Hyaluronan signaling during ozone-induced lung injury requires TLR4, MyD88, and TIRAP.

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Zhuowei Li

Full Text Available Ozone exposure is associated with exacerbation of reactive airways disease. We have previously reported that the damage-associated molecular pattern, hyaluronan, is required for the complete biological response to ambient ozone and that hyaluronan fragments signal through toll-like receptor 4 (TLR4. In this study, we further investigated the role of TLR4 adaptors in ozone-induced airway hyperresponsiveness (AHR and the direct response to hyaluronan fragments (HA. Using a murine model of AHR, C57BL/6J, TLR4-/-, MyD88-/-, and TIRAP-/- mice were characterized for AHR after exposure to either ozone (1 ppm × 3 h or HA fragments. Animals were characterized for AHR with methacholine challenge, cellular inflammation, lung injury, and production of pro-inflammatory cytokines. Ozone-exposed C57BL/6J mice developed cellular inflammation, lung injury, pro-inflammatory cytokines, and AHR, while mice deficient in TLR4, MyD88 or TIRAP demonstrated both reduced AHR and reduced levels of pro-inflammatory cytokines including TNFα, IL-1β, MCP-1, IL-6 and KC. The level of hyaluronan was increased after inhalation of ozone in each strain of mice. Direct challenge of mice to hyaluronan resulted in AHR in C57BL/6J mice, but not in TLR4-/-, MyD88-/-, or TIRAP-/- mice. HA-induced cytokine production in wild-type mice was significantly reduced in TLR4-/-, MyD88-/-, or TIRAP-/- mice. In conclusion, our findings support that ozone-induced airway hyperresponsiveness is dependent on the HA-TLR4-MyD88-TIRAP signaling pathway.

MyD88 expression in the rat dental follicle: Implications for osteoclastogenesis and tooth eruption

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Liu, Dawen; Yao, Shaomian; Wise, Gary E.

2010-01-01

Myeloid differentiation factor 88 (MyD88) is a key adaptor molecule in the interleukin-1 (IL-1) and IL-18 Toll-like receptor signaling pathway. Because it is present in dental follicle (DF) cells in vitro, the purpose of this study was to determine its chronological expression in vivo, as well as its possible role in osteoclastogenesis and tooth eruption. An oligo DNA microarray was used to determine gene expression of MyD88 in vivo in the DFs from the first mandibular molars of postnatal rats from days 1–11. The results showed that MyD88 was expressed maximally at day 3. Using siRNA to knock down MyD88 expression in the DF cells also reduced the gene expression of nuclear factor-kappa B-1 (NFKB1) and monocyte chemoattractant protein 1 (MCP-1). IL-1α up-regulated the expression of NFKB1, MCP-1 and receptor activator of nuclear factor kappa B ligand (RANKL), but knockdown of MyD88 nullified this IL-1α effect. Conditioned medium from DF cells with MyD88 knocked down reduced chemotactic activity for mononuclear cells and reduced osteoclastogenesis as opposed to controls. In conclusion, the maximal expression of MyD88 at day 3 in the DF may contribute to the major burst of osteoclastogenesis needed for eruption by up-regulating MCP-1 and RANKL expression. PMID:20662905

Rare variants in MYD88, IRAK4 and IKBKG and susceptibility to invasive pneumococcal disease: a population-based case-control study.

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Magda K Ellis

Full Text Available Although rare variants within the Toll-like receptor signalling pathway genes have been found to underlie human primary immunodeficiencies associated with selective predisposition to invasive pneumococcal disease (IPD, the contribution of variants in these genes to IPD susceptibility at the population level remains unknown. Complete re-sequencing of IRAK4, MYD88 and IKBKG genes was undertaken in 164 IPD cases from the UK and 164 geographically-matched population-based controls. 233 single-nucleotide variants (SNVs were identified, of which ten were in coding regions. Four rare coding variants were predicted to be deleterious, two variants in MYD88 and two in IRAK4. The predicted deleterious variants in MYD88 were observed as two heterozygote cases but not seen in controls. Frequencies of predicted deleterious IRAK4 SNVs were the same in cases and controls. Our findings suggest that rare, functional variants in MYD88, IRAK4 or IKBKG do not significantly contribute to IPD susceptibility in adults at the population level.

Downregulation of adaptor protein MyD88 compromises the angiogenic potential of B16 murine melanoma

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Araya, Paula; Nuñez, Nicolás Gonzalo; Mena, Hebe Agustina; Bocco, José Luis; Negrotto, Soledad; Maccioni, Mariana

2017-01-01

The mechanisms that link inflammatory responses to cancer development remain a subject of intense investigation, emphasizing the need to better understand the cellular and molecular pathways that create a tumor promoting microenvironment. The myeloid differentiation primary response protein MyD88 acts as a main adaptor molecule for the signaling cascades initiated from Toll-like receptors (TLRs) and the interleukin 1 receptor (IL-1R). MyD88 has been shown to contribute to tumorigenesis in many inflammation-associated cancer models. In this study, we sought to better define the role of MyD88 in neoplastic cells using a murine melanoma model. Herein, we have demonstrated that MyD88 expression is required to maintain the angiogenic switch that supports B16 melanoma growth. By knocking down MyD88 we reduced TLR-mediated NF-κB activation with no evident effects over cell proliferation and survival. In addition, MyD88 downregulation was associated with a decrease of HIF1α levels and its target gene VEGF, in correlation with an impaired capability to induce capillary sprouting and tube formation of endothelial cells. Melanomas developed from cells lacking MyD88 showed an enhanced secretion of chemoattractant ligands such as CCL2, CXCL10 and CXCL1 and have an improved infiltration of macrophages to the tumor site. Our results imply that cell-autonomous signaling through MyD88 is required to sustain tumor growth and underscore its function as an important positive modulator of tumor angiogenesis. PMID:28662055

Signaling through MyD88 regulates leukocyte recruitment after brain injury

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Babcock, Alicia A; Toft-Hansen, Henrik; Owens, Trevor

2008-01-01

hippocampus. We now show that significant leukocyte entry into the EC occurs within 3-12 h of stab injury. Whereas T cells showed small, gradual increases over 8 days, macrophage infiltration was pronounced and peaked within 12-24 h. MyD88 deficiency significantly reduced macrophage and T cell recruitment...... for TNF-alpha, IL-1beta, and CCL2, which increased >50-fold after stab injury in C57BL/6 mice at the time of peak expression, were severely reduced in injured MyD88 knockout mice. Leukocyte recruitment and gene expression were unaffected in TLR2-deficient or TLR4 mutant mice. No significant differences...... in gene expression were observed in mice lacking IL-1R or IL-18R. These data show that MyD88-dependent signaling mediates proinflammatory gene expression and leukocyte recruitment after CNS injury....

Epicutaneous exposure to nickel induces nickel allergy in mice via a MyD88-dependent and interleukin-1-dependent pathway

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Vennegaard, Marie T; Dyring-Andersen, Beatrice; Skov, Lone

2014-01-01

-lasting epicutaneous exposure to nickel. OBJECTIVE: To develop a mouse model reflecting nickel allergy in humans induced by epicutaneous exposure to nickel, and to investigate the mechanisms involved in such allergic responses. METHODS: Mice were exposed to NiCl2 on the dorsal side of the ears. Inflammation...... was evaluated by the swelling and cell infiltration of the ears. T cell responses were determined as numbers of CD4(+) and CD8(+) T cells in the draining lymph nodes. Localization of nickel was examined by dimethylglyoxime staining. RESULTS: Epicutaneous exposure to nickel results in prolonged localization...... of nickel in the epidermis, and induces nickel allergy in mice. The allergic response to nickel following epicutaneous exposure is MyD88-dependent and interleukin (IL)-1 receptor-dependent, but independent of toll-like receptor (TLR)-4. CONCLUSION: This new model for nickel allergy that reflects...

MyD88 But Not TRIF Is Essential for Osteoclastogenesis Induced by Lipopolysaccharide, Diacyl Lipopeptide, and IL-1α

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Sato, Nobuaki; Takahashi, Naoyuki; Suda, Koji; Nakamura, Midori; Yamaki, Mariko; Ninomiya, Tadashi; Kobayashi, Yasuhiro; Takada, Haruhiko; Shibata, Kenichiro; Yamamoto, Masahiro; Takeda, Kiyoshi; Akira, Shizuo; Noguchi, Toshihide; Udagawa, Nobuyuki

2004-01-01

Myeloid differentiation factor 88 (MyD88) plays essential roles in the signaling of the Toll/interleukin (IL)-1 receptor family. Toll–IL-1 receptor domain-containing adaptor inducing interferon-β (TRIF)-mediated signals are involved in lipopolysaccharide (LPS)-induced MyD88-independent pathways. Using MyD88-deficient (MyD88−/−) mice and TRIF-deficient (TRIF−/−) mice, we examined roles of MyD88 and TRIF in osteoclast differentiation and function. LPS, diacyl lipopeptide, and IL-1α stimulated osteoclastogenesis in cocultures of osteoblasts and hemopoietic cells obtained from TRIF−/− mice, but not MyD88−/− mice. These factors stimulated receptor activator of nuclear factor-κB ligand mRNA expression in TRIF−/− osteoblasts, but not MyD88−/− osteoblasts. LPS stimulated IL-6 production in TRIF−/− osteoblasts, but not TRIF−/− macrophages. LPS and IL-1α enhanced the survival of TRIF−/− osteoclasts, but not MyD88−/− osteoclasts. Diacyl lipopeptide did not support the survival of osteoclasts because of the lack of Toll-like receptor (TLR)6 in osteoclasts. Macrophages expressed both TRIF and TRIF-related adaptor molecule (TRAM) mRNA, whereas osteoblasts and osteoclasts expressed only TRIF mRNA. Bone histomorphometry showed that MyD88−/− mice exhibited osteopenia with reduced bone resorption and formation. These results suggest that the MyD88-mediated signal is essential for the osteoclastogenesis and function induced by IL-1 and TLR ligands, and that MyD88 is physiologically involved in bone turnover. PMID:15353553

Activation of TAK1 by MYD88 L265P drives malignant B-cell Growth in non-Hodgkin lymphoma

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Ansell, S M; Hodge, L S; Secreto, F J; Manske, M; Braggio, E; Price-Troska, T; Ziesmer, S; Li, Y; Johnson, S H; Hart, S N; Kocher, J-P A; Vasmatzis, G; Chanan-Kahn, A; Gertz, M; Fonseca, R; Dogan, A; Cerhan, J R; Novak, A J

2014-01-01

Massively parallel sequencing analyses have revealed a common mutation within the MYD88 gene (MYD88 L265P ) occurring at high frequencies in many non-Hodgkin lymphomas (NHLs) including the rare lymphoplasmacytic lymphoma, Waldenström's macroglobulinemia (WM). Using whole-exome sequencing, Sanger sequencing and allele-specific PCR, we validate the initial studies and detect the MYD88 L265P mutation in the tumor genome of 97% of WM patients analyzed (n=39). Due to the high frequency of MYD88 mutation in WM and other NHL, and its known effects on malignant B-cell survival, therapeutic targeting of MYD88 signaling pathways may be clinically useful. However, we are lacking a thorough characterization of the role of intermediary signaling proteins on the biology of MYD88 L265P -expressing B cells. We report here that MYD88 L265P signaling is constitutively active in both WM and diffuse large B-cell lymphoma cells leading to heightened MYD88 L265P , IRAK and TRAF6 oligomerization and NF-κB activation. Furthermore, we have identified the signaling protein, TAK1, to be an essential mediator of MYD88 L265P -driven signaling, cellular proliferation and cytokine secretion in malignant B cells. Our studies highlight the biological significance of MYD88 L265P in NHL and reveal TAK1 inhibition to be a potential therapeutic strategy for the treatment of WM and other diseases characterized by MYD88 L265P

TRAM is involved in IL-18 signaling and functions as a sorting adaptor for MyD88.

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Hidenori Ohnishi

Full Text Available MyD88, a Toll/interleukin-1 receptor homology (TIR domain-containing adaptor protein, mediates signals from the Toll-like receptors (TLR or IL-1/IL-18 receptors to downstream kinases. In MyD88-dependent TLR4 signaling, the function of MyD88 is enhanced by another TIR domain-containing adaptor, Mal/TIRAP, which brings MyD88 to the plasma membrane and promotes its interaction with the cytosolic region of TLR4. Hence, Mal is recognized as the "sorting adaptor" for MyD88. In this study, a direct interaction between MyD88-TIR and another membrane-sorting adaptor, TRAM/TICAM-2, was demonstrated in vitro. Cell-based assays including RNA interference experiments and TRAM deficient mice revealed that the interplay between MyD88 and TRAM in cells is important in mediating IL-18 signal transduction. Live cell imaging further demonstrated the co-localized accumulation of MyD88 and TRAM in the membrane regions in HEK293 cells. These findings suggest that TRAM serves as the sorting adaptor for MyD88 in IL-18 signaling, which then facilitates the signal transduction. The binding sites for TRAM are located in the TIR domain of MyD88 and actually overlap with the binding sites for Mal. MyD88, the multifunctional signaling adaptor that works together with most of the TLR members and with the IL-1/IL-18 receptors, can interact with two distinct sorting adaptors, TRAM and Mal, in a conserved manner in a distinct context.

Gliadin stimulation of murine macrophage inflammatory gene expression and intestinal permeability are MyD88-dependent: role of the innate immune response in Celiac disease.

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Thomas, Karen E; Sapone, Anna; Fasano, Alessio; Vogel, Stefanie N

2006-02-15

Recent studies have demonstrated the importance of TLR signaling in intestinal homeostasis. Celiac disease (CD) is an autoimmune enteropathy triggered in susceptible individuals by the ingestion of gliadin-containing grains. In this study, we sought to test the hypothesis that gliadin initiates this response by stimulating the innate immune response to increase intestinal permeability and by up-regulating macrophage proinflammatory gene expression and cytokine production. To this end, intestinal permeability and the release of zonulin (an endogenous mediator of gut permeability) in vitro, as well as proinflammatory gene expression and cytokine release by primary murine macrophage cultures, were measured. Gliadin and its peptide derivatives, 33-mer and p31-43, were found to be potent inducers of both a zonulin-dependent increase in intestinal permeability and macrophage proinflammatory gene expression and cytokine secretion. Gliadin-induced zonulin release, increased intestinal permeability, and cytokine production were dependent on myeloid differentiation factor 88 (MyD88), a key adapter molecule in the TLR/IL-1R signaling pathways, but were neither TLR2- nor TLR4-dependent. Our data support the following model for the innate immune response to gliadin in the initiation of CD. Gliadin interaction with the intestinal epithelium increases intestinal permeability through the MyD88-dependent release of zonulin that, in turn, enables paracellular translocation of gliadin and its subsequent interaction with macrophages within the intestinal submucosa. There, the interaction of gliadin with macrophages elicits a MyD88-dependent proinflammatory cytokine milieu that facilitates the interaction of T cells with APCs, leading ultimately to the Ag-specific adaptive immune response seen in patients with CD.

The -938C>A Polymorphism in MYD88 Is Associated with Susceptibility to Tuberculosis: A Pilot Study

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Kalliopi Aggelou

2016-01-01

Full Text Available Introduction. Tuberculosis (TB is a major disease worldwide, caused by Mycobacterium tuberculosis (MTB infection. The Toll-Like Receptor (TLR pathway plays a crucial role in the recognition of MTB. Aim. The present study aimed to investigate the involvement of myeloid differentiation primary response protein 88 (MYD88 gene polymorphisms in TB. Materials and Methods. A total of 103 TB cases and 92 control subjects were genotyped for the MYD88 -938C>A (rs4988453 and 1944C>G (rs4988457 polymorphisms. Results. The MYD88 -938CA and -938AA genotypes were associated with an increased risk for tuberculosis with odds ratio (OR of 5.71 (95% confidence intervals [CIs] 2.89–11.28, p=0.01. Conclusions. The MYD88 -938C>A genetic polymorphism is associated with increased susceptibility to TB and may serve as a marker to screen individuals who are at risk.

Identification and function of myeloid differentiation factor 88 (MyD88 in Litopenaeus vannamei.

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Shuang Zhang

Full Text Available Myeloid differentiation factor 88 (MyD88 is a universal and essential signaling protein in Toll-like receptor/interleukin-1 receptor-induced activation of nuclear factor-kappa B. In this study, two MyD88 protein variants (LvMyD88 and LvMyD88-1 were identified in Litopenaeus vannamei. The LvMyD88 cDNA is 1,848 bp in length and contains an open reading frame (ORF of 1,428 bp, whereas the LvMyD88-1 cDNA is 1,719 bp in length and has an ORF of 1,299 bp. Both variants encode proteins with death and Toll/interleukin-1 receptor domains and share 91% sequence identity. In healthy L. vannamei, the LvMyD88 genes were highly expressed in hemocytes but at a low level in the hepatopancreas. The LvMyD88s expression was induced in hemocytes after challenge with lipopolysaccharide, CpG-ODN2006, Vibrio parahaemolyticus, Staphyloccocus aureus, and white spot syndrome virus, but not by poly I∶C. Overexpression of LvMyD88 and LvMyD88-1 in Drosophila Schneider 2 cells led to activation of antimicrobial peptide genes and wsv069 (ie1, wsv303, and wsv371. These results suggested that LvMyD88 may play a role in antibacterial and antiviral response in L. vannamei. To our knowledge, this is the first report on MyD88 in shrimp and a variant of MyD88 gene in invertebrates.

Lack of MyD88 protects the immunodeficient host against fatal lung inflammation triggered by the opportunistic bacteria Burkholderia cenocepacia.

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Ventura, Grasiella M de C; Balloy, Viviane; Ramphal, Reuben; Khun, Huot; Huerre, Michel; Ryffel, Bernhard; Plotkowski, Maria-Cristina M; Chignard, Michel; Si-Tahar, Mustapha

2009-07-01

Burkholderia cenocepacia is an opportunistic pathogen of major concern for cystic fibrosis patients as well as immunocompromised cancer patients and transplant recipients. The mechanisms by which B. cenocepacia triggers a rapid health deterioration of the susceptible host have yet to be characterized. TLR and their key signaling intermediate MyD88 play a central role in the detection of microbial molecular patterns and in the initiation of an effective immune response. We performed a study to better understand the role of TLR-MyD88 signaling in B. cenocepacia-induced pathogenesis in the immunocompromised host, using an experimental murine model. The time-course of several dynamic parameters, including animal survival, bacterial load, and secretion of critical inflammatory mediators, was compared in infected and immunosuppressed wild-type and MyD88(-/-) mice. Notably, when compared with wild-type mice, infected MyD88(-/-) animals displayed significantly reduced levels of inflammatory mediators (including KC, TNF-alpha, IL-6, MIP-2, and G-CSF) in blood and lung airspaces. Moreover, despite a higher transient bacterial load in the lungs, immunosuppressed mice deficient in MyD88 had an unexpected survival advantage. Finally, we showed that this B. cenocepacia-induced life-threatening infection of wild-type mice involved the proinflammatory cytokine TNF-alpha and could be prevented by corticosteroids. Altogether, our findings demonstrate that a MyD88-dependent pathway can critically contribute to a detrimental host inflammatory response that leads to fatal pneumonia.

Experimental and natural infections in MyD88- and IRAK-4-deficient mice and humans

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von Bernuth, Horst; Picard, Capucine; Puel, Anne; Casanova, Jean-Laurent

2013-01-01

Most Toll-like-receptors (TLRs) and interleukin-1 receptors (IL-1Rs) signal via myeloid differentiation primary response 88 (MyD88) and interleukin-1 receptor-associated kinase 4 (IRAK-4). The combined roles of these two receptor families in the course of experimental infections have been assessed in MyD88- and IRAK-4-deficient mice for almost fifteen years. These animals have been shown to be susceptible to 46 pathogens: 27 bacteria, 8 viruses, 7 parasites, and 4 fungi. Humans with inborn MyD88 or IRAK-4 deficiency were first identified in 2003. They suffer from naturally occurring life-threatening infections caused by a small number of bacterial species, although the incidence and severity of these infections decrease with age. Mouse TLR- and IL-1R-dependent immunity mediated by MyD88 and IRAK-4 seems to be vital to combat a wide array of experimentally administered pathogens at most ages. By contrast, human TLR- and IL-1R-dependent immunity mediated by MyD88 and IRAK-4 seems to be effective in the natural setting against only a few bacteria and is most important in infancy and early childhood. The roles of TLRs and IL-1Rs in protective immunity deduced from studies in mutant mice subjected to experimental infections should therefore be reconsidered in the light of findings for natural infections in humans carrying mutations as discussed in this review. PMID:23255009

Andrographolide alleviates imiquimod-induced psoriasis in mice via inducing autophagic proteolysis of MyD88.

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Shao, Fenli; Tan, Tao; Tan, Yang; Sun, Yang; Wu, Xingxin; Xu, Qiang

2016-09-01

Psoriasis is a chronic inflammatory skin disease with excessive activation of toll-like receptors (TLRs), which play important roles in developing psoriasis. Targeting TLR signaling remains a challenge for treating psoriasis. Here, we found that andrographolide (Andro), a small-molecule natural product, alleviated imiquimod- but not interleukin 23 (IL-23)-induced psoriasis in mice with reducing expressions of IL-23 and IL-1β in the skin. The improvement in imiquimod-induced psoriasis by Andro was not observed in microtubule-associated protein 1 light chain 3 beta (MAP1LC3B) knockout mice. Furthermore, Andro inhibited mRNA expressions of IL-23, IL-6 and IL-1β but not CD80 and CD86 in bone-marrow derived dendritic cells (BMDCs) treated with lipopolysaccharide (LPS) in a MAP1LC3B-dependent manner. In addition, Andro inhibited imiquimod-induced mRNA expressions of IL-23, IL-6, IL-1β, CD80 and CD86 in BMDCs from mice. Interestingly, Andro induced a degradation of myeloid differentiation factor 88 (MyD88) and blocked the recruitment of TNF receptor-associated factor 6 (TRAF6) to MyD88 upon LPS stimulation in BMDCs from mice. Blockade of autophagic proteolysis using NH4Cl or MAP1LC3B(-/-) BMDCs abolished the Andro-induced MyD88 degradation. In conclusion, Andro controls activation of MyD88-dependent cytokines and alleviates psoriasis in mice via inducing autophagic proteolysis of MyD88, which could be a novel strategy to treat psoriasis. Copyright © 2016 Elsevier Inc. All rights reserved.

Amelioration of murine passive immune thrombocytopenia by IVIg and a therapeutic monoclonal CD44 antibody does not require the Myd88 signaling pathway.

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Andrew R Crow

Full Text Available Immune thrombocytopenia (ITP is an autoimmune bleeding disorder characterized by a low platelet count and the production of anti-platelet antibodies. The majority of ITP patients have antibodies to platelet integrin α(IIbβ₃ (GPIIbIIIa which can direct platelet phagocytosis by macrophages. One effective treatment for patients with ITP is intravenous immunoglobulin (IVIg which rapidly reverses thrombocytopenia. The exact mechanism of IVIg action in human patients is unclear, although in mouse models of passive ITP, IVIg can rapidly increase platelet counts in the absence of adaptive immunity. Another antibody therapeutic that can similarly increase platelet counts independent of adaptive immunity are CD44 antibodies. Toll-like receptors (TLRs are pattern recognition receptors which play a central role in helping direct the innate immune system. Dendritic cells, which are notable for their expression of TLRs, have been directly implicated in IVIg function as an initiator cell, while CD44 can associate with TLR2 and TLR4. We therefore questioned whether IVIg, or the therapeutic CD44 antibody KM114, mediate their ameliorative effects in a manner dependent upon normal TLR function. Here, we demonstrate that the TLR4 agonist LPS does not inhibit IVIg or KM114 amelioration of antibody-induced thrombocytopenia, and that these therapeutics do not ameliorate LPS-induced thrombocytopenia. IVIg was able to significantly ameliorate murine ITP in C3H/HeJ mice which have defective TLR4. All known murine TLRs except TLR3 utilize the Myd88 adapter protein to drive TLR signaling. Employing Myd88 deficient mice, we found that both IVIg and KM114 ameliorate murine ITP in Myd88 deficient mice to the same extent as normal mice. Thus both IVIg and anti-CD44 antibody can mediate their ameliorative effects in murine passive ITP independent of the Myd88 signaling pathway. These data help shed light on the mechanism of action of IVIg and KM114 in the amelioration of

Significance of TLR4/MyD88 expression in breast cancer

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Chen, Xiangjin; Zhao, Feng; Zhang, Huihao; Zhu, Youzhi; Wu, Kunlin; Tan, Guozheng

2015-01-01

Objective: To investigate the expression of TLR4/MyD88 in breast cancer, and explore the relationship between their expression and breast cancer tumor growth and invasion. Methods: We examined the protein expression of TLR4 and MyD88 in 60 cases of histologically confirmed breast cancer. The relationship of their protein expressions with clinical features including age at diagnosis, tumor size and stage, lymph node metastasis and distant metastasis were analyzed. Results: The IHC results showed that TLR4 and MyD88 were expressed in 63.3% (38/60) and 58.3% (35/60) of malignant breast tumors respectively. TLR4 expression in breast cancer were significantly higher than in fibroadenoma (n = 4, 20.0%) and adjacent normal tissues (n = 2, 10.0%) (P fibroadenoma (n = 4, 20.0%) and adjacent normal tissue (n = 3, 15.0%) (P fibroadenoma and adjacent normal tissues (P < 0.05). The protein expressions of TLR4 and MyD88 were also significantly associated with poor clinical features (P < 0.05). Conclusion: TLR4 and MyD88 expression might be associated with breast cancer growth and regional and distant metastases. PMID:26261595

The TLR13-MyD88-NF-κB signalling pathway of Cyclina sinensis plays vital roles in innate immune responses.

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Ren, Yipeng; Ding, Dan; Pan, Baoping; Bu, Wenjun

2017-11-01

Toll-like receptors, the best known pattern recognition receptors, play important roles in recognizing non-self molecules and binding pathogen-associated molecular patterns in the innate immune system. In the present research, the cDNA and protein characterization of the TLR signalling pathway genes including IRAK4, TRAK6 and IKKα (named CsIRAK4, CsTRAF6 and CsIKKα, respectively) with the typical motifs from Cyclina sinensis showed significant similarity with their homologues from other shellfish. Furthermore, the mRNA transcripts of these three genes are ubiquitously expressed in all tissues tested and are dominantly expressed in C. sinensis haemocytes (P sinensis by RNA interference and immune challenges. The results suggested the mRNA expression patterns of CsMyD88, CsIRAK4, CsTRAF6, CsIKKα, CsIκB, CsNF-κB, CsC-LYZ and CsAMP were all down-regulated (P sinensis haemocytes, revealing the involvement of the TLR13-MyD88-NF-κB signalling pathway in innate immunity by positively adjusting internal signalling factors and immune-related genes. In summary, a TLR13-MyD88-NF-κB signalling pathway exists and plays vital roles in innate immune responses in C. sinensis. These findings collectively lay the foundation for studying the functional characterization of internal signalling factors and establishing a regulatory network for the TLR signalling pathway in molluscs. Copyright © 2017 Elsevier Ltd. All rights reserved.

IL-1R and MyD88 signalling in CD4+ T cells promote Th17 immunity and atherosclerosis.

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Engelbertsen, Daniel; Rattik, Sara; Wigren, Maria; Vallejo, Jenifer; Marinkovic, Goran; Schiopu, Alexandru; Björkbacka, Harry; Nilsson, Jan; Bengtsson, Eva

2018-01-01

The role of CD4+ T cells in atherosclerosis has been shown to be dependent on cytokine cues that regulate lineage commitment into mature T helper sub-sets. In this study, we tested the roles of IL-1R1 and MyD88 signalling in CD4+ T cells in atherosclerosis. We transferred apoe-/-myd88+/+ or apoe-/-myd88-/- CD4+ T cells to T- and B-cell-deficient rag1-/-apoe-/- mice fed high fat diet. Mice given apoe-/-myd88-/- CD4+ T cells exhibited reduced atherosclerosis compared with mice given apoe-/-myd88+/+ CD4+ T cells. CD4+ T cells from apoe-/-myd88-/- produced less IL-17 but similar levels of IFN-γ. Treatment of human CD4+ T cells with a MyD88 inhibitor inhibited IL-17 secretion in vitro. Transfer of il1r1-/- CD4+ T cells recapitulated the phenotype seen by transfer of myd88-/- CD4+ T cells with reduced lesion development and a reduction in Th17 and IL-17 production compared with wild type CD4+ T cell recipients. Relative collagen content of lesions was reduced in mice receiving il1r1-/- CD4+ T cells. We demonstrate that both IL1R and MyD88 signalling in CD4+ T cells promote Th17 immunity, plaque growth and may regulate plaque collagen levels. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2017. For permissions please email: journals.permissions@oup.com.

Toll-like receptors and cancer: MYD88 mutation and inflammation

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James Q Wang

2014-07-01

Full Text Available Pattern recognition receptors (PRRs expressed on immune cells are crucial for the early detection of invading pathogens, in initiating early innate immune response and in orchestrating the adaptive immune response. PRRs are activated by specific pathogen-associated molecular patterns (PAMPs that are present in pathogenic microbes or nucleic acids of viruses or bacteria. However, inappropriate activation of these PRRs, such as the Toll-like receptors (TLRs, due to genetic lesions or chronic inflammation has been demonstrated to be a major cause of many haematological malignancies. Gain-of-function mutations in the TLR adaptor protein MYD88 found in 39% of the activated B cell type of diffuse large B cell lymphomas (ABC-DLBCL and almost 100% of Waldenström’s macroglobulinemia (WM further highlight the involvement of TLRs in these malignancies. MYD88 mutations result in the chronic activation of TLR signalling pathways, thus the constitutive activation of the transcription factor NFκB to promote cell survival and proliferation. These recent insights into TLR pathway driven malignancies warrant the need for a better understanding of TLRs in cancers and the development of novel anti-cancer therapies targeting TLRs. This review focuses on Toll-like receptors function and signalling in normal or inflammatory conditions, and how mutations can also hijack the TLR signalling pathways to give rise to cancer. Lastly, we discuss how potential therapeutic agents could be used to restore normal responses to TLRs and have long lasting anti-tumour effects.

MyD88 is crucial for the development of a protective CNS immune response to Toxoplasma gondii infection

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Torres Marbel

2013-02-01

Full Text Available Abstract Background Toxoplasmosis is one of the most common parasitic infections in humans. It can establish chronic infection and is characterized by the formation of tissue cysts in the brain. The cysts remain largely quiescent for the life of the host, but can reactivate and cause life-threatening toxoplasmic encephalitis in immunocompromised patients, such as those with AIDS, neoplastic diseases and organ transplants. Toll-like receptor (TLR adaptor MyD88 activation is required for the innate sensing of Toxoplasma gondii. Mice deficient in MyD88 have defective IL-12 and Th1 effector responses, and are highly susceptible to the acute phase of T. gondii infection. However, the role of this signaling pathway during cerebral infection is poorly understood and requires examination. Method MyD88-deficient mice and control mice were orally infected with T. gondii cysts. Cellular and parasite infiltration in the peripheral organs and in the brain were determined by histology and immunohistochemistry. Cytokine levels were determined by ELISA and chemokine mRNA levels were quantified by real-time PCR (qPCR. Results Thirteen days after infection, a higher parasite burden was observed but there was no histological change in the liver, heart, lungs and small intestine of MyD88−/− and MyD88+/+ mice. However, MyD88−/− mice compared to MyD88+/+ mice were highly susceptible to cerebral infection, displayed high parasite migration to the brain, severe neuropathological signs of encephalitis and succumbed within 2 weeks of oral infection. Susceptibility was primarily associated with lower expression of Th1 cytokines, especially IL-12, IFN-γ and TNF-α, significant decrease in the expression of CCL3, CCL5, CCL7 and CCL19 chemokines, marked defect of CD8+ T cells, and infiltration of CD11b+ and F4/80+ cells in the brain. Conclusion MyD88 is essential for the protection of mice during the cerebral installation of T. gondii infection. These results

The combination of maltose-binding protein and BCG-induced Th1 activation is involved in TLR2/9-mediated upregulation of MyD88-TRAF6 and TLR4-mediated downregulation of TRIF-TRAF3.

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Liu, Guomu; Zhai, Xiaoyu; Zhou, Hongyue; Yang, Xiaoyu; Zhang, Nannan; Tai, Guixiang; Ni, Weihua

2018-03-01

Our previous study demonstrated that maltose-binding protein (MBP) activated Th1 through the TLR2-mediated MyD88-dependent pathway and the TLR4-mediated TRIF-dependent pathway. The combination of MBP and BCG synergistically induced Th1 activation, and the TLR2/9-mediated MyD88-dependent pathway is involved in this process. To further explore this mechanism, we stimulated purified mouse CD4 + T cells with MBP and BCG in vitro. The results demonstrated that MBP combined with BCG synergistically increased IFN-γ production and TLR2/4/9 expression, suggesting the involvement of TLR2/4/9 in the combination-induced Th1 activation. Next, TLRs 2/4/9 were blocked to analyze the effects of TLRs on Th1 activation. The results demonstrated that MBP induced a low level of Th1 activation by upregulating TLR2-mediated MyD88-TRAF6 and TLR4-mediated TRIF-TRAF3 expression, whereas MBP combined with BCG induced synergistic Th1 activation, which was not only triggered by strong upregulation of TLR2/9-mediated MyD88-TRAF6 expression but also by shifting TLR4-mediated TRIF-TRAF3 into the TRIF-TRAF6 pathway. Moreover, we observed that a TLR4 antibody upregulated MyD88 expression and a TLR9 inhibitor downregulated TRIF expression, indicating that there was cross-talk between TLRs 2/4/9 in MBP combined with BCG-induced Th1 activation. Our findings may expand the knowledge regarding TLR cross-talk involved in regulating the Th1 response. Copyright © 2018 Elsevier Inc. All rights reserved.

The co-stimulatory effects of MyD88-dependent Toll-like receptor signaling on activation of murine γδ T cells.

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Jinping Zhang

Full Text Available γδ T cells express several different toll-like receptor (TLRs. The role of MyD88- dependent TLR signaling in TCR activation of murine γδ T cells is incompletely defined. Here, we report that Pam3CSK4 (PAM, TLR2 agonist and CL097 (TLR7 agonist, but not lipopolysaccharide (TLR4 agonist, increased CD69 expression and Th1-type cytokine production upon anti-CD3 stimulation of γδ T cells from young adult mice (6-to 10-week-old. However, these agonists alone did not induce γδ T cell activation. Additionally, we noted that neither PAM nor CL097 synergized with anti-CD3 in inducing CD69 expression on γδ T cells of aged mice (21-to 22-month-old. Compared to young γδ T cells, PAM and CL097 increased Th-1 type cytokine production with a lower magnitude from anti-CD3- stimulated, aged γδ T cells. Vγ1+ and Vγ4+ cells are two subpopulations of splenic γδ T cells. PAM had similar effects in anti-CD3-activated control and Vγ4+ subset- depleted γδ T cells; whereas CL097 induced more IFN-γ production from Vγ4+ subset-depleted γδ T cells than from the control group. Finally, we studied the role of MyD88-dependent TLRs in γδ T cell activation during West Nile virus (WNV infection. γδ T cell, in particular, Vγ1+ subset expansion was significantly reduced in both MyD88- and TLR7- deficient mice. Treatment with TLR7 agonist induced more Vγ1+ cell expansion in wild-type mice during WNV infection. In summary, these results suggest that MyD88-dependent TLRs provide co-stimulatory signals during TCR activation of γδ T cells and these have differential effects on distinct subsets.

Chronic intermittent hypoxia induces liver fibrosis in mice with diet-induced obesity via TLR4/MyD88/MAPK/NF-kB signaling pathways.

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Kang, Hyeon Hui; Kim, In Kyoung; Lee, Hye In; Joo, Hyonsoo; Lim, Jeong Uk; Lee, Jongmin; Lee, Sang Haak; Moon, Hwa Sik

2017-08-19

Obstructive sleep apnea (OSA) is associated with nonalcoholic fatty liver disease (NAFLD), and causes chronic intermittent hypoxia (CIH) during sleep. Inflammation is associated with the development of metabolic complications induced by CIH. Research suggests that innate immune mechanisms are involved in the pro-inflammatory pathways of liver fibrosis. The purpose of this study was to investigate whether innate immune responses induce liver fibrosis, and to evaluate mechanisms underlying hepatic inflammation related to CIH in a murine diet-induced obesity (DIO) model. Inflammatory and oxidative stress markers, TLR4, MyD88, Toll/interleukin-1-receptor-domain-containing adaptor-inducing interferon-β (TRIF), I-κB, NF-κB, p38 MAPK, c-JNK, and ERK activation, were measured in the serum and liver. As a result, α1(I)-collagen mRNA was significantly higher in DIO mice exposed to CIH than in the control groups. CIH mice exhibited liver fibrosis and significantly higher protein expression of TLR4, MyD88, phosphorylated (phospho-) I-κB, and phospho-ERK1/2 activation in the liver, and higher expression of NF-κB than that in the controls. TRIF, p38 MAPK, and JNK activation did not differ significantly between groups. We conclude that CIH in DIO mice leads to liver fibrosis via TLR4/MyD88/MAPK/NF-kB signaling pathways. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

EGCG Maintains Th1/Th2 Balance and Mitigates Ulcerative Colitis Induced by Dextran Sulfate Sodium through TLR4/MyD88/NF-κB Signaling Pathway in Rats

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Xue Bing

2017-01-01

Full Text Available Objective. To observe the protective effect of epigallocatechin gallate (EGCG on dextran sulfate sodium- (DSS- induced ulcerative colitis in rats and to explore the roles of TLR4/MyD88/NF-κB signaling pathway. Methods. Rat models of ulcerative colitis were established by giving DSS. EGCG (50 mg/kg/d was given to assess disease activity index. HE staining was applied to observe histological changes. ELISA and qPCR detected the expression of inflammatory factors. Flow cytometry was used to measure the percentage of CD4+IFN-γ+ and CD4+IL-4+ in the spleen and colon. TLR4 antagonist E5564 was given in each group. Flow cytometry was utilized to detect CD4+IFN-γ+ and CD4+IL-4+ cells. Immunohistochemistry, qPCR, and western blot assay were applied to measure the expression of TLR4, MyD88, and NF-κB. Results. EGCG improved the intestinal mucosal injury in rats, inhibited production of inflammatory factors, maintained the balance of Th1/Th2, and reduced the expression of TLR4, MyD88, and NF-κB. After TLR4 antagonism, the protective effect of EGCG on intestinal mucosal injury was weakened in rats with ulcerative colitis, and the expressions of inflammatory factors were upregulated. Conclusion. EGCG can inhibit the intestinal inflammatory response by reducing the severity of ulcerative colitis and maintaining the Th1/Th2 balance through the TLR4/MyD88/NF-κB signaling pathway.

Role of ESAT-6 in renal injury by regulating microRNA-155 expression via TLR4/MyD88 signaling pathway in mice with Mycobacterium tuberculosis infection.

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Zhou, Zhong-Qi; Wang, Zhi-Kui; Zhang, Lei; Ren, Yue-Qin; Ma, Zhong-Wei; Zhao, Nan; Sun, Fu-Yun

2017-08-31

The study aims to investigate the underlying mechanism involved in the early secretory antigenic target-6 (ESAT-6) in renal injury through regulation of the expression of miR-155 through the oll-like receptor (TLR)-4 (TLR4)/myeloid differentiation factor 88 (MyD88) signaling pathway in Mycobacterium tuberculosis (MTB)-infected mice. Sixty C57BL/6 mice with MTB-induced renal injury were randomly assigned into control, MTB, mimic, inhibitor, inhibitor + ESAT6, and inhibitor + ESAT6 + TAK242 groups. Body weight, the ratio of kidney weight to body weight (Kw/Bw), blood urea nitrogen (BUN), and serum creatinine (Scr) of mice were measured. Flow cytometry was used to detect renal activation in mice. Expressions of miR-155 and ESAT6 were detected by quantitative real-time PCR (qRT-PCR), and Western blotting was used to examine the expressions of ESAT6, TLR4, and MyD88. Expressions of tumor necrosis factor-α (TNF-α), interleukin-17 (IL-17), and interferon-γ (IFN-γ) were measured by qRT-PCR and ELISA. Compared with the control group, the BUN and Scr levels as well as the expression levels of miR-155 , TLR4, MyD88, TNF-α, IL-17, and IFN-γ increased, while Kw/Bw decreased in the MTB and mimic groups. In comparison with the MTB group, the above indexes except Kw/Bw were elevated in the mimic group, but were reduced in the inhibitor group, while the Kw/Bw dropped in the mimic group but increased in the inhibitor group. Compared with the inhibitor group, the Kw/Bw decreased while the rest of the indexes increased in the inhibitor + ESAT6 group. ESAT6 may induce renal injury by promoting miR-155 expression through the TLR-4/MyD88 signaling pathway in MTB-infected mice. © 2017 The Author(s).

Inhibition of IL-1R1/MyD88 signalling promotes mesenchymal stem cell-driven tissue regeneration.

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Martino, Mikaël M; Maruyama, Kenta; Kuhn, Gisela A; Satoh, Takashi; Takeuchi, Osamu; Müller, Ralph; Akira, Shizuo

2016-03-22

Tissue injury and the healing response lead to the release of endogenous danger signals including Toll-like receptor (TLR) and interleukin-1 receptor, type 1 (IL-1R1) ligands, which modulate the immune microenvironment. Because TLRs and IL-1R1 have been shown to influence the repair process of various tissues, we explored their role during bone regeneration, seeking to design regenerative strategies integrating a control of their signalling. Here we show that IL-1R1/MyD88 signalling negatively regulates bone regeneration, in the mouse. Furthermore, IL-1β which is released at the bone injury site, inhibits the regenerative capacities of mesenchymal stem cells (MSCs). Mechanistically, IL-1R1/MyD88 signalling impairs MSC proliferation, migration and differentiation by inhibiting the Akt/GSK-3β/β-catenin pathway. Lastly, as a proof of concept, we engineer a MSC delivery system integrating inhibitors of IL-1R1/MyD88 signalling. Using this strategy, we considerably improve MSC-based bone regeneration in the mouse, demonstrating that this approach may be useful in regenerative medicine applications.

Chlamydia abortus Pmp18.1 Induces IL-1β Secretion by TLR4 Activation through the MyD88, NF-κB, and Caspase-1 Signaling Pathways.

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Pan, Qing; Zhang, Qiang; Chu, Jun; Pais, Roshan; Liu, Shanshan; He, Cheng; Eko, Francis O

2017-01-01

The polymorphic membrane protein D (Pmp18D) is a 160-kDa outer membrane protein that is conserved and plays an important role in Chlamydia abortus pathogenesis. We have identified an N-terminal fragment of Pmp18D (designated Pmp18.1) as a possible subunit vaccine antigen. In this study, we evaluated the vaccine potential of Pmp18.1 by investigating its ability to induce innate immune responses in dendritic cells and the signaling pathway(s) involved in rPmp18.1-induced IL-1β secretion. We next investigated the immunomodulatory impact of VCG, in comparison with the more established Th1-promoting adjuvants, CpG and FL, on rPmp18.1-mediated innate immune activation. Finally, the effect of siRNA targeting TLR4, MyD88, NF-κB p50, and Caspase-1 mRNA in DCs on IL-1β cytokine secretion was also investigated. Bone marrow-derived dendritic cells (BMDCs) were stimulated with rPmp18.1 in the presence or absence of VCG or CpG or FL and the magnitude of cytokines produced was assessed using a multiplex cytokine ELISA assay. Expression of costimulatory molecules and Toll-like receptors (TLRs) was analyzed by flow cytometry. Quantitation of intracellular levels of myeloid differentiation factor 88 (MyD88), nuclear factor kappa beta (NF-κB p50/p65), and Caspase-1 was evaluated by Western immunoblotting analysis while NF-κB p65 nuclear translocation was assessed by confocal microscopy. The results showed DC stimulation with rPmp18.1 provoked the secretion of proinflammatory cytokines and upregulated expression of TLRs and co-stimulatory molecules associated with DC maturation. These responses were significantly ( p ≤ 0.001) enhanced by VCG but not CpG or FL. In addition, rPmp18.1 activated the expression of MyD88, NF-κB p50, and Caspase-1 as well as the nuclear expression of NF-κB p65 in treated DCs. Furthermore, targeting TLR4, MyD88, NF-κB p50, and Caspase-1 mRNA in BMDCs with siRNA significantly reduced their expression levels, resulting in decreased IL-1β cytokine

Chlamydia abortus Pmp18.1 Induces IL-1β Secretion by TLR4 Activation through the MyD88, NF-κB, and Caspase-1 Signaling Pathways

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Qing Pan

2017-12-01

Full Text Available The polymorphic membrane protein D (Pmp18D is a 160-kDa outer membrane protein that is conserved and plays an important role in Chlamydia abortus pathogenesis. We have identified an N-terminal fragment of Pmp18D (designated Pmp18.1 as a possible subunit vaccine antigen. In this study, we evaluated the vaccine potential of Pmp18.1 by investigating its ability to induce innate immune responses in dendritic cells and the signaling pathway(s involved in rPmp18.1-induced IL-1β secretion. We next investigated the immunomodulatory impact of VCG, in comparison with the more established Th1-promoting adjuvants, CpG and FL, on rPmp18.1-mediated innate immune activation. Finally, the effect of siRNA targeting TLR4, MyD88, NF-κB p50, and Caspase-1 mRNA in DCs on IL-1β cytokine secretion was also investigated. Bone marrow-derived dendritic cells (BMDCs were stimulated with rPmp18.1 in the presence or absence of VCG or CpG or FL and the magnitude of cytokines produced was assessed using a multiplex cytokine ELISA assay. Expression of costimulatory molecules and Toll-like receptors (TLRs was analyzed by flow cytometry. Quantitation of intracellular levels of myeloid differentiation factor 88 (MyD88, nuclear factor kappa beta (NF-κB p50/p65, and Caspase-1 was evaluated by Western immunoblotting analysis while NF-κB p65 nuclear translocation was assessed by confocal microscopy. The results showed DC stimulation with rPmp18.1 provoked the secretion of proinflammatory cytokines and upregulated expression of TLRs and co-stimulatory molecules associated with DC maturation. These responses were significantly (p ≤ 0.001 enhanced by VCG but not CpG or FL. In addition, rPmp18.1 activated the expression of MyD88, NF-κB p50, and Caspase-1 as well as the nuclear expression of NF-κB p65 in treated DCs. Furthermore, targeting TLR4, MyD88, NF-κB p50, and Caspase-1 mRNA in BMDCs with siRNA significantly reduced their expression levels, resulting in decreased IL-1�

Structural complementarity of Toll/interleukin-1 receptor domains in Toll-like receptors and the adaptors Mal and MyD88.

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Dunne, Aisling; Ejdeback, Mikael; Ludidi, Phumzile L; O'Neill, Luke A J; Gay, Nicholas J

2003-10-17

The Toll/interleukin 1 receptor (TIR) domain is a region found in the cytoplasmic tails of members of the Toll-like receptor/interleukin-1 receptor superfamily. The domain is essential for signaling and is also found in the adaptor proteins Mal (MyD88 adaptor-like) and MyD88, which function to couple activation of the receptor to downstream signaling components. Experimental structures of two Toll/interleukin 1 receptor domains reveal a alpha-beta-fold similar to that of the bacterial chemotaxis protein CheY, and other evidence suggests that the adaptors can make heterotypic interactions with both the receptors and themselves. Here we show that the purified TIR domains of Mal and MyD88 can form stable heterodimers and also that Mal homodimers and oligomers are dissociated in the presence of ATP. To identify structural features that may contribute to the formation of signaling complexes, we produced models of the TIR domains from human Toll-like receptor 4 (TLR4), Mal, and MyD88. We found that although the overall fold is conserved the electrostatic surface potentials are quite distinct. Docking studies of the models suggest that Mal and MyD88 bind to different regions in TLRs 2 and 4, a finding consistent with a cooperative role of the two adaptors in signaling. Mal and MyD88 are predicted to interact at a third non-overlapping site, suggesting that the receptor and adaptors may form heterotetrameric complexes. The theoretical model of the interactions is supported by experimental data from glutathione S-transferase pull-downs and co-immunoprecipitations. Neither theoretical nor experimental data suggest a direct role for the conserved proline in the BB-loop in the association of TLR4, Mal, and MyD88. Finally we show a sequence relationship between the Drosophila protein Tube and Mal that may indicate a functional equivalence of these two adaptors in the Drosophila and vertebrate Toll pathways.

Inflammation- and tumor-induced anorexia and weight loss require MyD88 in hematopoietic/myeloid cells but not in brain endothelial or neural cells.

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Ruud, Johan; Wilhelms, Daniel Björk; Nilsson, Anna; Eskilsson, Anna; Tang, Yan-Juan; Ströhle, Peter; Caesar, Robert; Schwaninger, Markus; Wunderlich, Thomas; Bäckhed, Fredrik; Engblom, David; Blomqvist, Anders

2013-05-01

Loss of appetite is a hallmark of inflammatory diseases. The underlying mechanisms remain undefined, but it is known that myeloid differentiation primary response gene 88 (MyD88), an adaptor protein critical for Toll-like and IL-1 receptor family signaling, is involved. Here we addressed the question of determining in which cells the MyD88 signaling that results in anorexia development occurs by using chimeric mice and animals with cell-specific deletions. We found that MyD88-knockout mice, which are resistant to bacterial lipopolysaccharide (LPS)-induced anorexia, displayed anorexia when transplanted with wild-type bone marrow cells. Furthermore, mice with a targeted deletion of MyD88 in hematopoietic or myeloid cells were largely protected against LPS-induced anorexia and displayed attenuated weight loss, whereas mice with MyD88 deletion in hepatocytes or in neural cells or the cerebrovascular endothelium developed anorexia and weight loss of similar magnitude as wild-type mice. Furthermore, in a model for cancer-induced anorexia-cachexia, deletion of MyD88 in hematopoietic cells attenuated the anorexia and protected against body weight loss. These findings demonstrate that MyD88-dependent signaling within the brain is not required for eliciting inflammation-induced anorexia. Instead, we identify MyD88 signaling in hematopoietic/myeloid cells as a critical component for acute inflammatory-driven anorexia, as well as for chronic anorexia and weight loss associated with malignant disease.

Critical role of macrophages and their activation via MyD88-NFκB signaling in lung innate immunity to Mycoplasma pneumoniae.

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Jen-Feng Lai

2010-12-01

Full Text Available Mycoplasma pneumoniae (Mp, a common cause of pneumonia, is associated with asthma; however, the mechanisms underlying this association remain unclear. We investigated the cellular immune response to Mp in mice. Intranasal inoculation with Mp elicited infiltration of the lungs with neutrophils, monocytes and macrophages. Systemic depletion of macrophages, but not neutrophils, resulted in impaired clearance of Mp from the lungs. Accumulation and activation of macrophages were decreased in the lungs of MyD88(-/- mice and clearance of Mp was impaired, indicating that MyD88 is a key signaling protein in the anti-Mp response. MyD88-dependent signaling was also required for the Mp-induced activation of NFκB, which was essential for macrophages to eliminate the microbe in vitro. Thus, MyD88-NFκB signaling in macrophages is essential for clearance of Mp from the lungs.

Limb Remote Ischemic Postconditioning Reduces Ischemia-Reperfusion Injury by Inhibiting NADPH Oxidase Activation and MyD88-TRAF6-P38MAP-Kinase Pathway of Neutrophils

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Gangling Chen

2016-11-01

Full Text Available Limb remote ischemic postconditioning (LRIP has been confirmed to reduce the ischemia-reperfusion injury but its mechanisms are still not clear. This study clarified the mechanism of LRIP based on the nicotinamide-adenine dinucleotide phosphate (NADPH oxidase and Myeloid differentiation factor 88 (MyD88-Tumor necrosis factor (TNF receptor-associated factor 6 (TRAF6-P38 pathway of neutrophils. Rat middle cerebral artery occlusion (MCAO model was used in this study. Ischemia-reperfusion injury was carried out by MCAO 1.5 h followed by 24 h reperfusion. LRIP operation was performed to the left femoral artery at 0, 1 or 3 h after reperfusion. Behavioral testing, including postural reflex test, vibrissae-elicited forelimb placing test and tail hang test, showed that LRIP operated at 0 h of reperfusion could significantly ameliorate these behavioral scores. Pathological examinations, infarct size, Myeloperoxidase (MPO activity showed that LRIP operated at 0 h of reperfusion could significantly ameliorate the pathological scores, reduce the infarct size and MPO activity in the brain and increase the MPO activity in the left leg. By using Neutrophil counting, immunofluorescence and real-time PCR techniques, we found that LRIP operated at 0 h of reperfusion could reduce neutrophil counts in the peripheral blood and downregulate the activation of neutrophil in the peripheral blood and rat brain. Western blots revealed that MyD88, TRAF6, p38 mitogen-activated protein kinase (p38-MAPK in neutrophils and the phosphorylation of p47phox (Ser 304 and Ser 345 in neutrophil could be downregulated by LRIP. Our study suggests that LRIP inhibits the number and activation of neutrophils in the rat brain and peripheral blood linked to down-regulating the activation of NADPH oxidase in neutrophils by MyD88/TRAF6/p38-MAPK pathway.

Dramatic Response with Single-Agent Ibrutinib in Multiply Relapsed Marginal Zone Lymphoma with MYD88L265P Mutation

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Ryan C. Lynch

2017-09-01

Full Text Available The B-cell receptor signaling pathway is important in the lymphomagenesis of many lymphomas, including marginal zone lymphoma (MZL. Herein we describe a case of extranodal MZL refractory to multiple lines of therapy. The presence of an IgM paraprotein prompted further evaluation, and the patient was found to have an MYD88L265P mutation. Treatment with ibrutinib led to a dramatic response with prompt resolution of symptoms and significant improvement in measurable sites of disease. The excellent response to ibrutinib in our patient with MYD88L265P-mutated refractory MZL supports a biological rationale for its use.

Utility of MYD88 in the Differential Diagnosis and Choice of Second-Line Therapy in a Case of Nonsecretory Lymphoplasmacytic Lymphoma versus Free Light Chain Waldenstrom’s Macroglobulinemia

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D. Kazmierski

2017-01-01

Full Text Available The MYD88 L265P somatic variant (MYD88 has a high prevalence in Waldenstrom’s Macroglobulinemia (WM, a form of lymphoplasmacytic lymphoma (LPL associated with monoclonal IgM. Although the role of MYD88 in WM was initially reported in 2012, it was not until 2016 that MYD88 testing was included in the National Cancer Care Network (NCCN Guidelines. We present a case illustrating the utility of MYD88 status in distinguishing atypical forms of WM from marginal zone lymphoma (MZL and in selecting second-line therapy with ibrutinib. In 2012, a 64-year-old male presented with dyspnea on exertion, a hemoglobin of 5.6 g/dL, a platelet count of 86,000, and monoclonal IgM kappa on serum immunofixation but no detectable M-spike. Bone marrow biopsy revealed 95% monoclonal B-lymphocytes with lymphoplasmacytic differentiation favoring a diagnosis of LPL/WM over MZL, with a favorable response to chemotherapy. This diagnosis was called into question 3 years later following relapse, and MZL was favored based on the lack of MYD88 mutation. One year later, however, repeat bone marrow biopsy detected the MYD88 mutation and therapy with ibrutinib yielded a favorable response. The distinction between certain lymphomas can be problematic and in this case MYD88 was helpful in clarifying a diagnosis of atypical LPL/WM from MZL and in selecting effective second-line therapy.

Pepsin Digest of Wheat Gliadin Fraction Increases Production of IL-1β via TLR4/MyD88/TRIF/MAPK/NF-κB Signaling Pathway and an NLRP3 Inflammasome Activation

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Palová-Jelínková, Lenka; Dáňová, Klára; Drašarová, Hana; Dvořák, Miloš; Funda, David P.; Fundová, Petra; Kotrbová-Kozak, Anna; Černá, Marie; Kamanová, Jana; Martin, Stefan F.; Freudenberg, Marina; Tučková, Ludmila

2013-01-01

Celiac disease (CD) is a gluten-responsive, chronic inflammatory enteropathy. IL-1 cytokine family members IL-1β and IL-18 have been associated with the inflammatory conditions in CD patients. However, the mechanisms of IL-1 molecule activation in CD have not yet been elucidated. We show in this study that peripheral blood mononuclear cells (PBMC) and monocytes from celiac patients responded to pepsin digest of wheat gliadin fraction (PDWGF) by a robust secretion of IL-1β and IL-1α and a slightly elevated production of IL-18. The analysis of the upstream mechanisms underlying PDWGF-induced IL-1β production in celiac PBMC show that PDWGF-induced de novo pro-IL-1β synthesis, followed by a caspase-1 dependent processing and the secretion of mature IL-1β. This was promoted by K+ efflux and oxidative stress, and was independent of P2X7 receptor signaling. The PDWGF-induced IL-1β release was dependent on Nod-like receptor family containing pyrin domain 3 (NLRP3) and apoptosis-associated speck like protein (ASC) as shown by stimulation of bone marrow derived dendritic cells (BMDC) from NLRP3−/− and ASC−/− knockout mice. Moreover, treatment of human PBMC as well as MyD88−/− and Toll-interleukin-1 receptor domain-containing adaptor-inducing interferon-β (TRIF)−/− BMDC illustrated that prior to the activation of caspase-1, the PDWGF-triggered signal constitutes the activation of the MyD88/TRIF/MAPK/NF-κB pathway. Moreover, our results indicate that the combined action of TLR2 and TLR4 may be required for optimal induction of IL-1β in response to PDWGF. Thus, innate immune pathways, such as TLR2/4/MyD88/TRIF/MAPK/NF-κB and an NLRP3 inflammasome activation are involved in wheat proteins signaling and may play an important role in the pathogenesis of CD. PMID:23658628

Kaempferol alleviates LPS-induced neuroinflammation and BBB dysfunction in mice via inhibiting HMGB1 release and down-regulating TLR4/MyD88 pathway.

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Cheng, Xiao; Yang, Ying-Lin; Yang, Huan; Wang, Yue-Hua; Du, Guan-Hua

2018-03-01

Kaempferol is a natural flavonoid with many biological activities including anti-oxidation and anti-inflammation. Nevertheless, its anti-neuroinflammation role and the relevant mechanism remain unclear. The present study was to investigate effects of kaempferol against LPS-induced neuroinflammation and blood-brain barrier dysfunction as well as the mechanism in mice. BALB/c mice were treated with LPS 5mg/kg to induce inflammation after pre-treatment with kaempferol 25, 50, or 100mg/kg for 7days. The results showed that kaempferol reduced the production of various pro-inflammatory factors and inflammatory proteins including IL-1β, IL-6, TNF-α, MCP-1, COX-2 and iNOS in brain tissues. In addition, kaempferol also protected BBB integrity and increased BBB related proteins including occludin-1, claudin-1 and CX43 in brain of LPS-induced mice. Furthermore, kaempferol significantly reduced HMGB1 level and suppressed TLR4/MyD88 inflammatory pathway in both transcription level and translation level. These results collectively suggested that kaempferol might be a promising neuroprotective agent for alleviating inflammatory responses and BBB dysfunction by inhibiting HMGB1 release and down-regulating TLR4/MyD88 inflammatory pathway. Copyright © 2018 Elsevier B.V. All rights reserved.

Decreased serum level of HMGB1 and MyD88 during human aging progress in healthy individuals.

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Fu, Guo-Xiang; Chen, Alex F; Zhong, Yuan; Zhao, Jian; Gu, Ying-Jia

2016-04-01

Previous studies have suggested that high mobility group box-1 protein (HMGB1) binds to the toll-like receptor 4 (TLR4) signaling mediates the progression of various inflammatory diseases. But the roles of HMGB1 and TLR4 in aging remain poorly unknown. In this study, we aimed to investigate the serum levels of HMGB1 and myeloid differentiation factor 88 (MyD88), which is one of TLR4's intracellular adaptor proteins during human aging process and their relevance with cathepsin B (CTSB). This research was conducted using the blood samples provided by healthy people (n = 90, 63 men and 27 women). Subjects were subdivided into groups with respect to age: young (about 25 years old, n = 30), middle age (about 40 years old, n = 30), and aged (above 65 years old, n = 30). Altered serum levels of HMGB1, MyD88 and CTSB were measured using an enzyme-linked immunosorbent assay. The serum levels of HMGB1 and MyD88 were significantly decreased in the aged group compared with those in the young group. Linear regression analysis showed that HMGB1 and MyD88 positively correlated with CTSB among the whole healthy people. A negative correlation was determined between MyD88 and age. The serum levels of HMGB1 and MyD88 significantly decreased with age. MyD88, but not HMGB1, was negatively correlated with age.

Appropriate Development of the Liver Treg Compartment Is Modulated by the Microbiota and Requires TGF-β and MyD88

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Ann Maria

2014-01-01

Full Text Available Neither the early postnatal development of the liver Treg compartment nor the factors that regulate its development has been characterized. We compared the early developmental patterns of Treg cell accumulation in murine liver, thymus, and spleen. A FoxP3EGFP reporter mouse was employed to identify Treg cells. Mononuclear cells were isolated from organs postnatally, stained for CD4, and examined by flow cytometry to enumerate FoxP3+CD4hi cells. To assess roles for TGF-β1, MyD88, and TLR2, gene-specific knockout pups were generated from heterozygous breeders. To test the role of commensal bacteria, pregnant dams were administered antibiotics during gestation and after parturition. The pattern of appearance of Treg cells differed in liver, spleen, and thymus. Notably, at 1-2 weeks, the frequency of CD4hi FoxP3+ T cells in liver exceeded that in spleen by 1.5- to 2-fold. The relative increase in liver Treg frequency was transient and was dependent upon TGF-β1 and MyD88, but not TLR2, and was abrogated by antibiotic treatment. A relative increase in liver Treg frequency occurs approximately 1-2 weeks after parturition that appears to be driven by colonization of the intestine with commensal bacteria and is mediated by a pathway that requires TGF-β1 and MyD88, but not TLR2.

Anti-Inflammatory Activity of Bee Venom in BV2 Microglial Cells: Mediation of MyD88-Dependent NF-κB Signaling Pathway.

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Im, Eun Ju; Kim, Su Jung; Hong, Seung Bok; Park, Jin-Kyu; Rhee, Man Hee

2016-01-01

Bee venom has long been used as a traditional folk medicine in Korea. It has been reportedly used for the treatment of arthritis, cancer, and inflammation. Although its anti-inflammatory activity in lipopolysaccharide- (LPS-) stimulated inflammatory cells has been reported, the exact mechanism of its anti-inflammatory action has not been fully elucidated. Therefore, the aim of this study was to investigate the anti-inflammatory mechanism of bee venom in BV2 microglial cells. We first investigated whether NO production in LPS-activated BV2 cells was inhibited by bee venom, and further iNOS mRNA and protein expressions were determined. The mRNA and protein levels of proinflammatory cytokines were examined using semiquantitative RT-PCR and immunoblotting, respectively. Moreover, modulation of the transcription factor NF-κB by bee venom was also investigated using a luciferase assay. LPS-induced NO production in BV2 microglial cells was significantly inhibited in a concentration-dependent manner upon pretreatment with bee venom. Bee venom markedly reduced the mRNA expression of COX-2, TNF-α, IL-1β, and IL-6 and suppressed LPS-induced activation of MyD88 and IRAK1 and phosphorylation of TAK1. Moreover, NF-κB translocation by IKKα/β phosphorylation and subsequent IκB-α degradation were also attenuated. Thus, collectively, these results indicate that bee venom exerts its anti-inflammatory activity via the IRAK1/TAK1/NF-κB signaling pathway.

Atorvastatin attenuates experimental contrast-induced acute kidney injury: a role for TLR4/MyD88 signaling pathway.

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Yue, Rongzheng; Zuo, Chuan; Zeng, Jing; Su, Baihai; Tao, Ye; Huang, Songmin; Zeng, Rui

2017-11-01

To investigate the protective effect of different atorvastatin doses on contrast-induced acute kidney injury and the related mechanism. Healthy male Sprague-Dawley (SD) rats were randomly divided into the blank control group, experimental control group and different-dose atorvastatin groups. A rat model of contrast-induced acute kidney injury was established. We detected changes in serum creatinine (Scr) and blood urea nitrogen (BUN) before and after model establishment, observed and scored renal tubular injury, analyzed rat renal cell apoptosis, and measure the expression of signal pathway proteins and downstream inflammatory factors. After contrast agent injection, the Scr and BUN levels of the experimental control group were significantly increased, the different doses applied in the atorvastatin group significantly reduced the Scr and BUN levels (p atorvastatin doses have protective effects on contrast-induced acute renal tubular injury in rats, possibly by targeting TLR4, suppressing TLR4 expression, regulating the TLR4/Myd88 signaling pathway, and inhibiting the expression of downstream inflammatory factors.

Signaling flux redistribution at toll-like receptor pathway junctions.

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Kumar Selvarajoo

Full Text Available Various receptors on cell surface recognize specific extracellular molecules and trigger signal transduction altering gene expression in the nucleus. Gain or loss-of-function mutations of one molecule have shown to affect alternative signaling pathways with a poorly understood mechanism. In Toll-like receptor (TLR 4 signaling, which branches into MyD88- and TRAM-dependent pathways upon lipopolysaccharide (LPS stimulation, we investigated the gain or loss-of-function mutations of MyD88. We predict, using a computational model built on the perturbation-response approach and the law of mass conservation, that removal and addition of MyD88 in TLR4 activation, enhances and impairs, respectively, the alternative TRAM-dependent pathway through signaling flux redistribution (SFR at pathway branches. To verify SFR, we treated MyD88-deficient macrophages with LPS and observed enhancement of TRAM-dependent pathway based on increased IRF3 phosphorylation and induction of Cxcl10 and Ifit2. Furthermore, increasing the amount of MyD88 in cultured cells showed decreased TRAM binding to TLR4. Investigating another TLR4 pathway junction, from TRIF to TRAF6, RIP1 and TBK1, the removal of MyD88-dependent TRAF6 increased expression of TRAM-dependent Cxcl10 and Ifit2. Thus, we demonstrate that SFR is a novel mechanism for enhanced activation of alternative pathways when molecules at pathway junctions are removed. Our data suggest that SFR may enlighten hitherto unexplainable intracellular signaling alterations in genetic diseases where gain or loss-of-function mutations are observed.

Sepsis Induces Hematopoietic Stem Cell Exhaustion and Myelosuppression through Distinct Contributions of TRIF and MYD88

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Huajia Zhang

2016-06-01

Full Text Available Toll-like receptor 4 (TLR4 plays a central role in host responses to bacterial infection, but the precise mechanism(s by which its downstream signaling components coordinate the bone marrow response to sepsis is poorly understood. Using mice deficient in TLR4 downstream adapters MYD88 or TRIF, we demonstrate that both cell-autonomous and non-cell-autonomous MYD88 activation are major causes of myelosuppression during sepsis, while having a modest impact on hematopoietic stem cell (HSC functions. In contrast, cell-intrinsic TRIF activation severely compromises HSC self-renewal without directly affecting myeloid cells. Lipopolysaccharide-induced activation of MYD88 or TRIF contributes to cell-cycle activation of HSC and induces rapid and permanent changes in transcriptional programs, as indicated by persistent downregulation of Spi1 and CebpA expression after transplantation. Thus, distinct mechanisms downstream of TLR4 signaling mediate myelosuppression and HSC exhaustion during sepsis through unique effects of MyD88 and TRIF.

MYD88 L265P Mutations Are Correlated with 6q Deletion in Korean Patients with Waldenström Macroglobulinemia

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Jung-Ah Kim

2014-01-01

Full Text Available Waldenström macroglobulinemia (WM is a malignant lymphoplasma-proliferative disorder with IgM monoclonal gammopathy. A recent whole-genome study identified MYD88 L265P as the key mutation in WM. We investigated MYD88 mutations in conjunction with cytogenetic study in 22 consecutive Korean WM patients. Conventional G-banding and interphase fluorescence in situ hybridization (FISH were performed at regions including 6q21 using bone marrow (BM aspirates. Sixteen patients were subjected to Sanger sequencing-based MYD88 mutation study. Five patients (28% showed cytogenetic aberrations in G-banding. The incidence of 6q21 deletion was 17% by conventional G-banding and 37% by FISH. Ten patients (45% showed cytogenetic aberrations using FISH: 6q deletion in eight (37% and IGH rearrangement in four (18%. Two patients had both the 6q deletion and IGH rearrangement, and two had only the IGH rearrangement. Eleven patients (69% presented with the MYD88 L265P mutation. MYD88 mutations were significantly associated with the presence of 6q deletions (P=0.037. Six patients with the 6q deletion for whom sequencing was possible were found to harbor MYD88 mutations. The MYD88 L265P mutation was also associated with increased lymphocyte burden in BM biopsy. This is the first report of high frequency MYD88 L265P mutations in Korean WM patients.

Curcumin attenuates acute inflammatory injury by inhibiting the TLR4/MyD88/NF-κB signaling pathway in experimental traumatic brain injury

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2014-01-01

neuron co-culture following treatment with curcumin after LPS stimulation. LPS increased TLR4 immunoreactivity and morphological activation in microglia and increased neuronal apoptosis, whereas curcumin normalized this upregulation. The increased protein levels of TLR4, MyD88 and NF-κB in microglia were attenuated by curcumin treatment. Conclusions Our results suggest that post-injury, curcumin administration may improve patient outcome by reducing acute activation of microglia/macrophages and neuronal apoptosis through a mechanism involving the TLR4/MyD88/NF-κB signaling pathway in microglia/macrophages in TBI. PMID:24669820

The attenuated inflammation of MPL is due to the lack of CD14-dependent tight dimerization of the TLR4/MD2 complex at the plasma membrane.

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Tanimura, Natsuko; Saitoh, Shin-Ichiroh; Ohto, Umeharu; Akashi-Takamura, Sachiko; Fujimoto, Yukari; Fukase, Koichi; Shimizu, Toshiyuki; Miyake, Kensuke

2014-06-01

TLR4/MD-2 senses lipid A, activating the MyD88-signaling pathway on the plasma membrane and the TRIF-signaling pathway after CD14-mediated TLR4/MD-2 internalization into endosomes. Monophosphoryl lipid A (MPL), a detoxified derivative of lipid A, is weaker than lipid A in activating the MyD88-dependent pathway. Little is known, however, about mechanisms underlying the attenuated activation of MyD88-dependent pathways. We here show that MPL was impaired in induction of CD14-dependent TLR4/MD-2 dimerization compared with lipid A. Impaired TLR4/MD-2 dimerization decreased CD14-mediated TNFα production. In contrast, MPL was comparable to lipid A in CD14-independent MyD88-dependent TNFα production and TRIF-dependent responses including cell surface CD86 up-regulation and IFNβ induction. Although CD86 up-regulation is dependent on TRIF signaling, it was induced by TLR4/MD-2 at the plasma membrane. These results revealed that the attenuated MPL responses were due to CD14-initiated responses at the plasma membrane, but not just to responses initiated by MyD88, that is, MPL was specifically unable to induce CD14-dependent TLR4/MD-2 dimerization that selectively enhances MyD88-mediated responses at the plasma membrane. © The Japanese Society for Immunology. 2013. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Positive Correlation between Enhanced Expression of TLR4/MyD88/NF-κB with Insulin Resistance in Placentae of Gestational Diabetes Mellitus.

