TLR4 Rs4986790 Research Articles

Study of the association of toll like-4 receptor gene polymorphism (rs1927911) and susceptibility to urinary tract infection among Iraqi patients

Introduction and Aim: Toll-like receptors (TLRs) are cell innate immune receptors that explore microbial molecules. A quantity of single-nucleotide polymorphisms (SNPs) within the TLR genes have been related with altered susceptibility to infectious, inflammatory, and allergic diseases suggesting: This study was conducted to investigate the relationship between genetic polymorphism in TLR4 gene (rs1927911) with susceptibility to UTIs in a sample of Iraqi patients. Materials and Methods: Blood samples were collected from 220 patients who had UTIs and 69 healthy subjects as control group, during the period from October 2021 to March 2022, at the Rheumatology Department of Clinic Consultant at Al-Yarmouk Hospital. The genetic study included an extraction of genomic DNA and PCR technique, to detect the role of TLR4 gene in the variation site (rs1927911) as a risk factor for the incidence of UTIs. Results: The results of genotype distribution and allele frequency of rs1927911 indicated, CC (18.6 %), CT (70 %) and TT (11.4 %) among the patients while 57.9 % for CC and 26.1%; 16% for CT, TT genotypes respectively of healthy control. Heterozygous (CT) may be considered as a risk factor among UTIs patients. The results revealed significant differences between UTIs patients and healthy controls in serum levels TLR4, IL-6 IL-2, TNF-alpha and IL-10 at P 0.005. In addition, the data found a highly significant difference of serum level of TLR4 protein with patients who have a carrier CT genotype. Conclusion: our findings indicated that the carriers of CT genotype of TLR4 (rs1927911) were significantly more prone to UTIs.

THE ROLE OF ANTIBACTERIAL THERAPY IN THE TREATMENT OF ATOPIC DERMATITIS IN CHILDREN

Background. Atopic dermatitis is one of the most common chronic inflammatory skin diseases. In atopic dermatitis, the diversity of the normal microflora decreases, so that the number of S. aureus prevails and the number of bacteria with antistaphylococcal activity decreases. The aim of this study was to evaluate the effectiveness of mupirocin 2% cream in the treatment of atopic dermatitis in children as an additional therapy, and the effect of the rs4696480 polymorphism in the TLR2 gene on the effectiveness of the treatment.Materials and methods. The study included patients with atopic dermatitis (n = 37), aged 1-18 years (7.9±4.9) from the Department of Allergy at Kyiv City Children Clinical Hospital №2. All children included in the study with atopic dermatitis had a positive S. aureus culture. Patients were randomized into two groups: a group receiving mupirocin 2% cream on the affected skin areas 2 times a day 10 days (group A) and a control group (group B). Children of the control group received only symptomatic therapy. The SCORAD score and CDLQI (children's dermatology life quality index) questionnaire score was recorded before and after the treatment, side effects were recorded during the study. Skin swab cultures were taken before and after treatment. Genotyping for TLR2 rs4696480 was performed by using Real-time PCR.This study was approved by the ethical committee of the O.Bogomolets National Medical University; all patients/parents of the children gave an informed consent to participate. Statistical processing of the obtained data was carried out using the statistical package IBM SPSS Statistics Base (version 22) and the software EZR version 1.32 (graphical interface of the R environment (version 2.13.0). The difference in the effectiveness of therapy between the two subgroups and the influence of genotype on the effectiveness of therapy was determined using the Student criterion for parametric data (T) and Wilcoxon W-test for nonparametric data (W). The dynamics of indicators in each group before and after treatment were evaluated using the Wilcoxon T-test (T-W). Eeffectiveness of S. aureus eradication was estimated using the odds ratio (OR) with a 95% confidence interval (CI). Results were considered statistically significant at the level of p < 0.05.Results. In both groups of children, improvement was observed in 10 days after the treatment. In group A, the improvement in ΔSCORAD was found to be 13.8 (T-W=190.0, p<0.001). In group B, there was also an improvement: ΔSCORAD 8.5 (T-W=153.0, p<0.001). The difference in ΔSCORAD scores between groups A and B was statistically significant (T=2.70, p=0.011). The decrease in CDLQI score after treatment was 31.3% in group A (T-W=190.0, p<0.001), and 18.3% in group B (T-W=171.0, p<0.001). The difference between these two groups was not statistically significant (W=334.0, p=0.409).After the treatment, the skin culture showed that in group A, 57.9% of patients were S. aureus-negative, in group B, only 22.2% of children had a negative culture (OR= 5.50, CI 1.32-22.86).We compared ΔSCORAD depending on the genotype of the rs4696480 polymorphism in TLR2 gene and found no difference in the two subgroups: In the subgroup with the AA genotype, ΔSCORAD was found to be 12.6±3.7; in the subgroup with the TT genotype ‒ 14.4±5.1 (T=0.84, p=0.413).Conclusions. Our results demonstrate the effectiveness of the use of topical mupirocin in the treatment of atopic dermatitis as an additional therapy.

