Intravenous Administration of Toll-Like Receptor Inhibitory Peptide 1 is Effective for the Treatment of Systemic Lupus Erythematosus in a Mus musculus Model

Objective@#Systemic lupus erythematosus (SLE) is a common chronic autoimmune inflammatory disease. According to recent studies, signaling through Toll-like receptor (TLR) protein, which promotes the production of inflammatory cytokines, leads to the development of SLE. TLR-inhibitory peptide 1 (TIP1) has been newly identified for the treatment of autoimmune diseases. @*Methods@#The effect of TIP1 was analyzed in an SLE mouse model (MRL/lpr). The mice in the control treatment group (n=5) were administered an intravenous injection of phosphate-buffered saline twice weekly, whereas the mice in the TIP1 treatment group (n=6) were administered an intravenous injection of TIP1 (1 nmol/g) twice weekly. MRL/mpj mice (n=5) were selected as normal controls. The mice were injected for 4 weeks between 14 and 18 weeks of age, followed by assays of their spleen, kidneys, lymph nodes, serum, and urine. @*Results@#The antinuclear antibody and inflammatory cytokine (interferon-α) in the serum as well as levels of albumin in the urine of the mice in the TIP1 treatment group had decreased when compared to those of mice in the control treatment group. Kidney inflammation in mice in the TIP1 treatment group was alleviated. The mRNA expression levels of TLR7- or TLR9-related downstream signaling molecules also decreased in all organs of the mice in the TIP1treatment group. @*Conclusion@#Intravenous treatment with TIP1 reduces symptoms and markers of inflammation in MRL/lpr mice. Hence, TIP1 is a promising medication for the treatment of SLE.

Intravenous Administration of Toll-Like Receptor Inhibitory Peptide 1 is Effective for the Treatment of Systemic Lupus Erythematosus in a Mus musculus Model

Objective@#Systemic lupus erythematosus (SLE) is a common chronic autoimmune inflammatory disease. According to recent studies, signaling through Toll-like receptor (TLR) protein, which promotes the production of inflammatory cytokines, leads to the development of SLE. TLR-inhibitory peptide 1 (TIP1) has been newly identified for the treatment of autoimmune diseases. @*Methods@#The effect of TIP1 was analyzed in an SLE mouse model (MRL/lpr). The mice in the control treatment group (n=5) were administered an intravenous injection of phosphate-buffered saline twice weekly, whereas the mice in the TIP1 treatment group (n=6) were administered an intravenous injection of TIP1 (1 nmol/g) twice weekly. MRL/mpj mice (n=5) were selected as normal controls. The mice were injected for 4 weeks between 14 and 18 weeks of age, followed by assays of their spleen, kidneys, lymph nodes, serum, and urine. @*Results@#The antinuclear antibody and inflammatory cytokine (interferon-α) in the serum as well as levels of albumin in the urine of the mice in the TIP1 treatment group had decreased when compared to those of mice in the control treatment group. Kidney inflammation in mice in the TIP1 treatment group was alleviated. The mRNA expression levels of TLR7- or TLR9-related downstream signaling molecules also decreased in all organs of the mice in the TIP1treatment group. @*Conclusion@#Intravenous treatment with TIP1 reduces symptoms and markers of inflammation in MRL/lpr mice. Hence, TIP1 is a promising medication for the treatment of SLE.

Middle East respiratory syndrome coronavirus: transmission, virology and therapeutic targeting to aid in outbreak control

Middle East respiratory syndrome coronavirus (MERS-CoV) causes high fever, cough, acute respiratory tract infection and multiorgan dysfunction that may eventually lead to the death of the infected individuals. MERS-CoV is thought to be transmitted to humans through dromedary camels. The occurrence of the virus was first reported in the Middle East and it subsequently spread to several parts of the world. Since 2012, about 1368 infections, including ~487 deaths, have been reported worldwide. Notably, the recent human-to-human \'superspreading' of MERS-CoV in hospitals in South Korea has raised a major global health concern. The fatality rate in MERS-CoV infection is four times higher compared with that of the closely related severe acute respiratory syndrome coronavirus infection. Currently, no drug has been clinically approved to control MERS-CoV infection. In this study, we highlight the potential drug targets that can be used to develop anti-MERS-CoV therapeutics.

