Comprehensive genotyping analysis of single nucleotide polymorphisms responsible for beef marbling in Japanese Black cattle.

BACKGROUND: Beef marbling is considered a desirable trait in the meat industry. Therefore, understanding the genetic factors that cause marbling is important. Previously, we performed a genome-wide association study to examine genetic factors associated with beef marbling in Japanese Black cattle and identified a candidate region between 10-30 Mbp on chromosome 7. We verified the effect of the SNPs in this region on beef marbling using linkage disequilibrium block analysis. We narrowed down the candidate region to a range of 15.8-16.1 Mbp. In this study, we comprehensively detected all of the SNPs in this region and verified their effects on beef marbling. RESULTS: Genome resequencing using four animals exhibiting high beef marbling standard (BMS) and four with low BMS revealed a total of 1,846 polymorphisms within the candidate region. Based on the annotation, we selected 13 SNPs exhibiting a moderate impact, as no high-impact SNPs were detected. All of the SNPs represented missense polymorphisms and were located in the following seven genes: RDH8, ANGPTL6, DNMT1, MRPL4, ICAM1, ICAM3, and ICAM5. Finally, we determined the effects of these SNPs on the BMS of a Japanese Black cattle population (n = 529). Analysis of variance revealed that the five SNPs were located in genes encoding the intercellular adhesion molecules (ICAM1, ICAM3, and ICAM5), and showed a highly significant association compared with the remainder (p < 0.01). The lowest p-value was observed for ICAM3_c.739G > A (p = 1.18E-04). Previous studies have suggested that intercellular adhesion molecules (ICAM) may be an upstream factor that regulates adipocyte differentiation. Therefore, considering the polymorphism and putative gene function, we suggest that ICAM1 is potentially responsible for beef marbling. c.470C > G and/or c.994G > A on ICAM1 may be responsible for this quantitative trait locus. CONCLUSIONS: Promising SNP candidates responsible for beef marbling were identified using extensive polymorphism verification in a previously reported QTL region. We aim to elucidate the mechanism of beef marbling in future studies by investigating how these polymorphisms alter protein structure and function.

Identification of Aedes aegypti salivary gland proteins interacting with human immune receptor proteins.

Mosquito saliva proteins modulate the human immune and hemostatic systems and control mosquito-borne pathogenic infections. One mechanism through which mosquito proteins may influence host immunity and hemostasis is their interactions with key human receptor proteins that may act as receptors for or coordinate attacks against invading pathogens. Here, using pull-down assays and proteomics-based mass spectrometry, we identified 11 Ae. aegypti salivary gland proteins (SGPs) (e.g., apyrase, Ae. aegypti venom allergen-1 [AaVA-1], neutrophil stimulating protein 1 [NeSt1], and D7 proteins), that interact with one or more of five human receptor proteins (cluster of differentiation 4 [CD4], CD14, CD86, dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin [DC-SIGN], and Toll-like receptor 4 [TLR4]). We focused on CD4- and DC-SIGN-interacting proteins and confirmed that CD4 directly interacts with AaVA-1, D7, and NeST1 recombinant proteins and that AaVA-1 showed a moderate interaction with DC-SIGN using ELISA. Bacteria responsive protein 1 (AgBR1), an Ae. aegypti saliva protein reported to enhance ZIKV infection in humans but that was not identified in our pull-down assay moderately interacts with CD4 in the ELISA assay. Functionally, we showed that AaVA-1 and NeST1 proteins promoted activation of CD4+ T cells. We propose the possible impact of these interactions and effects on mosquito-borne viral infections such as dengue, Zika, and chikungunya viruses. Overall, this study provides key insight into the vector-host (protein-protein) interaction network and suggests roles for these interactions in mosquito-borne viral infections.

Functional evaluation of various ICAM3 transcript variants in diffuse large B-Cell lymphoma.

