NAcht Leucine-rich-repeat Protein 1 Research Articles

Expression of Th17- and Treg-specific transcription factors in vitiligo patients.

Vitiligo is mainly considered an autoimmune skin disease as the number of IL-17 producing Th17 cells, involved in the development of autoimmune and inflammatory pathologies, increased in vitiligo skin. T regulatory cells (Tregs) seem to be altered during the disease. Thus, there must be some upstream molecular factors that regulate the cellular response to apoptotic and inflammatory stimuli. To investigate the expression of Th17- and Treg-specific transcription factors in PBMCs and to evaluate the correlation between these transcription factors and cytokines in vitiligo patients. We investigated 30 active NSV patients for Th17- and Treg-specific transcription factors RORγt (retinoic acid-related orphan receptor gamma t), FOXP3 (forkhead/winged helix), HELIOS, EOS, and IRF4 (Interferon Regulatory Factor 4) as well as apoptotic marker NALP1 (NACHT-leucine-rich-repeat protein 1) in PBMCs with RT-qPCR. Immunostaining was done for transcription factors and cytokines on skin sections. The mRNA level of FOXP3 was significantly lower in patients (0.76 fold, P<0.001), whereas RORγt was slight but notsignificantly increased (0.76 fold, P=0.06). Furthermore, NALP1 in lymphocytes was found to be increased in patients (0.69 fold, P<0.01). The immunostaining results revealed increased expression of RORγ, IL-17A, NALP1, and IL-1β in vitiligo skin when compared to normal healthy skin. Reduced FOXP3/RORγt mRNA ratio suggests thriving of the Th17 cell population in PBMCs of vitiligo patients. Increased NALP1 levels indicate the existence of an apoptotic phenomenon which correlates with the increased expression of IL-1β in vitiligo pathogenesis.

Curcumin ameliorates neuropathic pain by down-regulating spinal IL-1β via suppressing astroglial NALP1 inflammasome and JAK2-STAT3 signalling.

Curcumin has been shown to possess strong anti-inflammatory activity in many diseases. It has been demonstrated that the janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) cascade and the NAcht leucine-rich-repeat protein 1 (NALP1) inflammasome are important for the synthesis of Pro-Interleukin (IL)-1β and the processing of the inactive protein to its mature form, which plays an active role in the pathogenesis of neuropathic pain. The present study showed that repeated intraperitoneal injection of curcumin ameliorated SNI-induced mechanical and cold allodynia in a dose-dependent manner and inhibited the elevation of spinal mature IL-1β protein levels. Additionally, repeated curcumin treatment significantly inhibited the aggregation of the NALP1 inflammasome and the activation of the JAK2-STAT3 cascade in spinal astrocytes. Furthermore, the genetic down-regulation of NALP1 inflammasome activation by NALP1 siRNA and the pharmacological inhibition of the JAK2-STAT3 cascade by AG490 markedly inhibited IL-1β maturation and Pro-IL-1β synthesis, respectively, and reduced SNI-induced pain hypersensitivity. Our results suggest that curcumin attenuated neuropathic pain and down-regulated the production of spinal mature IL-1β by inhibiting the aggregation of NALP1 inflammasome and the activation of the JAK2-STAT3 cascade in astrocytes.

Single-cell imaging of inflammatory caspase dimerization reveals differential recruitment to inflammasomes.

