotype observed was due to knockout of CD14 in hematopoietic cells. Ovariectomy was used to remove the influence of female sex hormones, which completely abrogated the effect of CD14 knockout. These studies provide a novel treatment target and evidence of a new dichotomy in immune activation between sexes within the context of hypertensive disease where CD14 regulates immune cell activation and renal injury.

Increased risk of hypercholesterolemia in a French and Lebanese population due to an interaction between rs2569190 in CD14 and gender

RationaleSince identifying gender-specific genetic associations may have a significant impact on public health, we studied the interaction between rs2569190 in CD14 (cluster of differentiation 14) and gender in relation to the lipid traits in two independent populations. MethodsWe first tested the interaction in a discovery population (SFS, n = 956), then replicated it in an independent population (LGP, n = 460), followed by a meta-analysis (n = 1,416). Finally, stratification according to gender was conducted to test the association between rs2569190 and lipid traits. Binary multiple logistic regression models were used while correcting for many confounders. Power calculations were also performed. ResultsAn interaction between rs2569190 and gender, which increased the risk of total cholesterol levels in SFS, was found (OR = 2.151 and P = 0.05). This interaction was further replicated in the LGP (OR = 1.353 and P < 0.001), and the meta-analysis showed an overall significant interaction (OR = 1.436 and Pmeta = 0.02). Similarly, the meta-analysis showed an overall significant positive effect (OR = 1.204 and Pmeta = 0.004) for low-density lipoprotein cholesterol levels. Overall, 1,416 patients were evaluated, and the statistical heterogeneity was low, with I2 estimates ranging between 0% and 22.2%. In contrast, rs2569190 in CD14 did not show any significant interaction with gender influencing high-density lipoprotein levels and triglycerides levels in both populations. ConclusionAn interaction between rs2569190 in CD14 and gender increased the risk of hypercholesterolemia in two independent populations with a gender-specific effect in males.

Heparanase-2 protects from LPS-mediated endothelial injury by inhibiting TLR4 signalling

The endothelial glycocalyx and its regulated shedding are important to vascular health. Endo-β-D-glucuronidase heparanase-1 (HPSE1) is the only enzyme that can shed heparan sulfate. However, the mechanisms are not well understood. We show that HPSE1 activity aggravated Toll-like receptor 4 (TLR4)-mediated response of endothelial cells to LPS. On the contrary, overexpression of its endogenous inhibitor, heparanase-2 (HPSE2) was protective. The microfluidic chip flow model confirmed that HPSE2 prevented heparan sulfate shedding by HPSE1. Furthermore, heparan sulfate did not interfere with cluster of differentiation-14 (CD14)-dependent LPS binding, but instead reduced the presentation of the LPS to TLR4. HPSE2 reduced LPS-mediated TLR4 activation, subsequent cell signalling, and cytokine expression. HPSE2-overexpressing endothelial cells remained protected against LPS-mediated loss of cell-cell contacts. In vivo, expression of HPSE2 in plasma and kidney medullary capillaries was decreased in mouse sepsis model. We next applied purified HPSE2 in mice and observed decreases in TNFα and IL-6 plasma concentrations after intravenous LPS injections. Our data demonstrate the important role of heparan sulfate and the glycocalyx in endothelial cell activation and suggest a protective role of HPSE2 in microvascular inflammation. HPSE2 offers new options for protection against HPSE1-mediated endothelial damage and preventing microvascular disease.

Genetic polymorphisms in pattern recognition receptors are associated with allergic diseases through gene-gene interactions.

There is evidence that suggests variation in gene encoding pattern recognition receptors, the essential components of innate immunity, might be associated with atopic diseases. However, results have been inconclusive. The aim of the study was to determine the individual associations and possible interactive effects of the CD14 (cluster of differentiation 14), TLR4 (toll-like receptor 4) and TLR2 (toll-like receptor 2) polymorphisms on allergic diseases. The CD14 C-159T, TLR4 +896A/G and TLR2 A-16934T polymorphisms were identified in 115 children aged from 6 to 17 years. All subjects were selected using a detailed questionnaire which included questions on symptoms and each one underwent skin prick testing. All single-nucleotide polymorphisms (SNPs) were determined using real-time polymerase chain reaction (PCR) assays. There was no statistically significant correlation between the 3 polymorphisms (CD14 C-159T, TLR4 +896A/G and TLR2/-16934A/T) and either asthma, allergic rhinitis or atopy. We observed that children who were heterozygous or homozygous for both the CD14/-159T and TLR2/-16934A alleles had a 4-fold lower risk for asthma than children who were carriers of the T allele of CD14 but non-carriers of the A allele of TLR2, and an almost 3-fold lower risk for asthma when compared to all other groups. Concerning allergic rhinitis, a similar trend was observed. In addition, the presence of at least 1 A allele in the TLR2/-16934 polymorphism reduced the risk for asthma and allergic rhinitis, but only in children who were homozygous for the common A allele in the TLR4 +896 polymorphism. Our study supports the idea that the CD14, TLR2 and TLR4 polymorphisms may not be directly involved in the development of atopic diseases. However, our results suggest that their impact on the risk of asthma and allergic rhinitis might be modulated by gene-gene interactions.