Previous studies have identified several ICAM3 transcript variants and mainly investigated the function of the longest transcript of ICAM3 in various tumor progressions. However, the role of the other ICAM3 transcript variants remains unclear. Herein, we detected the expression of ICAM3 transcript variants 1-4 in DLBCL cells and tumor tissues, disclosed that variants 1, 3, and 4 were expressed in normal B cell lines and 3 DLBCL cell lines except SU-DHL-2 as well as tumor tissues, while variant 2 was not detected. Moreover, we found that ectopic expression of variants 1-4 enhanced cell proliferation by accelerating the cell cycle in SU-DHL2 cells in vitro. In addition, variants 1-4 overexpression showed no effects on SU-DHL2 cell apoptosis. Interestingly, the expression of variants 1, 3, and 4 promoted cell migration and EMT process while variant 2 had no effects. Collectively, the above results displayed the different roles of ICAM3 transcript variants in mediating DLBCL progression.

Pediatric ECMO: unfavorable outcomes are associated with inflammation and endothelial activation.

BACKGROUND: Inflammatory and endothelial activation responses during extracorporeal membrane oxygenation (ECMO) support in children are poorly understood. In this study, we aimed to determine if circulating inflammatory, endothelial activation, and fibrinolytic markers are associated with mortality and with neurologic outcomes in children on ECMO. METHODS: We conducted a secondary analysis of a two-center prospective observational study of 99 neonatal and pediatric ECMO patients. Inflammatory (interferon gamma [IFNγ], interleukin-6 [IL-6], IL-1ß, tumor necrosis factor alpha [TNFα]), endothelial activation (E-selectin, P-selectin, intercellular adhesion molecule-3 [ICAM-3], thrombomodulin [TM]), and fibrinolytic markers (tissue plasminogen activator [tPA], plasminogen activator inhibitor-1 [PAI-1]) were measured in plasma on days 1, 2, 3, 5, 7, and every third day thereafter during the ECMO course. RESULTS: All ECMO day 1 inflammatory biomarkers were significantly elevated in children with abnormal vs. normal neuroimaging. ECMO day 1 and peak levels of IL-6 and PAI-1 were significantly elevated in children who died compared to those who survived to hospital discharge. Tested biomarkers showed no significant association with long-term neurobehavioral outcomes measured using the Vineland Adaptive Behavioral Scales, Second Edition. CONCLUSIONS: High levels of circulating inflammatory, endothelial activation, and fibrinolytic markers are associated with mortality and abnormal neuroimaging in children on ECMO. IMPACT: The inflammatory, endothelial activation, and fibrinolytic profile of children on ECMO differs by primary indication for extracorporeal support. Proinflammatory biomarkers on ECMO day 1 are associated with abnormal neurologic imaging in children on ECMO in univariable but not multivariable models. In multivariable models, a pronounced proinflammatory and prothrombotic biomarker profile on ECMO day 1 and longitudinally was significantly associated with mortality. Further studies are needed to identify inflammatory, endothelial, and fibrinolytic profiles associated with increased risk for neurologic injury and mortality through potential mediation of bleeding and thrombosis.

The Phagocytic Code Regulating Phagocytosis of Mammalian Cells.

Mammalian phagocytes can phagocytose (i.e. eat) other mammalian cells in the body if they display certain signals, and this phagocytosis plays fundamental roles in development, cell turnover, tissue homeostasis and disease prevention. To phagocytose the correct cells, phagocytes must discriminate which cells to eat using a 'phagocytic code' - a set of over 50 known phagocytic signals determining whether a cell is eaten or not - comprising find-me signals, eat-me signals, don't-eat-me signals and opsonins. Most opsonins require binding to eat-me signals - for example, the opsonins galectin-3, calreticulin and C1q bind asialoglycan eat-me signals on target cells - to induce phagocytosis. Some proteins act as 'self-opsonins', while others are 'negative opsonins' or 'phagocyte suppressants', inhibiting phagocytosis. We review known phagocytic signals here, both established and novel, and how they integrate to regulate phagocytosis of several mammalian targets - including excess cells in development, senescent and aged cells, infected cells, cancer cells, dead or dying cells, cell debris and neuronal synapses. Understanding the phagocytic code, and how it goes wrong, may enable novel therapies for multiple pathologies with too much or too little phagocytosis, such as: infectious disease, cancer, neurodegeneration, psychiatric disease, cardiovascular disease, ageing and auto-immune disease.