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Hui Feng

Full Text Available Insulin resistance (IR is a critical factor of the pathophysiology of Gestational diabetes mellitus (GDM. Studies on key organs involved in IR, such as livers and adipose tissues, showed that Toll-like receptor 4 (TLR4 can regulate insulin sensitivity. As a maternal-fetal interface with multi-functions, placentae could contribute to the development of IR for GDM. Thus, we investigated the expressions of TLR4/Myeloid Differentiation factor 88 (MyD88/Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-kB in term placentae from 33 GDM women and 36 healthy pregnant women with normal glucose tolerance, evaluated local and systemic IR and furthermore identified the association between placental TLR4 and IR. TLR4 protein was expressed in various cells of term placenta, particularly in syncytiotrophoblast of villi. Compared with normal pregnancy, the expression of TLR4/MyD88/NF-kB pathway increased in the placenta of GDM (p<0.05, and these differences were more pronounced in the maternal section of the placenta and the syncytiotrophoblast of villi. In addition, more severe IR was observed in the placenta of GDM patients than the control group, evidenced with higher pIRS-1(ser312 (p<0.001 and lower IRS-1 (p<0.05 as well as pAkt proteins (p<0.01. The expression of TLR4 in placentae is positively correlated with local IR (pIRS-1: r = 0.76, p <0.001 and pAkt: r = -0.47, p <0.001 and maternal fasting (r = 0.42, p <0.01, one-hour (r = 0.52, p <0.01 and two-hour glucose (r = 0.54, p <0.01 at OGTT. We found an that enhanced expression of the TLR4-MyD88-NF-kB pathway occurs in GDM placentae, which positively correlates with heightened local IR in placentae and higher maternal hyperglycemia. The TLR4/MyD88/NF-kB pathway may play a potential role in the development of IR in placentae of GDM.

Role of a transductional-transcriptional processor complex involving MyD88 and IRF-7 in Toll-like receptor signaling

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Honda, Kenya; Yanai, Hideyuki; Mizutani, Tatsuaki; Negishi, Hideo; Shimada, Naoya; Suzuki, Nobutaka; Ohba, Yusuke; Takaoka, Akinori; Yeh, Wen-Chen; Taniguchi, Tadatsugu

2004-01-01

Toll-like receptor (TLR) activation is central to immunity, wherein the activation of the TLR9 subfamily members TLR9 and TLR7 results in the robust induction of type I IFNs (IFN-α/β) by means of the MyD88 adaptor protein. However, it remains unknown how the TLR signal “input” can be processed through MyD88 to “output” the induction of the IFN genes. Here, we demonstrate that the transcription factor IRF-7 interacts with MyD88 to form a complex in the cytoplasm. We provide evidence that this complex also involves IRAK4 and TRAF6 and provides the foundation for the TLR9-dependent activation of the IFN genes. The complex defined in this study represents an example of how the coupling of the signaling adaptor and effector kinase molecules together with the transcription factor regulate the processing of an extracellular signal to evoke its versatile downstream transcriptional events in a cell. Thus, we propose that this molecular complex may function as a cytoplasmic transductional-transcriptional processor. PMID:15492225

In Vivo Role of TLR2 and MyD88 Signaling in Eliciting Innate Immune Responses in Staphylococcal Endophthalmitis

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Talreja, Deepa; Singh, Pawan Kumar; Kumar, Ashok

2015-01-01

Purpose. The purpose of this study was to investigate the protective mechanisms evoked by TLR2 and MyD88 signaling in bacterial endophthalmitis in vivo. Methods. Endophthalmitis was induced in wild-type (WT), TLR2−/−, MyD88−/−, and Cnlp−/− mice by intravitreal injections of a laboratory strain (RN6390) and two endophthalmitis isolates of Staphylococcus aureus. Disease progression was monitored by assessing corneal and vitreous haze, bacterial burden, and retinal tissue damage. Levels of inflammatory cytokines/chemokines were determined using quantitative RT-PCR (qRT-PCR) and ELISA. Flow cytometry was used to assess neutrophil infiltration. Cathelicidin-related antimicrobial peptide (CRAMP) expression was determined by immunostaining and dot blot. Results. Eyes infected with either laboratory or clinical isolates exhibited higher levels of inflammatory mediators at the early stages of infection (≤24 hours) in WT mice than in TLR2−/− or MyD88−/− mice. However, their levels surpassed that of WT mice at the later stages of infection (>48 hours), coinciding with increased bacterial burden and retinal damage. Both TLR2−/− and MyD88−/− retinas produced reduced levels of CRAMP, and its deficiency (Cnlp−/−) rendered the mice susceptible to increased bacterial burden and retinal tissue damage as early as 1 day post infection. Analyses of inflammatory mediators and neutrophil levels in WT versus Cnlp−/− mice showed a trend similar to that observed in TLR2 and MyD88 KO mice. Furthermore, we observed that even a 10-fold lower infective dose of S. aureus was sufficient to cause endophthalmitis in TLR2−/− and MyD88−/− mice. Conclusions. TLR2 and MyD88 signaling plays an important role in protecting the retina from staphylococcal endophthalmitis by production of the antimicrobial peptide CRAMP. PMID:25678692

Bacterial lipoprotein-induced self-tolerance and cross-tolerance to LPS are associated with reduced IRAK-1 expression and MyD88-IRAK complex formation.

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Li, Chong Hui

2012-02-03

Tolerance to bacterial cell-wall components may represent an essential regulatory mechanism during bacterial infection. We have demonstrated previously that the inhibition of nuclear factor (NF)-kappaB and mitogen-activated protein kinase activation was present in bacterial lipoprotein (BLP) self-tolerance and its cross-tolerance to lipopolysaccharide (LPS). In this study, the effect of BLP-induced tolerance on the myeloid differentiation factor 88 (MyD88)-dependent upstream signaling pathway for NF-kappaB activation in vitro was examined further. When compared with nontolerant human monocytic THP-1 cells, BLP-tolerant cells had a significant reduction in tumor necrosis factor alpha (TNF-alpha) production in response to a high-dose BLP (86+\\/-12 vs. 6042+\\/-245 ng\\/ml, P < 0.01) or LPS (341+\\/-36 vs. 7882+\\/-318 ng\\/ml, P < 0.01) stimulation. The expression of Toll-like receptor 2 (TLR2) protein was down-regulated in BLP-tolerant cells, whereas no significant differences in TLR4, MyD88, interleukin-1 receptor-associated kinase 4 (IRAK-4), and TNF receptor-associated factor 6 expression were observed between nontolerant and BLP-tolerant cells, as confirmed by Western blot analysis. The IRAK-1 protein was reduced markedly in BLP-tolerant cells, although IRAK-1 mRNA expression remained unchanged as revealed by real-time reverse transcriptase-polymerase chain reaction analysis. Furthermore, decreased MyD88-IRAK immunocomplex formation, as demonstrated by immunoprecipitation, was observed in BLP-tolerant cells following a second BLP or LPS stimulation. BLP pretreatment also resulted in a marked inhibition in total and phosphorylated inhibitor of kappaB-alpha (IkappaB-alpha) expression, which was not up-regulated by subsequent BLP or LPS stimulation. These results demonstrate that in addition to the down-regulation of TLR2 expression, BLP tolerance is associated with a reduction in IRAK-1 expression, MyD88-IRAK association, and IkappaB-alpha phosphorylation. These

MyD88 and STING signaling pathways are required for IRF3-mediated IFN-β induction in response to Brucella abortus infection.

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Leonardo A de Almeida

Full Text Available Type I interferons (IFNs are cytokines that orchestrate diverse immune responses to viral and bacterial infections. Although typically considered to be most important molecules in response to viruses, type I IFNs are also induced by most, if not all, bacterial pathogens. In this study, we addressed the role of type I IFN signaling during Brucella abortus infection, a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. Herein, we have shown that B. abortus induced IFN-β in macrophages and splenocytes. Further, IFN-β induction by Brucella was mediated by IRF3 signaling pathway and activates IFN-stimulated genes via STAT1 phosphorylation. In addition, IFN-β expression induced by Brucella is independent of TLRs and TRIF signaling but MyD88-dependent, a pathway not yet described for Gram-negative bacteria. Furthermore, we have identified Brucella DNA as the major bacterial component to induce IFN-β and our study revealed that this molecule operates through a mechanism dependent on RNA polymerase III to be sensed probably by an unknown receptor via the adaptor molecule STING. Finally, we have demonstrated that IFN-αβR KO mice are more resistant to infection suggesting that type I IFN signaling is detrimental to host control of Brucella. This resistance phenotype is accompanied by increased IFN-γ and NO production by IFN-αβR KO spleen cells and reduced apoptosis.

MyD88 and STING Signaling Pathways Are Required for IRF3-Mediated IFN-β Induction in Response to Brucella abortus Infection

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de Almeida, Leonardo A.; Carvalho, Natalia B.; Oliveira, Fernanda S.; Lacerda, Thais L. S.; Vasconcelos, Anilton C.; Nogueira, Lucas; Bafica, Andre; Silva, Aristóbolo M.; Oliveira, Sergio C.

2011-01-01

Type I interferons (IFNs) are cytokines that orchestrate diverse immune responses to viral and bacterial infections. Although typically considered to be most important molecules in response to viruses, type I IFNs are also induced by most, if not all, bacterial pathogens. In this study, we addressed the role of type I IFN signaling during Brucella abortus infection, a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. Herein, we have shown that B. abortus induced IFN-β in macrophages and splenocytes. Further, IFN-β induction by Brucella was mediated by IRF3 signaling pathway and activates IFN-stimulated genes via STAT1 phosphorylation. In addition, IFN-β expression induced by Brucella is independent of TLRs and TRIF signaling but MyD88-dependent, a pathway not yet described for Gram-negative bacteria. Furthermore, we have identified Brucella DNA as the major bacterial component to induce IFN-β and our study revealed that this molecule operates through a mechanism dependent on RNA polymerase III to be sensed probably by an unknown receptor via the adaptor molecule STING. Finally, we have demonstrated that IFN-αβR KO mice are more resistant to infection suggesting that type I IFN signaling is detrimental to host control of Brucella. This resistance phenotype is accompanied by increased IFN-γ and NO production by IFN-αβR KO spleen cells and reduced apoptosis. PMID:21829705

A TIR domain variant of MyD88 adapter-like (Mal)/TIRAP results in loss of MyD88 binding and reduced TLR2/TLR4 signaling.

NARCIS (Netherlands)

Nagpal, K.; Plantinga, T.S.; Wong, J.; Monks, B.G.; Gay, N.J.; Netea, M.G.; Fitzgerald, K.A.; Golenbock, D.

2009-01-01

The adapter protein MyD88 adapter-like (Mal), encoded by TIR-domain containing adapter protein (Tirap) (MIM 606252), is the most polymorphic of the five adapter proteins involved in Toll-like receptor signaling, harboring eight non-synonymous single nucleotide polymorphisms in its coding region. We

Temporal Role for MyD88 in a Model of Brucella-Induced Arthritis and Musculoskeletal Inflammation.

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Lacey, Carolyn A; Mitchell, William J; Brown, Charles R; Skyberg, Jerod A

2017-03-01

Brucella spp. are facultative intracellular Gram-negative bacteria that cause the zoonotic disease brucellosis, one of the most common global zoonoses. Osteomyelitis, arthritis, and musculoskeletal inflammation are common focal complications of brucellosis in humans; however, wild-type (WT) mice infected systemically with conventional doses of Brucella do not develop these complications. Here we report C57BL/6 WT mice infected via the footpad with 10 3 to 10 6 CFU of Brucella spp. display neutrophil and monocyte infiltration of the joint space and surrounding musculoskeletal tissue. Joint inflammation is detectable as early as 1 day postinfection and peaks 1 to 2 weeks later, after which WT mice are able to slowly resolve inflammation. B and T cells were dispensable for the onset of swelling but required for resolution of joint inflammation and infection. At early time points, MyD88 -/- mice display decreased joint inflammation, swelling, and proinflammatory cytokine levels relative to WT mice. Subsequently, swelling of MyD88 -/- joints surpassed WT joint swelling, and resolution of joint inflammation was prolonged. Joint bacterial loads in MyD88 -/- mice were significantly greater than those in WT mice by day 3 postinfection and at all time points thereafter. In addition, MyD88 -/- joint inflammatory cytokine levels on day 3 and beyond were similar to WT levels. Collectively these data demonstrate MyD88 signaling mediates early inflammatory responses in the joint but also contributes to subsequent clearance of Brucella and resolution of inflammation. This work also establishes a mouse model for studying Brucella -induced arthritis, musculoskeletal complications, and systemic responses, which will lead to a better understanding of focal complications of brucellosis. Copyright © 2017 American Society for Microbiology.

Structure-Based Design and Synthesis of a Small Molecule that Exhibits Anti-inflammatory Activity by Inhibition of MyD88-mediated Signaling to Bacterial Toxin Exposure.

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Alam, Shahabuddin; Javor, Sacha; Degardin, Melissa; Ajami, Dariush; Rebek, Mitra; Kissner, Teri L; Waag, David M; Rebek, Julius; Saikh, Kamal U

2015-08-01

Both Gram-positive and Gram-negative pathogens or pathogen-derived components, such as staphylococcal enterotoxins (SEs) and endotoxin (LPS) exposure, activate MyD88-mediated pro-inflammatory cellular immunity for host defense. However, dysregulated MyD88-mediated signaling triggers exaggerated immune response that often leads to toxic shock and death. Previously, we reported a small molecule compound 1 mimicking BB-loop structure of MyD88 was capable of inhibiting pro-inflammatory response to SEB exposure in mice. In this study, we designed a dimeric structure compound 4210 covalently linked with compound 1 by a non-polar cyclohexane linker which strongly inhibited the production of pro-inflammatory cytokines in human primary cells to SEB (IC50 1-50 μm) or LPS extracted from Francisella tularensis, Escherichia coli, or Burkholderia mallei (IC50 10-200 μm). Consistent with cytokine inhibition, in a ligand-induced cell-based reporter assay, compound 4210 inhibited Burkholderia mallei or LPS-induced MyD88-mediated NF-kB-dependent expression of reporter activity (IC50 10-30 μm). Furthermore, results from a newly expressed MyD88 revealed that 4210 inhibited MyD88 dimer formation which is critical for pro-inflammatory signaling. Importantly, a single administration of compound 4210 in mice showed complete protection from lethal toxin challenge. Collectively, these results demonstrated that compound 4210 inhibits toxin-induced inflated pro-inflammatory immune signaling, thus displays a potential bacterial toxin therapeutic. © 2014 John Wiley & Sons A/S.

Bifidobacterium breve reduces apoptotic epithelial cell shedding in an exopolysaccharide and MyD88-dependent manner.

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Hughes, K R; Harnisch, L C; Alcon-Giner, C; Mitra, S; Wright, C J; Ketskemety, J; van Sinderen, D; Watson, A J M; Hall, L J

2017-01-01

Certain members of the microbiota genus Bifidobacterium are known to positively influence host well-being. Importantly, reduced bifidobacterial levels are associated with inflammatory bowel disease (IBD) patients, who also have impaired epithelial barrier function, including elevated rates of apoptotic extrusion of small intestinal epithelial cells (IECs) from villi-a process termed 'cell shedding'. Using a mouse model of pathological cell shedding, we show that mice receiving Bifidobacterium breve UCC2003 exhibit significantly reduced rates of small IEC shedding. Bifidobacterial-induced protection appears to be mediated by a specific bifidobacterial surface exopolysaccharide and interactions with host MyD88 resulting in downregulation of intrinsic and extrinsic apoptotic responses to protect epithelial cells under highly inflammatory conditions. Our results reveal an important and previously undescribed role for B. breve, in positively modulating epithelial cell shedding outcomes via bacterial- and host-dependent factors, supporting the notion that manipulation of the microbiota affects intestinal disease outcomes. © 2017 The Authors.

Analysis of gut microbial regulation of host gene expression along the length of the gut and regulation of gut microbial ecology through MyD88.

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Larsson, Erik; Tremaroli, Valentina; Lee, Ying Shiuan; Koren, Omry; Nookaew, Intawat; Fricker, Ashwana; Nielsen, Jens; Ley, Ruth E; Bäckhed, Fredrik

2012-08-01

The gut microbiota has profound effects on host physiology but local host-microbial interactions in the gut are only poorly characterised and are likely to vary from the sparsely colonised duodenum to the densely colonised colon. Microorganisms are recognised by pattern recognition receptors such as Toll-like receptors, which signal through the adaptor molecule MyD88. To identify host responses induced by gut microbiota along the length of the gut and whether these required MyD88, transcriptional profiles of duodenum, jejunum, ileum and colon were compared from germ-free and conventionally raised wild-type and Myd88-/- mice. The gut microbial ecology was assessed by 454-based pyrosequencing and viruses were analysed by PCR. The gut microbiota modulated the expression of a large set of genes in the small intestine and fewer genes in the colon but surprisingly few microbiota-regulated genes required MyD88 signalling. However, MyD88 was essential for microbiota-induced colonic expression of the antimicrobial genes Reg3β and Reg3γ in the epithelium, and Myd88 deficiency was associated with both a shift in bacterial diversity and a greater proportion of segmented filamentous bacteria in the small intestine. In addition, conventionally raised Myd88-/- mice had increased expression of antiviral genes in the colon, which correlated with norovirus infection in the colonic epithelium. This study provides a detailed description of tissue-specific host transcriptional responses to the normal gut microbiota along the length of the gut and demonstrates that the absence of MyD88 alters gut microbial ecology.

The absence of MyD88 has no effect on the induction of alternatively activated macrophage during Fasciola hepatica infection

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Luo HongLin

2011-11-01

Full Text Available Abstract Background Alternatively activated macrophages (AAMϕ play important roles in allergies and responses to parasitic infections. However, whether signaling through toll-like receptors (TLRs plays any role in AAMϕ induction when young Fasciola hepatica penetrates the liver capsule and migrates through the liver tissue is still unclear. Results The data show that the lack of myeloid differentiation factor 88 (MyD88 has no effect on the AAMϕ derived from the bone marrow (BMMϕ in vitro and does not impair the mRNA expression of arginase-1, resistin-like molecule (RELMα, and Ym1 in BMMϕs. The Th2 cytokine production bias in splenocytes was not significantly altered in F. hepatica-infected mice in the absence of MyD88 in vitro and in the pleural cavity lavage in vivo. In addition, MyD88-deficiency has no effect on the arginase production of the F. hepatica elicited macrophages (Fe Mϕs, production of RELMα and Ym1 proteins and mRNA expression of Ym1 and RELMα of macrophages in the peritoneal cavity 6 weeks post F. hepatica infection. Conclusions The absence of MyD88 has no effect on presence of AAMϕ 6 weeks post F. hepatica infection.

MiR-23a-5p modulates mycobacterial survival and autophagy during mycobacterium tuberculosis infection through TLR2/MyD88/NF-κB pathway by targeting TLR2.

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Gu, Xing; Gao, Yan; Mu, De-Guang; Fu, En-Qing

2017-05-15

Autophagy plays a pivotal role in activating the antimicrobial host defense against Mycobacterium tuberculosis (M.tb.). The emerging roles of microRNAs (miRNAs) in regulating immune responses have attracted increasing attention in recent years. Appreciating the potential of host-directed therapies designed to control autophagy during mycobacterial infection, we focused on the influence of miR-23a-5p on the activation of macrophage autophagy during M.tb. infection in bone marrow-derived macrophages (BMDMs) and murine RAW264.7 cells. Here, we demonstrated that M.tb.-infection of macrophages lead to markedly enhanced expression of miR-23a-5p in a time- and dose-dependent manner. Furthermore, forced expression of miR-23a-5p accelerated the survival rate of intracellular mycobacteria, while transfection with miR-23a-5p inhibitors attenuated mycobacterial survival. More importantly, overexpression of miR-23a-5p dramatically prevented M.tb.-induced activation of autophagy in macrophages, whereas inhibitors of miR-23a-5p remarkably accelerated M.tb.-induced autophagy. Mechanistically, miR-23a-5p is able to modulate TLR2/MyD88/NF-κB signaling activity by targeting TLR2 in RAW264.7 cells in response to M.tb.-infection. Collectively, these findings demonstrated that miR-23a-5p modulated the innate host defense by promoting mycobacteria survival and inhibiting the activation of autophagy against M.tb. through TLR2/MyD88/NF-κB pathway by targeting TLR2, which may provide a promising therapeutic target for tuberculosis. Copyright © 2017 Elsevier Inc. All rights reserved.

MyD88 mediates in vivo effector functions of alveolar macrophages in acute lung inflammatory responses to carbon nanotube exposure

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Frank, Evan A. [Division of Environmental Genetics and Molecular Toxicology, Department of Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267 (United States); Birch, M. Eileen [National Institute for Occupational Safety and Health, Cincinnati, OH 45213 (United States); Yadav, Jagjit S., E-mail: Jagjit.Yadav@uc.edu [Division of Environmental Genetics and Molecular Toxicology, Department of Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267 (United States)

2015-11-01

Carbon nanotubes (CNTs) are rapidly emerging as high-priority occupational toxicants. CNT powders contain fibrous particles that aerosolize readily in places of manufacture and handling, posing an inhalation risk for workers. Studies using animal models indicate that lung exposure to CNTs causes prolonged inflammatory responses and diffuse alveolar injury. The mechanisms governing CNT-induced lung inflammation are not fully understood but have been suggested to involve alveolar macrophages (AMs). In the current study, we sought to systematically assess the effector role of AMs in vivo in the induction of lung inflammatory responses to CNT exposures and investigate their cell type-specific mechanisms. Multi-wall CNTs characterized for various physicochemical attributes were used as the CNT type. Using an AM-specific depletion and repopulation approach in a mouse model, we unambiguously demonstrated that AMs are major effector cells necessary for the in vivo elaboration of CNT-induced lung inflammation. We further investigated in vitro AM responses and identified molecular targets which proved critical to pro-inflammatory responses in this model, namely MyD88 as well as MAPKs and Ca{sup 2} {sup +}/CamKII. We further demonstrated that MyD88 inhibition in donor AMs abrogated their capacity to reconstitute CNT-induced inflammation when adoptively transferred into AM-depleted mice. Taken together, this is the first in vivo demonstration that AMs act as critical effector cell types in CNT-induced lung inflammation and that MyD88 is required for this in vivo effector function. AMs and their cell type-specific mechanisms may therefore represent potential targets for future therapeutic intervention of CNT-related lung injury. - Highlights: • Demonstrated in vivo effector role of alveolar macrophages (AMs) in CNT toxicity • MyD88, MAPKs, and Ca{sup 2} {sup +}/CamKII are required for AM inflammatory responses in vitro. • MyD88 signaling is required for in vivo effector

Recurrent candidiasis and early-onset gastric cancer in a patient with a genetically defined partial MYD88 defect.

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Vogelaar, Ingrid P; Ligtenberg, Marjolijn J L; van der Post, Rachel S; de Voer, Richarda M; Kets, C Marleen; Jansen, Trees J G; Jacobs, Liesbeth; Schreibelt, Gerty; de Vries, I Jolanda M; Netea, Mihai G; Hoogerbrugge, Nicoline

2016-04-01

Gastric cancer is caused by both genetic and environmental factors. A woman who suffered from recurrent candidiasis throughout her life developed diffuse-type gastric cancer at the age of 23 years. Using whole-exome sequencing we identified a germline homozygous missense variant in MYD88. Immunological assays on peripheral blood mononuclear cells revealed an impaired immune response upon stimulation with Candida albicans, characterized by a defective production of the cytokine interleukin-17. Our data suggest that a genetic defect in MYD88 results in an impaired immune response and may increase gastric cancer risk.

Clone-specific MYD88 L265P and CXCR4 mutation status can provide clinical utility in suspected Waldenström macroglobulinemia/lymphoplasmacytic lymphoma.

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Burnworth, Bettina; Wang, Zhixing; Singleton, Timothy P; Bennington, Angela; Fritschle, Wayne; Bennington, Richard; Brodersen, Lisa Eidenschink; Wells, Denise A; Loken, Michael R; Zehentner, Barbara K

2016-12-01

MYD88 L265P, a diagnostic marker for lymphoplasmacytic lymphoma (LPL)/Waldenström macroglobulinemia (WM) can also be detected in other hematopoietic malignancies. We demonstrate a novel approach to increase the specificity of this marker for WM/LPL diagnosis by combining flow cytometric cell sorting with molecular analysis. Clonal B-lymphocyte and co-occurring clonal plasma cell populations of low-grade B-cell lymphomas were sorted by flow cytometry and analyzed for immunoglobulin gene rearrangements (PCR), and for MYD88 and CXCR4 mutations. Identical clonal origin was confirmed by PCR for 21 LPL/WM cases and MYD88 L265P was detected in both B-cell and plasma cell fractions. 9/20 other B-cell lymphomas with identical light chain restriction on B-cells and plasma cells were genotypically identical by PCR and MYD88 L265P was detected in both cell fractions in 7/9 whereas in 11/20 specimens with different clonal origin, MYD88 L265P was absent (5/11), or only found in B-lymphocytes (4/11), or plasma cells (2/11). CXCR4 mutations were detected in 17/39 cases, but missed in 63% of these without cell sorting. Confirming MYD88L265P in both B-cells and plasma cell fractions can provide a novel and powerful discriminator to distinguish LPL/WM from phenotypically similar disorders. Furthermore, this approach significantly increases CXCR4 detection sensitivity. Copyright © 2016 Elsevier Ltd. All rights reserved.

Patients with primary breast and primary female genital tract diffuse large B cell lymphoma have a high frequency of MYD88 and CD79B mutations.

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Cao, Xin-Xin; Li, Jian; Cai, Hao; Zhang, Wei; Duan, Ming-Hui; Zhou, Dao-Bin

2017-11-01

This study is to retrospectively evaluate the prevalence of MYD88 and CD79B mutations and the clinicopathologic characteristics of patients with primary diffuse large B cell lymphoma (DLBCL) of the female genital tract and breast. The characteristics, treatments, and outcomes of 19 patients diagnosed with primary DLBCL of the female genital tract and breast, who had formalin-fixed and paraffin-embedded tissues obtained from diagnostic samples diagnosed between January 2004 and June 2016, were analyzed retrospectively. Nineteen female patients (7 with primary breast and 12 with primary female genital tract DLBCL) were included in this retrospective study. Eleven patients (57.9%) carried a MYD88 mutation, including 10 with MYD8 L265P and 1 with the MYD88 L265S mutation. Seven patients (36.8%) harbored a CD79B mutation, which included two cases with CD79B Y196H, two cases with CD79B Y196N, one case with CD79B Y196D, one case with CD79B Y196F, and one case with CD79B Y196X. Four cases had both MYD88 and CD79B mutations. The clinicopathologic parameters, progression-free survival (PFS), and overall survival (OS) of the MYD88 mutation-carrying group were not significantly different from those of the MYD88 wild-type group except for higher LDH levels. Six patients received cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP), while 13 patients received rituximab plus CHOP, and 13 patients received central nervous system prophylaxis. The median OS and PFS were 73 and 56 months, respectively. Patients with primary breast and primary female genital tract DLBCL have a high frequency of MYD88 and CD79B mutations. The presence of these mutations does not affect survival but may offer additional therapeutic options.

TLR4/MyD88/NF-κB signaling and PPAR-γ within the paraventricular nucleus are involved in the effects of telmisartan in hypertension

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Li, Hong-Bao; Li, Xiang; Huo, Chan-Juan; Su, Qing; Guo, Jing [Department of Physiology and Pathophysiology, Key Laboratory of Environment and Genes Related to Diseases, Xi' an Jiaotong University School of Basic Medical Sciences, Xi' an Jiaotong University Health Science Center, Xi' an Jiaotong University Cardiovascular Research Center, Xi' an 710061 (China); Yuan, Zu-Yi [Department of Cardiovascular Medicine, First Affiliated Hospital of Medical College of Xi' an Jiaotong University, Xi' an 710061 (China); Zhu, Guo-Qing [Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Physiology, Nanjing Medical University, Nanjing 210029 (China); Shi, Xiao-Lian, E-mail: shxl@mail.xjtu.edu.cn [Department of Pharmacology, School of Basic Medical Sciences, Xi' an Jiaotong University Health Science Center, Xi' an 710061 (China); Liu, Jin-Jun, E-mail: jupet@163.com [Department of Physiology and Pathophysiology, Xi' an Jiaotong University School of Basic Medical Sciences, Xi' an Jiaotong University Health Science Center, Xi' an 710061 (China); Kang, Yu-Ming, E-mail: ykang@mail.xjtu.edu.cn [Department of Physiology and Pathophysiology, Key Laboratory of Environment and Genes Related to Diseases, Xi' an Jiaotong University School of Basic Medical Sciences, Xi' an Jiaotong University Health Science Center, Xi' an Jiaotong University Cardiovascular Research Center, Xi' an 710061 (China)

2016-08-15

Previous findings from our laboratory and others indicate that the main therapeutic effect of angiotensin II type 1 receptor (AT1-R) antagonists is to decrease blood pressure and exert anti-inflammatory effects in the cardiovascular system. In this study, we determined whether AT1-R antagonist telmisartan within the hypothalamic paraventricular nucleus (PVN) attenuates hypertension and hypothalamic inflammation via both the TLR4/MyD88/NF-κB signaling pathway and peroxisome proliferator-activated receptor-γ (PPAR-γ) in the PVN in hypertensive rats. Spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto (WKY) rats were treated for 4 weeks through bilateral PVN infusion with the AT1-R antagonist telmisartan (TEL, 10 μg/h), or losartan (LOS, 20 μg/h), or the PPAR-γ antagonist GW9662 (GW, 100 μg/h), or vehicle via osmotic minipump. Mean arterial pressure (MAP) was recorded by a tail-cuff occlusion method. PVN tissue and blood were collected for the measurement of AT1-R, PPAR-γ, pro-inflammatory cytokines (tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6), inducible nitric oxide synthase (iNOS), TLR4, MyD88, nuclear factor-kappa B (NF-κB) activity and plasma norepinephrine (NE), respectively. Hypertensive rats exhibited significantly higher level of AT1-R and lower level of PPAR-γ in the PVN. PVN treatment with TEL attenuated MAP, improved cardiac hypertrophy, reduced TNF-α, IL-1β, IL-6, iNOS levels, and plasma NE in SHR but not in WKY rats. These results were associated with reduced TLR4, MyD88 and NF-κB levels and increased PPAR-γ level in the PVN of hypertensive rats. Our findings suggest that TLR4/MyD88/NF-κB signaling and PPAR-γ within the PVN are involved in the beneficial effects of telmisartan in hypertension. - Highlights: • PVN infusion of TEL in spontaneously hypertensive rats is reported. • PVN infusion of TEL attenuates hypertension and proinflammatory cytokines in PVN. • PVN blockade of AT1-R attenuates

RNA-Seq Analysis of IL-1B and IL-36 Responses in Epidermal Keratinocytes Identifies a Shared MyD88-Dependent Gene Signature.

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Swindell, William R; Beamer, Maria A; Sarkar, Mrinal K; Loftus, Shannon; Fullmer, Joseph; Xing, Xianying; Ward, Nicole L; Tsoi, Lam C; Kahlenberg, Michelle J; Liang, Yun; Gudjonsson, Johann E

2018-01-01

IL-36 cytokines have recently emerged as mediators of inflammation in autoimmune conditions including psoriasis vulgaris (PsV) and generalized pustular psoriasis (GPP). This study used RNA-seq to profile the transcriptome of primary epidermal keratinocytes (KCs) treated with IL-1B, IL-36A, IL-36B, or IL-36G. We identified some early IL-1B-specific responses (8 h posttreatment), but nearly all late IL-1B responses were replicated by IL-36 cytokines (24 h posttreatment). Type I and II interferon genes exhibited time-dependent response patterns, with early induction (8 h) followed by no response or repression (24 h). Altogether, we identified 225 differentially expressed genes (DEGs) with shared responses to all 4 cytokines at both time points (8 and 24 h). These involved upregulation of ligands ( IL1A, IL1B , and IL36G ) and activating proteases ( CTSS ) but also upregulation of inhibitors such as IL1RN and IL36RN . Shared IL-1B/IL-36 DEGs overlapped significantly with genes altered in PsV and GPP skin lesions, as well as genes near GWAS loci linked to autoimmune and autoinflammatory diseases (e.g., PsV, psoriatic arthritis, inflammatory bowel disease, and primary biliary cholangitis). Inactivation of MyD88 adapter protein using CRISPR/Cas9 completely abolished expression responses of such DEGs to IL-1B and IL-36G stimulation. These results provide a global view of IL-1B and IL-36 expression responses in epidermal KCs with fine-scale characterization of time-dependent and cytokine-specific response patterns. Our findings support an important role for IL-1B and IL-36 in autoimmune or autoinflammatory conditions and show that MyD88 adaptor protein mediates shared IL-1B/IL-36 responses.

Inhibitors of MyD88-dependent proinflammatory cytokine production identified utilizing a novel RNA interference screening approach.

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John S Cho

2009-09-01

Full Text Available The events required to initiate host defenses against invading pathogens involve complex signaling cascades comprised of numerous adaptor molecules, kinases, and transcriptional elements, ultimately leading to the production of proinflammatory cytokines, such as tumor necrosis factor alpha (TNF-alpha. How these signaling cascades are regulated, and the proteins and regulatory elements participating are still poorly understood.We report here the development a completely random short-hairpin RNA (shRNA library coupled with a novel forward genetic screening strategy to identify inhibitors of Toll-like receptor (TLR dependent proinflammatory responses. We developed a murine macrophage reporter cell line stably transfected with a construct expressing diphtheria toxin-A (DT-A under the control of the TNF-alpha-promoter. Stimulation of the reporter cell line with the TLR ligand lipopolysaccharide (LPS resulted in DT-A induced cell death, which could be prevented by the addition of an shRNA targeting the TLR adaptor molecule MyD88. Utilizing this cell line, we screened a completely random lentiviral short hairpin RNA (shRNA library for sequences that inhibited TLR-mediated TNF-alpha production. Recovery of shRNA sequences from surviving cells led to the identification of unique shRNA sequences that significantly inhibited TLR4-dependent TNF-alpha gene expression. Furthermore, these shRNA sequences specifically blocked TLR2 but not TLR3-dependent TNF-alpha production.Thus, we describe the generation of novel tools to facilitate large-scale forward genetic screens in mammalian cells and the identification of potent shRNA inhibitors of TLR2 and TLR4- dependent proinflammatory responses.

Cloning and functional characterization of IRAK4 in large yellow croaker (Larimichthys crocea) that associates with MyD88 but impairs NF-κB activation.

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Zou, Peng Fei; Huang, Xue Na; Yao, Cui Luan; Sun, Qing Xue; Li, Ying; Zhu, Qian; Yu, Zhen Xing; Fan, Ze Jun

2017-04-01

As crucial signaling transducer in Toll-like receptor (TLR) and interleukin (IL)-1 receptor (IL-1R) signaling pathway, IL-1R-associated kinase 4 (IRAK4) mediates downstream signaling cascades and plays important roles in innate and adaptive immune responses. In the present study, an IRAK4 orthologue was characterized from large yellow croaker (Larimichthys crocea), named Lc-IRAK4, with a conservative N-terminal death domain and a C-terminal protein kinase domain. The genome of Lc-IRAK4 is structured into eleven exons and ten introns. Expression analysis indicated that Lc-IRAK4 was widely expressed in tested tissues, with the highest level in liver and weakest in muscle. Additionally, in the spleen, liver tissues and blood, it could be induced by poly I:C and LPS stimulation, but not be induced by Vibrio parahemolyticus infection. Fluorescence microscopy assays revealed that Lc-IRAK4 localized in the cytoplasm in HEK 293T cells. It was also determined that Lc-IRAK4 could interact with MyD88, whereas MyD88-mediated NF-κB activation was significantly impaired when co-transfected the two in HEK 293T cells. These findings collectively indicated that although Lc-IRAK4 was evolutionarily conserved in vertebrates, the exact function especially the signaling transduction mediated by IRAK4 in fish immune response was different from that in mammals, which impaired MyD88-mediated NF-κB activation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Retinal dendritic cell recruitment, but not function, was inhibited in MyD88 and TRIF deficient mice.

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Heuss, Neal D; Pierson, Mark J; Montaniel, Kim Ramil C; McPherson, Scott W; Lehmann, Ute; Hussong, Stacy A; Ferrington, Deborah A; Low, Walter C; Gregerson, Dale S

2014-08-13

Immune system cells are known to affect loss of neurons due to injury or disease. Recruitment of immune cells following retinal/CNS injury has been shown to affect the health and survival of neurons in several models. We detected close, physical contact between dendritic cells and retinal ganglion cells following an optic nerve crush, and sought to understand the underlying mechanisms. CD11c-DTR/GFP mice producing a chimeric protein of diphtheria toxin receptor (DTR) and GFP from a transgenic CD11c promoter were used in conjunction with mice deficient in MyD88 and/or TRIF. Retinal ganglion cell injury was induced by an optic nerve crush, and the resulting interactions of the GFPhi cells and retinal ganglion cells were examined. Recruitment of GFPhi dendritic cells to the retina was significantly compromised in MyD88 and TRIF knockout mice. GFPhi dendritic cells played a significant role in clearing fluorescent-labeled retinal ganglion cells post-injury in the CD11c-DTR/GFP mice. In the TRIF and MyD88 deficient mice, the resting level of GFPhi dendritic cells was lower, and their influx was reduced following the optic nerve crush injury. The reduction in GFPhi dendritic cell numbers led to their replacement in the uptake of fluorescent-labeled debris by GFPlo microglia/macrophages. Depletion of GFPhi dendritic cells by treatment with diphtheria toxin also led to their displacement by GFPlo microglia/macrophages, which then assumed close contact with the injured neurons. The contribution of recruited cells to the injury response was substantial, and regulated by MyD88 and TRIF. However, the presence of these adaptor proteins was not required for interaction with neurons, or the phagocytosis of debris. The data suggested a two-niche model in which resident microglia were maintained at a constant level post-optic nerve crush, while the injury-stimulated recruitment of dendritic cells and macrophages led to their transient appearance in numbers equivalent to or greater

Hepatitis C Virus NS3 Mediated Microglial Inflammation via TLR2/TLR6 MyD88/NF-κB Pathway and Toll Like Receptor Ligand Treatment Furnished Immune Tolerance.

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Ayilam Ramachandran Rajalakshmy

Full Text Available Recent evidence suggests the neurotrophic potential of hepatitis C virus (HCV. HCV NS3 protein is one of the potent antigens of this virus mediating inflammatory response in different cell types. Microglia being the immune surveillance cells in the central nervous system (CNS, the inflammatory potential of NS3 on microglia was studied. Role of toll like receptor (TLR ligands Pam2CSK3 and Pam3CSK4 in controlling the NS3 mediated microglial inflammation was studied using microglial cell line CHME3.IL (Interleukin-8, IL-6, TNF-α (Tumor nicrosis factor alpha and IL-1β gene expressions were measured by semi quantitative RT-PCR (reverse transcription-PCR. ELISA was performed to detect IL-8, IL-6, TNF-α, IL-1β and IL-10 secretion. FACS (Flourescent activated cell sorting was performed to quantify TLR1, TLR2, TLR6, MyD88 (Myeloid differntiation factor 88, IkB-α (I kappaB alpha and pNF-κB (phosphorylated nuclear factor kappaB expression. Immunofluorescence staining was performed for MyD88, TLR6 and NF-κB (Nuclear factor kappaB. Student's t-test or One way analysis of variance with Bonferoni post hoc test was performed and p < 0.05 was considered significant.Microglia responded to NS3 by secreting IL-8, IL-6, TNF-α and IL-1β via TLR2 or TLR6 mediated MyD88/NF-κB pathway. Transcription factor NF-κB was involved in activating the cytokine gene expression and the resultant inflammatory response was controlled by NF-κB inhibitor, Ro106-9920, which is known to down regulate pro-inflammatory cytokine secretion. Activation of the microglia by TLR agonists Pam3CSK4 and Pam2CSK4 induced immune tolerance against NS3. TLR ligand treatment significantly down regulated pro-inflammatory cytokine secretion in the microglia. IL-10 secretion was suggested as the possible mechanism by which TLR agonists induced immune tolerance. NS3 as such was not capable of self-inducing immune tolerance in microglia.In conclusion, NS3 protein was capable of activating

MyD88 Signaling Regulates Steady-State Migration of Intestinal CD103+ Dendritic Cells Independently of TNF-α and the Gut Microbiota

DEFF Research Database (Denmark)

Hägerbrand, Karin; Westlund, Jessica; Yrlid, Ulf

2015-01-01

Intestinal homeostasis and induction of systemic tolerance to fed Ags (i.e., oral tolerance) rely on the steady-state migration of smallintestinal lamina propria dendritic cells (DCs) into draining mesenteric lymph nodes (MLN). The majority of these migratory DCs express the α integrin chain CD103......, and in this study we demonstrate that the steady-state mobilization of CD103+ DCs into the MLN is in part governed by the IL-1R family/TLR signaling adaptor molecule MyD88. Similar to mice with complete MyD88 deficiency, specific deletion of MyD88 in DCs resulted in a 50–60% reduction in short-term accumulation......, the proportion and phenotypic composition of DCs were similar in mesentericlymph from germ-free and conventionally housed mice. Although TNF-α was required for CD103+ DC migration to the MLN after oral administration of the TLR7 agonist R848, it was not required for the steady-state migration of these cells...

Kaempferol ameliorates H9N2 swine influenza virus-induced acute lung injury by inactivation of TLR4/MyD88-mediated NF-κB and MAPK signaling pathways.

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Zhang, Ruihua; Ai, Xia; Duan, Yongjie; Xue, Man; He, Wenxiao; Wang, Cunlian; Xu, Tong; Xu, Mingju; Liu, Baojian; Li, Chunhong; Wang, Zhijun; Zhang, Ruihong; Wang, Guohua; Tian, Shufei; Liu, Huifeng

2017-05-01

Kaempferol, a very common type of dietary flavonoids, has been found to exert antioxidative and anti-inflammatory properties. The purpose of our investigation was designed to reveal the effect of kaempferol on H9N2 influenza virus-induced inflammation in vivo and in vitro. In vivo, BALB/C mice were infected intranasally with H9N2 influenza virus with or without kaempferol treatment to induce acute lung injury (ALI) model. In vitro, MH-S cells were infected with H9N2 influenza virus with or without kaempferol treatment. In vivo, kaempferol treatment attenuated pulmonary edema, the W/D mass ratio, pulmonary capillary permeability, myeloperoxidase (MPO) activity, and the numbers of inflammatory cells. Kaempferol reduced ROS and Malondialdehyde (MDA) production, and increased the superoxide dismutase (SOD) activity. Kaempferol also reduced overproduction of TNF-α, IL-1β and IL-6. In addition, kaempferol decreased the H9N2 viral titre. In vitro, ROS, MDA, TNF-α, IL-1β and IL-6 was also reduced by kaempferol. Moreover, our data showed that kaempferol significantly inhibited the upregulation of toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), phosphorylation level of IκBα and nuclear factor-κB (NF-κB) p65, NF-κB p65 DNA binding activity, and phosphorylation level of MAPKs, both in vivo and in vitro. These results suggest that kaempferol exhibits a protective effect on H9N2 virus-induced inflammation via suppression of TLR4/MyD88-mediated NF-κB and MAPKs pathways, and kaempferol may be considered as an effective drug for the potential treatment of influenza virus-induced ALI. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Multiple Roles of Myd88 in the Immune Response to the Plague F1-V Vaccine and in Protection against an Aerosol Challenge of Yersinia pestis CO92 in Mice

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Jennifer L. Dankmeyer

2014-01-01

Full Text Available The current candidate vaccine against Yersinia pestis infection consists of two subunit proteins: the capsule protein or F1 protein and the low calcium response V protein or V-antigen. Little is known of the recognition of the vaccine by the host’s innate immune system and how it affects the acquired immune response to the vaccine. Thus, we vaccinated Toll-like receptor (Tlr 2, 4, and 2/4-double deficient, as well as signal adaptor protein Myd88-deficient mice. We found that Tlr4 and Myd88 appeared to be required for an optimal immune response to the F1-V vaccine but not Tlr2 when compared to wild-type mice. However, there was a difference between the requirement for Tlr4 and MyD88 in vaccinated animals. When F1-V vaccinated Tlr4 mutant (lipopolysaccharide tolerant and Myd88-deficient mice were challenged by aerosol with Y. pestis CO92, all but one Tlr4 mutant mice survived the challenge, but no vaccinated Myd88-deficient mice survived the challenge. Spleens from these latter nonsurviving mice showed that Y. pestis was not cleared from the infected mice. Our results suggest that MyD88 appears to be important for both an optimal immune response to F1-V and in protection against a lethal challenge of Y. pestis CO92 in F1-V vaccinated mice.

Commensal bacteria-dependent select expression of CXCL2 contributes to periodontal tissue homeostasis.

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Zenobia, Camille; Luo, Xiao Long; Hashim, Ahmed; Abe, Toshiharu; Jin, Lijian; Chang, Yucheng; Jin, Zhi Chao; Sun, Jian Xun; Hajishengallis, George; Curtis, Mike A; Darveau, Richard P

2013-08-01

The oral and intestinal host tissues both carry a heavy microbial burden. Although commensal bacteria contribute to healthy intestinal tissue structure and function, their contribution to oral health is poorly understood. A crucial component of periodontal health is the recruitment of neutrophils to periodontal tissue. To elucidate this process, gingival tissues of specific-pathogen-free and germ-free wild-type mice and CXCR2KO and MyD88KO mice were examined for quantitative analysis of neutrophils and CXCR2 chemoattractants (CXCL1, CXCL2). We show that the recruitment of neutrophils to the gingival tissue does not require commensal bacterial colonization but is entirely dependent on CXCR2 expression. Strikingly, however, commensal bacteria selectively upregulate the expression of CXCL2, but not CXCL1, in a MyD88-dependent way that correlates with increased neutrophil recruitment as compared with germ-free conditions. This is the first evidence that the selective use of chemokine receptor ligands contributes to neutrophil homing to healthy periodontal tissue. © 2013 John Wiley & Sons Ltd.

Recurrent candidiasis and early-onset gastric cancer in a patient with a genetically defined partial MYD88 defect

NARCIS (Netherlands)

Vogelaar, Ingrid P.; Ligtenberg, Marjolijn J L; van der Post, Rachel S.; de Voer, Richarda M.; Kets, C. Marleen; Jansen, Trees J G; Jacobs, Liesbeth; Schreibelt, Gerty; de Vries, I. Jolanda M; Netea, Mihai G.; Hoogerbrugge, Nicoline; Lubinski, Jan; Jakubowska, Anna; Teodorczyk, Urszula; Schackert, Hans K.; Aalfs, Cora M.; Gómez García, Encarna B.; Ranzani, Guglielmina N.; Molinaro, Valeria; van Hest, Liselotte P.; Hes, Frederik J.; Holinski-Feder, Elke; Genuardi, Maurizio; Ausems, Margreet G E M; Sijmons, Rolf H.; Wagner, Anja; van der Kolk, Lizet E.; Pinheiro, Hugo; Oliveira, Carla; Bjørnevoll, Inga; Høberg Vetti, Hildegunn; Van Krieken, J. Han J M

2016-01-01

Gastric cancer is caused by both genetic and environmental factors. A woman who suffered from recurrent candidiasis throughout her life developed diffuse-type gastric cancer at the age of 23 years. Using whole-exome sequencing we identified a germline homozygous missense variant in MYD88.

DNA methylcytosine dioxygenase ten-eleven translocation 2 enhances lipopolysaccharide-induced cytokine expression in human dental pulp cells by regulating MyD88 hydroxymethylation.

Science.gov (United States)

Wang, Xinxuan; Feng, Zhihui; Li, Qimeng; Yi, Baicheng; Xu, Qiong

2018-04-13

Dental pulp inflammation is a bacterially driven inflammation process characterized by the local accumulation of cytokines/chemokines that participate in destructive processes in the pulp. Multiple mechanisms are involved in dental pulp inflammation, including epigenetic events, such as DNA methylation/demethylation. Ten-eleven translocation 2 (TET2) is a recently discovered DNA methylcytosine dioxygenase that plays important roles in inflammatory disease. However, its role in the inflammatory response of dental pulp is unknown. We observed elevated mRNA and protein levels of TET2 after lipopolysaccharide (LPS) stimulation in human dental pulp cells (hDPCs). To identify the effects of TET2 on cytokine expression, TET2 was knocked down and cytokines were detected using a cytokine antibody array after LPS stimulation. The protein expression of GM-CSF, IL-6, IL-8 and RANTES decreased in the LPS-induced hDPCs following TET2 knockdown. The downregulated expression levels of IL-6 and IL-8 were further confirmed by real-time quantitative polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). Additionally, the phosphorylation levels of IKK-α/β, p65 and IκBα of the NF-κB signaling pathway were decreased in the TET2-silenced group. Furthermore, the global 5-hydroxymethylcytosine (5hmC) level was significantly decreased and the genomic 5-methylcytosine (5mC) level was increased in the TET2-deficient hDPCs; TET2 depletion resulted in a decrease in the 5hmC level of the MyD88 promoter following LPS stimulation. These findings indicate that TET2 knockdown inhibits LPS-induced inflammatory response in hDPCs by downregulating MyD88 hydroxymethylation. Thus, TET2-dependent DNA demethylation might play an important role in dental pulp inflammation as an epigenetic regulator.

Taurine Attenuates Hepatic Inflammation in Chronic Alcohol-Fed Rats Through Inhibition of TLR4/MyD88 Signaling.

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Lin, Chao-Jen; Chiu, Chun-Ching; Chen, Yi-Chen; Chen, Mu-Lin; Hsu, Tsai-Ching; Tzang, Bor-Show

2015-12-01

Accumulating evidence indicates that overconsumption of ethanol contributes in many ways to the pathogenesis of hepatic injury. Although studies indicate that taurine decreases lipogenesis, oxidative stress, and inflammatory cytokines, the protective effect of taurine against alcohol-induced liver injury is still unclear. To clarify the precise signaling involved in the beneficial effect of taurine on alcohol-induced liver injury, rats were randomly divided into four treatment groups: (1) control (Ctl), (2) alcohol (Alc), (3) Alc+taurine (Tau), and (4) Alc+silymarin (Sil). The Tau and Sil groups had lower lymphocyte infiltration and significantly lower TLR-4/MyD88 and IκB/NFκB compared to the Alc group. The inducible nitric oxide synthase (iNOS), C-reactive protein (CRP), tumor necrosis factors (TNF)-α, interleukin (IL)-6, and IL-1β were also significantly lower in the Tau and Sil groups than in the Alc group. The experimental results indicated that hepatoprotection against alcohol-induced inflammation may be mediated by decreased TLR-4/MyD88 signaling.

Repeated stimulation by LPS promotes the senescence of DPSCs via TLR4/MyD88-NF-κB-p53/p21 signaling.

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Feng, Guijuan; Zheng, Ke; Cao, Tong; Zhang, Jinlong; Lian, Min; Huang, Dan; Wei, Changbo; Gu, Zhifeng; Feng, Xingmei

2018-02-26

Dental pulp stem cells (DPSCs), one type of mesenchymal stem cells, are considered to be a type of tool cells for regenerative medicine and tissue engineering. Our previous studies found that the stimulation with lipopolysaccharide (LPS) might introduce senescence of DPSCs, and this senescence would have a positive correlation with the concentration of LPS. The β-galactosidase (SA-β-gal) staining was used to evaluate the senescence of DPSCs and immunofluorescence to show the morphology of DPSCs. Our findings suggested that the activity of SA-β-gal has increased after repeated stimulation with LPS and the morphology of DPSCs has changed with the stimulation with LPS. We also found that LPS bound to the Toll-like receptor 4 (TLR4)/myeloid differentiation factor (MyD) 88 signaling pathway. Protein and mRNA expression of TLR4, MyD88 were enhanced in DPSCs with LPS stimulation, resulting in the activation of nuclear factor-κB (NF-κB) signaling, which exhibited the expression of p65 improved in the nucleus while the decreasing of IκB-α. Simultaneously, the expression of p53 and p21, the downstream proteins of the NF-κB signaling, has increased. In summary, DPSCs tend to undergo senescence after repeated stimulation in an inflammatory microenvironment. Ultimately, these findings may lead to a new direction for cell-based therapy in oral diseases and other regenerative medicines.

The use of droplet digital PCR in liquid biopsies: A highly sensitive technique for MYD88 p.(L265P) detection in cerebrospinal fluid.

Science.gov (United States)

Hiemcke-Jiwa, Laura S; Minnema, Monique C; Radersma-van Loon, Joyce H; Jiwa, N Mehdi; de Boer, Mirthe; Leguit, Roos J; de Weger, Roel A; Huibers, Manon M H

2018-04-01

The gold standard for diagnosis of central nervous system lymphomas still regards a stereotactic brain biopsy, with the risk of major complications for the patient. As tumor cells can be detected in cerebrospinal fluid (CSF), CSF analysis can be used as an alternative. In this respect, mutation analysis in CSF can be of added value to other diagnostic parameters such a cytomorphology and clonality analysis. A well-known example of targeted mutation analysis entails MYD88 p.(L265P) detection, which is present in the majority of Bing Neel syndrome and primary central nervous system lymphoma (PCNSL) patients. Unfortunately, tumor yield in CSF can be very low. Therefore, use of the highly sensitive droplet digital PCR (ddPCR) might be a suitable analysis strategy for targeted mutation detection. We analyzed 26 formalin fixed paraffin embedded (FFPE) samples (8 positive and 18 negative for MYD88 p.(L265P) mutation) by ddPCR, of which the results were compared with next generation sequencing (NGS). Subsequently, 32 CSF samples were analyzed by ddPCR. ddPCR and NGS results on FFPE material showed 100% concordance. Among the 32 CSF samples, 9 belonged to patients with lymphoplasmacytic lymphoma (LPL) and clinical suspicion of Bing Neel syndrome, and 3 belonged to patients with PCNSL. Nine of these samples tested positive for MYD88 p.(L265P) (8 LPL and 1 PCNSL). This study shows that sensitive MYD88 mutation analysis by ddPCR in CSF is highly reliable and can be applied even when DNA input is low. Therefore, ddPCR is of added value to current diagnostic parameters, especially when the available amount of DNA is limited. Copyright © 2017 John Wiley & Sons, Ltd.

Loss of BMI-1 dampens migration and EMT of colorectal cancer in inflammatory microenvironment through TLR4/MD-2/MyD88-mediated NF-κB signaling.

Science.gov (United States)

Ye, Kai; Chen, Qi-Wei; Sun, Ya-Feng; Lin, Jian-An; Xu, Jian-Hua

2018-02-01

Increasing evidence from various clinical and experimental studies has demonstrated that the inflammatory microenvironment created by immune cells facilitates tumor migration. Epithelial-mesenchymal transition (EMT) is involved in the progression of cancer invasion and metastasis in an inflammatory microenvironment. B-lymphoma Moloney murine leukemia virus insertion region 1 (BMI-1) acts as an oncogene in various tumors. Ectopic expression of Bmi-1 have an effect on EMT and invasiveness. The purpose of this study was to investigate the efficacy of BMI-1 on inflammation-induced tumor migration and EMT and the underlying mechanism. We observed that the expression of BMI-1, TNF-α, and IL-1β was significantly increased in HT29 and HCT116 cells after THP-1 Conditioned-Medium (THP-1-CM) stimulation. Additionally, inhibition of BMI-1 impeded cell invasion induced by THP-1-CM-stimulation in both HT29 and HCT116 cells. BMI-1 knockdown remarkably repressed THP-1-CM-induced EMT by regulating the expression of EMT biomarkers with an increase in E-cadherin accompanied by decrease in N-cadherin and vimentin. Furthermore, downregulation of BMI-1 dramatically impeded THP-1-CM-triggered Toll-like receptor 4(TLR4)/myeloid differentiation protein 2(MD-2)/myeloid differentiation factor 88(MyD88) activity by repressing the expression of the TLR4/MD-2 complex and MyD88. Further data demonstrated that knockout of BMI-1 also dampened NF-κB THP-1-CM-triggered activity. Taken all data together, our findings established that BMI-1 modulated TLR4/MD-2/MyD88 complex-mediated NF-κB signaling involved in inflammation-induced cancer cells invasion and EMT, and therefore, could be a potential chemopreventive agent against inflammation-associated colorectal cancer. Establishment of an inflammatory microenvironment. Suppression of BMI-1 reverses THP-1-CM-induced inflammatory cytokine production in CRC. Loss of BMI-1 suppressed TLR4/MD-2/MyD88 complex-mediated NF-κB signaling. © 2017 Wiley

Whole-body vibration improves the anti-inflammatory status in elderly subjects through toll-like receptor 2 and 4 signaling pathways.

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Rodriguez-Miguelez, Paula; Fernandez-Gonzalo, Rodrigo; Collado, Pilar S; Almar, Mar; Martinez-Florez, Susana; de Paz, José A; González-Gallego, Javier; Cuevas, María J

2015-09-01

Regular physical exercise has anti-inflammatory effects in elderly subjects. Yet, the inflammatory responses after whole body vibration (WBV) training, a popular exercise paradigm for the elderly, remain to be elucidated. This study assessed the effects of WBV training on the inflammatory response associated with toll-like receptors (TLRs) signaling pathways. Twenty-eight subjects were randomized to a training group (TG) or a control group (CG). TG followed an 8-week WBV training program. Blood samples were obtained before and after the training period in both groups. Peripheral blood mononuclear cells were isolated, and mRNA and protein levels of makers involved in the TLR2/TLR4 myeloid differentiation primary response gen 88 (MyD88) and TIR domain-containing adaptor inducing interferon (TRIF)-dependent pathways were analyzed. Plasma TNFα and C-reactive protein levels were also assessed. The WBV program reduced protein expression of TLR2, TLR4, MyD88, p65, TRIF and heat shock protein (HSP) 60, while HSP70 content increased. IL-10 mRNA level and protein concentration were upregulated, and TNFα protein content decreased, after WBV training. Plasma concentration of C-reactive protein and TNFα decreased in the TG. The current data suggest WBV may improve the anti-inflammatory status of elderly subjects through an attenuation of MyD88- and TRIF-dependent TLRs signaling pathways. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

mTOR inhibition in macrophages of asymptomatic HIV+ persons reverses the decrease in TLR4-mediated TNFα release through prolongation of MAPK pathway activation1

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Li, Xin; Han, Xinbing; Llano, Juliana; Bole, Medhavi; Zhou, Xiuqin; Swan, Katharine; Anandaiah, Asha; Nelson, Benjamin; Patel, Naimish R.; Reinach, Peter S.; Koziel, Henry; Tachado, Souvenir D.

2011-01-01

Toll-like receptor 4 (TLR4) mediated signaling is significantly impaired in macrophages from HIV+ persons predominantly due to altered MyD88-dependent pathway signaling caused in part by constitutive activation of PI3K. Here we assessed in these macrophages if the blunted increase in TLR4-mediated TNFα release induced by lipid A are associated with PI3K-induced upregulation of mammalian target of rapamycin (mTOR) activity. mTOR inhibition with rapamycin enhanced TLR4-mediated TNFα release, but instead suppressed anti-inflammatory IL-10 release. Targeted gene silencing of mTOR in macrophages resulted in lipid A-induced TNFα and IL-10 release patterns similar to those induced by rapamycin. Rapamycin restored MyD88-IRAK interaction in a dose-dependent manner. Targeted gene silencing of MyD88 (shRNA) and mTOR (RNAi) inhibition resulted in TLR4-mediated p70s6K activation and enhanced TNFα release, whereas IL-10 release was inhibited in both silenced and non-silenced HIV+ macrophages. Furthermore, mTOR inhibition augmented lipid A-induced TNFα release through enhanced and prolonged phosphorylation of ERK1/2 and JNK1/2 MAP kinases, which was associated with time-dependent MKP-1 destabilization. Taken together, impaired TLR4-mediated TNFα release in HIV+ macrophages is attributable in part to mTOR activation by constitutive PI3K expression in a MyD88-dependent signaling pathway. These changes result in MKP-1 stabilization, which shortens and blunts MAP kinase activation. mTOR inhibition may serve as a potential therapeutic target to upregulate macrophage innate immune host defense responsiveness in HIV+ persons. PMID:22025552

Hepatocyte MyD88 affects bile acids, gut microbiota and metabolome contributing to regulate glucose and lipid metabolism

DEFF Research Database (Denmark)

Duparc, Thibaut; Plovier, Hubert; Marrachelli, Vannina G

2017-01-01

performed microarrays and quantitative PCRs in the liver. In addition, we investigated the gut microbiota composition, bile acid profile and both liver and plasma metabolome. We analysed the expression pattern of genes in the liver of obese humans developing non-alcoholic steatohepatitis (NASH). RESULTS...... proliferator activator receptor-α, farnesoid X receptor (FXR), liver X receptors and STAT3) and bile acid profiles involved in glucose, lipid metabolism and inflammation. In addition to these alterations, the genetic deletion of MyD88 in hepatocytes changes the gut microbiota composition and their metabolomes...

The herpes simplex virus 1-encoded envelope glycoprotein B activates NF-κB through the Toll-like receptor 2 and MyD88/TRAF6-dependent signaling pathway.

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Mingsheng Cai

Full Text Available The innate immune response plays a critical role in the host defense against invading pathogens, and TLR2, a member of the Toll-like receptor (TLR family, has been implicated in the immune response and initiation of inflammatory cytokine secretion against several human viruses. Previous studies have demonstrated that infectious and ultraviolet-inactivated herpes simplex virus 1 (HSV-1 virions lead to the activation of nuclear factor kappa B (NF-κB and secretion of proinflammatory cytokines via TLR2. However, except for the envelope glycoprotein gH and gL, whether there are other determinants of HSV-1 responsible for TLR2 mediated biological effects is not known yet. Here, we demonstrated that the HSV-1-encoded envelope glycoprotein gB displays as molecular target recognized by TLR2. gB coimmunoprecipitated with TLR2, TLR1 and TLR6 in transfected and infected human embryonic kidney (HEK 293T cells. Treatment of TLR2-transfected HEK293T (HEK293T-TLR2 cells with purified gB results in the activation of NF-κB reporter, and this activation requires the recruitment of the adaptor molecules myeloid differentiation primary-response protein 88 (MyD88 and tumor necrosis factor receptor-associated factor 6 (TRAF6 but not CD14. Furthermore, activation of NF-κB was abrogated by anti-gB and anti-TLR2 blocking antibodies. In addition, the expression of interleukin-8 induced by gB was abrogated by the treatment of the human monocytic cell line THP-1 with anti-TLR2 blocking antibody or by the incubation of gB with anti-gB antibody. Taken together, these results indicate the importance and potency of HSV-1 gB as one of pathogen-associated molecular patterns (PAMPs molecule recognized by TLR2 with immediate kinetics.

Expression of myeloid differentiation factor 88 in neurons is not requisite for the induction of sickness behavior by interleukin-1β

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Braun Theodore P

2012-10-01

. Conclusion Sickness behavior is mediated by MyD88 and is dependent on the activity of cytokines within the brain. Our results demonstrate that MyD88 is not required in neurons or astrocytes to induce this behavioral response to IL-1β or LPS. This suggests that a non-Nestin expressing cell population responds to IL-1β in the CNS and transduces the signal to neurons controlling feeding and activity.

Primary Type3 (Non-ABC, Non-GCB Subtype of Extranodal Diffuse Large B-Cell Lymphoma of the Thyroid Bearing No MYD88 Mutation by Padlock Probe Hybridization

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Yukiko Nishi

2017-06-01

Full Text Available Primary extranodal malignant lymphoma of the thyroid is a rare entity composed of mostly neoplastic transformation of germinal center-like B cells (GCB or memory B cells. Other B-cell-type malignancies arising primarily in the thyroid have rarely been described. Immunohistochemical examination of autopsied primary malignant lymphoma of the thyroid in an 83-year-old Japanese female revealed the presence of a non-GCB subtype of diffuse large B-cell lymphoma (DLBCL without the typical codon 206 or 265 missense mutation of MYD88. The lack of the highly oncogenic MYD88 gene mutation, frequently observed in DLBCL of the activated B-cell (ABC subtype, and the detection of an extremely aggressive yet local clinical phenotype demonstrated that the present case was an exceptional entity of the type3 (non-GCB and non-ABC subtype.

Propofol Attenuates Airway Inflammation in a Mast Cell-Dependent Mouse Model of Allergic Asthma by Inhibiting the Toll-like Receptor 4/Reactive Oxygen Species/Nuclear Factor κB Signaling Pathway.

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Li, Hong-Yi; Meng, Jing-Xia; Liu, Zhen; Liu, Xiao-Wen; Huang, Yu-Guang; Zhao, Jing

2018-06-01

Propofol, an intravenous anesthetic agent widely used in clinical practice, is the preferred anesthetic for asthmatic patients. This study was designed to determine the protective effect and underlying mechanisms of propofol on airway inflammation in a mast cell-dependent mouse model of allergic asthma. Mice were sensitized by ovalbumin (OVA) without alum and challenged with OVA three times. Propofol was given intraperitoneally 0.5 h prior to OVA challenge. The inflammatory cell count and production of cytokines in the bronchoalveolar lavage fluid (BALF) were detected. The changes of lung histology and key molecules of the toll-like receptor 4 (TLR4)/reactive oxygen species (ROS)/NF-κB signaling pathway were also measured. The results showed that propofol significantly decreased the number of eosinophils and the levels of IL-4, IL-5, IL-6, IL-13, and TNF-α in BALF. Furthermore, propofol significantly attenuated airway inflammation, as characterized by fewer infiltrating inflammatory cells and decreased mucus production and goblet cell hyperplasia. Meanwhile, the expression of TLR4, and its downstream signaling adaptor molecules--myeloid differentiation factor 88 (MyD88) and NF-κB, were inhibited by propofol. The hydrogen peroxide and methane dicarboxylic aldehyde levels were decreased by propofol, and the superoxide dismutase activity was increased in propofol treatment group. These findings indicate that propofol may attenuate airway inflammation by inhibiting the TLR4/MyD88/ROS/NF-κB signaling pathway in a mast cell-dependent mouse model of allergic asthma.

Measurement of TLR-induced macrophage spreading by automated image analysis: differential role of Myd88 and MAPK in early and late responses

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Jens eWenzel

2011-10-01

Full Text Available Sensing of infectious danger by Toll-like receptors (TLR on macrophages causes not only a reprogramming of the transcriptome but also changes in the cytoskeleton important for cell spreading and motility. Since manual determination of cell contact areas from fluorescence microscopy pictures is very time consuming and prone to bias, we have developed and tested algorithms for automated measurement of macrophage spreading. The two-step method combines identification of cells by nuclear staining with DAPI and cell surface staining of the integrin CD11b. Automated image analysis correlated very well with manual annotation in resting macrophages and early after stimulation, whereas at later time points the automated cell segmentation algorithm and manual annotation showed slightly larger variation. The method was applied to investigate the impact of genetic or pharmacological inhibition of known TLR signaling components. Deificiency in the adapter protein Myd88 strongly reduced spreading activity at the late time points, but had no impact early after LPS stimulation. A similar effect was observed upon pharmacological inhibition of MEK1, the kinase activating the MAPK ERK1/2, indicating that ERK1/2 mediates Myd88-dependent macrophages spreading. In contrast, macrophages lacking the MAPK p38 were impaired in the initial spreading response but responded normally 8 – 24 h after stimulation. The dichotomy of p38 and ERK1/2 MAPK effects on early and late macrophage spreading raises the question which of the respective substrate proteins mediate(s cytoskeletal remodeling and spreading. The automated measurement of cell spreading described here increases the objectivity and greatly reduces the time required for such investigations and is therefore expected to facilitate larger through-put analysis of macrophage spreading, e.g. in siRNA knockdown screens.

Natural loss-of-function mutation of myeloid differentiation protein 88 disrupts its ability to form Myddosomes

NARCIS (Netherlands)

Nagpal, K.; Plantinga, T.S.; Sirois, C.M.; Monks, B.G.; Latz, E.; Netea, M.G.; Golenbock, D.T.

2011-01-01

Myeloid differentiation protein 88 (MyD88) is a key signaling adapter in Toll-like receptor (TLR) signaling. MyD88 is also one of the most polymorphic adapter proteins. We screened the reported nonsynonymous coding mutations in MyD88 to identify variants with altered function. In reporter assays, a

Dexmedetomidine Inhibits Inflammatory Reaction in Lung Tissues of Septic Rats by Suppressing TLR4/NF-κB Pathway

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Yuqing Wu

2013-01-01

and 20 μg/kg significantly decreased mortality and pulmonary inflammation of septic rats, as well as suppressed CLP-induced elevation of TNF-α and IL-6 and inhibited TLR4/MyD88 expression and NF-κB activation. These results suggest that dexmedetomidine may decrease mortality and inhibit inflammatory reaction in lung tissues of septic rats by suppressing TLR4/MyD88/NF-κB pathway.

Protective Effect of 2-Dodecyl-6-Methoxycyclohexa-2, 5-Diene-1, 4-Dione, Isolated from Averrhoa Carambola L., Against Palmitic Acid-Induced Inflammation and Apoptosis in Min6 Cells by Inhibiting the TLR4-MyD88-NF-κB Signaling Pathway

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Qiuqiao Xie

2016-09-01

Full Text Available Background/Aims: Studies have demonstrated that 2-dodecyl-6-methoxycyclohexa-2, 5-diene-1, 4-dione (DMDD, isolated from the roots of Averrhoa carambola L., has significant therapeutic potential for the treatment of diabetes. However, the protective effect of DMDD against pancreatic beta cell dysfunction has never been reported. We investigated whether DMDD protected against palmitic acid-induced dysfunction in pancreatic β-cell line Min6 cells by attenuating the inflammatory response and apoptosis and to shed light on its possible mechanism. Methods: Cell viability was assessed by CCK-8. Glucose-stimulated insulin secretion levels and inflammatory cytokines levels were examined by ELISA. Apoptosis was assessed by Annexin V-FITC/PI Flow cytometry assay, Hoechst 33342/PI double-staining assay, and Transmission electron microscopy assay. Relative quantitative real-time PCR and western blot were used to determine the expressions of genes and proteins. Results: Cell viability and glucose-stimulated insulin secretion levels were increased in DMDD-pretreated Min6 cells. DMDD inhibited inflammatory cytokines IL-6, TNF-α and MCP-1 generations in palmitic acid (PA-induced Min6 cells. Moreover, DMDD protected against PA-induced Min6 cells apoptosis and the expression of Cleaved-Caspase-3, -8 and -9 were down-regulated and the Bcl-2/Bax ratio was increased in DMDD-pretreated Min6 cells. In addition, the expression of TLR4, MyD88 and NF-κB were down-regulated in DMDD-pretreated Min6 cells and TAK-242-pretreated group cells. Conclusions: DMDD protected Min6 cells against PA-induced dysfunction by attenuating the inflammatory response and apoptosis, and its mechanism of this protection was associated with inhibiting the TLR4-MyD88-NF-κB signaling pathway.