Human αS1-casein induces IL-8 secretion by binding to the ecto-domain of the TLR4/MD2 receptor complex

BackgroundThe milk protein αS1-casein was recently reported to induce secretion of proinflammatory cytokines via Toll-like receptor 4 (TLR4). In this study, αS1-casein was identified as binder of theTLR4 ecto domain. MethodsIL-8 secretion after stimulation of TLR4/MD2 (myeloid differentiation factor 2)/CD14 (cluster of differentiation 14)-transfected HEK293 cells (TLR4+) and Mono Mac 6 cells (MM6) with recombinant αS1-casein, or LPS as control was monitored. Binding of αS1-casein to TLR4 was quantified by microscale thermophoresis (MST). ResultsαS1-casein induced secretion of IL-8 in TLR4+ cells and in MM6 cells with a six-times higher final IL-8 concentration in supernatants. IL-8 secretion was inhibited by intracellular TLR4-domain antagonist TAK-242 with an IC50-value of 259.6 nM, by ecto-domain TLR4 antagonistic mianserin with 10–51 μM and by anti-CD14-IgA. The binding constants (KD) of αS1-casein to the TLR4, MD2, and CD14 were 2.8 μM, 0.3 μM and 2.7 μM, respectively. Finally, αS1-casein showed a higher affinity to TLR4/MD2 (KD: 2.2 μM) compared to LPS (KD: 8.2 μM). ConclusionHuman αS1-casein induced proinflammatory effects are dependent upon binding to the TLR4 ectodomain and the presence of CD14. αS1-casein displayed stronger TLR4 agonistic activity than LPS via a different mode of action. General significanceBreast milk protein αS1-casein is a proinflammatory cytokine.

Plasma Biomarkers of Inflammation and Angiogenesis Predict Cerebral Cavernous Malformation Symptomatic Hemorrhage or Lesional Growth.

The clinical course of cerebral cavernous malformations is highly unpredictable, with few cross-sectional studies correlating proinflammatory genotypes and plasma biomarkers with prior disease severity. We hypothesize that a panel of 24 candidate plasma biomarkers, with a reported role in the physiopathology of cerebral cavernous malformations, may predict subsequent clinically relevant disease activity. Plasma biomarkers were assessed in nonfasting peripheral venous blood collected from consecutive cerebral cavernous malformation subjects followed for 1 year after initial sample collection. A first cohort (N=49) was used to define the best model of biomarker level combinations to predict a subsequent symptomatic lesional hemorrhagic expansion within a year after the blood sample. We generated the receiver operating characteristic curves and area under the curve for each biomarker individually and each weighted linear combination of relevant biomarkers. The best model to predict lesional activity was selected as that minimizing the Akaike information criterion. In this cohort, 11 subjects experienced symptomatic lesional hemorrhagic expansion (5 bleeds and 10 lesional growths) within a year after the blood draw. Subjects had lower soluble CD14 (cluster of differentiation 14; P=0.05), IL (interleukin)-6 (P=0.04), and VEGF (vascular endothelial growth factor; P=0.0003) levels along with higher plasma levels of IL-1β (P=0.008) and soluble ROBO4 (roundabout guidance receptor 4; P=0.03). Among the 31 weighted linear combinations of these 5 biomarkers, the best model (with the lowest Akaike information criterion value, 25.3) was the weighted linear combination including soluble CD14, IL-1β, VEGF, and soluble ROBO4, predicting a symptomatic hemorrhagic expansion with a sensitivity of 86% and specificity of 88% (area under the curve, 0.90; P<0.0001). We then validated our best model in the second sequential independent cohort (N=28). This is the first study reporting a predictive association between plasma biomarkers and subsequent cerebral cavernous malformation disease clinical activity. This may be applied in clinical prognostication and stratification of cases in clinical trials.