Ibuprofen mediates histone modification to diminish cancer cell stemness properties via a COX2-dependent manner.

BACKGROUND: The anticancer potential of ibuprofen has created a broad interest to explore the clinical benefits of ibuprofen in cancer therapy. However, the current understanding of the molecular mechanisms involved in the anticancer potential of ibuprofen remains limited. METHODS: Cancer stemness assays to validate ibuprofen function in vitro and in vivo. Histone modification assays to check the effect of ibuprofen on histone acetylation/methylation, as well as the activity of HDAC and KDM6A/B. Inhibitors' in vivo assays to evaluate therapeutic effects of various inhibitors' combination manners. RESULTS: In our in vitro studies, we report that ibuprofen diminishes cancer cell stemness properties that include reducing the ALDH + subpopulation, side population and sphere formation in three cancer types. In our in vivo studies, we report that ibuprofen decreases tumour growth, metastasis and prolongs survival. In addition, our results showed that ibuprofen inhibits inflammation-related stemness gene expression (especially ICAM3) identified by a high-throughput siRNA platform. In regard to the underlying molecular mechanism of action, we report that ibuprofen reduces HDACs and histone demethylase (KDM6A/B) expression that mediates histone acetylation and methylation, and suppresses gene expression via a COX2-dependent way. In regard to therapeutic strategies, we report that ibuprofen combined HDAC/HDM inhibitors prevents cancer progression in vivo. CONCLUSIONS: The aforementioned findings suggest a molecular model that explains how ibuprofen diminishes cancer cell stemness properties. These may provide novel targets for therapeutic strategies involving ibuprofen in the prevention of cancer progression.

Bottlebrush Glycopolymers from 2-Oxazolines and Acrylamides for Targeting Dendritic Cell-Specific Intercellular Adhesion Molecule-3-Grabbing Nonintegrin and Mannose-Binding Lectin.

Lectins are omnipresent carbohydrate binding proteins that are involved in a multitude of biological processes. Unearthing their binding properties is a powerful tool toward the understanding and modification of their functions in biological applications. Herein, we present the synthesis of glycopolymers with a brush architecture via a "grafting from" methodology. The use of a versatile 2-oxazoline inimer was demonstrated to open avenues for a wide range of 2-oxazoline/acrylamide bottle brush polymers utilizing aqueous Cu-mediated reversible deactivation radical polymerization (Cu-RDRP). The polymers in the obtained library were assessed for their thermal properties in aqueous solution and their binding toward the C-type animal lectins dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) and mannose-binding lectin (MBL) via surface plasmon resonance spectrometry. The encapsulation properties of a hydrophobic drug-mimicking compound demonstrated the potential use of glyco brush copolymers in biological applications.

Serum biomarker analysis in patients with premature ovarian insufficiency.

Premature ovarian insufficiency (POI) is a primary ovarian defect characterized by premature depletion of ovarian follicles before 40 years of age. The disorder has been attributed to various causes, but the study of altered proteins in serum levels as the cause is rare. Additionally, identifying novel biomarkers can contribute to more accurate diagnosis or prognosis of POI. In the present study, a solid-phase antibody array simultaneously detecting multiple proteins was used to analyze POI serum with menopausal and healthy fertile subjects as control groups. As a result, compared to the menopause and healthy fertile groups, eleven proteins, including Neurturin, Frizzled-5, Serpin D1, MMP-7, ICAM-3, IL-17F, IFN-gamma R1, IL-29, IL-17R, IL-17C and Soggy-1, were uniquely down-regulated, and Afamin was particularly up-regulated in POI serum. More importantly, all of these factors were firstly found to be associated with POI in this study, suggesting that these proteins may participate in the pathogenesis of POI and may be novel serum biomarkers for POI.