Protective Effect of 2-Dodecyl-6-Methoxycyclohexa-2, 5-Diene-1, 4-Dione, Isolated from Averrhoa Carambola L., Against Palmitic Acid-Induced Inflammation and Apoptosis in Min6 Cells by Inhibiting the TLR4-MyD88-NF-κB Signaling Pathway.

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Xie, Qiuqiao; Zhang, Shijun; Chen, Chunxia; Li, Juman; Wei, Xiaojie; Xu, Xiaohui; Xuan, Feifei; Chen, Ning; Pham, Thithaihoa; Qin, Ni; He, Junhui; Ye, Fangxing; Huang, Wansu; Huang, Renbin; Wen, Qingwei

2016-01-01

Studies have demonstrated that 2-dodecyl-6-methoxycyclohexa-2, 5-diene-1, 4-dione (DMDD), isolated from the roots of Averrhoa carambola L., has significant therapeutic potential for the treatment of diabetes. However, the protective effect of DMDD against pancreatic beta cell dysfunction has never been reported. We investigated whether DMDD protected against palmitic acid-induced dysfunction in pancreatic β-cell line Min6 cells by attenuating the inflammatory response and apoptosis and to shed light on its possible mechanism. Cell viability was assessed by CCK-8. Glucose-stimulated insulin secretion levels and inflammatory cytokines levels were examined by ELISA. Apoptosis was assessed by Annexin V-FITC/PI Flow cytometry assay, Hoechst 33342/PI double-staining assay, and Transmission electron microscopy assay. Relative quantitative real-time PCR and western blot were used to determine the expressions of genes and proteins. Cell viability and glucose-stimulated insulin secretion levels were increased in DMDD-pretreated Min6 cells. DMDD inhibited inflammatory cytokines IL-6, TNF-α and MCP-1 generations in palmitic acid (PA)-induced Min6 cells. Moreover, DMDD protected against PA-induced Min6 cells apoptosis and the expression of Cleaved-Caspase-3, -8 and -9 were down-regulated and the Bcl-2/Bax ratio was increased in DMDD-pretreated Min6 cells. In addition, the expression of TLR4, MyD88 and NF-κB were down-regulated in DMDD-pretreated Min6 cells and TAK-242-pretreated group cells. DMDD protected Min6 cells against PA-induced dysfunction by attenuating the inflammatory response and apoptosis, and its mechanism of this protection was associated with inhibiting the TLR4-MyD88-NF-κB signaling pathway. © 2016 The Author(s) Published by S. Karger AG, Basel.

Lipopolysaccharide stimulates p62-dependent autophagy-like aggregate clearance in hepatocytes.

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Chen, Christine; Deng, Meihong; Sun, Qian; Loughran, Patricia; Billiar, Timothy R; Scott, Melanie J

2014-01-01

Impairment of autophagy has been associated with liver injury. TLR4-stimulation by LPS upregulates autophagy in hepatocytes, although the signaling pathways involved remain elusive. The objective of this study was to determine the signaling pathway leading to LPS-stimulated autophagy in hepatocytes. Cell lysates from livers of wild type (WT; C57BL/6) mice given LPS (5 mg/kg-IP) and hepatocytes from WT, TLR4ko, and MyD88ko mice treated with LPS (100 ng/mL) up to 24 h were collected. LC3II, p62/SQSTM1, Nrf2, and beclin1 levels were determined by immunoblot, immunofluorescence, and qPCR. Autophagy-like activation was measured by GFP-LC3-puncta formation and LC3II-expression. Beclin1, Nrf2, p62, MyD88, and TIRAP were knocked-down using siRNA. LC3II-expression increased in both liver and hepatocytes after LPS and was dependent on TLR4. Beclin1 expression did not increase after LPS in hepatocytes and beclin1-knockdown did not affect LC3II levels. In hepatocytes given LPS, expression of p62 increased and p62 colocalized with LC3. p62-knockdown prevented LC3II puncta formation. LPS-induced LC3II/p62-puncta also required MyD88/TIRAP signaling and localization of both Nrf2 and NF κ B transcription factors to the nucleus to upregulate p62-expression. Therefore, TLR4-activation by LPS in hepatocytes induces a p62-mediated, not beclin1-mediated, autophagy-like clearance pathway that is hepatoprotective by clearing aggregate-prone or misfolded proteins from the cytosol and preserving energy homeostasis under stress.

Downstream Toll-like receptor signaling mediates adaptor-specific cytokine expression following focal cerebral ischemia

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Bolanle Famakin

2012-07-01

Full Text Available Abstract Background Deletion of some Toll-like receptors (TLRs affords protection against cerebral ischemia, but disruption of their known major downstream adaptors does not. To determine whether compensation in the production of downstream effectors by one pathway when the other is disrupted can explain these findings, we examined cytokine/chemokine expression and inflammatory infiltrates in wild-type (WT, MyD88−/− and TRIF-mutant mice following permanent middle cerebral artery occlusion (pMCAO. Methods Cytokine/chemokine expression was measured with a 25-plex bead array in the serum and brains of all three groups of mice at baseline (no surgery/naïve and at 3 hours and 24 hours following pMCAO. Brain inflammatory and neutrophil infiltrates were examined 24 hours following pMCAO. Results IL-6, keratinocyte chemoattractant (KC, granulocyte colony-stimulating factor (G-CSF and IL-10 were significantly decreased in MyD88−/− mice compared to WT mice following pMCAO. Significantly, decreased levels of the neutrophil chemoattractants KC and G-CSF corresponded with a trend toward fewer neutrophils in the brains of MyD88−/− mice. IP-10 was significantly decreased when either pathway was disrupted. MIP-1α was significantly decreased in TRIF-mutant mice, consistent with TRIF-dependent production. MyD88−/− mice showed elevations of a number of Th2 cytokines, such as IL-13, at baseline, which became significantly decreased following pMCAO. Conclusions Both MyD88 and TRIF mediate pathway-specific cytokine production following focal cerebral ischemia. Our results also suggest a compensatory Th2-type skew at baseline in MyD88−/− mice and a paradoxical switch to a Th1 phenotype following focal cerebral ischemia. The MyD88 pathway directs the expression of neutrophil chemoattractants following cerebral ischemia.

The IL-33-ST2-MyD88 axis promotes regulatory T cell proliferation in the murine liver.

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Xu, Lei; Li, Wei; Wang, Xiaofan; Zhang, Lina; Qi, Qianqian; Dong, Liyang; Wei, Chuan; Pu, Yanan; Li, Yalin; Zhu, Jifeng; Zhou, Sha; Liu, Feng; Chen, Xiaojun; Su, Chuan

2018-05-05

Hepatic Foxp3 + regulatory T (Treg) cells are crucial for maintaining local immune homeostasis in the liver. However, the environmental cues required for hepatic Treg cell homeostasis are unclear. In this study, we showed that the IL-33 receptor ST2 was preferentially expressed on Treg cells in the mouse liver, but it was more lowly expressed in the spleen, mesenteric lymph nodes, and blood. More importantly, we found that IL-33 promoted the proliferation of hepatic Treg cells through myeloid differentiation factor MyD88 signaling concomitant with increased CDK4 and cyclin D1 expression. These results suggested that IL-33 is a potential tissue-specific factor controlling Treg cell homeostasis via increased Treg proliferation in the liver. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Porphyromonas gingivalis Promotes Unrestrained Type I Interferon Production by Dysregulating TAM Signaling via MYD88 Degradation.

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Mizraji, Gabriel; Nassar, Maria; Segev, Hadas; Sharawi, Hafiz; Eli-Berchoer, Luba; Capucha, Tal; Nir, Tsipora; Tabib, Yaara; Maimon, Avraham; Dishon, Shira; Shapira, Lior; Nussbaum, Gabriel; Wilensky, Asaf; Hovav, Avi-Hai

2017-01-10

Whereas type I interferons (IFNs-I) were proposed to be elevated in human periodontitis, their role in the disease remains elusive. Using a bacterial-induced model of murine periodontitis, we revealed a prolonged elevation in IFN-I expression. This was due to the downregulation of TAM signaling, a major negative regulator of IFN-I. Further examination revealed that the expression of certain TAM components was reduced as a result of prolonged degradation of MYD88 by the infection. As a result of such prolonged IFN-I production, innate immunological functions of the gingiva were disrupted, and CD4 + T cells were constitutively primed by dendritic cells, leading to elevated RANKL expression and, subsequently, alveolar bone loss (ABL). Blocking IFN-I signaling restored proper immunological function and prevented ABL. Importantly, a loss of negative regulation on IFN-I expression by TAM signaling was also evident in periodontitis patients. These findings thus suggest a role for IFN-I in the pathogenesis of periodontitis. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Immune receptors involved in Streptococcus suis recognition by dendritic cells.

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Marie-Pier Lecours

Full Text Available Streptococcus suis is an important swine pathogen and an emerging zoonotic agent of septicemia and meningitis. Knowledge on host immune responses towards S. suis, and strategies used by this pathogen for subversion of these responses is scarce. The objective of this study was to identify the immune receptors involved in S. suis recognition by dendritic cells (DCs. Production of cytokines and expression of co-stimulatory molecules by DCs were shown to strongly rely on MyD88-dependent signaling pathways, suggesting that DCs recognize S. suis and become activated mostly through Toll-like receptor (TLR signaling. Supporting this fact, TLR2(-/- DCs were severely impaired in the release of several cytokines and the surface expression of CD86 and MHC-II. The release of IL-12p70 and CXC10, and the expression of CD40 were found to depend on signaling by both TLR2 and TLR9. The release of IL-23 and CXCL1 were partially dependent on NOD2. Finally, despite the fact that MyD88 signaling was crucial for DC activation and maturation, MyD88-dependent pathways were not implicated in S. suis internalization by DCs. This first study on receptors involved in DC activation by S. suis suggests a major involvement of MyD88 signaling pathways, mainly (but not exclusively through TLR2. A multimodal recognition involving a combination of different receptors seems essential for DC effective response to S. suis.

TLR-dependent control of Francisella tularensis infection and host inflammatory responses.

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Allison L Abplanalp

2009-11-01

Full Text Available Francisella tularensis is the causative agent of tularemia and is classified as a Category A select agent. Recent studies have implicated TLR2 as a critical element in the host protective response to F. tularensis infection, but questions remain about whether TLR2 signaling dominates the response in all circumstances and with all species of Francisella and whether F. tularensis PAMPs are predominantly recognized by TLR2/TLR1 or TLR2/TLR6. To address these questions, we have explored the role of Toll-like receptors (TLRs in the host response to infections with F. tularensis Live Vaccine Strain (LVS and F. tularensis subspecies (subsp. novicida in vivo.C57BL/6 (B6 control mice and TLR- or MyD88-deficient mice were infected intranasally (i.n. or intradermally (i.d. with F. tularensis LVS or with F. tularensis subsp. novicida. B6 mice survived >21 days following infection with LVS by both routes and survival of TLR1(-/-, TLR4(-/-, and TLR6(-/- mice infected i.n. with LVS was equivalent to controls. Survival of TLR2(-/- and MyD88(-/- mice, however, was significantly reduced compared to B6 mice, regardless of the route of infection or the subspecies of F. tularensis. TLR2(-/- and MyD88(-/- mice also showed increased bacterial burdens in lungs, liver, and spleen compared to controls following i.n. infection. Primary macrophages from MyD88(-/- and TLR2(-/- mice were significantly impaired in the ability to secrete TNF and other pro-inflammatory cytokines upon ex vivo infection with LVS. TNF expression was also impaired in vivo as demonstrated by analysis of bronchoalveolar lavage fluid and by in situ immunofluorescent staining.We conclude from these studies that TLR2 and MyD88, but not TLR4, play critical roles in the innate immune response to F. tularensis infection regardless of the route of infection or the subspecies. Moreover, signaling through TLR2 does not depend exclusively on TLR1 or TLR6 during F. tularensis LVS infection.

Gene expression disorders of innate antibacterial signaling pathway in pancreatic cancer patients: implications for leukocyte dysfunction and tumor progression

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Dąbrowska, Aleksandra; Lech, Gustaw; Słodkowski, Maciej; Słotwińska, Sylwia M.

2014-01-01

The study was carried out to investigate changes in gene expression of innate antibacterial signaling pathways in patients with pancreatic cancer. Expression of the following genes was measured in peripheral blood leukocytes of 55 patients with pancreatic adenocarcinoma using real-time polymerase chain reaction (RT-PCR): TLR4, NOD1, MyD88, TRAF6 and HMGB1. The levels of expression of TLR4, NOD1 and TRAF6 genes were significantly elevated (p = 0.007; p = 0.001 and p = 0.01, respectively), while MyD88 expression was markedly reduced (p = 0.0002), as compared to controls. Expression of TLR4 and NOD1 exceeded the normal level more than 3.5-fold and there was a significant correlation found between the expression of these genes (r = 0.558, p < 0.001). TLR4, NOD1 and MyD88 genes were expressed at a similar level both before and after surgery. No significant changes in the expression of HMGB1 gene were observed. The results of the study clearly indicate abnormal expression of genes belonging to innate antibacterial signaling pathways in peripheral blood leukocytes of patients with pancreatic cancer, which may lead to leukocyte dysfunction. Overexpression of TLR4, NOD1 and TRAF6 genes, and decreased MyD88 gene expression may contribute to chronic inflammation and tumor progression by up-regulation of the innate antibacterial response. The parameters tested are useful for monitoring innate immunity gene disorders and pancreatic cancer progression. PMID:26155170

Effects of Gui Zhi Ma Huang Ge Ban Tang on the TLR7 Pathway in Influenza Virus Infected Mouse Lungs in a Cold Environment.

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Qin, Hong-Qiong; Shi, Shan-Shan; Fu, Ying-Jie; Yan, Yu-Qi; Wu, Sha; Tang, Xiao-Long; Chen, Xiao-Yin; Hou, Guang-Hui; Jiang, Zhen-You

2018-01-01

We wished to investigate the effects of the traditional Chinese medicine Gui Zhi Ma Huang Ge Ban Tang on controlling influenza A virus (IAV) infection and improving inflammation in mouse lungs. Mice were maintained in normal and cold environments and infected with IAV by intranasal application, respectively. Real-time quantitative polymerase chain reaction was used to measure mRNA expression of TLR7, myeloid differentiation primary response 88 (MyD88), and nuclear factor-kappa B (NF- κ B)p65 in the TLR7 signaling pathway and virus replication in lungs. Western blotting was used to measure expression levels of TLR7, MyD88, and NF- κ B p65 proteins. Flow cytometry was used to detect the proportion of T-helper (Th)1/Th2 and Th17/T-regulatory (Treg) cells. Application of Gui Zhi Ma Huang Ge Ban Tang in influenza-infected mice in a cold environment showed (i) downregulation of TLR7, MyD88, and NF- κ Bp65; (ii) inhibition of transcriptional activities of promoters coding for TLR7, MyD88, and NF- κ Bp65; (iii) reduction in the proportion of Th1/Th2 and Th17/Treg cells. Gui Zhi Ma Huang Ge Ban Tang had a good therapeutic effect on mice infected with IAV, especially in the cold environment. It could reduce lung inflammation in mice significantly and elicit an anti-influenza effect by downregulating expression of the key factors in TLR7 signaling pathway.

H₂S protecting against lung injury following limb ischemia-reperfusion by alleviating inflammation and water transport abnormality in rats.

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Qi, Qi Ying Chun; Chen, Wen; Li, Xiao Ling; Wang, Yu Wei; Xie, Xiao Hua

2014-06-01

To investigate the effect of H₂S on lower limb ischemia-reperfusion (LIR) induced lung injury and explore the underlying mechanism. Wistar rats were randomly divided into control group, IR group, IR+ Sodium Hydrosulphide (NaHS) group and IR+ DL-propargylglycine (PPG) group. IR group as lung injury model induced by LIR were given 4 h reperfusion following 4 h ischemia of bilateral hindlimbs with rubber bands. NaHS (0.78 mg/kg) as exogenous H₂S donor and PPG (60 mg/kg) which can suppress endogenous H₂S production were administrated before LIR, respectively. The lungs were removed for histologic analysis, the determination of wet-to-dry weight ratios and the measurement of mRNA and protein levels of aquaporin-1 (AQP₁), aquaporin-5 (AQP₅) as indexes of water transport abnormality, and mRNA and protein levels of Toll-like receptor 4 (TLR₄), myeloid differentiation primary-response gene 88 (MyD88) and p-NF-κB as indexes of inflammation. LIR induced lung injury was accompanied with upregulation of TLR₄-Myd88-NF-κB pathway and downregulation of AQP1/AQP₅. NaHS pre-treatment reduced lung injury with increasing AQP₁/AQP₅ expression and inhibition of TLR₄-Myd88-NF-κB pathway, but PPG adjusted AQP₁/AQP₅ and TLR4 pathway to the opposite side and exacerbated lung injury. Endogenous H₂S, TLR₄-Myd88-NF-κB pathway and AQP₁/AQP₅ were involved in LIR induced lung injury. Increased H₂S would alleviate lung injury and the effect is at least partially depend on the adjustment of TLR₄-Myd88-NF-κB pathway and AQP₁/AQP₅ expression to reduce inflammatory reaction and lessen pulmonary edema. Copyright © 2014 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

A toll-like receptor 2 pathway regulates the Ppargc1a/b metabolic co-activators in mice with Staphylococcal aureus sepsis.

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Timothy E Sweeney

Full Text Available Activation of the host antibacterial defenses by the toll-like receptors (TLR also selectively activates energy-sensing and metabolic pathways, but the mechanisms are poorly understood. This includes the metabolic and mitochondrial biogenesis master co-activators, Ppargc1a (PGC-1α and Ppargc1b (PGC-1β in Staphylococcus aureus (S. aureus sepsis. The expression of these genes in the liver is markedly attenuated inTLR2(-/- mice and markedly accentuated in TLR4(-/- mice compared with wild type (WT mice. We sought to explain this difference by using specific TLR-pathway knockout mice to test the hypothesis that these co-activator genes are directly regulated through TLR2 signaling. By comparing their responses to S. aureus with WT mice, we found that MyD88-deficient and MAL-deficient mice expressed hepatic Ppargc1a and Ppargc1b normally, but that neither gene was activated in TRAM-deficient mice. Ppargc1a/b activation did not require NF-kβ, but did require an interferon response factor (IRF, because neither gene was activated in IRF-3/7 double-knockout mice in sepsis, but both were activated normally in Unc93b1-deficient (3d mice. Nuclear IRF-7 levels in TLR2(-/- and TLR4(-/- mice decreased and increased respectively post-inoculation and IRF-7 DNA-binding at the Ppargc1a promoter was demonstrated by chromatin immunoprecipitation. Also, a TLR2-TLR4-TRAM native hepatic protein complex was detected by immunoprecipitation within 6 h of S. aureus inoculation that could support MyD88-independent signaling to Ppargc1a/b. Overall, these findings disclose a novel MyD88-independent pathway in S. aureus sepsis that links TLR2 and TLR4 signaling in innate immunity to Ppargc1a/b gene regulation in a critical metabolic organ, the liver, by means of TRAM, TRIF, and IRF-7.

Complement C1q regulates LPS-induced cytokine production in bone marrow-derived dendritic cells.

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Yamada, Masahide; Oritani, Kenji; Kaisho, Tsuneyasu; Ishikawa, Jun; Yoshida, Hitoshi; Takahashi, Isao; Kawamoto, Shinichirou; Ishida, Naoko; Ujiie, Hidetoshi; Masaie, Hiroaki; Botto, Marina; Tomiyama, Yoshiaki; Matsuzawa, Yuji

2004-01-01

We show here that C1q suppresses IL-12p40 production in LPS-stimulated murine bone marrow-derived dendritic cells (BMDC). Serum IL-12p40 concentration of C1q-deficient mice was higher than that of wild-type mice after intraperitoneal LPS-injection. Because neither globular head of C1q (gC1q) nor collagen-like region of C1q (cC1q) failed to suppress LPS-induced IL-12p40 production, both gC1q and cC1q, and/or some specialized conformation of native C1q may be required for the inhibition. While C1q did not affect mRNA expression of Toll-like receptor 4 (TLR4), MD-2, and myeloid differentiation factor 88 (MyD88), BMDC treated with C1q showed the reduced activity of NF-kappaB and the delayed phosphorylation of p38, c-Jun N-terminal kinase, and extracellular signal-regulated kinase after LPS-stimulation. CpG oligodeoxynucleotide-induced IL-12p40 and TNF-alpha production, another MyD88-dependent TLR-mediated signal, was also suppressed by C1q treatment. Therefore, C1q is likely to suppress MyD88-dependent pathway in TLR-mediated signals. In contrast, C1q failed to suppress colony formation of B cells responding to LPS or LPS-induced CD40 and CD86 expression on BMDC in MyD88-deficient mice, indicating that inhibitory effects of C1q on MyD88-independent pathways may be limited. Taken together, C1q may regulate innate and adaptive immune systems via modification of signals mediated by interactions between invading pathogens and TLR.

Protective Effect of Phillyrin on Lethal LPS-Induced Neutrophil Inflammation in Zebrafish

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Liling Yang

2017-10-01

Full Text Available Background/Aims: Forsythia suspensa Vahl. (Oleaceae fruits are widely used in traditional Chinese medicine to treat pneumonia, typhoid, dysentery, ulcers and oedema. Antibacterial and anti-inflammatory activities have been reported for phillyrin (PHN, the main ingredient in Forsythia suspensa Vahl fruits, in vitro. However, the underlying mechanisms in vivo remain poorly defined. In this study, we discovered that PHN exerted potent anti-inflammatory effects in lethal LPS-induced neutrophil inflammation by suppressing the MyD88-dependent signalling pathway in zebrafish. Methods: LPS-yolk microinjection was used to induce a lethal LPS-infected zebrafish model. The effect of PHN on the survival of zebrafish challenged with lethal LPS was evaluated using survival analysis. The effect of PHN on neutrophil inflammation grading in vivo was assessed by tracking neutrophils with a transgenic line. The effects of PHN on neutrophil production and migration were analysed by SB+ cell counts during consecutive hours after modelling. Additionally, key cytokines and members of the MyD88 signalling pathway that are involved in inflammatory response were detected using quantitative RT-PCR. To assess gene expression changes during consecutive hours after modelling, the IL-1β, IL-6, TNF-α, MyD88, TRIF, ERK1/2, JNK, IκBa and NF-κB expression levels were measured. Results: PHN could protect zebrafish against a lethal LPS challenge in a dose-dependent manner, as indicated by decreased neutrophil infltration, reduced tissue necrosis and increased survival rates. Up-regulated IL-1β, IL-6 and TNF-α expression also showed the same tendencies of depression by PHN. Critically, PHN significantly inhibited the LPS-induced activation of MyD88, IκBa, and NF-κB but did not affect the expression of ERK1/2 MAPKs or JNK MAPKs in LPS-stimulated zebrafish. Additionally, PHN regulated the MyD88/IκBα/NF-κB signalling pathway by controlling IκBα, IL-1β, IL-6, and TNF

Anti-inflammatory activity of Odina wodier Roxb, an Indian folk remedy, through inhibition of toll-like receptor 4 signaling pathway.

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Ojha, Durbadal; Mukherjee, Hemanta; Mondal, Supriya; Jena, Aditya; Dwivedi, Ved Prakash; Mondal, Keshab C; Malhotra, Bharti; Samanta, Amalesh; Chattopadhyay, Debprasad

2014-01-01

Inflammation is part of self-limiting non-specific immune response, which occurs during bodily injury. In some disorders the inflammatory process becomes continuous, leading to the development of chronic inflammatory diseases including cardiovascular diseases, diabetes, cancer etc. Several Indian tribes used the bark of Odina wodier (OWB) for treating inflammatory disorders. Thus, we have evaluated the immunotherapeutic potential of OWB methanol extract and its major constituent chlorogenic acid (CA), using three popular in vivo antiinflammatory models: Carrageenan- and Dextran-induced paw edema, Cotton pellet granuloma, and Acetic acid-induced vascular permeability. To elucidate the possible anti-inflammatory mechanism of action we determine the level of major inflammatory mediators (NO, iNOS, COX-2-dependent prostaglandin E2 or PGE2), and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-12). Further, we determine the toll-like receptor 4 (TLR4), Myeloid differentiation primary response gene 88 (MyD88), c-Jun N-terminal kinases (JNK), nuclear factor kappa-B cells (NF-κB), and NF-kB inhibitor alpha (IK-Bα) by protein and mRNA expression, and Western blot analysis in drug treated LPS-induced murine macrophage model. Moreover, we determined the acute and sub-acute toxicity of OWB extract in BALB/c mice. Our study demonstrated a significant anti-inflammatory activity of OWB extract and CA along with the inhibition of TNF-α, IL-1β, IL-6 and IL-12 expressions. Further, the expression of TLR4, NF-κBp65, MyD88, iNOS and COX-2 molecules were reduced in drug-treated groups, but not in the LPS-stimulated untreated or control groups, Thus, our results collectively indicated that the OWB extract and CA can efficiently inhibit inflammation through the down regulation of TLR4/MyD88/NF-kB signaling pathway.

Anti-inflammatory activity of Odina wodier Roxb, an Indian folk remedy, through inhibition of toll-like receptor 4 signaling pathway.

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Durbadal Ojha

Full Text Available Inflammation is part of self-limiting non-specific immune response, which occurs during bodily injury. In some disorders the inflammatory process becomes continuous, leading to the development of chronic inflammatory diseases including cardiovascular diseases, diabetes, cancer etc. Several Indian tribes used the bark of Odina wodier (OWB for treating inflammatory disorders. Thus, we have evaluated the immunotherapeutic potential of OWB methanol extract and its major constituent chlorogenic acid (CA, using three popular in vivo antiinflammatory models: Carrageenan- and Dextran-induced paw edema, Cotton pellet granuloma, and Acetic acid-induced vascular permeability. To elucidate the possible anti-inflammatory mechanism of action we determine the level of major inflammatory mediators (NO, iNOS, COX-2-dependent prostaglandin E2 or PGE2, and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-12. Further, we determine the toll-like receptor 4 (TLR4, Myeloid differentiation primary response gene 88 (MyD88, c-Jun N-terminal kinases (JNK, nuclear factor kappa-B cells (NF-κB, and NF-kB inhibitor alpha (IK-Bα by protein and mRNA expression, and Western blot analysis in drug treated LPS-induced murine macrophage model. Moreover, we determined the acute and sub-acute toxicity of OWB extract in BALB/c mice. Our study demonstrated a significant anti-inflammatory activity of OWB extract and CA along with the inhibition of TNF-α, IL-1β, IL-6 and IL-12 expressions. Further, the expression of TLR4, NF-κBp65, MyD88, iNOS and COX-2 molecules were reduced in drug-treated groups, but not in the LPS-stimulated untreated or control groups, Thus, our results collectively indicated that the OWB extract and CA can efficiently inhibit inflammation through the down regulation of TLR4/MyD88/NF-kB signaling pathway.

STING-Dependent Cytosolic DNA Sensing Promotes Radiation-Induced Type I Interferon-Dependent Antitumor Immunity in Immunogenic Tumors.

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Deng, Liufu; Liang, Hua; Xu, Meng; Yang, Xuanming; Burnette, Byron; Arina, Ainhoa; Li, Xiao-Dong; Mauceri, Helena; Beckett, Michael; Darga, Thomas; Huang, Xiaona; Gajewski, Thomas F; Chen, Zhijian J; Fu, Yang-Xin; Weichselbaum, Ralph R

2014-11-20

Ionizing radiation-mediated tumor regression depends on type I interferon (IFN) and the adaptive immune response, but several pathways control I IFN induction. Here, we demonstrate that adaptor protein STING, but not MyD88, is required for type I IFN-dependent antitumor effects of radiation. In dendritic cells (DCs), STING was required for IFN-? induction in response to irradiated-tumor cells. The cytosolic DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS) mediated sensing of irradiated-tumor cells in DCs. Moreover, STING was essential for radiation-induced adaptive immune responses, which relied on type I IFN signaling on DCs. Exogenous IFN-? treatment rescued the cross-priming by cGAS or STING-deficient DCs. Accordingly, activation of STING by a second messenger cGAMP administration enhanced antitumor immunity induced by radiation. Thus radiation-mediated antitumor immunity in immunogenic tumors requires a functional cytosolic DNA-sensing pathway and suggests that cGAMP treatment might provide a new strategy to improve radiotherapy. Copyright © 2014 Elsevier Inc. All rights reserved.

Matrine pretreatment improves cardiac function in rats with diabetic cardiomyopathy via suppressing ROS/TLR-4 signaling pathway.

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Liu, Zhong-wei; Wang, Jun-kui; Qiu, Chuan; Guan, Gong-chang; Liu, Xin-hong; Li, Shang-jian; Deng, Zheng-rong

2015-03-01

Matrine is an alkaloid from Sophora alopecuroides L, which has shown a variety of pharmacological activities and potential therapeutic value in cardiovascular diseases. In this study we examined the protective effects of matrine against diabetic cardiomyopathy (DCM) in rats. Male SD rats were injected with streptozotocin (STZ) to induce DCM. One group of DCM rats was pretreated with matrine (200 mg·kg(-1)·d(-1), po) for 10 consecutive days before STZ injection. Left ventricular function was evaluated using invasive hemodynamic examination, and myocardiac apoptosis was assessed. Primary rat myocytes were used for in vitro experiments. Intracellular ROS generation, MDA content and GPx activity were determined. Real-time PCR and Western blotting were performed to detect the expression of relevant mRNAs and proteins. DCM rats exhibited abnormally elevated non-fasting blood glucose levels at 4 weeks after STZ injection, and LV function impairment at 16 weeks. The cardiac tissues of DCM rats showed markedly increased apoptosis, excessive ROS production, and activation of TLR-4/MyD-88/caspase-8/caspase-3 signaling. Pretreatment with matrine significantly decreased non-fasting blood glucose levels and improved LV function in DCM rats, which were associated with reducing apoptosis and ROS production, and suppressing TLR-4/MyD-88/caspase-8/caspase-3 signaling in cardiac tissues. Incubation in a high-glucose medium induced oxidative stress and activation of TLR-4/MyD-88 signaling in cultured myocytes in vitro, which were significantly attenuated by pretreatment with N-acetylcysteine. Excessive ROS production in DCM activates the TLR-4/MyD-88 signaling, resulting in cardiomyocyte apoptosis, whereas pretreatment with matrine improves cardiac function via suppressing ROS/TLR-4 signaling pathway.

Toll-like Receptor 4 Signaling Confers Cardiac Protection Against Ischemic Injury via Inducible Nitric Oxide Synthase- and Soluble Guanylate Cyclase-dependent Mechanisms

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Wang, E; Feng, Yan; Zhang, Ming; Zou, Lin; Li, Yan; Buys, Emmanuel S.; Huang, Peigen; Brouckaert, Peter; Chao, Wei

2011-01-01

Background Prior administration of a small dose of lipopolysaccharide confers a cardiac protection against ischemia-reperfusion injury. However, the signaling mechanisms that control the protection are incompletely understood. We tested the hypothesis that TLR4 mediates the ability of lipopolysaccharide to protect against cardiac ischemia-reperfusion injury through distinct intracellular pathways involving myeloid differentiation factor 88 (MyD88), TIR-domain-containing adaptor protein inducing interferon-β–mediated transcription-factor (Trif), inducible nitric-oxide synthase (iNOS), and soluble guanylate cyclase (sGC). Methods Wild-type mice and the genetically modified mice, i.e., TLR4-deficient (TLR4-def), TLR2 knockout (TLR2−/−), MyD88−/−, Trif−/−, iNOS−/−, and sGCα1−/−, were treated with normal saline or 0.1 mg/kg of lipopolysaccharide, intraperitoneally. Twenty-four hours later, isolated hearts were perfused in a Langendorff apparatus and subsequently subjected to 30 min of global ischemia and reperfusion for up to 60 min. Left ventricular function and myocardial infarction sizes were examined. Results Compared to saline-treated mice, lipopolysaccharide-treated mice had markedly improved left ventricular developed pressure and dP/dtmax (P < 0.01) and reduced MI sizes (37.2 ± 3.4% vs. 19.8 ± 4.9%, P < 0.01) after ischemia-reperfusion. The cardiac protective effect of lipopolysaccharide was abolished in the TLR4-def and MyD88−/− mice, but remained intact in TLR2−/− or Trif−/− mice. iNOS−/− mice or wild-type mice treated with the iNOS inhibitor 1400W failed to respond to the TLR4-induced nitric oxide production and were not protected by the lipopolysaccharide preconditioning. While sGC 1−/− mice had robust nitric oxide production in response to lipopolysaccharide, they were not protected by the TLR4-elicited cardiac protection. Conclusions TLR4 activation confers a potent cardiac protection against ischemia

Implication of TLR- but not of NOD2-signaling pathways in dendritic cell activation by group B Streptococcus serotypes III and V.

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Paul Lemire

Full Text Available Group B Streptococcus (GBS is an important agent of life-threatening invasive infection. It has been previously shown that encapsulated type III GBS is easily internalized by dendritic cells (DCs, and that this internalization had an impact on cytokine production. The receptors underlying these processes are poorly characterized. Knowledge on the mechanisms used by type V GBS to activate DCs is minimal. In this work, we investigated the role of Toll-like receptor (TLR/MyD88 signaling pathway, the particular involvement of TLR2, and that of the intracellular sensing receptor NOD2 in the activation of DCs by types III and V GBS. The role of capsular polysaccharide (CPS, one of the most important GBS virulence factors in bacterial-DC interactions was evaluated using non-encapsulated mutants. Despite differences in the role of CPS between types III and V GBS in bacterial internalization and intracellular survival, no major differences were observed in their capacity to modulate release of cytokines by DC. For both serotypes, CPS had a minor role in this response. Production of cytokines by DCs was shown to strongly rely on MyD88-dependent signaling pathways, suggesting that DCs recognize GBS and become activated mostly through TLR signaling. Yet, GBS-infected TLR2-/- DCs only showed a partial reduction in the production of IL-6 and CXCL1 compared to control DCs. Surprisingly, CXCL10 release by type III or type V GBS-infected DCs was MyD88-independent. No differences in DC activation were observed between NOD2-/- and control DCs. These results demonstrate the involvement of various receptors and the complexity of the cytokine production pathways activated by GBS upon DC infection.

Vitamin D inhibits lipopolysaccharide-induced inflammatory response potentially through the Toll-like receptor 4 signalling pathway in the intestine and enterocytes of juvenile Jian carp (Cyprinus carpio var. Jian).

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Jiang, Jun; Shi, Dan; Zhou, Xiao-Qiu; Yin, Long; Feng, Lin; Jiang, Wei-Dan; Liu, Yang; Tang, Ling; Wu, Pei; Zhao, Ye

2015-11-28

The present study was conducted to investigate the anti-inflammatory effect of vitamin D both in juvenile Jian carp (Cyprinus carpio var. Jian) in vivo and in enterocytes in vitro. In primary enterocytes, exposure to 10 mg lipopolysaccharide (LPS)/l increased lactate dehydrogenase activity in the culture medium (P<0·05) and resulted in a significant loss of cell viability (P<0·05). LPS exposure increased (P<0·05) the mRNA expression of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6 and IL-8), which was decreased by pre-treatment with 1,25-dihydroxyvitamin D (1,25D3) in a dose-dependent manner (P<0·05). Further results showed that pre-treatment with 1,25D3 down-regulated Toll-like receptor 4 (TLR4), myeloid differentiation primary response gene 88 (Myd88) and NF-κB p65 mRNA expression (P<0·05), suggesting potential mechanisms against LPS-induced inflammatory response. In vivo, intraperitoneal injection of LPS significantly increased TNF-α, IL-1β, IL-6 and IL-8 mRNA expression in the intestine of carp (P<0·05). Pre-treatment of fish with vitamin D3 protected the fish intestine from the LPS-induced increase of TNF-α, IL-1β, IL-6 and IL-8 mainly by downregulating TLR4, Myd88 and NF-κB p65 mRNA expression (P<0·05). These observations suggest that vitamin D could inhibit LPS-induced inflammatory response in juvenile Jian carp in vivo and in enterocytes in vitro. The anti-inflammatory effect of vitamin D is mediated at least in part by TLR4-Myd88 signalling pathways in the intestine and enterocytes of juvenile Jian carp.

The mucosal adjuvant cholera toxin B instructs non-mucosal dendritic cells to promote IgA production via retinoic acid and TGF-β.

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Anouk K Gloudemans

Full Text Available It is currently unknown how mucosal adjuvants cause induction of secretory immunoglobulin A (IgA, and how T cell-dependent (TD or -independent (TI pathways might be involved. Mucosal dendritic cells (DCs are the primary antigen presenting cells driving TI IgA synthesis, by producing a proliferation-inducing ligand (APRIL, B cell activating factor (BAFF, Retinoic Acid (RA, TGF-β or nitric oxide (NO. We hypothesized that the mucosal adjuvant Cholera Toxin subunit B (CTB could imprint non-mucosal DCs to induce IgA synthesis, and studied the mechanism of its induction. In vitro, CTB-treated bone marrow derived DCs primed for IgA production by B cells without the help of T cells, yet required co-signaling by different Toll-like receptor (TLR ligands acting via the MyD88 pathway. CTB-DC induced IgA production was blocked in vitro or in vivo when RA receptor antagonist, TGF-β signaling inhibitor or neutralizing anti-TGF-β was added, demonstrating the involvement of RA and TGF-β in promoting IgA responses. There was no major involvement for BAFF, APRIL or NO. This study highlights that synergism between CTB and MyD88-dependent TLR signals selectively imprints a TI IgA-inducing capacity in non-mucosal DCs, explaining how CTB acts as an IgA promoting adjuvant.

The mucosal adjuvant cholera toxin B instructs non-mucosal dendritic cells to promote IgA production via retinoic acid and TGF-β.

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Gloudemans, Anouk K; Plantinga, Maud; Guilliams, Martin; Willart, Monique A; Ozir-Fazalalikhan, Arifa; van der Ham, Alwin; Boon, Louis; Harris, Nicola L; Hammad, Hamida; Hoogsteden, Henk C; Yazdanbakhsh, Maria; Hendriks, Rudi W; Lambrecht, Bart N; Smits, Hermelijn H

2013-01-01

It is currently unknown how mucosal adjuvants cause induction of secretory immunoglobulin A (IgA), and how T cell-dependent (TD) or -independent (TI) pathways might be involved. Mucosal dendritic cells (DCs) are the primary antigen presenting cells driving TI IgA synthesis, by producing a proliferation-inducing ligand (APRIL), B cell activating factor (BAFF), Retinoic Acid (RA), TGF-β or nitric oxide (NO). We hypothesized that the mucosal adjuvant Cholera Toxin subunit B (CTB) could imprint non-mucosal DCs to induce IgA synthesis, and studied the mechanism of its induction. In vitro, CTB-treated bone marrow derived DCs primed for IgA production by B cells without the help of T cells, yet required co-signaling by different Toll-like receptor (TLR) ligands acting via the MyD88 pathway. CTB-DC induced IgA production was blocked in vitro or in vivo when RA receptor antagonist, TGF-β signaling inhibitor or neutralizing anti-TGF-β was added, demonstrating the involvement of RA and TGF-β in promoting IgA responses. There was no major involvement for BAFF, APRIL or NO. This study highlights that synergism between CTB and MyD88-dependent TLR signals selectively imprints a TI IgA-inducing capacity in non-mucosal DCs, explaining how CTB acts as an IgA promoting adjuvant.

Dual role of Fcγ receptors in host defense and disease in Borrelia burgdorferi-infected mice

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Alexia Anne Belperron

2014-06-01

Full Text Available Arthritis in mice infected with the Lyme disease spirochete, Borrelia burgdorferi, results from the influx of innate immune cells responding to the pathogen in the joint and is influenced in part by mouse genetics. Production of inflammatory cytokines by innate immune cells in vitro is largely mediated by Toll-like receptor (TLR interaction with Borrelia lipoproteins, yet surprisingly mice deficient in TLR2 or the TLR signaling molecule MyD88 still develop arthritis comparable to that seen in wild type mice after B. burgdorferi infection. These findings suggest that other, MyD88-independent inflammatory pathways can contribute to arthritis expression. Clearance of B. burgdorferi is dependent on the production of specific antibody and phagocytosis of the organism. As Fc receptors (FcγR are important for IgG-mediated clearance of immune complexes and opsonized particles by phagocytes, we examined the role that FcγR play in host defense and disease in B. burgdorferi-infected mice. B. burgdorferi-infected mice deficient in the Fc receptor common gamma chain (FcεRγ-/- mice harbored ~10 fold more spirochetes than similarly infected wild type mice, and this was associated with a transient increase in arthritis severity. While the elevated pathogen burdens seen in B. burgdorferi-infected MyD88-/- mice were not affected by concomitant deficiency in FcγR, arthritis was reduced in FcεRγ-/-MyD88-/- mice in comparison to wild type or single knockout mice. Gene expression analysis from infected joints demonstrated that absence of both MyD88 and FcγR lowers mRNA levels of proteins involved in inflammation, including Cxcl1 (KC, Xcr1 (Gpr5, IL-1beta, and C reactive protein. Taken together, our results demonstrate a role for FcγR-mediated immunity in limiting pathogen burden and arthritis in mice during the acute phase of B. burgdorferi infection, and further suggest that this pathway contributes to the arthritis that develops in B. burgdorferi

Multiple mechanisms involved in diabetes protection by lipopolysaccharide in non-obese diabetic mice

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Wang, Jun [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Department of Pharmacology, College of Medicine, Wuhan University of Science and Technology, Wuhan (China); Cao, Hui [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Wang, Hongjie [Section of Neurobiology, Torrey Pines Institute for Molecular Studies, Port Saint Lucie, FL (United States); Yin, Guoxiao; Du, Jiao; Xia, Fei; Lu, Jingli [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Xiang, Ming, E-mail: xiangming@mails.tjmu.edu.cn [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China)

2015-06-15

Toll-like receptor 4 (TLR4) activation has been proposed to be important for islet cell inflammation and eventually β cell loss in the course of type 1 diabetes (T1D) development. However, according to the “hygiene hypothesis”, bacterial endotoxin lipopolysaccharide (LPS), an agonist on TLR4, inhibits T1D progression. Here we investigated possible mechanisms for the protective effect of LPS on T1D development in non-obese diabetic (NOD) mice. We found that LPS administration to NOD mice during the prediabetic state neither prevented nor reversed insulitis, but delayed the onset and decreased the incidence of diabetes, and that a multiple-injection protocol is more effective than a single LPS intervention. Further, LPS administration suppressed spleen T lymphocyte proliferation, increased the generation of CD4{sup +}CD25{sup +}Foxp3{sup +} regulatory T cells (Tregs), reduced the synthesis of strong Th1 proinflammatory cytokines, and downregulated TLR4 and its downstream MyD88-dependent signaling pathway. Most importantly, multiple injections of LPS induced a potential tolerogenic dendritic cell (DC) subset with low TLR4 expression without influencing the DC phenotype. Explanting DCs from repeated LPS-treated NOD mice into NOD/SCID diabetic mice conferred sustained protective effects against the progression of diabetes in the recipients. Overall, these results suggest that multiple mechanisms are involved in the protective effects of LPS against the development of diabetes in NOD diabetic mice. These include Treg induction, down-regulation of TLR4 and its downstream MyD88-dependent signaling pathway, and the emergence of a potential tolerogenic DC subset. - Highlights: • Administration of lipopolysaccharide (LPS) prevented type 1 diabetes in NOD mice. • Downregulating TLR4 level and MyD88-dependent pathway contributed to protection of LPS. • LPS administration also hampered DC maturation and promoted Treg differentiation.

Multiple mechanisms involved in diabetes protection by lipopolysaccharide in non-obese diabetic mice

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Wang, Jun; Cao, Hui; Wang, Hongjie; Yin, Guoxiao; Du, Jiao; Xia, Fei; Lu, Jingli; Xiang, Ming

2015-01-01

Toll-like receptor 4 (TLR4) activation has been proposed to be important for islet cell inflammation and eventually β cell loss in the course of type 1 diabetes (T1D) development. However, according to the “hygiene hypothesis”, bacterial endotoxin lipopolysaccharide (LPS), an agonist on TLR4, inhibits T1D progression. Here we investigated possible mechanisms for the protective effect of LPS on T1D development in non-obese diabetic (NOD) mice. We found that LPS administration to NOD mice during the prediabetic state neither prevented nor reversed insulitis, but delayed the onset and decreased the incidence of diabetes, and that a multiple-injection protocol is more effective than a single LPS intervention. Further, LPS administration suppressed spleen T lymphocyte proliferation, increased the generation of CD4 + CD25 + Foxp3 + regulatory T cells (Tregs), reduced the synthesis of strong Th1 proinflammatory cytokines, and downregulated TLR4 and its downstream MyD88-dependent signaling pathway. Most importantly, multiple injections of LPS induced a potential tolerogenic dendritic cell (DC) subset with low TLR4 expression without influencing the DC phenotype. Explanting DCs from repeated LPS-treated NOD mice into NOD/SCID diabetic mice conferred sustained protective effects against the progression of diabetes in the recipients. Overall, these results suggest that multiple mechanisms are involved in the protective effects of LPS against the development of diabetes in NOD diabetic mice. These include Treg induction, down-regulation of TLR4 and its downstream MyD88-dependent signaling pathway, and the emergence of a potential tolerogenic DC subset. - Highlights: • Administration of lipopolysaccharide (LPS) prevented type 1 diabetes in NOD mice. • Downregulating TLR4 level and MyD88-dependent pathway contributed to protection of LPS. • LPS administration also hampered DC maturation and promoted Treg differentiation

Anti-Inflammatory Action of an Antimicrobial Model Peptide That Suppresses the TRIF-Dependent Signaling Pathway via Inhibition of Toll-Like Receptor 4 Endocytosis in Lipopolysaccharide-Stimulated Macrophages.

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Do-Wan Shim

Full Text Available Antimicrobial peptides (AMPs, also called host defense peptides, particularly those with amphipathic helical structures, are emerging as target molecules for therapeutic development due to their immunomodulatory properties. Although the antimicrobial activity of AMPs is known to be exerted primarily by permeation of the bacterial membrane, the mechanism underlying its anti-inflammatory activity remains to be elucidated. We report potent anti-inflammatory activity of WALK11.3, an antimicrobial model peptide with an amphipathic helical conformation, in lipopolysaccharide (LPS-stimulated RAW264.7 cells. This peptide inhibited the expression of inflammatory mediators, including nitric oxide, COX-2, IL-1β, IL-6, INF-β, and TNF-α. Although WALK11.3 did not exert a major effect on all downstream signaling in the MyD88-dependent pathway, toll-like receptor 4 (TLR4- mediated pro-inflammatory signals were markedly attenuated in the TRIF-dependent pathway due to inhibition of the phosphorylation of STAT1 by attenuation of IRF3 phosphorylation. WALK11.3 specifically inhibited the endocytosis of TLR4, which is essential for triggering TRIF-mediated signaling in macrophage cells. Hence, we suggest that specific interference with TLR4 endocytosis could be one of the major modes of the anti-inflammatory action of AMPs. Our designed WALK11 peptides, which possess both antimicrobial and anti-inflammatory activities, may be promising molecules for the development of therapies for infectious inflammation.

Protective Effect of 1,25-Dihydroxy Vitamin D3 on Pepsin-Trypsin-Resistant Gliadin-Induced Tight Junction Injuries.

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Dong, Shouquan; Singh, Tikka Prabhjot; Wei, Xin; Yao, Huang; Wang, Hongling

2018-01-01

Tight junction (TJ) injuries induced by pepsin-trypsin-resistant gliadin (PT-G) play an important role in the pathogenesis of celiac disease. Previously, 1,25-dihydroxy vitamin D3 (VD3) was reported to be a TJ regulator that attenuates lipopolysaccharide- and alcohol-induced TJ injuries. However, whether VD3 can attenuate PT-G-induced TJ injuries is unknown. The aim of this study was to evaluate the effects of VD3 on PT-G-induced TJ injuries. Caco-2 monolayers were used as in vitro models. After being cultured for 21 days, the monolayers were treated with PT-G plus different concentrations of VD3. Then, the changes in trans-epithelial electrical resistance and FITC-dextran 4000 (FD-4) flux were determined to evaluate the monolayer barrier function. TJ protein levels were measured to assess TJ injury severity, and myeloid differentiation factor 88 (MyD88) expression and zonulin release levels were determined to estimate zonulin release signaling pathway activity. Additionally, a gluten-sensitized mouse model was established as an in vivo model. After the mice were treated with VD3 for 7 days, we measured serum FD-4 concentrations, TJ protein levels, MyD88 expression, and zonulin release levels to confirm the effect of VD3. Both in vitro and in vivo, VD3 significantly attenuated the TJ injury-related increase in intestinal mucosa barrier permeability. Moreover, VD3 treatment up-regulated TJ protein expression levels and significantly decreased MyD88 expression and zonulin release levels. VD3 has protective effects against PT-G-induced TJ injuries both in vitro and in vivo, which may correlate with the disturbance of the MyD88-dependent zonulin release signaling pathway.

New advances of TMEM88 in cancer initiation and progression, with special emphasis on Wnt signaling pathway.

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Ge, Yun-Xuan; Wang, Chang-Hui; Hu, Fu-Yong; Pan, Lin-Xin; Min, Jie; Niu, Kai-Yuan; Zhang, Lei; Li, Jun; Xu, Tao

2018-01-01

Transmembrane protein 88 (TMEM88), a newly discovered protein localized on the cell membrane. Recent studies showed that TMEM88 was involved in the regulation of several types of cancer. TMEM88 was expressed at significantly higher levels in breast cancer (BC) cell line than in normal breast cell line with co-localized with Dishevelled (DVL) in the cytoplasm of BC cell line. TMEM88 silencing in the ovarian cancer cell line CP70 resulted in significant upregulation of Wnt downstream genes (c-Myc, cyclin-D1) and other Wnt target genes including JUN, PTIX2, CTNNB1 (β-catenin), further supporting that TMEM88 inhibits canonical Wnt signaling pathway. Wnt signaling pathway has been known to play important roles in many diseases, especially in cancer. For instance, hepatocellular carcinoma (HCC) has become one of the most common tumors harboring mutations in the Wnt signaling pathway. As the inhibitor of Wnt signaling, TMEM88 has been considered to act as an oncogene or a tumor suppressor. Up-regulated TMEM88 or gene therapy approaches could be an effective therapeutic approach against tumor as TMEM88 inhibits Wnt signaling through direct interaction with DVL. Here, we review the current knowledge on the functional role and potential clinical application of TMEM88 in the control of various cancers. Highlights Wnt signaling displays an important role in several pathogenesis of cancer. Wnt signaling pathway is activated during cancer development. TMEM88 has an impact on cancer by inhibiting canonical Wnt signaling. We discuss the importance and new applications of TMEM88 in cancer therapy. © 2017 Wiley Periodicals, Inc.

Gene structure, cDNA characterization and RNAi-based functional analysis of a myeloid differentiation factor 88 homolog in Tenebrio molitor larvae exposed to Staphylococcus aureus infection.

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Patnaik, Bharat Bhusan; Patnaik, Hongray Howrelia; Seo, Gi Won; Jo, Yong Hun; Lee, Yong Seok; Lee, Bok Luel; Han, Yeon Soo

2014-10-01

Myeloid differentiation factor 88 (MyD88), an intracellular adaptor protein involved in Toll/Toll-like receptor (TLR) signal processing, triggers activation of nuclear factor-kappaB (NF-κB) transcription factors. In the present study, we analyzed the gene structure and biological function of MyD88 in a coleopteran insect, Tenebrio molitor (TmMyD88). The TmMyD88 gene was 1380 bp in length and consisted of five exons and four introns. The 5'-flanking sequence revealed several putative transcription factor binding sites, such as STAT-4, AP-1, cJun, cfos, NF-1 and many heat shock factor binding elements. The cDNA contained a typical death domain, a conservative Toll-like interleukin-1 receptor (TIR) domain, and a C-terminal extension (CTE). The TmMyD88 TIR domain showed three significantly conserved motifs for interacting with the TIR domain of TLRs. TmMyD88 was grouped within the invertebrate cluster of the phylogenetic tree and shared 75% sequence identity with the TIR domain of Tribolium castaneum MyD88. Homology modeling of the TmMyD88 TIR domain revealed five parallel β-strands surrounded by five α-helices that adopted loop conformations to function as an adaptor. TmMyD88 expression was upregulated 7.3- and 4.79-fold after 12 and 6h, respectively, of challenge with Staphylococcus aureus and fungal β-1,3 glucan. Silencing of the TmMyD88 transcript by RNA interference led to reduced resistance of the host to infection by S. aureus. These results indicate that TmMyD88 is required for survival against Staphylococcus infection. Copyright © 2014 Elsevier Ltd. All rights reserved.

Ganoderma lucidum ethanol extract inhibits the inflammatory response by suppressing the NF-κB and toll-like receptor pathways in lipopolysaccharide-stimulated BV2 microglial cells.

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Yoon, Hyun-Min; Jang, Kyung-Jun; Han, Min Seok; Jeong, Jin-Woo; Kim, Gi Young; Lee, Jai-Heon; Choi, Yung Hyun

2013-03-01

Ganoderma lucidum is a traditional Oriental medicine that has been widely used as a tonic to promote longevity and health in Korea and other Asian countries. Although a great deal of work has been carried out on the therapeutic potential of this mushroom, the pharmacological mechanisms of its anti-inflammatory actions remain unclear. In this study, we evaluated the inhibitory effects of G. lucidum ethanol extract (EGL) on the production of inflammatory mediators and cytokines in lipopolysaccharide (LPS)-stimulated murine BV2 microglia. We also investigated the effects of EGL on the LPS-induced activation of nuclear factor kappaB (NF-κB) and upregulation of toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 (MyD88). Elevated levels of nitric oxide (NO), prostaglandin E(2) (PGE(2)) and pro-inflammatory cytokine production were detected in BV2 microglia following LPS stimulation. We identifed that EGL significantly inhibits the excessive production of NO, PGE(2) and pro-inflammatory cytokines, including interleukin (IL)-1β and tumor necrosis factor-α in a concentration-dependent manner without causing cytotoxicity. In addition, EGL suppressed NF-κB translocation and transcriptional activity by blocking IκB degradation and inhibiting TLR4 and MyD88 expression in LPS-stimulated BV2 cells. Our results indicate that the inhibitory effects of EGL on LPS-stimulated inflammatory responses in BV2 microglia are associated with the suppression of the NF-κB and TLR signaling pathways. Therefore, EGL may be useful in the treatment of neurodegenerative diseases by inhibiting inflammatory mediator responses in activated microglia.

Commensal Microbiota Are Required for Systemic Inflammation Triggered by Necrotic Dendritic Cells

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Jennifer A. Young

2013-06-01

Full Text Available The relationship between dendritic cells (DCs and commensal microflora in shaping systemic immune responses is not well understood. Here, we report that mice deficient for the Fas-associated death domain in DCs developed systemic inflammation associated with elevated proinflammatory cytokines and increased myeloid and B cells. These mice exhibited reduced DCs in gut-associated lymphoid tissues due to RIP3-dependent necroptosis, whereas DC functions remained intact. Induction of systemic inflammation required DC necroptosis and commensal microbiota signals that activated MyD88-dependent pathways in other cell types. Systemic inflammation was abrogated with the administration of broad-spectrum antibiotics or complete, but not DC-specific, deletion of MyD88. Thus, we have identified a previously unappreciated role for commensal microbiota in priming immune cells for inflammatory responses against necrotic cells. These studies demonstrate the impact intestinal microflora have on the immune system and their role in eliciting proper immune responses to harmful stimuli.

Suppression of the toll-like receptor 7-dependent type I interferon production pathway by autophagy resulting from enterovirus 71 and coxsackievirus A16 infections facilitates their replication.

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Song, Jie; Hu, Yajie; Li, Jiaqi; Zheng, Huiwen; Wang, Jingjing; Guo, Lei; Shi, Haijng; Liu, Longding

2018-01-01

Toll-like receptors (TLRs) act as molecular sentinels, detecting invading viral pathogens and triggering host innate immune responses, including autophagy. However, many viruses have evolved a series of strategies to manipulate autophagy for their own benefit. Enterovirus 71 (EV71) and coxsackievirus A16 (CA16), as the primary agents causing hand, foot and mouth disease (HFMD), can induce autophagy leading to their replication. Therefore, the objective of this study was to investigate whether enhanced viral replication caused by autophagy in EV71 and CA16 infections was associated with a TLR-related signaling pathway. Our results demonstrate that complete autophagy and incomplete autophagy were observed in human bronchial epithelial (16HBE) cells infected with EV71 and CA16. Moreover, suppression of autophagy by the pharmacological modulator 3-MA significantly and clearly decreased the survival rates and viral replication of EV71 and CA16 in 16HBE cells. Inhibition of autophagy also enhanced the expression of molecules related to the TLR7-dependent type I interferon (IFN-I) production pathway, such as TLR7, MyD88, IRF7 and IFN-α/β. Finally, immunofluorescence staining demonstrated that TLR7 endosome marker M6PR levels were clearly reduced in EV71- and CA16-infected cells, while they were markedly elevated in infected cells treated with 3-MA. These findings suggest that increased EV71 and CA16 replication meditated by autophagy in 16HBE cells might promote degradation of the endosome, leading to suppression of the TLR7-mediated IFN-I signaling pathway.

TLR9 played a more important role than TLR2 in the combination of maltose-binding protein and BCG-induced Th1 activation.

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Ni, Weihua; Wang, Fang; Liu, Guomu; Zhang, Nannan; Yuan, Hongyan; Jie, Jing; Tai, Guixiang

2016-11-01

Our previous study demonstrated that maltose-binding protein (MBP) combined with BCG induced synergistic mouse Th1 activation in vivo. Here, to explore the mechanism of MBP combined with BCG on Th1 activation, mouse purified CD4 + T cells were stimulated with MBP and BCG in vitro. The results showed that MBP combined with BCG synergistically increased IFN-γ production, accompanied with the upregulation of TLR2/9 expressions, suggesting that TLR2/9 were involved in the combination-induced Th1 activation. Next, TLR2 antibodies and TLR9 inhibitor were used to further analyze the effects of TLRs in Th1 activation. Results showed TLR2 antibody partly decreased MBP combined with BCG-induced IFN-γ production, MyD88 expression and IκB phosphorylation, indicating that TLR2-mediated MyD88-dependent pathway was involved in the MBP combined with BCG-induced Th1 activation. Moreover, MBP combined with BCG-induced Th1 activation was completely abrogated by TLR9 inhibitor, suggesting that TLR9-mediated MyD88-dependent pathway played a more important role than TLR2 in the combination-induced Th1 activation. Further study showed that TLR9 inhibitor downregulated TLR2 expression, suggesting that TLR9 signaling regulated TLR2 activation to favor Th1 resonse induced by MBP combined with BCG. Collectively, we demonstrated for the first time that the cross-talk of TLR2 and TLR9 triggered Th1 activation collaboratively and our findings provided valuable information about designing more effective adjuvant for cancer therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Andrographolide suppresses proliferation of human colon cancer SW620 cells through the TLR4/NF-κB/MMP-9 signaling pathway.

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Zhang, Rui; Zhao, Jian; Xu, Jian; Jiao, De-Xin; Wang, Jian; Gong, Zhi-Qiang; Jia, Jian-Hui

2017-10-01

Modern pharmacological research has revealed that andrographolide has various functions, including anti-bacterial, anti-inflammatory and anti-viral effects, immunoregulation, treating cardiovascular and cerebrovascular diseases, and prevention and treatment of alcoholic liver injury. The present study investigated whether andrographolide suppresses the proliferation of human colon cancer cell through the Toll-like receptor 4 (TLR4)/nuclear factor (NF)-κB/matrix metalloproteinase-9 (MMP-9) signaling pathway. The MTT assay and lactate dehydrogenase assay were used to evaluate the anticancer effects of andrographolide on cell proliferation and cytotoxicity in human colon cancer SW620 cells. Flow cytometry was used to analyze the anticancer effects of andrographolide on apoptosis by Annexin V-fluorescein isothiocyanate/propidium iodide kit. The effects of andrographolide on the activity of caspase-3/9 were measured using ELISA. Western blot analysis was also used to analyze the protein expression of TLR4, myeloid differentiation primary response gene 88 (MyD88), NF-κB-p65 and MMP-9. In the present study, it was found that andrographolide suppressed the cell proliferation, augmented cytotoxicity, evoked cell apoptosis and activated caspase-3/9 activities in human colon cancer SW620 cells. The results revealed that the anti-proliferation effects of andrographolide on the SW620 cells was associated with the inhibition of TLR4, MyD88, NF-κB-p65 and MMP-9 signaling activation. The results suggest that andrographolide is a promising drug for treatment of human colon cancer via suppression of the TLR4/NF-κB/MMP-9 signaling pathway.

Src-family-tyrosine kinase Lyn is critical for TLR2-mediated NF-κB activation through the PI 3-kinase signaling pathway.

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Toubiana, Julie; Rossi, Anne-Lise; Belaidouni, Nadia; Grimaldi, David; Pene, Frederic; Chafey, Philippe; Comba, Béatrice; Camoin, Luc; Bismuth, Georges; Claessens, Yann-Erick; Mira, Jean-Paul; Chiche, Jean-Daniel

2015-10-01

TLR2 has a prominent role in host defense against a wide variety of pathogens. Stimulation of TLR2 triggers MyD88-dependent signaling to induce NF-κB translocation, and activates a Rac1-PI 3-kinase dependent pathway that leads to transactivation of NF-κB through phosphorylation of the P65 NF-κB subunit. This transactivation pathway involves tyrosine phosphorylations. The role of the tyrosine kinases in TLR signaling is controversial, with discrepancies between studies using only chemical inhibitors and knockout mice. Here, we show the involvement of the tyrosine-kinase Lyn in TLR2-dependent activation of NF-κB in human cellular models, by using complementary inhibition strategies. Stimulation of TLR2 induces the formation of an activation cluster involving TLR2, CD14, PI 3-kinase and Lyn, and leads to the activation of AKT. Lyn-dependent phosphorylation of the p110 catalytic subunit of PI 3-kinase is essential to the control of PI 3-kinase biological activity upstream of AKT and thereby to the transactivation of NF-κB. Thus, Lyn kinase activity is crucial in TLR2-mediated activation of the innate immune response in human mononuclear cells. © The Author(s) 2015.

Microbiota promote secretory cell determination in the intestinal epithelium by modulating host Notch signaling.

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Troll, Joshua V; Hamilton, M Kristina; Abel, Melissa L; Ganz, Julia; Bates, Jennifer M; Stephens, W Zac; Melancon, Ellie; van der Vaart, Michiel; Meijer, Annemarie H; Distel, Martin; Eisen, Judith S; Guillemin, Karen

2018-02-23

Resident microbes promote many aspects of host development, although the mechanisms by which microbiota influence host tissues remain unclear. We showed previously that the microbiota is required for allocation of appropriate numbers of secretory cells in the zebrafish intestinal epithelium. Because Notch signaling is crucial for secretory fate determination, we conducted epistasis experiments to establish whether the microbiota modulates host Notch signaling. We also investigated whether innate immune signaling transduces microbiota cues via the Myd88 adaptor protein. We provide the first evidence that microbiota-induced, Myd88-dependent signaling inhibits host Notch signaling in the intestinal epithelium, thereby promoting secretory cell fate determination. These results connect microbiota activity via innate immune signaling to the Notch pathway, which also plays crucial roles in intestinal homeostasis throughout life and when impaired can result in chronic inflammation and cancer. © 2018. Published by The Company of Biologists Ltd.

The IkappaB kinase family phosphorylates the Parkinson's disease kinase LRRK2 at Ser935 and Ser910 during Toll-like receptor signaling.

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Nicolas Dzamko

Full Text Available Mutations in leucine-rich repeat kinase 2 (LRRK2 are strongly associated with late-onset autosomal dominant Parkinson's disease. LRRK2 is highly expressed in immune cells and recent work points towards a link between LRRK2 and innate immunity. Here we demonstrate that stimulation of the Toll-Like Receptor (TLR pathway by MyD88-dependent agonists in bone marrow-derived macrophages (BMDMs or RAW264.7 macrophages induces marked phosphorylation of LRRK2 at Ser910 and Ser935, the phosphorylation sites that regulate the binding of 14-3-3 to LRRK2. Phosphorylation of these residues is prevented by knock-out of MyD88 in BMDMs, but not the alternative TLR adaptor protein TRIF. Utilising both pharmacological inhibitors, including a new TAK1 inhibitor, NG25, and genetic models, we provide evidence that both the canonical (IKKα and IKKβ and IKK-related (IKKε and TBK1 kinases mediate TLR agonist induced phosphorylation of LRRK2 in vivo. Moreover, all four IKK members directly phosphorylate LRRK2 at Ser910 and Ser935 in vitro. Consistent with previous work describing Ser910 and Ser935 as pharmacodynamic biomarkers of LRRK2 activity, we find that the TLR independent basal phosphorylation of LRRK2 at Ser910 and Ser935 is abolished following treatment of macrophages with LRRK2 kinase inhibitors. However, the increased phosphorylation of Ser910 and Ser935 induced by activation of the MyD88 pathway is insensitive to LRRK2 kinase inhibitors. Finally, employing LRRK2-deficient BMDMs, we present data indicating that LRRK2 does not play a major role in regulating the secretion of inflammatory cytokines induced by activation of the MyD88 pathway. Our findings provide the first direct link between LRRK2 and the IKKs that mediate many immune responses. Further work is required to uncover the physiological roles that phosphorylation of LRRK2 by IKKs play in controlling macrophage biology and to determine how phosphorylation of LRRK2 by IKKs impacts upon the use of Ser

Human metapneumovirus M2-2 protein inhibits innate immune response in monocyte-derived dendritic cells.

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Junping Ren

Full Text Available Human metapneumovirus (hMPV is a leading cause of lower respiratory infection in young children, the elderly and immunocompromised patients. Repeated hMPV infections occur throughout life. However, immune evasion mechanisms of hMPV infection are largely unknown. Recently, our group has demonstrated that hMPV M2-2 protein, an important virulence factor, contributes to immune evasion in airway epithelial cells by targeting the mitochondrial antiviral-signaling protein (MAVS. Whether M2-2 regulates the innate immunity in human dendritic cells (DC, an important family of immune cells controlling antigen presenting, is currently unknown. We found that human DC infected with a virus lacking M2-2 protein expression (rhMPV-ΔM2-2 produced higher levels of cytokines, chemokines and IFNs, compared to cells infected with wild-type virus (rhMPV-WT, suggesting that M2-2 protein inhibits innate immunity in human DC. In parallel, we found that myeloid differentiation primary response gene 88 (MyD88, an essential adaptor for Toll-like receptors (TLRs, plays a critical role in inducing immune response of human DC, as downregulation of MyD88 by siRNA blocked the induction of immune regulatory molecules by hMPV. Since M2-2 is a cytoplasmic protein, we investigated whether M2-2 interferes with MyD88-mediated antiviral signaling. We found that indeed M2-2 protein associated with MyD88 and inhibited MyD88-dependent gene transcription. In this study, we also identified the domains of M2-2 responsible for its immune inhibitory function in human DC. In summary, our results demonstrate that M2-2 contributes to hMPV immune evasion by inhibiting MyD88-dependent cellular responses in human DC.

Phenotypic and pathologic evaluation of the myd mouse. A candidate model for facioscapulohumeral dystrophy

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Mathews, K.D.; Rapisarda, D.; Bailey, H.L. [Univ. of Iowa College of Medicine, Iowa City, IA (United States)] [and others

1995-07-01

Facioscapulohumeral dystrophy (FSHD) is an autosomal dominant disease of unknown pathogenesis which is characterized by weakness of the face and shoulder girdle. It is associated with a sensorineural hearing loss which may be subclinical. FSHD has been mapped to the distalmost portion of 4q35, although the gene has not yet been identified. Distal 4q has homology with a region of mouse chromosome 8 to which a mouse mutant, myodystrophy (myd), has been mapped. Muscle from homozygotes for the myd mutation appears dystrophic, showing degenerating and regenerating fibers, inflammatory infiltrates, central nuclei, and variation in fiber size. Brainstem auditory evoked potentials reveal a sensorineural hearing loss in myd homozygotes. Based on the homologous genetic map locations, and the phenotypic syndrome of dystrophic muscle with sensorineural hearing loss, we suggest that myd represents an animal model for the human disease FSHD. 28 refs., 4 figs.

IL-1RI (Interleukin-1 Receptor Type I Signalling is Essential for Host Defence and Hemichannel Activity During Acute Central Nervous System Bacterial Infection

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Juan Xiong

2012-03-01

Full Text Available Staphylococcus aureus is a common aetiological agent of bacterial brain abscesses. We have previously established that a considerable IL-1 (interleukin-1 response is elicited immediately following S. aureus infection, where the cytokine can exert pleiotropic effects on glial activation and blood–brain barrier permeability. To assess the combined actions of IL-1α and IL-1β during CNS (central nervous system infection, host defence responses were evaluated in IL-1RI (IL-1 receptor type I KO (knockout animals. IL-1RI KO mice were exquisitely sensitive to intracerebral S. aureus infection, as demonstrated by enhanced mortality rates and bacterial burdens within the first 24 h following pathogen exposure compared with WT (wild-type animals. Loss of IL-1RI signalling also dampened the expression of select cytokines and chemokines, concomitant with significant reductions in neutrophil and macrophage infiltrates into the brain. In addition, the opening of astrocyte hemichannels during acute infection was shown to be dependent on IL-1RI activity. Collectively, these results demonstrate that IL-1RI signalling plays a pivotal role in the genesis of immune responses during the acute stage of brain abscess development through S. aureus containment, inflammatory mediator production, peripheral immune cell recruitment, and regulation of astrocyte hemichannel activity. Taken in the context of previous studies with MyD88 (myeloid differentiation primary response gene 88 and TLR2 (Toll-like receptor 2 KO animals, the current report advances our understanding of MyD88-dependent cascades and implicates IL-1RI signalling as a major antimicrobial effector pathway during acute brain-abscess formation.

CD36 Differently Regulates Macrophage Responses to Smooth and Rough Lipopolysaccharide.

