Induction Of Toll-like Receptor Research Articles

OP0098 A Pattern of Toll-Like Receptor Activation Characterizes Lupus Patients with Antibodies to Ribonucleoprotein Complexes

BackgroundSystemic lupus is a highly heterogeneous disease. Along with production of autoantibodies against double stranded DNA, some patients also show reactivity for different ribonucleoprotein complexes (RNP), and they appear to...

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Toll-Like Receptor Activation in Immunity vs. Tolerance in Autoimmune Diabetes

Toll-like receptors: Key receptors of the innate immune system The human body constantly encounters a diverse spectrum of pathogens. To defend itself, a complex immune system has evolved consisting of two subdivisions: the innate and the adaptive immune systems. The innate immune system constitutes the so-called “first line of defense” and acts through highly conserved germ-line-encoded pattern recognition receptors (PRRs) (1). These receptors bind to common pathogen-associated molecular patterns (PAMPs), which are vital for the survival of the microorganisms and cannot be altered by mutations. Pattern recognition receptors were first discovered in the fruit fly Drosophila melanogaster. The PRRs in Drosophila, named Toll, play an important role in the recognition of microorganisms such as fungi as well as coordinating embryonic development of the dorso-ventral axis. Homologs of Toll in vertebrates are called Toll-like receptors (TLRs). The first TLR in humans was described in 1997 (2). To date, another 9 TLRs have been identified in humans and 13 in mice. Among these, only murine TLR10 is non-functional due to retrovirus insertion (3). Except for TLRs 3, 7, 8, and 9, which are expressed intracellularly, TLRs are located on the surface of innate and adaptive immune cells as well as on non-immune cells such as muscle cells, epithelial cells, adipocytes, and pancreatic beta cells (1). Cell-surface TLRs detect exogenous lipids, lipoproteins, and proteins from microbes; intracellular TLRs recognize bacterial and viral nucleic acids (3). Recognition of their cognate ligand triggers a complex signaling cascade that ultimately results in the induction of various pro-inflammatory chemokines and cytokines and the activation of the adaptive immune system. However, the activation of TLRs and the subsequent induction of inflammatory immune responses are not always beneficial to the host. TLRs are involved in the pathogenesis of various autoimmune and non-infective inflammatory diseases such as systemic lupus erythematosus, multiple sclerosis, arteriosclerosis, inflammatory bowel disease, diabetes, allergy, and cancer (4, 5). Their activation can either attenuate or boost the course of disease by inducing tolerance or triggering autoreactivity, respectively. The role of the TLRs in the pathogenesis of autoimmune-mediated diabetes, also referred to as type 1 diabetes (T1D), has been extensively studied (Table ​(Table1).1). T1D is a T-cell-mediated metabolic disorder with progressive destruction of insulin-producing pancreatic β cells (6). During the course of disease development, diabetogenic T-cells, macrophages, and dendritic cells will infiltrate the pancreatic islets and cause islet inflammation and eventually β cell loss. Table 1 TLR-related studies in the field of T1D. This opinion letter focuses on the beneficial and detrimental aspects of TLR induction in the course of T1D development.

Lactate Reduces Liver and Pancreatic Injury in Toll-Like Receptor– and Inflammasome-Mediated Inflammation via GPR81-Mediated Suppression of Innate Immunity

The NACHT, LRR, and pyrin domain-containing protein 3 (NLRP3) inflammasome induces inflammation in response to organ injury, but little is known about its regulation. Toll-like receptors (TLRs) provide the first signal required for activation of the inflammasome and stimulate aerobic glycolysis to generate lactate. We examined whether lactate and the lactate receptor, Gi-protein-coupled receptor 81 (GPR81), regulate TLR induction of signal 1 and limit inflammasome activation and organ injury. Primary mouse macrophages and human monocytes were incubated with TLR4 agonists and lactate and assayed for levels of pro-interleukin (IL)1β, NLRP3, and caspase-1 (CASP1); release of IL1β; and activation of nuclear factor-κB (NF-κB) and caspase-1. Small interfering RNAs were used to reduce levels of GPR81 and arrestin β-2 (ARRB2), and an NF-κB luciferase reporter transgene was transfected in RAW 264.7 cells. Cell lysates were analyzed by immunoprecipitation with an antibody against GPR81. Acute hepatitis was induced in C56BL/6N mice by administration of lipopolysaccharide and D-galactosamine. Acute pancreatitis was induced by administration of lipopolysaccharide and cerulein. Some mice were given intraperitoneal injections of sodium lactate or small interfering RNA against Gpr81. Activation of NF-κB in tissue macrophages was assessed in mice that expressed a reporter transgene. In macrophages and monocytes, increasing concentrations of lactate reduced TLR4-mediated induction of Il1B, Nlrp3, and Casp1; activation of NF-κB; release of IL1β; and cleavage of CASP1. GPR81 and ARRB2 physically interacted and were required for these effects. The administration of lactate reduced inflammation and organ injury in mice with immune hepatitis; this reduction required Gpr81 dependence invivo. Lactate also prevented activation of NF-κB in macrophages of mice, and, when given after injury, reduced the severity of acute pancreatitis and acute liver injury. Lactate negatively regulates TLR induction of the NLRP3 inflammasome and production of IL1β, via ARRB2 and GPR81. Lactate could be a promising immunomodulatory therapy for patients with acute organ injury.

