The innate immune response in patients who develop inflammatory bowel disease (IBD) may be abnormal. However, the exact role of Toll-like receptors (TLRs) / CD14 gene in the pathogenesis of IBD has not been fully elucidated. We aimed to investigate the association between polymorphisms of TLR1, 2, 4, 6, and CD14 gene and susceptibility to IBD in Korean population. A total 144 patients of IBD (99 patients with ulcerative colitis, 45 patients with Crohn's disease) and 178 healthy controls were enrolled. Using a PCR-RFLP, we evaluated mutations of TLR1 (Arg80Thr), TLR2 (Arg753Gln and Arg677Trp), TLR4 (Asp299Gly and Thr399Ile), TLR6 (Ser249Pro) genes and the -159 C/T promoter polymorphism of CD14 gene. No TLR polymorphisms were detected in Korean subjects. T allele and TT genotype frequencies of CD14 gene were significantly higher in IBD patients than in healthy controls. In subgroup analysis, T allelic frequency was higher in pancolitis phenotype of ulcerative colitis. In Korean population, the promoter polymorphism at -159 C/T of the CD14 gene is positively associated with IBD, both ulcerative colitis and Crohn's disease.
Adult ; Aged ; Alleles ; Antigens, CD14/*genetics ; Asian Continental Ancestry Group/*genetics ; Colitis, Ulcerative/genetics ; Crohn Disease/genetics ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; Humans ; Inflammatory Bowel Diseases/*genetics ; Male ; Middle Aged ; Phenotype ; Polymorphism, Single Nucleotide ; Promoter Regions, Genetic ; Republic of Korea ; Toll-Like Receptor 1/genetics ; Toll-Like Receptor 2/genetics ; Toll-Like Receptor 4/genetics ; Toll-Like Receptor 6/genetics ; Toll-Like Receptors/*genetics

Microglial cells were purified from a mixed neuroglia culture prepared from the neonatal chicken brain in vitro, and were infected with the vvMDV YL040920 isolate and an attenuated MDV vaccine strain CVI988/Rispens, respectively. The presence of cytopathic effect (CPE) was examined daily, and the MEQ expression in MDV-infected microglia was detected by immunohistochemistry assay. DNA replication of the MDV meq gene and transcription of the gB gene were determined by real-time quantitative PCR (qPCR) and qRT-PCR, respectively. The transcripts of Toll-like receptor (TLR) mRNA in microglia post MDV infection were quantified by qRT-PCR. The results of this study showed that both vvMDV YL040920 and attenuated vaccine strain CVI988/Rispens could infect microglia and produce characteristic CPE with plaque formation. The plaques were formed due to cells shedding at multi-sites, then quickly expanded and integrated. Furthermore, the MEQ protein was detected in nuclei of YL040920 and CVI988/ Rispens-infected microglia, and MDV meq DNA replication and gB gene transcription in MDV-infected microglia were also confirmed. Although both MDV DNA copies and gB transcripts were increased in the virus-infected microglia, the higher viral DNA load and gB transcript were observed for CVI988/Rispens than for YL040920 in vitro (P < or = 0.05/0.001). The transcriptions of TLR15 and TLR1LB gene were found to be up-regulated in microglia following MDV infection in vitro. Purified microglia infected with YL040920 was observed increased TLR15 and TLR1LB transcripts as early as 1 day post infection (dpi), and reached its peak level at 3 dpi, then decreased mildly at 5 dpi. For CVI988/Rispens, it induced an increase of TLR15 transcript as early as 1 dpi, and rose rapidly at 3 dpi, and then decreased slightly at 5 dpi. At the same time, CVI988/Rispens induced the increase of chTLR1LB transcript at 3 dpi and decreased at 5 dpi. By comparing the TLRs transcription between YL040920 and CVI988/Rispens-infected microglia, it was suggested that vvMDV YL040920 might induce more TLR15 transcript than the attenuated vaccine strain CVI988/Rispens (P < or = 0.01/0.001), while CVI988/Rispens induced more TLR1LB transcript than YL040920 (P < or = 0.001).
Animals ; Brain ; metabolism ; virology ; Chickens ; Gene Expression ; Herpesvirus 2, Gallid ; genetics ; physiology ; Marek Disease ; genetics ; metabolism ; virology ; Microglia ; metabolism ; virology ; Poultry Diseases ; genetics ; metabolism ; virology ; Toll-Like Receptor 1 ; genetics ; metabolism ; Toll-Like Receptors ; genetics ; metabolism ; Transcription, Genetic