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Rafał Biedroń

Full Text Available Lipopolysaccharide (LPS is the major pathogen-associated molecular pattern of Gram-negative bacterial infections, and includes smooth (S-LPS and rough (R-LPS chemotypes. Upon activation by LPS through CD14, TLR4/MD-2 heterodimers sequentially induce two waves of intracellular signaling for macrophage activation: the MyD88-dependent pathway from the plasma membrane and, following internalization, the TRIF-dependent pathway from endosomes. We sought to better define the role of scavenger receptors CD36 and CD204/SR-A as accessory LPS receptors that can contribute to pro-inflammatory and microbicidal activation of macrophages. We have found that CD36 differently regulates activation of mouse macrophages by S-LPS versus R-LPS. The ability of CD36 to substitute for CD14 in loading R-LPS, but not S-LPS onto TLR4/MD-2 allows CD14-independent macrophage responses to R-LPS. Conversely, S-LPS, but not R-LPS effectively stimulates CD14 binding to CD36, which favors S-LPS transfer from CD14 onto TLR4/MD-2 under conditions of low CD14 occupancy with S-LPS in serum-free medium. In contrast, in the presence of serum, CD36 reduces S-LPS binding to TLR4/MD-2 and the subsequent MyD88-dependent signaling, by mediating internalization of S-LPS/CD14 complexes. Additionally, CD36 positively regulates activation of TRIF-dependent signaling by both S-LPS and R-LPS, by promoting TLR4/MD-2 endocytosis. In contrast, we have found that SR-A does not function as a S-LPS receptor. Thus, by co-operating with CD14 in both R- and S-LPS loading onto TLR4/MD-2, CD36 can enhance the sensitivity of tissue-resident macrophages in detecting infections by Gram-negative bacteria. However, in later phases, following influx of serum to the infection site, the CD36-mediated negative regulation of MyD88-dependent branch of S-LPS-induced TLR4 signaling might constitute a mechanism to prevent an excessive inflammatory response, while preserving the adjuvant effect of S-LPS for adaptive

Radioresistance-related signaling pathways in nasopharyngeal carcinoma cells

International Nuclear Information System (INIS)

Guo Ya; Zhu Xiaodong; Qu Song; Su Fang; Wang Qi; Zhang Wei

2011-01-01

Objective: To study the difference of gene expression profile between the radioresistant human nasopharyngeal carcinoma cell line CNE-2R and CNE-2, and to screen the signaling pathway associated with radioresistance of nasopharyngeal carcinoma. Methods: The radioresistant nasopharyngeal carcinoma cell line CNE-2R was constructed from the original cell line CNE-2. CNE-2R and CNE-2 cells were cultured and administered with 60 Co γ-ray irradiation at the dose of 400 cGy for 15 times. Human-6v 3.0 whole genome expression profile was used to screen the differentially expressed genes. Bioinformatic analysis was used to identify the pathways related to radioresistance. Results: The number of the differentially expressed genes that were found in these 2 experiments was 374. The Kegg pathway and Biocarta pathway analysis of the differentially expressed genes showed the biological importance of Toll-like receptor signaling pathway and IL-1 R-mediated signal transduction pathway to the radioresistance of the CNE-2R cells and the significant differences of 13 genes in these 2 pathways,including JUN, MYD88, CCL5, CXCL10, STAT1, LY96, FOS, CCL3, IL-6, IL-8, IL-1α, IL-1β, and IRAK2 (t=13.47-66.57, P<0.05). Conclusions: Toll-like receptor signaling pathway and IL-1R-mediated signal transduction pathway might be related to the occurrence of radioresistance. (authors)

TLR4 Signaling Pathway Modulators as Potential Therapeutics in Inflammation and Sepsis

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Nikolay N. Kuzmich

2017-10-01

Full Text Available Toll-Like Receptor 4 (TLR4 signal pathway plays an important role in initiating the innate immune response and its activation by bacterial endotoxin is responsible for chronic and acute inflammatory disorders that are becoming more and more frequent in developed countries. Modulation of the TLR4 pathway is a potential strategy to specifically target these pathologies. Among the diseases caused by TLR4 abnormal activation by bacterial endotoxin, sepsis is the most dangerous one because it is a life-threatening acute system inflammatory condition that still lacks specific pharmacological treatment. Here, we review molecules at a preclinical or clinical phase of development, that are active in inhibiting the TLR4-MyD88 and TLR4-TRIF pathways in animal models. These are low-molecular weight compounds of natural and synthetic origin that can be considered leads for drug development. The results of in vivo studies in the sepsis model and the mechanisms of action of drug leads are presented and critically discussed, evidencing the differences in treatment results from rodents to humans.

The immune impact of mimic endoscopic retrograde appendicitis therapy and appendectomy on rabbits of acute appendicitis.

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Liu, Suqin; Pei, Fenghua; Wang, Xinhong; Li, Deliang; Zhao, Lixia; Song, Yanyan; Chen, Zhendong; Liu, Bingrong

2017-09-12

This study was conducted to evaluate the immune impact of mimic endoscopic retrograde appendicitis therapy and appendectomy on rabbits of acute suppurative appendicitis and to determine whether TLR4/MYD88/NF-κB signaling pathway was activated in this process. 48 rabbits were assigned into 4 groups: group I, the mimic endoscopic retrograde appendicitis therapy group; group II, the appendectomy group; group III, the model group; and group IV, the blank group. White blood cells decreased, while levels of C-reactive protein, tumor necrosis factor-α, interleukin-6, interleukin-4, and interleukin-10 increased on the 2 nd day in group I and II. IgA in feces decreased at 2 weeks, while fecal microbiota changed at 2 and 4 weeks after appendectomy. CD8 + cells in appendix of group I increased within 8 weeks. Upregulated expression of TLR4, MYD88, and nuclear NF-κB were detected on the 2 nd day in group I and II. Mimic endoscopic retrograde appendicitis therapy and appendectomy are effective ways for acute suppurative appendicitis. Mimic endoscopic retrograde appendicitis therapy was more preferable due to its advantage in maintaining intestinal immune function. TLR4/MYD88/NF-κB signaling pathway was activated in acute phase of appendicitis.

Support Care Facility Prague House, Freshford, Kilkenny

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Li, Chong Hui

2012-02-03

Tolerance to bacterial cell-wall components may represent an essential regulatory mechanism during bacterial infection. We have demonstrated previously that the inhibition of nuclear factor (NF)-kappaB and mitogen-activated protein kinase activation was present in bacterial lipoprotein (BLP) self-tolerance and its cross-tolerance to lipopolysaccharide (LPS). In this study, the effect of BLP-induced tolerance on the myeloid differentiation factor 88 (MyD88)-dependent upstream signaling pathway for NF-kappaB activation in vitro was examined further. When compared with nontolerant human monocytic THP-1 cells, BLP-tolerant cells had a significant reduction in tumor necrosis factor alpha (TNF-alpha) production in response to a high-dose BLP (86+\\/-12 vs. 6042+\\/-245 ng\\/ml, P < 0.01) or LPS (341+\\/-36 vs. 7882+\\/-318 ng\\/ml, P < 0.01) stimulation. The expression of Toll-like receptor 2 (TLR2) protein was down-regulated in BLP-tolerant cells, whereas no significant differences in TLR4, MyD88, interleukin-1 receptor-associated kinase 4 (IRAK-4), and TNF receptor-associated factor 6 expression were observed between nontolerant and BLP-tolerant cells, as confirmed by Western blot analysis. The IRAK-1 protein was reduced markedly in BLP-tolerant cells, although IRAK-1 mRNA expression remained unchanged as revealed by real-time reverse transcriptase-polymerase chain reaction analysis. Furthermore, decreased MyD88-IRAK immunocomplex formation, as demonstrated by immunoprecipitation, was observed in BLP-tolerant cells following a second BLP or LPS stimulation. BLP pretreatment also resulted in a marked inhibition in total and phosphorylated inhibitor of kappaB-alpha (IkappaB-alpha) expression, which was not up-regulated by subsequent BLP or LPS stimulation. These results demonstrate that in addition to the down-regulation of TLR2 expression, BLP tolerance is associated with a reduction in IRAK-1 expression, MyD88-IRAK association, and IkappaB-alpha phosphorylation. These

The potent suppressive effect of β-d-mannuronic acid (M2000) on molecular expression of the TLR/NF-kB Signaling Pathway in ankylosing spondylitis patients.

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Roozbehkia, Maryam; Mahmoudi, Mahdi; Aletaha, Somaye; Rezaei, Nima; Fattahi, Mohammad Javad; Jafarnezhad-Ansariha, Fahimeh; Barati, Anis; Mirshafiey, Abbas

2017-11-01

Ankylosing spondylitis (AS) is a chronic inflammatory rheumatic disease characterized by the inflammation of sacroiliac joints and axial skeleton. A combination of genetic, environmental and immunological factors are involved in AS's pathogenesis. TLRs are type I transmembrane glycoproteins that play a crucial role in the innate immune responses against invading pathogens. Observational studies have demonstrated a possible association between TLR dysregulation and AS. The β-d-mannuronic acid (M2000), as a novel NSAID with immunosuppressive property, has shown an inhibitory effect on Toll-like receptor (TLR) 2, 4 signaling in HEK293 cells. In the present study, we investigated the gene expression of Myd88, IKB-alpha, NF-kB and MAPK14 (genes of the TLR/NF-kB Signaling Pathway) in AS patients in comparison to healthy subjects and also the effect of β-d-mannuronic acid on disease activity and mRNA expression of these molecules in affected patients. We showed for the first time that the gene expression level of Myd88, IKB-alpha, NF-kB and MAPK14 was higher in AS patients in comparison to healthy subjects. Moreover we confirmed that the β-d-mannuronic acid not just reduced significantly the disease activity of AS individuals compared to placebo, but also it could significantly decrease the expression level of genes associated with TLR/NF-kB Signaling Pathway in treated patients with M2000. These results may provide a new therapeutic approach to attenuate inflammatory responses in AS patients, (Identified; IRCT 2013062213739N1). Copyright © 2017. Published by Elsevier B.V.

Leptospira santorosai Serovar Shermani detergent extract induces an increase in fibronectin production through a Toll-like receptor 2-mediated pathway.

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Tian, Ya-Chung; Hung, Cheng-Chieh; Li, Yi-Jung; Chen, Yung-Chang; Chang, Ming-Yang; Yen, Tzung-Hai; Hsu, Hsiang-Hao; Wu, Mai-Szu; Phillips, Aled; Yang, Chih-Wei

2011-03-01

Leptospirosis can activate inflammatory responses through Toll-like receptors (TLRs) and may cause renal tubulointerstitial fibrosis characterized by the accumulation of extracellular matrix (ECM). We have previously demonstrated that Leptospira santorosai serovar Shermani detergent extract stimulates ECM accumulation in vitro. The aim of this study was to examine the mechanistic basis of these previous observations and, in particular, to examine the potential involvement of TLRs. The addition of serovar Shermani detergent extract led to an increase in fibronectin gene expression and production. Inhibition of TLR2 but not TLR4 expression abrogated serovar Shermani detergent extract-mediated increases in fibronectin production. This response was also blocked by the knockdown of the gene expression of the TLR2 downstream transducers myeloid differentiation factor 88 (MyD88) and tumor necrosis factor receptor-associated factor 6 (TRAF6). Serovar Shermani detergent extract also activated nuclear factor-κB, and its inhibition by curcumin-attenuated serovar Shermani detergent extract induced increases in fibronectin production. These effects were also mimicked by the specific TLR2 agonist, Pam(3)CsK(4), a response that was also abrogated by the knockdown of MyD88 and TRAF6. Similarly, the administration of live leptospires to cells also induced fibronectin production that was blocked by inhibition of TLR2 and MyD88 expression. In conclusion, serovar Shermani detergent extract can induce fibronectin production through the TLR2-associated cascade, providing evidence of an association between TLRs and leptospirosis-mediated ECM deposition.

Toxoplasma gondii infection shifts dendritic cells into an amoeboid rapid migration mode encompassing podosome dissolution, secretion of TIMP-1, and reduced proteolysis of extracellular matrix.

Science.gov (United States)

Ólafsson, Einar B; Varas-Godoy, Manuel; Barragan, Antonio

2018-03-01

Dendritic cells (DCs) infected by Toxoplasma gondii rapidly acquire a hypermigratory phenotype that promotes systemic parasite dissemination by a "Trojan horse" mechanism in mice. Recent paradigms of leukocyte migration have identified the amoeboid migration mode of DCs as particularly suited for rapid locomotion in extracellular matrix and tissues. Here, we have developed a microscopy-based high-throughput approach to assess motility and matrix degradation by Toxoplasma-challenged murine and human DCs. DCs challenged with T. gondii exhibited dependency on metalloproteinase activity for hypermotility and transmigration but, strikingly, also dramatically reduced pericellular proteolysis. Toxoplasma-challenged DCs up-regulated expression and secretion of tissue inhibitor of metalloproteinases-1 (TIMP-1) and their supernatants impaired matrix degradation by naïve DCs and by-stander DCs dose dependently. Gene silencing of TIMP-1 by short hairpin RNA restored matrix degradation activity in Toxoplasma-infected DCs. Additionally, dissolution of podosome structures in parasitised DCs coincided with abrogated matrix degradation. Toxoplasma lysates inhibited pericellular proteolysis in a MyD88-dependent fashion whereas abrogated proteolysis persevered in Toxoplasma-infected MyD88-deficient DCs. This indicated that both TLR/MyD88-dependent and TLR/MyD88-independent signalling pathways mediated podosome dissolution and the abrogated matrix degradation. We report that increased TIMP-1 secretion and cytoskeletal rearrangements encompassing podosome dissolution are features of Toxoplasma-induced hypermigration of DCs with an impact on matrix degradation. Jointly, the data highlight how an obligate intracellular parasite orchestrates key regulatory cellular processes consistent with non-proteolytic amoeboid migration of the vehicle cells that facilitate its dissemination. © 2017 John Wiley & Sons Ltd.

Role of TLR4/NADPH oxidase/ROS-activated p38 MAPK in VCAM-1 expression induced by lipopolysaccharide in human renal mesangial cells

Directory of Open Access Journals (Sweden)

Lee I-Ta

2012-11-01

Full Text Available Abstract Background In bacteria-induced glomerulonephritis, Toll-like receptor 4 (TLR4 activation by lipopolysaccharide (LPS, a key component of the outer membranes of Gram-negative bacteria can increase oxidative stress and the expression of vascular cell adhesion molecule-1 (VCAM-1, which recruits leukocytes to the glomerular mesangium. However, the mechanisms underlying VCAM-1 expression induced by LPS are still unclear in human renal mesangial cells (HRMCs. Results We demonstrated that LPS induced VCAM-1 mRNA and protein levels associated with an increase in the promoter activity of VCAM-1, determined by Western blot, RT-PCR, and promoter assay. LPS-induced responses were inhibited by transfection with siRNAs of TLR4, myeloid differentiation factor 88 (MyD88, Nox2, Nox4, p47phox, c-Src, p38 MAPK, activating transcription factor 2 (ATF2, and p300 or pretreatment with the inhibitors of reactive oxygen species (ROS, edaravone, NADPH oxidase [apocynin (APO or diphenyleneiodonium chloride (DPI], c-Src (PP1, p38 MAPK (SB202190, and p300 (GR343. LPS induced NADPH oxidase activation, ROS production, and p47phox translocation from the cytosol to the membrane, which were reduced by PP1 or c-Src siRNA. We observed that LPS induced TLR4, MyD88, c-Src, and p47phox complex formation determined by co-immunoprecipitation and Western blot. We further demonstrated that LPS stimulated ATF2 and p300 phosphorylation and complex formation via a c-Src/NADPH oxidase/ROS/p38 MAPK pathway. Up-regulation of VCAM-1 led to enhancing monocyte adhesion to HRMCs challenged with LPS, which was inhibited by siRNAs of c-Src, p47phox, p38 MAPK, ATF2, and p300 or pretreatment with an anti-VCAM-1 neutralizing antibody. Conclusions In HRMCs, LPS-induced VCAM-1 expression was, at least in part, mediated through a TLR4/MyD88/ c-Src/NADPH oxidase/ROS/p38 MAPK-dependent p300 and ATF2 pathway associated with recruitment of monocyte adhesion to kidney. Blockade of these pathways may

The temperature dependent strontium isotope fractionation (δ88/86Sr) during calcium carbonate precipitation

International Nuclear Information System (INIS)

Fietzke, J.; Eisenhauer, A.

2006-01-01

Full text: In order to study the influence of stable isotope fractionation during inorganic and biologically controlled CaCO 3 precipitation we have developed the analytical principles for the measurement of strontium (Sr) isotope fractionation. We have established a measurement protocol for the application on a MC-ICP-MS (AXIOM) using the common bracketing standard technique. The Sr-standard CRM NBS987 was used as reference material for all measurements and to calculate the Sr fractionation. Latter value is expressed by the δ-notation defined as: δ 88/86 Sr = [( 88 Sr/ 86 Sr)sample / ( 88 Sr/ 86 Sr)standard ] * 1000 -1. A first set of experiments focused on the temperature dependency of Sr-isotope fractionation. For this purpose inorganically precipitated aragonite and calcite was prepared under controlled conditions in a temperature range from 10 to 50 o C. In addition, cultured and naturally grown corals were analyzed for their δ 88/86 Sr values. Repeated measurements of IAPSO seawater standard showed a mean δ 88/86 Sr value of 0.383 ± 0.008 (2 SEM) being the isotopically heaviest material measured so far. The first results of the inorganically precipitated aragonite and the natural corals revealed a clear temperature dependency of the δ 88/86 Sr values. For inorganic aragonite the slope of this correlation is about 0.0055 permil/ o C. However, for naturally grown corals (Pavona clavus) a 6 fold steeper slope of 0.033 permil/ o C was determined. This strong temperature dependency implies the potential to use stable Sr isotopes as a new marine (paleo)temperature proxy. (author)

Conditional Selection of Genomic Alterations Dictates Cancer Evolution and Oncogenic Dependencies.

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Mina, Marco; Raynaud, Franck; Tavernari, Daniele; Battistello, Elena; Sungalee, Stephanie; Saghafinia, Sadegh; Laessle, Titouan; Sanchez-Vega, Francisco; Schultz, Nikolaus; Oricchio, Elisa; Ciriello, Giovanni

2017-08-14

Cancer evolves through the emergence and selection of molecular alterations. Cancer genome profiling has revealed that specific events are more or less likely to be co-selected, suggesting that the selection of one event depends on the others. However, the nature of these evolutionary dependencies and their impact remain unclear. Here, we designed SELECT, an algorithmic approach to systematically identify evolutionary dependencies from alteration patterns. By analyzing 6,456 genomes from multiple tumor types, we constructed a map of oncogenic dependencies associated with cellular pathways, transcriptional readouts, and therapeutic response. Finally, modeling of cancer evolution shows that alteration dependencies emerge only under conditional selection. These results provide a framework for the design of strategies to predict cancer progression and therapeutic response. Copyright © 2017 Elsevier Inc. All rights reserved.

Tubulation of class II MHC compartments is microtubule dependent and involves multiple endolysosomal membrane proteins in primary dendritic cells.

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Vyas, Jatin M; Kim, You-Me; Artavanis-Tsakonas, Katerina; Love, J Christopher; Van der Veen, Annemarthe G; Ploegh, Hidde L

2007-06-01

Immature dendritic cells (DCs) capture exogenous Ags in the periphery for eventual processing in endolysosomes. Upon maturation by TLR agonists, DCs deliver peptide-loaded class II MHC molecules from these compartments to the cell surface via long tubular structures (endolysosomal tubules). The nature and rules that govern the movement of these DC compartments are unknown. In this study, we demonstrate that the tubules contain multiple proteins including the class II MHC molecules and LAMP1, a lysosomal resident protein, as well as CD63 and CD82, members of the tetraspanin family. Endolysosomal tubules can be stained with acidotropic dyes, indicating that they are extensions of lysosomes. However, the proper trafficking of class II MHC molecules themselves is not necessary for endolysosomal tubule formation. DCs lacking MyD88 can also form endolysosomal tubules, demonstrating that MyD88-dependent TLR activation is not necessary for the formation of this compartment. Endolysosomal tubules in DCs exhibit dynamic and saltatory movement, including bidirectional travel. Measured velocities are consistent with motor-based movement along microtubules. Indeed, nocodazole causes the collapse of endolysosomal tubules. In addition to its association with microtubules, endolysosomal tubules follow the plus ends of microtubules as visualized in primary DCs expressing end binding protein 1 (EB1)-enhanced GFP.

Postoperative ileus involves interleukin-1 receptor signaling in enteric glia.

Science.gov (United States)

Stoffels, Burkhard; Hupa, Kristof Johannes; Snoek, Susanne A; van Bree, Sjoerd; Stein, Kathy; Schwandt, Timo; Vilz, Tim O; Lysson, Mariola; Veer, Cornelis Van't; Kummer, Markus P; Hornung, Veit; Kalff, Joerg C; de Jonge, Wouter J; Wehner, Sven

2014-01-01

Postoperative ileus (POI) is a common consequence of abdominal surgery that increases the risk of postoperative complications and morbidity. We investigated the cellular mechanisms and immune responses involved in the pathogenesis of POI. We studied a mouse model of POI in which intestinal manipulation leads to inflammation of the muscularis externa and disrupts motility. We used C57BL/6 (control) mice as well as mice deficient in Toll-like receptors (TLRs) and cytokine signaling components (TLR-2(-/-), TLR-4(-/-), TLR-2/4(-/-), MyD88(-/-), MyD88/TLR adaptor molecule 1(-/-), interleukin-1 receptor [IL-1R1](-/-), and interleukin (IL)-18(-/-) mice). Bone marrow transplantation experiments were performed to determine which cytokine receptors and cell types are involved in the pathogenesis of POI. Development of POI did not require TLRs 2, 4, or 9 or MyD88/TLR adaptor molecule 2 but did require MyD88, indicating a role for IL-1R1. IL-1R1(-/-) mice did not develop POI; however, mice deficient in IL-18, which also signals via MyD88, developed POI. Mice given injections of an IL-1 receptor antagonist (anakinra) or antibodies to deplete IL-1α and IL-1β before intestinal manipulation were protected from POI. Induction of POI activated the inflammasome in muscularis externa tissues of C57BL6 mice, and IL-1α and IL-1β were released in ex vivo organ bath cultures. In bone marrow transplantation experiments, the development of POI required activation of IL-1 receptor in nonhematopoietic cells. IL-1R1 was expressed by enteric glial cells in the myenteric plexus layer, and cultured primary enteric glia cells expressed IL-6 and the chemokine monocyte chemotactic protein 1 in response to IL-1β stimulation. Immunohistochemical analysis of human small bowel tissue samples confirmed expression of IL-1R1 in the ganglia of the myenteric plexus. IL-1 signaling, via IL-1R1 and MyD88, is required for development of POI after intestinal manipulation in mice. Agents that interfere with

Tubulation of Class II MHC Compartments Is Microtubule Dependent and Involves Multiple Endolysosomal Membrane Proteins in Primary Dendritic Cells1

Science.gov (United States)

Vyas, Jatin M.; Kim, You-Me; Artavanis-Tsakonas, Katerina; Love, J. Christopher; Van der Veen, Annemarthe G.; Ploegh, Hidde L.

2009-01-01

Immature dendritic cells (DCs) capture exogenous Ags in the periphery for eventual processing in endolysosomes. Upon maturation by TLR agonists, DCs deliver peptide-loaded class II MHC molecules from these compartments to the cell surface via long tubular structures (endolysosomal tubules). The nature and rules that govern the movement of these DC compartments are unknown. In this study, we demonstrate that the tubules contain multiple proteins including the class II MHC molecules and LAMP1, a lysosomal resident protein, as well as CD63 and CD82, members of the tetraspanin family. Endolysosomal tubules can be stained with acidotropic dyes, indicating that they are extensions of lysosomes. However, the proper trafficking of class II MHC molecules themselves is not necessary for endolysosomal tubule formation. DCs lacking MyD88 can also form endolysosomal tubules, demonstrating that MyD88-dependent TLR activation is not necessary for the formation of this compartment. Endolysosomal tubules in DCs exhibit dynamic and saltatory movement, including bidirectional travel. Measured velocities are consistent with motor-based movement along microtubules. Indeed, nocodazole causes the collapse of endolysosomal tubules. In addition to its association with microtubules, endolysosomal tubules follow the plus ends of microtubules as visualized in primary DCs expressing end binding protein 1 (EB1)-enhanced GFP. PMID:17513769

Hypoxia-inducible factor-1α mediates the toll-like receptor 4 signaling pathway leading to anti-tumor effects in human hepatocellular carcinoma cells under hypoxic conditions.

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Zhang, Xiaoyu; Li, Shuchen; Li, Mingrong; Huang, Haiying; Li, Jingyuan; Zhou, Changwei

2016-08-01

Hypoxia-inducible factor-1α (HIF-1α) and toll-like receptor 4 (TLR4) are involved in numerous mechanisms of cancer biology, including cell proliferation and survival; however the interaction of the two factors under hypoxic conditions remains unclear. The present study investigated the in vitro mechanism that results in the suppression of tumor cell growth and cellular functions when HIF-1α is silenced. In the present study, the human hepatocellular carcinoma HepG2 cell line was transfected with short hairpin RNA (shRNA) against HIF-1α and cultured under hypoxic conditions (1% O 2 for 24 h). The expression of HIF-1α and various growth factors, including epidermal growth factor (EGF), hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2), were examined using quantitative polymerase chain reaction and immunoblotting. Tumor growth was measured using a Cell Counting Kit-8 assay and tumor activity was measured using tumor cell invasion and migration assays. Lipopolysaccharide and TAK-242 were used to activate and inhibit TLR4, respectively, to observe the role of TLR4 in the HIF-1α silenced tumor cells. The expression of TLR4 signaling pathway associates, including myeloid differentiation primary response gene 88 (MyD88), apoptosis signal-regulating kinase 1 (ASK1), p38 mitogen-activated protein kinases and HIF-1α, were analyzed by western blot assay. Under hypoxic conditions, silencing of HIF-1α expression suppressed tumor cell growth and regulated the expression of tumor growth-associated genes, including EGF, HGF, VEGF and FG2. Suppression of tumor cell invasion and migration was also observed in the HIF-1α silenced HepG2 cell line. In addition, TLR4 was identified to be involved in HIF-1α and MyD88 accumulation, and activation of ASK1 and p38 were demonstrated to be critical for TLR4-mediated HIF-1α pathway. In conclusion, silencing of HIF-1α expression may induce anti-tumor effects under hypoxic

Mechanisms Underlying the Immune Response Generated by an Oral Vibrio cholerae Vaccine

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Danylo Sirskyj

2016-07-01

Full Text Available Mechanistic details underlying the resulting protective immune response generated by mucosal vaccines remain largely unknown. We investigated the involvement of Toll-like receptor signaling in the induction of humoral immune responses following oral immunization with Dukoral, comparing wild type mice with TLR-2-, TLR-4-, MyD88- and Trif-deficient mice. Although all groups generated similar levels of IgG antibodies, the proliferation of CD4+ T-cells in response to V. cholerae was shown to be mediated via MyD88/TLR signaling, and independently of Trif signaling. The results demonstrate differential requirements for generation of immune responses. These results also suggest that TLR pathways may be modulators of the quality of immune response elicited by the Dukoral vaccine. Determining the critical signaling pathways involved in the induction of immune response to this vaccine would be beneficial, and could contribute to more precisely-designed versions of other oral vaccines in the future.

ST2 negatively regulates TLR2 signaling, but is not required for bacterial lipoprotein-induced tolerance.

LENUS (Irish Health Repository)

Liu, Jinghua

2010-05-15

Activation of TLR signaling is critical for host innate immunity against bacterial infection. Previous studies reported that the ST2 receptor, a member of the Toll\\/IL-1 receptor superfamily, functions as a negative regulator of TLR4 signaling and maintains LPS tolerance. However, it is undetermined whether ST2 negatively regulates TLR2 signaling and furthermore, whether a TLR2 agonist, bacterial lipoprotein (BLP)-induced tolerance is dependent on ST2. In this study, we show that BLP stimulation-induced production of proinflammatory cytokines and immunocomplex formation of TLR2-MyD88 and MyD88-IL-1R-associated kinase (IRAK) were significantly enhanced in ST2-deficient macrophages compared with those in wild-type controls. Furthermore, overexpression of ST2 dose-dependently attenuated BLP-induced NF-kappaB activation, suggesting a negative regulatory role of ST2 in TLR2 signaling. A moderate but significantly attenuated production of TNF-alpha and IL-6 on a second BLP stimulation was observed in BLP-pretreated, ST2-deficient macrophages, which is associated with substantially reduced IRAK-1 protein expression and downregulated TLR2-MyD88 and MyD88-IRAK immunocomplex formation. ST2-deficient mice, when pretreated with a nonlethal dose of BLP, benefitted from an improved survival against a subsequent lethal BLP challenge, indicating BLP tolerance develops in the absence of the ST2 receptor. Taken together, our results demonstrate that ST2 acts as a negative regulator of TLR2 signaling, but is not required for BLP-induced tolerance.

The role of Toll like receptors (TLRs) in the immunopathogenesis of pregnancy associated malaria.

OpenAIRE

Leandro Gustavo da Silva

2011-01-01

A malária asociada à gavidez pode gerar complicações para a mãe e para o feto. Receptores do tipo Toll (TLR) TLR2, TLR4 e TLR9, podem reconhecer componentes do Plasmódio. Estes receptores sinalizam via proteína MyD88. Contudo existem poucos dados sobre os TLR na malária placentária. Assim, o objetivo desse trabalho foi estudar o papel dos TLR2, 4, 9 e da MyD88 na malária placentária. Dentre fêmeas C57BL/6, TLR2-/-, TLR9-/- e MyD88-/-, a linhagem MyD88-/- apresentou maiores níveis de parasitem...

Using house dust extracts to understand the immunostimulatory activities of living environments.

Science.gov (United States)

Batzer, Glenda; Lam, Diane P; Paulus, Petra; Boasen, Jared; Ng, Nicholas; Horner, Anthony A

2007-01-01

Laboratory and epidemiological studies have provided indirect but compelling evidence that toll-like receptor (TLR) signaling pathways play an important role in host responsiveness to ambient immunostimulatory factors. Nonetheless, direct evidence is limited. This paper will present our experience investigating the innate immunostimulatory activities of sterile house dust extracts (HDEs). In initial studies, bone marrow derived dendritic cells (BMDDCs) were cultured with HDEs, and cytokine production and co-stimulatory molecule expression were evaluated. In additional experiments, the TLR dependence of these responses was determined. HDEs induced concentration-dependent BMDDC activation. Moreover, the relative bioactivities of HDEs correlated with their endotoxin content. Finally, HDE-mediated responses were found to be partially dependent on TLR2, TLR4, and TLR9 and almost completely dependent on MyD88. These investigations provide the first direct evidence that TLR signaling pathways play a key role in innate responsiveness to non-infectious factors ubiquitous in living environments.

Presence of Tube isoforms in Litopenaeus vannamei suggests various regulatory patterns of signal transduction in invertebrate NF-κB pathway.

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Li, Chaozheng; Chen, Yixiao; Weng, Shaoping; Li, Sedong; Zuo, Hongliang; Yu, Xiaoqiang; Li, Haoyang; He, Jianguo; Xu, Xiaopeng

2014-02-01

The toll-like receptor (TLR)/NF-κB signaling pathways play critical roles in the innate immune system. The intracellular signal transduction of most TLR pathways in invertebrate cells is triggered by formation of a heterotrimeric complex composed of MyD88, Tube and Pelle. In this study, we identified a Litopenaeus vannamei Pelle (LvPelle) and an isoform of L. vannamei Tube (LvTube) designated as LvTube-1. The interactions among LvPelle, LvTube/LvTube-1 and LvMyD88/LvMyD88-1 were elucidated and their functions during pathogen infections were investigated. Knockdowns of LvPelle and LvTube/LvTube-1 using RNAi strategy led to higher mortalities of shrimps during Vibrio parahemolyticus infection, and could reduce the genome copy number of white spot syndrome virus (WSSV) in the infected muscle tissue but did not affect the mortality caused by WSSV infection. The effects of LvPelle and LvTube/LvTube-1 on promoters containing NF-κB binding motifs were analyzed by dual-luciferase reporter assays and the results demonstrated that LvTube-1 could activate the NF-κB activity to significantly higher level than LvTube did. Moreover, tissue distributions of LvTube and LvTube-1 mRNAs and their expression profiles during pathogen and immune stimulant challenges were different, indicating that they could play different roles in immune responses. This is the first report of Tube isoforms in invertebrates. Together with our previous study on LvMyD88 isoforms, our results suggest that various isoforms of adaptor components may be involved in various regulatory patterns of signal transduction in invertebrate TLR/NF-κB pathway and this could be a strategy adopted by invertebrates to modulate immune responses. Copyright © 2013 Elsevier Ltd. All rights reserved.

Suppression of Adaptive Immune Cell Activation Does Not Alter Innate Immune Adipose Inflammation or Insulin Resistance in Obesity.

Directory of Open Access Journals (Sweden)

Manikandan Subramanian

Full Text Available Obesity-induced inflammation in visceral adipose tissue (VAT is a major contributor to insulin resistance and type 2 diabetes. Whereas innate immune cells, notably macrophages, contribute to visceral adipose tissue (VAT inflammation and insulin resistance, the role of adaptive immunity is less well defined. To address this critical gap, we used a model in which endogenous activation of T cells was suppressed in obese mice by blocking MyD88-mediated maturation of CD11c+ antigen-presenting cells. VAT CD11c+ cells from Cd11cCre+Myd88fl/fl vs. control Myd88fl/fl mice were defective in activating T cells in vitro, and VAT T and B cell activation was markedly reduced in Cd11cCre+Myd88fl/fl obese mice. However, neither macrophage-mediated VAT inflammation nor systemic inflammation were altered in Cd11cCre+Myd88fl/fl mice, thereby enabling a focused analysis on adaptive immunity. Unexpectedly, fasting blood glucose, plasma insulin, and the glucose response to glucose and insulin were completely unaltered in Cd11cCre+Myd88fl/fl vs. control obese mice. Thus, CD11c+ cells activate VAT T and B cells in obese mice, but suppression of this process does not have a discernible effect on macrophage-mediated VAT inflammation or systemic glucose homeostasis.

IgD class switching is initiated by microbiota and limited to mucosa-associated lymphoid tissue in mice

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Choi, Jin Huk; Wang, Kuan-wen; Zhang, Duanwu; Zhan, Xiaowei; Wang, Tao; Bu, Chun-Hui; Behrendt, Cassie L.; Zeng, Ming; Wang, Ying; Misawa, Takuma; Li, Xiaohong; Tang, Miao; Zhan, Xiaoming; Scott, Lindsay; Hildebrand, Sara; Murray, Anne R.; Moresco, Eva Marie Y.; Hooper, Lora V.; Beutler, Bruce

2017-01-01

Class-switch recombination (CSR) alters the Ig isotype to diversify antibody effector functions. IgD CSR is a rare event, and its regulation is poorly understood. We report that deficiency of 53BP1, a DNA damage-response protein, caused age-dependent overproduction of secreted IgD resulting from increased IgD CSR exclusively within B cells of mucosa-associated lymphoid tissues. IgD overproduction was dependent on activation-induced cytidine deaminase, hematopoietic MyD88 expression, and an intact microbiome, against which circulating IgD, but not IgM, was reactive. IgD CSR occurred via both alternative nonhomologous end-joining and homologous recombination pathways. Microbiota-dependent IgD CSR also was detected in nasal-associated lymphoid tissue of WT mice. These results identify a pathway, present in WT mice and hyperactivated in 53BP1-deficient mice, by which microbiota signal via Toll-like receptors to elicit IgD CSR. PMID:28137874

IgD class switching is initiated by microbiota and limited to mucosa-associated lymphoid tissue in mice.

Science.gov (United States)

Choi, Jin Huk; Wang, Kuan-Wen; Zhang, Duanwu; Zhan, Xiaowei; Wang, Tao; Bu, Chun-Hui; Behrendt, Cassie L; Zeng, Ming; Wang, Ying; Misawa, Takuma; Li, Xiaohong; Tang, Miao; Zhan, Xiaoming; Scott, Lindsay; Hildebrand, Sara; Murray, Anne R; Moresco, Eva Marie Y; Hooper, Lora V; Beutler, Bruce

2017-02-14

Class-switch recombination (CSR) alters the Ig isotype to diversify antibody effector functions. IgD CSR is a rare event, and its regulation is poorly understood. We report that deficiency of 53BP1, a DNA damage-response protein, caused age-dependent overproduction of secreted IgD resulting from increased IgD CSR exclusively within B cells of mucosa-associated lymphoid tissues. IgD overproduction was dependent on activation-induced cytidine deaminase, hematopoietic MyD88 expression, and an intact microbiome, against which circulating IgD, but not IgM, was reactive. IgD CSR occurred via both alternative nonhomologous end-joining and homologous recombination pathways. Microbiota-dependent IgD CSR also was detected in nasal-associated lymphoid tissue of WT mice. These results identify a pathway, present in WT mice and hyperactivated in 53BP1-deficient mice, by which microbiota signal via Toll-like receptors to elicit IgD CSR.

[TLR2 modulates Staphylococcus aureus-induced inflammatory response and autophagy in macrophages through PI3K signaling pathway].

Science.gov (United States)

Li, Shuai; Fang, Lei; Wang, Jiong; Liu, Rongyu

2017-09-01

Objective To investigate the molecular mechanisms of Toll-like receptor 2 (TLR2) taking part in inflammatory response in Staphylococcus aureus (SA)-induced asthma. Methods We established the cell inflammatory response model through stimulating mouse RAW264.7 macrophages with SA. The TLR2, myeloid differentiation factor 88 (MyD88), phosphoinositide-3 kinase (PI3K), nuclear factor κBp65 (NF-κBp65), phospho-NF-κBp65, beclin-1 and microtubule-associated protein 1 light chain 3B (LC3B) were detected by Western blot analysis after treatment with TLR2 small interfering RNA (siRNA) and 3-methyladenine (3-MA), and the tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) were determined by ELISA. In addition, the number of autolysosomes was observed by the laser scanning confocal microscope. Results SA-stimulated macrophages activated various signaling pathways including TLR2. TLR2 siRNA markedly repressed the expressions of PI3K, phospho-NF-κBp65, the autophagy protein beclin-1 and LC3B as well as the number of autolysosomes and the production of TNF- and IL-6. We also demonstrated that 3-MA had the same effect on autophagy and inflammation as TLR2 siRNA did. Conclusion TLR2 modulates SA-induced inflammatory response and autophagy in macrophages through PI3K signaling pathway.

The TLR3/TICAM-1 signal constitutively controls spontaneous polyposis through suppression of c-Myc in Apc Min/+ mice.

Science.gov (United States)

Ono, Junya; Shime, Hiroaki; Takaki, Hiromi; Takashima, Ken; Funami, Kenji; Yoshida, Sumito; Takeda, Yohei; Matsumoto, Misako; Kasahara, Masanori; Seya, Tsukasa

2017-10-17

Intestinal tumorigenesis is promoted by myeloid differentiation primary response gene 88 (MyD88) activation in response to the components of microbiota in Apc Min/+ mice. Microbiota also contains double-stranded RNA (dsRNA), a ligand for TLR3, which activates the toll-like receptor adaptor molecule 1 (TICAM-1, also known as TRIF) pathway. We established Apc Min/+ Ticam1 -/- mice and their survival was compared to survival of Apc Min/+ Myd88 -/- and wild-type (WT) mice. The properties of polyps were investigated using immunofluorescence staining and RT-PCR analysis. We demonstrate that TICAM-1 is essential for suppression of polyp formation in Apc Min/+ mice. TICAM-1 knockout resulted in shorter survival of mice compared to WT mice or mice with knockout of MyD88 in the Apc Min/+ background. Polyps were more frequently formed in the distal intestine of Apc Min/+ Ticam1 -/- mice than in Apc Min/+ mice. Infiltration of immune cells such as CD11b + and CD8α + cells into the polyps was detected histologically. CD11b and CD8α mRNAs were increased in polyps of Apc Min/+ Ticam1 -/- mice compared to Apc Min/+ mice. Gene expression of inducible nitric oxide synthase (iNOS), interferon (IFN)-γ, CXCL9 and IL-12p40 was increased in polyps of Apc Min/+ Ticam1 -/- mice. mRNA and protein expression of c-Myc, a critical transcription factor for inflammation-associated polyposis, were increased in polyps of Apc Min/+ Ticam1 -/- mice. A Lactobacillus strain producing dsRNA was detected in feces of Apc Min/+ mice. These results imply that the TLR3/TICAM-1 pathway inhibits polyposis through suppression of c-Myc expression and supports long survival in Apc Min/+ mice.

Combining MOD10A1 and MYD10A1 Images For Snow Cover Area Monitoring

Science.gov (United States)

Tekeli, A. E.

2008-12-01

MOD10A1 and MYD10A1 daily snow cover maps at 500 m resolution are available from MODIS sensors on Terra and Aqua satellites. Aqua obtains the image of same region approximately three hours after Terra over Turkey region. MODIS is an optic sensor and cloud cover degrades the usability of derived snow cover maps. Moreover, spectral similarity between clouds and snow complicates their separability in visible imagery. Fortunately, dynamic behavior of clouds enables their discrimination from snow stationary on the surface. Combined use of MOD10A1 and MYD10A1 images mostly reduces the cloud cover present in one image alone and provides better representation of surface snow cover. Comparison of merged images with in situ data indicated higher hit ratios. The individual comparison of MOD10A1 and MYD10A1 images with ground data each yielded 31% hit ratio whereas, the merged images provided 38%. One-day shifts in comparisons increased hit ratios to 52 % and 46% whereas and two-day shifts gave 77 % and 79 % for MOD10A1 and MYD10A1 respectively. Merged maps yielded 54% and 83% for one and two day shifts. The improvement provided by the merging technique is found to be 7% for the present day, 7 % for one- day and 5% for two-day shifts for the whole season. Monthly decomposition resulted 25% improvement as the maximum. The snow cover product obtained by merging Terra and Aqua satellites provided higher hit ratios, as expected.

A novel missense mutation in CCDC88C activates the JNK pathway and causes a dominant form of spinocerebellar ataxia.

Science.gov (United States)

Tsoi, Ho; Yu, Allen C S; Chen, Zhefan S; Ng, Nelson K N; Chan, Anne Y Y; Yuen, Liz Y P; Abrigo, Jill M; Tsang, Suk Ying; Tsui, Stephen K W; Tong, Tony M F; Lo, Ivan F M; Lam, Stephen T S; Mok, Vincent C T; Wong, Lawrence K S; Ngo, Jacky C K; Lau, Kwok-Fai; Chan, Ting-Fung; Chan, H Y Edwin

2014-09-01

Spinocerebellar ataxias (SCAs) are a group of clinically and genetically diverse and autosomal-dominant disorders characterised by neurological deficits in the cerebellum. At present, there is no cure for SCAs. Of the different distinct subtypes of autosomal-dominant SCAs identified to date, causative genes for only a fraction of them are currently known. In this study, we investigated the cause of an autosomal-dominant SCA phenotype in a family that exhibits cerebellar ataxia and pontocerebellar atrophy along with a global reduction in brain volume. Whole-exome analysis revealed a missense mutation c.G1391A (p.R464H) in the coding region of the coiled-coil domain containing 88C (CCDC88C) gene in all affected individuals. Functional studies showed that the mutant form of CCDC88C activates the c-Jun N-terminal kinase (JNK) pathway, induces caspase 3 cleavage and triggers apoptosis. This study expands our understanding of the cause of autosomal-dominant SCAs, a group of heterogeneous congenital neurological conditions in humans, and unveils a link between the JNK stress pathway and cerebellar atrophy. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Innate Immune Signalling Genetics of Pain, Cognitive Dysfunction and Sickness Symptoms in Cancer Pain Patients Treated with Transdermal Fentanyl

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Barratt, Daniel T.; Klepstad, Pål; Dale, Ola; Kaasa, Stein; Somogyi, Andrew A.

2015-01-01

Common adverse symptoms of cancer and chemotherapy are a major health burden; chief among these is pain, with opioids including transdermal fentanyl the mainstay of treatment. Innate immune activation has been implicated generally in pain, opioid analgesia, cognitive dysfunction, and sickness type symptoms reported by cancer patients. We aimed to determine if genetic polymorphisms in neuroimmune activation pathways alter the serum fentanyl concentration-response relationships for pain control, cognitive dysfunction, and other adverse symptoms, in cancer pain patients. Cancer pain patients (468) receiving transdermal fentanyl were genotyped for 31 single nucleotide polymorphisms in 19 genes: CASP1, BDNF, CRP, LY96, IL6, IL1B, TGFB1, TNF, IL10, IL2, TLR2, TLR4, MYD88, IL6R, OPRM1, ARRB2, COMT, STAT6 and ABCB1. Lasso and backward stepwise generalised linear regression were used to identify non-genetic and genetic predictors, respectively, of pain control (average Brief Pain Inventory fentanyl concentrations did not predict between-patient variability in these outcomes, nor did genetic factors predict pain control, sickness response or opioid adverse event complaint. Carriers of the MYD88 rs6853 variant were half as likely to have cognitive dysfunction (11/111) than wild-type patients (69/325), with a relative risk of 0.45 (95% CI: 0.27 to 0.76) when accounting for major non-genetic predictors (age, Karnofsky functional score). This supports the involvement of innate immune signalling in cognitive dysfunction, and identifies MyD88 signalling pathways as a potential focus for predicting and reducing the burden of cognitive dysfunction in cancer pain patients. PMID:26332828

Comprehensive logic based analyses of Toll-like receptor 4 signal transduction pathway.

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Mahesh Kumar Padwal

Full Text Available Among the 13 TLRs in the vertebrate systems, only TLR4 utilizes both Myeloid differentiation factor 88 (MyD88 and Toll/Interleukin-1 receptor (TIR-domain-containing adapter interferon-β-inducing Factor (TRIF adaptors to transduce signals triggering host-protective immune responses. Earlier studies on the pathway combined various experimental data in the form of one comprehensive map of TLR signaling. But in the absence of adequate kinetic parameters quantitative mathematical models that reveal emerging systems level properties and dynamic inter-regulation among the kinases/phosphatases of the TLR4 network are not yet available. So, here we used reaction stoichiometry-based and parameter independent logical modeling formalism to build the TLR4 signaling network model that captured the feedback regulations, interdependencies between signaling kinases and phosphatases and the outcome of simulated infections. The analyses of the TLR4 signaling network revealed 360 feedback loops, 157 negative and 203 positive; of which, 334 loops had the phosphatase PP1 as an essential component. The network elements' interdependency (positive or negative dependencies in perturbation conditions such as the phosphatase knockout conditions revealed interdependencies between the dual-specific phosphatases MKP-1 and MKP-3 and the kinases in MAPK modules and the role of PP2A in the auto-regulation of Calmodulin kinase-II. Our simulations under the specific kinase or phosphatase gene-deficiency or inhibition conditions corroborated with several previously reported experimental data. The simulations to mimic Yersinia pestis and E. coli infections identified the key perturbation in the network and potential drug targets. Thus, our analyses of TLR4 signaling highlights the role of phosphatases as key regulatory factors in determining the global interdependencies among the network elements; uncovers novel signaling connections; identifies potential drug targets for

Roles of Toll-like receptors in allogeneic islet transplantation.

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Ro, Han; Hong, Juho; Kim, Beom Seok; Lee, Eun Won; Kim, Myung-Gyu; Han, Kyu Hyun; Yeom, Hye-Jung; Lee, Eun Mi; Jeong, Jong Cheol; Oh, Kook-Hwan; Ahn, Curie; Yang, Jaeseok

2012-11-27

Toll-like receptors (TLRs) are involved in the rejection of solid organ allografts. However, the roles of TLRs in islets are still controversial. We investigated the roles of TLRs in donor islets together with those in recipients in allogeneic islet transplantation. To assess the roles of TLRs in either donor islets or recipients, allogeneic islet transplantation was performed using myeloid differentiation factor 88 (MyD88)-knockout (KO), TLR4-KO, or Toll/interleukin-1 receptor domain-containing adaptor-inducing interferon-β (TRIF)-KO mice. Both polyriboinosinic polyribocytidylic acid and lipopolysaccharide (LPS) stimulation induced the mRNA expression of regulated and normal T cell expressed and secreted, interferon-γ-inducible protein-10, monocyte chemotactic protein-1, interleukin-8, and inducible nitric oxide synthase in murine islets, whereas the induction was attenuated in TRIF-KO, interferon-β promoter stimulator-1-KO, and TLR4-KO mice. When islets from MyD88-KO, TLR4-KO, or TRIF-KO C57BL/6 mice were transplanted to BALB/c recipients, graft survival was not better than that of wild-type (WT) islets. However, the survival of the MyD88-KO islet allograft was significantly prolonged when combined with anti-CD40L. In parallel, LPS stimulation in donor islets interfered with anti-CD40L blockade-mediated long-term survival of islet allografts in TLR4-KO recipients. LPS stimulation increased the perigraft infiltration of both T cells and macrophages. Then again, when islets from WT BALB/c mice were transplanted to MyD88-KO, TRIF-KO, or WT C57BL/6 mice, there was no difference in graft survival, although some of the MyD88-KO recipients obtained long-term graft survival. However, anti-CD40L prolonged graft survival significantly in MyD88-KO recipients. The absence of MyD88 in either donors or recipients decreased the perigraft infiltration of inflammatory cells when combined with anti-CD40L. TLRs in both donor islets and recipients are involved in islet allograft

Genetics Home Reference: MyD88 deficiency

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... C, Sirvent N, de Ricaud D, Camcioglu Y, Vasconcelos J, Guedes M, Vitor AB, Rodrigo C, Almazán ... Chrabieh M, Mustapha IB, Ghandil P, Camcioglu Y, Vasconcelos J, Sirvent N, Guedes M, Vitor AB, Herrero- ...

Novel synthetic Medea selfish genetic elements drive population replacement in Drosophila; a theoretical exploration of Medea-dependent population suppression.

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Akbari, Omar S; Chen, Chun-Hong; Marshall, John M; Huang, Haixia; Antoshechkin, Igor; Hay, Bruce A

2014-12-19

Insects act as vectors for diseases of plants, animals, and humans. Replacement of wild insect populations with genetically modified individuals unable to transmit disease provides a potentially self-perpetuating method of disease prevention. Population replacement requires a gene drive mechanism in order to spread linked genes mediating disease refractoriness through wild populations. We previously reported the creation of synthetic Medea selfish genetic elements able to drive population replacement in Drosophila. These elements use microRNA-mediated silencing of myd88, a maternally expressed gene required for embryonic dorso-ventral pattern formation, coupled with early zygotic expression of a rescuing transgene, to bring about gene drive. Medea elements that work through additional mechanisms are needed in order to be able to carry out cycles of population replacement and/or remove existing transgenes from the population, using second-generation elements that spread while driving first-generation elements out of the population. Here we report the synthesis and population genetic behavior of two new synthetic Medea elements that drive population replacement through manipulation of signaling pathways involved in cellular blastoderm formation or Notch signaling, demonstrating that in Drosophila Medea elements can be generated through manipulation of diverse signaling pathways. We also describe the mRNA and small RNA changes in ovaries and early embryos associated from Medea-bearing females. Finally, we use modeling to illustrate how Medea elements carrying genes that result in diapause-dependent female lethality could be used to bring about population suppression.

Novel Synthetic Medea selfish genetic elements drive population replacement in Drosophila, and a theoretical exploration of Medea-dependent population suppression

Science.gov (United States)

Akbari, Omar S.; Chen, Chun-Hong; Marshall, John M.; Huang, Haixia; Antoshechkin, Igor; Hay, Bruce A.

2013-01-01

Insects act as vectors for diseases of plants, animals and humans. Replacement of wild insect populations with genetically modified individuals unable to transmit disease provides a potentially self-perpetuating method of disease prevention. Population replacement requires a gene drive mechanism in order to spread linked genes mediating disease refractoriness through wild populations. We previously reported the creation of synthetic Medea selfish genetic elements able to drive population replacement in Drosophila. These elements use microRNA-mediated silencing of myd88, a maternally expressed gene required for embryonic dorso-ventral pattern formation, coupled with early zygotic expression of a rescuing transgene, to bring about gene drive. Medea elements that work through additional mechanisms are needed in order to be able to carry out cycles of population replacement and/or remove existing transgenes from the population, using second-generation elements that spread while driving first-generation elements out of the population. Here we report the synthesis and population genetic behavior of two new synthetic Medea elements that drive population replacement through manipulation of signaling pathways involved in cellular blastoderm formation or Notch signaling, demonstrating that in Drosophila Medea elements can be generated through manipulation of diverse signaling pathways. We also describe the mRNA and small RNA changes in ovaries and early embryos associated from Medea-bearing females. Finally, we use modeling to illustrate how Medea elements carrying genes that result in diapause-dependent female lethality could be used to bring about population suppression. PMID:23654248

Capsular Polysaccharide is a Main Component of Mycoplasma ovipneumoniae in the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells

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Zhongjia Jiang

2017-01-01

Full Text Available Mycoplasma ovipneumoniae (M. ovipneumoniae is characterized as an etiological agent of primary atypical pneumonia that specifically infects sheep and goat. In an attempt to better understand the pathogen-host interaction between the invading M. ovipneumoniae and airway epithelial cells, we investigated the host inflammatory responses against capsular polysaccharide (designated as CPS of M. ovipneumoniae using sheep bronchial epithelial cells cultured in an air-liquid interface (ALI model. Results showed that CPS derived from M. ovipneumoniae could activate toll-like receptor- (TLR- mediated inflammatory responses, along with an elevated expression of nuclear factor kappa B (NF-κB, activator protein-1 (AP-1, and interferon regulatory factor 3 (IRF3 as well as various inflammatory-associated mediators, representatively including proinflammatory cytokines, such as IL1β, TNFα, and IL8, and anti-inflammatory cytokines such as IL10 and TGFβ of TLR signaling cascade. Mechanistically, the CPS-induced inflammation was TLR initiated and was mediated by activations of both MyD88-dependent and MyD88-independent signaling pathways. Of importance, a blockage of CPS with specific antibody led a significant reduction of M. ovipneumoniae-induced inflammatory responses in sheep bronchial epithelial cells. These results suggested that CPS is a key virulent component of M. ovipneumoniae, which may play a crucial role in the inflammatory response induced by M. ovipneumoniae infections.

Capsular Polysaccharide is a Main Component of Mycoplasma ovipneumoniae in the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells.

Science.gov (United States)

Jiang, Zhongjia; Song, Fuyang; Li, Yanan; Xue, Di; Deng, Guangcun; Li, Min; Liu, Xiaoming; Wang, Yujiong

2017-01-01

Mycoplasma ovipneumoniae ( M. ovipneumoniae ) is characterized as an etiological agent of primary atypical pneumonia that specifically infects sheep and goat. In an attempt to better understand the pathogen-host interaction between the invading M. ovipneumoniae and airway epithelial cells, we investigated the host inflammatory responses against capsular polysaccharide (designated as CPS) of M. ovipneumoniae using sheep bronchial epithelial cells cultured in an air-liquid interface (ALI) model. Results showed that CPS derived from M. ovipneumoniae could activate toll-like receptor- (TLR-) mediated inflammatory responses, along with an elevated expression of nuclear factor kappa B (NF- κ B), activator protein-1 (AP-1), and interferon regulatory factor 3 (IRF3) as well as various inflammatory-associated mediators, representatively including proinflammatory cytokines, such as IL1 β , TNF α , and IL8, and anti-inflammatory cytokines such as IL10 and TGF β of TLR signaling cascade. Mechanistically, the CPS-induced inflammation was TLR initiated and was mediated by activations of both MyD88-dependent and MyD88-independent signaling pathways. Of importance, a blockage of CPS with specific antibody led a significant reduction of M. ovipneumoniae -induced inflammatory responses in sheep bronchial epithelial cells. These results suggested that CPS is a key virulent component of M. ovipneumoniae , which may play a crucial role in the inflammatory response induced by M. ovipneumoniae infections.

Selective recognition of 6-mercaptopurine based on luminescent metal-organic frameworks Fe-MIL-88NH₂.

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Sun, Zhengjuan; Liu, Yali; Li, Yuanfang

2015-03-15

A novel and rapid spectrofluorometry method for the recognition of 6-mercaptopurine (6-MP) has been developed based on luminescent metal-organic frameworks Fe-MIL-88NH2 as fluorescent probe. The strong fluorescence of Fe-MIL-88NH2 at 430 nm could be quenched by 6-MP directly, and the Fe-MIL-88NH2 shows high selectivity for 6-MP compared to other thiol-containing amino acids such as homocysteine (Hcy), cysteine (Cys), glutathione (GSH), etc. Under optimal conditions, the relative fluorescence intensity was linearly proportional to the concentration of 6-MP in the range of 5-600 μM with the detection limit at 1.17 μM (S/N=3). Furthermore, the present approach has been successfully applied to the determination of 6-MP in human serum samples. The possible fluorescence quenching mechanism has also been investigated, where it is revealed that the quenching was attributed to competition of absorption of the light source energy as well as electron transfer between Fe-MIL-88NH2 and 6-MP. Copyright © 2014 Elsevier B.V. All rights reserved.

Junction detection and pathway selection

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Peck, Alex N.; Lim, Willie Y.; Breul, Harry T.

1992-02-01

The ability to detect junctions and make choices among the possible pathways is important for autonomous navigation. In our script-based navigation approach where a journey is specified as a script of high-level instructions, actions are frequently referenced to junctions, e.g., `turn left at the intersection.' In order for the robot to carry out these kind of instructions, it must be able (1) to detect an intersection (i.e., an intersection of pathways), (2) know that there are several possible pathways it can take, and (3) pick the pathway consistent with the high level instruction. In this paper we describe our implementation of the ability to detect junctions in an indoor environment, such as corners, T-junctions and intersections, using sonar. Our approach uses a combination of partial scan of the local environment and recognition of sonar signatures of certain features of the junctions. In the case where the environment is known, we use additional sensor information (such as compass bearings) to help recognize the specific junction. In general, once a junction is detected and its type known, the number of possible pathways can be deduced and the correct pathway selected. Then the appropriate behavior for negotiating the junction is activated.

Innate Immune Signalling Genetics of Pain, Cognitive Dysfunction and Sickness Symptoms in Cancer Pain Patients Treated with Transdermal Fentanyl.

Directory of Open Access Journals (Sweden)

Daniel T Barratt

Full Text Available Common adverse symptoms of cancer and chemotherapy are a major health burden; chief among these is pain, with opioids including transdermal fentanyl the mainstay of treatment. Innate immune activation has been implicated generally in pain, opioid analgesia, cognitive dysfunction, and sickness type symptoms reported by cancer patients. We aimed to determine if genetic polymorphisms in neuroimmune activation pathways alter the serum fentanyl concentration-response relationships for pain control, cognitive dysfunction, and other adverse symptoms, in cancer pain patients. Cancer pain patients (468 receiving transdermal fentanyl were genotyped for 31 single nucleotide polymorphisms in 19 genes: CASP1, BDNF, CRP, LY96, IL6, IL1B, TGFB1, TNF, IL10, IL2, TLR2, TLR4, MYD88, IL6R, OPRM1, ARRB2, COMT, STAT6 and ABCB1. Lasso and backward stepwise generalised linear regression were used to identify non-genetic and genetic predictors, respectively, of pain control (average Brief Pain Inventory < 4, cognitive dysfunction (Mini-Mental State Examination ≤ 23, sickness response and opioid adverse event complaint. Serum fentanyl concentrations did not predict between-patient variability in these outcomes, nor did genetic factors predict pain control, sickness response or opioid adverse event complaint. Carriers of the MYD88 rs6853 variant were half as likely to have cognitive dysfunction (11/111 than wild-type patients (69/325, with a relative risk of 0.45 (95% CI: 0.27 to 0.76 when accounting for major non-genetic predictors (age, Karnofsky functional score. This supports the involvement of innate immune signalling in cognitive dysfunction, and identifies MyD88 signalling pathways as a potential focus for predicting and reducing the burden of cognitive dysfunction in cancer pain patients.

Qi-Dong-Huo-Xue-Yin Inhibits Inflammation in Acute Lung Injury in Mice via Toll-Like Receptor 4/Caveolin-1 Signaling

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Li-Ying Xu

2018-01-01

Full Text Available Acute lung injury (ALI is a critical illness with no current effective treatment. Caveolin-1 indirectly activates inflammation-associated signaling pathways by inhibiting endothelial nitric oxide synthase (eNOS. This induces an imbalance between pro- and anti-inflammatory cytokine levels, which are involved in the pathogenesis of ALI. The compound Chinese prescription Qi-Dong-Huo-Xue-Yin (QDHXY is efficacious for ALI treatment via an anti-inflammatory effect; however, the exact underlying mechanism is unknown. Therefore, we explored the protective effect of QDHXY against lipopolysaccharide- (LPS- induced ALI in mice. Histopathological changes in mouse lung tissues were studied. Furthermore, alterations in the serum levels of pro- and anti-inflammatory cytokines were investigated. The levels of tumor necrosis factor- (TNF-α, interleukin- (IL- 6, IL-1β, and interferon-γ-induced protein 10 in bronchoalveolar lavage fluid were measured. Additionally, the expression levels of myeloid differentiation factor 88 (MyD88, caveolin-1, and eNOS were assessed. QDHXY significantly reduced lung infiltration with inflammatory cells and the production of serum pro- and anti-inflammatory cytokines and inhibited the expression of TNF-α, IL-1β, caveolin-1, and MyD88 but not eNOS. These indicate that QDHXY significantly improved the balance between pro- and anti-inflammatory cytokine levels, possibly by inhibiting the caveolin-1 signaling pathway. Therefore, QDHXY may be a potential treatment for ALI.

The Effects of Agaricus blazei Murill Polysaccharides on Cadmium-Induced Apoptosis and the TLR4 Signaling Pathway of Peripheral Blood Lymphocytes in Chicken.

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Liu, Wenjing; Ge, Ming; Hu, Xuequan; Lv, Ai; Ma, Dexing; Huang, Xiaodan; Zhang, Ruili

2017-11-01

In this study, we investigated the effects of Agaricus blazei Murill polysaccharides (ABP) on cadmium (Cd)-induced apoptosis and the TLR4 signaling pathway of chicken peripheral blood lymphocytes (PBLs). Seven-day-old healthy chickens were randomly divided into four groups, and each group contained 20 males. The cadmium-supplemented diet group (Cd group) was fed daily with full feed that contained 140 mg cadmium chloride (CdCl 2 )/kg and 0.2 mL saline. The A. blazei Murill polysaccharide diet group (ABP group) was fed daily with full feed with 0.2 mL ABP solution (30 mg/mL) by oral gavage. The cadmium-supplemented plus A. blazei Murill polysaccharide diet group (Cd + ABP group) was fed daily with full feed containing 140 mg CdCl 2 /kg and 0.2 mL ABP solution (30 mg/mL) by gavage. The control group was fed daily with full feed with 0.2 mL saline per day. We measured the apoptosis rate and messenger RNA (mRNA) levels of apoptosis genes (caspase-3, Bax, and Bcl-2), the mRNA levels of TLR4 and TLR4 signaling pathway-related factors (MyD88, TRIF, NF-κB, and IRF3), the TLR4 protein expression, and the concentrations of inflammatory cytokines (IL-1β, IL-6, and TNF-α) in chicken PBLs. The results showed that the PBL apoptosis rate was significantly increased, the mRNA levels of caspase-3 and Bax were significantly increased, while that of Bcl-2 was significantly reduced. The Bax/Bcl-2 ratio was significantly increased in the Cd group at 20, 40, and 60 days after treatment compared with that in the control group. After treatment with ABP, the above changes were clearly suppressed. At the same time, ABP reduced the concentrations of IL-1β, IL-6, and TNF-α induced by Cd. We also found that ABP inhibited the TLR4 mRNA level and protein expression and inhibited the mRNA levels of MyD88, TRIF, NF-κB, and IRF3. The results demonstrated that Cd could induce apoptosis, activate the TLR4 signaling pathway, and induce the expression of inflammatory cytokines in

DMPD: Crosstalk among Jak-STAT, Toll-like receptor, and ITAM-dependent pathways inmacrophage activation. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 17502339 Crosstalk among Jak-STAT, Toll-like receptor, and ITAM-dependent pathways inmacrophage...May 14. (.png) (.svg) (.html) (.csml) Show Crosstalk among Jak-STAT, Toll-like receptor, and ITAM-dependent pathways inmacrophage...T, Toll-like receptor, and ITAM-dependent pathways inmacrophage activation. Authors Hu X, Chen J, Wang L, Iv

Waldenström macroglobulinemia: biology, genetics, and therapy

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Paludo J

2016-07-01

Full Text Available Jonas Paludo,1,2 Stephen M Ansell,1 1Division of Hematology, 2Division of Medical Oncology, Mayo Clinic, Rochester, MN, USA Abstract: Waldenström macroglobulinemia (WM is a distinct clinicopathologic entity characterized by the presence of a lymphoplasmacytic lymphoma, a non-Hodgkin lymphoma, and IgM monoclonal gammopathy. WM is an indolent, uncommon malignancy mostly affecting the elderly. Patient outcomes have modestly improved since the introduction of rituximab to conventional cytotoxic chemotherapy more than 20 years ago. However, the pivotal discovery of the somatic MYD88 L265P mutation, harbored by most patients with WM, and the somatic CXCR4 WHIM mutations, similar to germline CXCR4 mutations seen in the warts, hypogammaglobulinemia, immunodeficiency, and myelokathexis (WHIM syndrome, present in approximately one-third of patients with WM, has fundamentally changed our understanding of this disease and expanded the potential therapeutic targets. Within this new paradigm, ibrutinib emerged as a promising new drug. Ibrutinib targets Bruton’s tyrosine kinase, a downstream protein in the B-cell receptor pathway that is overactivated by the MYD88 L265P mutation. A seminal Phase II trial of ibrutinib in previously treated WM patients showed impressive response rates and confirmed the effects of MYD88 L265P and CXCR4 WHIM mutations in response to therapy. Ibrutinib is the first and only US Food and Drug Administration–approved drug specifically for the treatment of WM. However, before ibrutinib can be established as the standard of care for WM, long-term data regarding efficacy and safety are required. Further research to address ibrutinib resistance and cost-effectiveness is also imperative before ibrutinib can gain widespread acceptance. This review will cover the present pathophysiologic understanding of WM in light of the recent MYD88 and CXCR4 discovery, as well as current and emergent treatment regimens with focus on ibrutinib

Effects of total glucosides of paeony on immune regulatory toll-like receptors TLR2 and 4 in the kidney from diabetic rats.

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Xu, Xing-xin; Qi, Xiang-Ming; Zhang, Wei; Zhang, Chao-Qun; Wu, Xiao-Xu; Wu, Yong-Gui; Wang, Kun; Shen, Ji-Jia

2014-05-15

TLRs are a family of receptors that play a critical role in the pathogenesis of diabetic nephropathy. TGP have been shown to have anti-inflammatory and immuno-regulatory activities. However, the relation between TGP and TLRs on diabetic nephropathy remains unknown. In this study, we examined effects of TGP on immune regulatory TLR2 and 4 in the kidney from streptozotocin-induced diabetic rats. TGP decreased the levels of 24h urinary albumin excretion rate significantly in diabetic rats. Western blot analysis showed that TGP significantly inhibited the expression of TLR2 and 4, MyD88, p-IRAK1, NF-κB p65, p-IRF3, TNF-α and IL-1β. Quantitative real-time PCR analysis showed that the significantly increased levels of TLR2 and 4, and MyD88mRNA in the kidneys of diabetic rats were significantly suppressed by TGP treatment. Macrophages infiltration were also markedly increased in the kidneys of the diabetic rats, but were significantly inhibited by TGP in a dose-dependent manner. These results suggest that TGP has protective effects on several pharmacological targets in the progress of diabetic nephropathy by selectively blocking TLRs activation in vivo. Copyright © 2013 Elsevier GmbH. All rights reserved.

The Combination of Early and Rapid Type I IFN, IL-1α, and IL-1β Production Are Essential Mediators of RNA-Like Adjuvant Driven CD4+ Th1 Responses

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Madera, Rachel F.; Wang, Jennifer P.; Libraty, Daniel H.

2011-01-01

There is a growing need for novel vaccine adjuvants that can provide safe and potent T-helper type 1 (Th1) activity. RNA-like immune response modifiers (IRMs) are candidate T-cell adjuvants that skew acquired immune responses towards a Th1 phenotype. We set out to delineate the essential signaling pathways by which the RNA-like IRMs, resiquimod (R-848) and polyinosinic:polycytidylic acid (poly I:C), augment CD4+ T-helper 1 (Th1) responses. Highly purified murine conventional dendritic cells (cDCs) and conventional CD4+ T-cells were co-cultured in allogeneic and MHC congenic mixed leukocyte reactions. The activation of CD4+ Th1 cells was examined utilizing cells from mice deficient in specific RNA-sensing pattern recognition receptors and signaling mediators. R-848 and poly I:C stimulation of Type I interferon production and signaling in cDCs was essential but not sufficient for driving CD4+ Th1 responses. The early and rapid production of IL-1α and IL-1β was equally critical for the optimal activation of Th1 CD4+ T-cells. R-848 activation of Toll-like receptor 7/MyD88-dependent signaling in cDCs led to a rapid upregulation of pro-IL-1α and pro-IL-1β production compared to poly I:C activation of MyD88-independent signaling pathways. The in vitro data show that CD4+ T-cell adjuvant activity of RNA-like IRMs is mediated by a critical combination of early and rapid Type I interferon, IL-1α and IL-1β production. These results provide important insights into the key signaling pathways responsible for RNA-like IRM CD4+ Th1 activation. A better understanding of the critical signaling pathways by which RNA-like IRMs stimulate CD4+ Th1 responses is relevant to the rational design of improved vaccine adjuvants. PMID:22206014

The combination of early and rapid type I IFN, IL-1α, and IL-1β production are essential mediators of RNA-like adjuvant driven CD4+ Th1 responses.

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Rachel F Madera

Full Text Available There is a growing need for novel vaccine adjuvants that can provide safe and potent T-helper type 1 (Th1 activity. RNA-like immune response modifiers (IRMs are candidate T-cell adjuvants that skew acquired immune responses towards a Th1 phenotype. We set out to delineate the essential signaling pathways by which the RNA-like IRMs, resiquimod (R-848 and polyinosinic:polycytidylic acid (poly I:C, augment CD4+ T-helper 1 (Th1 responses. Highly purified murine conventional dendritic cells (cDCs and conventional CD4+ T-cells were co-cultured in allogeneic and MHC congenic mixed leukocyte reactions. The activation of CD4+ Th1 cells was examined utilizing cells from mice deficient in specific RNA-sensing pattern recognition receptors and signaling mediators. R-848 and poly I:C stimulation of Type I interferon production and signaling in cDCs was essential but not sufficient for driving CD4+ Th1 responses. The early and rapid production of IL-1α and IL-1β was equally critical for the optimal activation of Th1 CD4+ T-cells. R-848 activation of Toll-like receptor 7/MyD88-dependent signaling in cDCs led to a rapid upregulation of pro-IL-1α and pro-IL-1β production compared to poly I:C activation of MyD88-independent signaling pathways. The in vitro data show that CD4+ T-cell adjuvant activity of RNA-like IRMs is mediated by a critical combination of early and rapid Type I interferon, IL-1α and IL-1β production. These results provide important insights into the key signaling pathways responsible for RNA-like IRM CD4+ Th1 activation. A better understanding of the critical signaling pathways by which RNA-like IRMs stimulate CD4+ Th1 responses is relevant to the rational design of improved vaccine adjuvants.

