Ancient familial Mediterranean fever mutations in human pyrin and resistance to Yersinia pestis.

Familial Mediterranean fever (FMF) is an autoinflammatory disease caused by homozygous or compound heterozygous gain-of-function mutations in MEFV, which encodes pyrin, an inflammasome protein. Heterozygous carrier frequencies for multiple MEFV mutations are high in several Mediterranean populations, suggesting that they confer selective advantage. Among 2,313 Turkish people, we found extended haplotype homozygosity flanking FMF-associated mutations, indicating evolutionarily recent positive selection of FMF-associated mutations. Two pathogenic pyrin variants independently arose >1,800 years ago. Mutant pyrin interacts less avidly with Yersinia pestis virulence factor YopM than with wild-type human pyrin, thereby attenuating YopM-induced interleukin (IL)-1ß suppression. Relative to healthy controls, leukocytes from patients with FMF harboring homozygous or compound heterozygous mutations and from asymptomatic heterozygous carriers released heightened IL-1ß specifically in response to Y. pestis. Y. pestis-infected MefvM680I/M680I FMF knock-in mice exhibited IL-1-dependent increased survival relative to wild-type knock-in mice. Thus, FMF mutations that were positively selected in Mediterranean populations confer heightened resistance to Y. pestis.

InflammamiRs in focus: Delivery strategies and therapeutic approaches.

microRNAs (miRNAs) are small non-protein-coding RNAs which are essential regulators of host genome expression at the post-transcriptional level. There is evidence of dysregulated miRNA expression patterns in a wide variety of diseases, such as autoimmune and inflammatory conditions. These miRNAs have been termed "inflammamiRs." When working with miRNAs, the method followed, the approach to treat or diagnosis, and the selected biological material are very crucial. Demonstration of the role of miRNAs in particular disease phenotypes facilitates their evaluation as potential and effective therapeutic tools. A growing number of reports suggest the significant utility of miRNAs and other small RNA drugs in clinical medicine. Most miRNAs seem promising therapeutic options, but some features associated with miRNA therapy like off-target effect, effective dosage, or differential delivery methods, mainly caused by the short target's sequence, make miRNA therapies challenging. In this review, we aim to discuss some of the inflammamiRs in diseases associated with inflammatory pathways and the challenge of identifying the most potent therapeutic candidates and provide a perspective on achieving safe and targeted delivery of miRNA therapeutics. We also discuss the status of inflammamiRs in clinical trials.

Anti-LAMP-2 Antibody Seropositivity in Children with Primary Systemic Vasculitis Affecting Medium- and Large-Sized Vessels.

Chronic primary systemic vasculitis (PSV) comprises a group of heterogeneous diseases that are broadly classified by affected blood vessel size, clinical traits and the presence (or absence) of anti-neutrophil cytoplasmic antibodies (ANCA) against proteinase 3 (PR3) and myeloperoxidase (MPO). In small vessel vasculitis (SVV), ANCA are not present in all patients, and they are rarely detected in patients with vasculitis involving medium (MVV) and large (LVV) blood vessels. Some studies have demonstrated that lysosome-associated membrane protein-2 (LAMP-2/CD107b) is a target of ANCA in SVV, but its presence and prognostic value in childhood MVV and LVV is not known. This study utilized retrospective sera and clinical data obtained from 90 children and adolescents with chronic PSV affecting small (SVV, n = 53), medium (MVV, n = 16), and large (LVV, n = 21) blood vessels. LAMP-2-ANCA were measured in time-of-diagnosis sera using a custom electrochemiluminescence assay. The threshold for seropositivity was established in a comparator cohort of patients with systemic autoinflammatory disease. The proportion of LAMP-2-ANCA-seropositive individuals and sera concentrations of LAMP-2-ANCA were assessed for associations with overall and organ-specific disease activity at diagnosis and one-year follow up. This study demonstrated a greater time-of-diagnosis prevalence and sera concentration of LAMP-2-ANCA in MVV (52.9% seropositive) and LVV (76.2%) compared to SVV (45.3%). Further, LAMP-2-ANCA-seropositive individuals had significantly lower overall, but not organ-specific, disease activity at diagnosis. This did not, however, result in a greater reduction in disease activity or the likelihood of achieving inactive disease one-year after diagnosis. The results of this study demonstrate particularly high prevalence and concentration of LAMP-2-ANCA in chronic PSV that affects large blood vessels and is seronegative for traditional ANCA. Our findings invite reconsideration of roles for autoantigens other than MPO and PR3 in pediatric vasculitis, particularly in medium- and large-sized blood vessels.

The pyrin inflammasome aggravates inflammatory cell migration in patients with familial Mediterranean fever.