Melatonin from slow-release implants did not influence the gene expression of the lipopolysaccharide receptor complex in the choroid plexus of seasonally anoestrous adult ewes subjected or not to a systemic inflammatory stimulus

This study was designed to determine the effect of melatonin from continuous slow-release implants on (i) the mRNA expression for the lipopolysaccharide (LPS) receptor complex, which includes toll-like receptor 4 (TLR4), myeloid differentiation-2 factor (MD-2) and cluster of differentiation-14 (CD14) in the choroid plexus (CP) and (ii) on the plasma concentration of LPS binding protein (LBP) in ewes subjected or not to a systemic inflammatory stimulus (intravenous injection of LPS). Experiments were conducted in ovariectomized, oestradiol-implanted ewes during seasonal anoestrus (May/June). In experiment 1, the ewes were divided into four groups: sham-implanted and placebo-treated (C/C), melatonin-implanted and placebo-treated (M/C, Melovine 18mg), sham-implanted and LPS-treated for 3h (C/LPS, 400ng/kg of body mass) and melatonin-implanted and LPS-treated for 3h (M/LPS). In experiment 2, the ovariectomized ewes were melatonin-implanted or sham-implanted and treated with LPS for 6h (M/LPS and C/LPS, respectively). Plasma melatonin concentrations reached 4.5±1.5pg/ml in the C/C group, 151.4±56.4pg/ml in the M/C group, 7.8±4.3pg/ml in the C/LPS group and 240.6±93.0pg/ml in the M/LPS group one month after melatonin or sham implantation. Plasma cortisol concentrations remained at basal level throughout the experiment in the control ewes (C/C and M/C), whereas in the LPS-treated ewes, mean cortisol concentrations increased from 13.7±2.4ng/ml to 92.0±10.2ng/ml and from 15.4±5.9ng/ml to 100.0±10.7ng/ml in the C/LPS and M/LPS groups, respectively. The LPS treatment significantly (p≤0.05) increased the mRNA expression for TLR4 and CD14 but had no effect on the MD-2 mRNA expression in the C/LPS and M/LPS groups. Plasma LBP concentration was not affected within 3h of the LPS-treatment (experiment 1) but increased significantly at 4–6h after LPS-treatment in the C/LPS and M/LPS groups (experiment 2). Melatonin from the slow-release implants did not change the mRNA expression of all the examined genes or the plasma LPB concentration. These results allow us to infer that melatonin implantation does not disturb the CP potential to detect the specific microbial components derived from gram-negative bacteria.

Elevated CD14 (Cluster of Differentiation 14) and Toll-Like Receptor (TLR) 4 Signaling Deteriorate Periapical Inflammation in TLR2 Deficient Mice.

Apical periodontitis (periapical lesions) is an infection-induced chronic inflammation in the jaw, ultimately resulting in the destruction of apical periodontal tissue. Toll-like receptors (TLRs) are prominent in the initial recognition of pathogens. Our previous study showed that TLR4 signaling is proinflammatory in periapical lesions induced by a polymicrobial endodontic infection. In contrast, the functional role of TLR2 in regulation of periapical tissue destruction is still not fully understood. Using TLR2 deficient (KO), TLR2/TLR4 double deficient (dKO), and wild-type (WT) mice, we demonstrate that TLR2 KO mice are highly responsive to polymicrobial infection-induced periapical lesion caused by over activation of TLR4 signal transduction pathway that resulted in elevation of NF-kB (nuclear factor kappa B) and proinflammatory cytokine production. The altered TLR4 signaling is caused by TLR2 deficiency-dependent elevation of CD14 (cluster of differentiation 14), which is a co-receptor of TLR4. Indeed, neutralization of CD14 strikingly suppresses TLR2 deficiency-dependent inflammation and tissue destruction in vitro and in vivo. Our findings suggest that a network of TLR2, TLR4, and CD14 is a key factor in regulation of polymicrobial dentoalveolar infection and subsequent tissue destruction. Anat Rec, 299:1281-1292, 2016. © 2016 Wiley Periodicals, Inc.

The Role of Different Monocyte Subsets in the Pathogenesis of Atherosclerosis and Acute Coronary Syndromes.