CT findings of muscular dystrophy; Limb girdle type (LG), myotonic type (MYD) and Duchenne type (DMD)

Energy Technology Data Exchange (ETDEWEB)

Saitoh, Hiroshi (Tokushima Univ. (Japan). School of Medicine)

1991-07-01

CT scans of muscles in patients with limb girdle type (LG), myotonic type (MYD) and Duchenne type (DMD) dystrophies were obtained at five different body levels: the neck, L3 vertebral body, pelvic girdle, thigh and lower leg. CT numbers, cross sectional areas (CSA) and %CSA of muscle or fat were evaluated in each muscle. The characteristic CT patterns for each type of muscular dystrophy were obtained. Compared with DMD, the gracilis and soleus were more severely damaged in LG and the biceps femoris remained relatively preserved among the hamstrings. In addition, the multifidus of the neck and sternocleidomastoid also were more severely damaged in MYD. This study suggests that CT scan will be useful in the differential diagnosis of these types of muscular dystrophy as well as in planning appropriate rehabilitation and detecting damaged muscles. (author).

Diclofenac pretreatment modulates exercise-induced inflammation in skeletal muscle of rats through the TLR4/NF-κB pathway.

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Barcelos, Rômulo Pillon; Bresciani, Guilherme; Cuevas, Maria José; Martínez-Flórez, Susana; Soares, Félix Alexandre Antunes; González-Gallego, Javier

2017-07-01

Nonsteroidal anti-inflammatory drugs, such as diclofenac, are widely used to treat inflammation and pain in several conditions, including sports injuries. This study analyzes the influence of diclofenac on the toll-like receptor-nuclear factor kappa B (TLR-NF-κB) pathway in skeletal muscle of rats submitted to acute eccentric exercise. Twenty male Wistar rats were divided into 4 groups: control-saline, control-diclofenac, exercise-saline, and exercise-diclofenac. Diclofenac or saline were administered for 7 days prior to an acute eccentric exercise bout. The inflammatory status was evaluated through mRNA levels of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), interleukin-6 (IL-6), IL-1β, and tumor necrosis factor alpha (TNF-α), and protein content of COX-2, IL-6, and TNF-α in vastus lateralis muscle. Data obtained showed that a single bout of eccentric exercise significantly increased COX-2 gene expression. Similarly, mRNA expression and protein content of other inflammation-related genes also increased after the acute exercise. However, these effects were attenuated in the exercise + diclofenac group. TLR4, myeloid differentiation primary response gene 88 (MyD88), and p65 were also upregulated after the acute eccentric bout and the effect was blunted by the anti-inflammatory drug. These findings suggest that pretreatment with diclofenac may represent an effective tool to ameliorate the pro-inflammatory status induced by acute exercise in rat skeletal muscle possibly through an attenuation of the TLR4-NF-κB signaling pathway.

Intraflagellar transport 88 (IFT88) is crucial for craniofacial development in mice and is a candidate gene for human cleft lip and palate.

Science.gov (United States)

Tian, Hua; Feng, Jifan; Li, Jingyuan; Ho, Thach-Vu; Yuan, Yuan; Liu, Yang; Brindopke, Frederick; Figueiredo, Jane C; Magee, William; Sanchez-Lara, Pedro A; Chai, Yang

2017-03-01

Ciliopathies are pleiotropic human diseases resulting from defects of the primary cilium, and these patients often have cleft lip and palate. IFT88 is required for the assembly and function of the primary cilia, which mediate the activity of key developmental signaling pathways. Through whole exome sequencing of a family of three affected siblings with isolated cleft lip and palate, we discovered that they share a novel missense mutation in IFT88 (c.915G > C, p.E305D), suggesting this gene should be considered a candidate for isolated orofacial clefting. In order to evaluate the function of IFT88 in regulating craniofacial development, we generated Wnt1-Cre;Ift88fl/fl mice to eliminate Ift88 specifically in cranial neural crest (CNC) cells. Wnt1-Cre;Ift88fl/flpups died at birth due to severe craniofacial defects including bilateral cleft lip and palate and tongue agenesis, following the loss of the primary cilia in the CNC-derived palatal mesenchyme. Loss of Ift88 also resulted in a decrease in neural crest cell proliferation during early stages of palatogenesis as well as a downregulation of the Shh signaling pathway in the palatal mesenchyme. Importantly, Osr2KI-Cre;Ift88fl/flmice, in which Ift88 is lost specifically in the palatal mesenchyme, exhibit isolated cleft palate. Taken together, our results demonstrate that IFT88 has a highly conserved function within the primary cilia of the CNC-derived mesenchyme in the lip and palate region in mice and is a strong candidate as an orofacial clefting gene in humans. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

[Polymorphisms of TLR7 rs3853839 and rs179010 are associated with susceptibility to and severity of hand, foot and mouth disease caused by enterovirus 71 in male children].

Science.gov (United States)

Li, Yaping; Zhai, Song; Li, Mei; Wang, Yuan; Lu, Tong; Deng, Huiling; Zhang, Xin; Dang, Shuangsuo

2017-07-01

Objective To investigate whether the polymorphisms of TLR7/MyD88 signaling pathway is associated with the susceptibility to and severity of hand, foot and mouth disease (HFMD) caused by enterovirus 71 (EV71) in children. Methods We collected 180 EV71 HFMD cases and 201 healthy controls from both the Second Affiliated Hospital of Xi'an Jiaotong University and Xi'an Children's Hospital. The genotypes including rs3853839, rs179010 of TLR7, and rs7744 of MyD88 were detected in the 381 samples by SNPscan kit. Results The susceptibility risk (OR=2.343, 95%CI:1.516-3.621) and severity risk (OR=1.939, 95%CI: 1.064-3.521) of TLR7 rs3853839 allele C significantly increased in the male children with EV71 HFMD. Also, the susceptibility risk (OR=1.701, 95%CI: 1.142-2.535) and severity risk (OR=1.852, 95%CI: 1.038-3.305) of TLR7 rs179010 allele T significantly increased in the male children with EV71 HFMD. But there was no significant difference in the distribution of TLR7 rs179010 and rs3853839 genes between female children with EV71 HFMD and female controls. There was no correlation between the genetic polymorphisms of MyD88 rs7744 and the susceptibility to and severity of EV71 HFMD in the children. Conclusion Polymorphisms of TLR7 rs3853839 and rs179010 are correlated to the susceptibility to and severity of EV71 HFMD in male children.

Hydrogen peroxide induces activation of insulin signaling pathway via AMP-dependent kinase in podocytes

International Nuclear Information System (INIS)

Piwkowska, Agnieszka; Rogacka, Dorota; Angielski, Stefan; Jankowski, Maciej

2012-01-01

Highlights: ► H 2 O 2 activates the insulin signaling pathway and glucose uptake in podocytes. ► H 2 O 2 induces time-dependent changes in AMPK phosphorylation. ► H 2 O 2 enhances insulin signaling pathways via AMPK activation. ► H 2 O 2 stimulation of glucose uptake is AMPK-dependent. -- Abstract: Podocytes are cells that form the glomerular filtration barrier in the kidney. Insulin signaling in podocytes is critical for normal kidney function. Insulin signaling is regulated by oxidative stress and intracellular energy levels. We cultured rat podocytes to investigate the effects of hydrogen peroxide (H 2 O 2 ) on the phosphorylation of proximal and distal elements of insulin signaling. We also investigated H 2 O 2 -induced intracellular changes in the distribution of protein kinase B (Akt). Western blots showed that H 2 O 2 (100 μM) induced rapid, transient phosphorylation of the insulin receptor (IR), the IR substrate-1 (IRS1), and Akt with peak activities at 5 min (Δ 183%, P 2 O 2 >. Furthermore, H 2 O 2 inhibited phosphorylation of the phosphatase and tensin homologue (PTEN; peak activity at 10 min; Δ −32%, P 2 O 2 on IR phosphorylation by about 40% (from 2.07 ± 0.28 to 1.28 ± 0.12, P 2 O 2 increased glucose uptake in podocytes (from 0.88 ± 0.04 to 1.29 ± 0.12 nmol/min/mg protein, P 2 O 2 activated the insulin signaling pathway and glucose uptake via AMPK in cultured rat podocytes. This signaling may play a potential role in the prevention of insulin resistance under conditions associated with oxidative stress.

The down-stream effects of mannan-induced lectin complement pathway activation depend quantitatively on alternative pathway amplification

DEFF Research Database (Denmark)

Harboe, Morten; Garred, Peter; Karlstrøm, Ellen

2009-01-01

Complement activation plays an important role in human pathophysiology. The effect of classical pathway activation is largely dependent on alternative pathway (AP) amplification, whereas the role of AP for the down-stream effect of mannan-induced lectin pathway (LP) activation is poorly understood...... that AP amplification is quantitatively responsible for the final effect of initial specific LP activation. TCC generation on the solid phase was distinctly but less inhibited by anti-fD. C2 bypass of the LP pathway could be demonstrated, and AP amplification was also essential during C2 bypass in LP...... as shown by complete inhibition of TCC generation in C2-deficient serum by anti-fD and anti-properdin antibodies. In conclusion, the down-stream effect of LP activation depends strongly on AP amplification in normal human serum and in the C2 bypass pathway....

Dexamethasone, rituximab and cyclophosphamide for relapsed and/or refractory and treatment-naïve patients with Waldenstrom macroglobulinemia.

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Paludo, Jonas; Abeykoon, Jithma P; Kumar, Shaji; Shreders, Amanda; Ailawadhi, Sikander; Gertz, Morie A; Kourelis, Taxiarchis; King, Rebecca L; Reeder, Craig B; Leung, Nelson; Kyle, Robert A; Buadi, Francis K; Habermann, Thomas M; Dingli, David; Witzig, Thomas E; Dispenzieri, Angela; Lacy, Martha Q; Go, Ronald S; Lin, Yi; Gonsalves, Wilson I; Warsame, Rahma; Lust, John A; Rajkumar, S Vincent; Ansell, Stephen M; Kapoor, Prashant

2017-10-01

The management of Waldenström macroglobulinaemia (WM) relies predominantly on small trials, one of which has demonstrated activity of dexamethasone, rituximab and cyclophosphamide (DRC) in the frontline setting. We report on the efficacy of DRC, focusing on relapsed/refractory (R/R) patients. Ibrutinib, a recently approved agent in WM demonstrated limited activity in patients with MYD88 WT genotype. Herein, we additionally report on the activity of DRC based on the MYD88 L265P mutation status. Of 100 WM patients evaluated between January 2007 and December 2014 who received DRC, 50 had R/R WM. The overall response rate (ORR) was 87%. The median progression-free survival (PFS) and time-to-next-therapy (TTNT) were 32 (95% confidence interval [CI]: 15-51) and 50 (95% CI: 35-60) months, respectively. In the previously untreated cohort (n = 50), the ORR was 96%, and the median PFS and TTNT were 34 months (95% CI: 23-not reached [NR]) and NR (95% CI: 37-NR), respectively. Twenty-five (86%) of 29 genotyped patients harbored MYD88 L265P . The response rates and outcomes were independent of MYD88 mutation status. Grade ≥3 adverse effects included neutropenia (20%), thrombocytopenia (7%) and infections (3%). Similar to the frontline setting, DRC is an effective and well-tolerated salvage regimen for WM. In contrast to ibrutinib, DRC offers a less expensive, fixed-duration option, with preliminary data suggesting efficacy independent of the patients' MYD88 status. © 2017 John Wiley & Sons Ltd.

Biglycan, a novel trigger of Th1 and Th17 cell recruitment into the kidney.

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Nastase, Madalina-Viviana; Zeng-Brouwers, Jinyang; Beckmann, Janet; Tredup, Claudia; Christen, Urs; Radeke, Heinfried H; Wygrecka, Malgorzata; Schaefer, Liliana

2017-12-15

Th1 and Th17 cells, T helper (Th) subtypes, are key inducers of renal fibrosis. The molecular mechanisms of their recruitment into the kidney, however, are not well understood. Here, we show that biglycan, a proteoglycan of the extracellular matrix, acting in its soluble form as a danger signal, stimulates autonomously the production of Th1 and Th17 chemoattractants CXCL10 and CCL20 in macrophages. In the presence of IFNγ, biglycan synergistically stimulates CXCL9. In macrophages deficient for TLR2, TLR4, and their adaptor molecules MyD88 or TRIF, we identified highly selective mechanisms of biglycan-dependent Th1/17 chemoattraction. Thus, the expression of CXCL9 and CXCL10, common chemoattractants for CXCR3-positive Th1 and Th17 cells, is triggered in a biglycan-TLR4/TRIF-dependent manner. By contrast, biglycan induces CCL20 chemokine production, responsible for CCR6-positive Th17 cell recruitment, in a TLR2/4/MyD88-dependent manner. Importantly, at the onset of diabetes mellitus and lupus nephritis we provide evidence for biglycan-dependent recruitment of Th1 and Th17 cells, IFNγ and IL-17 production, and development of albuminuria in mice lacking or overexpressing soluble biglycan. Furthermore, by genetic ablation of Cxcl10 we showed in vivo involvement of this chemokine in biglycan-dependent recruitment of Th1 and Th17 cells into the kidney. Finally, a positive correlation of biglycan and CXCL10/CXCL9 levels was detected in plasma from patients with diabetic nephropathy and lupus nephritis. Taken together, we identified biglycan as a novel trigger of Th1 and Th17 cell recruitment into the kidney and we postulate that interfering with biglycan/TLR/TRIF/MyD88-signaling might provide novel therapeutic avenues for renal fibrosis. Copyright © 2017 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

Flaviviridae virus nonstructural proteins 5 and 5A mediate viral immune evasion and are promising targets in drug development.

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Chen, Shun; Yang, Chao; Zhang, Wei; Mahalingam, Suresh; Wang, Mingshu; Cheng, Anchun

2018-05-06

Infections with viruses in the Flaviviridae family have a vast global and economic impact because of the high morbidity and mortality. The pathogenesis of Flaviviridae infections is very complex and not fully understood because these viruses can inhibit multiple immune pathways including the complement system, NK cells, and IFN induction and signalling pathways. The non-structural (NS) 5 and 5A proteins of Flaviviridae viruses are highly conserved and play an important role in resisting host immunity through various evasion mechanisms. This review summarizes the strategies used by the NS5 and 5A proteins of Flaviviridae viruses for evading the innate immune response by inhibiting pattern recognition receptor (PRR) signalling pathways (TLR/MyD88, IRF7), suppressing interferon (IFN) signalling pathways (IFN-γRs, STAT1, STAT2), and impairing the function of IFN-stimulated genes (ISGs) (e.g. protein kinase R [PKR], oligoadenylate synthase [OAS]). All of these immune evasion mechanisms depend on the interaction of NS5 or NS5A with cellular proteins, such as MyD88 and IRF7, IFN-αRs, IFN-γRs, STAT1, STAT2, PKR and OAS. NS5 is the most attractive target for the discovery of broad spectrum compounds against Flaviviridae virus infection. The methyltransferase (MTase) and RNA-dependent RNA polymerase (RdRp) activities of NS5 are the main therapeutic targets for antiviral drugs against Flaviviridae virus infection. Based on our site mapping, the sites involved in immune evasion provide some potential and promising targets for further novel antiviral therapeutics. Copyright © 2018 Elsevier Inc. All rights reserved.

Analysis of the effect of Qizhuyigan on liver function in a mouse ...

African Journals Online (AJOL)

expenses, the severe toxic side effects of drugs, and viral drug resistance, ... Recently, the pharmacological mechanisms and the clinical ... saline at 4°C, and dried with filter paper. Samples (0.2 g) .... resistance to copper in hydroids linked to hormesis. Mar. Environ ... provide a MyD88-dependent negative signal for Th2 cell.

The effect of n-3/n-6 polyunsaturated fatty acids on acute reflux esophagitis in rats.

Science.gov (United States)

Zhuang, Ze-Hao; Xie, Jing-Jing; Wei, Jing-Jing; Tang, Du-Peng; Yang, Li-Yong

2016-10-04

Polyunsaturated fatty acids (PUFAs) play various roles in inflammation. However, the effect of PUFAs in the development of reflux esophagitis (RE) is unclear. This study is to investigate the potential effect of n-3/n-6 PUFAs on acute RE in rats along with the underlying protective mechanisms. Forty Sprague Dawley rats were randomly divided into four groups (n = 10 in each group). RE model was established by pyloric clip and section ligation. Fish oil- and soybean oil-based fatty emulsion (n-3 and n-6 groups), or normal saline (control and sham operation groups) was injected intraperitoneally 2 h prior to surgery and 24 h postoperatively (2 mL/kg, respectively). The expressions of interleukin (IL)-1β, IL-8, IL-6 and myeloid differentiation primary response gene 88 (MyD88) in esophageal tissues were evaluated by Western blot and immunohistochemistry after 72 h. The malondialdehyde (MDA) and superoxide dismutase (SOD) expression in the esophageal tissues were determined to assess the oxidative stress. The mildest macroscopic/microscopic esophagitis was found in the n-3 group (P < 0.05). The expression of IL-1β, IL-8, IL-6 and MyD88 were increased in all RE groups, while the lowest and highest expression were found in n-3 and n-6 group, respectively (P < 0.05). The MDA levels were increased in all groups (P < 0.05), in an ascending trend from n-3, n-6 groups to control group. The lowest and highest SOD levels were found in the control and n-3 group, respectively (P < 0.05). n-3 PUFAs may reduce acute RE in rats, which may be due to inhibition of the MyD88-NF-kB pathway and limit oxidative damage.

Waldenström macroglobulinemia: Progress in evidence-based medicine and updates of consensus panel recommendation

Directory of Open Access Journals (Sweden)

Jian HOU

2013-09-01

Full Text Available Waldenström macroglobulinemia (WM is a distinct B-cell lymphoproliferative disorder primarily characterized by infiltration of lymphoplasmacytic cells into the bone marrow, along with demonstration of monoclonal immunoglobulinemia M(IgM. Recent studies have demonstrated the MYD88 L265P somatic mutation contributes to the pathogenesis of WM. Mutated MYD88 activates NF-κB through a series of signals, which results in abnormal propagation of B cells. Clinical manifestation mainly consisted of tissue injuries attributable to tissue infiltration and monoclonal IgM. WM should be differentiated from lymphomas producing monoclonal IgM, and detection of MYD88 L265P mutation provides a new way to differentiate WM from other similar diseases. At present recommended treatments for WM include alkylating agents, nucleoside analogues, bortezomib, rituximab etc.

Thymic commitment of regulatory T cells is a pathway of TCR-dependent selection that isolates repertoires undergoing positive or negative selection.

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Coutinho, A; Caramalho, I; Seixas, E; Demengeot, J

2005-01-01

The seminal work of Le Douarin and colleagues (Ohki et al. 1987; Ohki et al. 1988; Salaun et al. 1990; Coutinho et al. 1993) first demonstrated that peripheral tissue-specific tolerance is centrally established in the thymus, by epithelial stromal cells (TEC). Subsequent experiments have shown that TEC-tolerance is dominant and mediated by CD4 regulatory T cells (Treg) that are generated intrathymically by recognition of antigens expressed on TECs (Modigliani et al. 1995; Modigliani et al. 1996a). From these and other observations, in 1996 Modigliani and colleagues derived a general model for the establishment and maintenance of natural tolerance (MM96) (Modigliani et al. 1996b), with two central propositions: (1) T cell receptor (TCR)-dependent sorting of emergent repertoires generates TEC-specific Treg displaying the highest TCR self-affinities below deletion thresholds, thus isolating repertoires undergoing positive and negative selection; (2) Treg are intrathymically committed (and activated) for a unique differentiative pathway with regulatory effector functions. The model explained the embryonic/perinatal time window of natural tolerance acquisition, by developmental programs determining (1) TCR multireactivity, (2) the cellular composition in the thymic stroma (relative abundance of epithelial vs hemopoietic cells), and (3) the dynamics of peripheral lymphocyte pools, built by accumulation of recent thymic emigrants (RTE) that remain recruitable to regulatory functions. We discuss here the MM96 in the light of recent results demonstrating the promiscuous expression of tissue-specific antigens by medullary TECs (Derbinski et al. 2001; Anderson et al. 2002; Gotter et al. 2004) and indicating that Treg represent a unique differentiative pathway (Fontenot et al. 2003; Hori et al. 2003; Khattri et al. 2003), which is adopted by CD4 T cells with high avidity for TEC-antigens (Bensinger et al. 2001; Jordan et al. 2001; Apostolou et al. 2002). In the likelihood that

HSV-1 ICP0: An E3 Ubiquitin Ligase That Counteracts Host Intrinsic and Innate Immunity

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Mirna Perusina Lanfranca

2014-05-01

Full Text Available The herpes simplex virus type 1 (HSV-1 encoded E3 ubiquitin ligase, infected cell protein 0 (ICP0, is required for efficient lytic viral replication and regulates the switch between the lytic and latent states of HSV-1. As an E3 ubiquitin ligase, ICP0 directs the proteasomal degradation of several cellular targets, allowing the virus to counteract different cellular intrinsic and innate immune responses. In this review, we will focus on how ICP0’s E3 ubiquitin ligase activity inactivates the host intrinsic defenses, such as nuclear domain 10 (ND10, SUMO, and the DNA damage response to HSV-1 infection. In addition, we will examine ICP0’s capacity to impair the activation of interferon (innate regulatory mediators that include IFI16 (IFN γ-inducible protein 16, MyD88 (myeloid differentiation factor 88, and Mal (MyD88 adaptor-like protein. We will also consider how ICP0 allows HSV-1 to evade activation of the NF-κB (nuclear factor kappa B inflammatory signaling pathway. Finally, ICP0’s paradoxical relationship with USP7 (ubiquitin specific protease 7 and its roles in intrinsic and innate immune responses to HSV-1 infection will be discussed.

microRNA-124 negatively regulates TLR signaling in alveolar macrophages in response to mycobacterial infection.

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Ma, Chunyan; Li, Yong; Li, Min; Deng, Guangcun; Wu, Xiaoling; Zeng, Jin; Hao, Xiujing; Wang, Xiaoping; Liu, Jing; Cho, William C S; Liu, Xiaoming; Wang, Yujiong

2014-11-01

The emerging roles of microRNAs (miRNAs) in regulating immune responses have attracted increasing attention in recent years; and the alveolar macrophages (AMs) are the main targets of mycobacterial infection, which play a pivotal role in the pathogenesis of Mycobacterium tuberculosis infection. However, the immunoregulatory role of miRNAs in AMs has not been fully demonstrated. In this study, we find that miR-124 is up-regulated in the peripheral leukocytes of patients with pulmonary tuberculosis; furthermore, the expression miR-124 can be induced upon Mycobacterium bovis Bacillus Calmette-Guerin (BCG) infection in both RAW264.7 AM cells in vitro and murine AMs in vivo. Mechanistically, miR-124 is able to modulate toll-like receptor (TLR) signaling activity in RAW264.7 cells in response to BCG infection. In this regard, multiple components of TLR signaling cascade, including the TLR6, myeloid differentiation factor 88 (MyD88), TNFR-associated factor 6 and tumor necrosis factor-α are directly targeted by miR-124. In addition, both overexpression of TLR signaling adaptor MyD88 and BCG infection are able to augment miR-124 transcription, while MyD88 expression silenced by small interfering RNA dramatically suppresses miR-124 expression in AMs in vitro. Moreover, the abundance of miR-124 transcript in murine AMs of MyD88 deficient mice is significantly less than that of their wild-type or heterozygous littermates; and the BCG infection fails to induce miR-124 expression in the lung of MyD88 deficient mouse. These results indicate a negative regulatory role of miR-124 in fine-tuning inflammatory response in AMs upon mycobacterial infection, in part through a mechanism by directly targeting TLR signaling. Copyright © 2014 Elsevier Ltd. All rights reserved.

Caspase-1 from Human Myeloid-Derived Suppressor Cells Can Promote T Cell-Independent Tumor Proliferation.

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Zeng, Qi; Fu, Juan; Korrer, Michael; Gorbounov, Mikhail; Murray, Peter J; Pardoll, Drew; Masica, David L; Kim, Young J

2018-05-01

Immunosuppressive myeloid-derived suppressive cells (MDSCs) are characterized by their phenotypic and functional heterogeneity. To better define their T cell-independent functions within the tumor, sorted monocytic CD14 + CD11b + HLA-DR low/- MDSCs (mMDSC) from squamous cell carcinoma patients showed upregulated caspase-1 activity, which was associated with increased IL1β and IL18 expression. In vitro studies demonstrated that mMDSCs promoted caspase-1-dependent proliferation of multiple squamous carcinoma cell lines in both human and murine systems. In vivo , growth rates of B16, MOC1, and Panc02 were significantly blunted in chimeric mice adoptively transferred with caspase-1 null bone marrow cells under T cell-depleted conditions. Adoptive transfer of wild-type Gr-1 + CD11b + MDSCs from tumor-bearing mice reversed this antitumor response, whereas caspase-1 inhibiting thalidomide-treated MDSCs phenocopied the antitumor response found in caspase-1 null mice. We further hypothesized that MDSC caspase-1 activity could promote tumor-intrinsic MyD88-dependent carcinogenesis. In mice with wild-type caspase-1, MyD88-silenced tumors displayed reduced growth rate, but in chimeric mice with caspase-1 null bone marrow cells, MyD88-silenced tumors did not display differential tumor growth rate. When we queried the TCGA database, we found that caspase-1 expression is correlated with overall survival in squamous cell carcinoma patients. Taken together, our findings demonstrated that caspase-1 in MDSCs is a direct T cell-independent mediator of tumor proliferation. Cancer Immunol Res; 6(5); 566-77. ©2018 AACR . ©2018 American Association for Cancer Research.

Electroacupuncture Improved Hippocampal Neurogenesis following Traumatic Brain Injury in Mice through Inhibition of TLR4 Signaling Pathway

Directory of Open Access Journals (Sweden)

Yuqin Ye

2017-01-01

Full Text Available The protective role of electroacupuncture (EA treatment in diverse neurological diseases such as ischemic stroke is well acknowledged. However, whether and how EA act on hippocampal neurogenesis following traumatic brain injury (TBI remains poorly understood. This study aims to investigate the effect of EA on hippocampal neurogenesis and neurological functions, as well as its underlying association with toll-like receptor 4 (TLR4 signaling in TBI mice. BrdU/NeuN immunofluorescence was performed to label newborn neurons in the hippocampus after EA treatment. Water maze test and neurological severity score were used to evaluate neurological function posttrauma. The hippocampal level of TLR4 and downstream molecules and inflammatory cytokines were, respectively, detected by Western blot and enzyme-linked immunosorbent assay. EA enhanced hippocampal neurogenesis and inhibited TLR4 expression at 21, 28, and 35 days after TBI, but the beneficial effects of EA on posttraumatic neurogenesis and neurological functions were attenuated by lipopolysaccharide-induced TLR4 activation. In addition, EA exerted an inhibitory effect on both TLR4/Myd88/NF-κB and TLR4/TRIF/NF-κB pathways, as well as the inflammatory cytokine expression in the hippocampus following TBI. In conclusion, EA promoted hippocampal neurogenesis and neurological recovery through inhibition of TLR4 signaling pathway posttrauma, which may be a potential approach to improve the outcome of TBI.

Angular dependent torque measurements on CaFe0.88Co0.12AsF

Science.gov (United States)

Xiao, H.; Gao, B.; Ma, Y. H.; Li, X. J.; Mu, G.; Hu, T.

2016-08-01

Out-of-plane angular dependent torque measurements were performed on CaFe0.88Co0.12AsF (Ca1 1 1 1) single crystals. In the normal state, the torque data shows \\sin 2θ angular dependence and H 2 magnetic field dependence, as a result of paramagnetism. In the mixed state, the torque signal is a combination of the vortex torque and paramagnetic torque, and the former allows the determination of the anisotropy parameter γ. At T   =  11.5 K, γ (11.5 K ≃ 0.5 T c)  =  19.1, which is similar to the result of SmFeAsO0.8F0.2, γ ≃ 23 at T≃ 0.4{{T}\\text{c}} . So the 11 1 1 is more anisotropic compared to 11 and 122 families of iron-based superconductors. This may suggest that the electronic coupling between layers in 1 1 1 1 is less effective than in 11 and 122 families.

Functional analysis of the Borrelia burgdorferi bba64 gene product in murine infection via tick infestation.

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Toni G Patton

Full Text Available Borrelia burgdorferi, the causative agent of Lyme borreliosis, is transmitted to humans from the bite of Ixodes spp. ticks. During the borrelial tick-to-mammal life cycle, B. burgdorferi must adapt to many environmental changes by regulating several genes, including bba64. Our laboratory recently demonstrated that the bba64 gene product is necessary for mouse infectivity when B. burgdorferi is transmitted by an infected tick bite, but not via needle inoculation. In this study we investigated the phenotypic properties of a bba64 mutant strain, including 1 replication during tick engorgement, 2 migration into the nymphal salivary glands, 3 host transmission, and 4 susceptibility to the MyD88-dependent innate immune response. Results revealed that the bba64 mutant's attenuated infectivity by tick bite was not due to a growth defect inside an actively feeding nymphal tick, or failure to invade the salivary glands. These findings suggested there was either a lack of spirochete transmission to the host dermis or increased susceptibility to the host's innate immune response. Further experiments showed the bba64 mutant was not culturable from mouse skin taken at the nymphal bite site and was unable to establish infection in MyD88-deficient mice via tick infestation. Collectively, the results of this study indicate that BBA64 functions at the salivary gland-to-host delivery interface of vector transmission and is not involved in resistance to MyD88-mediated innate immunity.

Time-dependent LXR/RXR pathway modulation characterizes capillary remodeling in inflammatory corneal neovascularization.

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Mukwaya, Anthony; Lennikov, Anton; Xeroudaki, Maria; Mirabelli, Pierfrancesco; Lachota, Mieszko; Jensen, Lasse; Peebo, Beatrice; Lagali, Neil

2018-05-01

Inflammation in the normally immune-privileged cornea can initiate a pathologic angiogenic response causing vision-threatening corneal neovascularization. Inflammatory pathways, however, are numerous, complex and are activated in a time-dependent manner. Effective resolution of inflammation and associated angiogenesis in the cornea requires knowledge of these pathways and their time dependence, which has, to date, remained largely unexplored. Here, using a model of endogenous resolution of inflammation-induced corneal angiogenesis, we investigate the time dependence of inflammatory genes in effecting capillary regression and the return of corneal transparency. Endogenous capillary regression was characterized by a progressive thinning and remodeling of angiogenic capillaries and inflammatory cell retreat in vivo in the rat cornea. By whole-genome longitudinal microarray analysis, early suppression of VEGF ligand-receptor signaling and inflammatory pathways preceded an unexpected later-phase preferential activation of LXR/RXR, PPARα/RXRα and STAT3 canonical pathways, with a concurrent attenuation of LPS/IL-1 inhibition of RXR function and Wnt/β-catenin signaling pathways. Potent downstream inflammatory cytokines such as Cxcl5, IL-1β, IL-6 and Ccl2 were concomitantly downregulated during the remodeling phase. Upstream regulators of the inflammatory pathways included Socs3, Sparc and ApoE. A complex and coordinated time-dependent interplay between pro- and anti-inflammatory signaling pathways highlights a potential anti-inflammatory role of LXR/RXR, PPARα/RXRα and STAT3 signaling pathways in resolving inflammatory corneal angiogenesis.

Molecular cloning, characterization and expression analysis of TLR9, MyD88 and TRAF6 genes in common carp (Cyprinus carpio)

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Induction of innate immune pathways is critical for early host defense but there is limited understanding of how teleost fish recognize pathogen molecules and activate these pathways. In mammals, cells of the innate immune system detect pathogenic molecular structures using pattern recognition rece...

Structure-Based Design of Potent and Selective 3-Phosphoinositide-Dependent Kinase-1 (PDK1) Inhibitors

Energy Technology Data Exchange (ETDEWEB)

Medina, Jesus R.; Becker, Christopher J.; Blackledge, Charles W.; Duquenne, Celine; Feng, Yanhong; Grant, Seth W.; Heerding, Dirk; Li, William H.; Miller, William H.; Romeril, Stuart P.; Scherzer, Daryl; Shu, Arthur; Bobko, Mark A.; Chadderton, Antony R.; Dumble, Melissa; Gardiner, Christine M.; Gilbert, Seth; Liu, Qi; Rabindran, Sridhar K.; Sudakin, Valery; Xiang, Hong; Brady, Pat G.; Campobasso, Nino; Ward, Paris; Axten, Jeffrey M. (GSKPA)

2014-10-02

Phosphoinositide-dependent protein kinase-1(PDK1) is a master regulator of the AGC family of kinases and an integral component of the PI3K/AKT/mTOR pathway. As this pathway is among the most commonly deregulated across all cancers, a selective inhibitor of PDK1 might have utility as an anticancer agent. Herein we describe our lead optimization of compound 1 toward highly potent and selective PDK1 inhibitors via a structure-based design strategy. The most potent and selective inhibitors demonstrated submicromolar activity as measured by inhibition of phosphorylation of PDK1 substrates as well as antiproliferative activity against a subset of AML cell lines. In addition, reduction of phosphorylation of PDK1 substrates was demonstrated in vivo in mice bearing OCl-AML2 xenografts. These observations demonstrate the utility of these molecules as tools to further delineate the biology of PDK1 and the potential pharmacological uses of a PDK1 inhibitor.

Downregulation of toll-like receptor-mediated signalling pathways in oral lichen planus.

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Sinon, Suraya H; Rich, Alison M; Parachuru, Venkata P B; Firth, Fiona A; Milne, Trudy; Seymour, Gregory J

2016-01-01

The objective of this study was to investigate the expression of Toll-like receptors (TLR) and TLR-associated signalling pathway genes in oral lichen planus (OLP). Initially, immunohistochemistry was used to determine TLR expression in 12 formalin-fixed archival OLP tissues with 12 non-specifically inflamed oral tissues as controls. RNA was isolated from further fresh samples of OLP and non-specifically inflamed oral tissue controls (n = 6 for both groups) and used in qRT(2)-PCR focused arrays to determine the expression of TLRs and associated signalling pathway genes. Genes with a statistical significance of ±two-fold regulation (FR) and a P-value < 0.05 were considered as significantly regulated. Significantly more TLR4(+) cells were present in the inflammatory infiltrate in OLP compared with the control tissues (P < 0.05). There was no statistically significant difference in the numbers of TLR2(+) and TLR8(+) cells between the groups. TLR3 was significantly downregulated in OLP (P < 0.01). TLR8 was upregulated in OLP, but the difference between the groups was not statistically significant. The TLR-mediated signalling-associated protein genes MyD88 and TIRAP were significantly downregulated (P < 0.01 and P < 0.05), as were IRAK1 (P < 0.05), MAPK8 (P < 0.01), MAP3K1 (P < 0.05), MAP4K4 (P < 0.05), REL (P < 0.01) and RELA (P < 0.01). Stress proteins HMGB1 and the heat shock protein D1 were significantly downregulated in OLP (P < 0.01). These findings suggest a downregulation of TLR-mediated signalling pathways in OLP lesions. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Functional Relevance of Different Basal Ganglia Pathways Investigated in a Spiking Model with Reward Dependent Plasticity

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Pierre Berthet

2016-07-01

Full Text Available The brain enables animals to behaviourally adapt in order to survive in a complex and dynamic environment, but how reward-oriented behaviours are achieved and computed by its underlying neural circuitry is an open question. To address this concern, we have developed a spiking model of the basal ganglia (BG that learns to dis-inhibit the action leading to a reward despite ongoing changes in the reward schedule. The architecture of the network features the two pathways commonly described in BG, the direct (denoted D1 and the indirect (denoted D2 pathway, as well as a loop involving striatum and the dopaminergic system. The activity of these dopaminergic neurons conveys the reward prediction error (RPE, which determines the magnitude of synaptic plasticity within the different pathways. All plastic connections implement a versatile four-factor learning rule derived from Bayesian inference that depends upon pre- and postsynaptic activity, receptor type and dopamine level. Synaptic weight updates occur in the D1 or D2 pathways depending on the sign of the RPE, and an efference copy informs upstream nuclei about the action selected. We demonstrate successful performance of the system in a multiple-choice learning task with a transiently changing reward schedule. We simulate lesioning of the various pathways and show that a condition without the D2 pathway fares worse than one without D1. Additionally, we simulate the degeneration observed in Parkinson’s disease (PD by decreasing the number of dopaminergic neurons during learning. The results suggest that the D1 pathway impairment in PD might have been overlooked. Furthermore, an analysis of the alterations in the synaptic weights shows that using the absolute reward value instead of the RPE leads to a larger change in D1.

Inhibition of HMGB1 reduces rat spinal cord astrocytic swelling and AQP4 expression after oxygen-glucose deprivation and reoxygenation via TLR4 and NF-κB signaling in an IL-6-dependent manner.

Science.gov (United States)

Sun, Lin; Li, Man; Ma, Xun; Feng, Haoyu; Song, Junlai; Lv, Cong; He, Yajun

2017-11-25

Spinal cord astrocyte swelling is an important component to spinal cord edema and is associated with poor functional recovery as well as therapeutic resistance after spinal cord injury (SCI). High mobility group box-1 (HMGB1) is a mediator of inflammatory responses in the central nervous system and plays a critical role after SCI. Given this, we sought to identify both the role and underlying mechanisms of HMGB1 in cellular swelling and aquaporin 4 (AQP4) expression in cultured rat spinal cord astrocytes after oxygen-glucose deprivation/reoxygenation (OGD/R). The post-natal day 1-2 Sprague-Dawley rat spinal cord astrocytes were cultured in vitro, and the OGD/R model was induced. We first investigated the effects of OGD/R on spinal cord astrocytic swelling and HMGB1 and AQP4 expression, as well as HMGB1 release. We then studied the effects of HMGB1 inhibition on cellular swelling, HMGB1 and AQP4 expression, and HMGB1 release. The roles of both toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) signaling pathway and interleukin-6 (IL-6) in reducing cellular swelling resulting from HMGB1 inhibition in spinal cord astrocytes after OGD/R were studied. Intergroup data were compared using one-way analysis of variance (ANOVA) followed by Dunnett's test. The OGD/R increased spinal cord astrocytic swelling and HMGB1 and AQP4 expression, as well as HMGB1 release. Inhibition of HMGB1 using either HMGB1 shRNA or ethyl pyruvate resulted in reduced cellular volume, mitochondrial and endoplasmic reticulum swelling, and lysosome number and decreased upregulation of both HMGB1 and AQP4 in spinal cord astrocytes, as well as HMGB1 release. The HMGB1 effects on spinal cord astrocytic swelling and AQP4 upregulation after OGD/R were mediated-at least in part-via activation of TLR4, myeloid differentiation primary response gene 88 (MyD88), and NF-κB. These activation effects can be repressed by TLR4 inhibition using CLI-095 or C34, or by NF-κB inhibition using BAY 11

Developmental pathways from child maltreatment to adolescent marijuana dependence: Examining moderation by FKBP5

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Handley, Elizabeth D.; Rogosch, Fred A.; Cicchetti, Dante

2015-01-01

The current study examined the prospective association between child maltreatment and the development of substance use disorder (SUD) in adolescence with the aim of investigating pathways underlying this relation, as well as genetic moderation of these developmental mechanisms. Specifically, we tested whether youth who experienced maltreatment prior to age 8 were at risk for the development of marijuana dependence in adolescence by way of a childhood externalizing pathway and a childhood internalizing pathway. Moreover, we tested whether variation in FKBP5 CATT haplotype moderated these pathways. The participants were 326 children (n=179 maltreated; n=147 nonmaltreated) assessed across two waves of data collection (childhood: ages 7–9 and adolescence: ages 15–18). Results indicated that higher levels of child externalizing symptoms significantly mediated the effect of child maltreatment on adolescent marijuana dependence symptoms for individuals with 1–2 copies of the FKBP5 CATT haplotype only. We did not find support for an internalizing pathway from child maltreatment to adolescent marijuana dependence, nor did we find evidence of moderation of the internalizing pathway by FKBP5 haplotype variation. Findings extend previous research by demonstrating that whether a maltreated child will traverse an externalizing pathway toward SUD in adolescence is dependent on FKBP5 genetic variation. PMID:26535939

Gliadin induces an increase in intestinal permeability and zonulin release by binding to the chemokine receptor CXCR3.

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Lammers, Karen M; Lu, Ruliang; Brownley, Julie; Lu, Bao; Gerard, Craig; Thomas, Karen; Rallabhandi, Prasad; Shea-Donohue, Terez; Tamiz, Amir; Alkan, Sefik; Netzel-Arnett, Sarah; Antalis, Toni; Vogel, Stefanie N; Fasano, Alessio

2008-07-01

Celiac disease is an immune-mediated enteropathy triggered by gliadin, a component of the grain protein gluten. Gliadin induces an MyD88-dependent zonulin release that leads to increased intestinal permeability, a postulated early element in the pathogenesis of celiac disease. We aimed to establish the molecular basis of gliadin interaction with intestinal mucosa leading to intestinal barrier impairment. Alpha-gliadin affinity column was loaded with intestinal mucosal membrane lysates to identify the putative gliadin-binding moiety. In vitro experiments with chemokine receptor CXCR3 transfectants were performed to confirm binding of gliadin and/or 26 overlapping 20mer alpha-gliadin synthetic peptides to the receptor. CXCR3 protein and gene expression were studied in intestinal epithelial cell lines and human biopsy specimens. Gliadin-CXCR3 interaction was further analyzed by immunofluorescence microscopy, laser capture microscopy, real-time reverse-transcription polymerase chain reaction, and immunoprecipitation/Western blot analysis. Ex vivo experiments were performed using C57BL/6 wild-type and CXCR3(-/-) mouse small intestines to measure intestinal permeability and zonulin release. Affinity column and colocalization experiments showed that gliadin binds to CXCR3 and that at least 2 alpha-gliadin 20mer synthetic peptides are involved in this binding. CXCR3 is expressed in mouse and human intestinal epithelia and lamina propria. Mucosal CXCR3 expression was elevated in active celiac disease but returned to baseline levels following implementation of a gluten-free diet. Gliadin induced physical association between CXCR3 and MyD88 in enterocytes. Gliadin increased zonulin release and intestinal permeability in wild-type but not CXCR3(-/-) mouse small intestine. Gliadin binds to CXCR3 and leads to MyD88-dependent zonulin release and increased intestinal permeability.

IFT88 influences chondrocyte actin organization and biomechanics.

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Wang, Z; Wann, A K T; Thompson, C L; Hassen, A; Wang, W; Knight, M M

2016-03-01

Primary cilia are microtubule based organelles which control a variety of signalling pathways important in cartilage development, health and disease. This study examines the role of the intraflagellar transport (IFT) protein, IFT88, in regulating fundamental actin organisation and mechanics in articular chondrocytes. The study used an established chondrocyte cell line with and without hypomorphic mutation of IFT88 (IFT88(orpk)). Confocal microscopy was used to quantify F-actin and myosin IIB organisation. Viscoelastic cell and actin cortex mechanics were determined using micropipette aspiration with actin dynamics visualised in live cells transfected with LifeACT-GFP. IFT88(orpk) cells exhibited a significant increase in acto-myosin stress fibre organisation relative to wild-type (WT) cells in monolayer and an altered response to cytochalasin D. Rounded IFT88(orpk) cells cultured in suspension exhibited reduced cortical actin expression with reduced cellular equilibrium modulus. Micropipette aspiration resulted in reduced membrane bleb formation in IFT88(orpk) cells. Following membrane blebbing, IFT88(orpk) cells exhibited slower reformation of the actin cortex. IFT88(orpk) cells showed increased actin deformability and reduced cortical tension confirming that IFT regulates actin cortex mechanics. The reduced cortical tension is also consistent with the reduced bleb formation. This study demonstrates for the first time that the ciliary protein IFT88 regulates fundamental actin organisation and the stiffness of the actin cortex leading to alterations in cell deformation, mechanical properties and blebbing in an IFT88 chondrocyte cell line. This adds to the growing understanding of the role of primary cilia and IFT in regulating cartilage biology. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Arctigenin inhibits osteoclast differentiation and function by suppressing both calcineurin-dependent and osteoblastic cell-dependent NFATc1 pathways.

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Yamashita, Teruhito; Uehara, Shunsuke; Udagawa, Nobuyuki; Li, Feng; Kadota, Shigetoshi; Esumi, Hiroyasu; Kobayashi, Yasuhiro; Takahashi, Naoyuki

2014-01-01

Arctigenin, a lignan-derived compound, is a constituent of the seeds of Arctium lappa. Arctigenin was previously shown to inhibit osteoclastogenesis; however, this inhibitory mechanism has yet to be elucidated. Here, we showed that arctigenin inhibited the action of nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), a key transcription factor for osteoclastogenesis. NFATc1 in osteoclast precursors was activated through two distinct pathways: the calcineurin-dependent and osteoblastic cell-dependent pathways. Among the several lignan-derived compounds examined, arctigenin most strongly inhibited receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclast-like cell formation in mouse bone marrow macrophage (BMM) cultures, in which the calcineurin-dependent NFATc1 pathway was activated. Arctigenin suppressed neither the activation of nuclear factor κB and mitogen-activated protein kinases nor the up-regulation of c-Fos expression in BMMs treated with RANKL. However, arctigenin suppressed RANKL-induced NFATc1 expression. Interestingly, the treatment of osteoclast-like cells with arctigenin converted NFATc1 into a lower molecular weight species, which was translocated into the nucleus even in the absence of RANKL. Nevertheless, arctigenin as well as cyclosporin A (CsA), a calcineurin inhibitor, suppressed the NFAT-luciferase reporter activity induced by ionomycin and phorbol 12-myristate 13-acetate in BMMs. Chromatin immunoprecipitation analysis confirmed that arctigenin inhibited the recruitment of NFATc1 to the promoter region of the NFATc1 target gene. Arctigenin, but not CsA suppressed osteoclast-like cell formation in co-cultures of osteoblastic cells and bone marrow cells, in which the osteoblastic cell-dependent NFATc1 pathway was activated. The forced expression of constitutively active NFATc1 rescued osteoclastogenesis in BMM cultures treated with CsA, but not that treated with arctigenin. Arctigenin also suppressed the pit

Arctigenin Inhibits Osteoclast Differentiation and Function by Suppressing Both Calcineurin-Dependent and Osteoblastic Cell-Dependent NFATc1 Pathways

Science.gov (United States)

Yamashita, Teruhito; Uehara, Shunsuke; Udagawa, Nobuyuki; Li, Feng; Kadota, Shigetoshi; Esumi, Hiroyasu; Kobayashi, Yasuhiro; Takahashi, Naoyuki

2014-01-01

Arctigenin, a lignan-derived compound, is a constituent of the seeds of Arctium lappa. Arctigenin was previously shown to inhibit osteoclastogenesis; however, this inhibitory mechanism has yet to be elucidated. Here, we showed that arctigenin inhibited the action of nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), a key transcription factor for osteoclastogenesis. NFATc1 in osteoclast precursors was activated through two distinct pathways: the calcineurin-dependent and osteoblastic cell-dependent pathways. Among the several lignan-derived compounds examined, arctigenin most strongly inhibited receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclast-like cell formation in mouse bone marrow macrophage (BMM) cultures, in which the calcineurin-dependent NFATc1 pathway was activated. Arctigenin suppressed neither the activation of nuclear factor κB and mitogen-activated protein kinases nor the up-regulation of c-Fos expression in BMMs treated with RANKL. However, arctigenin suppressed RANKL-induced NFATc1 expression. Interestingly, the treatment of osteoclast-like cells with arctigenin converted NFATc1 into a lower molecular weight species, which was translocated into the nucleus even in the absence of RANKL. Nevertheless, arctigenin as well as cyclosporin A (CsA), a calcineurin inhibitor, suppressed the NFAT-luciferase reporter activity induced by ionomycin and phorbol 12-myristate 13-acetate in BMMs. Chromatin immunoprecipitation analysis confirmed that arctigenin inhibited the recruitment of NFATc1 to the promoter region of the NFATc1 target gene. Arctigenin, but not CsA suppressed osteoclast-like cell formation in co-cultures of osteoblastic cells and bone marrow cells, in which the osteoblastic cell-dependent NFATc1 pathway was activated. The forced expression of constitutively active NFATc1 rescued osteoclastogenesis in BMM cultures treated with CsA, but not that treated with arctigenin. Arctigenin also suppressed the pit

Arctigenin inhibits osteoclast differentiation and function by suppressing both calcineurin-dependent and osteoblastic cell-dependent NFATc1 pathways.

Directory of Open Access Journals (Sweden)

Teruhito Yamashita

Full Text Available Arctigenin, a lignan-derived compound, is a constituent of the seeds of Arctium lappa. Arctigenin was previously shown to inhibit osteoclastogenesis; however, this inhibitory mechanism has yet to be elucidated. Here, we showed that arctigenin inhibited the action of nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1, a key transcription factor for osteoclastogenesis. NFATc1 in osteoclast precursors was activated through two distinct pathways: the calcineurin-dependent and osteoblastic cell-dependent pathways. Among the several lignan-derived compounds examined, arctigenin most strongly inhibited receptor activator of nuclear factor κB ligand (RANKL-induced osteoclast-like cell formation in mouse bone marrow macrophage (BMM cultures, in which the calcineurin-dependent NFATc1 pathway was activated. Arctigenin suppressed neither the activation of nuclear factor κB and mitogen-activated protein kinases nor the up-regulation of c-Fos expression in BMMs treated with RANKL. However, arctigenin suppressed RANKL-induced NFATc1 expression. Interestingly, the treatment of osteoclast-like cells with arctigenin converted NFATc1 into a lower molecular weight species, which was translocated into the nucleus even in the absence of RANKL. Nevertheless, arctigenin as well as cyclosporin A (CsA, a calcineurin inhibitor, suppressed the NFAT-luciferase reporter activity induced by ionomycin and phorbol 12-myristate 13-acetate in BMMs. Chromatin immunoprecipitation analysis confirmed that arctigenin inhibited the recruitment of NFATc1 to the promoter region of the NFATc1 target gene. Arctigenin, but not CsA suppressed osteoclast-like cell formation in co-cultures of osteoblastic cells and bone marrow cells, in which the osteoblastic cell-dependent NFATc1 pathway was activated. The forced expression of constitutively active NFATc1 rescued osteoclastogenesis in BMM cultures treated with CsA, but not that treated with arctigenin. Arctigenin also suppressed the

A gene expression signature of RAS pathway dependence predicts response to PI3K and RAS pathway inhibitors and expands the population of RAS pathway activated tumors.

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Loboda, Andrey; Nebozhyn, Michael; Klinghoffer, Rich; Frazier, Jason; Chastain, Michael; Arthur, William; Roberts, Brian; Zhang, Theresa; Chenard, Melissa; Haines, Brian; Andersen, Jannik; Nagashima, Kumiko; Paweletz, Cloud; Lynch, Bethany; Feldman, Igor; Dai, Hongyue; Huang, Pearl; Watters, James

2010-06-30

Hyperactivation of the Ras signaling pathway is a driver of many cancers, and RAS pathway activation can predict response to targeted therapies. Therefore, optimal methods for measuring Ras pathway activation are critical. The main focus of our work was to develop a gene expression signature that is predictive of RAS pathway dependence. We used the coherent expression of RAS pathway-related genes across multiple datasets to derive a RAS pathway gene expression signature and generate RAS pathway activation scores in pre-clinical cancer models and human tumors. We then related this signature to KRAS mutation status and drug response data in pre-clinical and clinical datasets. The RAS signature score is predictive of KRAS mutation status in lung tumors and cell lines with high (> 90%) sensitivity but relatively low (50%) specificity due to samples that have apparent RAS pathway activation in the absence of a KRAS mutation. In lung and breast cancer cell line panels, the RAS pathway signature score correlates with pMEK and pERK expression, and predicts resistance to AKT inhibition and sensitivity to MEK inhibition within both KRAS mutant and KRAS wild-type groups. The RAS pathway signature is upregulated in breast cancer cell lines that have acquired resistance to AKT inhibition, and is downregulated by inhibition of MEK. In lung cancer cell lines knockdown of KRAS using siRNA demonstrates that the RAS pathway signature is a better measure of dependence on RAS compared to KRAS mutation status. In human tumors, the RAS pathway signature is elevated in ER negative breast tumors and lung adenocarcinomas, and predicts resistance to cetuximab in metastatic colorectal cancer. These data demonstrate that the RAS pathway signature is superior to KRAS mutation status for the prediction of dependence on RAS signaling, can predict response to PI3K and RAS pathway inhibitors, and is likely to have the most clinical utility in lung and breast tumors.

A gene expression signature of RAS pathway dependence predicts response to PI3K and RAS pathway inhibitors and expands the population of RAS pathway activated tumors

Directory of Open Access Journals (Sweden)

Paweletz Cloud

2010-06-01

Full Text Available Abstract Background Hyperactivation of the Ras signaling pathway is a driver of many cancers, and RAS pathway activation can predict response to targeted therapies. Therefore, optimal methods for measuring Ras pathway activation are critical. The main focus of our work was to develop a gene expression signature that is predictive of RAS pathway dependence. Methods We used the coherent expression of RAS pathway-related genes across multiple datasets to derive a RAS pathway gene expression signature and generate RAS pathway activation scores in pre-clinical cancer models and human tumors. We then related this signature to KRAS mutation status and drug response data in pre-clinical and clinical datasets. Results The RAS signature score is predictive of KRAS mutation status in lung tumors and cell lines with high (> 90% sensitivity but relatively low (50% specificity due to samples that have apparent RAS pathway activation in the absence of a KRAS mutation. In lung and breast cancer cell line panels, the RAS pathway signature score correlates with pMEK and pERK expression, and predicts resistance to AKT inhibition and sensitivity to MEK inhibition within both KRAS mutant and KRAS wild-type groups. The RAS pathway signature is upregulated in breast cancer cell lines that have acquired resistance to AKT inhibition, and is downregulated by inhibition of MEK. In lung cancer cell lines knockdown of KRAS using siRNA demonstrates that the RAS pathway signature is a better measure of dependence on RAS compared to KRAS mutation status. In human tumors, the RAS pathway signature is elevated in ER negative breast tumors and lung adenocarcinomas, and predicts resistance to cetuximab in metastatic colorectal cancer. Conclusions These data demonstrate that the RAS pathway signature is superior to KRAS mutation status for the prediction of dependence on RAS signaling, can predict response to PI3K and RAS pathway inhibitors, and is likely to have the most clinical

density-dependent selection revisited

Indian Academy of Sciences (India)

Unknown

is a more useful way of looking at density-dependent selection, and then go on ... these models was that the condition for maintenance of ... In a way, their formulation may be viewed as ... different than competition among species, and typical.

Size- and charge selectivity of glomerular filtration in Type 1 (insulin-dependent) diabetic patients with and without albuminuria

DEFF Research Database (Denmark)

Deckert, T; Kofoed-Enevoldsen, A; Vidal, P

1993-01-01

Albuminuria is the first clinical event in the development of diabetic nephropathy. We assessed glomerular charge- and size selectivity in 51 patients with Type 1 (insulin-dependent) diabetes mellitus of juvenile onset and 11 healthy individuals. Patients were allocated to five groups. The urinary...... techniques and tubular protein reabsorption by excretion of beta 2-microglobulin. Charge selectivity was estimated from the IgG/IgG4 selectivity index. Size selectivity was measured by dextran clearance. Dextran was measured by refractive index detection after fractionation (2 A fractions in the range 26...... macromolecular pathways in the development of diabetic nephropathy....

Involvement of caspase-dependent and -independent apoptotic pathways in cisplatin-induced apoptosis

Science.gov (United States)

Liu, Lei; Zhang, Yingjie; Wang, Xianwang

2009-02-01

Cisplatin, an efficient anticancer agent, can trigger multiple apoptotic pathways in cancer cells. However, the signal transduction pathways in response to cisplatin-based chemotherapy are complicated, and the mechanism is not fully understood. In current study, we showed that, during cisplatin-induced apoptosis of human lung adenocarcinoma cells, both the caspase-dependent and -independent pathways were activated. Herein, we reported that after cisplatin treatment, the activities of caspase-9/-3 were sharply increased; pre-treatment with Z-LEHD-fmk (inhibitor of caspase-9), Z-DEVD-fmk (inhibitor of caspase-3), and Z-VAD-fmk (a pan-caspase inhibitor) increased cell viability and decreased apoptosis, suggesting that caspase-mediated apoptotic pathway was activated following cisplatin treatment. Confocal imaging of the cells transfected with AIF-GFP demonstrated that AIF release occurred about 9 h after cisplatin treatment. The event proceeded progressively over time, coinciding with a nuclear translocation and lasting for more than 2 hours. Down-regulation of AIF by siRNA also significantly increased cell viability and decreased apoptosis, these results suggested that AIF-mediated caspase-independent apoptotic pathway was involved in cispatin-induced apoptosis. In conclusion, the current study demonstrated that both caspase-dependent and -independent apoptotic pathways were involved in cisplatin-induced apoptosis in human lung adenocarcinoma cells.

Tenebrio molitor Gram-negative-binding protein 3 (TmGNBP3) is essential for inducing downstream antifungal Tenecin 1 gene expression against infection with Beauveria bassiana JEF-007.

Science.gov (United States)

Yang, Yi-Ting; Lee, Mi Rong; Lee, Se Jin; Kim, Sihyeon; Nai, Yu-Shin; Kim, Jae Su

2017-05-23

The Toll signaling pathway is responsible for defense against both Gram-positive bacteria and fungi. Gram-negative binding protein 3 (GNBP3) has a strong affinity for the fungal cell wall component, β-1,3-glucan, which can activate the prophenoloxidase (proPO) cascade and induce the Toll signaling pathway. Myeloid differentiation factor 88 (MyD88) is an intracellular adaptor protein involved in the Toll signaling pathway. In this study, we monitored the response of 5 key genes (TmGNBP3, TmMyD88, and Tenecin 1, 2, and 3) in the Toll pathway of the mealworm Tenebrio molitor immune system against the fungus Beauveria bassiana JEF-007 using RT-PCR. TmGNBP3, Tenecin 1, and Tenecin 2 were significantly upregulated after fungal infection. To better understand the roles of the Toll signaling pathway in the mealworm immune system, TmGNBP3 and TmMyD88 were knocked down by RNAi silencing. Target gene expression levels decreased at 2 d postknockdown and were dramatically reduced at 6 d post-dsRNA injection. Therefore, mealworms were compromised by B. bassiana JEF-007 at 6 d post-dsRNA injection. Silencing of TmMyD88 and TmGNBP3 resulted in reduced resistance of the host to fungal infection. Particularly, reducing TmGNBP3 levels obviously downregulated Tenecin 1 and Tenecin 2 expression levels, whereas silencing TmMyD88 expression resulted in decreased Tenecin 2 expression. These results indicate that TmGNBP3 is essential to induce downstream antifungal peptide Tenecin 1 expression against B. bassiana JEF-007. © 2017 Institute of Zoology, Chinese Academy of Sciences.

Cyclooxygenase-2 Selectively Controls Renal Blood Flow Through a Novel PPARβ/δ-Dependent Vasodilator Pathway.

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Kirkby, Nicholas S; Sampaio, Walkyria; Etelvino, Gisele; Alves, Daniele T; Anders, Katie L; Temponi, Rafael; Shala, Fisnik; Nair, Anitha S; Ahmetaj-Shala, Blerina; Jiao, Jing; Herschman, Harvey R; Xiaomeng, Wang; Wahli, Walter; Santos, Robson A; Mitchell, Jane A

2018-02-01

Cyclooxygenase-2 (COX-2) is an inducible enzyme expressed in inflammation and cancer targeted by nonsteroidal anti-inflammatory drugs. COX-2 is also expressed constitutively in discreet locations where its inhibition drives gastrointestinal and cardiovascular/renal side effects. Constitutive COX-2 expression in the kidney regulates renal function and blood flow; however, the global relevance of the kidney versus other tissues to COX-2-dependent blood flow regulation is not known. Here, we used a microsphere deposition technique and pharmacological COX-2 inhibition to map the contribution of COX-2 to regional blood flow in mice and compared this to COX-2 expression patterns using luciferase reporter mice. Across all tissues studied, COX-2 inhibition altered blood flow predominantly in the kidney, with some effects also seen in the spleen, adipose, and testes. Of these sites, only the kidney displayed appreciable local COX-2 expression. As the main site where COX-2 regulates blood flow, we next analyzed the pathways involved in kidney vascular responses using a novel technique of video imaging small arteries in living tissue slices. We found that the protective effect of COX-2 on renal vascular function was associated with prostacyclin signaling through PPARβ/δ (peroxisome proliferator-activated receptor-β/δ). These data demonstrate the kidney as the principle site in the body where local COX-2 controls blood flow and identifies a previously unreported PPARβ/δ-mediated renal vasodilator pathway as the mechanism. These findings have direct relevance to the renal and cardiovascular side effects of drugs that inhibit COX-2, as well as the potential of the COX-2/prostacyclin/PPARβ/δ axis as a therapeutic target in renal disease. © 2018 The Authors.

Brain Pathways to Recovery from Alcohol Dependence

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Cui, Changhai; Noronha, Antonio; Warren, Kenneth; Koob, George F.; Sinha, Rajita; Thakkar, Mahesh; Matochik, John; Crews, Fulton T.; Chandler, L. Judson; Pfefferbaum, Adolf; Becker, Howard C.; Lovinger, David; Everitt, Barry; Egli, Mark; Mandyam, Chitra; Fein, George; Potenza, Marc N.; Harris, R. Adron; Grant, Kathleen A.; Roberto, Marisa; Meyerhoff, Dieter J.; Sullivan, Edith V.

2015-01-01

This article highlights the research presentations at the satellite symposium on “Brain Pathways to Recovery from Alcohol Dependence” held at the 2013 Society for Neuroscience Annual Meeting. The purpose of this symposium was to provide an up to date overview of research efforts focusing on understanding brain mechanisms that contribute to recovery from alcohol dependence. A panel of scientists from the alcohol and addiction research field presented their insights and perspectives on brain mechanisms that may underlie both recovery and lack of recovery from alcohol dependence. The four sessions of the symposium encompassed multilevel studies exploring mechanisms underlying relapse and craving associated with sustained alcohol abstinence, cognitive function deficit and recovery, and translational studies on preventing relapse and promoting recovery. Gaps in our knowledge and research opportunities were also discussed. PMID:26074423

Hydrogen peroxide induces activation of insulin signaling pathway via AMP-dependent kinase in podocytes

Energy Technology Data Exchange (ETDEWEB)

Piwkowska, Agnieszka, E-mail: apiwkowska@cmdik.pan.pl [Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdansk (Poland); Rogacka, Dorota; Angielski, Stefan [Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdansk (Poland); Jankowski, Maciej [Mossakowski Medical Research Centre, Polish Academy of Sciences, Laboratory of Molecular and Cellular Nephrology, Gdansk (Poland); Medical University of Gdansk, Department of Therapy Monitoring and Pharmacogenetics (Poland)

2012-11-09

Highlights: Black-Right-Pointing-Pointer H{sub 2}O{sub 2} activates the insulin signaling pathway and glucose uptake in podocytes. Black-Right-Pointing-Pointer H{sub 2}O{sub 2} induces time-dependent changes in AMPK phosphorylation. Black-Right-Pointing-Pointer H{sub 2}O{sub 2} enhances insulin signaling pathways via AMPK activation. Black-Right-Pointing-Pointer H{sub 2}O{sub 2} stimulation of glucose uptake is AMPK-dependent. -- Abstract: Podocytes are cells that form the glomerular filtration barrier in the kidney. Insulin signaling in podocytes is critical for normal kidney function. Insulin signaling is regulated by oxidative stress and intracellular energy levels. We cultured rat podocytes to investigate the effects of hydrogen peroxide (H{sub 2}O{sub 2}) on the phosphorylation of proximal and distal elements of insulin signaling. We also investigated H{sub 2}O{sub 2}-induced intracellular changes in the distribution of protein kinase B (Akt). Western blots showed that H{sub 2}O{sub 2} (100 {mu}M) induced rapid, transient phosphorylation of the insulin receptor (IR), the IR substrate-1 (IRS1), and Akt with peak activities at 5 min ({Delta} 183%, P < 0.05), 3 min ({Delta} 414%, P < 0.05), and 10 min ({Delta} 35%, P < 0.05), respectively. Immunostaining cells with an Akt-specific antibody showed increased intensity at the plasma membrane after treatment with H{sub 2}O{sub 2}>. Furthermore, H{sub 2}O{sub 2} inhibited phosphorylation of the phosphatase and tensin homologue (PTEN; peak activity at 10 min; {Delta} -32%, P < 0.05) and stimulated phosphorylation of the AMP-dependent kinase alpha subunit (AMPK{alpha}; 78% at 3 min and 244% at 10 min). The stimulation of AMPK was abolished with an AMPK inhibitor, Compound C (100 {mu}M, 2 h). Moreover, Compound C significantly reduced the effect of H{sub 2}O{sub 2} on IR phosphorylation by about 40% (from 2.07 {+-} 0.28 to 1.28 {+-} 0.12, P < 0.05). In addition, H{sub 2}O{sub 2} increased glucose uptake in podocytes

The Protective Effects of the Supercritical-Carbon Dioxide Fluid Extract of Chrysanthemum indicum against Lipopolysaccharide-Induced Acute Lung Injury in Mice via Modulating Toll-Like Receptor 4 Signaling Pathway

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Xiao-Li Wu

2014-01-01

Full Text Available The supercritical-carbon dioxide fluid extract of Chrysanthemum indicum Linné. (CFE has been demonstrated to be effective in suppressing inflammation. The aim of this study is to investigate the preventive action and underlying mechanisms of CFE on acute lung injury (ALI induced by lipopolysaccharide (LPS in mice. ALI was induced by intratracheal instillation of LPS into lung, and dexamethasone was used as a positive control. Results revealed that pretreatment with CFE abated LPS-induced lung histopathologic changes, reduced the wet/dry ratio and proinflammatory cytokines productions (TNF-α, IL-1β, and IL-6, inhibited inflammatory cells migrations and protein leakages, suppressed the levels of MPO and MDA, and upregulated the abilities of antioxidative enzymes (SOD, CAT, and GPx. Furthermore, the pretreatment with CFE downregulated the activations of NF-κB and the expressions of TLR4/MyD88. These results suggested that CFE exerted potential protective effects against LPS-induced ALI in mice and was a potential therapeutic drug for ALI. Its mechanisms were at least partially associated with the modulations of TLR4 signaling pathways.

Benchmarking pathway interaction network for colorectal cancer to identify dysregulated pathways

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Q. Wang

Full Text Available Different pathways act synergistically to participate in many biological processes. Thus, the purpose of our study was to extract dysregulated pathways to investigate the pathogenesis of colorectal cancer (CRC based on the functional dependency among pathways. Protein-protein interaction (PPI information and pathway data were retrieved from STRING and Reactome databases, respectively. After genes were aligned to the pathways, each pathway activity was calculated using the principal component analysis (PCA method, and the seed pathway was discovered. Subsequently, we constructed the pathway interaction network (PIN, where each node represented a biological pathway based on gene expression profile, PPI data, as well as pathways. Dysregulated pathways were then selected from the PIN according to classification performance and seed pathway. A PIN including 11,960 interactions was constructed to identify dysregulated pathways. Interestingly, the interaction of mRNA splicing and mRNA splicing-major pathway had the highest score of 719.8167. Maximum change of the activity score between CRC and normal samples appeared in the pathway of DNA replication, which was selected as the seed pathway. Starting with this seed pathway, a pathway set containing 30 dysregulated pathways was obtained with an area under the curve score of 0.8598. The pathway of mRNA splicing, mRNA splicing-major pathway, and RNA polymerase I had the maximum genes of 107. Moreover, we found that these 30 pathways had crosstalks with each other. The results suggest that these dysregulated pathways might be used as biomarkers to diagnose CRC.

Evolution of branched regulatory genetic pathways: directional selection on pleiotropic loci accelerates developmental system drift.

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Johnson, Norman A; Porter, Adam H

2007-01-01

Developmental systems are regulated by a web of interacting loci. One common and useful approach in studying the evolution of development is to focus on classes of interacting elements within these systems. Here, we use individual-based simulations to study the evolution of traits controlled by branched developmental pathways involving three loci, where one locus regulates two different traits. We examined the system under a variety of selective regimes. In the case where one branch was under stabilizing selection and the other under directional selection, we observed "developmental system drift": the trait under stabilizing selection showed little phenotypic change even though the loci underlying that trait showed considerable evolutionary divergence. This occurs because the pleiotropic locus responds to directional selection and compensatory mutants are then favored in the pathway under stabilizing selection. Though developmental system drift may be caused by other mechanisms, it seems likely that it is accelerated by the same underlying genetic mechanism as that producing the Dobzhansky-Muller incompatibilities that lead to speciation in both linear and branched pathways. We also discuss predictions of our model for developmental system drift and how different selective regimes affect probabilities of speciation in the branched pathway system.

Differential selection on carotenoid biosynthesis genes as a function of gene position in the metabolic pathway: a study on the carrot and dicots.

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Jérémy Clotault

Full Text Available Selection of genes involved in metabolic pathways could target them differently depending on the position of genes in the pathway and on their role in controlling metabolic fluxes. This hypothesis was tested in the carotenoid biosynthesis pathway using population genetics and phylogenetics.Evolutionary rates of seven genes distributed along the carotenoid biosynthesis pathway, IPI, PDS, CRTISO, LCYB, LCYE, CHXE and ZEP, were compared in seven dicot taxa. A survey of deviations from neutrality expectations at these genes was also undertaken in cultivated carrot (Daucus carota subsp. sativus, a species that has been intensely bred for carotenoid pattern diversification in its root during its cultivation history. Parts of sequences of these genes were obtained from 46 individuals representing a wide diversity of cultivated carrots. Downstream genes exhibited higher deviations from neutral expectations than upstream genes. Comparisons of synonymous and nonsynonymous substitution rates between genes among dicots revealed greater constraints on upstream genes than on downstream genes. An excess of intermediate frequency polymorphisms, high nucleotide diversity and/or high differentiation of CRTISO, LCYB1 and LCYE in cultivated carrot suggest that balancing selection may have targeted genes acting centrally in the pathway.Our results are consistent with relaxed constraints on downstream genes and selection targeting the central enzymes of the carotenoid biosynthesis pathway during carrot breeding history.