ICAM3-Fc Outperforms Receptor-Specific Antibodies Targeted Nanoparticles to Dendritic Cells for Cross-Presentation.

Optimal targeting of nanoparticles (NP) to dendritic cells (DCs) receptors to deliver cancer-specific antigens is key to the efficient induction of anti-tumour immune responses. Poly (lactic-co-glycolic acid) (PLGA) nanoparticles containing tètanus toxoid and gp100 melanoma-associated antigen, toll-like receptor adjuvants were targeted to the DC-SIGN receptor in DCs by specific humanized antibodies or by ICAM3-Fc fusion proteins, which acts as the natural ligand. Despite higher binding and uptake efficacy of anti-DC-SIGN antibody-targeted NP vaccines than ICAM3-Fc ligand, no difference were observed in DC activation markers CD80, CD83, CD86 and CCR7 induced. DCs loaded with NP coated with ICAM3-Fc appeared more potent in activating T cells via cross-presentation than antibody-coated NP vaccines. This fact could be very crucial in the design of new cancer vaccines.

The Cross-Talk between miR-511-3p and C-Type Lectin Receptors on Dendritic Cells Affects Dendritic Cell Function.

MicroRNAs are small, noncoding RNAs that function as posttranscriptional modulators of gene expression by binding target mRNAs and inhibiting translation. They are therefore crucial regulators of several biological as well as immunological events. Recently, miR-511-3p has been implicated in the development and differentiation of APCs, such as dendritic cells (DCs), and regulating several human diseases. Interestingly, miR-511-3p is embedded within the human MRC1 gene that encodes the mannose receptor. In this study, we sought to examine the impact of miR-511-3p up- or downregulation on human DC surface phenotype, cytokine profile, immunogenicity (using IDO activity as a surrogate), and downstream T cell polarization. Using gene silencing and a selection of microRNA mimics, we could successfully suppress or induce the expression of miR-511-3p in DCs. Consequently, we show for the first time, to our knowledge, that inhibition and/or overexpression of miR-511-3p has opposing effects on the expression levels of two key C-type lectin receptors, namely the mannose receptor and DC-specific ICAM 3 nonintegrin at protein and mRNA levels, thereby affecting C-type lectin receptor-induced modulation of IDO activity in DCs. Furthermore, we show that downregulation of miR-511-3p drives an anti-inflammatory DC response characterized by IL-10 production. Interestingly, the miR-511-3plow DCs also promoted IL-4 secretion and suppressed IL-17 in cocultures with autologous T cells. Together, our data highlight the potential role of miR-511 in regulating DC function and downstream events leading to Th polarization and immune modulation.

Transcriptomics and Immunological Analyses Reveal a Pro-Angiogenic and Anti-Inflammatory Phenotype for Decidual Endothelial Cells.

BACKGROUND: In pregnancy, excessive inflammation and break down of immunologic tolerance can contribute to miscarriage. Endothelial cells (ECs) are able to orchestrate the inflammatory processes by secreting pro-inflammatory mediators and bactericidal factors by modulating leakiness and leukocyte trafficking, via the expression of adhesion molecules and chemokines. The aim of this study was to analyse the differences in the phenotype between microvascular ECs isolated from decidua (DECs) and ECs isolated from human skin (ADMECs). METHODS: DECs and ADMECs were characterized for their basal expression of angiogenic factors and adhesion molecules. A range of immunological responses was evaluated, such as vessel leakage, reactive oxygen species (ROS) production in response to TNF-α stimulation, adhesion molecules expression and leukocyte migration in response to TNF-α and IFN-γ stimulation. RESULTS: DECs produced higher levels of HGF, VEGF-A and IGFBP3 compared to ADMECs. DECs expressed adhesion molecules, ICAM-2 and ICAM-3, and a mild response to TNF-α was observed. Finally, DECs produced high levels of CXCL9/MIG and CXCL10/IP-10 in response to IFN-γ and selectively recruited Treg lymphocytes. CONCLUSION: DEC phenotype differs considerably from that of ADMECs, suggesting that DECs may play an active role in the control of immune response and angiogenesis at the foetal-maternal interface.