A case-control study was carried out that involved 203 individuals diagnosed with pulmonary tuberculosis (PTB) and 203 healthy subjects. Genotyping of TLR1 rs5743551 and rs5743618 polymorphisms was done using polymerase chain reaction-restriction fragments length polymorphism assay. We found that TLR1 rs5743551 variant affected the risk of PTB in the codominant (OR=3.28, 95% CI=1.98-5.45, P<0.0001, GA vs. GG; OR=1.86, 95% CI=1.05-3.28, P=0.033, AA vs. GG) and dominant (OR=2.69, 95% CI=1.67-4.34, P<0.0001, GA+AA vs. GG) inheritance models tested. The A allele was associated with a higher risk of PTB than the G allele (OR=1.33, 95% CI=1.01-1.75, P=0.049). The TG genotype of the rs5743618 variant significantly increased the risk of PTB compared to the risk associated with the TT genotype (OR=3.29, 95% CI=1.82-5.97, P<0.0001). The G allele was associated with a higher risk of PTB than the T allele (OR=3.00, 95% CI=1.69-5.31, P=0.0001). Our findings revealed that TLR1 rs5743551 and rs5743618 polymorphisms affected the risk of PTB in an Iranian population sample. Additional studies with larger sample sizes and involving subjects of different ethnicities are required to validate our present findings.
Adult ; Case-Control Studies ; Female ; Humans ; Iran ; epidemiology ; Male ; Middle Aged ; Polymorphism, Genetic ; Risk Factors ; Toll-Like Receptor 1 ; genetics ; Tuberculosis, Pulmonary ; epidemiology ; genetics

TLR2 activity plays an important role in the pathogenesis of autoimmune diseases, tumor carcinogenesis and cardio-cerebrovascular diseases. To establish a TLR2 receptor-based cell screening model, NF-kappaB promoter-driven luciferase reporter plasmids were transfected into human embryonic kidney cells (HEK293) stably expressing human TLR2 and co-receptors CD14, TLR1 and TLR6. Single clones were then isolated and characterized. Using this screening system, a human TLR2-binding peptide C8 was obtained from the Ph.D.-7 Phage Display Peptide Library through biopanning and rapid analysis of selective interactive ligands (BRASIL). The binding characteristic of C8 with human TLR2 was evaluated by ELISA, flow cytometry and immunofluorescence. The NF-kappaB luciferase activity assay showed that C8 could activate the TLR2/TLR1 signaling pathway and induce the production of cytokines TNF-alpha and IL-6. In conclusion, the TLR2 receptor-based cell screening system is successfully established and a new TLR2-binding peptide is identified by using this system.
Bacteriophages ; Drug Evaluation, Preclinical ; Genes, Reporter ; HEK293 Cells ; Humans ; Interleukin-6 ; metabolism ; Lipopolysaccharide Receptors ; metabolism ; Luciferases ; genetics ; metabolism ; Peptide Library ; Peptides ; metabolism ; pharmacology ; Promoter Regions, Genetic ; Protein Binding ; Signal Transduction ; drug effects ; Toll-Like Receptor 1 ; metabolism ; Toll-Like Receptor 2 ; metabolism ; Toll-Like Receptor 6 ; metabolism ; Transfection ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo explore the expression of YKL-40, TLR4 and NF-κB in chronic rhinosinusitis (CRS) with or without nasal polyps (CRSwNP and CRSsNP), and to investigate their expressional correlation and the potential role in pathogenesis of CRS.