Regular Exercise Enhances the Immune Response Against Microbial Antigens Through Up-Regulation of Toll-like Receptor Signaling Pathways

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Qishi Zheng

2015-09-01

Full Text Available Background/Aims: Regular physical exercise can enhance resistance to many microbial infections. However, little is known about the mechanism underlying the changes in the immune system induced by regular exercise. Methods: We recruited members of a university badminton club as the regular exercise (RE group and healthy sedentary students as the sedentary control (SC group. We investigated the distribution of peripheral blood mononuclear cell (PBMC subsets and functions in the two groups. Results: There were no significant differences in plasma cytokine levels between the RE and SC groups in the true resting state. However, enhanced levels of IFN-γ, TNF-α, IL-6, IFN-α and IL-12 were secreted by PBMCs in the RE group following microbial antigen stimulation, when compared to the SC group. In contrast, the levels of TNF-α and IL-6 secreted by PBMC in the RE group were suppressed compared with those in SC group following non-microbial antigen stimulation (concanavalin A or α-galactosylceramide. Furthermore, PBMC expression of TLR2, TLR7 and MyD88 was significantly increased in the RE group in response to microbial antigen stimulation. Conclusion: Regular exercise enhances immune cell activation in response to pathogenic stimulation leading to enhanced cytokine production mediated via the TLR signaling pathways.

Hyperglycemia Induces Toll-Like Receptor-2 and -4 Expression and Activity in Human Microvascular Retinal Endothelial Cells: Implications for Diabetic Retinopathy

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Uthra Rajamani

2014-01-01

Full Text Available Diabetic retinopathy (DR causes visual impairment in working age adults and hyperglycemia-mediated inflammation is central in DR. Toll-like receptors (TLRs play a key role in innate immune responses and inflammation. However, scanty data is available on their role in DR. Hence, in this study, we examined TLR2 and TLR4 mRNA and protein expression and activity in hyperglycemic human retinal endothelial cells (HMVRECs. HMVRECs were treated with hyperglycemia (HG or euglycemia and mRNA and protein levels of TLR-2, TLR-4, MyD88, IRF3, and TRIF as well as NF-κB p65 activation were measured. IL-8, IL-1β, TNF-α and MCP-1, ICAM-1, and VCAM-1 as well as monocyte adhesion to HMVRECs were also assayed. HG (25 mM significantly induced TLR2 and TLR4 mRNA and protein in HMVRECs. It also increased both MyD88 and non-MyD88 pathways, nuclear factor-κB (NF-κB, biomediators, and monocyte adhesion. This inflammation was attenuated by TLR-4 or TLR-2 inhibition, and dual inhibition by a TLR inhibitory peptide as well as TLR2 and 4 siRNA. Additionally, antioxidant treatment reduced TLR-2 and TLR4 expression and downstream inflammatory markers. Collectively, our novel data suggest that hyperglycemia induces TLR-2 and TLR-4 activation and downstream signaling mediating increased inflammation possibly via reactive oxygen species (ROS and could contribute to DR.

Polysaccharide from Lentinus edodes inhibits the immunosuppressive function of myeloid-derived suppressor cells.

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Hao Wu

Full Text Available Reversing the function of immune suppressor cells may improve the efficacy of cancer therapy. Here, we have isolated a novel polysaccharide MPSSS (577.2 Kd from Lentinus edodes and examined its effects on differentiation and function of myeloid-derived suppressor cells (MDSCs. MPSSS is composed of glucose (75.0%, galactose (11.7%, mannose (7.8%, and xylose (0.4%. In vivo, it inhibits the growth of McgR32 tumor cells, which is correlated with a reduced percentage of MDSCs in peripheral blood. In vitro, it induces both morphological and biophysical changes in MDSCs. Importantly, MPSSS up-regulates MHC II and F4/80 expression on MDSCs, and reverses their inhibition effect on CD4(+ T cells in a dose-dependent manner. The mechanism study shows that MPSSS may stimulate MDSCs through a MyD88 dependent NF-κB signaling pathway. Together, we demonstrated for the first time that MPSSS stimulates the differentiation of MDSCs and reverses its immunosuppressive functions, shedding new light on developing novel anti-cancer strategies by targeting MDSCs.

Toll-like receptor 7 cooperates with IL-4 in activated B cells through antigen receptor or CD38 and induces class switch recombination and IgG1 production.

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Tsukamoto, Yumiko; Nagai, Yoshinori; Kariyone, Ai; Shibata, Takuma; Kaisho, Tsuneyasu; Akira, Shizuo; Miyake, Kensuke; Takatsu, Kiyoshi

2009-04-01

IL-4 and 8-mercaptoguanosine (8-SGuo) stimulation of CD38-activated B cells induces mu to gamma1 class switch recombination (CSR) at the DNA level leading to a high level of IgG1 production. Although some of signaling events initiated by IL-4 in activated B cells have been characterized, the involvement of TLR/MyD88 and Btk pathway in IL-4-dependent mu to gamma1 CSR has not been thoroughly evaluated. In this study, we characterized receptors for 8-SGuo and differential roles of 8-SGuo and IL-4 in the induction and mu to gamma1 CSR and IgG1 production. The role of TLR7 and MyD88 in 8-SGuo-induced AID expression and mu to gamma1 CSR was documented, as 8-SGuo did not act on CD38-stimulated splenic B cells from Tlr7(-/-) and Myd88(-/-) mice. CD38-activated B cells from Btk-deficient mice failed to respond to TLR7 ligands for the AID expression and CSR, indicating that Btk is also indispensable for the system. Stimulation of CD38-activated B cells with 8-SGuo induced significant AID expression and DNA double strand breaks, but IL-4 stimulation by itself did not trigger mu to gamma1 CSR. Intriguingly, the mu to gamma1 CSR in the B cells stimulated with CD38 and 8-SGuo totally depends on IL-4 stimulation. Similar results were obtained in the activated B cells through BCR and loxoribine, a well-known TLR7 ligand, in place of 8-SGuo. In vivo administration of TLR7 ligand and anti-CD38 antibody induced the generation of CD138(+) IgG1(+) cells. These results indicate that TLR7 is a receptor for 8-SGuo and plays an essential role in the AID and Blimp-1 expression; however it is not enough to complete mu to gamma1 CSR in CD38-activated B cells. IL-4 may be required for the induction of DNA repair system together with AID for the completion of CSR.

Dependency of climate change and carbon cycle on CO2 emission pathways

International Nuclear Information System (INIS)

Nohara, Daisuke; Yoshida, Yoshikatsu; Misumi, Kazuhiro; Ohba, Masamichi

2013-01-01

Previous research has indicated that the response of globally average temperature is approximately proportional to cumulative CO 2 emissions, yet evidence of the robustness of this relationship over a range of CO 2 emission pathways is lacking. To address this, we evaluate the dependency of climate and carbon cycle change on CO 2 emission pathways using a fully coupled climate–carbon cycle model. We design five idealized pathways (including an overshoot scenario for cumulative emissions), each of which levels off to final cumulative emissions of 2000 GtC. The cumulative emissions of the overshoot scenario reach 4000 GtC temporarily, subsequently reducing to 2000 GtC as a result of continuous negative emissions. Although we find that responses of climatic variables and the carbon cycle are largely independent of emission pathways, a much weakened Atlantic meridional overturning circulation (AMOC) is projected in the overshoot scenario despite cessation of emissions. This weakened AMOC is enhanced by rapid warming in the Arctic region due to considerable temporary elevation of atmospheric CO 2 concentration and induces the decline of surface air temperature and decrease of precipitation over the northern Atlantic and Europe region. Moreover, the weakened AMOC reduces CO 2 uptake by the Atlantic and Arctic oceans. However, the weakened AMOC contributes little to the global carbon cycle. In conclusion, although climate variations have been found to be dependent on emission pathways, the global carbon cycle is relatively independent of these emission pathways, at least superficially. (letter)

Temperature dependence of muonium spin exchange with O2 in the range 88 K to 478 K

International Nuclear Information System (INIS)

Senba, M.; Garner, D.M.; Arseneau, D.J.; Fleming, D.G.

1984-01-01

The authors have extended an earlier study of the spin exchange reactions of Mu with O 2 in the range 295 K to 478 K, to a low temperature region down to 88 K. From 135 K to 296 K, the spin depolarization rate constant was found to vary according to the relative velocity of the colliding species, which indicates that the spin exchange cross section of Mu-O 2 is temperature independent in this range. However, it was found that below 105 K and above 400 K, the spin depolarization rate constant tends to have stronger temperature dependences. (Auth.)

Flavivirus internalization is regulated by a size-dependent endocytic pathway.

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Hackett, Brent A; Cherry, Sara

2018-04-17

Flaviviruses enter host cells through the process of clathrin-mediated endocytosis, and the spectrum of host factors required for this process are incompletely understood. Here we found that lymphocyte antigen 6 locus E (LY6E) promotes the internalization of multiple flaviviruses, including West Nile virus, Zika virus, and dengue virus. Perhaps surprisingly, LY6E is dispensable for the internalization of the endogenous cargo transferrin, which is also dependent on clathrin-mediated endocytosis for uptake. Since viruses are substantially larger than transferrin, we reasoned that LY6E may be required for uptake of larger cargoes and tested this using transferrin-coated beads of similar size as flaviviruses. LY6E was indeed required for the internalization of transferrin-coated beads, suggesting that LY6E is selectively required for large cargo. Cell biological studies found that LY6E forms tubules upon viral infection and bead internalization, and we found that tubule formation was dependent on RNASEK, which is also required for flavivirus internalization, but not transferrin uptake. Indeed, we found that RNASEK is also required for the internalization of transferrin-coated beads, suggesting it functions upstream of LY6E. These LY6E tubules resembled microtubules, and we found that microtubule assembly was required for their formation and flavivirus uptake. Since microtubule end-binding proteins link microtubules to downstream activities, we screened the three end-binding proteins and found that EB3 promotes virus uptake and LY6E tubularization. Taken together, these results highlight a specialized pathway required for the uptake of large clathrin-dependent endocytosis cargoes, including flaviviruses. Copyright © 2018 the Author(s). Published by PNAS.

Phase-dependent quantum interference between different pathways in bichromatic harmonic generation

International Nuclear Information System (INIS)

Jun, Cai; Li-Ming, Wang; Hao-Xue, Qiao

2009-01-01

This paper studies the harmonic generation of the hydrogen atom subjected to a collinear bichromatic laser field by numerically solving the time-dependent Schrödinger equation using the split-operator pseudo-spectral method. By adding a frequency variation to the additional field, the contributions of different pathways to particular order harmonic generation can be isolated. The quantum interference pattern between harmonic pathways, which influences the harmonic intensity, is found to be either constructive or destructive with respect to different relative phase of the two field components. Detailed description of up to the 35th-order harmonics and the harmonic pathways for a wide range of field parameters is presented. (atomic and molecular physics)

Revealing mechanisms of selective, concentration-dependent potentials of 4-hydroxy-2-nonenal to induce apoptosis in cancer cells through inactivation of membrane-associated catalase.

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Bauer, Georg; Zarkovic, Neven

2015-04-01

Tumor cells generate extracellular superoxide anions and are protected against superoxide anion-mediated intercellular apoptosis-inducing signaling by the expression of membrane-associated catalase. 4-Hydroxy-2-nonenal (4-HNE), a versatile second messenger generated during lipid peroxidation, has been shown to induce apoptosis selectively in malignant cells. The findings described in this paper reveal the strong, concentration-dependent potential of 4-HNE to specifically inactivate extracellular catalase of tumor cells both indirectly and directly and to consequently trigger apoptosis in malignant cells through superoxide anion-mediated intercellular apoptosis-inducing signaling. Namely, 4-HNE caused apoptosis selectively in NOX1-expressing tumor cells through inactivation of their membrane-associated catalase, thus reactivating subsequent intercellular signaling through the NO/peroxynitrite and HOCl pathways, followed by the mitochondrial pathway of apoptosis. Concentrations of 4-HNE of 1.2 µM and higher directly inactivated membrane-associated catalase of tumor cells, whereas at lower concentrations, 4-HNE triggered a complex amplificatory pathway based on initial singlet oxygen formation through H2O2 and peroxynitrite interaction. Singlet-oxygen-dependent activation of the FAS receptor and caspase-8 increased superoxide anion generation by NOX1 and amplification of singlet oxygen generation, which allowed singlet-oxygen-dependent inactivation of catalase. 4-HNE and singlet oxygen cooperate in complex autoamplificatory loops during this process. The finding of these novel anticancer pathways may be useful for understanding the role of 4-HNE in the control of malignant cells and for the optimization of ROS-dependent therapeutic approaches including antioxidant treatments. Copyright © 2015 Elsevier Inc. All rights reserved.

'CM 88' - A multiple disease resistant chickpea mutant variety

International Nuclear Information System (INIS)

Haq, M.A.; Hassan, Mahmudul; Sadiq, M.

2001-01-01

Full text: Chickpea is the most important grain legume crop of Pakistan. Ascochyta blight (Ascochyta rabiei) and Fusarium wilt (Fusarium oxysporum F. sp cicer) are most serious diseases, having the potential to devastate a crop. A multiple disease resistant and high yielding mutant CM 88 has been developed through 100 Gy gamma irradiation treatment of variety 'C 727'. This was once a widely grown and popular variety, which lost its resistance to Ascochyta and was replaced. The selection of mutants was performed in the M2 generation grown in the Ascochyta blight nursery and sixteen mutants were selected. In the subsequent generations CM 88 proved resistant to both Ascochyta blight and Fusarium wilt, and exhibited superiority in agronomic characteristics. CM 88 was also tested for many years in the various yield trials on research stations and farmers fields throughout the country. In these trials it out yielded both the parent and standard varieties. The mutant CM 88 has been approved by the Punjab Seed Council on 27 October 1994 for general cultivation in the Punjab Province, especially the Thal area which accounts for more than 70% of the area under chickpea cultivation. (author)

Lipids Derived from Virulent Francisella tularensis Broadly Inhibit Pulmonary Inflammation via Toll-Like Receptor 2 and Peroxisome Proliferator-Activated Receptor α

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Crane, Deborah D.; Ireland, Robin; Alinger, Joshua B.; Small, Pamela

2013-01-01

Francisella tularensis is a Gram-negative facultative intracellular pathogen that causes an acute lethal respiratory disease in humans. The heightened virulence of the pathogen is linked to its unique ability to inhibit Toll-like receptor (TLR)-mediated inflammatory responses. The bacterial component and mechanism of this inhibition are unknown. Here we show that lipids isolated from virulent but not attenuated strains of F. tularensis are not detected by host cells, inhibit production of proinflammatory cytokines by primary macrophages in response to known TLR ligands, and suppress neutrophil recruitment in vivo. We further show that lipid-mediated inhibition of inflammation is dependent on TLR2, MyD88, and the nuclear hormone and fatty acid receptor peroxisome proliferator-activated receptor α (PPARα). Pathogen lipid-mediated interference with inflammatory responses through the engagement of TLR2 and PPARα represents a novel manipulation of host signaling pathways consistent with the ability of highly virulent F. tularensis to efficiently evade host immune responses. PMID:23925884

Ethnicity-dependent influence of innate immune genetic markers on morphine PCA requirements and adverse effects in postoperative pain.

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Somogyi, Andrew A; Sia, Alex T; Tan, Ene-Choo; Coller, Janet K; Hutchinson, Mark R; Barratt, Daniel T

2016-11-01

Although several genetic factors have been associated with postsurgical morphine requirements, those involving the innate immune system and cytokines have not been well investigated. The aim of this study was to investigate the contribution of genetic variability in innate immune signalling pathways to variability in morphine dosage after elective caesarean section under spinal anaesthesia in 133 Indian, 230 Malay, and 598 Han Chinese women previously studied. Twenty single nucleotide polymorphisms in 14 genes involved in glial activation (TLR2, TLR4, MYD88, MD2), inflammatory signalling (IL2, IL6, IL10, IL1B, IL6R, TNFA, TGFB1, CRP, CASP1), and neuronal regulation (BDNF) were newly investigated, in addition to OPRM1, COMT, and ABCB1 genetic variability identified previously. Postsurgical patient-controlled analgesia morphine use (mg/24 hours) was binned into 6 normally distributed groups and scored 0 to 5 to facilitate step-down multiple linear regression analysis of genetic predictors, controlling for ethnicity and nongenetic variables. Ethnicity, OPRM1 rs1799971 (increased), TLR2 rs3804100 (decreased), and an interaction between ethnicity and IL1B rs1143634 (increased), predicted 9.8% of variability in morphine use scores in the entire cohort. In the Indian cohort, 14.5% of the variance in morphine use score was explained by IL1B rs1143634 (increased) and TGFB1 rs1800469 (decreased). In Chinese patients, the incidence of postsurgical pain was significantly higher in variant COMT rs4680 genotypes (P = 0.0007) but not in the Malay or Indian cohorts. Innate immune genetics may contribute to variability in postsurgical opioid requirements in an ethnicity-dependent manner.

Human cerebral venous outflow pathway depends on posture and central venous pressure

DEFF Research Database (Denmark)

Gisolf, J; van Lieshout, J J; van Heusden, K

2004-01-01

Internal jugular veins are the major cerebral venous outflow pathway in supine humans. In upright humans the positioning of these veins above heart level causes them to collapse. An alternative cerebral outflow pathway is the vertebral venous plexus. We set out to determine the effect of posture...... and during a Valsalva manoeuvre in both body positions, correlate highly with model simulation of the jugular cross-sectional area (R(2) = 0.97). The results suggest that the cerebral venous flow distribution depends on posture and CVP: in supine humans the internal jugular veins are the primary pathway...

Control of neuropeptide expression by parallel activity-dependent pathways in caenorhabditis elegans

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Rojo Romanos, Teresa; Petersen, Jakob Gramstrup; Pocock, Roger

2017-01-01

Monitoring of neuronal activity within circuits facilitates integrated responses and rapid changes in behavior. We have identified a system in Caenorhabditis elegans where neuropeptide expression is dependent on the ability of the BAG neurons to sense carbon dioxide. In C. Elegans, CO 2 sensing...... is predominantly coordinated by the BAG-expressed receptor-type guanylate cyclase GCY-9. GCY-9 binding to CO 2 causes accumulation of cyclic GMP and opening of the cGMP-gated TAX-2/TAX-4 cation channels; provoking an integrated downstream cascade that enables C. Elegans to avoid high CO 2. Here we show that c...... that expression of flp-19::GFP is controlled in parallel to GCY-9 by the activity-dependent transcription factor CREB (CRH-1) and the cAMP-dependent protein kinase (KIN-2) signaling pathway. We therefore show that two parallel pathways regulate neuropeptide gene expression in the BAG sensory neurons: the ability...

The canonical Wnt signaling pathway promotes chondrocyte differentiation in a Sox9-dependent manner

International Nuclear Information System (INIS)

Yano, Fumiko; Kugimiya, Fumitaka; Ohba, Shinsuke; Ikeda, Toshiyuki; Chikuda, Hirotaka; Ogasawara, Toru; Ogata, Naoshi; Takato, Tsuyoshi; Nakamura, Kozo; Kawaguchi, Hiroshi; Chung, Ung-il

2005-01-01

To better understand the role of the canonical Wnt signaling pathway in cartilage development, we adenovirally expressed a constitutively active (Canada) or a dominant negative (dn) form of lymphoid enhancer factor-1 (LEF-1), the main nuclear effector of the pathway, in undifferentiated mesenchymal cells, chondrogenic cells, and primary chondrocytes, and examined the expression of markers for chondrogenic differentiation and hypertrophy. caLEF-1 and LiCl, an activator of the canonical pathway, promoted both chondrogenic differentiation and hypertrophy, whereas dnLEF-1 and the gene silencing of β-catenin suppressed LiCl-promoted effects. To investigate whether these effects were dependent on Sox9, a master regulator of cartilage development, we stimulated Sox9-deficient ES cells with the pathway. caLEF-1 and LiCl promoted both chondrogenic differentiation and hypertrophy in wild-type, but not in Sox9-deficient, cells. The response of Sox9-deficient cells was restored by the adenoviral expression of Sox9. Thus, the canonical Wnt signaling pathway promotes chondrocyte differentiation in a Sox9-dependent manner

Innate immune responses induced by lipopolysaccharide and lipoteichoic acid in primary goat mammary epithelial cells.

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Bulgari, Omar; Dong, Xianwen; Roca, Alfred L; Caroli, Anna M; Loor, Juan J

2017-01-01

Innate immune responses induced by in vitro stimulation of primary mammary epithelial cells (MEC) using Gram-negative lipopolysaccharide (LPS) and Gram-positive lipoteichoic acid (LTA) bacterial cell wall components are well- characterized in bovine species. The objective of the current study was to characterize the downstream regulation of the inflammatory response induced by Toll-like receptors in primary goat MEC (pgMEC). We performed quantitative real-time RT-PCR (qPCR) to measure mRNA levels of 9 genes involved in transcriptional regulation or antibacterial activity: Toll-like receptor 2 ( TLR2 ), Toll-like receptor 4 ( TLR4 ), prostaglandin-endoperoxide synthase 2 ( PTGS2 ), interferon induced protein with tetratricopeptide repeats 3 ( IFIT3 ), interferon regulatory factor 3 ( IRF3 ), myeloid differentiation primary response 88 ( MYD88 ), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 ( NFKB1 ), Toll interacting protein ( TOLLIP ), and lactoferrin ( LTF ). Furthermore, we analyzed 7 cytokines involved in Toll-like receptor signaling pathways: C-C motif chemokine ligand 2 ( CCL2 ), C-C motif chemokine ligand 5 ( CCL5 ), C-X-C motif chemokine ligand 6 ( CXCL6 ), interleukin 8 ( CXCL8 ), interleukin 1 beta ( IL1B ), interleukin 6 ( IL6 ), and tumor necrosis factor alpha ( TNF ). Stimulation of pgMEC with LPS for 3 h led to an increase in expression of CCL2 , CXCL6 , IL6 , CXCL8 , PTGS2 , IFIT3 , MYD88 , NFKB1 , and TLR4 ( P  < 0.05). Except for IL6 , and PTGS2 , the same genes had greater expression than controls at 6 h post-LPS ( P  < 0.05). Expression of CCL5 , PTGS2 , IFIT3 , NFKB1 , TLR4 , and TOLLIP was greater than controls after 3 h of incubation with LTA ( P  < 0.05). Compared to controls, stimulation with LTA for 6 h led to greater expression of PTGS2 , IFIT3 , NFKB1 , and TOLLIP ( P  < 0.05) whereas the expression of CXCL6 , CXCL8 , and TLR4 was lower ( P  < 0.05). At 3 h incubation with both toxins

Flaxseed Oil Attenuates Intestinal Damage and Inflammation by Regulating Necroptosis and TLR4/NOD Signaling Pathways Following Lipopolysaccharide Challenge in a Piglet Model.

Science.gov (United States)

Zhu, Huiling; Wang, Haibo; Wang, Shuhui; Tu, Zhixiao; Zhang, Lin; Wang, Xiuying; Hou, Yongqing; Wang, Chunwei; Chen, Jie; Liu, Yulan

2018-05-01

Flaxseed oil is a rich source of α-linolenic acid (ALA), which is the precursor of the long-chain n-3 polyunsaturated fatty acids (PUFAs), including docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA). This study investigates the protective effect of flaxseed oil against intestinal injury induced by lipopolysaccharide (LPS). Twenty-four weaned pigs were used in a 2 × 2 factorial experiment with dietary treatment (5% corn oil vs 5% flaxseed oil) and LPS challenge (saline vs LPS). On day 21 of the experiment, pigs were administrated with LPS or saline. At 2 h and 4 h post-administration, blood samples were collected. After the blood harvest at 4 h, all piglets were slaughtered and intestinal samples were collected. Flaxseed oil supplementation led to the enrichment of ALA, EPA, and total n-3 PUFAs in intestine. Flaxseed oil improved intestinal morphology, jejunal lactase activity, and claudin-1 protein expression. Flaxseed oil downregulated the mRNA expression of intestinal necroptotic signals. Flaxseed oil also downregulated the mRNA expression of intestinal toll-like receptors 4 (TLR4) and its downstream signals myeloid differentiation factor 88 (MyD88), nuclear factor kappa B (NF-κB), and nucleotide-binding oligomerization domain proteins 1, 2 (NOD1, NOD2) and its adapter molecule, receptor-interacting protein kinase 2 (RIPK2). These results suggest that dietary addition of flaxseed oil enhances intestinal integrity and barrier function, which is involved in modulating necroptosis and TLR4/NOD signaling pathways. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

C5a receptor (CD88) blockade protects against MPO-ANCA GN.

Science.gov (United States)

Xiao, Hong; Dairaghi, Daniel J; Powers, Jay P; Ertl, Linda S; Baumgart, Trageen; Wang, Yu; Seitz, Lisa C; Penfold, Mark E T; Gan, Lin; Hu, Peiqi; Lu, Bao; Gerard, Norma P; Gerard, Craig; Schall, Thomas J; Jaen, Juan C; Falk, Ronald J; Jennette, J Charles

2014-02-01

Necrotizing and crescentic GN (NCGN) with a paucity of glomerular immunoglobulin deposits is associated with ANCA. The most common ANCA target antigens are myeloperoxidase (MPO) and proteinase 3. In a manner that requires activation of the alternative complement pathway, passive transfer of antibodies to mouse MPO (anti-MPO) induces a mouse model of ANCA NCGN that closely mimics human disease. Here, we confirm the importance of C5aR/CD88 in the mediation of anti-MPO-induced NCGN and report that C6 is not required. We further demonstrate that deficiency of C5a-like receptor (C5L2) has the reverse effect of C5aR/CD88 deficiency and results in more severe disease, indicating that C5aR/CD88 engagement enhances inflammation and C5L2 engagement suppresses inflammation. Oral administration of CCX168, a small molecule antagonist of human C5aR/CD88, ameliorated anti-MPO-induced NCGN in mice expressing human C5aR/CD88. These observations suggest that blockade of C5aR/CD88 might have therapeutic benefit in patients with ANCA-associated vasculitis and GN.

Selective maintenance of multi-state systems with structural dependence

International Nuclear Information System (INIS)

Dao, Cuong D.; Zuo, Ming J.

2017-01-01

This paper studies the selective maintenance problem for multi-state systems with structural dependence. Each component can be in one of multiple working levels and several maintenance actions are possible to a component in a maintenance break. The components structurally form multiple hierarchical levels and dependence groups. A directed graph is used to represent the precedence relations of components in the system. A selective maintenance optimization model is developed to maximize the system reliability in the next mission under time and cost constraints. A backward search algorithm is used to determine the assembly sequence for a selective maintenance scenario. The maintenance model helps maintenance managers in determining the best combination of maintenance activities to maximize the probability of successfully completing the next mission. Examples showing the use of the proposed method are presented. - Highlights: • A selective maintenance model for multi-state systems is proposed considering both economic and structural dependence. • Structural dependence is modeled as precedence relationship when disassembling components for maintenance. • Resources for disassembly and maintenance are evaluated using a backward search algorithm. • Maintenance strategies with and without structural dependence are analyzed. • Ignoring structural dependence may lead to over-estimation of system reliability.

A nuclear-receptor-dependent phosphatidylcholine pathway with antidiabetic effects.

Science.gov (United States)

Lee, Jae Man; Lee, Yoon Kwang; Mamrosh, Jennifer L; Busby, Scott A; Griffin, Patrick R; Pathak, Manish C; Ortlund, Eric A; Moore, David D

2011-05-25

Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (also known as NR5A2) regulates bile acid biosynthesis. Structural studies have identified phospholipids as potential LRH-1 ligands, but their functional relevance is unclear. Here we show that an unusual phosphatidylcholine species with two saturated 12 carbon fatty acid acyl side chains (dilauroyl phosphatidylcholine (DLPC)) is an LRH-1 agonist ligand in vitro. DLPC treatment induces bile acid biosynthetic enzymes in mouse liver, increases bile acid levels, and lowers hepatic triglycerides and serum glucose. DLPC treatment also decreases hepatic steatosis and improves glucose homeostasis in two mouse models of insulin resistance. Both the antidiabetic and lipotropic effects are lost in liver-specific Lrh-1 knockouts. These findings identify an LRH-1 dependent phosphatidylcholine signalling pathway that regulates bile acid metabolism and glucose homeostasis.

Interleukin-33 promoting Th1 lymphocyte differentiation dependents on IL-12.

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Komai-Koma, Mousa; Wang, Eryi; Kurowska-Stolarska, Mariola; Li, Dong; McSharry, Charles; Xu, Damo

2016-03-01

The pro-Th2 cytokine IL-33 is now emerging as an important Th1 cytokine-IFN-γ inducer in murine CD4(+) T cells that is essential for protective cell-mediated immunity against viral infection in mice. However, whether IL-33 can promote human Th1 cell differentiation and how IL-33 polarizes Th1 cells is less understood. We assessed the ability of IL-33 to induce Th1 cell differentiation and IFN-γ production in vitro and in vivo. We report here that IL-33 alone had no ability in Th1 cell polarization. However it potentiated IL-12-mediated Th1 cell differentiation and IFN-γ production in TCR-stimulated murine and human CD4(+) T cells in vitro and in vivo. IL-33 promoted Th1 cell development via MyD88 and synergized with IL-12 to enhance St2 and IL-12R expression in CD4(+) T cells. These data therefore provide a novel mechanism for Th1 cell differentiation and optimal induction of a Type 1 response. Thus, IL-33 is capable of inducing IL-12-dependent Th1 cell differentiation in human and mouse CD4(+) T cells. Copyright © 2015 The Authors. Published by Elsevier GmbH.. All rights reserved.

Intercohort density dependence drives brown trout habitat selection

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Ayllón, Daniel; Nicola, Graciela G.; Parra, Irene; Elvira, Benigno; Almodóvar, Ana

2013-01-01

Habitat selection can be viewed as an emergent property of the quality and availability of habitat but also of the number of individuals and the way they compete for its use. Consequently, habitat selection can change across years due to fluctuating resources or to changes in population numbers. However, habitat selection predictive models often do not account for ecological dynamics, especially density dependent processes. In stage-structured population, the strength of density dependent interactions between individuals of different age classes can exert a profound influence on population trajectories and evolutionary processes. In this study, we aimed to assess the effects of fluctuating densities of both older and younger competing life stages on the habitat selection patterns (described as univariate and multivariate resource selection functions) of young-of-the-year, juvenile and adult brown trout Salmo trutta. We observed all age classes were selective in habitat choice but changed their selection patterns across years consistently with variations in the densities of older but not of younger age classes. Trout of an age increased selectivity for positions highly selected by older individuals when their density decreased, but this pattern did not hold when the density of younger age classes varied. It suggests that younger individuals are dominated by older ones but can expand their range of selected habitats when density of competitors decreases, while older trout do not seem to consider the density of younger individuals when distributing themselves even though they can negatively affect their final performance. Since these results may entail critical implications for conservation and management practices based on habitat selection models, further research should involve a wider range of river typologies and/or longer time frames to fully understand the patterns of and the mechanisms underlying the operation of density dependence on brown trout habitat

Cell Free DNA of Tumor Origin Induces a 'Metastatic' Expression Profile in HT-29 Cancer Cell Line.

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István Fűri

Full Text Available Epithelial cells in malignant conditions release DNA into the extracellular compartment. Cell free DNA of tumor origin may act as a ligand of DNA sensing mechanisms and mediate changes in epithelial-stromal interactions.To evaluate and compare the potential autocrine and paracrine regulatory effect of normal and malignant epithelial cell-related DNA on TLR9 and STING mediated pathways in HT-29 human colorectal adenocarcinoma cells and normal fibroblasts.DNA isolated from normal and tumorous colonic epithelia of fresh frozen surgically removed tissue samples was used for 24 and 6 hour treatment of HT-29 colon carcinoma and HDF-α fibroblast cells. Whole genome mRNA expression analysis and qRT-PCR was performed for the elements/members of TLR9 signaling pathway. Immunocytochemistry was performed for epithelial markers (i.e. CK20 and E-cadherin, DNA methyltransferase 3a (DNMT3a and NFκB (for treated HDFα cells.Administration of tumor derived DNA on HT29 cells resulted in significant (p<0.05 mRNA level alteration in 118 genes (logFc≥1, p≤0.05, including overexpression of metallothionein genes (i.e. MT1H, MT1X, MT1P2, MT2A, metastasis-associated genes (i.e. TACSTD2, MACC1, MALAT1, tumor biomarker (CEACAM5, metabolic genes (i.e. INSIG1, LIPG, messenger molecule genes (i.e. DAPP, CREB3L2. Increased protein levels of CK20, E-cadherin, and DNMT3a was observed after tumor DNA treatment in HT-29 cells. Healthy DNA treatment affected mRNA expression of 613 genes (logFc≥1, p≤0.05, including increased expression of key adaptor molecules of TLR9 pathway (e.g. MYD88, IRAK2, NFκB, IL8, IL-1β, STING pathway (ADAR, IRF7, CXCL10, CASP1 and the FGF2 gene.DNA from tumorous colon epithelium, but not from the normal epithelial cells acts as a pro-metastatic factor to HT-29 cells through the overexpression of pro-metastatic genes through TLR9/MYD88 independent pathway. In contrast, DNA derived from healthy colonic epithelium induced TLR9 and STING signaling

IL-27 Induced by Select Candida spp. via TLR7/NOD2 Signaling and IFN-β Production Inhibits Fungal Clearance

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Patin, Emmanuel C.; Jones, Adam V.; Thompson, Aiysha; Clement, Mathew; Liao, Chia-Te; Griffiths, James S.; Wallace, Leah E.; Bryant, Clare E.; Lang, Roland; Rosenstiel, Philip; Humphreys, Ian R.; Taylor, Philip R.

2016-01-01

Candida spp. elicit cytokine production downstream of various pathogen recognition receptors, including C-type lectin-like receptors, TLRs, and nucleotide oligomerization domain (NOD)–like receptors. IL-12 family members IL-12p70 and IL-23 are important for host immunity against Candida spp. In this article, we show that IL-27, another IL-12 family member, is produced by myeloid cells in response to selected Candida spp. We demonstrate a novel mechanism for Candida parapsilosis–mediated induction of IL-27 in a TLR7-, MyD88-, and NOD2-dependent manner. Our data revealed that IFN-β is induced by C. parapsilosis, which in turn signals through the IFN-α/β receptor and STAT1/2 to induce IL-27. Moreover, IL-27R (WSX-1)–deficient mice systemically infected with C. parapsilosis displayed enhanced pathogen clearance compared with wild-type mice. This was associated with increased levels of proinflammatory cytokines in the serum and increased IFN-γ and IL-17 responses in the spleens of IL-27R–deficient mice. Thus, our data define a novel link between C. parapsilosis, TLR7, NOD2, IFN-β, and IL-27, and we have identified an important role for IL-27 in the immune response against C. parapsilosis. Overall, these findings demonstrate an important mechanism for the suppression of protective immune responses during infection with C. parapsilosis, which has potential relevance for infections with other fungal pathogens. PMID:27259855

Effect of Short-Circuit Pathways on Water Quality in Selected Confined Aquifers (Invited)

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McMahon, P. B.

2010-12-01

Confined aquifers in the United States generally contain fewer anthropogenic contaminants than unconfined aquifers because confined aquifers often contain water recharged prior to substantial human development and redox conditions are more reducing, which favors degradation of common contaminants like nitrate and chlorinated solvents. Groundwater in a confined part of the High Plains aquifer near York, Nebraska had an adjusted radiocarbon age of about 2,000 years, and groundwater in a confined part of the Floridan aquifer near Tampa, Florida had apparent ages greater than 60 years on the basis of tritium measurements. Yet compounds introduced more recently into the environment (anthropogenic nitrate and volatile organic compounds) were detected in selected public-supply wells completed in both aquifers. Depth-dependent measurements of flow and chemistry in the pumping supply wells, groundwater age dating, numerical modeling of groundwater flow, and other monitoring data indicated that the confined aquifers sampled by the supply wells were connected to contaminated unconfined aquifers by short-circuit pathways. In the High Plains aquifer, the primary pathways appeared to be inactive irrigation wells screened in both the unconfined and confined aquifers. In the Floridan aquifer, the primary pathways were karst sinkholes and conduits. Heavy pumping in both confined systems exacerbated the problem by reducing the potentiometric surface and increasing groundwater velocities, thus enhancing downward gradients and reducing reaction times for processes like denitrification. From a broader perspective, several confined aquifers in the U.S. have experienced large declines in their potentiometric surfaces because of groundwater pumping and this could increase the potential for contamination in those aquifers, particularly where short-circuit pathways connect them to shallower, contaminated sources of water, such as was observed in York and Tampa.

Negative regulation of Toll-like receptor signalling 

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Halina Antosz

2013-04-01

Full Text Available The mechanism of innate immunity is based on the pattern recognition receptors (PRR that recognize molecular patterns associated with pathogens (PAMPs. Among PRR receptors Toll-like receptors (TLR are distinguished. As a result of contact with pathogens, TLRs activate specific intracellular signaling pathways. It happens through proteins such as adaptor molecules, e.g. MyD88, TIRAP, TRIF, TRAM, and IPS-1, which participate in the cascade activation of kinases (IKK, MAP, RIP-1, TBK-1 as well as transcription factors (NF-κB, AP-1 and regulatory factor (IRF3. The result of this activation is the production of active proinflammatory cytokines, chemokines, interferons and enzymes. The PRR pathways are controlled by extra – and intracellular molecules to prevent overexpression of PRR. They include soluble receptors (sTLR, transmembrane proteins (ST2, SIGIRR, RP105, TRAIL-R and intracellular inhibitors (SOCS-1, SOCS-3, sMyD88, TOLLIP, IRAK-M, SARM, A20, β-arrestin, CYLD, SHP. These molecules maintain the balance between activation and inhibition and ensure balancing of the beneficial and adverse effects of antigen recognition.

Genome-Wide Gene Set Analysis for Identification of Pathways Associated with Alcohol Dependence

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Biernacka, Joanna M.; Geske, Jennifer; Jenkins, Gregory D.; Colby, Colin; Rider, David N.; Karpyak, Victor M.; Choi, Doo-Sup; Fridley, Brooke L.

2013-01-01

It is believed that multiple genetic variants with small individual effects contribute to the risk of alcohol dependence. Such polygenic effects are difficult to detect in genome-wide association studies that test for association of the phenotype with each single nucleotide polymorphism (SNP) individually. To overcome this challenge, gene set analysis (GSA) methods that jointly test for the effects of pre-defined groups of genes have been proposed. Rather than testing for association between the phenotype and individual SNPs, these analyses evaluate the global evidence of association with a set of related genes enabling the identification of cellular or molecular pathways or biological processes that play a role in development of the disease. It is hoped that by aggregating the evidence of association for all available SNPs in a group of related genes, these approaches will have enhanced power to detect genetic associations with complex traits. We performed GSA using data from a genome-wide study of 1165 alcohol dependent cases and 1379 controls from the Study of Addiction: Genetics and Environment (SAGE), for all 200 pathways listed in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Results demonstrated a potential role of the “Synthesis and Degradation of Ketone Bodies” pathway. Our results also support the potential involvement of the “Neuroactive Ligand Receptor Interaction” pathway, which has previously been implicated in addictive disorders. These findings demonstrate the utility of GSA in the study of complex disease, and suggest specific directions for further research into the genetic architecture of alcohol dependence. PMID:22717047

Distribution of events of positive selection and population differentiation in a metabolic pathway: the case of asparagine N-glycosylation

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Dall’Olio Giovanni

2012-06-01

Full Text Available Abstract Background Asparagine N-Glycosylation is one of the most important forms of protein post-translational modification in eukaryotes. This metabolic pathway can be subdivided into two parts: an upstream sub-pathway required for achieving proper folding for most of the proteins synthesized in the secretory pathway, and a downstream sub-pathway required to give variability to trans-membrane proteins, and involved in adaptation to the environment and innate immunity. Here we analyze the nucleotide variability of the genes of this pathway in human populations, identifying which genes show greater population differentiation and which genes show signatures of recent positive selection. We also compare how these signals are distributed between the upstream and the downstream parts of the pathway, with the aim of exploring how forces of population differentiation and positive selection vary among genes involved in the same metabolic pathway but subject to different functional constraints. Results Our results show that genes in the downstream part of the pathway are more likely to show a signature of population differentiation, while events of positive selection are equally distributed among the two parts of the pathway. Moreover, events of positive selection are frequent on genes that are known to be at bifurcation points, and that are identified as being in key position by a network-level analysis such as MGAT3 and GCS1. Conclusions These findings indicate that the upstream part of the Asparagine N-Glycosylation pathway has lower diversity among populations, while the downstream part is freer to tolerate diversity among populations. Moreover, the distribution of signatures of population differentiation and positive selection can change between parts of a pathway, especially between parts that are exposed to different functional constraints. Our results support the hypothesis that genes involved in constitutive processes can be expected to show

TRX-E-002-1 Induces c-Jun-Dependent Apoptosis in Ovarian Cancer Stem Cells and Prevents Recurrence In Vivo.

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Alvero, Ayesha B; Heaton, Andrew; Lima, Eydis; Pitruzzello, Mary; Sumi, Natalia; Yang-Hartwich, Yang; Cardenas, Carlos; Steinmacher, Sahra; Silasi, Dan-Arin; Brown, David; Mor, Gil

2016-06-01

Chemoresistance is a major hurdle in the management of patients with epithelial ovarian cancer and is responsible for its high mortality. Studies have shown that chemoresistance is due to the presence of a subgroup of cancer cells with stemness properties and a high capacity for tumor repair. We have developed a library of super-benzopyran analogues to generate potent compounds that can induce cell death in chemoresistant cancer stem cells. TRX-E-002-1 is identified as the most potent analogue and can induce cell death in all chemoresistant CD44(+)/MyD88(+) ovarian cancer stem cells tested (IC50 = 50 nmol/L). TRX-E-002-1 is also potent against spheroid cultures formed from cancer stem cells, chemosensitive CD44(-)/MyD88(-) ovarian cancer cells, and heterogeneous cultures of ovarian cancer cells. Cell death was associated with the phosphorylation and increased levels of c-Jun and induction of caspases. In vivo, TRX-E-002-1 given as daily intraperitoneal monotherapy at 100 mg/kg significantly decreased intraperitoneal tumor burden compared with vehicle control. When given in combination with cisplatin, animals receiving the combination of cisplatin and TRX-E-002-1 showed decreased tumor burden compared with each monotherapy. Finally, TRX-E-002-1 given as maintenance treatment after paclitaxel significantly delayed disease recurrence. Our results suggest that TRX-E-002-1 may fill the current need for better therapeutic options in the control and management of recurrent ovarian cancer and may help improve patient survival. Mol Cancer Ther; 15(6); 1279-90. ©2016 AACR. ©2016 American Association for Cancer Research.

Role of CSL-dependent and independent Notch signaling pathways in cell apoptosis.

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Zeng, Chong; Xing, Rui; Liu, Jing; Xing, Feiyue

2016-01-01

Apoptosis is a normally biological phenomenon in various organisms, involving complexly molecular mechanisms with a series of signaling processes. Notch signaling is found evolutionarily conserved in many species, playing a critical role in embryonic development, normal tissue homeostasis, angiogenesis and immunoregulation. The focus of this review is on currently novel advances about roles of CSL-dependent and independent Notch signaling pathways in cell apoptosis. The CSL can bind Notch intracellular domain (NIC) to act as a switch in mediating transcriptional activation or inactivation of the Notch signaling pathway downstream genes in the nucleus. It shows that CSL-dependent signaling regulates the cell apoptosis through Hes-1-PTEN-AKT-mTOR signaling, but rather the CSL-independent signaling mediates the cell apoptosis possibly via NIC-mTORC2-AKT-mTOR signaling, providing a new insight into apoptotic mechanisms.

Ligand-selective activation of heterologously-expressed mammalian olfactory receptor.

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Ukhanov, K; Bobkov, Y; Corey, E A; Ache, B W

2014-10-01

Mammalian olfactory receptors (ORs) appear to have the capacity to couple to multiple G protein-coupled signaling pathways in a ligand-dependent selective manner. To better understand the mechanisms and molecular range of such ligand selectivity, we expressed the mouse eugenol OR (mOR-EG) in HEK293T cells together with Gα15 to monitor activation of the phospholipase-C (PLC) signaling pathway and/or Gαolf to monitor activation of the adenylate cyclase (AC) signaling pathway, resulting in intracellular Ca(2+) release and/or Ca(2+) influx through a cyclic nucleotide-gated channel, respectively. PLC-dependent responses differed dynamically from AC-dependent responses, allowing them to be distinguished when Gα15 and Gαolf were co-expressed. The dynamic difference in readout was independent of the receptor, the heterologous expression system, and the ligand concentration. Of 17 reported mOR-EG ligands tested, including eugenol, its analogs, and structurally dissimilar compounds (mousse cristal, nootkatone, orivone), some equally activated both signaling pathways, some differentially activated both signaling pathways, and some had no noticeable effect even at 1-5mM. Our findings argue that mOR-EG, when heterologously expressed, can couple to two different signaling pathways in a ligand selective manner. The challenge now is to determine the potential of mOR-EG, and perhaps other ORs, to activate multiple signaling pathways in a ligand selective manner in native ORNs. Copyright © 2014 Elsevier Ltd. All rights reserved.

Two Neisseria meningitidis strains with different ability to stimulate toll-like receptor 4 through the MyD88-independent pathway

DEFF Research Database (Denmark)

Mogensen, T.H.; Paludan, Søren Riis; Kilian, Mogens

2006-01-01

Neisseria meningitidis causes acute severe diseases, including sepsis and meningitis, and more benign manifestations such as chronic meningococcemia or colonization of the upper respiratory tract. The inflammatory response, which contributes to the pathogenesis of meningococcal disease......, is initiated by pattern recognition receptors, among which Toll-like receptors (TLR)s have been ascribed a particularly important role. We have previously demonstrated that N. meningitidis induce proinflammatory cytokine expression through TLR2 and TLR4. Here we characterize the molecular basis...... for differential activation of the inflammatory response by two N. meningitidis strains. This difference was due to differential ability to activate signal transduction through TLR4, as HEK293 cells expressing TLR4 produced significantly different levels of interleukin-8 in response to these strains. At the level...

Constraint-based modeling and kinetic analysis of the Smad dependent TGF-beta signaling pathway.

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Zhike Zi

Full Text Available BACKGROUND: Investigation of dynamics and regulation of the TGF-beta signaling pathway is central to the understanding of complex cellular processes such as growth, apoptosis, and differentiation. In this study, we aim at using systems biology approach to provide dynamic analysis on this pathway. METHODOLOGY/PRINCIPAL FINDINGS: We proposed a constraint-based modeling method to build a comprehensive mathematical model for the Smad dependent TGF-beta signaling pathway by fitting the experimental data and incorporating the qualitative constraints from the experimental analysis. The performance of the model generated by constraint-based modeling method is significantly improved compared to the model obtained by only fitting the quantitative data. The model agrees well with the experimental analysis of TGF-beta pathway, such as the time course of nuclear phosphorylated Smad, the subcellular location of Smad and signal response of Smad phosphorylation to different doses of TGF-beta. CONCLUSIONS/SIGNIFICANCE: The simulation results indicate that the signal response to TGF-beta is regulated by the balance between clathrin dependent endocytosis and non-clathrin mediated endocytosis. This model is useful to be built upon as new precise experimental data are emerging. The constraint-based modeling method can also be applied to quantitative modeling of other signaling pathways.

Calcineurin inhibitors recruit protein kinases JAK2 and JNK, TLR signaling and the UPR to activate NF-κB-mediated inflammatory responses in kidney tubular cells

International Nuclear Information System (INIS)

González-Guerrero, Cristian; Ocaña-Salceda, Carlos; Berzal, Sergio; Carrasco, Susana; Fernández-Fernández, Beatriz

2013-01-01

The calcineurin inhibitors (CNIs) cyclosporine (CsA) and tacrolimus are key drugs in current immunosuppressive regimes for solid organ transplantation. However, they are nephrotoxic and promote death and profibrotic responses in tubular cells. Moreover, renal inflammation is observed in CNI nephrotoxicity but the mechanisms are poorly understood. We have now studied molecular pathways leading to inflammation elicited by the CNIs in cultured and kidney tubular cells. Both CsA and tacrolimus elicited a proinflammatory response in tubular cells as evidenced by a transcriptomics approach. Transcriptomics also suggested several potential pathways leading to expression of proinflammatory genes. Validation and functional studies disclosed that in tubular cells, CNIs activated protein kinases such as the JAK2/STAT3 and TAK1/JNK/AP-1 pathways, TLR4/Myd88/IRAK signaling and the Unfolded Protein Response (UPR) to promote NF-κB activation and proinflammatory gene expression. CNIs also activated an Nrf2/HO-1-dependent compensatory response and the Nrf2 activator sulforaphane inhibited JAK2 and JNK activation and inflammation. A murine model of CsA nephrotoxicity corroborated activation of the proinflammatory pathways identified in cell cultures. Human CNIs nephrotoxicity was also associated with NF-κB, STAT3 and IRE1α activation. In conclusion, CNIs recruit several intracellular pathways leading to previously non-described proinflammatory actions in renal tubular cells. Identification of these pathways provides novel clues for therapeutic intervention to limit CNIs nephrotoxicity. - Highlights: • Molecular mechanisms modulating CNI renal inflammation were investigated. • Kinases, immune receptors and ER stress mediate the inflammatory response to CNIs. • Several intracellular pathways activate NF-κB in CNIs-treated tubular cells. • A NF-κB-dependent cytokine profile characterizes CNIs-induced inflammation. • CNI nephrotoxicity was associated to inflammatory

Microglia Induce Neurotoxic IL-17+ γδ T Cells Dependent on TLR2, TLR4, and TLR9 Activation.

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Katja Derkow

Full Text Available Interleukin-17 (IL-17 acts as a key regulator in central nervous system (CNS inflammation. γδ T cells are an important innate source of IL-17. Both IL-17+ γδ T cells and microglia, the major resident immune cells of the brain, are involved in various CNS disorders such as multiple sclerosis and stroke. Also, activation of Toll-like receptor (TLR signaling pathways contributes to CNS damage. However, the mechanisms underlying the regulation and interaction of these cellular and molecular components remain unclear.In this study, we investigated the crosstalk between γδ T cells and microglia activated by TLRs in the context of neuronal damage. To this end, co-cultures of IL-17+ γδ T cells, neurons, and microglia were analyzed by immunocytochemistry, flow cytometry, ELISA and multiplex immunoassays.We report here that IL-17+ γδ T cells but not naïve γδ T cells induce a dose- and time-dependent decrease of neuronal viability in vitro. While direct stimulation of γδ T cells with various TLR ligands did not result in up-regulation of CD69, CD25, or in IL-17 secretion, supernatants of microglia stimulated by ligands specific for TLR2, TLR4, TLR7, or TLR9 induced activation of γδ T cells through IL-1β and IL-23, as indicated by up-regulation of CD69 and CD25 and by secretion of vast amounts of IL-17. This effect was dependent on the TLR adaptor myeloid differentiation primary response gene 88 (MyD88 expressed by both γδ T cells and microglia, but did not require the expression of TLRs by γδ T cells. Similarly to cytokine-primed IL-17+ γδ T cells, IL-17+ γδ T cells induced by supernatants derived from TLR-activated microglia also caused neurotoxicity in vitro. While these neurotoxic effects required stimulation of TLR2, TLR4, or TLR9 in microglia, neuronal injury mediated by bone marrow-derived macrophages did not require TLR signaling. Neurotoxicity mediated by IL-17+ γδ T cells required a direct cell-cell contact between T

Interleukin-34 Regulates Th1 and Th17 Cytokine Production by Activating Multiple Signaling Pathways through CSF-1R in Chicken Cell Lines

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Anh Duc Truong

2018-06-01

Full Text Available Interleukin-34 (IL-34 is a newly recognized cytokine with functions similar to macrophage colony-stimulating factor 1. It is expressed in macrophages and fibroblasts, where it induces cytokine production; however, the mechanism of chicken IL-34 (chIL-34 signaling has not been identified to date. The aim of this study was to analyze the signal transduction pathways and specific biological functions associated with chIL-34 in chicken macrophage (HD11 and fibroblast (OU2 cell lines. We found that IL-34 is a functional ligand for the colony-stimulating factor receptor (CSF-1R in chicken cell lines. Treatment with chIL-34 increased the expression of Th1 and Th17 cytokines through phosphorylation of tyrosine and serine residues in Janus kinase (JAK 2, tyrosine kinase 2 (TYK2, signal transducer and activator of transcription (STAT 1, STAT3, and Src homology 2-containing tyrosine phosphatase 2 (SHP-2, which also led to phosphorylation of NF-κB1, p-mitogen-activated protein kinase kinase kinase 7 (TAK1, MyD88, suppressor of cytokine signaling 1 (SOCS1, and extracellular signal-regulated kinase 1 and 2 (ERK1/2. Taken together, these results suggest that chIL-34 functions by binding to CSF-1R and activating the JAK/STAT, nuclear factor κ B (NF-κB, and mitogen-activated protein kinase signaling pathways; these signaling events regulate cytokine expression and suggest roles for chIL-34 in innate and adaptive immunity.

The hedgehog pathway gene shifted functions together with the hmgcr-dependent isoprenoid biosynthetic pathway to orchestrate germ cell migration.

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Girish Deshpande

Full Text Available The Drosophila embryonic gonad is assembled from two distinct cell types, the Primordial Germ Cells (PGCs and the Somatic Gonadal Precursor cells (SGPs. The PGCs form at the posterior of blastoderm stage embryos and are subsequently carried inside the embryo during gastrulation. To reach the SGPs, the PGCs must traverse the midgut wall and then migrate through the mesoderm. A combination of local repulsive cues and attractive signals emanating from the SGPs guide migration. We have investigated the role of the hedgehog (hh pathway gene shifted (shf in directing PGC migration. shf encodes a secreted protein that facilitates the long distance transmission of Hh through the proteoglycan matrix after it is released from basolateral membranes of Hh expressing cells in the wing imaginal disc. shf is expressed in the gonadal mesoderm, and loss- and gain-of-function experiments demonstrate that it is required for PGC migration. Previous studies have established that the hmgcr-dependent isoprenoid biosynthetic pathway plays a pivotal role in generating the PGC attractant both by the SGPs and by other tissues when hmgcr is ectopically expressed. We show that production of this PGC attractant depends upon shf as well as a second hh pathway gene gγ1. Further linking the PGC attractant to Hh, we present evidence indicating that ectopic expression of hmgcr in the nervous system promotes the release/transmission of the Hh ligand from these cells into and through the underlying mesodermal cell layer, where Hh can contact migrating PGCs. Finally, potentiation of Hh by hmgcr appears to depend upon cholesterol modification.

GP88 (PC-Cell Derived Growth Factor, progranulin stimulates proliferation and confers letrozole resistance to aromatase overexpressing breast cancer cells

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Sabnis Gauri

2011-06-01

Full Text Available Abstract Background Aromatase inhibitors (AI that inhibit breast cancer cell growth by blocking estrogen synthesis have become the treatment of choice for post-menopausal women with estrogen receptor positive (ER+ breast cancer. However, some patients display de novo or acquired resistance to AI. Interactions between estrogen and growth factor signaling pathways have been identified in estrogen-responsive cells as one possible reason for acquisition of resistance. Our laboratory has characterized an autocrine growth factor overexpressed in invasive ductal carcinoma named PC-Cell Derived Growth Factor (GP88, also known as progranulin. In the present study, we investigated the role GP88 on the acquisition of resistance to letrozole in ER+ breast cancer cells Methods We used two aromatase overexpressing human breast cancer cell lines MCF-7-CA cells and AC1 cells and their letrozole resistant counterparts as study models. Effect of stimulating or inhibiting GP88 expression on proliferation, anchorage-independent growth, survival and letrozole responsiveness was examined. Results GP88 induced cell proliferation and conferred letrozole resistance in a time- and dose-dependent fashion. Conversely, naturally letrozole resistant breast cancer cells displayed a 10-fold increase in GP88 expression when compared to letrozole sensitive cells. GP88 overexpression, or exogenous addition blocked the inhibitory effect of letrozole on proliferation, and stimulated survival and soft agar colony formation. In letrozole resistant cells, silencing GP88 by siRNA inhibited cell proliferation and restored their sensitivity to letrozole. Conclusion Our findings provide information on the role of an alternate growth and survival factor on the acquisition of aromatase inhibitor resistance in ER+ breast cancer.

Diosgenin induces apoptosis in IGF-1-stimulated human thyrocytes through two caspase-dependent pathways

International Nuclear Information System (INIS)

Mu, Shumin; Tian, Xingsong; Ruan, Yongwei; Liu, Yuantao; Bian, Dezhi; Ma, Chunyan; Yu, Chunxiao; Feng, Mei; Wang, Furong; Gao, Ling; Zhao, Jia-jun

2012-01-01

Highlights: ► Diosgenin induces apoptosis in IGF-1-treated thyrocytes through two caspase pathways. ► Diosgenin inhibits FLIP and activates caspase-8 in FAS related-pathway. ► Diosgenin increases ROS, regulates the ratio of Bax/Bcl-2 in mitochondrial pathway. -- Abstract: Insulin-like growth factor-1 (IGF-1) is a growth factor of the thyroid that has been shown in our previous study to possess proliferative and antiapoptotic effects in FRTL-5 cell lines through the upregulation of cyclin D and Fas-associated death domain-like interleukin-1-converting enzyme (FLICE)-inhibitory protein (FLIP). Diosgenin, a natural steroid sapogenin from plants, has been shown to induce apoptosis in many cell lines, with the exception of thyroid cells. In this report, we investigated the apoptotic effect and mechanism of diosgenin in IGF-1-stimulated primary human thyrocytes. Primary human thyrocytes were preincubated with or without IGF-1 for 24 h and subsequently exposed to varying concentrations of diosgenin for different times. We found that diosgenin induced apoptosis in human thyrocytes pretreated with IGF-1 in a dose-dependent manner through the activation of caspase cascades. Moreover, diosgenin inhibited FLIP and activated caspase-8 in the FAS-related apoptotic pathway. Diosgenin increased the production of ROS, regulated the balance of Bax and Bcl-2 and cleaved caspase-9 in the mitochondrial apoptotic pathway. These results indicate that diosgenin induces apoptosis in IGF-1-stimulated primary human thyrocytes through two caspase-dependent pathways.

Selective Ionic Transport Pathways in Phosphorene.

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Nie, Anmin; Cheng, Yingchun; Ning, Shoucong; Foroozan, Tara; Yasaei, Poya; Li, Wen; Song, Boao; Yuan, Yifei; Chen, Lin; Salehi-Khojin, Amin; Mashayek, Farzad; Shahbazian-Yassar, Reza

2016-04-13

Despite many theoretical predictions indicating exceptionally low energy barriers of ionic transport in phosphorene, the ionic transport pathways in this two-dimensional (2D) material has not been experimentally demonstrated. Here, using in situ aberration-corrected transmission electron microscopy (TEM) and density functional theory, we studied sodium ion transport in phosphorene. Our high-resolution TEM imaging complemented by electron energy loss spectroscopy demonstrates a precise description of anisotropic sodium ions migration along the [100] direction in phosphorene. This work also provides new insight into the effect of surface and the edge sites on the transport properties of phosphorene. According to our observation, the sodium ion transport is preferred in zigzag edge rather than the armchair edge. The use of this highly selective ionic transport property may endow phosphorene with new functionalities for novel chemical device applications.

P-MAPA immunotherapy potentiates the effect of cisplatin on serous ovarian carcinoma through targeting TLR4 signaling.

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de Almeida Chuffa, Luiz Gustavo; de Moura Ferreira, Grazielle; Lupi, Luiz Antonio; da Silva Nunes, Iseu; Fávaro, Wagner José

2018-01-17

Toll-like receptors (TLRs) are transmembrane proteins expressed on the surface of ovarian cancer (OC) and immune cells. Identifying the specific roles of the TLR-mediated signaling pathways in OC cells is important to guide new treatments. Because immunotherapies have emerged as the adjuvant treatment for patients with OC, we investigated the effect of a promising immunotherapeutic strategy based on protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride (P-MAPA) combined with cisplatin (CIS) on the TLR2 and TLR4 signaling pathways via myeloid differentiation factor 88 (MyD88) and TLR-associated activator of interferon (TRIF) in an in vivo model of OC. Tumors were chemically induced by a single injection of 100 μg of 7,12-dimethylbenz(a)anthracene (DMBA) directly under the left ovarian bursa in Fischer 344 rats. After the rats developed serous papillary OC, they were given P-MAPA, CIS or the combination P-MAPA+CIS as therapies. To understand the effects of the treatments, we assessed the tumor size, histopathology, and the TLR2- and TLR4-mediated inflammatory responses. Although CIS therapy was more effective than P-MAPA in reducing the tumor size, P-MAPA immunotherapy significantly increased the expressions of TLR2 and TLR4. More importantly, the combination of P-MAPA with CIS showed a greater survival rate compared to CIS alone, and exhibited a significant reduction in tumor volume compared to P-MAPA alone. The combination therapy also promoted the increase in the levels of the following OC-related proteins: TLR4, MyD88, TRIF, inhibitor of phosphorylated NF-kB alpha (p-IkBα), and nuclear factor kappa B (NF-kB p65) in both cytoplasmic and nuclear sites. While P-MAPA had no apparent effect on tumor necrosis factor alpha (TNF-α) and interleukin (IL)-6, it seems to increase interferon-γ (IFN-γ), which may induce the Thelper (Th1)-mediated immune response. Collectively, our results suggest that P-MAPA immunotherapy combined with cisplatin

Hepatitis C virus NS2 protein activates cellular cyclic AMP-dependent pathways

International Nuclear Information System (INIS)

Kim, Kyoung Mi; Kwon, Shi-Nae; Kang, Ju-Il; Lee, Song Hee; Jang, Sung Key; Ahn, Byung-Yoon; Kim, Yoon Ki

2007-01-01

Chronic infection of the hepatitis C virus (HCV) leads to liver cirrhosis and cancer. The mechanism leading to viral persistence and hepatocellular carcinoma, however, has not been fully understood. In this study, we show that the HCV infection activates cellular cAMP-dependent pathways. Expression of a luciferase reporter gene controlled by a basic promoter with the cAMP response element (CRE) was significantly elevated in human hepatoma Huh-7 cells infected with the HCV JFH1. Analysis with viral subgenomic replicons indicated that the HCV NS2 protein is responsible for the effect. Furthermore, the level of cellular transcripts whose stability is known to be regulated by cAMP was specifically reduced in cells harboring NS2-expressing replicons. These results allude to the HCV NS2 protein having a novel function of regulating cellular gene expression and proliferation through the cAMP-dependent pathway

Calcium-Dependent Protein Kinases in Phytohormone Signaling Pathways

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Wuwu Xu

2017-11-01

Full Text Available Calcium-dependent protein kinases (CPKs/CDPKs are Ca2+-sensors that decode Ca2+ signals into specific physiological responses. Research has reported that CDPKs constitute a large multigene family in various plant species, and play diverse roles in plant growth, development, and stress responses. Although numerous CDPKs have been exhaustively studied, and many of them have been found to be involved in plant hormone biosynthesis and response mechanisms, a comprehensive overview of the manner in which CDPKs participate in phytohormone signaling pathways, regulating nearly all aspects of plant growth, has not yet been undertaken. In this article, we reviewed the structure of CDPKs and the mechanism of their subcellular localization. Some CDPKs were elucidated to influence the intracellular localization of their substrates. Since little work has been done on the interaction between CDPKs and cytokinin signaling pathways, or on newly defined phytohormones such as brassinosteroids, strigolactones and salicylic acid, this paper mainly focused on discussing the integral associations between CDPKs and five plant hormones: auxins, gibberellins, ethylene, jasmonates, and abscisic acid. A perspective on future work is provided at the end.

Grizzly bear habitat selection is scale dependent.

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Ciarniello, Lana M; Boyce, Mark S; Seip, Dale R; Heard, Douglas C

2007-07-01

The purpose of our study is to show how ecologists' interpretation of habitat selection by grizzly bears (Ursus arctos) is altered by the scale of observation and also how management questions would be best addressed using predetermined scales of analysis. Using resource selection functions (RSF) we examined how variation in the spatial extent of availability affected our interpretation of habitat selection by grizzly bears inhabiting mountain and plateau landscapes. We estimated separate models for females and males using three spatial extents: within the study area, within the home range, and within predetermined movement buffers. We employed two methods for evaluating the effects of scale on our RSF designs. First, we chose a priori six candidate models, estimated at each scale, and ranked them using Akaike Information Criteria. Using this method, results changed among scales for males but not for females. For female bears, models that included the full suite of covariates predicted habitat use best at each scale. For male bears that resided in the mountains, models based on forest successional stages ranked highest at the study-wide and home range extents, whereas models containing covariates based on terrain features ranked highest at the buffer extent. For male bears on the plateau, each scale estimated a different highest-ranked model. Second, we examined differences among model coefficients across the three scales for one candidate model. We found that both the magnitude and direction of coefficients were dependent upon the scale examined; results varied between landscapes, scales, and sexes. Greenness, reflecting lush green vegetation, was a strong predictor of the presence of female bears in both landscapes and males that resided in the mountains. Male bears on the plateau were the only animals to select areas that exposed them to a high risk of mortality by humans. Our results show that grizzly bear habitat selection is scale dependent. Further, the

Steroidal androgens and nonsteroidal, tissue-selective androgen receptor modulator, S-22, regulate androgen receptor function through distinct genomic and nongenomic signaling pathways.

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Narayanan, Ramesh; Coss, Christopher C; Yepuru, Muralimohan; Kearbey, Jeffrey D; Miller, Duane D; Dalton, James T

2008-11-01

Androgen receptor (AR) ligands are important for the development and function of several tissues and organs. However, the poor oral bioavailability, pharmacokinetic properties, and receptor cross-reactivity of testosterone, coupled with side effects, place limits on its clinical use. Selective AR modulators (SARMs) elicit anabolic effects in muscle and bone, sparing reproductive organs like the prostate. However, molecular mechanisms underlying the tissue selectivity remain ambiguous. We performed a variety of in vitro studies to compare and define the molecular mechanisms of an aryl propionamide SARM, S-22, as compared with dihydrotestosterone (DHT). Studies indicated that S-22 increased levator ani muscle weight but decreased the size of prostate in rats. Analysis of the upstream intracellular signaling events indicated that S-22 and DHT mediated their actions through distinct pathways. Modulation of these pathways altered the recruitment of AR and its cofactors to the PSA enhancer in a ligand-dependent fashion. In addition, S-22 induced Xenopus laevis oocyte maturation and rapid phosphorylation of several kinases, through pathways distinct from steroids. These studies reveal novel differences in the molecular mechanisms by which S-22, a nonsteroidal SARM, and DHT mediate their pharmacological effects.

Replacing Escherichia coli NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH) with a NADP-dependent enzyme from Clostridium acetobutylicum facilitates NADPH dependent pathways.

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Martínez, Irene; Zhu, Jiangfeng; Lin, Henry; Bennett, George N; San, Ka-Yiu

2008-11-01

Reactions requiring reducing equivalents, NAD(P)H, are of enormous importance for the synthesis of industrially valuable compounds such as carotenoids, polymers, antibiotics and chiral alcohols among others. The use of whole-cell biocatalysis can reduce process cost by acting as catalyst and cofactor regenerator at the same time; however, product yields might be limited by cofactor availability within the cell. Thus, our study focussed on the genetic manipulation of a whole-cell system by modifying metabolic pathways and enzymes to improve the overall production process. In the present work, we genetically engineered an Escherichia coli strain to increase NADPH availability to improve the productivity of products that require NADPH in its biosynthesis. The approach involved an alteration of the glycolysis step where glyceraldehyde-3-phosphate (GAP) is oxidized to 1,3 bisphophoglycerate (1,3-BPG). This reaction is catalyzed by NAD-dependent endogenous glyceraldehyde-3-phosphate dehydrogenase (GAPDH) encoded by the gapA gene. We constructed a recombinant E. coli strain by replacing the native NAD-dependent gapA gene with a NADP-dependent GAPDH from Clostridium acetobutylicum, encoded by the gene gapC. The beauty of this approach is that the recombinant E. coli strain produces 2 mol of NADPH, instead of NADH, per mole of glucose consumed. Metabolic flux analysis showed that the flux through the pentose phosphate (PP) pathway, one of the main pathways that produce NADPH, was reduced significantly in the recombinant strain when compared to that of the parent strain. The effectiveness of the NADPH enhancing system was tested using the production of lycopene and epsilon-caprolactone as model systems using two different background strains. The recombinant strains, with increased NADPH availability, consistently showed significant higher productivity than the parent strains.

Comparisons of CAP88PC version 2.0 default parameters to site specific inputs

International Nuclear Information System (INIS)

Lehto, M. A.; Courtney, J. C.; Charter, N.; Egan, T.

2000-01-01

The effects of varying the input for the CAP88PC Version 2.0 program on the total effective dose equivalents (TEDEs) were determined for hypothetical releases from the Hot Fuel Examination Facility (HFEF) located at the Argonne National Laboratory site on the Idaho National Engineering and Environmental Laboratory (INEEL). Values for site specific meteorological conditions and agricultural production parameters were determined for the 80 km radius surrounding the HFEF. Four nuclides, 3 H, 85 Kr, 129 I, and 137 Cs (with its short lived progeny, 137m Ba) were selected for this study; these are the radioactive materials most likely to be released from HFEF under normal or abnormal operating conditions. Use of site specific meteorological parameters of annual precipitation, average temperature, and the height of the inversion layer decreased the TEDE from 137 Cs- 137m Ba up to 36%; reductions for other nuclides were less than 3%. Use of the site specific agricultural parameters reduced TEDE values between 7% and 49%, depending on the nuclide. Reductions are associated with decreased committed effective dose equivalents (CEDEs) from the ingestion pathway. This is not surprising since the HFEF is located well within the INEEL exclusion area, and the surrounding area closest to the release point is a high desert with limited agricultural diversity. Livestock and milk production are important in some counties at distances greater than 30 km from the HFEF

Diosgenin induces apoptosis in IGF-1-stimulated human thyrocytes through two caspase-dependent pathways

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Mu, Shumin [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Hospital Affiliated to Shandong Traditional Chinese Medicine University, Jinan 250011 (China); Institute of Endocrinology, Shandong Academy of Clinical Medicine, Jinan 250021 (China); Tian, Xingsong; Ruan, Yongwei [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Liu, Yuantao [The Second Hospital of Shandong University, Jinan 250033 (China); Bian, Dezhi [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Jining Medical College, Jining 272013 (China); Ma, Chunyan [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Yu, Chunxiao [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Institute of Endocrinology, Shandong Academy of Clinical Medicine, Jinan 250021 (China); Feng, Mei [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Wang, Furong [Shandong University of Traditional Chinese Medicine, Jinan 250011 (China); Gao, Ling [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Institute of Endocrinology, Shandong Academy of Clinical Medicine, Jinan 250021 (China); Zhao, Jia-jun, E-mail: jjzhao@medmail.com.cn [Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Institute of Endocrinology, Shandong Academy of Clinical Medicine, Jinan 250021 (China)

2012-02-10

Highlights: Black-Right-Pointing-Pointer Diosgenin induces apoptosis in IGF-1-treated thyrocytes through two caspase pathways. Black-Right-Pointing-Pointer Diosgenin inhibits FLIP and activates caspase-8 in FAS related-pathway. Black-Right-Pointing-Pointer Diosgenin increases ROS, regulates the ratio of Bax/Bcl-2 in mitochondrial pathway. -- Abstract: Insulin-like growth factor-1 (IGF-1) is a growth factor of the thyroid that has been shown in our previous study to possess proliferative and antiapoptotic effects in FRTL-5 cell lines through the upregulation of cyclin D and Fas-associated death domain-like interleukin-1-converting enzyme (FLICE)-inhibitory protein (FLIP). Diosgenin, a natural steroid sapogenin from plants, has been shown to induce apoptosis in many cell lines, with the exception of thyroid cells. In this report, we investigated the apoptotic effect and mechanism of diosgenin in IGF-1-stimulated primary human thyrocytes. Primary human thyrocytes were preincubated with or without IGF-1 for 24 h and subsequently exposed to varying concentrations of diosgenin for different times. We found that diosgenin induced apoptosis in human thyrocytes pretreated with IGF-1 in a dose-dependent manner through the activation of caspase cascades. Moreover, diosgenin inhibited FLIP and activated caspase-8 in the FAS-related apoptotic pathway. Diosgenin increased the production of ROS, regulated the balance of Bax and Bcl-2 and cleaved caspase-9 in the mitochondrial apoptotic pathway. These results indicate that diosgenin induces apoptosis in IGF-1-stimulated primary human thyrocytes through two caspase-dependent pathways.

Bendamustine and rituximab (BR) versus dexamethasone, rituximab, and cyclophosphamide (DRC) in patients with Waldenström macroglobulinemia.

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Paludo, Jonas; Abeykoon, Jithma P; Shreders, Amanda; Ansell, Stephen M; Kumar, Shaji; Ailawadhi, Sikander; King, Rebecca L; Koehler, Amber B; Reeder, Craig B; Buadi, Francis K; Dispenzieri, Angela; Lacy, Martha Q; Dingli, David; Witzig, Thomas E; Go, Ronald S; Gonsalves, Wilson I; Kourelis, Taxiarchis; Warsame, Rahma; Leung, Nelson; Habermann, Thomas M; Hayman, Suzanne; Lin, Yi; Kyle, Robert A; Rajkumar, S Vincent; Gertz, Morie A; Kapoor, Prashant

2018-04-03

The treatment approaches for Waldenstrom macroglobulinemia (WM) are largely based upon information from single-arm phase II trials, without comparative data. We compared the efficacy of two commonly used regimens in routine practice (bendamustine-rituximab (BR) and dexamethasone, rituximab plus cyclophosphamide (DRC)) and evaluated their activity with respect to the patients' MYD88 L265P mutation status. Of 160 consecutive patients, 60 received BR (43 with relapsed/refractory WM) and 100 received DRC (50 had relapsed/refractory WM). In the treatment-naïve setting, overall response rate (ORR) was 93% with BR versus 96% with DRC (p = 0.55). Two-year progression-free survival (PFS) with BR and DRC was 88 and 61%, respectively (p = 0.07). In salvage setting, ORR was 95% with BR versus 87% with DRC, p = 0.45; median PFS with BR was 58 versus 32 months with DRC (2-year PFS was 66 versus 53%; p = 0.08). Median disease-specific survival was not reached with BR versus 166 months with DRC (p = 0.51). The time-to-event endpoints and depth of response were independent of the MYD88 mutation status. Grade ≥ 3 adverse events of both regimens were comparable. A trend for longer PFS was observed with BR although the regimens have comparable toxicities. The activity of BR and DRC appears to be unaffected by patients' MYD88 mutation status.

Whole-genome sequencing identifies recurrent mutations in chronic lymphocytic leukaemia

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Puente, Xose S.; Pinyol, Magda; Quesada, Víctor; Conde, Laura; Ordóñez, Gonzalo R.; Villamor, Neus; Escaramis, Georgia; Jares, Pedro; Beà, Sílvia; González-Díaz, Marcos; Bassaganyas, Laia; Baumann, Tycho; Juan, Manel; López-Guerra, Mónica; Colomer, Dolors; Tubío, José M. C.; López, Cristina; Navarro, Alba; Tornador, Cristian; Aymerich, Marta; Rozman, María; Hernández, Jesús M.; Puente, Diana A.; Freije, José M. P.; Velasco, Gloria; Gutiérrez-Fernández, Ana; Costa, Dolors; Carrió, Anna; Guijarro, Sara; Enjuanes, Anna; Hernández, Lluís; Yagüe, Jordi; Nicolás, Pilar; Romeo-Casabona, Carlos M.; Himmelbauer, Heinz; Castillo, Ester; Dohm, Juliane C.; de Sanjosé, Silvia; Piris, Miguel A.; de Alava, Enrique; Miguel, Jesús San; Royo, Romina; Gelpí, Josep L.; Torrents, David; Orozco, Modesto; Pisano, David G.; Valencia, Alfonso; Guigó, Roderic; Bayés, Mónica; Heath, Simon; Gut, Marta; Klatt, Peter; Marshall, John; Raine, Keiran; Stebbings, Lucy A.; Futreal, P. Andrew; Stratton, Michael R.; Campbell, Peter J.; Gut, Ivo; López-Guillermo, Armando; Estivill, Xavier; Montserrat, Emili; López-Otín, Carlos; Campo, Elías

2012-01-01

Chronic lymphocytic leukaemia (CLL), the most frequent leukaemia in adults in Western countries, is a heterogeneous disease with variable clinical presentation and evolution1,2. Two major molecular subtypes can be distinguished, characterized respectively by a high or low number of somatic hypermutations in the variable region of immunoglobulin genes3,4. The molecular changes leading to the pathogenesis of the disease are still poorly understood. Here we performed whole-genome sequencing of four cases of CLL and identified 46 somatic mutations that potentially affect gene function. Further analysis of these mutations in 363 patients with CLL identified four genes that are recurrently mutated: notch 1 (NOTCH1), exportin 1 (XPO1), myeloid differentiation primary response gene 88 (MYD88) and kelch-like 6 (KLHL6). Mutations in MYD88 and KLHL6 are predominant in cases of CLL with mutated immunoglobulin genes, whereas NOTCH1 and XPO1 mutations are mainly detected in patients with unmutated immunoglobulins. The patterns of somatic mutation, supported by functional and clinical analyses, strongly indicate that the recurrent NOTCH1, MYD88 and XPO1 mutations are oncogenic changes that contribute to the clinical evolution of the disease. To our knowledge, this is the first comprehensive analysis of CLL combining whole-genome sequencing with clinical characteristics and clinical outcomes. It highlights the usefulness of this approach for the identification of clinically relevant mutations in cancer. PMID:21642962

Pathway Interaction Network Analysis Identifies Dysregulated Pathways in Human Monocytes Infected by Listeria monocytogenes

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Wufeng Fan

2017-01-01

Full Text Available In our study, we aimed to extract dysregulated pathways in human monocytes infected by Listeria monocytogenes (LM based on pathway interaction network (PIN which presented the functional dependency between pathways. After genes were aligned to the pathways, principal component analysis (PCA was used to calculate the pathway activity for each pathway, followed by detecting seed pathway. A PIN was constructed based on gene expression profile, protein-protein interactions (PPIs, and cellular pathways. Identifying dysregulated pathways from the PIN was performed relying on seed pathway and classification accuracy. To evaluate whether the PIN method was feasible or not, we compared the introduced method with standard network centrality measures. The pathway of RNA polymerase II pretranscription events was selected as the seed pathway. Taking this seed pathway as start, one pathway set (9 dysregulated pathways with AUC score of 1.00 was identified. Among the 5 hub pathways obtained using standard network centrality measures, 4 pathways were the common ones between the two methods. RNA polymerase II transcription and DNA replication owned a higher number of pathway genes and DEGs. These dysregulated pathways work together to influence the progression of LM infection, and they will be available as biomarkers to diagnose LM infection.

Examining the evidence for major histocompatibility complex-dependent mate selection in humans and nonhuman primates

Czech Academy of Sciences Publication Activity Database

Winternitz, Jamie Caroline; Abbate, J. L.

2015-01-01

Roč. 6, 13 May (2015), s. 73-88 ISSN 1179-7274 Institutional support: RVO:68081766 Keywords : major histocompatibility complex * sexual selection * olfaction * facial attraction * parasite resistance * inbreeding avoidance Subject RIV: EB - Genetics ; Molecular Biology

An intermediate state of T7 RNA polymerase provides another pathway of nucleotide selection

International Nuclear Information System (INIS)

Wang Zhan-Feng; Liu Yu-Ru; Wang Peng-Ye; Xie Ping

2017-01-01

Phage T7 RNA polymerase is a single-subunit transcription enzyme, transcribing template DNA to RNA. Nucleoside triphosphate (NTP) selection and translocation are two critical steps of the transcription elongation. Here, using all-atom molecular dynamics simulations, we found that between pre- and post-translocation states of T7 RNA polymerase an intermediate state exists, where the O helix C-terminal residue tyrosine 639, which plays important roles in translocation, locates between its pre- and post-translocation positions and the side chain of the next template DNA nucleotide has moved into the active site. NTP selection in this intermediate state was studied, revealing that the selection in the intermediate state can be achieved relying on the effect of Watson–Crick interaction between NTP and template DNA nucleotide, effect of stability of the components near the active site such as the nascent DNA–RNA hybrid and role of tyrosine 639. This indicates that another NTP-selection pathway can also exist besides the main pathway where NTP selection begins at the post-translocation state upon the entry of NTP. (paper)

The C-terminus of H-Ras as a target for the covalent binding of reactive compounds modulating Ras-dependent pathways.

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Clara L Oeste

2011-01-01

Full Text Available Ras proteins are crucial players in differentiation and oncogenesis and constitute important drug targets. The localization and activity of Ras proteins are highly dependent on posttranslational modifications at their C-termini. In addition to an isoprenylated cysteine, H-Ras, but not other Ras proteins, possesses two cysteine residues (C181 and C184 in the C-terminal hypervariable domain that act as palmitoylation sites in cells. Cyclopentenone prostaglandins (cyPG are reactive lipidic mediators that covalently bind to H-Ras and activate H-Ras dependent pathways. Dienone cyPG, such as 15-deoxy-Δ(12,14-PGJ(2 (15d-PGJ(2 and Δ(12-PGJ(2 selectively bind to the H-Ras hypervariable domain. Here we show that these cyPG bind simultaneously C181 and C184 of H-Ras, thus potentially altering the conformational tendencies of the hypervariable domain. Based on these results, we have explored the capacity of several bifunctional cysteine reactive small molecules to bind to the hypervariable domain of H-Ras proteins. Interestingly, phenylarsine oxide (PAO, a widely used tyrosine phosphatase inhibitor, and dibromobimane, a cross-linking agent used for cysteine mapping, effectively bind H-Ras hypervariable domain. The interaction of PAO with H-Ras takes place in vitro and in cells and blocks modification of H-Ras by 15d-PGJ(2. Moreover, PAO treatment selectively alters H-Ras membrane partition and the pattern of H-Ras activation in cells, from the plasma membrane to endomembranes. These results identify H-Ras as a novel target for PAO. More importantly, these observations reveal that small molecules or reactive intermediates interacting with spatially vicinal cysteines induce intramolecular cross-linking of H-Ras C-terminus potentially contributing to the modulation of Ras-dependent pathways.

Developmental pathways from child maltreatment to adolescent marijuana dependence: Examining moderation by FK506 binding protein 5 gene (FKBP5).

Science.gov (United States)

Handley, Elizabeth D; Rogosch, Fred A; Cicchetti, Dante

2015-11-01

The current study examined the prospective association between child maltreatment and the development of substance use disorder in adolescence with the aim of investigating pathways underlying this relation, as well as genetic moderation of these developmental mechanisms. Specifically, we tested whether youth who experienced maltreatment prior to age 8 were at risk for the development of marijuana dependence in adolescence by way of a childhood externalizing pathway and a childhood internalizing pathway. Moreover, we tested whether variation in FK506 binding protein 5 gene (FKBP5) CATT haplotype moderated these pathways. The participants were 326 children (n =179 maltreated; n = 147 nonmaltreated) assessed across two waves of data collection (childhood: ages 7-9 and adolescence: ages 15-18). Results indicated that higher levels of child externalizing symptoms significantly mediated the effect of child maltreatment on adolescent marijuana dependence symptoms for individuals with one or two copies of the FKBP5 CATT haplotype only. We did not find support for an internalizing pathway from child maltreatment to adolescent marijuana dependence, nor did we find evidence of moderation of the internalizing pathway by FKBP5 haplotype variation. Findings extend previous research by demonstrating that whether a maltreated child will traverse an externalizing pathway toward substance use disorder in adolescence is dependent on FKBP5 genetic variation.

Probiotic Bifidobacterium breve induces IL-10-producing Tr1 cells in the colon.

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Seong Gyu Jeon

Full Text Available Specific intestinal microbiota has been shown to induce Foxp3(+ regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103(+ dendritic cells (DCs mediated B. breve-induced development of IL-10-producing T cells. CD103(+ DCs from Il10(-/-, Tlr2(-/-, and Myd88(-/- mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103(+ DCs failed to induce IL-10 production from co-cultured Il27ra(-/- T cells. B. breve treatment of Tlr2(-/- mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103(+ DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4(+ T cells from wild-type mice, but not Il10(-/- mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.

Probiotic Bifidobacterium breve induces IL-10-producing Tr1 cells in the colon.

Science.gov (United States)

Jeon, Seong Gyu; Kayama, Hisako; Ueda, Yoshiyasu; Takahashi, Takuya; Asahara, Takashi; Tsuji, Hirokazu; Tsuji, Noriko M; Kiyono, Hiroshi; Ma, Ji Su; Kusu, Takashi; Okumura, Ryu; Hara, Hiromitsu; Yoshida, Hiroki; Yamamoto, Masahiro; Nomoto, Koji; Takeda, Kiyoshi

2012-01-01

Specific intestinal microbiota has been shown to induce Foxp3(+) regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103(+) dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103(+) DCs from Il10(-/-), Tlr2(-/-), and Myd88(-/-) mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103(+) DCs failed to induce IL-10 production from co-cultured Il27ra(-/-) T cells. B. breve treatment of Tlr2(-/-) mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103(+) DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4(+) T cells from wild-type mice, but not Il10(-/-) mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.

Toll-Like Receptor 4 Activation Contributes to Diabetic Bladder Dysfunction in a Murine Model of Type 1 Diabetes.

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Szasz, Theodora; Wenceslau, Camilla F; Burgess, Beth; Nunes, Kenia P; Webb, R Clinton

2016-12-01

Diabetic bladder dysfunction (DBD) is a common urological complication of diabetes. Innate immune system activation via Toll-like receptor 4 (TLR4) leads to inflammation and oxidative stress and was implicated in diabetes pathophysiology. We hypothesized that bladder hypertrophy and hypercontractility in DBD is mediated by TLR4 activation. Wild-type (WT) and TLR4 knockout (TLR4KO) mice were made diabetic by streptozotocin (STZ) treatment, and bladder contractile function and TLR4 pathway expression were evaluated. Immunohistochemistry confirmed the expression of TLR4 in human and mouse bladder. Recombinant high-mobility group box protein 1 (HMGB1) increased bladder TLR4 and MyD88 expression and enhanced contractile response to electrical field stimulation. Bladder expression of TLR4 and MyD88 and serum expression of HMGB1 were increased in STZ compared with control mice. Carbachol (CCh)-mediated contraction was increased in bladders from STZ mice, and TLR4 inhibitor CLI-095 attenuated this increase. Induction of diabetes by STZ in WT mice increased bladder weight and contractile responses to CCh and to electrical field stimulation. TLR4KO mice were not protected from STZ-induced diabetes; however, despite levels of hyperglycemia similar to those of WT STZ mice, TLR4KO STZ mice were protected from diabetes-induced bladder hypertrophy and hypercontractility. These data suggest that TLR4 activation during diabetes mediates DBD-associated bladder hypertrophy and hypercontractility. © 2016 by the American Diabetes Association.

Brucella TIR-like protein TcpB/Btp1 specifically targets the host adaptor protein MAL/TIRAP to promote infection.

Science.gov (United States)

Li, Wenna; Ke, Yuehua; Wang, Yufei; Yang, Mingjuan; Gao, Junguang; Zhan, Shaoxia; Xinying, Du; Huang, Liuyu; Li, Wenfeng; Chen, Zeliang; Li, Juan

2016-08-26

Brucella spp. are known to avoid host immune recognition and weaken the immune response to infection. Brucella like accomplish this by employing two clever strategies, called the stealth strategy and hijacking strategy. The TIR domain-containing protein (TcpB/Btp1) of Brucella melitensis is thought to be involved in inhibiting host NF-κB activation by binding to adaptors downstream of Toll-like receptors. However, of the five TIR domain-containing adaptors conserved in mammals, whether MyD88 or MAL, even other three adaptors, are specifically targeted by TcpB has not been identified. Here, we confirmed the effect of TcpB on B.melitensis virulence in mice and found that TcpB selectively targets MAL. By using siRNA against MAL, we found that TcpB from B.melitensis is involved in intracellular survival and that MAL affects intracellular replication of B.melitensis. Our results confirm that TcpB specifically targets MAL/TIRAP to disrupt downstream signaling pathways and promote intra-host survival of Brucella spp. Copyright © 2016 Elsevier Inc. All rights reserved.

Toll-Like Receptor 9-Dependent AMPKα Activation Occurs via TAK1 and Contributes to RhoA/ROCK Signaling and Actin Polymerization in Vascular Smooth Muscle Cells.

Science.gov (United States)

McCarthy, Cameron G; Wenceslau, Camilla F; Ogbi, Safia; Szasz, Theodora; Webb, R Clinton

2018-04-01

Traditionally, Toll-like receptor 9 (TLR9) signals through an MyD88-dependent cascade that results in proinflammatory gene transcription. Recently, it was reported that TLR9 also participates in a stress tolerance signaling cascade in nonimmune cells. In this noncanonical pathway, TLR9 binds to and inhibits sarcoplasmic/endoplasmic reticulum Ca 2+ -ATPase 2 (SERCA2), modulating intracellular calcium handling, and subsequently resulting in the activation of 5'-AMP-activated protein kinase α (AMPK α ). We have previously reported that TLR9 causes increased contraction in isolated arteries; however, the mechanisms underlying this vascular dysfunction need to be further clarified. Therefore, we hypothesized that noncanonical TLR9 signaling was also present in vascular smooth muscle cells (VSMCs) and that it mediates enhanced contractile responses through SERCA2 inhibition. To test these hypotheses, aortic microsomes, aortic VSMCs, and isolated arteries from male Sprague-Dawley rats were incubated with vehicle or TLR9 agonist (ODN2395). Despite clear AMPK α activation after treatment with ODN2395, SERCA2 activity was unaffected. Alternatively, ODN2395 caused the phosphorylation of AMPK α via transforming growth factor β -activated kinase 1 (TAK1), a kinase involved in TLR9 inflammatory signaling. Downstream, we hypothesized that that TLR9 activation of AMPK α may be important in mediating actin cytoskeleton reorganization. ODN2395 significantly increased the filamentous-to-globular actin ratio, as well as indices of RhoA/Rho-associated protein kinase (ROCK) activation, with the latter being prevented by AMPK α inhibition. In conclusion, AMPK α phosphorylation after TLR9 activation in VSMCs appears to be an extension of traditional inflammatory signaling via TAK1, as opposed to SERCA2 inhibition and the noncanonical pathway. Nonetheless, TLR9-AMPK α signaling can mediate VSMC function via RhoA/ROCK activation and actin polymerization. Copyright © 2018 by The

Subversion of innate defenses by the interplay between DENV and pre-existing enhancing antibodies: TLRs signaling collapse.

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Naphak Modhiran

Full Text Available BACKGROUND: The phenomenon of antibody dependent enhancement as a major determinant that exacerbates disease severity in DENV infections is well accepted. While the detailed mechanism of antibody enhanced disease severity is unclear, evidence suggests that it is associated with both increased DENV infectivity and suppression of the type I IFN and pro-inflammatory cytokine responses. Therefore, it is imperative for us to understand the intracellular mechanisms altered during ADE infection to decipher the mechanism of severe pathogenesis. METHODOLOGY/PRINCIPAL FINDINGS: In this present work, qRT-PCR, immunoblotting and gene array analysis were conducted to determine whether DENV-antibody complex infection exerts a suppressive effect on the expression and/or function of the pathogen recognition patterns, focusing on the TLR-signaling pathway. We show here that FcγRI and FcγRIIa synergistically facilitated entry of DENV-antibody complexes into monocytic THP-1 cells. Ligation between DENV-antibody complexes and FcR not only down regulated TLRs gene expression but also up regulated SARM, TANK, and negative regulators of the NF-κB pathway, resulting in suppression of innate responses but increased viral production. These results were confirmed by blocking with anti-FcγRI or anti-FcγRIIa antibodies which reduced viral production, up-regulated IFN-β synthesis, and increased gene expression in the TLR-dependent signaling pathway. The negative impact of DENV-ADE infection on the TLR-dependent pathway was strongly supported by gene array screening which revealed that both MyD88-dependent and -independent signaling molecules were down regulated during DENV-ADE infection. Importantly, the same phenomenon was seen in PBMC of secondary DHF/DSS patients but not in PBMC of DF patients. CONCLUSIONS/SIGNIFICANCE: Our present work demonstrates the mechanism by which DENV uses pre-existing immune mediators to defeat the principal activating pathway of innate

Synaptic Mechanisms Generating Orientation Selectivity in the ON Pathway of the Rabbit Retina.

Science.gov (United States)

Venkataramani, Sowmya; Taylor, W Rowland

2016-03-16

Neurons that signal the orientation of edges within the visual field have been widely studied in primary visual cortex. Much less is known about the mechanisms of orientation selectivity that arise earlier in the visual stream. Here we examine the synaptic and morphological properties of a subtype of orientation-selective ganglion cell in the rabbit retina. The receptive field has an excitatory ON center, flanked by excitatory OFF regions, a structure similar to simple cell receptive fields in primary visual cortex. Examination of the light-evoked postsynaptic currents in these ON-type orientation-selective ganglion cells (ON-OSGCs) reveals that synaptic input is mediated almost exclusively through the ON pathway. Orientation selectivity is generated by larger excitation for preferred relative to orthogonal stimuli, and conversely larger inhibition for orthogonal relative to preferred stimuli. Excitatory orientation selectivity arises in part from the morphology of the dendritic arbors. Blocking GABAA receptors reduces orientation selectivity of the inhibitory synaptic inputs and the spiking responses. Negative contrast stimuli in the flanking regions produce orientation-selective excitation in part by disinhibition of a tonic NMDA receptor-mediated input arising from ON bipolar cells. Comparison with earlier studies of OFF-type OSGCs indicates that diverse synaptic circuits have evolved in the retina to detect the orientation of edges in the visual input. A core goal for visual neuroscientists is to understand how neural circuits at each stage of the visual system extract and encode features from the visual scene. This study documents a novel type of orientation-selective ganglion cell in the retina and shows that the receptive field structure is remarkably similar to that of simple cells in primary visual cortex. However, the data indicate that, unlike in the cortex, orientation selectivity in the retina depends on the activity of inhibitory interneurons. The

Elucidation of Altered Pathways in Tumor-Initiating Cells of Triple-Negative Breast Cancer: A Useful Cell Model System for Drug Screening.

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Christensen, Anne G; Ehmsen, Sidse; Terp, Mikkel G; Batra, Richa; Alcaraz, Nicolas; Baumbach, Jan; Noer, Julie B; Moreira, José; Leth-Larsen, Rikke; Larsen, Martin R; Ditzel, Henrik J

2017-08-01

A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation

Succinate modulates Ca(2+) transient and cardiomyocyte viability through PKA-dependent pathway.

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Aguiar, Carla J; Andrade, Vanessa L; Gomes, Enéas R M; Alves, Márcia N M; Ladeira, Marina S; Pinheiro, Ana Cristina N; Gomes, Dawidson A; Almeida, Alvair P; Goes, Alfredo M; Resende, Rodrigo R; Guatimosim, Silvia; Leite, M Fatima

2010-01-01

GPR91 is an orphan G-protein-coupled receptor (GPCR) that has been characterized as a receptor for succinate, a citric acid cycle intermediate, in several tissues. In the heart, the role of succinate is unknown. We now report that rat ventricular cardiomyocytes express GPR91. We found that succinate, through GPR91, increases the amplitude and the rate of decline of global Ca(2+) transient, by increasing the phosphorylation levels of ryanodine receptor and phospholamban, two well known Ca(2+) handling proteins. The effects of succinate on Ca(2+) transient were abolished by pre-treatment with adenylyl cyclase and cAMP-dependent protein kinase (PKA) inhibitors. Direct PKA activation by succinate was further confirmed using a FRET-based A-kinase activity reporter. Additionally, succinate decreases cardiomyocyte viability through a caspase-3 activation pathway, effect also prevented by PKA inhibition. Taken together, these observations show that succinate acts as a signaling molecule in cardiomyocytes, modulating global Ca(2+) transient and cell viability through a PKA-dependent pathway. 2009 Elsevier Ltd. All rights reserved.

Bullatacin Triggered ABCB1-Overexpressing Cell Apoptosis via the Mitochondrial-Dependent Pathway

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Yong-Ju Liang

2009-01-01

Full Text Available This paper was to explore bullatacin-mediated multidrug-resistant cell apoptosis at extremely low concentration. To investigate its precise mechanisms, the pathway of cell apoptosis induced by bullatacin was examined. Bullatacin causes an upregulation of ROS and a downregulation of ΔΨm in a concentration-dependent manner in ABCB1-overexpressing KBv200 cells. In addition, cleavers of caspase-9, caspase-3, and PARP were observed following the release of cytochrome c from mitochondria after bullatacin treatment. However, neither cleavage of caspase-8 nor change of expression level of bcl-2, bax and Fas was observed by the same treatment. Pretreating KBv200 cells with N-acetylcysteine, an antioxidant modulator, resulted in a significant reduction of ROS generation and cell apoptosis induced by bullatacin. Bullatacin-induced apoptosis was antagonized by z-LEHD-fmk, a caspase-9 inhibitor, but not by z-IETD-fmk, a caspase-8 inhibitor. These implied that apoptosis of KBv200 cells induced by bullatacin was associated with the mitochondria-dependent pathway that was limited to activation of apical caspase-9.

Backup pathways of NHEJ in cells of higher eukaryotes: Cell cycle dependence

International Nuclear Information System (INIS)

Iliakis, George

2009-01-01

DNA double-strand breaks (DSBs) induced by ionizing radiation (IR) in cells of higher eukaryotes are predominantly repaired by a pathway of non-homologous end joining (NHEJ) utilizing Ku, DNA-PKcs, DNA ligase IV, XRCC4 and XLF/Cernunnos (D-NHEJ) as central components. Work carried out in our laboratory and elsewhere shows that when this pathway is chemically or genetically compromised, cells do not shunt DSBs to homologous recombination repair (HRR) but instead use another form of NHEJ operating as a backup (B-NHEJ). Here I review our efforts to characterize this repair pathway and discuss its dependence on the cell cycle as well as on the growth conditions. I present evidence that B-NHEJ utilizes ligase III, PARP-1 and histone H1. When B-NHEJ is examined throughout the cell cycle, significantly higher activity is observed in G2 phase that cannot be attributed to HRR. Furthermore, the activity of B-NHEJ is compromised when cells enter the plateau phase of growth. Together, these observations uncover a repair pathway with unexpected biochemical constitution and interesting cell cycle and growth factor regulation. They generate a framework for investigating the mechanistic basis of HRR contribution to DSB repair.

Superconducting fluctuation effect in CaFe0.88Co0.12AsF

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Xiao, H.; Gao, B.; Ma, Y. H.; Li, X. J.; Mu, G.; Hu, T.

2016-11-01

Out-of-plane angular dependent torque measurements were performed on CaFe0.88Co0.12AsF single crystals. Superconducting fluctuations, featured by magnetic field enhanced and exponential temperature dependent diamagnetism, are observed above the superconducting transition temperature T c, which is similar to that of cuprate superconductors, but less pronounced. In addition, the ratio of T c versus superfluid density follows well the Uemura line of high-T c cuprates, which suggests the exotic nature of the superconductivity in CaFe0.88Co0.12AsF.

Listeria monocytogenes alters mast cell phenotype, mediator and osteopontin secretion in a listeriolysin-dependent manner.

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Catherine E Jobbings

Full Text Available Whilst mast cells participate in the immune defence against the intracellular bacterium Listeria monocytogenes, there is conflicting evidence regarding the ability of L. monocytogenes to infect mast cells. It is known that the pore-forming toxin listeriolysin (LLO is important for mast cell activation, degranulation and the release of pro-inflammatory cytokines. Mast cells, however, are a potential source of a wide range of cytokines, chemokines and other mediators including osteopontin, which contributes to the clearing of L. monocytogenes infections in vivo, although its source is unknown. We therefore aimed to resolve the controversy of mast cell infection by L. monocytogenes and investigated the extent of mediator release in response to the bacterium. In this paper we show that the infection of bone marrow-derived mast cells by L. monocytogenes is inefficient and LLO-independent. LLO, however, is required for calcium-independent mast cell degranulation as well as for the transient and selective downregulation of cell surface CD117 (c-kit on mast cells. We demonstrate that in addition to the key pro-inflammatory cytokines TNF-α and IL-6, mast cells release a wide range of other mediators in response to L. monocytogenes. Osteopontin, IL-2, IL-4, IL-13 and granulocyte macrophage colony-stimulating factor (GM-CSF, and chemokines including CCL2, CCL3, CCL4 and CCL5 are released in a MyD88-dependent manner. The wide range of mediators released by mast cells in response to L. monocytogenes may play an important role in the recruitment and activation of a variety of immune cells in vivo. The cocktail of mediators, however, is unlikely to skew the immune response to a particular effector response. We propose that mast cells provide a hitherto unreported source of osteopontin, and may provide an important role in co-ordinating the immune response during Listeria infection.

Cloning, annotation and expression analysis of mycoparasitism-related genes in Trichoderma harzianum 88.

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Yao, Lin; Yang, Qian; Song, Jinzhu; Tan, Chong; Guo, Changhong; Wang, Li; Qu, Lianhai; Wang, Yun

2013-04-01

Trichoderma harzianum 88, a filamentous soil fungus, is an effective biocontrol agent against several plant pathogens. High-throughput sequencing was used here to study the mycoparasitism mechanisms of T. harzianum 88. Plate confrontation tests of T. harzianum 88 against plant pathogens were conducted, and a cDNA library was constructed from T. harzianum 88 mycelia in the presence of plant pathogen cell walls. Randomly selected transcripts from the cDNA library were compared with eukaryotic plant and fungal genomes. Of the 1,386 transcripts sequenced, the most abundant Gene Ontology (GO) classification group was "physiological process". Differential expression of 19 genes was confirmed by real-time RT-PCR at different mycoparasitism stages against plant pathogens. Gene expression analysis revealed the transcription of various genes involved in mycoparasitism of T. harzianum 88. Our study provides helpful insights into the mechanisms of T. harzianum 88-plant pathogen interactions.

How I treat Waldenström macroglobulinemia.

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Treon, Steven P

2015-08-06

Waldenström macroglobulinemia (WM) is a B-cell neoplasm manifested by the accumulation of clonal immunoglobulin (Ig)M-secreting lymphoplasmacytic cells. MYD88 and CXCR4 warts, hypogammaglobulinemia, infections, myelokathexis syndrome-like somatic mutations are present in >90% and 30% to 35% of WM patients, respectively, and impact disease presentation, treatment outcome, and overall survival. Familial predisposition is common in WM. Asymptomatic patients should be observed. Patients with disease-related hemoglobin ibrutinib. In the salvage setting, an alternative frontline regimen, ibrutinib, everolimus, or stem cell transplantation can be considered. Investigational therapies under development for WM include agents that target MYD88, CXCR4, BCL2, and CD27/CD70 signaling, novel proteasome inhibitors, and chimeric antigen receptor-modified T-cell therapy. © 2015 by The American Society of Hematology.

Malaria-induced NLRP12/NLRP3-dependent caspase-1 activation mediates inflammation and hypersensitivity to bacterial superinfection.

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Marco A Ataide

2014-01-01

Full Text Available Cyclic paroxysm and high fever are hallmarks of malaria and are associated with high levels of pyrogenic cytokines, including IL-1β. In this report, we describe a signature for the expression of inflammasome-related genes and caspase-1 activation in malaria. Indeed, when we infected mice, Plasmodium infection was sufficient to promote MyD88-mediated caspase-1 activation, dependent on IFN-γ-priming and the expression of inflammasome components ASC, P2X7R, NLRP3 and/or NLRP12. Pro-IL-1β expression required a second stimulation with LPS and was also dependent on IFN-γ-priming and functional TNFR1. As a consequence of Plasmodium-induced caspase-1 activation, mice produced extremely high levels of IL-1β upon a second microbial stimulus, and became hypersensitive to septic shock. Therapeutic intervention with IL-1 receptor antagonist prevented bacterial-induced lethality in rodents. Similar to mice, we observed a significantly increased frequency of circulating CD14(+CD16(-Caspase-1(+ and CD14(dimCD16(+Caspase-1(+ monocytes in peripheral blood mononuclear cells from febrile malaria patients. These cells readily produced large amounts of IL-1β after stimulation with LPS. Furthermore, we observed the presence of inflammasome complexes in monocytes from malaria patients containing either NLRP3 or NLRP12 pyroptosomes. We conclude that NLRP12/NLRP3-dependent activation of caspase-1 is likely to be a key event in mediating systemic production of IL-1β and hypersensitivity to secondary bacterial infection during malaria.

Metabolic Engineering to Develop a Pathway for the Selective Cleavage of Carbon-Nitrogen Bonds

Energy Technology Data Exchange (ETDEWEB)

John J. Kilbane II

2005-10-01

The objective of the project is to develop a biochemical pathway for the selective cleavage of C-N bonds in molecules found in petroleum. Specifically a novel biochemical pathway will be developed for the selective cleavage of C-N bonds in carbazole. The cleavage of the first C-N bond in carbazole is accomplished by the enzyme carbazole dioxygenase, that catalyzes the conversion of carbazole to 2-aminobiphenyl-2,3-diol. The genes encoding carbazole dioxygenase were cloned from Sphingomonas sp. GTIN11 and from Pseudomonas resinovorans CA10. The selective cleavage of the second C-N bond has been challenging, and efforts to overcome that challenge have been the focus of recent research in this project. Enrichment culture experiments succeeded in isolating bacterial cultures that can metabolize 2-aminobiphenyl, but no enzyme capable of selectively cleaving the C-N bond in 2-aminobiphenyl has been identified. Aniline is very similar to the structure of 2-aminobiphenyl and aniline dioxygenase catalyzes the conversion of aniline to catechol and ammonia. For the remainder of the project the emphasis of research will be to simultaneously express the genes for carbazole dioxygenase and for aniline dioxygenase in the same bacterial host and then to select for derivative cultures capable of using carbazole as the sole source of nitrogen.

A second component of the SltA-dependent cation tolerance pathway in Aspergillus nidulans.

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Mellado, Laura; Calcagno-Pizarelli, Ana Maria; Lockington, Robin A; Cortese, Marc S; Kelly, Joan M; Arst, Herbert N; Espeso, Eduardo A

2015-09-01

The transcriptional response to alkali metal cation stress is mediated by the zinc finger transcription factor SltA in Aspergillus nidulans and probably in other fungi of the pezizomycotina subphylum. A second component of this pathway has been identified and characterized. SltB is a 1272 amino acid protein with at least two putative functional domains, a pseudo-kinase and a serine-endoprotease, involved in signaling to the transcription factor SltA. Absence of SltB activity results in nearly identical phenotypes to those observed for a null sltA mutant. Hypersensitivity to a variety of monovalent and divalent cations, and to medium alkalinization are among the phenotypes exhibited by a null sltB mutant. Calcium homeostasis is an exception and this cation improves growth of sltΔ mutants. Moreover, loss of kinase HalA in conjunction with loss-of-function sltA or sltB mutations leads to pronounced calcium auxotrophy. sltA sltB double null mutants display a cation stress sensitive phenotype indistinguishable from that of single slt mutants showing the close functional relationship between these two proteins. This functional relationship is reinforced by the fact that numerous mutations in both slt loci can be isolated as suppressors of poor colonial growth resulting from certain null vps (vacuolar protein sorting) mutations. In addition to allowing identification of sltB, our sltB missense mutations enabled prediction of functional regions in the SltB protein. Although the relationship between the Slt and Vps pathways remains enigmatic, absence of SltB, like that of SltA, leads to vacuolar hypertrophy. Importantly, the phenotypes of selected sltA and sltB mutations demonstrate that suppression of null vps mutations is not dependent on the inability to tolerate cation stress. Thus a specific role for both SltA and SltB in the VPS pathway seems likely. Finally, it is noteworthy that SltA and SltB have a similar, limited phylogenetic distribution, being restricted to

Essential role of TGF-beta/Smad pathway on statin dependent vascular smooth muscle cell regulation.

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Juan Rodríguez-Vita

Full Text Available BACKGROUND: The 3-hydroxy-3-methylglutaryl CoA reductase inhibitors (also called statins exert proven beneficial effects on cardiovascular diseases. Recent data suggest a protective role for Transforming Growth Factor-beta (TGF-beta in atherosclerosis by regulating the balance between inflammation and extracellular matrix accumulation. However, there are no studies about the effect of statins on TGF-beta/Smad pathway in atherosclerosis and vascular cells. METHODOLOGY: In cultured vascular smooth muscle cells (VSMCs statins enhanced Smad pathway activation caused by TGF-beta. In addition, statins upregulated TGF-beta receptor type II (TRII, and increased TGF-beta synthesis and TGF-beta/Smad-dependent actions. In this sense, statins, through Smad activation, render VSMCs more susceptible to TGF-beta induced apoptosis and increased TGF-beta-mediated ECM production. It is well documented that high doses of statins induce apoptosis in cultured VSMC in the presence of serum; however the precise mechanism of this effect remains to be elucidated. We have found that statins-induced apoptosis was mediated by TGF-beta/Smad pathway. Finally, we have described that RhoA inhibition is a common intracellular mechanisms involved in statins effects. The in vivo relevance of these findings was assessed in an experimental model of atherosclerosis in apolipoprotein E deficient mice: Treatment with Atorvastatin increased Smad3 phosphorylation and TRII overexpression, associated to elevated ECM deposition in the VSMCs within atheroma plaques, while apoptosis was not detected. CONCLUSIONS: Statins enhance TGF-beta/Smad pathway, regulating ligand levels, receptor, main signaling pathway and cellular responses of VSMC, including apoptosis and ECM accumulation. Our findings show that TGF-beta/Smad pathway is essential for statins-dependent actions in VSMCs.

Nucleolus-derived mediators in oncogenic stress response and activation of p53-dependent pathways.

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Stępiński, Dariusz

2016-08-01

Rapid growth and division of cells, including tumor ones, is correlated with intensive protein biosynthesis. The output of nucleoli, organelles where translational machineries are formed, depends on a rate of particular stages of ribosome production and on accessibility of elements crucial for their effective functioning, including substrates, enzymes as well as energy resources. Different factors that induce cellular stress also often lead to nucleolar dysfunction which results in ribosome biogenesis impairment. Such nucleolar disorders, called nucleolar or ribosomal stress, usually affect cellular functioning which in fact is a result of p53-dependent pathway activation, elicited as a response to stress. These pathways direct cells to new destinations such as cell cycle arrest, damage repair, differentiation, autophagy, programmed cell death or aging. In the case of impaired nucleolar functioning, nucleolar and ribosomal proteins mediate activation of the p53 pathways. They are also triggered as a response to oncogenic factor overexpression to protect tissues and organs against extensive proliferation of abnormal cells. Intentional impairment of any step of ribosome biosynthesis which would direct the cells to these destinations could be a strategy used in anticancer therapy. This review presents current knowledge on a nucleolus, mainly in relation to cancer biology, which is an important and extremely sensitive element of the mechanism participating in cellular stress reaction mediating activation of the p53 pathways in order to counteract stress effects, especially cancer development.

Flexible metabolic pathway construction using modular and divisible selection gene regulators

DEFF Research Database (Denmark)

Rugbjerg, Peter; Myling-Petersen, Nils; Sommer, Morten Otto Alexander

2015-01-01

Genetic selections are important to biological engineering. Although selectable traits are limited,currently each trait only permits simultaneous introduction of a single DNA fragment. Complex pathwayand strain construction however depends on rapid, combinatorial introduction of many genes thaten...

Arctigenin Inhibits Osteoclast Differentiation and Function by Suppressing Both Calcineurin-Dependent and Osteoblastic Cell-Dependent NFATc1 Pathways

OpenAIRE

Yamashita, Teruhito; Uehara, Shunsuke; Udagawa, Nobuyuki; Li, Feng; Kadota, Shigetoshi; Esumi, Hiroyasu; Kobayashi, Yasuhiro; Takahashi, Naoyuki

2014-01-01

Arctigenin, a lignan-derived compound, is a constituent of the seeds of Arctium lappa. Arctigenin was previously shown to inhibit osteoclastogenesis; however, this inhibitory mechanism has yet to be elucidated. Here, we showed that arctigenin inhibited the action of nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), a key transcription factor for osteoclastogenesis. NFATc1 in osteoclast precursors was activated through two distinct pathways: the calcineurin-dependent and osteoblasti...

A non-canonical RNA degradation pathway suppresses RNAi-dependent epimutations in the human fungal pathogen Mucor circinelloides.

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Calo, Silvia; Nicolás, Francisco E; Lee, Soo Chan; Vila, Ana; Cervantes, Maria; Torres-Martinez, Santiago; Ruiz-Vazquez, Rosa M; Cardenas, Maria E; Heitman, Joseph

2017-03-01

Mucorales are a group of basal fungi that includes the casual agents of the human emerging disease mucormycosis. Recent studies revealed that these pathogens activate an RNAi-based pathway to rapidly generate drug-resistant epimutant strains when exposed to stressful compounds such as the antifungal drug FK506. To elucidate the molecular mechanism of this epimutation pathway, we performed a genetic analysis in Mucor circinelloides that revealed an inhibitory role for the non-canonical RdRP-dependent Dicer-independent silencing pathway, which is an RNAi-based mechanism involved in mRNA degradation that was recently identified. Thus, mutations that specifically block the mRNA degradation pathway, such as those in the genes r3b2 and rdrp3, enhance the production of drug resistant epimutants, similar to the phenotype previously described for mutation of the gene rdrp1. Our genetic analysis also revealed two new specific components of the epimutation pathway related to the quelling induced protein (qip) and a Sad-3-like helicase (rnhA), as mutations in these genes prevented formation of drug-resistant epimutants. Remarkably, drug-resistant epimutant production was notably increased in M. circinelloides f. circinelloides isolates from humans or other animal hosts. The host-pathogen interaction could be a stressful environment in which the phenotypic plasticity provided by the epimutant pathway might provide an advantage for these strains. These results evoke a model whereby balanced regulation of two different RNAi pathways is determined by the activation of the RNAi-dependent epimutant pathway under stress conditions, or its repression when the regular maintenance of the mRNA degradation pathway operates under non-stress conditions.

A non-canonical RNA degradation pathway suppresses RNAi-dependent epimutations in the human fungal pathogen Mucor circinelloides.

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Silvia Calo

2017-03-01

Full Text Available Mucorales are a group of basal fungi that includes the casual agents of the human emerging disease mucormycosis. Recent studies revealed that these pathogens activate an RNAi-based pathway to rapidly generate drug-resistant epimutant strains when exposed to stressful compounds such as the antifungal drug FK506. To elucidate the molecular mechanism of this epimutation pathway, we performed a genetic analysis in Mucor circinelloides that revealed an inhibitory role for the non-canonical RdRP-dependent Dicer-independent silencing pathway, which is an RNAi-based mechanism involved in mRNA degradation that was recently identified. Thus, mutations that specifically block the mRNA degradation pathway, such as those in the genes r3b2 and rdrp3, enhance the production of drug resistant epimutants, similar to the phenotype previously described for mutation of the gene rdrp1. Our genetic analysis also revealed two new specific components of the epimutation pathway related to the quelling induced protein (qip and a Sad-3-like helicase (rnhA, as mutations in these genes prevented formation of drug-resistant epimutants. Remarkably, drug-resistant epimutant production was notably increased in M. circinelloides f. circinelloides isolates from humans or other animal hosts. The host-pathogen interaction could be a stressful environment in which the phenotypic plasticity provided by the epimutant pathway might provide an advantage for these strains. These results evoke a model whereby balanced regulation of two different RNAi pathways is determined by the activation of the RNAi-dependent epimutant pathway under stress conditions, or its repression when the regular maintenance of the mRNA degradation pathway operates under non-stress conditions.

Modulation of the intestinal microbiota alters colitis-associated colorectal cancer susceptibility.

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Joshua M Uronis

2009-06-01

Full Text Available It is well established that the intestinal microbiota plays a key role in the pathogenesis of Crohn's disease (CD and ulcerative colitis (UC collectively referred to as inflammatory bowel disease (IBD. Epidemiological studies have provided strong evidence that IBD patients bear increased risk for the development of colorectal cancer (CRC. However, the impact of the microbiota on the development of colitis-associated cancer (CAC remains largely unknown. In this study, we established a new model of CAC using azoxymethane (AOM-exposed, conventionalized-Il10(-/- mice and have explored the contribution of the host intestinal microbiota and MyD88 signaling to the development of CAC. We show that 8/13 (62% of AOM-Il10(-/- mice developed colon tumors compared to only 3/15 (20% of AOM- wild-type (WT mice. Conventionalized AOM-Il10(-/- mice developed spontaneous colitis and colorectal carcinomas while AOM-WT mice were colitis-free and developed only rare adenomas. Importantly, tumor multiplicity directly correlated with the presence of colitis. Il10(-/- mice mono-associated with the mildly colitogenic bacterium Bacteroides vulgatus displayed significantly reduced colitis and colorectal tumor multiplicity compared to Il10(-/- mice. Germ-free AOM-treated Il10(-/- mice showed normal colon histology and were devoid of tumors. Il10(-/-; Myd88(-/- mice treated with AOM displayed reduced expression of Il12p40 and Tnfalpha mRNA and showed no signs of tumor development. We present the first direct demonstration that manipulation of the intestinal microbiota alters the development of CAC. The TLR/MyD88 pathway is essential for microbiota-induced development of CAC. Unlike findings obtained using the AOM/DSS model, we demonstrate that the severity of chronic colitis directly correlates to colorectal tumor development and that bacterial-induced inflammation drives progression from adenoma to invasive carcinoma.

Glutamate alleviates muscle protein loss by modulating TLR4, NODs, Akt/FOXO and mTOR signaling pathways in LPS-challenged piglets.

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Ping Kang

Full Text Available The experiment was conducted to study the effect of the glutamate (Glu on muscle protein loss through toll-like receptor 4 (TLR4, nucleotide-binding oligomerization domain proteins (NODs, Akt/Forkhead Box O (Akt/FOXO and mammalian target of rapamycin (mTOR signaling pathways in LPS-challenged piglets. Twenty-four weaned piglets were assigned into four treatments: (1 Control; (2 LPS+0% Glu; (3 LPS + 1.0% Glu; (4 LPS + 2.0% Glu. The experiment was lasted for 28 days. On d 28, the piglets in the LPS challenged groups were injected with LPS on 100 μg/kg body weight (BW, and the piglets in the control group were injected with the same volume of 0.9% NaCl solution. After 4 h LPS or saline injection, the piglets were slaughtered and the muscle samples were collected. Glu supplementation increased the protein/DNA ratio in gastrocnemius muscle, and the protein content in longissimus dorsi (LD muscle after LPS challenge (P<0.05. In addition, Glu supplementation decreased TLR4, IL-1 receptor-associated kinase (IRAK 1, receptor-interacting serine/threonine-protein kinase (RIPK 2, and nuclear factor-κB (NF-κB mRNA expression in gastrocnemius muscle (P<0.05, MyD88 mRNA expression in LD muscle, and FOXO1 mRNA expression in LD muscle (P<0.05. Moreover, Glu supplementation increased p-Akt/t-Akt ratio (P<0.05 in gastrocnemius muscle, and p-4EBP1/t-4EBP1 ratio in both gastrocnemius and LD muscles (P<0.05. Glu supplementation in the piglets' diets might be an effective strategy to alleviate LPS-induced muscle protein loss, which might be due to suppressing the mRNA expression of TLR4 and NODs signaling-related genes, and modulating Akt/FOXO and mTOR signaling pathways.

Exosome RNA Released by Hepatocytes Regulates Innate Immune Responses to Hepatitis B Virus Infection

Directory of Open Access Journals (Sweden)

Takahisa Kouwaki

2016-08-01

Full Text Available The innate immune system is essential for controlling viral infection. Hepatitis B virus (HBV persistently infects human hepatocytes and causes hepatocellular carcinoma. However, the innate immune response to HBV infection in vivo remains unclear. Using a tree shrew animal model, we showed that HBV infection induced hepatic interferon (IFN-γ expression during early infection. Our in vitro study demonstrated that hepatic NK cells produced IFN-γ in response to HBV only in the presence of hepatic F4/80+ cells. Moreover, extracellular vesicles released from HBV-infected hepatocytes contained viral nucleic acids and induced NKG2D ligand expression in macrophages by stimulating MyD88, TICAM-1, and MAVS-dependent pathways. In addition, depletion of exosomes from extracellular vesicles markedly reduced NKG2D ligand expression, suggesting the importance of exosomes for NK cell activation. In contrast, infection of hepatocytes with HBV increased immunoregulatory microRNA levels in extracellular vesicles and exosomes, which were transferred to macrophages, thereby suppressing IL-12p35 mRNA expression in macrophages to counteract the host innate immune response. IFN-γ increased the hepatic expression of DDX60 and augmented the DDX60-dependent degradation of cytoplasmic HBV RNA. Our results elucidated the crucial role of exosomes in antiviral innate immune response against HBV.

Dependency of a therapy-resistant state of cancer cells on a lipid peroxidase pathway.

Science.gov (United States)

Viswanathan, Vasanthi S; Ryan, Matthew J; Dhruv, Harshil D; Gill, Shubhroz; Eichhoff, Ossia M; Seashore-Ludlow, Brinton; Kaffenberger, Samuel D; Eaton, John K; Shimada, Kenichi; Aguirre, Andrew J; Viswanathan, Srinivas R; Chattopadhyay, Shrikanta; Tamayo, Pablo; Yang, Wan Seok; Rees, Matthew G; Chen, Sixun; Boskovic, Zarko V; Javaid, Sarah; Huang, Cherrie; Wu, Xiaoyun; Tseng, Yuen-Yi; Roider, Elisabeth M; Gao, Dong; Cleary, James M; Wolpin, Brian M; Mesirov, Jill P; Haber, Daniel A; Engelman, Jeffrey A; Boehm, Jesse S; Kotz, Joanne D; Hon, Cindy S; Chen, Yu; Hahn, William C; Levesque, Mitchell P; Doench, John G; Berens, Michael E; Shamji, Alykhan F; Clemons, Paul A; Stockwell, Brent R; Schreiber, Stuart L

2017-07-27

Plasticity of the cell state has been proposed to drive resistance to multiple classes of cancer therapies, thereby limiting their effectiveness. A high-mesenchymal cell state observed in human tumours and cancer cell lines has been associated with resistance to multiple treatment modalities across diverse cancer lineages, but the mechanistic underpinning for this state has remained incompletely understood. Here we molecularly characterize this therapy-resistant high-mesenchymal cell state in human cancer cell lines and organoids and show that it depends on a druggable lipid-peroxidase pathway that protects against ferroptosis, a non-apoptotic form of cell death induced by the build-up of toxic lipid peroxides. We show that this cell state is characterized by activity of enzymes that promote the synthesis of polyunsaturated lipids. These lipids are the substrates for lipid peroxidation by lipoxygenase enzymes. This lipid metabolism creates a dependency on pathways converging on the phospholipid glutathione peroxidase (GPX4), a selenocysteine-containing enzyme that dissipates lipid peroxides and thereby prevents the iron-mediated reactions of peroxides that induce ferroptotic cell death. Dependency on GPX4 was found to exist across diverse therapy-resistant states characterized by high expression of ZEB1, including epithelial-mesenchymal transition in epithelial-derived carcinomas, TGFβ-mediated therapy-resistance in melanoma, treatment-induced neuroendocrine transdifferentiation in prostate cancer, and sarcomas, which are fixed in a mesenchymal state owing to their cells of origin. We identify vulnerability to ferroptic cell death induced by inhibition of a lipid peroxidase pathway as a feature of therapy-resistant cancer cells across diverse mesenchymal cell-state contexts.

Deciphering the contribution of human meningothelial cells to the inflammatory and antimicrobial response at the meninges.

Science.gov (United States)

Royer, Pierre-Joseph; Rogers, Andrew J; Wooldridge, Karl G; Tighe, Patrick; Mahdavi, Jafar; Rittig, Michael G; Ala'Aldeen, Dlawer

2013-11-01

We have investigated the response of primary human meningothelial cells to Neisseria meningitidis. Through a transcriptome analysis, we provide a comprehensive examination of the response of meningothelial cells to bacterial infection. A wide range of chemokines are elicited which act to attract and activate the main players of innate and adaptive immunity. We showed that meningothelial cells expressed a high level of Toll-like receptor 4 (TLR4), and, using a gene silencing strategy, we demonstrated the contribution of this pathogen recognition receptor in meningothelial cell activation. Secretion of interleukin-6 (IL-6), CXCL10, and CCL5 was almost exclusively TLR4 dependent and relied on MyD88 and TRIF adaptor cooperation. In contrast, IL-8 induction was independent of the presence of TLR4, MyD88, and TRIF. Transcription factors NF-κB p65, p38 mitogen-activated protein kinase (MAPK), Jun N-terminal protein kinase (JNK1), IRF3, and IRF7 were activated after contact with bacteria. Interestingly, the protein kinase IRAK4 was found to play a minor role in the meningothelial cell response to Neisseria infection. Our work highlights the role of meningothelial cells in the development of an immune response and inflammation in the central nervous system (CNS) in response to meningococcal infection. It also sheds light on the complexity of intracellular signaling after TLR triggering.

Brain-derived neurotrophic factor reduces inflammation and hippocampal apoptosis in experimental Streptococcus pneumoniae meningitis.

Science.gov (United States)

Xu, Danfeng; Lian, Di; Wu, Jing; Liu, Ying; Zhu, Mingjie; Sun, Jiaming; He, Dake; Li, Ling

2017-08-04

-inflammatory factor IL-10. Moreover, BDNF pretreatment increased TrkB expression, activated downstream phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling, and inhibited the myeloid differentiation primary response gene 88 (MyD88)/NF-κB-signaling pathway. These data suggested that BDNF administration exerted anti-inflammatory and anti-apoptotic effects on an experimental pneumococcal meningitis model via modulation of MyD88/NF-κB- and PI3K/AKT-signaling pathways. Our results indicated that treatment with exogenous BDNF might constitute a potential therapeutic strategy for the treatment of bacterial meningitis.

Research Article Polymorphism detection of promoter region of IFN ...

Indian Academy of Sciences (India)

MRT Pack 24 DVDs

Introduction. Improving animal health is considered to be the basic goal of the animal breeding industry ... agriculture sector for providing the invaluable supply of proteins, fats, minerals and vitamins as well as ..... 2011 MyD88 deficiency leads.

Scale-Dependent Habitat Selection and Size-Based Dominance in Adult Male American Alligators.

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Bradley A Strickland

Full Text Available Habitat selection is an active behavioral process that may vary across spatial and temporal scales. Animals choose an area of primary utilization (i.e., home range then make decisions focused on resource needs within patches. Dominance may affect the spatial distribution of conspecifics and concomitant habitat selection. Size-dependent social dominance hierarchies have been documented in captive alligators, but evidence is lacking from wild populations. We studied habitat selection for adult male American alligators (Alligator mississippiensis; n = 17 on the Pearl River in central Mississippi, USA, to test whether habitat selection was scale-dependent and individual resource selectivity was a function of conspecific body size. We used K-select analysis to quantify selection at the home range scale and patches within the home range to determine selection congruency and important habitat variables. In addition, we used linear models to determine if body size was related to selection patterns and strengths. Our results indicated habitat selection of adult male alligators was a scale-dependent process. Alligators demonstrated greater overall selection for habitat variables at the patch level and less at the home range level, suggesting resources may not be limited when selecting a home range for animals in our study area. Further, diurnal habitat selection patterns may depend on thermoregulatory needs. There was no relationship between resource selection or home range size and body size, suggesting size-dependent dominance hierarchies may not have influenced alligator resource selection or space use in our sample. Though apparent habitat suitability and low alligator density did not manifest in an observed dominance hierarchy, we hypothesize that a change in either could increase intraspecific interactions, facilitating a dominance hierarchy. Due to the broad and diverse ecological roles of alligators, understanding the factors that influence their

Scale-dependent habitat selection and size-based dominance in adult male American alligators

Science.gov (United States)

Strickland, Bradley A.; Vilella, Francisco; Belant, Jerrold L.

2016-01-01

Habitat selection is an active behavioral process that may vary across spatial and temporal scales. Animals choose an area of primary utilization (i.e., home range) then make decisions focused on resource needs within patches. Dominance may affect the spatial distribution of conspecifics and concomitant habitat selection. Size-dependent social dominance hierarchies have been documented in captive alligators, but evidence is lacking from wild populations. We studied habitat selection for adult male American alligators (Alligator mississippiensis; n = 17) on the Pearl River in central Mississippi, USA, to test whether habitat selection was scale-dependent and individual resource selectivity was a function of conspecific body size. We used K-select analysis to quantify selection at the home range scale and patches within the home range to determine selection congruency and important habitat variables. In addition, we used linear models to determine if body size was related to selection patterns and strengths. Our results indicated habitat selection of adult male alligators was a scale-dependent process. Alligators demonstrated greater overall selection for habitat variables at the patch level and less at the home range level, suggesting resources may not be limited when selecting a home range for animals in our study area. Further, diurnal habitat selection patterns may depend on thermoregulatory needs. There was no relationship between resource selection or home range size and body size, suggesting size-dependent dominance hierarchies may not have influenced alligator resource selection or space use in our sample. Though apparent habitat suitability and low alligator density did not manifest in an observed dominance hierarchy, we hypothesize that a change in either could increase intraspecific interactions, facilitating a dominance hierarchy. Due to the broad and diverse ecological roles of alligators, understanding the factors that influence their social dominance

TLR9 is required for MAPK/NF-κB activation but does not cooperate with TLR2 or TLR6 to induce host resistance to Brucella abortus.

Science.gov (United States)

Gomes, Marco Túlio; Campos, Priscila Carneiro; Pereira, Guilherme de Sousa; Bartholomeu, Daniella Castanheira; Splitter, Gary; Oliveira, Sergio Costa

2016-05-01

Brucella abortus is a Gram-negative intracellular bacterial pathogen that causes a zoonosis of worldwide occurrence, leading to undulant fever in humans and abortion in domestic animals. B. abortus is recognized by several pattern-recognition receptors triggering pathways during the host innate immune response. Therefore, here, we determined the cooperative role of TLR9 with TLR2 or TLR6 receptors in sensing Brucella Furthermore, we deciphered the host innate immune response against B. abortus or its DNA, emphasizing the role of TLR9-MAPK/NF-κB signaling pathways in the production of proinflammatory cytokines. TLR9 is required for the initial host control of B. abortus, but this TLR was dispensable after 6 wk of infection. The susceptibility of TLR9(-/-)-infected animals to Brucella paralleled with lower levels of IFN-γ produced by mouse splenocytes stimulated with this pathogen compared with wild-type cells. However, no apparent cooperative interplay was observed between TLR2-TLR9 or TLR6-TLR9 receptors to control infection. Moreover, B. abortus or its DNA induced activation of MAPK/NF-κB pathways and production of IL-12 and TNF-α by macrophages partially dependent on TLR9 but completely dependent on MyD88. In addition, B. abortus-derived CpG oligonucleotides required TLR9 to promote IL-12 and TNF-α production by macrophages. By confocal microscopy, we demonstrated that TLR9 redistributed and colocalized with lysosomal-associated membrane protein-1 upon Brucella infection. Thus, B. abortus induced TLR9 traffic, leading to cell signaling activation and IL-12 and TNF-α production. Although TLR9 recognized Brucella CpG motifs, our results suggest a new pathway of B. abortus DNA-activating macrophages independent of TLR9. © Society for Leukocyte Biology.

How can we model selectively neutral density dependence in evolutionary games.

Science.gov (United States)

Argasinski, Krzysztof; Kozłowski, Jan

2008-03-01

The problem of density dependence appears in all approaches to the modelling of population dynamics. It is pertinent to classic models (i.e., Lotka-Volterra's), and also population genetics and game theoretical models related to the replicator dynamics. There is no density dependence in the classic formulation of replicator dynamics, which means that population size may grow to infinity. Therefore the question arises: How is unlimited population growth suppressed in frequency-dependent models? Two categories of solutions can be found in the literature. In the first, replicator dynamics is independent of background fitness. In the second type of solution, a multiplicative suppression coefficient is used, as in a logistic equation. Both approaches have disadvantages. The first one is incompatible with the methods of life history theory and basic probabilistic intuitions. The logistic type of suppression of per capita growth rate stops trajectories of selection when population size reaches the maximal value (carrying capacity); hence this method does not satisfy selective neutrality. To overcome these difficulties, we must explicitly consider turn-over of individuals dependent on mortality rate. This new approach leads to two interesting predictions. First, the equilibrium value of population size is lower than carrying capacity and depends on the mortality rate. Second, although the phase portrait of selection trajectories is the same as in density-independent replicator dynamics, pace of selection slows down when population size approaches equilibrium, and then remains constant and dependent on the rate of turn-over of individuals.

Green Tea Extract Supplementation Induces the Lipolytic Pathway, Attenuates Obesity, and Reduces Low-Grade Inflammation in Mice Fed a High-Fat Diet

Directory of Open Access Journals (Sweden)

Cláudio A. Cunha

2013-01-01

Full Text Available The aim of this study was to evaluate the effects of green tea Camellia sinensis extract on proinflammatory molecules and lipolytic protein levels in adipose tissue of diet-induced obese mice. Animals were randomized into four groups: CW (chow diet and water; CG (chow diet and water + green tea extract; HW (high-fat diet and water; HG (high-fat diet and water + green tea extract. The mice were fed ad libitum with chow or high-fat diet and concomitantly supplemented (oral gavage with 400 mg/kg body weight/day of green tea extract (CG and HG, resp.. The treatments were performed for eight weeks. UPLC showed that in 10 mg/mL green tea extract, there were 15 μg/mg epigallocatechin, 95 μg/mg epigallocatechin gallate, 20.8 μg/mg epicatechin gallate, and 4.9 μg/mg gallocatechin gallate. Green tea administered concomitantly with a high-fat diet increased HSL, ABHD5, and perilipin in mesenteric adipose tissue, and this was associated with reduced body weight and adipose tissue gain. Further, we observed that green tea supplementation reduced inflammatory cytokine TNFα levels, as well as TLR4, MYD88, and TRAF6 proinflammatory signalling. Our results show that green tea increases the lipolytic pathway and reduces adipose tissue, and this may explain the attenuation of low-grade inflammation in obese mice.

Selection of Models for Ingestion Pathway and Relocation

International Nuclear Information System (INIS)

Blanchard, A.; Thompson, J.M.

1998-01-01

The area in which intermediate phase protective actions (such as food interdiction and relocation) may be needed following postulated accidents at three Savannah River Site nonreactor nuclear facilities will be determined by modeling. The criteria used to select dispersion/deposition models are presented. Several models are considered, including ARAC, MACCS, HOTSPOT, WINDS (coupled with PUFF-PLUME), and UFOTRI. Although ARAC and WINDS are expected to provide more accurate modeling of atmospheric transport following an actual release, analyses consistent with regulatory guidance for planning purposes may be accomplished with comparatively simple dispersion models such as HOTSPOT and UFOTRI. A recommendation is made to use HOTSPOT for non-tritium facilities and UFOTRI for tritium facilities. The most recent Food and Drug Administration Derived Intervention Levels (August 1998) are adopted as evaluation guidelines for ingestion pathways

Selection of Models for Ingestion Pathway and Relocation

International Nuclear Information System (INIS)

Blanchard, A.; Thompson, J.M.

1999-01-01

The area in which intermediate phase protective actions (such as food interdiction and relocation) may be needed following postulated accidents at three Savannah River Site nonreactor nuclear facilities will be determined by modeling. The criteria used to select dispersion/deposition models are presented. Several models are considered, including ARAC, MACCS, HOTSPOT, WINDS (coupled with PUFF-PLUME), and UFOTRI. Although ARAC and WINDS are expected to provide more accurate modeling of atmospheric transport following an actual release, analyses consistent with regulatory guidance for planning purposes may be accomplished with comparatively simple dispersion models such as HOTSPOT and UFOTRI. A recommendation is made to use HOTSPOT for non-tritium facilities and UFOTRI for tritium facilities. The most recent Food and Drug Administration Derived Intervention Levels (August 1998) are adopted as evaluation guidelines for ingestion pathways

METABOLIC ENGINEERING TO DEVELOP A PATHWAY FOR THE SELECTIVE CLEAVAGE OF CARBON-NITROGEN BONDS

Energy Technology Data Exchange (ETDEWEB)

John J. Kilbane III

2003-12-01

The objective of the project is to develop biochemical pathways for the selective cleavage of C-N bonds in molecules found in petroleum. The initial phase of the project will focus on the isolation or development of an enzyme capable of cleaving the C-N bond in aromatic amides, specifically 2-aminobiphenyl. The objective of the second phase of the research will be to construct a biochemical pathway for the selective removal of nitrogen from carbazole by combining the carA genes from Sphingomonas sp. GTIN11 with the gene(s) encoding an appropriate amidase. The objective of the final phase of the project will be to develop derivative CN bond cleaving enzymes that have broader substrate ranges and to demonstrate the use of such strains to selectively remove nitrogen from petroleum. The project is on schedule and no major difficulties have been encountered. During the first year of the project (October, 2002-September, 2003) enrichment culture experiments have resulted in the isolation of promising cultures that may be capable of cleaving C-N bonds in aromatic amides, several amidase genes have been cloned and are currently undergoing directed evolution to obtain derivatives that can cleave C-N bonds in aromatic amides, and the carA genes from Sphingomonas sp. GTIN11, and Pseudomonas resinovorans CA10 were cloned in vectors capable of replicating in Escherichia coli. Future research will address expression of these genes in Rhodococcus erythropolis. Enrichment culture experiments and directed evolution experiments continue to be a main focus of research activity and further work is required to obtain an appropriate amidase that will selectively cleave C-N bonds in aromatic substrates. Once an appropriate amidase gene is obtained it must be combined with genes encoding an enzyme capable of converting carbazole to 2'aminobiphenyl-2,3-diol: specifically carA genes. The carA genes from two sources have been cloned and are ready for construction of C-N bond cleavage

IL-33-induced alterations in murine intestinal function and cytokine responses are MyD88, STAT6, and IL-13-dependent

Science.gov (United States)

IL-33 is a recently identified cytokine member of the IL-1 family. The biological activities of IL-33 are associated with promotion of Th2 and inhibition of Th1/Th17 immune responses. Exogenous IL-33 induces a typical “type 2” immune response in the gastrointestinal tract, yet the underlying mechani...

Examining the evidence for major histocompatibility complex-dependent mate selection in humans and nonhuman primates

Czech Academy of Sciences Publication Activity Database

Winternitz, Jamie Caroline; Abbate, J. L.

2015-01-01

Roč. 6, 13 May (2015), s. 73-88 ISSN 1179-7274 R&D Projects: GA MŠk(CZ) EE2.3.30.0048 Institutional support: RVO:67985939 Keywords : secual selection * olfaction * facial attraction * inbreeding avoidance * parasite resistance Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Genetics and heredity (medical genetics to be 3)

9 CFR 381.88 - Parasites.

Science.gov (United States)

2010-01-01

... 9 Animals and Animal Products 2 2010-01-01 2010-01-01 false Parasites. 381.88 Section 381.88 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE AGENCY... § 381.88 Parasites. Organs or other parts of carcasses which are found to be infested with parasites, or...

21 CFR 808.88 - Pennsylvania.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Pennsylvania. 808.88 Section 808.88 Food and Drugs... and Local Exemptions § 808.88 Pennsylvania. (a) The following Pennsylvania medical device requirements... the condition that, in enforcing this requirement, Pennsylvania apply the definition of “used hearing...

34 CFR 668.88 - Hearing.

Science.gov (United States)

2010-07-01

... 34 Education 3 2010-07-01 2010-07-01 false Hearing. 668.88 Section 668.88 Education Regulations of... Proceedings § 668.88 Hearing. (a) A hearing is an orderly presentation of arguments and evidence conducted by a hearing official. (b) If the hearing official, the designated department official who brought a...

Alternative pathways of thromboplastin-dependent activation of human factor X in plasma

International Nuclear Information System (INIS)

Marlar, R.A.; Griffin, J.H.

1981-01-01

To determine the interrelationships of the major coagulation pathways, the activation of 3H-labeled factor X in normal and various deficient human plasmas was evaluated when clotting was triggered by dilute rabbit or human thromboplastin. Various dilutions of thromboplastin and calcium were added to plasma samples containing 3H-factor X, and the time course of factor X activation was determined. At a 1/250 dilution of rabbit brain thromboplastin, the rate of factor X activation in plasmas deficient in factor VIII or factor IX was 10% of the activation rate of normal plasma or of factor XI deficient plasma. Reconstitution of the deficient plasmas with factors VIII or IX, respectively, reconstituted normal factor X activation. Similar results were obtained when various dilutions of human thromboplastin replaced the rabbit thromboplastin. From these plasma experiments, it is inferred that the dilute thromboplastin-dependent activation of factor X requires factors VII, IX, and VIII. An alternative extrinsic pathway that involves factors IX and VIII may be the physiologic extrinsic pathway and hence help to explain the consistent clinical observations of bleeding diatheses in patients deficient in factors IX or VIII

Ubiquitin-coated nanodiamonds bind to autophagy receptors for entry into the selective autophagy pathway.

Science.gov (United States)

Liu, Kuang-Kai; Qiu, Wei-Ru; Naveen Raj, Emmanuel; Liu, Huei-Fang; Huang, Hou-Syun; Lin, Yu-Wei; Chang, Chien-Jen; Chen, Ting-Hua; Chen, Chinpiao; Chang, Huan-Cheng; Hwang, Jenn-Kang; Chao, Jui-I

2017-01-02

Selective macroautophagy/autophagy plays a pivotal role in the processing of foreign pathogens and cellular components to maintain homeostasis in human cells. To date, numerous studies have demonstrated the uptake of nanoparticles by cells, but their intracellular processing through selective autophagy remains unclear. Here we show that carbon-based nanodiamonds (NDs) coated with ubiquitin (Ub) bind to autophagy receptors (SQSTM1 [sequestosome 1], OPTN [optineurin], and CALCOCO2/NDP52 [calcium binding and coiled-coil domain 2]) and are then linked to MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) for entry into the selective autophagy pathway. NDs are ultimately delivered to lysosomes. Ectopically expressed SQSTM1-green fluorescence protein (GFP) could bind to the Ub-coated NDs. By contrast, the Ub-associated domain mutant of SQSTM1 (ΔUBA)-GFP did not bind to the Ub-coated NDs. Chloroquine, an autophagy inhibitor, prevented the ND-containing autophagosomes from fusing with lysosomes. Furthermore, autophagy receptors OPTN and CALCOCO2/NDP52, involved in the processing of bacteria, were found to be involved in the selective autophagy of NDs. However, ND particles located in the lysosomes of cells did not induce mitotic blockage, senescence, or cell death. Single ND clusters in the lysosomes of cells were observed in the xenografted human lung tumors of nude mice. This study demonstrated for the first time that Ub-coated nanoparticles bind to autophagy receptors for entry into the selective autophagy pathway, facilitating their delivery to lysosomes.