CD30, CD15, CD50, and PAX5 Expressions as Diagnostic Markers for Hodgkin Lymphoma (HL) and Systemic Anaplastic Large Cell Lymphoma (sALCL).

BACKGROUND: the expression of CD30, CD15, CD50, and PAX5 are used to help in confirming diagnosis of HL and sALCL; however data on the proportion of these markers have not been available. The study was aimed to identify the proportion of CD30, CD15, CD50 and PAX5 expressions and characteristics of patients with HL and sALCL at Dharmais National Cancer Center Hospital between 2005 and 2015. METHODS: a retrospective observational study was conducted using data from medical records and histopathological results of HL and sALCL adult patients who sought treatment at the hospital between 2005 and 2015. Immunohistochemistry (IHC) examinations were performed and data on the proportion of positive CD30, CD15, CD50, and PAX5 expressions were analyzed descriptively. RESULTS: a total of 45 patients were recruited in this study, with the majority (42 patients, 93.3%) were HL patients and only 6.7% were sALCL patients. The median age of HL patients was younger than sALCL patients; 35 (18-72 years old) versus 54 (49-61 years old). Moreover, the immunohistochemistry examination demonstrated that the positive CD15, CD30, CD50, and PAX5 expressions were found respectively in 37.5%, 88.9%, 31.2%, and 31.2% patients with HL; while in patients with sALCL, in spite of their small sample size, positive CD30, CD15, CD50 and PAX5 expressions were found in 100%; 66,7%; 50%; and 50%, respectively. Overall, CD15, CD50, and PAX5 positive expressions were found in 39.5%, 32.4%, and 32.4% patients who had HL and sALCL; while positive expression of CD30 was found in 89.5% of them. CONCLUSION: present study shows that almost 90% patients have positive CD30 expression;  while the positive expressions of CD15, CD50, and PAX5 are found in less than 40% patients. It indicates that CD30 is an important diagnostic marker for HL and sALCL and it may improve treatment strategy.

Elevations of inflammatory proteins in neonatal blood are associated with obesity and overweight among 2-year-old children born extremely premature.

BackgroundChildhood obesity is associated with elevated blood concentrations of inflammation markers. It is not known to what extent inflammation precedes the development of obesity.MethodsIn a cohort of 882 infants born before 28 weeks of gestation, we examined relationships between concentrations of 25 inflammation-related proteins in blood obtained during the first two postnatal weeks and body mass index at 2 years of age.ResultsAmong children delivered for spontaneous indications (n=734), obesity was associated with elevated concentrations of four proteins (IL-1ß, IL-6, TNF-R1, and MCP-1) on the first postnatal day; one protein (IL-6) on postnatal day 7; and two proteins (ICAM-3 and VEGF-R1) on postnatal day 14. Among children delivered for maternal or fetal indications (n=148), obesity was associated with elevated concentrations of seven proteins on the 14th postnatal day. In multivariable models in the spontaneous indications subsample, elevated IL-6 on day 1 predicted obesity (odds ratio: 2.9; 95% confidence limits: 1.2, 6.8), whereas elevated VCAM-1 on day 14 predicted overweight at 2 years of age (odds ratio: 2.3; 95% confidence limits: 1.2, 4.3).ConclusionsIn this cohort, neonatal systemic inflammation preceded the onset of obesity, suggesting that inflammation might contribute to the development of obesity.

Pathological Findings in Myasthenia Gravis Patients with Thymic Hyperplasia and Thymoma.