METHODSThe specimens were obtained from sinus mucosa and inferior turbinate mucosa of the patients with informed consent. The different expression of YKL-40, TLR4 and NF-κB among each group was detected by real time RT-PCR and immunohistochemistry (S-P method). SPSS 17.0 software was used to analyze the data.

RESULTSmRNA level: The relative expression of YKL-40 in CRSwNP group (0.91±0.17) was higher than those in the control group (0.49±0.09), the difference was significant (t=2.12, P<0.05). The relative expression of TLR4 in CRSsNP group (0.88±0.19) and CRSwNP group (0.67±0.13) were lower than those in control group (1.48±0.14), the differences were significant (t value was -4.11, -2.48, all P<0.05). The relative expression of NF-κB in CRSsNP group (0.69±0.13) and CRSwNP group (0.72±0.14) were lower than those in control group (1.20±0.15), the differences were significant (t value was 2.33, 2.27, all P<0.05). Protein level: The expression of YKL-40 in CRSwNP group was stronger than that in CRSsNP group and control group (U value was 72.5 and 73, all P<0.01). The expression of TLR4 in CRSwNP group and CRSsNP group was weaker than that in control group (U value was 62 and 38, all P<0.01). There was a negative correlation between YKL-40 and TLR4 (rmRNA=-0.741, P<0.01; rprotein=-0.46, P<0.05) in CRSwNP group.

CONCLUSIONSThe expression of YKL-40 in pantients with CRSwNP is higher than those in healthy control and CRSsNP patients. There was a negative correlation between YKL-40 and TLR4. Both of them may be involved in the pathogenesis of CRSwNP.

Adipokines ; genetics ; metabolism ; Chitinase-3-Like Protein 1 ; Chronic Disease ; Humans ; Immunohistochemistry ; Lectins ; genetics ; metabolism ; NF-kappa B ; Nasal Mucosa ; Nasal Polyps ; RNA, Messenger ; Rhinitis ; metabolism ; Sinusitis ; metabolism ; Toll-Like Receptor 4 ; metabolism ; Turbinates

Cells, Cultured ; Enzyme-Linked Immunosorbent Assay ; Female ; Fetal Blood ; Humans ; Infant, Newborn ; Lipopolysaccharides ; Lymphocytes ; metabolism ; Male ; RNA, Messenger ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Suppressor of Cytokine Signaling 1 Protein ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; genetics ; metabolism ; Time Factors ; Toll-Like Receptor 2 ; genetics ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

This study aimed to explore the possible effect of Xixin Decoction(XXD) on the learning and memory ability of Alzheimer's disease(AD) model senescence-accelerated mouse-prone 8(SAMP8) and the related mechanism in enhancing neuroprotective effect and reducing neuroinflammation. Forty SAMP8 were randomly divided into a model group(10 mL·kg~(-1)·d~(-1)), a probiotics group(0.39 g·kg~(-1)·d~(-1)), a high-dose group of XXD granules(H-XXD, 5.07 g·kg~(-1)·d~(-1)), a medium-dose group of XXD granules(M-XXD, 2.535 g·kg~(-1)·d~(-1)), and a low-dose group of XXD granules(L-XXD, 1.267 5 g·kg~(-1)·d~(-1)). Eight senescence-accelerated mouse-resistant 1(SAMR1) of the same age and strain were assigned to the control group(10 mL·kg~(-1)·d~(-1)). After ten weeks of intragastric administration, the Morris water maze was used to test the changes in spatial learning and memory ability of mice after treatment. Meanwhile, immunofluorescence staining was used to detect the positive expression of receptor for advanced glycation end products(AGER), Toll-like receptor 1(TLR1), and Toll-like receptor 2(TLR2) in the hippocampal CA1 region of mice. Western blot was employed to test the protein expression levels of silencing information regulator 2 related enzyme 1(SIRT1), AGER, TLR1, and TLR2 in the hippocampus of mice. Enzyme linked immunosorbent assay(ELISA) was applied to assess the levels of Aβ_(1-42) in the hippocampus of mice and the levels of nuclear factor κB p65(NF-κB p65), NOD-like receptor protein 3(NLRP3), tumor necrosis factor-α(TNF-α), and interleukin-1β(IL-1β) in the serum and hippocampus of mice. Compared with the model group, XXD significantly improved the spatial learning and memory ability of SAMP8, increased the expression of neuroprotective factors in the hippocampus, decreased the levels of neuroinflammatory factors, and inhibited the expression of Aβ_(1-42). In particular, H-XXD significantly increased the expression of SIRT1 in the hippocampus of mice, reduced the expression levels of NF-κB p65, NLRP3, TNF-α, and IL-1β in the serum and hippocampus of mice, and decreased the expression of AGER, TLR1, and TLR2 in the hippocampus of mice(P<0.05 or P<0.01). XXD may improve the spatial learning and memory ability of AD model SAMP8 by enhancing the neuroprotective effect and inhibiting neuroinflammation.
Humans ; Neuroprotective Agents/therapeutic use* ; Sirtuin 1/metabolism* ; Toll-Like Receptor 2/metabolism* ; NF-kappa B/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Neuroinflammatory Diseases ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Toll-Like Receptor 1/metabolism* ; Alzheimer Disease/genetics* ; Hippocampus

