• IMMUNE NETWORK
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• 제13권5호
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• pp.194-198
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• 2013

Recent papers have shown that the initial event in the pathogenesis of autoimmune type 1 diabetes (T1D) comprises sensing of molecular patterns released from apoptotic ${\beta}$-cells by innate immune receptors such as toll-like receptor (TLR). We have reported that apoptotic ${\beta}$-cells undergoing secondary necrosis called 'late apoptotic' ${\beta}$-cells stimulate dendritic cells (DCs) and induce diabetogenic T cell priming through TLR2. The role of other innate immune receptors such as TLR7 or TLR9 in the initiation of T1D has also been suggested. We hypothesized that TLR2 blockade could inhibit T1D at the initial step of T1D. Indeed, when a TLR2 agonist, $Pam3CSK_4$ was administered chronically, the development of T1D in nonobese diabetic (NOD) mice was inhibited. Diabetogenic T cell priming by DCs was attenuated by chronic treatment with $Pam3CSK_4$, indicating DC tolerance. For the treatment of established T1D, immune tolerance alone is not enough because ${\beta}$-cell mass is critically reduced. We employed TLR2 tolerance in conjunction with islet transplantation, which led to reversal of newly established T1D. Dipeptidyl peptidase 4 (DPP4) inhibitors are a new class of anti-diabetic agents that have beneficial effects on ${\beta}$-cells. We investigated whether a combination of DPP4 inhibition and TLR2 tolerization could reverse newly established T1D without islet transplantation. We could achieve normoglycemia by TLR2 tolerization in combination with DPP4 inhibition but not by TLR2 tolerization or DPP4 inhibition alone. ${\beta}$-cell mass was significantly increased by combined treatment with TLR2 tolerization and DPP4 inhibition. These results suggest the possibility that a novel strategy of TLR tolerization will be available for the inhibition or treatment of established T1D when combined with measures increasing critically reduced ${\beta}$-cell mass of T1D patients such as DPP4 inhibition or stem cell technology.

• Biomolecules & Therapeutics
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• 제17권2호
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• pp.144-150
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• 2009

Reactive oxygen species play a role in signal transduction and in many human diseases. B-cell activating factor (BAFF) plays a role for mature B cell generation and maintenance and for the incidence of autoimmune diseases. We previously reported that BAFF expression was induced by ROS. In this study, we investigated whether ROS-induced BAFF expression was affected by toll-like receptor (TLR) 4 or myeloid differentiation primary response gene (MyD) 88. BAFF expression was increased by serum deprivation that is an experimental modification to produce ROS. In contrast, TLR4 and MyD88 were decreased by serum deprivation. Although ROS production was decreased in TLR4-nonfunctional or MyD88-deficient splenocytes as compared to that in control mice, serum deprivation increased ROS production and augmented BAFF expression in both cells. $50{\mu}M\;H_2O_2$ also increased BAFF expression in TLR4-deficient or MyD88-deficient splenocytes. Collectively, results show that BAFF expression may be mediated by TLR4 or MyD88-independent manner and TLR4 or MyD88 may not be required in BAFF expression.

• Parasites, Hosts and Diseases
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• 제53권4호
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• pp.431-438
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• 2015

In Trichinella spiralis infection, type 2 helper T (Th2) cell-related and regulatory T ($T_{reg}$) cell-related immune responses are the most important immune events. In order to clarify which Toll-like receptors (TLRs) are closely associated with these responses, we analyzed the expression of mouse TLR genes in the small intestine and muscle tissue during T. spiralis infection. In addition, the expression of several chemokine- and cytokine-encoding genes, which are related to Th2 and $T_{reg}$ cell mediated immune responses, were analyzed in mouse embryonic fibroblasts (MEFs) isolated from myeloid differentiation factor 88 (MyD88)/TIR-associated proteins (TIRAP) and Toll receptor-associated activator of interferons (TRIF) adapter protein deficient and wild type (WT) mice. The results showed significantly increased TLR4 and TLR9 gene expression in the small intestine after 2 weeks of T. spiralis infection. In the muscle, TLR1, TLR2, TLR5, and TLR9 gene expression significantly increased after 4 weeks of infection. Only the expression of the TLR4 and TLR9 genes was significantly elevated in WT MEF cells after treatment with excretory-secretory (ES) proteins. Gene expression for Th2 chemokine genes were highly enhanced by ES proteins in WT MEF cells, while this elevation was slightly reduced in MyD88/$TIRAP^{-/-}$ MEF cells, and quite substantially decreased in $TRIF^{-/-}$ MEF cells. In contrast, IL-10 and $TGF-{\beta}$ expression levels were not elevated in MyD88/$TIRAP^{-/-}$ MEF cells. In conclusion, we suggest that TLR4 and TLR9 might be closely linked to Th2 cell and $T_{reg}$ cell mediated immune responses, although additional data are needed to convincingly prove this observation.