The inflammation underlying both atherosclerosis and acute coronary syndromes is strongly related to monocyte-related actions. However, different monocyte subsets can play differential roles in the formation and destabilization of atherosclerotic plaque as well as healing of damaged myocardial tissue. Monocytes are currently being divided into three functionally distinct subsets with different levels of CD14 (cluster of differentiation 14) and CD16 expression. Thus, there are classical CD14++CD16-, intermediate CD14++CD16+ and non-classical CD14+CD16++ monocytes. Here, we summarize the current knowledge on complex activities of different monocyte subsets in atherosclerosis and acute coronary syndromes. Moreover, we discuss which monocyte subsets can serve either as predictive biomarkers of cardiovascular risk or as potential targets used in atherosclerosis and its complications.

Microglial Heparan Sulfate Proteoglycans Facilitate the Cluster-of-Differentiation 14 (CD14)/Toll-like Receptor 4 (TLR4)-Dependent Inflammatory Response

Microglia rapidly mount an inflammatory response to pathogens in the central nervous system (CNS). Heparan sulfate proteoglycans (HSPGs) have been attributed various roles in inflammation. To elucidate the relevance of microglial HSPGs in a pro-inflammatory response we isolated microglia from mice overexpressing heparanase (Hpa-tg), the HS-degrading endoglucuronidase, and challenged them with lipopolysaccharide (LPS), a bacterial endotoxin. Prior to LPS-stimulation, the LPS-receptor cluster-of-differentiation 14 (CD14) and Toll-like receptor 4 (TLR4; essential for the LPS response) were similarly expressed in Ctrl and Hpa-tg microglia. However, compared with Ctrl microglia, Hpa-tg cells released significantly less tumor necrosis factor-α (TNFα), essentially failed to up-regulate interleukin-1β (IL1β) and did not initiate synthesis of proCD14. Isolated primary astroyctes expressed TLR4, but notably lacked CD14 and in contrast to microglia, LPS challenge induced a similar TNFα response in Ctrl and Hpa-tg astrocytes, while neither released IL1β. The astrocyte TNFα-induction was thus attributed to CD14-independent TLR4 activation and was unaffected by the cells HS status. Equally, the suppressed LPS-response in Hpa-tg microglia indicated a loss of CD14-dependent TLR4 activation, suggesting that microglial HSPGs facilitate this process. Indeed, confocal microscopy confirmed interactions between microglial HS and CD14 in LPS-stimulated microglia and a potential HS-binding motif in CD14 was identified. We conclude that microglial HSPGs facilitate CD14-dependent TLR4 activation and that heparanase can modulate this mechanism.

Phylogenetic analysis and comparative study of Sahiwal cattle CD14 gene

Cluster of Differentiation-14 (CD14) gene is one of the strong candidate genes for mastitis resistance trait. Bovine CD14 gene spans over 2630 bp having 2 exons. Present investigation is based on phylogenetic analysis and comparative study of CD14 gene in Sahiwal cattle. A total of 100 lactating Sahiwal cattle maintained at NDRI, Karnal (Haryana) were included in the present study. Sequence analysis revealed that Sahiwal cattle CD14 gene sequence is 73 to 99% identical with several species. Phylogenetic analysis indicated that Sahiwal cattle (Bos indicus) and Bos taurus are genetically the most similar (99%) followed by Bubalus bubalis. The predicted peptide sequence revealed 373 amino acids precursor corresponding to coding sequence of CD14 gene of Sahiwal cattle. Amino acid length of CD14 protein was found different in reported species. BLAST analysis also revealed that Sahiwal cattle CD14 protein is 73 to 100% similar with other reported species. The sites for leucine-rich repeats, leucine rich nuclear export signal, putative N-linked glycosylation, O-linked glycosylation and disulphide bridges were predicted and comparative study was done.

Alterations in hepatic miRNA expression during negative energy balance in postpartum dairy cattle.