The aim of this study was to investigate the influence of opiate abuse on the expression of Toll-like receptor 9 (TLR9) in the peripheral blood mononuclear cells (PBMCs) of HIV-1-infected patients and to elucidate possible mechanisms involved in the enhancement of HIV-1 replication by opiate abuse. A total of 200 participants were enrolled in the study by random selection from methadone treatment centers and voluntary HIV counseling and testing centers in the cities of Nanning, Liuzhou, and Qinzhou. These participants included 50 HIV-positive opiate abusers (Opiates HIV(+) group), 50 HIV-negative opiate abusers (Opiates HIV(-) group), 50 HIV-positive subjects who were not opiate abusers (Non-opiates HIV (+) group), and 50 HIV-negative subjects who were not opiate abusers (Control group). PBMCs were isolated from the peripheral blood samples from the subjects and the expression levels of TLR9 mRNA and protein were determined by q-PCR and western blot respectively. There was no significant difference among the four groups in age, gender, nationality, domicile, marital status, educational background or duration of drug abuse (P > 0.05). The median viral loads of the Opiates HIV(+) were significantly higher than those of the Non-Opiates HIV(+) groups (4.450 x 10(3) and 3.977 x 10(3) copies/mL respectively, P < 0.05). The relative expression levels of TLR9 mRNA in the Opiates HIV(+), Non-Opiates HIV(+), Opiates HIV(-) and Control groups were (2.13 +/- 1.59) x 10(-3), (3.66 +/- 2.22) x 10(-3), (1.96 +/- 1.42) x 10(-3) and (7.66 +/- 4.87) x 10(-3), respectively. The expression of TLR9 mRNA was significantly lower in both HIV-1-infected and -uninfected groups of opiate abusers compared with groups of non-abusers (P < 0.05). There was no significant difference in TLR9 mRNA expression levels between the Opiates HIV(+) group and the Opiates HIV(-) group (P > 0.05). However, in the non-opiate groups, the expression levels of TLR9 mRNA in the HIV(+) group were significantly lower than that of the control group (P< 0.05). Western blot results confirmed that the expression of TLR9 protein was lower in the Opiates HIV(+), Non-Opiates HIV(+), and Opiates HIV(-) groups compared to the control group. These results suggest that opiate abuse can decrease the expression of TLR9 in PBMCs, which may result in the enhancement of HIV-1 infection and replication due to a decline in immune response mediated by the TLR9 pathway.
Adolescent ; Adult ; Female ; HIV Infections ; genetics ; metabolism ; virology ; HIV-1 ; physiology ; Humans ; Leukocytes, Mononuclear ; metabolism ; Male ; Middle Aged ; Opioid-Related Disorders ; genetics ; metabolism ; Toll-Like Receptor 9 ; genetics ; metabolism ; Young Adult

OBJECTIVETo investigate the protective mechanisms of Radix Salviae miltiorrhizae (RSM) on chronic alcoholic liver injury in mice.