The human inflammatory caspases, including caspase-1, -4, -5 and -12, are considered as key regulators of innate immunity protecting from sepsis and numerous inflammatory diseases. Caspase-1 is activated by proximity-induced dimerization following recruitment to inflammasomes but the roles of the remaining inflammatory caspases in inflammasome assembly are unclear. Here, we use caspase bimolecular fluorescence complementation to visualize the assembly of inflammasomes and dimerization of inflammatory caspases in single cells. We observed caspase-1 dimerization induced by the coexpression of a range of inflammasome proteins and by lipospolysaccharide (LPS) treatment in primary macrophages. Caspase-4 and -5 were only dimerized by select inflammasome proteins, whereas caspase-12 dimerization was not detected by any investigated treatment. Strikingly, we determined that certain inflammasome proteins could induce heterodimerization of caspase-1 with caspase-4 or -5. Caspase-5 homodimerization and caspase-1/-5 heterodimerization was also detected in LPS-primed primary macrophages in response to cholera toxin subunit B. The subcellular localization and organization of the inflammasome complexes varied markedly depending on the upstream trigger and on which caspase or combination of caspases were recruited. Three-dimensional imaging of the ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain)/caspase-1 complexes revealed a large spherical complex of ASC with caspase-1 dimerized on the outer surface. In contrast, NALP1 (NACHT leucine-rich repeat protein 1)/caspase-1 complexes formed large filamentous structures. These results argue that caspase-1, -4 or -5 can be recruited to inflammasomes under specific circumstances, often leading to distinctly organized and localized complexes that may impact the functions of these proteases.

Involvement of the spinal NALP1 inflammasome in neuropathic pain and aspirin-triggered-15-epi-lipoxin A4 induced analgesia

Neuroinflammation plays an important role in nerve-injury-induced neuropathic pain, but the explicit molecular mechanisms of neuroinflammation in neuropathic pain remain unclear. As one of the most critical inflammatory cytokines, interleukin-1β (IL-1β) has been regarded as broadly involved in the pathology of neuropathic pain. The inflammasome caspase-1 platform is one primary mechanism responsible for the maturation of IL-1β. Lipoxins, a type of endogenous anti-inflammatory lipid, have proved to be effective in relieving neuropathic pain behaviors. The present study was designed to examine whether the inflammasome caspase-1 IL-1β platform is involved in chronic constriction injury (CCI)-induced neuropathic pain and in lipoxin-induced analgesia. After rats were subjected to the CCI surgery, mature IL-1β was significantly increased in the ipsilateral spinal cord, and the inflammasome platform consisting of NALP1 (NAcht leucine-rich-repeat protein 1), caspase-1 and ASC (apoptosis-associated speck-like protein containing a caspase-activating recruitment domain) was also activated in spinal astrocytes and neurons, especially at the superficial laminae of the spinal dorsal horn; The aspirin-triggered-15-epi-lipoxin A4 (ATL), which shares the potent actions of the endogenous lipoxins, was administered to the CCI rats. Repeated intrathecal injection with ATL markedly attenuated the CCI-induced thermal hyperalgesia and significantly inhibited NALP1 inflammasome activation, caspase-1 cleavage, and IL-1β maturation. These results suggested that spinal NALP1 inflammasome was involved in the CCI-induced neuropathic pain and that the analgesic effect of ATL was associated with suppressing NALP1 inflammasome activation.

Polymorphisms in NLRP1 gene and susceptibility to autoimmune thyroid disease

The autoimmune thyroid disorders, or AITDs, comprise 2 related disorders, Graves' disease and Hashimoto thyroiditis. In AITD, immune system produces antibodies against autothyroid antigens. The etiology of AITDs involves a complex interaction between genetic predisposing factors and environmental triggering factors. Variations in NACHT leucine-rich repeat protein 1(NLRP1) gene a key regulator of the innate immunity have been shown to confer risk for vitiligo and several autoimmune diseases. In this study we hypothesize that variants in NLRP1 gene might be involved in the susceptibility to autoimmune thyroid disease. Five single nucleotide polymorphisms (SNPs) in NLRP1 were genotyped in 207 AITD patients and 220 normal controls. We found that NLRP1 rs12150220 T allele (OR = 1.273, 95% CI: 0.971–1.670, p = 0.040) and NLRP1 rs2670660 G allele (OR = 1.264, 95% CI: 0.965–1.656, p = 0.044) were significantly associated with AITD compared with controls. These results suggest that NLRP1 may be involved in the pathogenesis of AITD.