Impact of Toll-Like Receptor 2 and 9 Gene Polymorphisms on COVID-19: Susceptibility, Severity, and Thrombosis.

Toll-like receptors (TLRs) play an important role in activation of innate and adaptive immune responses. We aimed to detect the association between TLR2 rs5743708 G>A and TLR9 rs5743836 C>T variants and COVID-19 disease susceptibility, severity, and thrombosis by using neutrophil extracellular traps (NETs). We included 100 adult COVID-19 patients as well as 100 age- and gender-matched normal controls. Participants were genotyped for TLR2 rs5743708 and TLR9 rs5743836. Citrullinated Histone (H3) was detected as an indicator of NETs. The mutant (G/A and C/C) genotypes and (A and C) alleles of TLR2 rs5743708 and TLR9 rs5743836, respectively, have been significantly related to a higher risk of COVID-19 infection, representing a significant risk factor for the severity of COVID-19. There was no significant association between the two variants and citrullinated histone (H3). TLR2 rs5743708 and TLR9 rs5743836 variants have been significantly related to a higher risk and severity of COVID-19 infection but had no effect on thrombus formation.

Association of TLR4 and TLR9 Gene Polymorphisms with the Risk and Progression of Cervical Lesions in HPV-Infected Women

Aim: Toll-like receptors involved in tumor-associated inflammatory response, this study aimed to investigate the role of TLR4 and TLR9 gene polymorphisms in the risk and progression of HPV-related cervical lesions. Materials & methods: A total of 220 cervical lesion patients and 227 healthy controls were enrolled. Single-nucleotide polymorphisms were genotyped using PCR-restriction fragment length polymorphism. Results: A significantly decreased risk of cervical lesions was observed to be associated with the TLR4 rs10116253 (C), rs1927911 (T) and rs10759931 (G) mutant alleles. rs187084-rs1927911-HPV-16/18 was the best interaction model to affect cervical lesion risk. Conclusion: TLR4 rs10116253, rs1927911 and rs10759931 were potential biomarkers for cervical lesion susceptibility.

Evaluation of toll-like receptors 2 and 4 polymorphism and intestinal microbiota in children with food allergies.

Mutual regulation between immune system and gut microbiota is achieved through several mechanisms including the engagement of toll-like receptors (TLRs) which is expressed on numerous cell types. In this study we aimed to explore the association between food allergies and TLR gene polymorphisms in association with gut microbiota. Toll-like receptors polymorphism frequencies and some bacteria in the gut microbiota in 130 infants aged 1-24 months with egg and/or milk allergy in a prospective cohort were compared with 110 non-food allergic controls. Four candidate polymorphisms (TLR2 rs1898830/rs5743708 and TLR4 rs4986790/rs4986791) were genotyped by allelic discrimination polymerase chain rection (PCR) method. Gut microbiota analysis was achieved by using high-throughput sequencing. The TLR4 rs4986790 (Asp299Gly) single nucleotide polymorphism (SNP) major/minor allele frequency was 0.788/0.212 in food allergy patients and 0.719/0.280 in controls (p=0.017). There was a statistically significant difference between groups in terms of genotype frequencies (AA, AG, GG). Gut microbiota analysis revealed increased Firmicutes phylum in stool of the patients with food allergy. Except for TLR4 rs4986791 (Thr399lle) allele, the other TLR polymorphisms were not associated with food allergies in children. When the bacteria in the intestinal microbiota and TLR2 and TLR4 gene polymorphisms were compared; we determined a statistically significant increase in Bifidobacterium concentration in the intestinal microbiota in TLR4 rs4986791 CT heterozygous genotype (p=0.004). This study demonstrated a partial role of TLR4 gene polymorphism and gut microbiota in the development of food allergies. Future work in this area will be required to clarify the roles of different microbial strains that modulate gut microbiota composition and function in conjunction with TLR transcription pathways.