G-CSF regulates hematopoietic stem cell activity, in part, through activation of Toll-like receptor signaling

Recent studies demonstrate that inflammatory signals regulate hematopoietic stem cells (HSCs). Granulocyte-colony stimulating factor (G-CSF) is often induced with infection and plays a key role in the stress granulopoiesis response. However, its effects on HSCs are less clear. Herein, we show that treatment with G-CSF induces expansion and increased quiescence of phenotypic HSCs, but causes a marked, cell-autonomous HSC repopulating defect associated with induction of toll-like receptor (TLR) expression and signaling. The G-CSF-mediated expansion of HSCs is reduced in mice lacking TLR2, TLR4 or the TLR signaling adaptor MyD88. Induction of HSC quiescence is abrogated in mice lacking MyD88 or in mice treated with antibiotics to suppress intestinal flora. Finally, loss of TLR4 or germ free conditions mitigates the G-CSF-mediated HSC repopulating defect. These data suggest that low level TLR agonist production by commensal flora contributes to the regulation of HSC function and that G-CSF negatively regulates HSCs, in part, by enhancing TLR signaling.

Pharmacologic Suppression of Inflammation by a Diphenyldifluoroketone, EF24, in a Rat Model of Fixed-Volume Hemorrhage Improves Survival

An exaggerated release of proinflammatory cytokines and accompanying inflammation contributes to the development of multiple organ failure after hemorrhagic shock. Here, we tested the nuclear factor (NF) κ-light-chain-enhancer of activated B cell (NF-κB)-mediated transcriptional control of inflammatory pathways as a target in the management of hemorrhage-induced inflammation. We performed a study in a rat model of fixed-volume hemorrhage to investigate the anti-inflammatory effects of the diphenyldifluoroketone EF24 [3,5-bis(2-fluorobenzylidene)piperidin-4-one], an NF-κB inhibitor, in lung tissue. EF24 treatment (0.4 mg/kg) significantly prevented the upregulation of inflammatory biomarkers in rats subjected to 50% hemorrhage and preserved the pulmonary histology in hemorrhaged rats. The lung tissue from treated rats showed marked suppression of the hemorrhage-mediated induction of Toll-like receptor 4, phospho-p65 NF-κB, inducible nitric-oxide synthase, heme oxygenase-1, and cyclooxygenase-2 (COX-2). The hemorrhage-induced COX-2 activity was also significantly inhibited by the EF24 treatment. At the same time, EF24 induced nuclear factor (erythroid-derived 2)-like 2-mediated protective mechanisms against oxidative stress. EF24 also reduced hemorrhage-induced lung myeloperoxidase activity. The plasma levels of proinflammatory tumor necrosis factor-α, interleukin (IL)-6, IL-1α, and IL-1β were lower in EF24-treated rats than in untreated rats. Moreover, there was a significant reduction in the pulmonary expression of high-mobility group B1 protein. These biochemical effects were accompanied by a significant improvement in the survival of rats administered with EF24 as compared with the rats receiving vehicle control (P < 0.05). Overall, the results suggest that EF24 attenuates hemorrhage-induced inflammation and could serve as a salutary anti-inflammatory agent in resuscitation strategies.