Genetic Predictors of Glucocorticoid Response in Pediatric Patients With Inflammatory Bowel Diseases

Glucocorticoids (GCs) are used in moderate-to-severe inflammatory bowel diseases (IBD) but their effect is often unpredictable. To determine the influence of 4 polymorphisms in the GC receptor [nuclear receptor subfamily 3, group C, member 1 (NR3C1)], interleukin-1β (IL-1β), and NACHT leucine-rich-repeat protein 1 (NALP1) genes, on the clinical response to steroids in pediatric patients with IBD. One hundred fifty-four young IBD patients treated with GCs for at least 30 days and with a minimum follow-up of 1 year were genotyped. The polymorphisms considered are the BclI in the NR3C1 gene, C-511T in IL-1β gene, and Leu155His and rs2670660/C in NALP1 gene. Patients were grouped as responder, dependant, and resistant to GCs. The relation between GC response and the genetic polymorphisms considered was examined using univariate, multivariate, and Classification and Regression Tree (CART) analysis. Univariate analysis showed that BclI polymorphism was more frequent in responders compared with dependant patients (P=0.03) and with the combined dependant and resistant groups (P=0.02). Moreover, the NALP1 Leu155His polymorphism was less frequent in the GC responsive group compared with resistant (P=0.0059) and nonresponder (P=0.02) groups. Multivariate analysis comparing responders and nonresponders confirmed an association between BclI mutated genotype and steroid response (P=0.030), and between NALP1 Leu155His mutant variant and nonresponders (P=0.033). An association between steroid response and male sex was also observed (P=0.034). In addition, Leu155His mutated genotype was associated with steroid resistance (P=0.034). Two CART analyses supported these findings by showing that BclI and Leu155His polymorphisms had the greatest effect on steroid response (permutation P value=0.046). The second CART analysis also identified age of disease onset and male sex as important variables affecting response. These results confirm that genetic and demographic factors may affect the response to GCs in young patients with IBD and strengthen the importance of studying high-order interactions for predicting response.

Update on childhood vitiligo

This review addresses recent changes in the understanding and the treatment of vitiligo vulgaris. Two target genes for vitiligo have been identified, NACHT-leucine-rich-repeat protein-1 (NALP1), part of the inflammasome cascade, and tyrosinase, the enzyme that produces melanin. Identification of reactive oxidation species has furthered the understanding of melanocyte destruction. Comorbid autoimmune disease, including thyroid autoimmunity seen in 25%, is genetically linked to generalized vitiligo and is noted in both childhood vitiligo patients and their families. Screening for vitamin deficiencies and concurrent autoimmunity can be beneficial to the overall health of the child with vitiligo. About half of all vitiligo vulgaris patients have onset of their illness during childhood, causing increased psychological stress during the formative years. Fortunately, therapy has improved as well, with the development of newer topical agents for vitiligo, including topical calcineurin inhibitors; new topical combinations such as topical corticosteroids and calcipotriene; and new technological advances including narrowband ultraviolet B and excimer laser. A cyclic approach to therapy should be used wherein topical agents are altered every 6-8 months and technology is used as an alternative after achievement of maximal topical response. With cyclic therapy and early disease intervention, good cosmetic outcomes may be achievable, particularly in localized cases.

Initiation and Progression of Axonopathy in Experimental Autoimmune Encephalomyelitis

Axonal loss is the principal cause of chronic disability in multiple sclerosis and experimental autoimmune encephalomyelitis (EAE). In C57BL/6 mice with EAE induced by immunization with myelin oligodendrocyte glycoprotein peptide 35-55, the first evidences of axonal damage in spinal cord were in acute subpial and perivascular foci of infiltrating neutrophils and lymphocytes and included intra-axonal accumulations of the endovesicular Toll-like receptor TLR8, and the inflammasome protein NAcht leucine-rich repeat protein 1 (NALP1). Later in the course of this illness, focal inflammatory infiltrates disappeared from the spinal cord, but there was persistent activation of spinal cord innate immunity and progressive, bilaterally symmetric loss of small-diameter corticospinal tract axons. These results support the hypothesis that both contact-dependent and paracrine interactions of systemic inflammatory cells with axons and an innate immune-mediated neurodegenerative process contribute to axonal loss in this multiple sclerosis model.