Negative regulatory approaches to the attenuation of Toll-like receptor signaling

Toll-like receptors (TLRs) are pivotal components of the innate immune response, which is responsible for eradicating invading microorganisms through the induction of inflammatory molecules. These receptors are also involved in responding to harmful endogenous molecules and have crucial roles in the activation of the innate immune system and shaping the adaptive immune response. However, TLR signaling pathways must be tightly regulated because undue TLR stimulation may disrupt the fine balance between pro- and anti-inflammatory responses. Such disruptions may harm the host through the development of autoimmune and inflammatory diseases, such as rheumatoid arthritis and systemic lupus erythematosus. Several studies have investigated the regulatory pathways of TLRs that are essential for modulating proinflammatory responses. These studies reported several pathways and molecules that act individually or in combination to regulate immune responses. In this review, we have summarized recent advancements in the elucidation of the negative regulation of TLR signaling. Moreover, this review covers the modulation of TLR signaling at multiple levels, including adaptor complex destabilization, phosphorylation and ubiquitin-mediated degradation of signal proteins, manipulation of other receptors, and transcriptional regulation. Lastly, synthetic inhibitors have also been briefly discussed to highlight negative regulatory approaches in the treatment of inflammatory diseases.

Retraction: A Fosmid Cloning Strategy for Detecting the Widest Possible Spectrum of Microbes from the International Space Station Drinking Water System

No abstract available.

Systems Biological Approaches Reveal Non-additive Responses and Multiple Crosstalk Mechanisms between TLR and GPCR Signaling

A variety of ligands differ in their capacity to bind the receptor, elicit gene expression, and modulate physiological responses. Such receptors include Toll-like receptors (TLRs), which recognize various patterns of pathogens and lead to primary innate immune activation against invaders, and G-protein coupled receptors (GPCRs), whose interaction with their cognate ligands activates heterotrimeric G proteins and regulates specific downstream effectors, including immuno-stimulating molecules. Once TLRs are activated, they lead to the expression of hundreds of genes together and bridge the arm of innate and adaptive immune responses. We characterized the gene expression profile of Toll-like receptor 4 (TLR4) in RAW 264.7 cells when it bound with its ligand, 2-keto-3-deoxyoctonate (KDO), the active part of lipopolysaccharide. In addition, to determine the network communications among the TLR, Janus kinase (JAK)/signal transducer and activator of transcription (STAT), and GPCR, we tested RAW 264.7 cells with KDO, interferon-beta, or cAMP analog 8-Br. The ligands were also administered as a pair of double and triple combinations.

A Fosmid Cloning Strategy for Detecting the Widest Possible Spectrum of Microbes from the International Space Station Drinking Water System

In this study, fosmid cloning strategies were used to assess the microbial populations in water from the International Space Station (ISS) drinking water system (henceforth referred to as Prebiocide and Tank A water samples). The goals of this study were: to compare the sensitivity of the fosmid cloning strategy with that of traditional culture-based and 16S rRNA-based approaches and to detect the widest possible spectrum of microbial populations during the water purification process. Initially, microbes could not be cultivated, and conventional PCR failed to amplify 16S rDNA fragments from these low biomass samples. Therefore, randomly primed rolling-circle amplification was used to amplify any DNA that might be present in the samples, followed by size selection by using pulsed-field gel electrophoresis. The amplified high-molecular-weight DNA from both samples was cloned into fosmid vectors. Several hundred clones were randomly selected for sequencing, followed by Blastn/Blastx searches. Sequences encoding specific genes from Burkholderia, a species abundant in the soil and groundwater, were found in both samples. Bradyrhizobium and Mesorhizobium, which belong to rhizobia, a large community of nitrogen fixers often found in association with plant roots, were present in the Prebiocide samples. Ralstonia, which is prevalent in soils with a high heavy metal content, was detected in the Tank A samples. The detection of many unidentified sequences suggests the presence of potentially novel microbial fingerprints. The bacterial diversity detected in this pilot study using a fosmid vector approach was higher than that detected by conventional 16S rRNA gene sequencing.

Gene Expression Analysis of Galpha13-/- Knockout Mouse Embryos Reveals Perturbations in Galpha13 Signaling Related to Angiogenesis and Hypoxia

Angiogenesis is regulated by a large number of molecules and complex signaling mechanisms. The G protein Galpha13 is a part of this signaling mechanism as an endothelial cell movement regulator. Gene expression analysis of Galpha13 knockout mouse embryos was carried out to identify the role of Galpha13 in angiogenesis signaling during embryonic development. Hypoxia-inducible response factors including those acting as regulators of angiogenesis were over expressed, while genes related to the cell cycle, DNA replication, protein modification and cell-cell dissociation were under expressed. Functional annotation and network analysis indicate that Galpha13 -/- embryonic mice were exposed to hypoxic conditions. The present analysis of the time course highlighted the significantly high levels of disorder in the development of the cardiovascular system. The data suggested that hypoxia-inducible factors including those associated with angiogenesis and abnormalities related to endothelial cell division contributed to the developmental failure of Galpha13 knockout mouse embryos.