Relationship of Toll-Like Receptors 2 and 4 Gene Polymorphisms with Essential Hypertension in Chinese Han Population

Objective: There are numerous studies suggesting that genetic polymor-phisms of inflammation factors Toll-like receptors 2 and 4 (TLR2, TLR4) might play a role in the pathophysiological process of hypertension. In this study, we evaluated the association in a sample of members of the Chinese Han population. Method: We selected four single nucleotide polymor-phisms (SNP) of TLR2 (rs3804099, rs3804100, rs7656411) and TLR4 (rs1927906) genes, and measured the distributions of genotypic and allelic frequencies in 1063 participants, including 391 essential hypertension pa-tients and 672 controls. Result: No significant differences in the genotypic and allelic frequencies of the four SNPs were detected between cases and controls. However, three haplotypes, CCG, TTG and TTT of TLR2, were significantly associated with a decrease in the risk of essential hyperten-sion (OR: 0.512, 95% CI: 0.397 - 0.660, P P = 0.0038; OR: 0.797, 95% CI: 0.667 - 0.952, P = 0.0122, respectively). Inversely, the risk of essential hypertension increased sig-nificantly in patients with the CTG, TCG or TCT haplotypes (OR: 2.924, 95% CI: 2.157 - 3.963, P P P TLR2 might have profound effects on the development of essential hypertension in the Chinese Han population.

The association of rs4696480 polymorphism in TLR2 gene and Staphylococcus aureus skin colonization in children with atopic dermatitis

Topicality. Up to 90% of patients with atopic dermatitis are colonized with S. aureus, with S. aureus predominance being unique to atopic dermatitis. TLR2 play a role in the presentation of S. aureus antigens in a course of atopic dermatits. Purpose - to investigate the association of rs4696480 polymorphism in TLR2 gene and S. aureus skin colonization. Materials and methods. The study included 101 patients with eczema and 84 healthy children. Skin swab cultures were taken. Subjects were classified as carriers if the cultures were positive, while those with culture found to be negative were classified as non-carriers. Genotyping for TLR2 rs4696480 was performed by using Real-time PCR. Results. We determined the prevalence of S. aureus carriage in a cohort study of atopic dermatitis patients in Ukraine. Skin culture for the presence of S. aureus was performed in 82 patients: 45.1% children had positive culture for S. aureus, 54.9% had a negative result. SCORing Atopic Dermatitis (SCORAD) index was significantly higher in S. aureus carriers (p<0.001). There was no difference in genotype distribution among patients and control group (OR=1.021 (95% CI 0.507-2.054) for AT genotype, OR=0.880 (95% CI 0.398-1.947) for TT genotype, р>0.05). AA genotype was significantly more frequent among S. aureus carriers (OR=2.745 (95% CI 0.865-8.708) for AT genotype, OR=7.000 (95% CI 1.852-26.462) for TT genotype. To our knowledge, the association of T16934A (rs4696480) and S. aureus colonization of lesion skin in children with atopic dermatitis have not been studied before. Conclusions. AA variant of TLR2 rs4696480 polymorphism predisposes to S. aureus colonization of skin in children with atopic dermatits. The research was carried out in accordance with the principles of the Helsinki Declaration. The study protocol was approved by the Local Ethics Committee of the participating institution. The informed consent of the patient was obtained for conducting the studies. No conflict of interests was declared by the author.

Impact of polymorphisms in genes orchestrating innate immune responses on replication kinetics of Torque teno virus after kidney transplantation.