A coding polymorphism in NALP1 confers risk for autoimmune Addison's disease and type 1 diabetes

Variants in the gene encoding NACHT leucine-rich-repeat protein 1 (NALP1), an important molecule in innate immunity, have recently been shown to confer risk for vitiligo and associated autoimmunity. We hypothesized that sequence variants in this gene may be involved in susceptibility to a wider spectrum of autoimmune diseases. Investigating large patient cohorts from six different autoimmune diseases, that is autoimmune Addison's disease (n=333), type 1 diabetes (n=1086), multiple sclerosis (n=502), rheumatoid arthritis (n=945), systemic lupus erythematosus (n=156) and juvenile idiopathic arthritis (n=505), against 3273 healthy controls, we analyzed four single nucleotide polymorphisms (SNPs) in NALP1. The major allele of the coding SNP rs12150220 revealed significant association with autoimmune Addison's disease compared with controls (OR=1.25, 95% CI: 1.06-1.49, P=0.007), and with type 1 diabetes (OR=1.15, 95% CI: 1.04-1.27, P=0.005). Trends toward the same associations were seen in rheumatoid arthritis, systemic lupus erythematosus and, although less obvious, multiple sclerosis. Patients with juvenile idiopathic arthritis did not show association with NALP1 gene variants. The results indicate that NALP1 and the innate immune system may be implicated in the pathogenesis of many autoimmune disorders, particularly organ-specific autoimmune diseases.

A Molecular Platform in Neurons Regulates Inflammation after Spinal Cord Injury

Vigorous immune responses are induced in the immune privileged CNS by injury and disease, but the molecular mechanisms regulating innate immunity in the CNS are poorly defined. The inflammatory response initiated by spinal cord injury (SCI) involves activation of interleukin-1beta (IL-1beta) that contributes to secondary cell death. In the peripheral immune response, the inflammasome activates caspase-1 to process proinflammatory cytokines, but the regulation of trauma-induced inflammation in the CNS is not clearly understood. Here we show that a molecular platform [NALP1 (NAcht leucine-rich-repeat protein 1) inflammasome] consisting of caspase-1, caspase-11, ASC (apoptosis-associated speck-like protein containing a caspase-activating recruitment domain), and NALP1 is expressed in neurons of the normal rat spinal cord and forms a protein assembly with the X-linked inhibitor of apoptosis protein (XIAP). Moderate cervical contusive SCI induced processing of IL-1beta, IL-18, activation of caspase-1, cleavage of XIAP, and promoted assembly of the multiprotein complex. Anti-ASC neutralizing antibodies administered to injured rats entered spinal cord neurons via a mechanism that was sensitive to carbenoxolone. Therapeutic neutralization of ASC reduced caspase-1 activation, XIAP cleavage, and interleukin processing, resulting in significant tissue sparing and functional improvement. Thus, rat spinal cord neurons contain a caspase-1, pro-ILbeta, and pro-IL-18 activating complex different from the human NALP1 inflammasome that constitutes an important arm of the innate CNS inflammatory response after SCI.

Expression of NALP1 in cerebellar granule neurons stimulates apoptosis

NAcht Leucine-rich-repeat Protein 1 (NALP1) contains a putative nucleotide binding site, a region of leucine-rich repeats, and death domain folds at both termini providing protein/protein association functions such as caspase recruitment. We report here that NALP1 gene expression was induced in primary cerebellar granule neurons (CGN) upon injury. Up-regulation of NALP1 was also observed in a model of transient focal ischemia induced by middle cerebral artery occlusion. We investigated the biological consequence of over-expression of NALP1 in both HeLa cells and in CGN. Expression of recombinant NALP1 stimulated cell death in both HeLa cells and CGN by an apoptotic mechanism, demonstrated by the induction of apoptotic nuclear morphology and activation of the apoptotic enzyme caspase-3. Also described here are studies on the mechanism of action studies including deletion analyses and investigations of nucleotide binding, which begin to elucidate a regulatory function for NALP1 in neuronal apoptosis.