METHODSThe chronic alcoholic liver injury mouse model was established. The morphologic change of hepatic tissue was observed with hematoxylin-eosin (HE) staining; the levels of toll-like receptor-4 (TLR-4) mRNA in hepatic tissue and hemeoxygenase-1 (HO-1) mRNA were determined using reverse transcription polymerase chain reaction (RT-PCR) technique; and the expression of TLR-4 protein was determined by immunohistochemistry method.

RESULTSRSM could alleviate the fatty degeneration and adiponecrosis of hepatic cells induced by alcohol, down-regulate the expressions of TLR-4 mRNA and HO-1 mRNA, and significantly decrease the number of TLR-4 positive cells.

CONCLUSIONRSM could prevent liver injury from alcohol by way of influencing TLR-4 signal transcription.

Animals ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; Hepatitis, Alcoholic ; drug therapy ; pathology ; Liver ; metabolism ; Male ; Membrane Glycoproteins ; biosynthesis ; genetics ; Membrane Proteins ; Mice ; Phytotherapy ; RNA, Messenger ; biosynthesis ; genetics ; Receptors, Cell Surface ; biosynthesis ; genetics ; Salvia miltiorrhiza ; Toll-Like Receptor 4 ; Toll-Like Receptors

BACKGROUNDToll-like receptor 4 (TLR4) is a crucial receptor in the innate immune system and noninfectious immune responses. It has been reported that TLR4 participates in the pathological course of ischemia/reperfusion (I/R) injury. However, the role of TLR4 in the process of I/R injury after cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) is still unknown. In this study, we investigated the effects of TLR4 mutation on survival and neurological outcome in a mouse model of CA/CPR.

METHODSA model of potassium-induced CA was performed on TLR4-mutant mice (C3H/HeJ) and wild-type mice (C3H/HeN). After 3 min of untreated CA, resuscitation was attempted with chest compression, ventilation, and intravenous epinephrine. Behavioral tests were performed on mice on day 3 after CPR. The morphological changes in hippocampal neurons were assessed by light and electron microscopy. Expressions of TLR4 and intercellular adhesion molecule-1 (ICAM-1) were detected by Western blot. Levels of tumor necrosis factor-α (TNF-α) and myeloperoxidase (MPO) were measured with enzyme-linked immunosorbent assay (ELISA).

RESULTSOn day 3 after resuscitation the overall mortality was 33.33% in C3H/HeJ group compared with 53.33% in C3H/HeN group (P < 0.05). And there was much higher central tendency in C3H/HeJ group than C3H/HeN group during open field test (P < 0.05). Meanwhile, the percentage of nonviable neurons was 21.16% in C3H/HeJ group compared with 53.11% in C3H/HeN group (P < 0.05). And there were significantly lower levels of hippocampal TNF-α and MPO in C3H/HeJ mice (TNF-α: 6.85±1.19 ng/mL, MPO: 0.33±0.11 U/g) than C3H/HeN mice (TNF-α: 11.36±2.12 ng/mL, MPO: 0.54±0.17 U/g) (all P < 0.01). CPR also significantly increased the expressions of TLR4 and ICAM-1 in C3H/HeN group. However, the expression of ICAM-1 was much lower in C3H/HeJ group than in C3H/HeN group after CPR (P < 0.01).

CONCLUSIONTLR4 signaling is involved in brain damage and in inflammation triggered by CA/CPR.

Animals ; Blotting, Western ; Brain ; immunology ; metabolism ; Cardiopulmonary Resuscitation ; methods ; Heart Arrest ; genetics ; metabolism ; therapy ; Intercellular Adhesion Molecule-1 ; metabolism ; Male ; Mice ; Mutation ; Peroxidase ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism