Regulatory Role of rno-miR-30b-5p in IL-10 and Toll-like Receptor 4 Expressions of T Lymphocytes in Experimental Autoimmune Uveitis In Vitro

Uveitis is a serious eye disease that usually damages young adult’s health. MicroRNAs (miRNAs) are a class of small noncoding RNAs which regulate messenger RNA (mRNA) expression. It is predicted that rno-miR-30b-5p can regulate the expressions of interleukin-10 (IL-10) and Toll-like receptor 4 (TLR4). In this study, the regulatory role of rno-miR-30b-5p in IL-10 and TLR4 gene expressions was validated using luciferase activity assay. Further, the inflammatory manifestation of the anterior segment and pathological examination of the eye were explored in experimental autoimmune uveitis (EAU) rats. Meanwhile, the levels of rno-miR-30b-5p in eye tissues, spleen, and lymph nodes were measured using quantitative PCR (Q-PCR). IL-10 and TLR4 in spleen and lymph nodes were further separately determined by using Q-PCR and Enzyme-Linked Immunosorbent Assay (ELISA). Moreover, rno-miR-30b-5p mimic and its inhibitor were separately transfected into purified T cells, and the levels of IL-10 and TLR4 were detected using PCR, flow cytometry, and ELISA techniques. Results indicate that rno-miR-30b-5p was downregulated in spleen, lymph nodes, and eye tissues whereas the expressions of IL-10 and TLR4 at mRNA and protein levels were upregulated. The levels of IL-10 and TLR4 were negatively correlated to rno-miR-30b-5p levels. The result of in vitro cell transfection experiment indicates that IL-10 and TLR4 expressions were inhibited at mRNA and protein levels after T cells incubated with rno-miR-30b-5p mimic. However, the IL-10 and TLR4 mRNA levels were upregulated in purified T cells from spleen and lymph nodes after treatment with miR-30b-5p antagonist. In addition, there was no evident change of IL-10 and TLR4 proteins in spleen and lymph node T cells between EAU control and negative treatment groups. Flow cytometry analysis revealed that rno-miR-30b-5p mimic could reduce the number of both IL-10 and TLR4 positive cells, whereas rno-miR-30b-5p inhibitor could increase the number of IL-10 and TLR4 positive cells. Our study demonstrates that rno-miR-30b-5p influences the development of uveitis by regulating the level of IL-10 and TLR4 positive cells, thereby playing a role in the pathogenesis of uveitis.

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Detrimental Role of Heat Shock Protein 60&70 and Toll-like Receptor 4 in Skeletal Muscle Ischaemia In Vitro

Journal of Vascular Surgery ◽

10.1016/j.jvs.2013.02.134 ◽

2013 ◽

Vol 57 (5) ◽

pp. 77S

Author(s):

Ali Navi ◽

Rebekah Yu ◽

Xu Shi-Wen ◽

Sidney Shaw ◽

George Hamilton ◽

...

Keyword(s):

Skeletal Muscle ◽

Heat Shock ◽

Heat Shock Protein ◽

Heat Shock Protein 60 ◽

Toll Like Receptor ◽

Toll Like Receptor 4 ◽

Muscle Ischaemia

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Abstract 1766: Diet-Induced Obesity Causes Inflammation by Activating Toll-Like Receptor 4 Signaling and Downregulates AMPK in Heart

Circulation ◽

10.1161/circ.118.suppl_18.s_385-a ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Hwi Jin Ko ◽

Dae Young Jung ◽

Zhexi Ma ◽

Jason K Kim

Keyword(s):