Solubility-Parameter-Guided Solvent Selection to Initiate Ostwald Ripening for Interior Space-Tunable Structures with Architecture-Dependent Electrochemical Performance.

Science.gov (United States)

Mao, Baoguang; Guo, Donglei; Qin, Jinwen; Meng, Tao; Wang, Xin; Cao, Minhua

2018-01-08

Despite significant advancement in preparing various hollow structures by Ostwald ripening, one common problem is the intractable uncontrollability of initiating Ostwald ripening due to the complexity of the reaction processes. Here, a new strategy on Hansen solubility parameter (HSP)-guided solvent selection to initiate Ostwald ripening is proposed. Based on this comprehensive principle for solvent optimization, N,N-dimethylformamide (DMF) was screened out, achieving accurate synthesis of interior space-tunable MoSe 2 spherical structures (solid, core-shell, yolk-shell and hollow spheres). The resultant MoSe 2 structures exhibit architecture-dependent electrochemical performances towards hydrogen evolution reaction and sodium-ion batteries. This pre-solvent selection strategy can effectively provide researchers great possibility in efficiently synthesizing various hollow structures. This work paves a new pathway for deeply understanding Ostwald ripening. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

TLR4-NOX4-AP-1 signaling mediates lipopolysaccharide-induced CXCR6 expression in human aortic smooth muscle cells

International Nuclear Information System (INIS)

Patel, Devang N.; Bailey, Steven R.; Gresham, John K.; Schuchman, David B.; Shelhamer, James H.; Goldstein, Barry J.; Foxwell, Brian M.; Stemerman, Michael B.; Maranchie, Jodi K.; Valente, Anthony J.; Mummidi, Srinivas; Chandrasekar, Bysani

2006-01-01

CXCL16 is a transmembrane non-ELR CXC chemokine that signals via CXCR6 to induce aortic smooth muscle cell (ASMC) proliferation. While bacterial lipopolysaccharide (LPS) has been shown to stimulate CXCL16 expression in SMC, its effects on CXCR6 are not known. Here, we demonstrate that LPS upregulates CXCR6 mRNA, protein, and surface expression in human ASMC. Inhibition of TLR4 with neutralizing antibodies or specific siRNA interference blocked LPS-mediated CXCR6 expression. LPS stimulated both AP-1 (c-Fos, c-Jun) and NF-κB (p50 and p65) activation, but only inhibition of AP-1 attenuated LPS-induced CXCR6 expression. Using dominant negative expression vectors and siRNA interference, we demonstrate that LPS induces AP-1 activation via MyD88, TRAF6, ERK1/2, and JNK signaling pathways. Furthermore, the flavoprotein inhibitor diphenyleniodonium chloride significantly attenuated LPS-mediated AP-1-dependent CXCR6 expression, as did inhibition of NOX4 NADPH oxidase by siRNA. Finally, CXCR6 knockdown inhibited CXCL16-induced ASMC proliferation. These results demonstrate that LPS-TLR4-NOX4-AP-1 signaling can induce CXCR6 expression in ASMC, and suggest that the CXCL16-CXCR6 axis may be an important proinflammatory pathway in the pathogenesis of atherosclerosis

Nature of the bifunctional chelating agent used for radioimmunotherapy with yttrium-88 monoclonal antibodies: critical factors in determining in vivo survival and organ toxicity

Energy Technology Data Exchange (ETDEWEB)

Kozak, R.W.; Raubitschek, A.; Mirzadeh, S.; Brechbiel, M.W.; Junghaus, R.; Gansow, O.A.; Waldmann, T.A. (Center for Biologics Evaluation and Research, FDA, Bethesda, MD (USA))

1989-05-15

One factor that is critical to the potential effectiveness of radioimmunotherapy is the design of radiometal-chelated antibodies that will be stable in vivo. Stability in vivo depends on the condition that both the chelate linkage and radiolabeling procedures not alter antibody specificity and biodistribution. In addition, synthesis and selection of the chelating agent is critical for each radiometal in order to prevent inappropriate release of the radiometal in vivo. In the present study, we compare the in vivo stability of seven radioimmunoconjugates that use different polyaminocarboxylate chelating agents to complex yttrium-88 to the mouse anti-human interleukin-2 receptor monoclonal antibody, anti-Tac. Chelate linkage and radiolabeling procedures did not alter the immunospecificity of anti-Tac. In order to assess whether yttrium was inappropriately released from the chelate-coupled antibody in vivo, iodine-131-labeled and yttrium-88 chelate-coupled antibodies were simultaneously administered to the same animals to correlate the decline in yttrium and radioiodinated antibody activity. The four stable yttrium-88 chelate-coupled antibodies studied displayed similar iodine-131 and yttrium-88 activity, indicating minimal elution of yttrium-88 from the complex. In contrast, the unstable yttrium-88 chelate-coupled antibodies had serum yttrium-88 activities that declined much more rapidly than their iodine-131 activities, suggesting loss of the radiolabel yttrium-88 from the chelate. Furthermore, high rates of yttrium-88 elution correlated with deposition in bone. Four chelating agents emerged as promising immunotherapeutic reagents: isothiocyanate benzyl DTPA and its derivatives 1B3M, MX, and 1M3B.

Berberine regulates neurite outgrowth through AMPK-dependent pathways by lowering energy status

Energy Technology Data Exchange (ETDEWEB)

Lu, Jiaqi; Cao, Yuanzhao; Cheng, Kuoyuan; Xu, Bo; Wang, Tianchang; Yang, Qi; Yang, Qin [State Key Laboratory of Natural and Biomimetic Drugs, Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing (China); Feng, Xudong, E-mail: xudong.feng@childrens.harvard.edu [Department of Medicine, Children' s Hospital Boston, Harvard Medical School, 300 Longwood Ave, Boston, MA 02115 (United States); Xia, Qing, E-mail: xqing@hsc.pku.edu.cn [State Key Laboratory of Natural and Biomimetic Drugs, Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing (China)

2015-06-10

As a widely used anti-bacterial agent and a metabolic inhibitor as well as AMP-activated protein kinase (AMPK) activator, berberine (BBR) has been shown to cross the blood–brain barrier. Its efficacy has been investigated in various disease models of the central nervous system. Neurite outgrowth is critical for nervous system development and is a highly energy-dependent process regulated by AMPK-related pathways. In the present study, we aimed to investigate the effects of BBR on AMPK activation and neurite outgrowth in neurons. The neurite outgrowth of primary rat cortical neurons at different stages of polarization was monitored after exposure of BBR. Intracellular energy level, AMPK activation and polarity-related pathways were also inspected. The results showed that BBR suppressed neurite outgrowth and affected cytoskeleton stability in the early stages of neuronal polarization, which was mediated by lowered energy status and AMPK activation. Liver kinase B1 and PI3K–Akt–GSK3β signaling pathways were also involved. In addition, mitochondrial dysfunction and endoplasmic reticulum stress contributed to the lowered energy status induced by BBR. This study highlighted the knowledge of the complex activities of BBR in neurons and corroborated the significance of energy status during the neuronal polarization. - Highlights: • BBR inhibited neurite outgrowth in early stages of neuronal development. • Lowered neuronal energy status was induced by BBR treatment. • Neuronal energy stress induced by BBR activated AMPK-related pathways. • BBR induced mitochondrial dysfunction and endoplasmic reticulum stress.

Berberine regulates neurite outgrowth through AMPK-dependent pathways by lowering energy status

International Nuclear Information System (INIS)

Lu, Jiaqi; Cao, Yuanzhao; Cheng, Kuoyuan; Xu, Bo; Wang, Tianchang; Yang, Qi; Yang, Qin; Feng, Xudong; Xia, Qing

2015-01-01

As a widely used anti-bacterial agent and a metabolic inhibitor as well as AMP-activated protein kinase (AMPK) activator, berberine (BBR) has been shown to cross the blood–brain barrier. Its efficacy has been investigated in various disease models of the central nervous system. Neurite outgrowth is critical for nervous system development and is a highly energy-dependent process regulated by AMPK-related pathways. In the present study, we aimed to investigate the effects of BBR on AMPK activation and neurite outgrowth in neurons. The neurite outgrowth of primary rat cortical neurons at different stages of polarization was monitored after exposure of BBR. Intracellular energy level, AMPK activation and polarity-related pathways were also inspected. The results showed that BBR suppressed neurite outgrowth and affected cytoskeleton stability in the early stages of neuronal polarization, which was mediated by lowered energy status and AMPK activation. Liver kinase B1 and PI3K–Akt–GSK3β signaling pathways were also involved. In addition, mitochondrial dysfunction and endoplasmic reticulum stress contributed to the lowered energy status induced by BBR. This study highlighted the knowledge of the complex activities of BBR in neurons and corroborated the significance of energy status during the neuronal polarization. - Highlights: • BBR inhibited neurite outgrowth in early stages of neuronal development. • Lowered neuronal energy status was induced by BBR treatment. • Neuronal energy stress induced by BBR activated AMPK-related pathways. • BBR induced mitochondrial dysfunction and endoplasmic reticulum stress

METABOLIC ENGINEERING TO DEVELOP A PATHWAY FOR THE SELECTIVE CLEAVAGE OF CARBON-NITROGEN BONDS

Energy Technology Data Exchange (ETDEWEB)

John J. Kilbane II

2004-10-01

The objective of the project is to develop biochemical pathways for the selective cleavage of C-N bonds in molecules found in petroleum. The initial phase of the project was focused on the isolation or development of an enzyme capable of cleaving the C-N bond in aromatic amides, specifically 2-aminobiphenyl. The objective of the second phase of the research will be to construct a biochemical pathway for the selective removal of nitrogen from carbazole by combining the carA genes from Sphingomonas sp. GTIN11 with the gene(s) encoding an appropriate deaminase. The objective of the final phase of the project will be to develop derivative C-N bond cleaving enzymes that have broader substrate ranges and to demonstrate the use of such strains to selectively remove nitrogen from petroleum. During the first year of the project (October, 2002-September, 2003) enrichment culture experiments resulted in the isolation of microbial cultures that utilize aromatic amides as sole nitrogen sources, several amidase genes were cloned and were included in directed evolution experiments to obtain derivatives that can cleave C-N bonds in aromatic amides, and the carA genes from Sphingomonas sp. GTIN11, and Pseudomonas resinovorans CA10 were cloned in vectors capable of replicating in Escherichia coli. During the second year of the project (October, 2003-September, 2004) enrichment culture experiments succeeded in isolating a mixed bacterial culture that can utilize 2-aminobiphenyl as a sole nitrogen source, directed evolution experiments were focused on the aniline dioxygenase enzyme that is capable of deaminating aniline, and expression vectors were constructed to enable the expression of genes encoding C-N bond cleaving enzymes in Rhodococcus hosts. The construction of a new metabolic pathway to selectively remove nitrogen from carbazole and other molecules typically found in petroleum should lead to the development of a process to improve oil refinery efficiency by reducing the

Optimal processing pathway selection for microalgae-based biorefinery under uncertainty

DEFF Research Database (Denmark)

Rizwan, Muhammad; Zaman, Muhammad; Lee, Jay H.

2015-01-01

We propose a systematic framework for the selection of optimal processing pathways for a microalgaebased biorefinery under techno-economic uncertainty. The proposed framework promotes robust decision making by taking into account the uncertainties that arise due to inconsistencies among...... and shortage in the available technical information. A stochastic mixed integer nonlinear programming (sMINLP) problem is formulated for determining the optimal biorefinery configurations based on a superstructure model where parameter uncertainties are modeled and included as sampled scenarios. The solution...... the accounting of uncertainty are compared with respect to different objectives. (C) 2015 Elsevier Ltd. All rights reserved....

The TIR-domain containing adaptor TRAM is required for TLR7 mediated RANTES production.

Directory of Open Access Journals (Sweden)

Enda Shevlin

Full Text Available Toll-like receptor 7 (TLR7 plays a vital role in the immune response to ssRNA viruses such as human rhinovirus (HRV and Influenza, against which there are currently no treatments or vaccines with long term efficacy available. Clearly, a more comprehensive understanding of the TLR7 signaling axis will contribute to its molecular targeting. TRIF related adaptor molecule (TRAM plays a vital role in TLR4 signaling by recruiting TRIF to TLR4, followed by endosomal trafficking of the complex and initiation of IRF3 dependent type I interferon production as well as NF-κB dependent pro-inflammatory cytokine production. Towards understanding the molecular mechanisms that regulate TLR7 functionality, we found that TRAM(-/- murine macrophages exhibited a transcriptional and translational impairment in TLR7 mediated RANTES, but not TNFα, production. Suppression of TRAM expression in human macrophages also resulted in an impairment in TLR7 mediated CCL5 and IFN-β, but not TNFα, gene induction. Furthermore, suppression of endogenous human TRAM expression in human macrophages significantly impaired RV16 induced CCL5 and IFNβ, but not TNFα gene induction. Additionally, TRAM-G2A dose-dependently inhibited TLR7 mediated activation of CCL5, IFNβ and IFNα reporter genes. TLR7-mediated phosphorylation and nuclear translocation of IRF3 was impaired in TRAM(-/- cells. Finally, co-immunoprecipitation studies indicated that TRAM physically interacts with MyD88 upon TLR7 stimulation, but not under basal conditions. Our results clearly demonstrate that TRAM plays a, hitherto unappreciated, role in TLR7 signaling through a novel signaling axis containing, but not limited to, MyD88, TRAM and IRF3 towards the activation of anti-viral immunity.

Ofloxacin induces apoptosis in microencapsulated juvenile rabbit chondrocytes by caspase-8-dependent mitochondrial pathway

International Nuclear Information System (INIS)

Sheng Zhiguo; Cao Xiaojuan; Peng Shuangqing; Wang Changyong; Li Qianqian; Wang Yimei; Liu Mifeng

2008-01-01

Quinolones (QNs)-induced arthropathy is an important toxic effect in immature animals leading to restriction of their therapeutic use in pediatrics. However, the exact mechanism still remains unclear. Recently, we have demonstrated that ofloxacin, a typical QN, induces apoptosis of alginate microencapsulated juvenile rabbit joint chondrocytes by disturbing the β 1 integrin functions and inactivating the ERK/MAPK signaling pathway. In this study, we extend our initial observations to further elucidate the mechanism(s) of ofloxacin-induced apoptosis by utilizing specific caspase inhibitors. Pretreatment with both caspase-9-specific inhibitor zLEHD-fmk and caspase-8 inhibitor zIETD-fmk attenuated ofloxacin-induced apoptosis and activation of caspase-3 of chondrocyte in a concentration-dependent manner, as determined by fluorescent dye staining, enzyme activity assay and immunoblotting. Furthermore, the activation of caspase-9, -8 and -3 stimulated by ofloxacin was significantly inhibited in the presence of zIETD-fmk while pretreatment with zLEHD-fmk only blocked the activation of caspase-9 and -3. Ofloxacin also stimulated a concentration-dependent translocation of cytochrome c from mitochondria into the cytosol and a decrease of mitochondrial transmembrane potential, which was completely inhibited by zIETD-fmk. In addition, ofloxacin was found to increase the level of Bax, tBid, p53 in a concentration- and time-dependent manner. Taken together, The current results indicate that the caspase-8-dependent mitochondrial pathway is primarily involved in the ofloxacin-induced apoptosis of microencapsulated juvenile rabbit joint chondrocytes

C5a Receptor (CD88) Blockade Protects against MPO-ANCA GN

OpenAIRE

Xiao, Hong; Dairaghi, Daniel J.; Powers, Jay P.; Ertl, Linda S.; Baumgart, Trageen; Wang, Yu; Seitz, Lisa C.; Penfold, Mark E.T.; Gan, Lin; Hu, Peiqi; Lu, Bao; Gerard, Norma P.; Gerard, Craig; Schall, Thomas J.; Jaen, Juan C.

2013-01-01

Necrotizing and crescentic GN (NCGN) with a paucity of glomerular immunoglobulin deposits is associated with ANCA. The most common ANCA target antigens are myeloperoxidase (MPO) and proteinase 3. In a manner that requires activation of the alternative complement pathway, passive transfer of antibodies to mouse MPO (anti-MPO) induces a mouse model of ANCA NCGN that closely mimics human disease. Here, we confirm the importance of C5aR/CD88 in the mediation of anti-MPO–induced NCGN and report th...

Increased toll-like receptors and p53 levels regulate apoptosis and angiogenesis in non-muscle invasive bladder cancer: mechanism of action of P-MAPA biological response modifier.

Science.gov (United States)

Garcia, Patrick Vianna; Seiva, Fábio Rodrigues Ferreira; Carniato, Amanda Pocol; de Mello Júnior, Wilson; Duran, Nelson; Macedo, Alda Maria; de Oliveira, Alexandre Gabarra; Romih, Rok; Nunes, Iseu da Silva; Nunes, Odilon da Silva; Fávaro, Wagner José

2016-07-07

The new modalities for treating patients with non-muscle invasive bladder cancer (NMIBC) for whom BCG (Bacillus Calmette-Guerin) has failed or is contraindicated are recently increasing due to the development of new drugs. Although agents like mitomycin C and BCG are routinely used, there is a need for more potent and/or less-toxic agents. In this scenario, a new perspective is represented by P-MAPA (Protein Aggregate Magnesium-Ammonium Phospholinoleate-Palmitoleate Anhydride), developed by Farmabrasilis (non-profit research network). This study detailed and characterized the mechanisms of action of P-MAPA based on activation of mediators of Toll-like Receptors (TLRs) 2 and 4 signaling pathways and p53 in regulating angiogenesis and apoptosis in an animal model of NMIBC, as well as, compared these mechanisms with BCG treatment. Our results demonstrated the activation of the immune system by BCG (MyD88-dependent pathway) resulted in increased inflammatory cytokines. However, P-MAPA intravesical immunotherapy led to distinct activation of TLRs 2 and 4-mediated innate immune system, resulting in increased interferons signaling pathway (TRIF-dependent pathway), which was more effective in the NMIBC treatment. Interferon signaling pathway activation induced by P-MAPA led to increase of iNOS protein levels, resulting in apoptosis and histopathological recovery. Additionally, P-MAPA immunotherapy increased wild-type p53 protein levels. The increased wild-type p53 protein levels were fundamental to NO-induced apoptosis and the up-regulation of BAX. Furthermore, interferon signaling pathway induction and increased p53 protein levels by P-MAPA led to important antitumor effects, not only suppressing abnormal cell proliferation, but also by preventing continuous expansion of tumor mass through suppression of angiogenesis, which was characterized by decreased VEGF and increased endostatin protein levels. Thus, P-MAPA immunotherapy could be considered an important therapeutic

Potent Anti-Inflammatory Activity of Pyrenocine A Isolated from the Marine-Derived Fungus Penicillium paxilli Ma(G)K

Science.gov (United States)

Toledo, Thaís Regina; Dejani, Naiara N.; Monnazzi, Luis Gustavo Silva; Kossuga, Miriam H.; Berlinck, Roberto G. S.; Sette, Lara D.; Medeiros, Alexandra I.

2014-01-01

Very little is known about the immunomodulatory potential of secondary metabolites isolated from marine microorganisms. In the present study, we characterized pyrenocine A, which is produced by the marine-derived fungus Penicillium paxilli Ma(G)K and possesses anti-inflammatory activity. Pyrenocine A was able to suppress, both pretreatment and posttreatment, the LPS-induced activation of macrophages via the inhibition of nitrite production and the synthesis of inflammatory cytokines and PGE2. Pyrenocine A also exhibited anti-inflammatory effects on the expression of receptors directly related to cell migration (Mac-1) as well as costimulatory molecules involved in lymphocyte activation (B7.1). Nitrite production was inhibited by pyrenocine A in macrophages stimulated with CpG but not Poly I:C, suggesting that pyrenocine A acts through the MyD88-dependent intracellular signaling pathway. Moreover, pyrenocine A is also able to inhibit the expression of genes related to NFκB-mediated signal transduction on macrophages stimulated by LPS. Our results indicate that pyrenocine A has promissory anti-inflammatory properties and additional experiments are necessary to confirm this finding in vivo model. PMID:24574582

Innate immune response of human plasmacytoid dendritic cells to poxvirus infection is subverted by vaccinia E3 via its Z-DNA/RNA binding domain.

Directory of Open Access Journals (Sweden)

Hua Cao

Full Text Available Plasmacytoid dendritic cells (pDCs play important roles in antiviral innate immunity by producing type I interferon (IFN. In this study, we assess the immune responses of primary human pDCs to two poxviruses, vaccinia and myxoma virus. Vaccinia, an orthopoxvirus, was used for immunization against smallpox, a contagious human disease with high mortality. Myxoma virus, a Leporipoxvirus, causes lethal disease in rabbits, but is non-pathogenic in humans. We report that myxoma virus infection of human pDCs induces IFN-α and TNF production, whereas vaccinia infection does not. Co-infection of pDCs with myxoma virus plus vaccinia blocks myxoma induction effects. We find that heat-inactivated vaccinia (Heat-VAC; by incubating the virus at 55°C for 1 h gains the ability to induce IFN-α and TNF in primary human pDCs. Induction of IFN-α in pDCs by myxoma virus or Heat-VAC is blocked by chloroquine, which inhibits endosomal acidification required for TLR7/9 signaling, and by inhibitors of cellular kinases PI3K and Akt. Using purified pDCs from genetic knockout mice, we demonstrate that Heat-VAC-induced type I IFN production in pDCs requires the endosomal RNA sensor TLR7 and its adaptor MyD88, transcription factor IRF7 and the type I IFN feedback loop mediated by IFNAR1. These results indicate that (i vaccinia virus, but not myxoma virus, expresses inhibitor(s of the poxvirus sensing pathway(s in pDCs; and (ii Heat-VAC infection fails to produce inhibitor(s but rather produces novel activator(s, likely viral RNA transcripts that are sensed by the TLR7/MyD88 pathway. Using vaccinia gene deletion mutants, we show that the Z-DNA/RNA binding domain at the N-terminus of the vaccinia immunomodulatory E3 protein is an antagonist of the innate immune response of human pDCs to poxvirus infection and TLR agonists. The myxoma virus ortholog of vaccinia E3 (M029 lacks the N-terminal Z-DNA/RNA binding domain, which might contribute to the immunostimulating

Innate Immune Response of Human Plasmacytoid Dendritic Cells to Poxvirus Infection Is Subverted by Vaccinia E3 via Its Z-DNA/RNA Binding Domain

Science.gov (United States)

Dai, Peihong; Wang, Weiyi; Li, Hao; Yuan, Jianda; Wang, Fangjin; Fang, Chee-Mun; Pitha, Paula M; Liu, Jia; Condit, Richard C; McFadden, Grant; Merghoub, Taha; Houghton, Alan N; Young, James W; Shuman, Stewart; Deng, Liang

2012-01-01

Plasmacytoid dendritic cells (pDCs) play important roles in antiviral innate immunity by producing type I interferon (IFN). In this study, we assess the immune responses of primary human pDCs to two poxviruses, vaccinia and myxoma virus. Vaccinia, an orthopoxvirus, was used for immunization against smallpox, a contagious human disease with high mortality. Myxoma virus, a Leporipoxvirus, causes lethal disease in rabbits, but is non-pathogenic in humans. We report that myxoma virus infection of human pDCs induces IFN-α and TNF production, whereas vaccinia infection does not. Co-infection of pDCs with myxoma virus plus vaccinia blocks myxoma induction effects. We find that heat-inactivated vaccinia (Heat-VAC; by incubating the virus at 55°C for 1 h) gains the ability to induce IFN-α and TNF in primary human pDCs. Induction of IFN-α in pDCs by myxoma virus or Heat-VAC is blocked by chloroquine, which inhibits endosomal acidification required for TLR7/9 signaling, and by inhibitors of cellular kinases PI3K and Akt. Using purified pDCs from genetic knockout mice, we demonstrate that Heat-VAC-induced type I IFN production in pDCs requires the endosomal RNA sensor TLR7 and its adaptor MyD88, transcription factor IRF7 and the type I IFN feedback loop mediated by IFNAR1. These results indicate that (i) vaccinia virus, but not myxoma virus, expresses inhibitor(s) of the poxvirus sensing pathway(s) in pDCs; and (ii) Heat-VAC infection fails to produce inhibitor(s) but rather produces novel activator(s), likely viral RNA transcripts that are sensed by the TLR7/MyD88 pathway. Using vaccinia gene deletion mutants, we show that the Z-DNA/RNA binding domain at the N-terminus of the vaccinia immunomodulatory E3 protein is an antagonist of the innate immune response of human pDCs to poxvirus infection and TLR agonists. The myxoma virus ortholog of vaccinia E3 (M029) lacks the N-terminal Z-DNA/RNA binding domain, which might contribute to the immunostimulating properties of

CSF Proteomics Identifies Specific and Shared Pathways for Multiple Sclerosis Clinical Subtypes.

Directory of Open Access Journals (Sweden)

Timucin Avsar

Full Text Available Multiple sclerosis (MS is an immune-mediated, neuro-inflammatory, demyelinating and neurodegenerative disease of the central nervous system (CNS with a heterogeneous clinical presentation and course. There is a remarkable phenotypic heterogeneity in MS, and the molecular mechanisms underlying it remain unknown. We aimed to investigate further the etiopathogenesis related molecular pathways in subclinical types of MS using proteomic and bioinformatics approaches in cerebrospinal fluids of patients with clinically isolated syndrome, relapsing remitting MS and progressive MS (n=179. Comparison of disease groups with controls revealed a total of 151 proteins that are differentially expressed in clinically different MS subtypes. KEGG analysis using PANOGA tool revealed the disease related pathways including aldosterone-regulated sodium reabsorption (p=8.02x10-5 which is important in the immune cell migration, renin-angiotensin (p=6.88x10-5 system that induces Th17 dependent immunity, notch signaling (p=1.83x10-10 pathway indicating the activated remyelination and vitamin digestion and absorption pathways (p=1.73x10-5. An emerging theme from our studies is that whilst all MS clinical forms share common biological pathways, there are also clinical subtypes specific and pathophysiology related pathways which may have further therapeutic implications.

Selective mutism: a consensus based care pathway of good practice.

Science.gov (United States)

Keen, D V; Fonseca, S; Wintgens, A

2008-10-01

Selective mutism (SM) now acknowledged as an anxiety condition, tends to be a poorly understood, highly complex and vastly under-recognised clinical entity. Children with SM are a vulnerable group as the condition is not the remit of any one professional group. This inevitably leads to delay in formal diagnosis and management. There is a lack of systematic research on which to base guidelines for management. To develop, agree and validate key principles underlying the management of SM through a consensus process involving international experts, in order to create a local care pathway. A local multi-agency consultation process developed 11 statements, which were felt to be the key principles underpinning a potential care pathway for managing SM. Thirteen recognised experts from North America, Europe and Australia participated in a modified Delphi process involving two rounds using a Likert-scale and free commentary. Both quantitative and qualitative analyses were used in the validation or revision of the statements at each stage. Response rates were 100% for Round 1 and 84.6% for Round 2. Despite the differing professional backgrounds and service contexts, by successive revision and/or revalidation of statements, it was possible to arrive at a consensus about key principles relating to early recognition, assessment and intervention. The agreed key principles are presented together with the resulting local care pathway. Through a Delphi process, agreement was reached by a multidisciplinary group of professionals, on key principles that underpin the timely identification, assessment and management of children with SM. These include the potential for staff in school/preschool settings to identify SM and that intervention programmes should generally be based in these settings. Children with SM should receive assessment for possible coexisting disorders, whether developmental, emotional or behavioural and additional specific intervention given for these. Agreement was

Erythrocyte signal transduction pathways, their oxygenation dependence and functional significance.

Science.gov (United States)

Barvitenko, Nadezhda N; Adragna, Norma C; Weber, Roy E

2005-01-01

Erythrocytes play a key role in human and vertebrate metabolism. Tissue O2 supply is regulated by both hemoglobin (Hb)-O2 affinity and erythrocyte rheology, a key determinant of tissue perfusion. Oxygenation-deoxygenation transitions of Hb may lead to re-organization of the cytoskeleton and signalling pathways activation/deactivation in an O2-dependent manner. Deoxygenated Hb binds to the cytoplasmic domain of the anion exchanger band 3, which is anchored to the cytoskeleton, and is considered a major mechanism underlying the oxygenation-dependence of several erythrocyte functions. This work discusses the multiple modes of Hb-cytoskeleton interactions. In addition, it reviews the effects of Mg2+, 2,3-diphosphoglycerate, NO, shear stress and Ca2+, all factors accompanying the oxygenation-deoxygenation cycle in circulating red cells. Due to the extensive literature on the subject, the data discussed here, pertain mainly to human erythrocytes whose O2 affinity is modulated by 2,3-diphosphoglycerate, ectothermic vertebrate erythrocytes that use ATP, and to bird erythrocytes that use inositol pentaphosphate. Copyright 2005 S. Karger AG, Basel.

4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to inhibit hepatocellular carcinoma cells

International Nuclear Information System (INIS)

Fu, Meili; Wan, Fuqiang; Li, Zhengling; Zhang, Fenghua

2016-01-01

The aim of the present study is to investigate the potential anti-hepatocellular carcinoma (HCC) cell activity by 4SC-202, a novel class I HDAC inhibitor (HDACi). The associated signaling mechanisms were also analyzed. We showed that 4SC-202 treatment induced potent cytotoxic and proliferation–inhibitory activities against established HCC cell lines (HepG2, HepB3, SMMC-7721) and patient-derived primary HCC cells. Further, adding 4SC-202 in HCC cells activated mitochondrial apoptosis pathway, which was evidenced by mitochondrial permeability transition pore (mPTP) opening, cytochrome C cytosol release and caspase-3/-9 activation. Inhibition of this apoptosis pathway, by caspase-3/-9 inhibitors, mPTP blockers, or by shRNA-mediated knockdown of cyclophilin-D (Cyp-D, a key component of mPTP), significantly attenuated 4SC-202-induced HCC cell death and apoptosis. Reversely, over-expression of Cyp-D enhanced 4SC-202's sensitivity in HCC cells. Further studies showed that 4SC-202 induced apoptosis signal-regulating kinase 1 (ASK1) activation, causing it translocation to mitochondria and physical association with Cyp-D. This mitochondrial ASK1-Cyp-D complexation appeared required for mediating 4SC-202-induced apoptosis activation. ASK1 stable knockdown by targeted-shRNAs largely inhibited 4SC-202-induced mPTP opening, cytochrome C release, and following HCC cell apoptotic death. Together, we suggest that 4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to potently inhibit human HCC cells. - Highlights: • 4SC-202 exerts potent anti-proliferative and cytotoxic activity against established/primary HCC cells. • SC-202-induced anti-HCC cell activity relies on caspase-dependent apoptosis activation. • 4SC-202 activates Cyp-D-dependent mitochondrial apoptosis pathway in HCC cells. • 4SC-202 activates ASK1 in HCC cells, causing it translocation to mitochondria. • Mitochondrial ASK1-Cyp-D complexation mediates 4SC-202's activity in HCC cells.

4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to inhibit hepatocellular carcinoma cells

Energy Technology Data Exchange (ETDEWEB)

Fu, Meili, E-mail: fumeilidrlinyi@tom.com [Department of Infectious Disease, Linyi People' s Hospital, Linyi 276000 (China); Wan, Fuqiang [Department of Head and Neck Surgery, Linyi Tumor Hospital, Linyi 276000 (China); Li, Zhengling [Department of Nursing, Tengzhou Central People' s Hospital, Tengzhou 277500 (China); Zhang, Fenghua [Department of Operating Room, Linyi People' s Hospital, Linyi 276000 (China)

2016-03-04

The aim of the present study is to investigate the potential anti-hepatocellular carcinoma (HCC) cell activity by 4SC-202, a novel class I HDAC inhibitor (HDACi). The associated signaling mechanisms were also analyzed. We showed that 4SC-202 treatment induced potent cytotoxic and proliferation–inhibitory activities against established HCC cell lines (HepG2, HepB3, SMMC-7721) and patient-derived primary HCC cells. Further, adding 4SC-202 in HCC cells activated mitochondrial apoptosis pathway, which was evidenced by mitochondrial permeability transition pore (mPTP) opening, cytochrome C cytosol release and caspase-3/-9 activation. Inhibition of this apoptosis pathway, by caspase-3/-9 inhibitors, mPTP blockers, or by shRNA-mediated knockdown of cyclophilin-D (Cyp-D, a key component of mPTP), significantly attenuated 4SC-202-induced HCC cell death and apoptosis. Reversely, over-expression of Cyp-D enhanced 4SC-202's sensitivity in HCC cells. Further studies showed that 4SC-202 induced apoptosis signal-regulating kinase 1 (ASK1) activation, causing it translocation to mitochondria and physical association with Cyp-D. This mitochondrial ASK1-Cyp-D complexation appeared required for mediating 4SC-202-induced apoptosis activation. ASK1 stable knockdown by targeted-shRNAs largely inhibited 4SC-202-induced mPTP opening, cytochrome C release, and following HCC cell apoptotic death. Together, we suggest that 4SC-202 activates ASK1-dependent mitochondrial apoptosis pathway to potently inhibit human HCC cells. - Highlights: • 4SC-202 exerts potent anti-proliferative and cytotoxic activity against established/primary HCC cells. • SC-202-induced anti-HCC cell activity relies on caspase-dependent apoptosis activation. • 4SC-202 activates Cyp-D-dependent mitochondrial apoptosis pathway in HCC cells. • 4SC-202 activates ASK1 in HCC cells, causing it translocation to mitochondria. • Mitochondrial ASK1-Cyp-D complexation mediates 4SC-202's activity in HCC cells.

Characterization of cellular immune response and innate immune signaling in human and nonhuman primate primary mononuclear cells exposed to Burkholderia mallei.

Science.gov (United States)

Alam, Shahabuddin; Amemiya, Kei; Bernhards, Robert C; Ulrich, Robert G; Waag, David M; Saikh, Kamal U

2015-01-01

Burkholderia pseudomallei infection causes melioidosis and is often characterized by severe sepsis. Although rare in humans, Burkholderia mallei has caused infections in laboratory workers, and the early innate cellular response to B. mallei in human and nonhuman primates has not been characterized. In this study, we examined the primary cellular immune response to B. mallei in PBMC cultures of non-human primates (NHPs), Chlorocebus aethiops (African Green Monkeys), Macaca fascicularis (Cynomolgus macaque), and Macaca mulatta (Rhesus macaque) and humans. Our results demonstrated that B. mallei elicited strong primary pro-inflammatory cytokines (IFN-γ, TNF-α, IL-1β, and IL-6) equivalent to the levels of B. pseudomallei in primary PBMC cultures of NHPs and humans. When we examined IL-1β and other cytokine responses by comparison to Escherichia coli LPS, African Green Monkeys appears to be most responsive to B. mallei than Cynomolgus or Rhesus. Characterization of the immune signaling mechanism for cellular response was conducted by using a ligand induced cell-based reporter assay, and our results demonstrated that MyD88 mediated signaling contributed to the B. mallei and B. pseudomallei induced pro-inflammatory responses. Notably, the induced reporter activity with B. mallei, B. pseudomallei, or purified LPS from these pathogens was inhibited and cytokine production was attenuated by a MyD88 inhibitor. Together, these results show that in the scenario of severe hyper-inflammatory responses to B. mallei infection, MyD88 targeted therapeutic intervention may be a successful strategy for therapy. Published by Elsevier Ltd.

MAPK pathway activation by chronic lead-exposure increases vascular reactivity through oxidative stress/cyclooxygenase-2-dependent pathways

Energy Technology Data Exchange (ETDEWEB)

Simões, Maylla Ronacher, E-mail: yllars@hotmail.com [Dept. of Physiological Sciences, Federal University of Espirito Santo, Vitória, ES CEP 29040-091 (Brazil); Department of Pharmacology, Universidad Autonoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), Madrid (Spain); Aguado, Andrea [Department of Pharmacology, Universidad Autonoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), Madrid (Spain); Fiorim, Jonaína; Silveira, Edna Aparecida; Azevedo, Bruna Fernandes; Toscano, Cindy Medice [Dept. of Physiological Sciences, Federal University of Espirito Santo, Vitória, ES CEP 29040-091 (Brazil); Zhenyukh, Olha; Briones, Ana María [Department of Pharmacology, Universidad Autonoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), Madrid (Spain); Alonso, María Jesús [Dept. of Biochemistry, Physiology and Molecular Genetics, Universidad Rey Juan Carlos, Alcorcón (Spain); Vassallo, Dalton Valentim [Dept. of Physiological Sciences, Federal University of Espirito Santo, Vitória, ES CEP 29040-091 (Brazil); Health Science Center of Vitória-EMESCAM, Vitória, ES CEP 29045-402 (Brazil); Salaices, Mercedes, E-mail: mercedes.salaices@uam.es [Department of Pharmacology, Universidad Autonoma de Madrid, Instituto de Investigación Hospital Universitario La Paz (IdiPAZ), Madrid (Spain)

2015-03-01

Chronic exposure to low lead concentration produces hypertension; however, the underlying mechanisms remain unclear. We analyzed the role of oxidative stress, cyclooxygenase-2-dependent pathways and MAPK in the vascular alterations induced by chronic lead exposure. Aortas from lead-treated Wistar rats (1st dose: 10 μg/100 g; subsequent doses: 0.125 μg/100 g, intramuscular, 30 days) and cultured aortic vascular smooth muscle cells (VSMCs) from Sprague Dawley rats stimulated with lead (20 μg/dL) were used. Lead blood levels of treated rats attained 21.7 ± 2.38 μg/dL. Lead exposure increased systolic blood pressure and aortic ring contractile response to phenylephrine, reduced acetylcholine-induced relaxation and did not affect sodium nitroprusside relaxation. Endothelium removal and L-NAME left-shifted the response to phenylephrine more in untreated than in lead-treated rats. Apocynin and indomethacin decreased more the response to phenylephrine in treated than in untreated rats. Aortic protein expression of gp91(phox), Cu/Zn-SOD, Mn-SOD and COX-2 increased after lead exposure. In cultured VSMCs lead 1) increased superoxide anion production, NADPH oxidase activity and gene and/or protein levels of NOX-1, NOX-4, Mn-SOD, EC-SOD and COX-2 and 2) activated ERK1/2 and p38 MAPK. Both antioxidants and COX-2 inhibitors normalized superoxide anion production, NADPH oxidase activity and mRNA levels of NOX-1, NOX-4 and COX-2. Blockade of the ERK1/2 and p38 signaling pathways abolished lead-induced NOX-1, NOX-4 and COX-2 expression. Results show that lead activation of the MAPK signaling pathways activates inflammatory proteins such as NADPH oxidase and COX-2, suggesting a reciprocal interplay and contribution to vascular dysfunction as an underlying mechanisms for lead-induced hypertension. - Highlights: • Lead-exposure increases oxidative stress, COX-2 expression and vascular reactivity. • Lead exposure activates MAPK signaling pathway. • ROS and COX-2 activation by

Probabilistic pathway construction.

Science.gov (United States)

Yousofshahi, Mona; Lee, Kyongbum; Hassoun, Soha

2011-07-01

Expression of novel synthesis pathways in host organisms amenable to genetic manipulations has emerged as an attractive metabolic engineering strategy to overproduce natural products, biofuels, biopolymers and other commercially useful metabolites. We present a pathway construction algorithm for identifying viable synthesis pathways compatible with balanced cell growth. Rather than exhaustive exploration, we investigate probabilistic selection of reactions to construct the pathways. Three different selection schemes are investigated for the selection of reactions: high metabolite connectivity, low connectivity and uniformly random. For all case studies, which involved a diverse set of target metabolites, the uniformly random selection scheme resulted in the highest average maximum yield. When compared to an exhaustive search enumerating all possible reaction routes, our probabilistic algorithm returned nearly identical distributions of yields, while requiring far less computing time (minutes vs. years). The pathways identified by our algorithm have previously been confirmed in the literature as viable, high-yield synthesis routes. Prospectively, our algorithm could facilitate the design of novel, non-native synthesis routes by efficiently exploring the diversity of biochemical transformations in nature. Copyright © 2011 Elsevier Inc. All rights reserved.

Dissecting Bacterial Cell Wall Entry and Signaling in Eukaryotic Cells: an Actin-Dependent Pathway Parallels Platelet-Activating Factor Receptor-Mediated Endocytosis.

Science.gov (United States)

Loh, Lip Nam; Gao, Geli; Tuomanen, Elaine I

2017-01-03

The Gram-positive bacterial cell wall (CW) peptidoglycan-teichoic acid complex is released into the host environment during bacterial metabolism or death. It is a highly inflammatory Toll-like receptor 2 (TLR2) ligand, and previous in vivo studies have demonstrated its ability to recapitulate pathological features of pneumonia and meningitis. We report that an actin-dependent pathway is involved in the internalization of the CW by epithelial and endothelial cells, in addition to the previously described platelet-activating factor receptor (PAFr)-dependent uptake pathway. Unlike the PAFr-dependent pathway, which is mediated by clathrin and dynamin and does not lead to signaling, the alternative pathway is sensitive to 5-(N-ethyl-N-isopropyl) amiloride (EIPA) and engenders Rac1, Cdc42, and phosphatidylinositol 3-kinase (PI3K) signaling. Upon internalization by this macropinocytosis-like pathway, CW is trafficked to lysosomes. Intracellular CW trafficking is more complex than previously recognized and suggests multiple points of interaction with and without innate immune signaling. Streptococcus pneumoniae is a major human pathogen infecting the respiratory tract and brain. It is an established model organism for understanding how infection injures the host. During infection or bacterial growth, bacteria shed their cell wall (CW) into the host environment and trigger inflammation. A previous study has shown that CW enters and crosses cell barriers by interacting with a receptor on the surfaces of host cells, termed platelet-activating factor receptor (PAFr). In the present study, by using cells that are depleted of PAFr, we identified a second pathway with features of macropinocytosis, which is a receptor-independent fluid uptake mechanism by cells. Each pathway contributes approximately the same amount of cell wall trafficking, but the PAFr pathway is silent, while the new pathway appears to contribute to the host inflammatory response to CW insult. Copyright © 2017

HSP60 mediates the neuroprotective effects of curcumin by suppressing microglial activation.

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Ding, Feijia; Li, Fan; Li, Yunhong; Hou, Xiaolin; Ma, Yi; Zhang, Nan; Ma, Jiao; Zhang, Rui; Lang, Bing; Wang, Hongyan; Wang, Yin

2016-08-01

Curcumin has anti-inflammatory and antioxidant properties and has been widely used to treat or prevent neurodegenerative diseases. However, the mechanisms underlying the neuroprotective effects of curcumin are not well known. In the present study, the effect of curcumin on lipopolysaccharide (LPS)-stimulated BV2 mouse microglia cells was investigated using enzyme-linked immunosorbent assays of the culture medium and western blotting of cell lysates. The results showed that curcumin significantly inhibited the LPS-induced expression and release of heat shock protein 60 (HSP60) in the BV2 cells. The level of heat shock factor (HSF)-1 was upregulated in LPS-activated BV2 microglia, indicating that the increased expression of HSP60 was driven by HSF-1 activation. However, the increased HSF-1 level was downregulated by curcumin. Extracellular HSP60 is a ligand of Toll-like receptor 4 (TLR-4), and the level of the latter was increased in the LPS-activated BV2 microglia and inhibited by curcumin. The activation of TLR-4 is known to be associated with the activation of myeloid differentiation primary response 88 (MyD88) and nuclear factor (NF)-κB, with the subsequent production of proinflammatory and neurotoxic factors. In the present study, curcumin demonstrated marked suppression of the LPS-induced expression of MyD88, NF-κB, caspase-3, inducible nitric oxide synthase, tumor necrosis factor-α, interleukin (IL)-1β and IL-6 in the microglia. These results indicate that curcumin may exert its neuroprotective and anti-inflammatory effects by inhibiting microglial activation through the HSP60/TLR-4/MyD88/NF-κB signaling wpathway. Therefore, curcumin may be useful for the treatment of neurodegenerative diseases that are associated with microglial activation.

RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.

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Julia F Pielage

2008-03-01

Full Text Available Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of approximately 80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa-induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections.

Intracellular expression of toll-like receptor 4 in neuroblastoma cells and their unresponsiveness to lipopolysaccharide

International Nuclear Information System (INIS)

Hassan, Ferdaus; Islam, Shamima; Tumurkhuu, Gantsetseg; Naiki, Yoshikazu; Koide, Naoki; Mori, Isamu; Yoshida, Tomoaki; Yokochi, Takashi

2006-01-01

Recently it has been reported that, toll-like receptors (TLRs) are expressed on a series of tumor cells, such as colon cancer, breast cancer, prostate cancer, melanoma and lung cancer. Although some cancer cells like melanoma cells are known to respond to lipopolysaccharide (LPS) via TLR4, not all cancer cells are positive for TLR4. There is little information on the expression and function of TLR4 in neuroblastoma cells. In this study, we investigated the expression of TLR4 in human neuroblastoma NB-1 cell line. Expression and localization of TLR4 were detected by reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometric analysis, respectively. Activation of nuclear factor (NF)-κB by LPS was detected by degradation of IκB-α and NF-κB luciferase assay. Activation and expression of mitogen-activated protein (MAP) kinase and interferon regulatory factor (IRF)-3 was detected by immunoblot analysis. Human NB-1 neuroblastoma cells expressed intracellular form of TLR4, but not the cell surface form. Further, NB-1 cells express CD14, MD2 and MyD88, which are required for LPS response. However, LPS did not significantly induce NF-κB activation in NB-1 cells although it slightly degraded IκB-α. NB-1 cells expressed no IRF-3, which plays a pivotal role on the MyD88-independent pathway of LPS signaling. Collectively, NB-1 cells are capable to avoid their response to LPS. Although human NB-1 neuroblastoma cells possessed all the molecules required for LPS response, they did not respond to LPS. It might be responsible for intracellular expression of TLR4 or lack of IRF-3

Endocytic pathways involved in PLGA nanoparticle uptake by grapevine cells and role of cell wall and membrane in size selection.

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Palocci, Cleofe; Valletta, Alessio; Chronopoulou, Laura; Donati, Livia; Bramosanti, Marco; Brasili, Elisa; Baldan, Barbara; Pasqua, Gabriella

2017-12-01

PLGA NPs' cell uptake involves different endocytic pathways. Clathrin-independent endocytosis is the main internalization route. The cell wall plays a more prominent role than the plasma membrane in NPs' size selection. In the last years, many studies on absorption and cell uptake of nanoparticles by plants have been conducted, but the understanding of the internalization mechanisms is still largely unknown. In this study, polydispersed and monodispersed poly(lactic-co-glycolic) acid nanoparticles (PLGA NPs) were synthesized, and a strategy combining the use of transmission electron microscopy (TEM), confocal analysis, fluorescently labeled PLGA NPs, a probe for endocytic vesicles (FM4-64), and endocytosis inhibitors (i.e., wortmannin, ikarugamycin, and salicylic acid) was employed to shed light on PLGA NP cell uptake in grapevine cultured cells and to assess the role of the cell wall and plasma membrane in size selection of PLGA NPs. The ability of PLGA NPs to cross the cell wall and membrane was confirmed by TEM and fluorescence microscopy. A strong adhesion of PLGA NPs to the outer side of the cell wall was observed, presumably due to electrostatic interactions. Confocal microscopy and treatment with endocytosis inhibitors suggested the involvement of both clathrin-dependent and clathrin-independent endocytosis in cell uptake of PLGA NPs and the latter appeared to be the main internalization pathway. Experiments on grapevine protoplasts revealed that the cell wall plays a more prominent role than the plasma membrane in size selection of PLGA NPs. While the cell wall prevents the uptake of PLGA NPs with diameters over 50 nm, the plasma membrane can be crossed by PLGA NPs with a diameter of 500-600 nm.

49 CFR 393.88 - Television receivers.

Science.gov (United States)

2010-10-01

... 49 Transportation 5 2010-10-01 2010-10-01 false Television receivers. 393.88 Section 393.88... NECESSARY FOR SAFE OPERATION Miscellaneous Parts and Accessories § 393.88 Television receivers. Any motor vehicle equipped with a television viewer, screen or other means of visually receiving a television...

In vitro expression of Sec-dependent pathway and type 4B secretion system in Piscirickettsia salmonis.

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Cortés, Marcos; Sánchez, Patricio; Ruiz, Pamela; Haro, Ronie; Sáez, Jerson; Sánchez, Fabián; Hernández, Mauricio; Oliver, Cristian; Yáñez, Alejandro J

2017-09-01

Piscirickettsia salmonis is an intracellular bacterium and the causative agent of Piscirickettsiosis, a disease responsible for considerable mortalities in the Chilean salmon farming industry. Currently, P. salmonis protein translocation across the membrane and the mechanisms by which virulence factors are delivered to host cells are poorly understood. However, it is known that Gram-negative bacteria possess several mechanisms that transport proteins to the periplasmic and extracellular compartments. The aim of this study was to evaluate the expressional changes of several genes in the P. salmonis Sec-dependent pathway and type 4B secretion system during in vitro infection. Genes homologous and the main proteins belonging to Sec-dependent pathway and Type 4 Dot/Icm secretion system were found in the genome and proteome of P. salmonis AUSTRAL-005 strain. Additionally, several genes of these protein transport mechanisms were overexpressed during in vitro P. salmonis infection in SHK-1 cell line. The obtained data indicate that the Sec-dependent pathway and Type 4B secretion system are biologically active during P. salmonis infection. These mechanisms could contribute to the recycling of proteins into the inner and outer bacterial membrane and in translocate virulence factors to infected cell, which would favor the structural integrity and virulence of this bacterium. Copyright © 2017 Elsevier Ltd. All rights reserved.

Study to elucidate formation pathways of selected roast-smelling odorants upon extrusion cooking.

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Davidek, Tomas; Festring, Daniel; Dufossé, Thierry; Novotny, Ondrej; Blank, Imre

2013-10-30

The formation pathways of the N-containing roast-smelling compounds 2-acetyl-1-pyrroline, 2-acetyl-1(or 3),4,5,6-tetrahydropyridine, and their structural analogues 2-propionyl-1-pyrroline and 2-propionyl-1(or 3),4,5,6-tetrahydropyridine were studied upon extrusion cooking using the CAMOLA approach. The samples were produced under moderate extrusion conditions (135 °C, 20% moisture, 400 rpm) employing a rice-based model recipe enriched with flavor precursors ([U-(13)C6]-D-glucose, D-glucose, glycine, L-proline, and L-ornithine). The obtained data indicate that the formation of these compounds upon extrusion follows pathways similar to those reported for nonsheared model systems containing D-glucose and L-proline. 2-Acetyl-1-pyrroline is formed (i) by acylation of 1-pyrroline via C2 sugar fragments (major pathway) and (ii) via ring-opening of 1-pyrroline incorporating C3 sugar fragments (minor pathway), whereas 2-propionyl-1-pyrroline incorporates exclusively C3 sugar fragments. 2-Acetyl-1(or 3),4,5,6-tetrahydropyridine and the corresponding propionyl analogue incorporate C3 and C4 sugar fragments, respectively. In addition, it has been shown that the formation of 2-acetyl-1-pyrroline in low-moisture systems depends on the pH value of the reaction mixture.

50 CFR 18.88 - Cross-examination.

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2010-10-01

... 50 Wildlife and Fisheries 6 2010-10-01 2010-10-01 false Cross-examination. 18.88 Section 18.88... PLANTS (CONTINUED) MARINE MAMMALS Notice and Hearing on Section 103 Regulations § 18.88 Cross-examination... examination; and (2) Exclude cross-examination questions that are immaterial, irrelevant or unduly repetitious...

50 CFR 260.88 - Political activity.

Science.gov (United States)

2010-10-01

... part in political management or in political campaigns. Political activities in city, county, State, or... 50 Wildlife and Fisheries 7 2010-10-01 2010-10-01 false Political activity. 260.88 Section 260.88... Products for Human Consumption Miscellaneous § 260.88 Political activity. All inspectors and licensed...

Human cerebral venous outflow pathway depends on posture and central venous pressure

Science.gov (United States)

Gisolf, J; van Lieshout, J J; van Heusden, K; Pott, F; Stok, W J; Karemaker, J M

2004-01-01

Internal jugular veins are the major cerebral venous outflow pathway in supine humans. In upright humans the positioning of these veins above heart level causes them to collapse. An alternative cerebral outflow pathway is the vertebral venous plexus. We set out to determine the effect of posture and central venous pressure (CVP) on the distribution of cerebral outflow over the internal jugular veins and the vertebral plexus, using a mathematical model. Input to the model was a data set of beat-to-beat cerebral blood flow velocity and CVP measurements in 10 healthy subjects, during baseline rest and a Valsalva manoeuvre in the supine and standing position. The model, consisting of 2 jugular veins, each a chain of 10 units containing nonlinear resistances and capacitors, and a vertebral plexus containing a resistance, showed blood flow mainly through the internal jugular veins in the supine position, but mainly through the vertebral plexus in the upright position. A Valsalva manoeuvre while standing completely re-opened the jugular veins. Results of ultrasound imaging of the right internal jugular vein cross-sectional area at the level of the laryngeal prominence in six healthy subjects, before and during a Valsalva manoeuvre in both body positions, correlate highly with model simulation of the jugular cross-sectional area (R2 = 0.97). The results suggest that the cerebral venous flow distribution depends on posture and CVP: in supine humans the internal jugular veins are the primary pathway. The internal jugular veins are collapsed in the standing position and blood is shunted to an alternative venous pathway, but a marked increase in CVP while standing completely re-opens the jugular veins. PMID:15284348

Identification of a novel Gnao-mediated alternate olfactory signaling pathway in murine OSNs

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Paul eScholz

2016-03-01

Full Text Available It is generally agreed that in olfactory sensory neurons (OSNs, the binding of odorant molecules to their specific olfactory receptor (OR triggers a cAMP-dependent signaling cascade, activating cyclic-nucleotide gated (CNG channels. However, considerable controversy dating back more than 20 years has surrounded the question of whether alternate signaling plays a role in mammalian olfactory transduction. In this study, we demonstrate a specific alternate signaling pathway in Olfr73-expressing OSNs. Methylisoeugenol (MIEG and at least one other known weak Olfr73 agonist (Raspberry Ketone trigger a signaling cascade independent from the canonical pathway, leading to the depolarization of the cell. Interestingly, this pathway is mediated by Gnao activation, leading to Cl- efflux; however, the activation of adenylyl cyclase III (ACIII, the recruitment of Ca2+ from extra-or intracellular stores, and phosphatidylinositol 3-kinase-dependent signaling (PI signaling are not involved. Furthermore, we demonstrated that our newly identified pathway coexists with the canonical olfactory cAMP pathway in the same OSN and can be triggered by the same OR in a ligand-selective manner. We suggest that this pathway might reflect a mechanism for odor recognition predominantly used in early developmental stages before olfactory cAMP signaling is fully developed. Taken together, our findings support the existence of at least one odor-induced alternate signal transduction pathway in native OSNs mediated by Olfr73 in a ligand-selective manner.

Selective maintenance for multi-state series–parallel systems under economic dependence

International Nuclear Information System (INIS)

Dao, Cuong D.; Zuo, Ming J.; Pandey, Mayank

2014-01-01

This paper presents a study on selective maintenance for multi-state series–parallel systems with economically dependent components. In the selective maintenance problem, the maintenance manager has to decide which components should receive maintenance activities within a finite break between missions. All the system reliabilities in the next operating mission, the available budget and the maintenance time for each component from its current state to a higher state are taken into account in the optimization models. In addition, the components in series–parallel systems are considered to be economically dependent. Time and cost savings will be achieved when several components are simultaneously repaired in a selective maintenance strategy. As the number of repaired components increases, the saved time and cost will also increase due to the share of setting up between components and another additional reduction amount resulting from the repair of multiple identical components. Different optimization models are derived to find the best maintenance strategy for multi-state series–parallel systems. A genetic algorithm is used to solve the optimization models. The decision makers may select different components to be repaired to different working states based on the maintenance objective, resource availabilities and how dependent the repair time and cost of each component are

19 CFR 141.88 - Computed value.

Science.gov (United States)

2010-04-01

... 19 Customs Duties 2 2010-04-01 2010-04-01 false Computed value. 141.88 Section 141.88 Customs... (CONTINUED) ENTRY OF MERCHANDISE Invoices § 141.88 Computed value. When the port director determines that information as to computed value is necessary in the appraisement of any class or kind of merchandise, he...

A fluorescent glycolipid-binding peptide probe traces cholesterol dependent microdomain-derived trafficking pathways.

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Steffen Steinert

Full Text Available BACKGROUND: The uptake and intracellular trafficking of sphingolipids, which self-associate into plasma membrane microdomains, is associated with many pathological conditions, including viral and toxin infection, lipid storage disease, and neurodegenerative disease. However, the means available to label the trafficking pathways of sphingolipids in live cells are extremely limited. In order to address this problem, we have developed an exogenous, non-toxic probe consisting of a 25-amino acid sphingolipid binding domain, the SBD, derived from the amyloid peptide Abeta, and conjugated by a neutral linker with an organic fluorophore. The current work presents the characterization of the sphingolipid binding and live cell trafficking of this novel probe, the SBD peptide. SBD was the name given to a motif originally recognized by Fantini et al in a number of glycolipid-associated proteins, and was proposed to interact with sphingolipids in membrane microdomains. METHODOLOGY/PRINCIPAL FINDINGS: In accordance with Fantini's model, optimal SBD binding to membranes depends on the presence of sphingolipids and cholesterol. In synthetic membrane binding assays, SBD interacts preferentially with raft-like lipid mixtures containing sphingomyelin, cholesterol, and complex gangliosides in a pH-dependent manner, but is less glycolipid-specific than Cholera toxin B (CtxB. Using quantitative time-course colocalization in live cells, we show that the uptake and intracellular trafficking route of SBD is unlike that of either the non-raft marker Transferrin or the raft markers CtxB and Flotillin2-GFP. However, SBD traverses an endolysosomal route that partially intersects with raft-associated pathways, with a major portion being diverted at a late time point to rab11-positive recycling endosomes. Trafficking of SBD to acidified compartments is strongly disrupted by cholesterol perturbations, consistent with the regulation of sphingolipid trafficking by cholesterol

Stochastic Modeling of the Clathrin-dependent and -independent Endocytic Pathways

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Deng, Hua; Dutta, Prashanta; Liu, Jin

2017-11-01

Endocytosis is one of the important processes that bioparticles use to enter the cells. During endocytosis the membrane-bound vesicles are formed by the invagination of plasma membrane as a result of interactions among many proteins and cytoskeletons. The clathrin-mediated endocytosis is one of the most significant form of endocytosis, where the dynamic assembly of clathrin-coated pits play a critical role. While herpes simplex virus-1 has recently shown to infect cell by a novel phagocytosis-like endocytic pathway where actin polymerization may facilitate the viral entry. In this work, we propose a stochastic model for both clathrin-dependent and -independent endocytic pathways based on Monte Carlo simulations. The important roles of clathrin coating and actin cytoskeleton as well as the impact of other biological parameters are studied. Our preliminary results indicate that there exist an intermediate particle size and ligand density that maximize the internalization efficiency. Below a critical size or surface ligand density, it is difficult for the entry of a single particle, which means clustering may needed for more efficient internalization. We also find that lower membrane bending rigidity may help promote the bioparticle entry. Research reported in this publication was supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number R01GM122081.

Regulation of energy partitioning and alternative electron transport pathways during cold acclimation of lodgepole pine is oxygen dependent.

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Savitch, Leonid V; Ivanov, Alexander G; Krol, Marianna; Sprott, David P; Oquist, Gunnar; Huner, Norman P A

2010-09-01

Second year needles of Lodgepole pine (Pinus contorta L.) were exposed for 6 weeks to either simulated control summer ['summer'; 25 °C/250 photon flux denisty (PFD)], autumn ('autumn'; 15°C/250 PFD) or winter conditions ('winter'; 5 °C/250 PFD). We report that the proportion of linear electron transport utilized in carbon assimilation (ETR(CO2)) was 40% lower in both 'autumn' and 'winter' pine when compared with the 'summer' pine. In contrast, the proportion of excess photosynthetic linear electron transport (ETR(excess)) not used for carbon assimilation within the total ETR(Jf) increased by 30% in both 'autumn' and 'winter' pine. In 'autumn' pine acclimated to 15°C, the increased amounts of 'excess' electrons were directed equally to 21  kPa O2-dependent and 2  kPa O2-dependent alternative electron transport pathways and the fractions of excitation light energy utilized by PSII photochemistry (Φ(PSII)), thermally dissipated through Φ(NPQ) and dissipated by additional quenching mechanism(s) (Φ(f,D)) were similar to those in 'summer' pine. In contrast, in 'winter' needles acclimated to 5 °C, 60% of photosynthetically generated 'excess' electrons were utilized through the 2  kPa O2-dependent electron sink and only 15% by the photorespiratory (21  kPa O2) electron pathway. Needles exposed to 'winter' conditions led to a 3-fold lower Φ(PSII), only a marginal increase in Φ(NPQ) and a 2-fold higher Φ(f,D), which was O2 dependent compared with the 'summer' and 'autumn' pine. Our results demonstrate that the employment of a variety of alternative pathways for utilization of photosynthetically generated electrons by Lodgepole pine depends on the acclimation temperature. Furthermore, dissipation of excess light energy through constitutive non-photochemical quenching mechanisms is O2 dependent.

Movement reveals scale dependence in habitat selection of a large ungulate

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Northrup, Joseph; Anderson, Charles R.; Hooten, Mevin B.; Wittemyer, George

2016-01-01

Ecological processes operate across temporal and spatial scales. Anthropogenic disturbances impact these processes, but examinations of scale dependence in impacts are infrequent. Such examinations can provide important insight to wildlife–human interactions and guide management efforts to reduce impacts. We assessed spatiotemporal scale dependence in habitat selection of mule deer (Odocoileus hemionus) in the Piceance Basin of Colorado, USA, an area of ongoing natural gas development. We employed a newly developed animal movement method to assess habitat selection across scales defined using animal-centric spatiotemporal definitions ranging from the local (defined from five hour movements) to the broad (defined from weekly movements). We extended our analysis to examine variation in scale dependence between night and day and assess functional responses in habitat selection patterns relative to the density of anthropogenic features. Mule deer displayed scale invariance in the direction of their response to energy development features, avoiding well pads and the areas closest to roads at all scales, though with increasing strength of avoidance at coarser scales. Deer displayed scale-dependent responses to most other habitat features, including land cover type and habitat edges. Selection differed between night and day at the finest scales, but homogenized as scale increased. Deer displayed functional responses to development, with deer inhabiting the least developed ranges more strongly avoiding development relative to those with more development in their ranges. Energy development was a primary driver of habitat selection patterns in mule deer, structuring their behaviors across all scales examined. Stronger avoidance at coarser scales suggests that deer behaviorally mediated their interaction with development, but only to a degree. At higher development densities than seen in this area, such mediation may not be possible and thus maintenance of sufficient

Transcutaneous immunization with a novel imiquimod nanoemulsion induces superior T cell responses and virus protection.

Science.gov (United States)

Lopez, Pamela Aranda; Denny, Mark; Hartmann, Ann-Kathrin; Alflen, Astrid; Probst, Hans Christian; von Stebut, Esther; Tenzer, Stefan; Schild, Hansjörg; Stassen, Michael; Langguth, Peter; Radsak, Markus P

2017-09-01

Transcutaneous immunization (TCI) is a novel vaccination strategy utilizing the skin associated lymphatic tissue to induce immune responses. TCI using a cytotoxic T lymphocyte (CTL) epitope and the Toll-like receptor 7 (TLR7) agonist imiquimod mounts strong CTL responses by activation and maturation of skin-derived dendritic cells (DCs) and their migration to lymph nodes. However, TCI based on the commercial formulation Aldara only induces transient CTL responses that needs further improvement for the induction of durable therapeutic immune responses. Therefore we aimed to develop a novel imiquimod solid nanoemulsion (IMI-Sol) for TCI with superior vaccination properties suited to induce high quality T cell responses for enhanced protection against infections. TCI was performed by applying a MHC class I or II restricted epitope along with IMI-Sol or Aldara (each containing 5% Imiquimod) on the shaved dorsum of C57BL/6, IL-1R, Myd88, Tlr7 or Ccr7 deficient mice. T cell responses as well as DC migration upon TCI were subsequently analyzed by flow cytometry. To determine in vivo efficacy of TCI induced immune responses, CTL responses and frequency of peptide specific T cells were evaluated on day 8 or 35 post vaccination and protection in a lymphocytic choriomeningitis virus (LCMV) infection model was assessed. TCI with the imiquimod formulation IMI-Sol displayed equal skin penetration of imiquimod compared to Aldara, but elicited superior CD8 + as well as CD4 + T cell responses. The induction of T-cell responses induced by IMI-Sol TCI was dependent on the TLR7/MyD88 pathway and independent of IL-1R. IMI-Sol TCI activated skin-derived DCs in skin-draining lymph nodes more efficiently compared to Aldara leading to enhanced protection in a LCMV infection model. Our data demonstrate that IMI-Sol TCI can overcome current limitations of previous imiquimod based TCI approaches opening new perspectives for transcutaneous vaccination strategies and allowing the use of this

40 CFR 763.88 - Assessment.

Science.gov (United States)

2010-07-01

... ACBM in the school building. (2) Each accredited inspector providing a written assessment shall sign... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Assessment. 763.88 Section 763.88 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) TOXIC SUBSTANCES CONTROL ACT ASBESTOS...

Diclofenac pretreatment effects on the toll-like receptor 4/nuclear factor kappa B-mediated inflammatory response to eccentric exercise in rat liver.

Science.gov (United States)

Barcelos, Rômulo Pillon; Bresciani, Guilherme; Rodriguez-Miguelez, Paula; Cuevas, Maria José; Soares, Félix Alexandre Antunes; Barbosa, Nilda Vargas; González-Gallego, Javier

2016-03-01

Acute exercise is a stress stimulus that may cause cell damage through the activation of the toll-like receptor (TLR)4 pathway, resulting in the translocation of nuclear factor kappa B (NF-κB) into the cell nucleus and the upregulation of inflammatory genes. Nonsteroidal anti-inflammatory drugs, such as diclofenac, are often prescribed to counteract exercise-induced inflammation. This study analyzed effects of diclofenac pretreatment on the TLR4/NF-κB pathway in rat liver after an acute eccentric exercise. Twenty male Wistar rats were divided in four groups: control-saline, control-diclofenac, exercise-saline and exercise-diclofenac. The rats received saline or diclofenac (10mg/kg) for 7days prior to an eccentric exercise bout. After exercise there was an increase in TLR4, myeloid differentiation primary response gene 88 (MyD88), TIR domain-containing adaptor inducing interferon (TRIF) and p65 NF-κB subunit protein levels. Exercise also resulted in increased mRNA and protein expression of interleukin (IL)-6, inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF)-α. Proinflammatory effects of exercise were prevented by the administration of diclofenac, which blunted the activation of the TLR4/NF-κB pathway and the inflammatory response in the liver of exercised rats. Results from the present study highlight the role of TLR4 as a target for anti-inflammatory interventions. Copyright © 2016 Elsevier Inc. All rights reserved.

Role of toll-like receptor 4 on the immune escape of human oral squamous cell carcinoma and resistance of cisplatin-induced apoptosis

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Sun Zujun

2012-05-01

Full Text Available Abstract Background Toll-like receptor 4 (TLR4 is expressed on immune cells as a sensor that recognizes lipopolysaccharide (LPS, a microbial conserved component. It has recently been determined that the expression of TLR4 is also found in various types of tumor cells. Cisplatin is a widely used chemotherapeutic agent for oral squamous cell carcinoma (OSCC treatment. However, the mechanisms responsible for cisplatin resistance are not well understood. Results The present study was designed to elucidate the role of TLR4 expression in human OSCC regarding immune escape and apoptotic resistance to cisplatin. TLR4 and the myeloid differentiation primary response gene 88 (MyD88 were highly expressed in OSCC cell lines. Upon LPS stimulation both NF-κB and p38 MAPK pathways were activated in OSCC cell lines, followed by the production of large quantities of IL-6, IL-8 and VEGF compared with human immortalized oral epithelia cells (HIOECs. OSCC cell lines were found to be resistant to cisplatin-mediated apoptosis after pretreatment with LPS. Conclusions Our results suggested that TLR4 was functionally expressed in human OSCC cells and development of resistance to cisplatin in human OSCC might occur through the mechanism involving TLR4 and its signaling pathway. Suppression of TLR4 and its signaling pathway might thus elevate sensitivity to cisplatin and potentially help improve the prognosis of patients with OSCC.

A novel approach to select differential pathways associated with hypertrophic cardiomyopathy based on gene co‑expression analysis.

Science.gov (United States)

Chen, Xiao-Min; Feng, Ming-Jun; Shen, Cai-Jie; He, Bin; Du, Xian-Feng; Yu, Yi-Bo; Liu, Jing; Chu, Hui-Min

2017-07-01

The present study was designed to develop a novel method for identifying significant pathways associated with human hypertrophic cardiomyopathy (HCM), based on gene co‑expression analysis. The microarray dataset associated with HCM (E‑GEOD‑36961) was obtained from the European Molecular Biology Laboratory‑European Bioinformatics Institute database. Informative pathways were selected based on the Reactome pathway database and screening treatments. An empirical Bayes method was utilized to construct co‑expression networks for informative pathways, and a weight value was assigned to each pathway. Differential pathways were extracted based on weight threshold, which was calculated using a random model. In order to assess whether the co‑expression method was feasible, it was compared with traditional pathway enrichment analysis of differentially expressed genes, which were identified using the significance analysis of microarrays package. A total of 1,074 informative pathways were screened out for subsequent investigations and their weight values were also obtained. According to the threshold of weight value of 0.01057, 447 differential pathways, including folding of actin by chaperonin containing T‑complex protein 1 (CCT)/T‑complex protein 1 ring complex (TRiC), purine ribonucleoside monophosphate biosynthesis and ubiquinol biosynthesis, were obtained. Compared with traditional pathway enrichment analysis, the number of pathways obtained from the co‑expression approach was increased. The results of the present study demonstrated that this method may be useful to predict marker pathways for HCM. The pathways of folding of actin by CCT/TRiC and purine ribonucleoside monophosphate biosynthesis may provide evidence of the underlying molecular mechanisms of HCM, and offer novel therapeutic directions for HCM.