Differential Expression of IFN-γ, IL-10, TLR1, and TLR2 and Their Potential Effects on Downgrading Leprosy Reaction and Erythema Nodosum Leprosum.

Leprosy reactions are acute immunological events that occur during the evolution of chronic infectious disease causing neural damage and disabilities. A study using blood samples of 17 leprosy reaction patients and 17 reaction-free was carried out by means of associations between antigens, receptors, and expression of cytokines, using path analysis providing new insights into the immunological mechanisms involved in triggering leprosy reactions. Toll-like receptors (TLR) such as TLR1 and TLR2, presented balanced expression in the reaction-free multibacillary (MB) group (TLR1: 1.01 ± 0.23, TLR2: 1.22 ± 0.18; p = 0.267). On the other hand, downgrading type 1 reaction (T1R) (TLR1: 1.24 ± 0.17, TLR2: 2.88 ± 0.37; p = 0.002) and erythema nodosum leprosum (ENL) (TLR1: 1.93 ± 0.17, TLR2: 2.81 ± 0.15; p = 0.004) revealed an unbalance in relation to the expression of these receptors. When the path analysis was approached, it was noted that interleukin 10 (IL-10) expression showed a dependence relation with phenolic glycolipid I (PGL-I) in downgrading T1R (direct effect = 0.503 > residual effect = 0.364), whereas in ENL, such relationship occurred with lipoarabinomannan (LAM) (direct effect = 0.778 > residual effect = 0.280). On the contrary, in the reaction-free leprosy group, interferon-gamma (IFN-γ) levels were dependent on the association between TLR2 and TLR1 (0.8735). The high TLR2 expression associated with IL-10 levels, in the leprosy reaction groups, may be hypothetically related to the formation of TLR2/2 homodimers and/or TLR2/6 heterodimers linked to evasion mechanisms in downgrading reactions and pathophysiology of ENL.

Evaluation of Toll-like receptor expression profile in patients with psoriasis vulgaris.

TLRs are thought to play a role in the pathophysiology of such dermatological diseases as leprosy, acne and psoriasis. The study included 20 patients with plaque psoriasis, as well as 20 healthy age- and gender-matched control subjects. Real-time polymerase chain reaction evaluation was made of the messenger RNA expression of TLRs 1-10 in lesional tissue and peripheral blood mononuclear cell samples in psoriasis patients. TLR 3, 5, 6, 7, 9 and 10 lesional tissue mRNA expressions were increased significantly when compared to the expression levels in the PBMCs of the same patients (p = 0.0082, p = 0.0176, p = 0.0239, p = 0.0261, p = 0.0223, p = 0.0206). A comparison of the TLR expression in the PBMCs of healthy subjects and the PBMCs of patients with psoriasis showed a significant increase in the TLR 1, 8 and 10 mRNA expressions in the patient group (p < 0.0001, p < 0.0001, p = 0.0035). The TLR 5 mRNA expression was significantly higher in the control group than in the patient group (p = 0.0037). To the best of our knowledge, this is the first study in literature to evaluate mRNA TLR expression levels in the lesional tissue and PBMCs of patients with psoriasis.

The TLR1 gene is associated with higher protection from leprosy in women.

Leprosy is an infectious disease with a complex genetic and immunological background. Polymorphisms in genes that encode cytokines and receptors involved in the immune response, such as the Toll-like receptor 1 (TLR1), may be associated with disease risk. We hypothesized that polymorphisms in innate immunity genes confer susceptibility to leprosy that differs between women and men. In this study, we investigate sex differences in the association between a single nucleotide polymorphism (SNP) in TLR1 and Nucleotide-binding oligomerization domain containing 2 (NOD2) genes and leprosy susceptibility in 256 clinically classified leprosy patients and 233 control subjects in a Brazilian population. Our results showed no association between the SNP rs8057341 in NOD2 and leprosy in this population. However, the heterozygous genotype of the TLR1 SNP (rs4833095) showed a statistically significant association in women (OR = 0.54, P = 0.02). Our findings suggest that the TLR1 polymorphism was associated with an increased protection from leprosy in women.