Glucose Metabolism ◽

Whole Body ◽

Chow Diet ◽

Toll Like Receptor ◽

Toll Like Receptor 4 ◽

Myocardial Glucose Metabolism ◽

Cardiac Inflammation ◽

Protein Levels ◽

Diet Induced Obesity

Increasing evidence implicates the role of inflammation in diabetes and complications. Macrophages are shown to infiltrate adipose tissue in obesity, and inflammatory cytokines alter glucose metabolism in peripheral organs. Male C57BL/6 mice were fed high-fat diet (HFD; 55% fat by calories) or chow diet for 6 weeks, and heart samples were taken for analysis (n = 5~7). Chronic HFD increased whole body fat mass, measured by 1 H-MRS, by 3-fold, and elevated plasma IL-6 and TNF-α levels by 40%. Diet-induced obesity caused inflammation in heart and increased macrophage-specific CD68 levels by 5-fold (Fig. 1) (* P < 0.05 vs Chow). Diet-induced cardiac inflammation was associated with significant increases in toll-like receptor 4 (TLR4) and MyD88 levels in heart (Fig. 2). HFD also increased cardiomyocyte SOCS3 levels by more than 3-fold (Fig. 3). Myocardial glucose metabolism was measured using intravenous injection of 2-[ 14 C]deoxyglucose in awake mice (n = 6). Chronic HFD reduced myocardial glucose uptake by 50%, and this was associated with significant reductions in total GLUT4 and GLUT1 protein levels. Further, Thr 172 phosphorylation of AMPK, a critical regulator of energy balance, was markedly reduced in heart following HFD (Fig. 4). These results demonstrate that diet-induced obesity causes macrophage infiltration and inflammation in heart by increasing TLR4 signaling in cardiomyocytes. Similar to the effects of inflammation on peripheral glucose metabolism, diet-induced cardiac inflammation reduced myocardial glucose metabolism by downregulating AMPK and GLUT protein levels. Thus, our findings underscore an important role of inflammation in diabetic heart.

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Development of a Secondary Immune Response toMycobacterium tuberculosisIs Independent of Toll-Like Receptor 2

Infection and Immunity ◽

10.1128/iai.01076-10 ◽

2010 ◽

Vol 79 (3) ◽

pp. 1118-1123 ◽

Cited By ~ 11

Author(s):

Amanda McBride ◽

Kamlesh Bhatt ◽

Padmini Salgame

Keyword(s):

Immune Response ◽

T Cells ◽

Host Resistance ◽

Toll Like Receptor ◽

Response Phenotype ◽

Toll Like Receptor 2 ◽

Antigen Presenting ◽

Memory Immune Response

ABSTRACTPublished work indicates that the contribution of Toll-like receptor 2 (TLR2) to host resistance during acuteMycobacterium tuberculosisinfection is marginal. However, in these studies, TLR2 participation in the memory immune response toM. tuberculosiswas not determined. The substantialin vitroevidence thatM. tuberculosisstrongly triggers TLR2 on dendritic cells and macrophages to bring about either activation or inhibition of antigen-presenting cell (APC) functions, along with accumulating evidence that memory T cell development can be calibrated by TLR signals, led us to question the role of TLR2 in host resistance to secondary challenge withM. tuberculosis. To address this question, a memory immunity model was employed, and the response of TLR2-deficient (TLR2 knockout [TLR2KO]) mice following a secondary exposure toM. tuberculosiswas compared to that of wild-type (WT) mice based on assessment of the bacterial burden, recall response, phenotype of recruited T cells, and granulomatous response. We found that upon rechallenge withM. tuberculosis, both WT and TLR2KO immune mice displayed similarly enhanced resistance to infection in comparison to their naïve counterparts. The frequencies ofM. tuberculosis-specific gamma interferon (IFN-γ)-producing T cells, the phenotypes of recruited T cells, and the granulomatous responses were also similar between WT and TLR2KO immune mice. Together, the findings from this study indicate that TLR2 signaling does not influence memory immunity toM. tuberculosis.

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TREM2, Driving the Microglial Polarization, Has a TLR4 Sensitivity Profile After Subarachnoid Hemorrhage

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.693342 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yangchun Hu ◽

Chao Li ◽

Xiaojian Wang ◽

Weiwei Chen ◽

Yu Qian ◽

...

Keyword(s):

Subarachnoid Hemorrhage ◽

Neuroprotective Effect ◽

Neurological Dysfunction ◽

Toll Like Receptor ◽

Toll Like Receptor 4 ◽

Microglial Polarization ◽

The Relationship

Increasing evidence suggests that triggering receptor expressed on myeloid cells 2 (TREM2) is implicated in the pathophysiology of neuroinflammation. The aim here was to investigate the neuroprotective role of TREM2 and its regulatory mechanism after subarachnoid hemorrhage (SAH). TREM2 siRNA was administered to measure the detrimental role of TREM2 in mediating microglial polarization in vivo and in vitro after experimental SAH. The relationship between Toll-like receptor 4 (TLR4) signaling and TREM2 was further explored. The soluble TREM2 from the cerebrospinal fluid (CSF) of patients with SAH was detected. The results showed that TREM2 mainly located in the microglia and presented a markedly delayed elevation after SAH. TREM2 knockdown triggered increased pro-inflammatory productions, aggravated microglial activities, and further exacerbated neurological dysfunction after SAH. Significantly, TLR4 knockout increased the expression of TREM2, accompanied by ameliorated neuroinflammation and improved neurological function. Corresponding to different clinical Hunt–Hess grades, obviously enhanced accumulation of soluble TREM2 was detected in the CSF of patients with SAH. TREM2 played a pivotal role in mediating microglial polarization after SAH, and the neuroprotective effect of TREM2 might be potentially suppressed by the hyperactive TLR4 in the early phase of SAH. Pharmacological targeting of TREM2 may be a promising strategy for SAH therapy.