Induction of Toll-like receptor 4 signaling in avian macrophages inhibits infectious laryngotracheitis virus replication in a nitric oxide dependent way

LPS is one of the pathogen associated molecular patterns that activates Toll-like receptor 4 (TLR4) signaling pathway eliciting antiviral host responses in mammals although information on such responses in avian species is scarce. Our objectives were to characterize the LPS induced innate responses particularly the expression of LPS receptors (TLR4, CD14) in avian macrophages and observe whether TLR4 mediated induction of NO can elicit antiviral response against infectious laryngotracheitis virus (ILTV) replication. We found that LPS was capable of inducing the expression of TLR4, CD14 and NO production but not the type 1 interferons in an avian macrophage cell line, MQ-NCSU. We also showed that TLR4 mediated NO production can lead to antiviral response against ILTV replication when MQ-NCSU cells were treated with LPS and the resultant supernatant was then transferred to ILTV replicating cells to assess antiviral activity. Antiviral activity of NO was blocked by a selective inhibitor, S-methylisothiourea sulfhate that inhibits inducible NO synthase. This observation confirms that the antiviral activity is positively correlated with NO production. The data show that LPS can be a potential innate immune stimulant that can be used against ILTV infection in chickens that require further evaluation in vivo.

Augmented hepatic Toll‐like receptors by fatty acids trigger the pro‐inflammatory state of non‐alcoholic fatty liver disease in mice

There is considerable evidence that intestinal microbiota are involved in the development of metabolic syndromes and, consequently, with the development of non-alcoholic fatty liver disease (NAFLD). Toll-like receptors (TLRs) are essential for the recognition of microbiota. However, the induction mechanism of TLR signals through the gut-liver axis for triggering the development of non-alcoholic steatohepatitis (NASH) or NAFLD remains unclear. In this study, we investigated the role of palmitic acid (PA) in triggering the development of a pro-inflammatory state of NAFLD. Non-alcoholic fatty liver disease was induced in mice fed a high fat diet (HFD). The mice were killed and the expression of TLRs, tumor necrosis factor (TNF), interleukin (IL)-1β, and phospho-interleukin-1 receptor-associated kinase 1 in the liver and small intestine were assessed. In addition, primary hepatocytes and Kupffer cells were treated with PA, and the direct effects of PA on TLRs induction by these cells were evaluated. The expression of inflammatory cytokines such as TNF, IL-1β, and TLR-2, -4, -5, and -9 was increased in the liver, but decreased in the small intestine of HFD-fed mice in vivo. In addition, the expression of TLRs in primary hepatocytes and Kupffer cells was increased by treatment with PA. In the development of the pro-inflammatory state of NAFLD, PA triggers the expression of TLRs, which contribute to the induction of inflammatory cytokines through TLR signals by intestinal microbiota.

Induction of Toll‐Like Receptor Expression by Porphyromonas gingivalis

Toll-like receptors (TLRs) play pivotal roles in host immune responses and have been suggested to be involved in the development of many infectious diseases. In this study, the mRNA expression levels of TLR2, TLR4, and TLR9 and their relationship with periodontopathic bacteria in periodontal tissue are examined. Furthermore, the mechanism of TLR induction by Porphyromonas gingivalis is investigated in human gingival fibroblasts (HGFs). Gingival tissue and subgingival plaque samples were collected from 19 patients with chronic periodontitis (CP) and 16 control individuals without periodontitis. Gene expression levels in the tissues and in HGFs were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). The numbers of periodontopathic bacteria were determined by quantitative real-time PCR. The expression levels of TLR2 and TLR9 were significantly higher in the tissues of patients with CP compared to the tissues of control individuals. The mRNA levels of TLR2 and TLR9, but not TLR4, were positively correlated with the number of P. gingivalis in subgingival plaque. P. gingivalis sonicated extract, P. gingivalis lipopolysaccharide, P. gingivalis DNA, and tumor necrosis factor-α(TNF-α) could significantly upregulate the mRNA expression of TLR2 in HGFs. Furthermore, P. gingivalis-mediated TLR2 expression was suppressed by TNF-α antibody. This study suggests that P. gingivalis infection induces TLR2 and TLR9 upregulation in patients with CP. P. gingivalis-induced TLR2 expression in HGFs is partially dependent on TNF-α and may lead to sensitization of HGFs to bacterial components encountered in the periodontal microenvironment.