Polymorphisms in genes TLR1, 2 and 4 are associated with differential cytokine and chemokine serum production in patients with leprosy

BACKGROUND Leprosy or hansen’s disease is a spectral disease whose clinical forms mostly depends on host’s immune and genetic factors. Different Toll-like receptors (TLR) variants have been described associated with leprosy, but with some lack of replication across different populations. OBJECTIVES To evaluate the role of polymorphisms in genes TLR1, TLR2 and TLR4 and susceptibility to leprosy in a genetic case control study; to verify the association between genotypes of these markers and the immunological profile in the serum of patients with leprosy. METHODS Pre-designed TaqMan® assays were used to genotype markers at TLR1 (rs4833095, rs5743551), TLR2 (rs7656411, rs3804099) and TLR4 (rs1927914, rs1927911). A panel of cytokines and chemokines was accessed by enzime-linked immunosorbent assay (ELISA) test in the serum of a subgroup of patients with and without leprosy reactions. FINDINGS Our results show an association between the T allele of rs3804099 at the TLR2 gene and increased risk for leprosy per se [Odds ratio (OR) = 1.296, p = 0,022]. In addition, evaluating the association between different genotypes of the TLR1, 2 and 4 markers and cytokine/chemokine serological levels, IL-17 appears as an immunological marker regulated by the polymorphism of the three TLR genes evaluated, whereas different TLR1 genotypes were associated with differential production of IL-12p40 and MCP-1(CCL2). Furthermore, other relevant serum markers such as CXCL-10 and IL-6 seemed to be regulated by TLR2 variants and IL-1β was related to TLR4 genotypes. MAIN CONCLUSIONS All together our data points that the tested TLR markers may have a regulatory role in the immunity against Mycobacterium leprae, by driving the host’s production of key cytokines and chemokines involved in the pathogenesis of this disease.

Polymorphisms in genes TLR1, 2 and 4 are associated with differential cytokine and chemokine serum production in patients with leprosy.

BACKGROUND: Leprosy or hansen's disease is a spectral disease whose clinical forms mostly depends on host's immune and genetic factors. Different Toll-like receptors (TLR) variants have been described associated with leprosy, but with some lack of replication across different populations. OBJECTIVES: To evaluate the role of polymorphisms in genes TLR1, TLR2 and TLR4 and susceptibility to leprosy in a genetic case control study; to verify the association between genotypes of these markers and the immunological profile in the serum of patients with leprosy. METHODS: Pre-designed TaqMan® assays were used to genotype markers at TLR1 (rs4833095, rs5743551), TLR2 (rs7656411, rs3804099) and TLR4 (rs1927914, rs1927911). A panel of cytokines and chemokines was accessed by enzime-linked immunosorbent assay (ELISA) test in the serum of a subgroup of patients with and without leprosy reactions. FINDINGS: Our results show an association between the T allele of rs3804099 at the TLR2 gene and increased risk for leprosy per se [Odds ratio (OR) = 1.296, p = 0,022]. In addition, evaluating the association between different genotypes of the TLR1, 2 and 4 markers and cytokine/chemokine serological levels, IL-17 appears as an immunological marker regulated by the polymorphism of the three TLR genes evaluated, whereas different TLR1 genotypes were associated with differential production of IL-12p40 and MCP-1(CCL2). Furthermore, other relevant serum markers such as CXCL-10 and IL-6 seemed to be regulated by TLR2 variants and IL-1ß was related to TLR4 genotypes. MAIN CONCLUSIONS: All together our data points that the tested TLR markers may have a regulatory role in the immunity against Mycobacterium leprae, by driving the host's production of key cytokines and chemokines involved in the pathogenesis of this disease.

Red squirrels in the British Isles are infected with leprosy bacilli.

Leprosy, caused by infection with Mycobacterium leprae or the recently discovered Mycobacterium lepromatosis, was once endemic in humans in the British Isles. Red squirrels in Great Britain (Sciurus vulgaris) have increasingly been observed with leprosy-like lesions on the head and limbs. Using genomics, histopathology, and serology, we found M. lepromatosis in squirrels from England, Ireland, and Scotland, and M. leprae in squirrels from Brownsea Island, England. Infection was detected in overtly diseased and seemingly healthy animals. Phylogenetic comparisons of British and Irish M. lepromatosis with two Mexican strains from humans show that they diverged from a common ancestor around 27,000 years ago, whereas the M. leprae strain is closest to one that circulated in Medieval England. Red squirrels are thus a reservoir for leprosy in the British Isles.

Haplotype structure and positive selection at TLR1.