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THE ROLE OF LYMPHOCYTES IN THE SENSITIZATION OF RATS TO RENAL HOMOGRAFTS

Journal of Experimental Medicine ◽

10.1084/jem.122.2.347 ◽

1965 ◽

Vol 122 (2) ◽

pp. 347-360 ◽

Cited By ~ 78

Author(s):

S. Strober ◽

J. L. Gowans

Keyword(s):

Lymph Nodes ◽

Thoracic Duct ◽

Femoral Vein ◽

F1 Hybrid ◽

Duct Cells ◽

Degree Of Sensitization ◽

Spleen And Lymph Nodes

In order to study the role of blood-borne small lymphocytes in the sensitization of rats to renal homografts 2 techniques for the perfusion of isolated rat kidneys were employed: (a) the in vitro perfusion of kidneys with thoracic duct cells suspended in either an artificial medium or in blood; the perfusates were then injected into rats syngeneic with the lymphocyte donors; (b) the in vivo perfusion of kidneys with blood issuing from the femoral artery and returning to the femoral vein of living rats. The degree of sensitization conferred on the recipients by the perfusates was assessed by applying a skin homograft from the kidney donor and scoring the epithelial necrosis at 6 days. The in vitro experiments indicated that parental strain thoracic duct cells, which had passed through an F1 hybrid kidney could confer upon a parental rat sensitivity to an F1 skin graft. Several perfusions with radioactively labelled lymphocytes showed that the injected cells migrated to the lymph nodes and spleen of the recipients Labelled large pyroninophilic cells were occasionally seen in the spleen and lymph nodes of recipients, and it was suggested that these had arisen from the injected cells. Although the in vitro perfusions with blood indicated that renal homografts might sensitize their hosts within 1 hour, the in vivo perfusions suggested that about 5 to 12 hours were required. The more rapid sensitization in vitro was possibly due to the more frequent opportunity for contact between lymphocytes and kidney vascular endothelium which was afforded by the conditions in vitro.

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ACTIVITY OF HOST-DERIVED T CELLS WHICH DIFFERENTIATE IN NUDE MICE GRAFTED WITH CO-ISOGENIC OR ALLOGENEIC THYMUSES

Journal of Experimental Medicine ◽

10.1084/jem.139.5.1215 ◽

1974 ◽

Vol 139 (5) ◽

pp. 1215-1227 ◽

Cited By ~ 50

Author(s):

Berenice Kindred ◽

Francis Loor

Keyword(s):

T Cells ◽

T Cell ◽

Lymph Nodes ◽

Nude Mice ◽

Precursor Cells ◽

Donor Strain ◽

Skin Grafts ◽

Host Type ◽

Spleen And Lymph Nodes

If nude mice are grafted with a neonatal thymus, host type precursor cells develop within the graft thymus and after about 6 wk the T-cell population of the thymus, spleen, and lymph nodes is of host type. However, immunological responsiveness produced in nude mice in this manner is incomplete: (a) the ability to react to T-cell mitogens in vitro is greater than in untreated nudes but lower than in normal mice; (b) the response to T-cell dependent antigens is less than normal; and (c) the rejection of skin grafts is slower than in normal animals. Whether host precursor cells which differentiate in an allogeneic thymus are able to reject skin grafts from thymus donor strain appears to depend on the strain combination used.

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Platelet-bound lipopolysaccharide enhances Fc receptor–mediated phagocytosis of IgG-opsonized platelets

Blood ◽

10.1182/blood-2006-12-062695 ◽

2007 ◽

Vol 109 (11) ◽

pp. 4803-4805 ◽

Cited By ~ 91

Author(s):

John W. Semple ◽

Rukhsana Aslam ◽

Michael Kim ◽

Edwin R. Speck ◽

John Freedman

Keyword(s):