Tumor progression locus 2/Cot is required for activation of extracellular regulated kinase in liver injury and toll-like receptor–induced TIMP-1 gene transcription in hepatic stellate cells in mice

Toll-like receptors (TLRs) function as key regulators of liver fibrosis and are able to modulate the fibrogenic actions of nonparenchymal liver cells. The fibrogenic signaling events downstream of TLRs on Kupffer cells (KCs) and hepatic stellate cells (HSCs) are poorly defined. Here, we describe the MAP3K tumor progression locus 2 (Tpl2) as being important for the activation of extracellular regulated kinase (ERK) signaling in KCs and HSCs responding to stimulation of TLR4 and TLR9. KCs lacking Tpl2 display defects with TLR induction of cytokines interleukin (IL)-1β, IL-10, and IL-23. tpl2(-/-) HSCs were unable to increase expression of fibrogenic genes IL-1β and tissue inhibitor of metalloproteinase 1 (TIMP-1), with the latter being the result of defective stimulation of TIMP-1 promoter activity by TLRs. To determine the in vivo relevance of Tpl2 signaling in liver fibrosis, we compared the fibrogenic responses of wild-type (WT) and tpl2(-/-) mice in three distinct models of chronic liver injury. In the carbon tetrachloride and methionine-choline-deficient diet models, we observed a significant reduction in fibrosis in mice lacking Tpl2, compared to WT controls. However, in the bile duct ligation model, there was no effect of tpl2 deletion, which may reflect a lesser role for HSCs in wounding response to biliary injury. We conclude that Tpl2 is an important signal transducer for TLR activation of gene expression in KCs and HSCs by the ERK pathway and that suppression of its catalytic activity may be a route toward suppressing fibrosis caused by hepatocellular injuries. (HEPATOLOGY 2013).

High Fat Diet-Induced Gut Microbiota Exacerbates Inflammation and Obesity in Mice via the TLR4 Signaling Pathway

Background & AimsWhile it is widely accepted that obesity is associated with low-grade systemic inflammation, the molecular origin of the inflammation remains unknown. Here, we investigated the effect of endotoxin-induced inflammation via TLR4 signaling pathway at both systemic and intestinal levels in response to a high-fat diet.MethodsC57BL/6J and TLR4-deficient C57BL/10ScNJ mice were maintained on a low-fat (10 kcal % fat) diet (LFD) or a high–fat (60 kcal % fat) diet (HFD) for 8 weeks.ResultsHFD induced macrophage infiltration and inflammation in the adipose tissue, as well as an increase in the circulating proinflammatory cytokines. HFD increased both plasma and fecal endotoxin levels and resulted in dysregulation of the gut microbiota by increasing the Firmicutes to Bacteriodetes ratio. HFD induced the growth of Enterobecteriaceae and the production of endotoxin in vitro. Furthermore, HFD induced colonic inflammation, including the increased expression of proinflammatory cytokines, the induction of Toll-like receptor 4 (TLR4), iNOS, COX-2, and the activation of NF-κB in the colon. HFD reduced the expression of tight junction-associated proteins claudin-1 and occludin in the colon. HFD mice demonstrated higher levels of Akt and FOXO3 phosphorylation in the colon compared to the LFD mice. While the body weight of HFD-fed mice was significantly increased in both TLR4-deficient and wild type mice, the epididymal fat weight and plasma endotoxin level of HFD-fed TLR4-deficient mice were 69% and 18% of HFD-fed wild type mice, respectively. Furthermore, HFD did not increase the proinflammatory cytokine levels in TLR4-deficient mice.ConclusionsHFD induces inflammation by increasing endotoxin levels in the intestinal lumen as well as in the plasma by altering the gut microbiota composition and increasing its intestinal permeability through the induction of TLR4, thereby accelerating obesity.