Toll-like receptor 1, when dimerized with Toll-like receptor 2, is a cell surface receptor that, upon recognition of bacterial lipoproteins, activates the innate immune system. Variants in TLR1 associate with the risk of a variety of medical conditions and diseases, including sepsis, leprosy, tuberculosis, and others. The foremost of these is rs5743618 c.2079T>G(p.(Ile602Ser)), the derived allele of which is associated with reduced risk of sepsis, leprosy, and other diseases. Interestingly, 602Ser, which shows signatures of selection, inhibits TLR1 surface trafficking and subsequent activation of NFκB upon recognition of a ligand. This suggests that reduced TLR1 activity may be beneficial for human health. To better understand TLR1 variation and its link to human health, we have typed all 7 high-frequency missense variants (>5% in at least one population) along with 17 other variants in and around TLR1 in 2548 individuals from 56 populations from around the globe. We have also found additional signatures of selection on missense variants not associated with rs5743618, suggesting that there may be multiple functional alleles under positive selection in this gene.

Toll-like receptor 1 N248S single-nucleotide polymorphism is associated with leprosy risk and regulates immune activation during mycobacterial infection.

Conflicting findings about the association between leprosy and TLR1 variants N248S and I602S have been reported. Here, we performed case-control and family based studies, followed by replication in 2 case-control populations from Brazil, involving 3162 individuals. Results indicated an association between TLR1 248S and leprosy in the case-control study (SS genotype odds ratio [OR], 1.81; P = .004) and the family based study (z = 2.02; P = .05). This association was consistently replicated in other populations (combined OR, 1.51; P < .001), corroborating the finding that 248S is a susceptibility factor for leprosy. Additionally, we demonstrated that peripheral blood mononuclear cells (PBMCs) carrying 248S produce a lower tumor necrosis factor/interleukin-10 ratio when stimulated with Mycobacterium leprae but not with lipopolysaccharide or PAM3cysK4. The same effect was observed after infection of PBMCs with the Moreau strain of bacillus Calmette-Guerin but not after infection with other strains. Finally, molecular dynamics simulations indicated that the Toll-like receptor 1 structure containing 248S amino acid is different from the structure containing 248N. Our results suggest that TLR1 248S is associated with an increased risk for leprosy, consistent with its hypoimmune regulatory function.

Toll-like receptor 1 N248S single-nucleotide polymorphism is associated with leprosy risk and regulates immune activation during mycobacterial infection

Conflicting findings about the association between leprosy and TLR1 variants N248S and I602S have been reported. Here, we performed case-control and family based studies, followed by replication in 2 case-control populations from Brazil, involving 3162 individuals. Results indicated an association between TLR1 248S and leprosy in the case-control study (SS genotype odds ratio [OR], 1.81; P = .004) and the family based study (z = 2.02; P = .05). This association was consistently replicated in other populations (combined OR, 1.51; P < .001), corroborating the finding that 248S is a susceptibility factor for leprosy. Additionally, we demonstrated that peripheral blood mononuclear cells (PBMCs) carrying 248S produce a lower tumor necrosis factor/interleukin-10 ratio when stimulated with Mycobacterium leprae but not with lipopolysaccharide or PAM3cysK4. The same effect was observed after infection of PBMCs with the Moreau strain of bacillus Calmette-Guerin but not after infection with other strains. Finally, molecular dynamics simulations indicated that the Toll-like receptor 1 structure containing 248S amino acid is different from the structure containing 248N. Our results suggest that TLR1 248S is associated with an increased risk for leprosy, consistent with its hypoimmune regulatory function.

Differential trafficking of TLR1 I602S underlies host protection against pathogenic mycobacteria.

We recently identified I602S as a frequent single-nucleotide polymorphism of human TLR1 that greatly inhibits cell surface trafficking, confers hyporesponsiveness to TLR1 agonists, and protects against the mycobacterial diseases leprosy and tuberculosis. Because mycobacteria are known to manipulate the TLR system to their advantage, we hypothesize that the hyporesponsive 602S variant may confer protection by enabling the host to overcome this immune subversion. We report that primary human monocytes and macrophages from homozygous TLR1 602S individuals are resistant to mycobacterial-induced downregulation of macrophage MHC class II, CD64, and IFN-γ responses compared with individuals who harbor the TLR1 602I variant. Additionally, when challenged with mycobacterial agonists, macrophages from TLR1 602S/S individuals resist induction of host arginase-1, an enzyme that depletes cellular arginine stores required for the production of antimicrobial reactive nitrogen intermediates. The differences in cell activation mediated by TLR1 602S and TLR1 602I are observed upon stimulation with soluble mycobacterial-derived agonists but not with whole mycobacterial cells. Taken together, these results suggest that the TLR1 602S variant protects against mycobacterial disease by preventing soluble mycobacterial products, perhaps released from granulomas, from disarming myeloid cells prior to their encounter with whole mycobacteria.

Leprosy and the adaptation of human toll-like receptor 1.