• Asian Pacific Journal of Cancer Prevention
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• 제13권6호
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• pp.2807-2812
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• 2012

The purpose of this study was to examine the effect of a Toll-like receptor 5 (TLR5) agonist, CBLB502, on the growth and radiosensitivity of A549 lung cancer cells in vivo. Expression of myeloid differentiation factor 88 (MyD88) or TLR5 was stably knocked down in human lung cancer cells (A549) using lentivirus expressing short hairpin RNA targeting human MyD88 or TLR5. Lack of MyD88 or TLR5 expression enhanced tumor growth in mouse xenografts of A549 lung cancer cells. CBLB502 inhibited the growth of A549 lung cancer cells, not A549-MyD88-KD cells in vivo in the murine xenograft model. Our results showed that the inhibition of A549 by CBLB502 in vivo was realized through regulating the expression of neutrophil recruiting cytokines and neutrophil infiltration. Finally, we found that activation of TLR5 signaling did not affect the radiosensitivity of tumors in vivo.

• BMB Reports
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• 제49권6호
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• pp.305-310
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• 2016

Toll-like receptors (TLRs) play a critical role in the innate immune response against pathogens. Each TLR recognizes specific pathogen-associated molecular patterns, after which they activate the adaptor protein MyD88 or TRIF-assembled signaling complex to produce immune mediators, including inflammatory cytokines and type I IFNs. Although the activation of TLR is important for host defense, its uncontrolled activation can damage the host. During the past decade, numerous studies have demonstrated that GSK3β is a key regulator of inflammatory cytokine production in MyD88-mediated TLR signaling via TLR2 and TLR4. Recently, GSK3β has also been implicated in the TRIF-dependent signaling pathway via TLR3. In this review, we describe current advances on the regulatory role of GSK3β in immune responses associated with various TLRs. A better understanding of the role of GSK3β in TLR signaling might lead to more effective anti-inflammatory interventions.

• 미생물학회지
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• 제43권3호
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• pp.166-172
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• 2007

ERM 단백질은 23S rRNA의 A2058에 methylation시킴으로써 macrolide-lincosamide- streptogramin B $(MLS_B)$계 항생제의 부착을 저해하여 항생제의 활성을 억제하는 내성 인자 단백질로 monomethylase와 dimethylase로 나누어진다. Dimethylase와 비교되는 monomethylase의 특성을 밝히기 위해 dimethylase (ErmSF)와 monomethylase (TlrD)를 동시에 보유한 Streptomyces fradiae에서 tlrD를 클론하고 대장균에서 최초로 대략생산을 시도하여 $37^{\circ}C$에서 세포내 전체 단백질의 55%를 차지할 정도로 대량생산된 불용성 단백질을 얻어내었다. 그러나 ErmSF와는 달리 낮은 온도에서 대량생산된 단백질이 용해성 단백질로 전환되지 않고 불용성 단백질로 남아있었다. Thioredoxin과 샤페론인 GroESL은 모두 ErmSF의 경우와 마찬가지로 용해성 단백질로의 전환에 도움을 주지 않았다. 이러한 차이점은 천재까지 전혀 밝혀지지 않은 단백질내의 구조적 특성에 의한 monomethylase와 dimethylase의 차이점을 밝힐 수 있다는 가능성을 말해주는 것으로 추정된다. 그러나 ErmSF의 경우와 동일하게 SDS-PAGE에서 검색되지 않은 미량의 발현된 용해성 단백질이 TlrD를 함유한 세포에 항생제에 대한 내성을 나타내게 하였고 이렇게 발현된 내성은 monomethylase에 의한 내성에서 기대되는 내성과 일치하였다.

• 한국식품과학회지
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• 제39권2호
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• pp.175-180
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• 2007

TLRs는 병원균이 숙주의 몸 속에 들어 왔을 때, 병원균들이 가지고 있는 독특한 구조를 인식하여 선천성 면역반응과 뒤이어 후천성 면역반응을 유도하는 중요한 역할을 한다. 우리는 이번 실험을 통하여 curcumin이 선행연구에서 밝혀낸 TLR4 뿐만 아니라 TLR2와 TLR6 그리고 TLR3를 또한 분자학적인 타깃으로 할 수 있다는 것을 알아내었다. Curcumin이 MALP-2(TLR2,6 agonist)에 의해서 유도된 IRAK-1 degradation을 억제시켰다. 이러한 결과는 curcumin의 분자학적인 타깃이 IRAK-1위에 놓여 있으며, TLR2와 TLR6가 될 것이라는 가능성을 제시해 준다고 할 수 있다. 또한 curcumin은 viral 자극제인 poly[I:C](TLR3 agonist)에 의해서 유도된 IRF3나 $NF-{\kappa}B$ 활성화를 억제하였지만, TRIF에 의해서 유도된 IRF3 활성화는 억제시키지를 못하였다. 이러한 결과 또한 TLR3 자체가 curcumin의 분자학적인 타깃이라는 가능성을 제시해 준다고 할 수 있겠다. 이러한 결과를 종합해 볼때, curcumin의 분자학적인 타깃이 $IKK{\beta}$ 이외에 모든 TLRs가 될 수 있다는 가능성을 제시해 준다고 할 수 있겠다. 이러한 결과는 curcumin이 그람음성균 뿐만이 아니라 바이러스나 박테리아 등 여러 병원균들로부터 유도되는 염증반응이나 만성적인 질병들을 조절할 수 있다는 것을 보여주는 결과라 할 수 있겠다.

• Journal of Microbiology and Biotechnology
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• 제33권9호
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• pp.1213-1227
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• 2023

Fetal growth restriction (FGR) is a prevalent obstetric condition. This study aimed to investigate the role of Toll-like receptor 9 (TLR9) in regulating the inflammatory response and gut microbiota structure in FGR. An FGR animal model was established in rats, and ODN1668 and hydroxychloroquine (HCQ) were administered. Changes in gut microbiota structure were assessed using 16S rRNA sequencing, and fecal microbiota transplantation (FMT) was conducted. HTR-8/Svneo cells were treated with ODN1668 and HCQ to evaluate cell growth. Histopathological analysis was performed, and relative factor levels were measured. The results showed that FGR rats exhibited elevated levels of TLR9 and myeloid differentiating primary response gene 88 (MyD88). In vitro experiments demonstrated that TLR9 inhibited trophoblast cell proliferation and invasion. TLR9 upregulated lipopolysaccharide (LPS), LPS-binding protein (LBP), interleukin (IL)-1β and tumor necrosis factor (TNF)-α while downregulating IL-10. TLR9 activated the TARF3-TBK1-IRF3 signaling pathway. In vivo experiments showed HCQ reduced inflammation in FGR rats, and the relative cytokine expression followed a similar trend to that observed in vitro. TLR9 stimulated neutrophil activation. HCQ in FGR rats resulted in changes in the abundance of Eubacterium_coprostanoligenes_group at the family level and the abundance of Eubacterium_coprostanoligenes_group and Bacteroides at the genus level. TLR9 and associated inflammatory factors were correlated with Bacteroides, Prevotella, Streptococcus, and Prevotellaceae_Ga6A1_group. FMT from FGR rats interfered with the therapeutic effects of HCQ. In conclusion, our findings suggest that TLR9 regulates the inflammatory response and gut microbiota structure in FGR, providing new insights into the pathogenesis of FGR and suggesting potential therapeutic interventions.

• Clinical and Experimental Reproductive Medicine
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• 제50권1호
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• pp.44-52
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• 2023

Objective: The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. Methods: Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. Results: The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1β, IL-6, IL-12, interferon α (IFN-α), IFN-β, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. Conclusion: The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

• Molecules and Cells
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• 제38권1호
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• pp.26-32
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• 2015

Toll-like receptors (TLR) 7 and 9 transduce a cellular signal through the MyD88-dependent pathway and induce the production of inflammatory mediators against microbial nucleotide components. The repeated stimulation of TLR4 leads to endotoxin tolerance, but the molecular mechanisms of tolerance induced through the costimulation of individual TLR has not yet been established, although endosomal TLRs share signaling pathways with TLR4. In the present study, mouse macrophages were simultaneously stimulated with the TLR7 agonist, gardiquimod (GDQ), and the TLR9 agonist, CpG ODN 1826, to examine the mechanism and effector functions of macrophage tolerance. Compared with individual stimulation, the costimulation of both TLRs reduced the secretion of TNF-${\alpha}$ and IL-6 through the delayed activation of the NF-${\kappa}B$ pathway; notably, IL-10 remained unchanged in costimulated macrophages. This tolerance reflected the early induction of suppressor of cytokine signaling-1 (SOCS-1), according to the detection of elevated TNF-${\alpha}$ secretion and restored NF-${\kappa}B$ signaling in response to the siRNA-mediated abrogation of SOCS-1 signaling. In addition, the restimulation of each TLRs using the same ligand significantly reduced the expression of both TLRs in endosomes. These findings revealed that the costimulation of TLR7 and TLR9 induced macrophage tolerance via SOCS-1, and the restimulation of each receptor or both TLR7 and TLR9 downregulated TLR expression through a negative feedback mechanisms that protects the host from excessive inflammatory responses. Moreover, the insufficient and impaired immune response in chronic viral infection might also reflect the repeated and simultaneous stimulation of those endosomal TLRs.