Gene expression profiling of light-induced retinal degeneration in phototransduction gene knockout mice

Exposure to light can induce photoreceptor cell death and exacerbate retinal degeneration. In this study, mice with genetic knockout of several genes, including rhodopsin kinase (Rhok-/-), arrestin (Sag-/-), transducin (Gnat1-/-), c-Fos (c-Fos-/-) and arrestin/transducin (Sag-/-/Gnat1-/-), were examined. We measured the expression levels of thousands of genes in order to investigate their roles in phototransduction signaling in light-induced retinal degeneration using DNA microarray technology and then further explored the gene network using pathway analysis tools. Several cascades of gene components were induced or inhibited as a result of corresponding gene knockout under specific light conditions. Transducin deletion blocked the apoptotic signaling induced by exposure to low light conditions, and it did not require c-Fos/AP-1. Deletion of c-Fos blocked the apoptotic signaling induced by exposure to high intensity light. In the present study, we identified many gene transcripts that are essential for the initiation of light-induced rod degeneration and proposed several important networks that are involved in pro- and anti-apoptotic signaling. We also demonstrated the different cascades of gene components that participate in apoptotic signaling under specific light conditions.

Toll-like receptor signal transduction

Toll-like receptors (TLRs) are the archetypal pattern recognition receptors in sensing exogenous pathogens. Activation of TLRs is a first line of defense of the immune system, leading to the activation and recruitment of neutrophils and macrophages to sites of infection and enhances antimicrobial activity. The TLR signaling through different intracellular molecules, such as MAP kinases and IkappaB kinases which are conserved signaling elements for many receptors, leads to a distinct set of proinflammatory gene expressions. However, how these pathways differentially and precisely control the transcription of identical genes remains largely unknown. Our review focuses on the details of up-to- date signaling molecules including negative regulators and their role in controlling innate immune response. We also stress the importance of developing systemic approaches for the global understanding of TLR signaling so that appropriate drug therapeutic targets can be identified for regulating inflammatory diseases.

Statistical Analysis of Gene Expression in Innate Immune Responses: Dynamic Interactions between MicroRNA and Signaling Molecules

MicroRNAs (miRNAs) are known to negatively control protein-coding genes by binding to messenger RNA (mRNA) in the cytoplasm. In innate immunity, the role of miRNA gene silencing is largely unknown. In this study, we performed microarray-based experiments using lipopolysaccharide (LPS)-stimulated macrophages derived from wild-type, MyD88 knockout (KO), TRIF KO, and MyD88/TRIF double KO mice. We employed a statistical approach to determine the importance of the commonality and specificity of miRNA binding sites among groups of temporally co-regulated genes. We demonstrate that both commonality and specificity are irrelevant to define a priori groups of co-downregulated genes. In addition, analyzing the various experimental conditions, we suggest that miRNA regulation may not only be a late-phase process (after transcription) but can also occur even early (1h) after stimulation in knockout conditions. This further indicates the existence of dynamic interactions between miRNA and signaling molecules/ transcription factor regulation; this is another proof for the need of shifting from a 'hard-wired' paradigm of gene regulation to a dynamical one in which the gene co-regulation is established on a case-by-case basis.

Fluorescence Quenching Causes Systematic Dye Bias in Microarray Experiments Using Cyanine Dye

The development of microarray technology has facilitated the understanding of gene expression profiles. Despite its convenience, the cause of dye-bias that confounds data interpretation in dual-color DNA microarray experiments is not well known. In order to economize time and money, it is necessary to identify the cause of dye bias, since designing dye-swaps to reduce the dye-specific bias tends to be very expensive. Hence, we sought to determine the reliable cause of systematic dye bias after treating murine macrophage RAW 264.7 cells with 2-keto-3-deoxyoctonate (KDO), interferon-beta (IFN-beta), and 8-bromoadenosine (8-BR). To find the cause of systematic dye bias from the point of view of fluorescence quenching, we examined the correlation between systematic dye bias and the proportion of each nucleotide in mRNA and oligonucleotide probe sequence. Cy3-dye bias was highly correlated with the proportion of adenines. Our results support the fact that systematic dye bias is affected by fluorescence quenching of each feature. In addition, we also found that the strength of fluorescence quenching is based on not only dye-dye interactions but also dye-nucleotide interactions as well.