Thymectomy is routinely carried out in patients with myasthenia gravis (MG) and thymomas. However, there is still a dispute as to whether MG patients with thymic hyperplasia should undergo thymectomy. We aimed to investigate the pathological findings in the thymus in patients with co-existing MG and thymic hyperplasia or thymomas treated with thymectomy, as well as effects of immunosuppression. Thirty-three patients with MG were selected and grouped accordingly: patients with no thymic abnormalities, patients with thymic hyperplasia, and patients with thymomas. All patients were treated with methylprednisolone alongside immunosuppression. A separate cohort of 24 MG patients with thymic hyperplasia or thymomas and treated with thymectomy were selected. As controls, 5 patients with thymomas or thymic carcinoma without MG were selected. Expression of CD5, extracellular regulated protein kinases1/2 mitogen activated protein kinase (ERK1/2MAPKs) and CD95 ligand (FasL) in the thymus was examined. Methylprednisolone and immunosuppressive therapy are highly effective in MG patients with normal thymus tissue and MG patients with thymic hyperplasia compared to MG patients with thymomas alone. CD5 expression was highest in MG patients with thymic hyperplasia, correlating with expression of ERK1/2MAPKs. FasL expression was similar across all groups. Thymomas may be distinguished from thymic hyperplasia by expression of CD5 and ERK1/2MAPKs. Thymectomy is the preferred treatment for MG patients with thymomas but may not be necessary in MG patients with thymic hyperplasia who are treated with immunosuppressive therapy.

Pathogen-specific B-cell receptors drive chronic lymphocytic leukemia by light-chain-dependent cross-reaction with autoantigens.

Several lines of evidence indirectly suggest that antigenic stimulation through the B-cell receptor (BCR) supports chronic lymphocytic leukemia (CLL) development. In addition to self-antigens, a number of microbial antigens have been proposed to contribute to the selection of the immunoglobulins expressed in CLL. How pathogen-specific BCRs drive CLL development remains, however, largely unexplored. Here, we utilized mouse models of CLL pathogenesis to equip B cells with virus-specific BCRs and study the effect of antigen recognition on leukemia growth. Our results show that BCR engagement is absolutely required for CLL development. Unexpectedly, however, neither acute nor chronic exposure to virus-derived antigens influenced leukemia progression. Rather, CLL clones preferentially selected light chains that, when paired with virus-specific heavy chains, conferred B cells the ability to recognize a broad range of autoantigens. Taken together, our results suggest that pathogens may drive CLL pathogenesis by selecting and expanding pathogen-specific B cells that cross-react with one or more self-antigens.

Increased Cell Adhesion Molecules, PECAM-1, ICAM-3, or VCAM-1, Predict Increased Risk for Flare in Patients With Quiescent Inflammatory Bowel Disease.

GOALS AND BACKGROUND: Predicting the risk of flare-ups for patients with inflammatory bowel disease (IBD) is difficult. Alterations in gut endothelial regulation of mucosal immune homeostasis might be early events leading to flares in IBD. Cell adhesion molecules (CAMs), in particular, are important in maintaining endothelial integrity and regulating the migration of leukocytes into the gut. STUDY: We evaluated the mRNA expression of various tight junction proteins, with an emphasis on CAMs, in 40 patients with IBD in clinical remission. Patients were retrospectively assessed at 6, 12, and 24 months after baseline colonoscopy, and at the end of all available follow-up (maximum 65 mo), for flare events to determine whether baseline mRNA expression was associated with subsequent flares. RESULTS: At all follow-up points, the baseline expression of platelet endothelial cell adhesion molecule-1 (PECAM-1), ICAM-3, and VCAM-1 was significantly higher in patients who flared than in those who did not (2.4-fold elevation, P=0.012 for PECAM-1; 1.9-fold increased, P=0.03 for ICAM-3; and 1.4-fold increased, P=0.02 for VCAM-1). PECAM-1 and ICAM-3 expression was significantly increased in patients who flared as early as 6 months after baseline colonoscopy. In contrast, there were no significant differences between patients with and without flares in baseline expression of other CAMs (ESAM, ICAM-1, ICAM-2, E-selectin, P-selectin, and MadCAM1). CONCLUSIONS: Increased expression of PECAM-1, ICAM-3, and VCAM-1 in colonic biopsies from patients with IBD in clinical remission is associated with subsequent flares. This suggests that increases in the expression of these proteins may be early events that lead to flares in patients with IBD.