BackgroundNegative energy balance (NEB), an altered metabolic state, occurs in early postpartum dairy cattle when energy demands to support lactation exceed energy intake. During NEB the liver undergoes oxidative stress and increased breakdown of fatty acids accompanied by changes in gene expression. It is now known that micro RNAs (miRNA) can have a role in mediating such alterations in gene expression through repression or degradation of target mRNAs. miRNA expression is known to be altered by metabolism and environmental factors and miRNAs are implicated in expression modulation of metabolism related genes.ResultsmiRNA expression was profiled in the liver of moderate yielding dairy cattle under severe NEB (SNEB) and mild NEB (MNEB) using the Affymetrix Gene Chip miRNA_2.0 array with 679 probe sets for Bos-taurus miRNAs. Ten miRNAs were found to be differentially expressed using the ‘samr’ statistical package (delta = 0.6) at a q-value FDR of < 12%. Five miRNAs including miR-17-5p, miR-31, miR-140, miR-1281 and miR-2885 were validated using RT-qPCR, to be up-regulated under SNEB. Liver diseases associated with these miRNAs include non-alcoholic fatty liver (NAFLD) and hepatocellular carcinoma (HCC). miR-140 and miR-17-5p are known to show differential expression under oxidative stress. A total of 32 down-regulated putative target genes were also identified among 418 differentially expressed hepatic genes previously reported for the same animal model. Among these, GPR37 (G protein-coupled receptor 37), HEYL (hairy/enhancer-of-split related with YRPW motif-like), DNJA1, CD14 (Cluster of differentiation 14) and GNS (glucosamine (N-acetyl)-6-sulfatase) are known to be associated with hepatic metabolic disorders. In addition miR-140 and miR-2885 have binding sites on the most down-regulated of these genes, FADS2 (Fatty acid desaturase 2) which encodes an enzyme critical in lipid biosynthesis. Furthermore, HNF3-gamma (Hepatocyte nuclear factor 3-gamma), a hepatic transcription factor (TF) that is involved in IGF-1 expression regulation and maintenance of glucose homeostasis is a putative target of miR-31.ConclusionsThis study shows that SNEB affects liver miRNA expression and these miRNAs have putative targets in hepatic genes down-regulated under this condition. This study highlights the potential role of miRNAs in transcription regulation of hepatic gene expression during SNEB in dairy cattle.

CD14 is a coreceptor of Toll-like receptors 7 and 9

Recognition of pathogens by the innate immune system requires proteins that detect conserved molecular patterns. Nucleic acids are recognized by cytoplasmic sensors as well as by endosomal Toll-like receptors (TLRs). It has become evident that TLRs require additional proteins to be activated by their respective ligands. In this study, we show that CD14 (cluster of differentiation 14) constitutively interacts with the MyD88-dependent TLR7 and TLR9. CD14 was necessary for TLR7- and TLR9-dependent induction of proinflammatory cytokines in vitro and for TLR9-dependent innate immune responses in mice. CD14 associated with TLR9 stimulatory DNA in precipitation experiments and confocal imaging. The absence of CD14 led to reduced nucleic acid uptake in macrophages. Additionally, CD14 played a role in the stimulation of TLRs by viruses. Using various types of vesicular stomatitis virus, we showed that CD14 is dispensable for viral uptake but is required for the triggering of TLR-dependent cytokine responses. These data show that CD14 has a dual role in nucleic acid-mediated TLR activation: it promotes the selective uptake of nucleic acids, and it acts as a coreceptor for endosomal TLR activation.

Polymorphisms of genes encoding tumor necrosis factor-alpha, interleukin-10, cluster of differentiation-14 and interleukin-1ra in critically ill patients

Purpose The aim of the study was to determine whether distributions of tumor necrosis factor (TNF)- α 308, interleukin (IL)-10 1082, CD14 159, and IL-1ra gene intron 2 genotypes in critically ill patients are associated with outcome, underlying cause of sepsis, and type of microorganism. Materials and Methods Blood samples from 106 critically ill white patients were genotyped by method based on polymerase chain reaction for TNF- α 308, IL-10 1082, CD14 159, and IL-1ra gene intron 2. Results All patients with TNF- α 308AA genotype survived; relative risk (RR) of death in patients with AG was 3.250 and with GG, 1.923 ( P < .01). In patients with Gram-positive sepsis, IL-10 1082AA and then AG genotypes were the most frequent ones (odds ratio [OR], 18.67 and 7.20, respectively; P < .01). When comparing IL-10 1082AA with AG, RR of pancreatitis was 1.80 and OR was 3.40. When AA and GG were compared, RR was 7.33 and OR was 20.00. In patients with GG, RR of peritonitis was 4.07 and OR was 5.88 ( P < .01). In patients with Gram-positive sepsis, CD14 159CT was the most frequent one with OR of 5.25. Distribution of 6 IL-1ra gene intron 2 genotypes showed no significant association. Conclusions Distribution of TNF- α 308 genotypes is associated with outcome, IL-10 1082 with type of microorganism and underlying cause of sepsis, and CD14 159 with type of microorganism.