Leprosy is an infectious disease caused by the obligate intracellular pathogen Mycobacterium leprae and remains endemic in many parts of the world. Despite several major studies on susceptibility to leprosy, few genomic loci have been replicated independently. We have conducted an association analysis of more than 1,500 individuals from different case-control and family studies, and observed consistent associations between genetic variants in both TLR1 and the HLA-DRB1/DQA1 regions with susceptibility to leprosy (TLR1 I602S, case-control P = 5.7 x 10(-8), OR = 0.31, 95% CI = 0.20-0.48, and HLA-DQA1 rs1071630, case-control P = 4.9 x 10(-14), OR = 0.43, 95% CI = 0.35-0.54). The effect sizes of these associations suggest that TLR1 and HLA-DRB1/DQA1 are major susceptibility genes in susceptibility to leprosy. Further population differentiation analysis shows that the TLR1 locus is extremely differentiated. The protective dysfunctional 602S allele is rare in Africa but expands to become the dominant allele among individuals of European descent. This supports the hypothesis that this locus may be under selection from mycobacteria or other pathogens that are recognized by TLR1 and its co-receptors. These observations provide insight into the long standing host-pathogen relationship between human and mycobacteria and highlight the key role of the TLR pathway in infectious diseases.

Polymorphism N248S in the human Toll-like receptor 1 gene is related to leprosy and leprosy reactions.

We investigated the association between a polymorphism of a key innate immunity receptor, Toll-like receptor 1 (TLR1) N248S, and susceptibility to leprosy and its clinical presentation. TLR1 N248S has been shown elsewhere to diminish TLR1 signaling and subsequent leprosy disease. The homozygous genotype SS was more frequent (P=.012) and the heterozygous SN genotype was less frequent (P=.015) in patients with leprosy than in control subjects. Additional observed differences in allelic frequency in patients who experienced reversal reactions and/or erythema nodosum leprosum reactions indicates that altered TLR1 function, or at least a TLR1 N248S-linked trait, may affect the progression from infection to disease as well as the disease course and the risk of debilitating reactional episodes in this population.

Human TLR1 deficiency is associated with impaired mycobacterial signaling and protection from leprosy reversal reaction.

Toll-like receptors (TLRs) are important regulators of the innate immune response to pathogens, including Mycobacterium leprae, which is recognized by TLR1/2 heterodimers. We previously identified a transmembrane domain polymorphism, TLR1_T1805G, that encodes an isoleucine to serine substitution and is associated with impaired signaling. We hypothesized that this TLR1 SNP regulates the innate immune response and susceptibility to leprosy. In HEK293 cells transfected with the 1805T or 1805G variant and stimulated with extracts of M. leprae, NF-kappaB activity was impaired in cells with the 1805G polymorphism. We next stimulated PBMCs from individuals with different genotypes for this SNP and found that 1805GG individuals had significantly reduced cytokine responses to both whole irradiated M. leprae and cell wall extracts. To investigate whether TLR1 variation is associated with clinical presentations of leprosy or leprosy immune reactions, we examined 933 Nepalese leprosy patients, including 238 with reversal reaction (RR), an immune reaction characterized by a Th1 T cell cytokine response. We found that the 1805G allele was associated with protection from RR with an odds ratio (OR) of 0.51 (95% CI 0.29-0.87, p = 0.01). Individuals with 1805 genotypes GG or TG also had a reduced risk of RR in comparison to genotype TT with an OR of 0.55 (95% CI 0.31-0.97, p = 0.04). To our knowledge, this is the first association of TLR1 with a Th1-mediated immune response. Our findings suggest that TLR1 deficiency influences adaptive immunity during leprosy infection to affect clinical manifestations such as nerve damage and disability.

Cutting edge: A common polymorphism impairs cell surface trafficking and functional responses of TLR1 but protects against leprosy.

TLRs constitute an essential family of pattern recognition molecules that, through direct recognition of conserved microbial components, initiate inflammatory responses following infection. In this role, TLR1 enables host responses to a variety of bacteria, including pathogenic species of mycobacteria. In this study, we report that I602S, a common single nucleotide polymorphism within TLR1, is associated with aberrant trafficking of the receptor to the cell surface and diminished responses of blood monocytes to bacterial agonists. When expressed in heterologous systems, the TLR1 602S variant, but not the TLR1 602I variant, exhibits the expected deficiencies in trafficking and responsiveness. Among white Europeans, the 602S allele represents the most common single nucleotide polymorphism affecting TLR function identified to date. Surprisingly, the 602S allele is associated with a decreased incidence of leprosy, suggesting that Mycobacterium leprae subverts the TLR system as a mechanism of immune evasion.