Background: Torque teno virus (TTV) DNAemia has been proposed as a surrogate marker of immunosuppression after kidney transplantation (KT), under the assumption that the control of viral replication is mainly exerted by T-cell-mediated immunity. However, Tthe impact on post-transplant TTV kinetics of single genetic polymorphisms (SNPs) in genes orchestrating innate responses remains unknown. We aimed to characterize the potential association between 14 of these SNPs and TTV DNA levels in a single-center cohort of KT recipients. Methods: Plasma TTV DNAemia was quantified by real-time PCR in 221KT recipients before transplantation (baseline) and regularly through the first 12 post-transplant months. We performed genotyping of the following SNPs: CTLA4 (rs5742909, rs231775), TLR3 (rs3775291), TLR9 (rs5743836, rs352139), CD209 (rs735240, rs4804803), IFNL3 (rs12979860, rs8099917), TNF (rs1800629), IL10 (rs1878672, rs1800872), IL12B (rs3212227) and IL17A (rs2275913). Results: The presence of the minor G allele of CD209 (rs4804803) in the homozygous state was associated with undetectable TTV DNAemia at the pre-transplant assessment (adjusted odds ratio: 36.96; 95% confidence interval: 4.72-289.67; p-value = 0.001). After applying correction for multiple comparisons, no significant differences across SNP genotypes were observed for any of the variables of post-transplant TTV DNAemia analyzed (mean and peak values, areas under the curve during discrete periods, or absolute increments from baseline to day 15 and months 1, 3, 6 and 12 after transplantation). Conclusion: The minor G allele of CD209 (rs4804803) seems to exert a recessive protective effect against TTV infection in non-immunocompromised patients. However, no associations were observed between the SNPs analyzed and post-transplant kinetics of TTV DNAemia. These negative results would suggest that post-transplant TTV replication is mainly influenced by immunosuppressive therapy rather than by underlying genetic predisposition, reinforcing its clinical application as a biomarker of adaptive immunity.

E6020, a TLR4 Agonist Adjuvant, Enhances Both Antibody Titers and Isotype Switching in Response to Immunization with Hapten-Protein Antigens and Is Diminished in Mice with TLR4 Signaling Insufficiency.

The mechanisms by which TLR4-based adjuvants enhance immunogenicity are not fully understood. We have taken advantage of a novel knock-in mouse strain that homozygously expresses two single-nucleotide polymorphisms (SNPs) that are homologous to human TLR4 (rs4986790 and rs4986791) and have been associated with LPS hyporesponsiveness in vivo and in vitro. TLR4-SNP (coexpressing mutations D298G/N397I in TLR4) mice that recapitulate the human phenotype were compared with wild-type (WT) mice for their hapten-specific Ab responses after immunization with hapten 4-hydroxy-3-nitrophenyl acetyl (NP) NP-Ficoll or NP-OVA in the absence or presence of a water-soluble TLR4 analog adjuvant, E6020. IgM and IgG anti-NP responses were comparable in WT and TLR4-SNP mice after immunization with either NP-Ficoll or NP-OVA only. E6020 significantly yet transiently improved the IgM and IgG anti-NP responses of both WT and TLR4-SNP mice to NP-Ficoll (T-independent), with modestly enhanced Ab production in WT mice. In contrast, T-dependent (NP-OVA), adjuvant-enhanced responses showed sustained elevation of NP-specific Ab titers in WT mice, intermediate responses in TLR4-SNP mice, and negligible enhancement in TLR4-/- mice. E6020-enhanced early humoral responses in WT and TLR4-SNP mice to NP-OVA favored an IgG1 response. After a second immunization, however, the immune responses of TLR4-SNP mice remained IgG1 dominant, whereas WT mice reimmunized with NP-OVA and E6020 exhibited increased anti-NP IgG2c titers and a sustained increase in the IgG1 and IgG2c production by splenocytes. These findings indicate that E6020 increases and sustains Ab titers and promotes isotype class switching, as evidenced by reduced titers and IgG1-dominant immune responses in mice with TLR4 insufficiency.

Association of TLR7 and TLR9 genes polymorphisms in Egyptian patients with systemic lupus erythematosus

IntroductionSystemic lupus erythematosus (SLE) is a chronic, inflammatory, multiorgan, systemic autoimmune disease. It is characterized by the high production of autoantibodies against nuclear compounds. TLRs (toll-like receptors 7/9) are pattern-recognition receptors that recognize nucleic acids and induce proinflammatory responses by activating NF-kB and producing type I interferon, which play a role in eliciting innate/adaptive immune responses and developing chronic inflammation. TLR7 and TLR9 single nucleotide polymorphisms (SNPs) have been linked to systemic lupus erythematosus in numerous studies (SLE). In this work, we wanted to evaluate and analyze single nucleotide polymorphisms (SNPs) in the TLR7 (rs3853839) and TLR9 (rs187084) genes among Egyptian SLE patients and healthy controls. MethodWhole blood samples were taken from 100 SLE patients and 100 controls; DNA was extracted and then processed for TLR7 rs3853839 and TLR9 rs187084 single nucleotide polymorphisms analysis by real-time polymerase chain reaction technology and restriction fragment-length polymorphism. We also assessed the association between TLR 7 and TLR 9 genes polymorphism with SLE clinical parameters. ResultsOur results showed that TLR7 rs3853839 CG genotypes and G allele were significantly associated with SLE. Also, TLR7 rs3853839 genotypes and alleles were significantly associated with nephritis, arthritis, oral ulcers, and thrombocytopenia.Whereas genotypes and alleles of TLR9 were not significantly associated with the risk nor the clinical characteristics of SLE except for malar rash. ConclusionIn the investigated Egyptian cohort, our findings suggest that TLR7 rs3853839 gene polymorphisms increase the risk for SLE development and play a role in developing clinical characteristics, especially nephritis.