Sterol Intermediates of Cholesterol Biosynthesis Inhibit Hair Growth and Trigger an Innate Immune Response in Cicatricial Alopecia

Primary cicatricial alopecia (PCA) is a group of inflammatory hair disorders that cause scarring and permanent hair loss. Previous studies have implicated PPARγ, a transcription factor that integrates lipogenic and inflammatory signals, in the pathogenesis of PCA. However, it is unknown what triggers the inflammatory response in these disorders, whether the inflammation is a primary or secondary event in disease pathogenesis, and whether the inflammatory reaction reflects an autoimmune process. In this paper, we show that the cholesterol biosynthetic pathway is impaired in the skin and hair follicles of PCA patients. Treatment of hair follicle cells with BM15766, a cholesterol biosynthesis inhibitor, or 7-dehydrocholesterol (7-DHC), a sterol precursor, stimulates the expression of pro-inflammatory chemokine genes. Painting of mouse skin with 7-DHC or BM15766 inhibits hair growth, causes follicular plugging and induces the infiltration of inflammatory cells into the interfollicular dermis. Our results demonstrate that cholesterologenic changes within hair follicle cells trigger an innate immune response that is associated with the induction of toll-like receptor (TLR) and interferon (IFN) gene expression, and the recruitment of macrophages that surround the hair follicles and initiate their destruction. These findings reveal a previously unsuspected role for cholesterol precursors in PCA pathogenesis and identify a novel link between sterols and inflammation that may prove transformative in the diagnosis and treatment of these disorders.

Alpha-Melanocyte Stimulating Hormone Preserves Islet Graft Survival Through Down-Regulation of Toll-Like Receptors

The induction of Toll-like receptors (TLRs) in β cells is involved in β-cell death and graft rejection after transplantation. This study investigated the ability of alpha-melanocyte stimulating hormone (α-MSH) to protect pancreatic islets and improve graft survival through regulation of TLRs. To test the effect of α-MSH on TLR regulation, we first isolated pancreatic islets from rats pretreated with/without α-MSH and assayed inflammatory cytokines and insulin release, and measured the expression of TLRs. Pancreatic islets were transplanted into the kidney capsule of a diabetes mellitus (DM) mouse with and without prior injection of α-MSH. The blood glucose levels were measured and TLR4 expression in transplanted kidney tissue was assessed. Islet morphology, including size and total mass, was improved in the α-MSH group compared to the control group. The expression of TLRs as well as nitric oxide and monocyte chemoattractant protein 1 production were decreased in islets isolated from α-MSH–treated rats. In DM mice, the normoglycemic ratio was higher in the α-MSH–treated group than in the sham group. Moreover, the high levels of TLR4 expression observed in DM kidney tissue were significantly decreased in islet-transplanted tissue with α-MSH. This study showed that α-MSH protects pancreatic islets from cell death and dysfunction through downregulation of TLRs. In conclusion, α-MSH could contribute to improved islet graft survival and function in pancreatic islet transplantation.

Deficiencies In Toll Like Receptors 4 Impair The Functional Recovery In A Mouse Model Of Spinal Cord Trauma

Toll‐like receptors (TLRs) are a family of molecules that play a critical role in innate immunity. TLRs can be activated by heat shock proteins (HSPs), necrotic cells, extracellular matrix, and oxidized lipids – all of which are found in the injured spinal cord. Here, using a model of spinal cord injury, we show dramatic induction of TLRs and molecules associated with TLR signaling. The injury was produced by extradural compression of the spinal cord using an aneurysm clip with a closing force of 24 g to the dura via a four‐level T5‐T8 laminectomy in mice. Our present data show that the deficiency of TLR4 in C57BL/10ScN mice (TLR4 knockout) significantly impairs the normal progression of SCI/repair and functional recovery compared with wild type (WT) animals. We evaluated the following points: histological damage, motor recovery, nuclear Factor‐κB expression, cytokines production, inducible nitric oxide synthase (iNOS) expression, neutrophil infiltration, glial fibrillary acidic protein (GFAP) and CD11β expression, nitrotyrosine and poly‐ADP‐ribose (PAR) formation, glial cell‐derived neurotrophic factor (GDNF) expression, apoptosis (Caspase 3, Bax and Bcl‐2 expression). Capable of both protective and pathological roles, TLR4 can be helpful or harmful under varying neurological conditions. When and why TLR4 initiates beneficial outcome is still largely unknown, but progress in the field suggests that this question will receive much attention, as TLR4 is a useful and druggable target.