Flow Cytometry ◽

Toll Like Receptor ◽

Autoimmune Thrombocytopenia ◽

Toll Like Receptor 4 ◽

Platelet Destruction ◽

Gram Negative ◽

Synergistic Increase

Abstract Platelets express Toll-like receptor 4 (TLR4), and this has been shown to be responsible for the thrombocytopenia induced by lipopolysaccharide (LPS) administration in vivo. We studied the role of LPS in mediating platelet phagocytosis by THP-1 cells in vitro by flow cytometry. Opsonization of platelets with an IgG monoclonal (W6/32) antibody or with IgG autoantibody-positive sera from patients with autoimmune thrombocytopenia (AITP) significantly enhanced platelet phagocytosis (P < .001). In contrast, platelet phagocytosis did not occur if platelets were bound with only LPS. If, however, the LPS-bound platelets were also opsonized with either W6/32 or autoantibody-positive sera with titers greater than 4, there was a significant and synergistic increase in Fc-dependent platelet phagocytosis (P < .001, P = .003, P = .048, and P = .047). These results suggest that, in the presence of antiplatelet antibodies, bacterial products can significantly alter platelet phagocytosis, and this may have relevance to how Gram-negative infections enhance platelet destruction in some patients with AITP.

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Toll-Like Receptor-Dependent Immunomodulatory Activity of Pycnogenol®

Nutrients ◽

10.3390/nu11020214 ◽

2019 ◽

Vol 11 (2) ◽

pp. 214 ◽

Cited By ~ 2

Author(s):

Annelies Verlaet ◽

Nieke van der Bolt ◽

Ben Meijer ◽

Annelies Breynaert ◽

Tania Naessens ◽

...

Keyword(s):

Partial Agonist ◽

Immunomodulatory Activity ◽

Toll Like Receptor ◽

Gastrointestinal Digestion ◽

Toll Like Receptor 4 ◽

Tlr Agonists ◽

Anti Inflammatory ◽

Formation Of Complexes

Background: Pycnogenol® (PYC), an extract of French maritime pine bark, is widely used as a dietary supplement. PYC has been shown to exert anti-inflammatory actions via inhibiting the Toll-like receptor 4 (TLR4) pathway. However, the role of the other receptors from the TLR family in the immunomodulatory activity of PYC has not been described so far. Aim: The aim of this study was to investigate whether PYC might exert its immunomodulatory properties through cell membrane TLRs (TLR1/2, TLR5, and TLR2/6) other than TLR4. Moreover, the effect of gastrointestinal metabolism on the immunomodulatory effects of PYC was investigated. Findings: We showed that intact non-metabolized PYC dose-dependently acts as an agonist of TLR1/2 and TLR2/6 and as a partial agonist of TLR5. PYC on its own does not agonize or antagonize TLR4. However, after the formation of complexes with lipopolysaccharides (LPS), it is a potent activator of TLR4 signaling. Gastrointestinal metabolism of PYC revealed the immunosuppressive potential of the retentate fraction against TLR1/2 and TLR2/6 when compared to the control fraction containing microbiota and enzymes only. The dialyzed fraction containing PYC metabolites revealed the capacity to induce anti-inflammatory IL-10 secretion. Finally, microbially metabolized PYC affected the colonic microbiota composition during in vitro gastrointestinal digestion. Conclusions: This study showed that gastrointestinal metabolism of PYC reveals its biological activity as a potential inhibitor of TLRs signaling. The results suggest that metabolized PYC acts as a partial agonist of TLR1/2 and TLR2/6 in the presence of the microbiota-derived TLR agonists (retentate fraction) and that it possesses anti-inflammatory potential reflected by the induction of IL-10 from THP-1 macrophages (dialysate fraction).

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Macrophages and Uveitis in Experimental Animal Models

Mediators of Inflammation ◽

10.1155/2015/671417 ◽

2015 ◽

Vol 2015 ◽

pp. 1-10 ◽

Cited By ~ 11

Author(s):

Salvador Mérida ◽

Elena Palacios ◽

Amparo Navea ◽

Francisco Bosch-Morell

Keyword(s):

Animal Models ◽

Experimental Animal ◽

Macrophage Polarization ◽

Toll Like Receptor ◽

Toll Like Receptor 4 ◽

Experimental Animal Models ◽

Leading Role ◽

Antigen Presenting ◽

Experimental Autoimmune

Resident and infiltrated macrophages play relevant roles in uveitis as effectors of innate immunity and inductors of acquired immunity. They are major effectors of tissue damage in uveitis and are also considered to be potent antigen-presenting cells. In the last few years, experimental animal models of uveitis have enabled us to enhance our understanding of the leading role of macrophages in eye inflammation processes, including macrophage polarization in experimental autoimmune uveoretinitis and the major role of Toll-like receptor 4 in endotoxin-induced uveitis. This improved knowledge should guide advantageous iterative research to establish mechanisms and possible therapeutic targets for human uveitis resolution.

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