Toll-like receptor and matrix metalloproteinase single-nucleotide polymorphisms, haplotypes, and polygenic risk score differentiated between tuberculosis disease and infection

ObjectivesThe association of toll-like receptors (TLRs) and matrix metalloproteinases (MMPs) single-nucleotide polymorphisms (SNPs) among latent tuberculosis (TB) infection and active TB remained less studied. MethodsWe recruited participants with TB disease (active TB) (n = 400) and TB infection (latent TB infection) (n = 203) in this study. We genotyped SNPs in TLR1, TLR2, TLR4, MMP1, MMP8, MMP9, MMP12, and tissue inhibitor of MMP2. Single-variant analysis and haplotype analysis were performed, and a polygenic risk score (PRS) was created. ResultsWe found that SNPs in TLR1 (rs5743580, rs5743551), TLR2 (rs3804100), and MMP8 (rs2508383) were associated with different TB disease status risks. TLR1 rs5743580 was associated with a higher risk of TB disease status in genotypic, recessive, and additive models. TLR2 rs3804100 polymorphisms demonstrated significant association with TB disease status in genotypic, dominant, and additive models. In the haplotype analysis, the TLR1 haplotype was associated with a higher risk of TB disease, and the MMP12 haplotype was associated with a lower risk of TB disease. A PRS using 3 SNPs was associated with a higher risk of TB disease. ConclusionThis study revealed that SNP variants in TLR1, TLR2, and MMP8 differed among TB infection and disease. Haplotypes and PRS could potentially help predict TB disease status.

The Toll-like Receptor 4 Polymorphism Asp299Gly Is Associated with an Increased Risk of Ovarian Cancer.

Ovarian cancer (OC) is one of the most common cancers threatening women’s lives around the world. Epithelial ovarian tumors represent the most common ovarian neoplasms. Most OC patients are diagnosed at the advanced stage, and there is an urgent need to identify novel biomarkers of the disease. Single-nucleotide polymorphisms (SNPs) in TLR genes may serve as crucial markers of cancer susceptibility. We investigated the frequency of TLR polymorphisms in a group of 200 women, including 70 with OC. Four SNPs, two each in TLR4 (rs4986790 and rs4986791) and TLR9 (rs187084 and rs5743836), were analyzed using polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP). The digested fragments were separated and identified by multicapillary electrophoresis. The load quantification of human papillomavirus (HPV) types 16/18 was determined using a digital droplet PCR method. We found an increased frequency of heterozygous genotype and minor allele of the TLR4 rs4986790 SNP in women with OC compared with healthy controls, and this result remained highly significant after Bonferroni’s correction for multiple testing (p < 0.0001). No evidence of linkage disequilibrium was found with any of the examined TLR SNPs. The findings suggest that the TLR4 Asp299Gly polymorphism could be a genetic risk factor for the development of OC.

Exploring the significant role of TLR 7 gene polymorphism with severity infection of COVID-19 patients in Wasit province

Toll-like receptor 7 (TLR7 genes was involved in the host immune response against viral infections including SARS-COV-2. This study aimed to investigate the association between the TLR7 (rs179008) polymorphisms with the prognosis and susceptibility to COVID-19 pneumonia accompanying SARSCOV-2 infection. This case-control study included 120 individuals: 80 COVID-19 patients (severe and mild) and 40 controls. Polymorphisms (TLR7 rs179008) were genotyped by Sanger sequencing using ABI3730XL, automated DNA sequencer, by Macrogen Corporation – Korea after sent PCR products. This study also investigated predictors of mortality in COVID-19 severity through logistic regression. The mutant‘T/T' genotypes and the ‘T' alleles of TLR7 (rs179008) polymorphisms were significantly associated with increased risk of COVID-19 severity. Our study illustrated that the males harbouring the TT genotype of TLR7 rs 170008 polymorphism could be more susceptible to COVID-19 pneumonia than females having the same genotype.

TLR3 polymorphisms in HIV infected individuals naïve to ART.

TLR3 polymorphisms affect the risk of HIV infection and modify the disease course. Consequently, we analyzed the association of TLR3 polymorphism (rs5743312, rs3775296, and rs3775291) with susceptilbity to HIV-1 acquisition and disease progression. This is a cross-sectional study. Genotyping of TLR3 polymorphisms were completed by the utilization of the PCR-RFLP technique in 153 HIV naive subjects and 158 healthy controls. A haplotype is a physical grouping of genomic variants that tend to be inherited together. The TCC haplotype was increased in HIV infected individuals compared with healthy controls (0.05% versus 0.03%). TLR3 rs3775291CT genotype was associated to the early stage of HIV infection (OR=2.19, P=0.04), with a higher occurrence in advance stage of HIV infection when contrasted with healthy controls (41.2% versus 32.3%). TLR3 rs3775296 CA genotype was likely to be associated with intermediate stage of HIV infection (19.5% versus 31.6%, OR=0.42, P=0.06). TLR3 rs5743312TT genotype was used to be greater prevalence in advanced stage of HIV infection compared with healthy controls (2.9% versus 1.9%). TLR3 rs3775296CA genotype was less prevalent in HIV subjects devouring tobacco when contrasted with non-users (9.1% versus 34.9%, OR=0.25, P=0.09). TLR3 rs3775296AA and rs3775291CT and TT genotypes have been overrepresented in HIV subjects using alcohol when contrasted with non-users (5.6% versus 1.1%, OR=1.83, P=0.67; 50.0% versus 42.2%, OR=1.84, P=0.31; 5.6% versus 3.3%, OR=2.70, P=0.50). In multivariate examination, rs5743312TT genotype showed a greater risk for HIV infection (OR=1.86, P=0.50). TLR3 rs3775291 C/T polymorphism may assist the risk of disease progression in alcohol consumers. TLR3 rs3775291 CT genotype may enhance the disease progression whereas the TLR3 rs3775296 CA genotype may protect for disease progression.

Polymorphisms in the Genes Coding for TLRs, NLRs and RLRs Are Associated with Clinical Parameters of Patients with Acute Myeloid Leukemia.

Toll-like receptors (TLRs), NOD-like receptors (NLRs), and RIG-I-like receptors (RLRs) are major elements of the innate immune system that recognize pathogen-associated molecular patterns. Single-nucleotide polymorphisms (SNPs) in the TLR, NLR, and RLR genes may lead to an imbalance in the production of pro- and anti-inflammatory cytokines, changes in susceptibility to infections, the development of diseases, and carcinogenesis. Acute myeloid leukemia (AML) is a bone marrow malignancy characterized by uncontrolled proliferation of transformed myeloid precursors. We retrospectively analyzed 90 AML patients. We investigated the effect of fifteen SNPs located in the genes coding for RLR1 (rs9695310, rs10738889, rs10813831), NOD1 (rs2075820, rs6958571), NOD2 (rs2066845, rs2066847, rs2066844), TLR3 (rs5743305, rs3775296, 3775291), TLR4 (rs4986791, rs4986790), and TLR9 (rs187084, rs5743836). We observed that TLR4 rs4986791, TLR9 rs5743836, and NOD2 rs2066847 were associated with CRP levels, while RLR-1 rs10738889 was associated with LDH level. Furthermore, we found TLR3 rs5743305 AA to be more common in patients with infections. We also found TLR9 rs187084 C to be associated with more favorable risk, and RLR-1 rs9695310 GG with higher age at diagnosis. In conclusion, the current study showed that SNPs in the genes encoding TLRs, NLRs, and RLRs may be potential biomarkers in patients with AML.

Polygenic Innate Immunity Score to Predict the Risk of Cytomegalovirus Infection in CMV D+/R- Transplant Recipients. A Prospective Multicenter Cohort Study.