Induction of toll-like receptor (TLR) 2, and MyD88-dependent TLR- signaling in response to ligand stimulation and bacterial infections in the Indian major carp, mrigal (Cirrhinus mrigala)

Toll-like receptor 2 (TLR2) is a member of TLR family. It recognizes a wide range of bacteria and their products, and is involved in inducing innate immune responses. In this article, we reported inductive expression of TLR2 and myeloid differentiation primary response gene 88 (MyD88)-dependent signaling in the Indian major carp, mrigal (Cirrhinus mrigala) which is highly commercially important fish species in the Indian subcontinent. Ontogeny analysis of TLR2, MyD88 and TRAF6 (TNF receptor associated factor 6) genes by quantitative real-time PCR (qRT-PCR) revealed constitutive expression of these genes in all embryonic developmental stages, indicating their involvement in embryonic innate immune defense system in fish. Tissue specific expression analysis of these genes by qRT-PCR showed their wide distribution in various organs and tissues. Highest expression of TLR2 was in gill, MyD88 in liver and TRAF6 was in kidney. Inductive expression of TLR2, MyD88 and TRAF6 genes were observed following peptidoglycan (PGN)-treatment, and Streptococcus uberis and Aeromonas hydrophila infections. Expression of interleukin (IL)-8 and TNF-α in various organs were significantly enhanced by PGN-treatment and bacterial infections, and were closely associated with TLR2 induction. These findings together highlighted the contribution of TLR2 in augmenting innate immunity in fish, and indicated it's important role in immune surveillance of various organs during pathogenic invasion. This study will enrich the information in understanding the innate immune mechanism in fish, and will be helpful in developing preventive measures against infectious diseases in fish.

Abstract P209: MyD88 Mediated Inflammatory Signaling Involved in Post-MI CaMKII Activation and Cardiac Hypertrophy

Toll-like receptors (TLRs) and calmodulin kinase II (CaMKII) participate in pathological responses to myocardial infarction (MI), including activation of nuclear factor kappa B (NF-κB) transcription and complement factor B expression (Singh JCI 2009). We tested the effect of MyD88 deficiency on the adverse cardiac outcome after MI. MyD88 knock out (MyD88-/-) hearts had significantly reduced hypertrophy, hypertrophic gene expression, and inflammatory gene expression compared to WT control hearts. Cultured cardiomyocytes from expressing MyD88 and a NF-κB reporter demonstrated robust induction of NF-κB upon TLR induction by bacterial lipopolysaccharides (LPS). In contrast, cardiomyocytes from MyD88-/- mice failed to induce NF-κB upon LPS treatment. An alternative MyD88-independent pathway for NF-κB remained functional in WT and MyD88-/- cells. Both LPS treatment and MI increased a Ca2+/calmodulin independet activated form of CaMKII (CaMKIIox) in cultured cardiomyocytes and mouse hearts, respectively. MyD88-/- hearts, however, did not display increased CaMKIIox upon MI. Taken together, we interpret these results to show that: (1) MyD88 participates in MI mediated hypertrophic and inflammatory gene expression, (2) CaMKII enhances NF-kB activation in cardiomyocytes by a MyD88-dependent mechanism, and (3) CaMKIIox plays a role in MyD88 dependent signaling to induce expression of proinflammatory genes in post-MI hearts.

Toll-like receptors 1–9 are elevated in livers with fructose-induced hepatic steatosis

Studies in animals and human subjects indicate that gut-derived bacterial endotoxins may play a critical role in the development of non-alcoholic fatty liver disease (NAFLD). In the present study, we investigated if the liver is also sensitised by other microbial components during the onset of fructose-induced steatosis in a mouse model. C57BL/6 mice were either fed with 30 % fructose solution or tap water (control) with or without antibiotics for 8 weeks. Expression of toll-like receptors (TLR)1-9, TNF-α, inducible NO synthase (iNOS), myeloid differentiation factor 88 (MyD88) and number of F4/80 positive cells in the liver were assessed. Occludin protein, DNA of microbiota in the small and large intestine and retinol binding protein 4 (RBP4) in plasma were analysed using Western blot, DNA fingerprinting and ELISA, respectively. F4/80 positive cells were determined by immunohistochemistry. The accumulation of TAG found in the livers of fructose-fed mice was associated with a significant induction of TLR 1-4 and 6-8. Plasma RBP4 concentration and hepatic mRNA expression levels of TNF-α, iNOS, MyD88 and number of F4/80 positive cells of fructose-fed animals were significantly higher than those of controls; however, these effects of fructose were attenuated in antibiotic-treated mice. Whereas protein concentration of occludin was lower in the duodenum of fructose-treated mice, no systematic alterations of microbiota were found in this part of the intestine. Taken together, these data support the hypothesis that (1) an increased intestinal translocation of microbial components and (2) an increased number of F4/80 positive cells and induction of several TLR and dependent pathways (e.g. MyD88 and iNOS) may be involved in the onset of fructose-induced NAFLD.

Mechanisms of Sperm Storage in the Female Reproductive Tract: an Interspecies Comparison

Once semen has been collected for artificial insemination, it is diluted into extenders designed to prevent its deterioration over the period prior to insemination. If the semen is not frozen, the extenders provide protection for a period of a few hours to a few days, depending on species. Despite the efforts of biotechnologists to increase the duration of storage without compromising fertility, there has been relatively little progress for many years. However, comparative studies in diverse species have revealed that long-term sperm storage (up to months and years) within the female reproductive tract is relatively commonplace in reptiles, fishes, birds and amphibians. Even among mammals, some species of bat have evolved mechanisms for storing spermatozoa for several months in the uterus or oviduct so that they can mate in the autumn but postpone fertilization until the spring. We currently know little about the mechanisms that support such long-term sperm storage, mainly because evidence from such species is either absent or fragmentary. Nevertheless, parallels between mammalian and other systems, where spermatozoa are sequestered in sperm storage tubules, suggest that the enclosure of spermatozoa within pockets of epithelial cells may be sufficient to achieve long-term sperm storage. In addition, recent evidence from sperm-storing bats has suggested an alternative, or additional, hypothesis that the modulation of apoptosis within epithelial cells is important in controlling sperm survival. Despite a lack of direct experimental evidence from a wide variety of species, I propose that there is now enough evidence to warrant investigation of these hypotheses.

MyD88 Deficiency Ameliorates β-Amyloidosis in an Animal Model of Alzheimer's Disease

The accumulation of β-amyloid protein (Aβ) in the brain is thought to be a primary etiologic event in Alzheimer's disease (AD). Fibrillar Aβ plaques, a hallmark of AD abnormality, are closely associated with activated microglia. Activated microglia have contradictory roles in the pathogenesis of AD, being either neuroprotective (by clearing harmful Aβ and repairing damaged tissues) or neurotoxic (by producing proinflammatory cytokines and reactive oxygen species). Aβ aggregates can activate microglia by interacting with multiple toll-like receptors (TLRs), the pattern-recognition receptors of the innate immune system. Because the adapter protein MyD88 is essential for the downstream signaling of all TLRs, except TLR3, we investigated the effects of MyD88 deficiency (MyD88(-/-)) on Aβ accumulation and microglial activation in an AD mouse model. MyD88 deficiency decreased Aβ load and microglial activation in the brain. The decrease in Aβ load in an MyD88(-/-) AD mouse model was associated with increased and decreased protein expression of apolipoprotein E (apoE) and CX3CR1, respectively, compared with that in an MyD88 wild-type AD mouse model. These results suggest that MyD88 deficiency may reduce Aβ load by enhancing the phagocytic capability of microglia through fractalkine (the ligand of CX3CR1) signaling and by promoting apoE-mediated clearance of Aβ from the brain. These findings also suggest that chronic inflammatory responses induced by Aβ accumulation via the MyD88-dependent signaling pathway exacerbate β-amyloidosis in AD.

Induction of toll-like receptor 2 positive antigen-presenting cells in spleen of pristane-induced arthritis in rats

Toll-like receptors (TLRs) have been found to contribute to the pathogenesis of rheumatoid arthritis (RA). The aim of this study is to investigate the regulation and potential role of TLR2 in spleen of pristane-induced arthritis (PIA) rat, which can be used to further understand the mechanisms of RA. Arthritis in DA rats was induced by pristane. TLR2 expression in spleen was detected by real-time quantitative PCR and western blotting, and TLR2 expression at both mRNA and protein levels was upregulated in PIA rats. Peptidoglycan (PGN) was systemically administrated to PIA rats, and arthritis severity was evaluated macroscopically and microscopically. Results showed that systemic administration of PGN to PIA rats obviously deteriorated arthritis severity. TLR2 expression on splenocytes and different types of immune cells was measured by flow cytometry. And it was found that TLR2 was mainly expressed on antigen-presenting cells (APCs) of spleen, and the proportion of TLR2(+) dendritic cells and macrophages in spleen of PIA rats was increased remarkably. Thus, we conclude that the induction of TLR2(+) APCs in spleen may participate in the maintenance of PIA.