Several genetic polymorphisms of the innate immune system have been described to increase the risk of cytomegalovirus (CMV) infection in transplant patients. The aim of this study was to assess the impact of a polygenic score to predict CMV infection and disease in high risk CMV transplant recipients (heart, liver, kidney or pancreas). On hundred and sixteen CMV-seronegative recipients of grafts from CMV-seropositive donors undergoing heart, liver, and kidney or pancreas transplantation from 7 centres were prospectively included for this purpose during a 2-year period. All recipients received 100-day prophylaxis with valganciclovir. CMV infection occurred in 61 patients (53%) at 163 median days from transplant, 33 asymptomatic replication (28%) and 28 CMV disease (24%). Eleven patients (9%) had recurrent CMV infection. Clinically and/or functionally relevant single nucleotide polymorphisms (SNPs) from TLR2, TLR3, TLR4, TLR7, TLR9, AIM2, MBL2, IL28, IFI16, MYD88, IRAK2 and IRAK4 were assessed by real time polymerase chain reaction (RT-PCR) or sequence-based typing (PCR-SBT). A polygenic score including the TLR4 (rs4986790/rs4986791), TLR9 (rs3775291), TLR3 (rs3775296), AIM2 (rs855873), TLR7 (rs179008), MBL (OO/OA/XAO), IFNL3/IL28B (rs12979860) and IFI16 (rs6940) SNPs was built based on the risk of CMV infection and disease. The CMV score predicted the risk of CMV disease with an AUC of the model of 0.68, with sensitivity and specificity of 64.3 and 71.6%, respectively. Even though further studies are needed to validate this score, its use would represent an effective model to develop more robust scores predicting the risk of CMV disease in donor/recipient mismatch (D+/R-) transplant recipients.

Influence of single-nucleotide polymorphisms in TLR3 (rs3775291) and TLR9 (rs352139) on the risk of CMV infection in kidney transplant recipients.

Risk stratification for cytomegalovirus (CMV) infection after kidney transplantation (KT) remains to be determined. Since endosomal toll-like receptors (TLRs) are involved in viral sensing, we investigated the impact of common single-nucleotide polymorphisms (SNPs) located within TLR3 and TLR9 genes on the occurrence of overall and high-level (≥1,000 IU/ml) CMV infection in a cohort of 197 KT recipients. Homozygous carriers of the minor allele of TLR3 (rs3775291) had higher infection-free survival compared with reference allele carriers (60.0% for TT versus 42.3% for CC/CT genotypes; P-value = 0.050). Decreased infection-free survival was observed with the minor allele of TLR9 (rs352139) (38.2% for TC/CC versus 59.3% for TT genotypes; P-value = 0.004). After multivariable adjustment, the recessive protective effect of the TLR3 (rs3775291) TT genotype was confirmed (adjusted hazard ratio [aHR]: 0.327; 95% CI: 0.167–0.642; P-value = 0.001), as was the dominant risk-conferring effect of TLR9 (rs352139) TC/CC genotypes (aHR: 1.865; 95% CI: 1.170–2.972; P-value = 0.009). Carriers of the TLR9 (rs352139) TC/CC genotypes showed lower CMV-specific interferon-γ-producing CD4+ T-cell counts measured by intracellular cytokine staining compared with the TT genotype (median of 0.2 versus 0.7 cells/μl; P-value = 0.003). In conclusion, TLR3/TLR9 genotyping may inform CMV infection risk after KT.

Toll-like receptor-1, -2, and -6 genotypes in relation to salivary human beta-defensin-1, -2, -3 and human neutrophilic peptide-1.

To examine whether functional gene polymorphisms of toll-like receptor (TLR)1, TLR2, and TLR6 are related to the salivary concentrations of human beta-defensins (hBDs)-1, -2, -3, and human neutrophilic peptide (HNP)-1. Polymorphisms of TLR1 (rs5743618), TLR2 (rs5743708), and TLR6 (rs5743810) were genotyped by PCR-based pyrosequencing from the salivary samples of 230 adults. Salivary hBD-1, -2, -3, and HNP-1 concentrations were measured using enzyme-linked immunosorbent assay. General and periodontal health examinations, including panoramic radiography, were available for all participants. The genotype frequencies for wild types and variant types were as follows: 66.5% and 33.5% for TLR1, 95.5% and 4.5% for TLR2, and 25.1% and 74.9% for TLR6, respectively. The TLR2 heterozygote variant group exhibited higher salivary hBD-2 concentrations than the TLR2 wild-type group (p= .038). On the contrary, elevated hBD-2 concentrations were detected in the TLR6 wild-type group compared with the TLR6 heterozygote and homozygote variant group (p= .028). The associations between TLR6 genotypes and salivary hBD-2 concentrations remained significant after adjusting them for periodontal status, age, and smoking. hBD-2 concentrations in saliva are related to TLR2 and TLR6 polymorphisms, but only the TLR6 genotype seems to exhibit an independent association with the salivary hBD-2 concentrations.

Polymorphisms in toll-like receptor 3 and 4 genes as prognostic and outcome biomarkers in melanoma patients.

Melanoma is one of the most aggressive tumors, and in the setting of rising incidence and mortality, there is an urgent need to identify new prognostic markers. Toll-like receptors (TLRs), are aberrantly expressed in numerous cancers, including melanoma. TLR signaling provides a microenvironment that is involved in antitumor immune response, chronic inflammation, cancer cell proliferation and evasion of immune destruction. In the present study, we investigated whether single nucleotide polymorphisms (SNPs) in TLR3 and TLR4 genes are associated with clinicopathologic features, progression and survival of melanoma patients. The study was conducted on 120 melanoma patients. DNA extracted from peripheral blood was genotyped for TLR3 polymorphisms rs5743312 and rs3775291 (L412F) and TLR4 polymorphisms rs4986790 (D299G) and rs4986791 (T399I), by TaqMan Real-Time PCR Assays. Kaplan-Meier survival curves were compared by the log-rank test. TLR3 polymorphism L412F was associated with a higher mitotic index ( P = 0.035). TLR4 D299G and T399I polymorphisms were associated with indicators of melanoma severity, nodal metastases ( P = 0.005 and P = 0.007, respectively) and advanced stage III ( P = 0.005 and P = 0.004, respectively). Cox regression analysis showed that the presence of tumor-infiltrating lymphocytes (TILs) predicted better overall survival (HR = 0.318; P = 0.004). TLR4 T399I polymorphism was significantly associated with worse survival, P = 0.025. The overall survival rates were significantly lower for patients carrying variant allele T of TLR4 T399I SNP (TC and TT genotypes combined) ( P = 0.008, log-rank test), compared to wild-type genotype CC. Our findings indicate that TLR4 polymorphisms T399I (rs4986791) and D299G (rs4986790) could be potential prognostic and survival markers for melanoma patients.

Role of cytokine and Toll-like receptor genes in pathogenesis of inborn heart disease

Sporadic congenital heart disease (CHD) may result from immune disorders in the mother – embryo system and/or constitutional disorders in regulatory systems, including those associated with TLR receptors, cytokines and their receptors. The aim of our study was to investigate associations between cytokine and TLR genes and sporadic congenital heart disease in children. In the main group, 188 children with sporadic (without family history) congenital heart defects were examined. Separate groups of CHD were identified: septal CHD – 98 children; valvular heart disease – 17 children; Fallot tetralogy – 15 children; aorta coarctation – 10 children; fetal drains – 32 children; single ventricle affection – 9 children, and anomalous drainage of v. pulmonalis was diagnosed in 7 children. The control group included 103 age- and sex-matched healthy children. We have determined gene polymorphisms of five genes encoding cytokines and their receptors (IL6 rs1800796, IL6 rs2069827, IL6R rs2228145, IL6R rs2229238, IL8 rs4073, IL10 rs1800871, IL10 rs1800896, IL10 rs1800872, TNF rs1800629, TNF rs361525, TNF rs1799964), four genes Toll-like receptors (TLR: TLR1 rs5743611, TLR1 rs5743551, TLR2 rs5743708, TLR2 rs3804099, TLR4 rs4986791, TLR4 rs4986790, TLR6 rs3775073, TLR6 rs5743810). The dbSNP, SNPinfo, SNPnexus databases were used to select and design test systems. Stepwise logistic regression was the main method of statistical analysis. Clinical diagnosis of congenital heart defects is associated with immune regulatory genes. In particular, the missense mutation TLR6 rs5743810, which was a predictor of congenital valvular heart disease, is of particular importance. Development of congenital heart valve defects and aortic coarctation is associated with intergenic interactions of TLR2 rs5743708 with TLR6 rs5743810, and TLR2 rs5743708 with TLR6 rs3775073, respectively. For congenital heart valve defects, such polymorphic regions are as follows: IL6 rs2069827, IL6R rs2229238, and IL8 rs4073, for aortic coarctation – IL6R rs2228145, IL8 rs4073. Development of septal congenital heart defects is associated with general contribution of polymorphic variants of the TLR genes and cytokines to this pathology. A missense mutation of the TLR4 rs4986790 gene and a TNF rs1799964 mutation leading to increased synthesis of the TNFα molecule, may have a combined effect on this process. In general, contribution of TLR and cytokine genes interactions to the CHD development seems to be not significant.