BACKGROUND: Familial Mediterranean fever (FMF) is an autoinflammatory disease caused by pathogenic variants of the MEFV gene, which encodes pyrin. Leukocyte migration to serosal sites is a key event during inflammation in FMF. The pyrin inflammasome is a multiprotein complex involved in inflammation. Here, we aimed to determine the relationship between inflammatory cell migration and the pyrin inflammasome in FMF patients. METHODS: Monocytes were isolated from blood samples collected from patients with FMF, healthy controls, and a patient with cryopyrin-associated periodic syndrome (CAPS), which served as a disease control. Inflammasome proteins were analyzed under inflammasome activation and inhibition by western blotting. Cell migration assays were performed with the isolated primary monocytes as well as THP-1 monocytes and THP-1-derived macrophages. RESULTS: When the pyrin inflammasome was suppressed, migration of monocytes from FMF patients was significantly decreased compared to the migration of monocytes from the CAPS patient and healthy controls. Cell line experiments showed a relationship between pyrin inflammasome activation and cell migration. CONCLUSIONS: These findings suggest that the increased cell migration in FMF is due to the presence of more active pyrin inflammasome. This study contributes to our understanding of the role of pyrin in inflammatory cell migration through inflammasome formation. IMPACT: The pyrin inflammasome may play a role in inflammatory cell migration. FMF patients show a pyrin inflammasome-dependent increase in inflammatory cell migration. Correlations between the pyrin inflammasome and cell migration were observed in both THP-1 monocytes and THP-1-derived macrophages.

Mitochondrial Dysfunction in Cyanotic Congenital Heart Disease: A Promising Therapeutic Approach for the Future.

Congenital heart disease (CHD) is one of the most specific and yet challenging fields of heart surgery. Apart from the known clinical approaches, including surgery, a significant scale of regenerative therapeutic options is available, which increase the number of cardiomyocytes and restore cardiac function. Although it has been revealed in recent years that mitochondrial transplantation can be used as a promising treatment option in this disease group, there is no clinical evidence for the significance of mitochondrial function in myocardial tissue of patients with CHD regarding cardiac surgery. In this study, mitochondrial morphology and function, myocardial fibrosis, and myocyte atypia were evaluated in myocardial biopsy tissue of pediatric patients with cyanotic and acyanotic CHD, five from each group. After histopathological evaluation of myocardial tissue specimens, mitochondrial morphology and network were analyzed by immunofluorescence staining using an anti-Tom20 antibody, electron transport chain complexes of myocardium were examined by cytochrome c oxidase/succinate dehydrogenase staining, and the amount of ATP was measured by bioluminescence assay. In addition, cardiac markers have been tested to be reviewed as a potential indicator for postoperative follow-up. Myocyte atypia and fibrosis were classified on a scale of 1 to 4. In this study, unlike patients with acyanotic CHD, alterations in mitochondrial network and reduction in ATP production were detected in all pediatric patients with cyanotic CHD. A statistically significant correlation was also determined between mitochondrial dysfunction and cardiac markers. These findings may be assumed as a promising pathway for evaluating the relationship between mitochondrial dysfunction and cyanotic CHD.

The Common miRNA Signatures Associated with Mitochondrial Dysfunction in Different Muscular Dystrophies.

Secondary mitochondrial damage in skeletal muscles is a common feature of different neuromuscular disorders, which fall outside the mitochondrial cytopathies. The common cause of mitochondrial dysfunction and structural changes in skeletal muscle tissue remains to be discovered. Although they are associated with different clinical, genetic, and pathologic backgrounds, the pathomechanisms underlying neuromuscular disorders might be attributed to the complex interaction and cross talk between mitochondria and the associated miRNAs. This study aimed to identify the common miRNA signatures that are associated with mitochondrial damage in different muscular dystrophies (MDs; Duchenne muscular dystrophy, megaconial congenital muscular dystrophy, Ullrich congenital muscular dystrophy, and α-dystroglycanopathy). The miRNome profiles of skeletal muscle biopsies acquired from four different MD groups and control individuals were analyzed by miRNA microarray. We identified 17 common up-regulated miRNAs in all of the tested MD groups. A specific bioinformatics approach identified 10 of these miRNAs to be specifically related to the mitochondrial pathways. Six miRNAs, miR-134-5p, miR-199a-5p, miR-382-5p, miR-409-3p, miR-497-5p, and miR-708-5p, were associated with the top four mitochondrial pathways and were thus selected as priority candidates for further validation by quantitative real-time PCR analysis. We demonstrate, for the first time, common up-regulated miRNAs that are associated with mitochondrial damage in different MD groups, therefore contributing to the pathophysiology. Our findings may open a new gate toward therapeutics.

Inflammation-related differentially expressed common miRNAs in systemic autoinflammatory disorders patients can regulate the clinical course.

OBJECTIVES: Systemic autoinflammatory diseases (SAIDs) are caused by the malfunctioning of the innate immune system factors. Clinical heterogeneity and undefined pathobiology are common phenomena among SAIDs. In this study, we aimed to assess the involvement of microRNAs in regulating these complex diseases. METHODS: The expression pattern of different miRNAs was compared between SAID patients with high autoinflammatory disease activity index (AIDAI) score and with low AIDAI score, and their role in inflammation-related pathways was investigated. Differentially expressed miRNAs were determined using the Multi Experiment Viewer (MEV) and Transcriptome Analysis Console (TAC) analysis tools using miRNA microarray. Potential targets of miRNAs were enriched for inflammation-related genes and validated using qRT-PCR analysis. RESULTS: Upon performing microarray analysis, 40 differentially expressed miRNAs were identified between mild familial Mediterranean fever (FMF) patients and severe SAID patients. Thereafter, 21 of 40 miRNAs were found to be potentially involved in inflammatory pathways, of which, 8 were further validated through qRT-PCR. The targets of these 8 miRNAs (miR-29b-3p, miR-29c-3p, miR-30e-3p, miR-130b-3p, miR-148a-3p, miR-186-5p, miR-197-3p, and miR-374b-5p) belonged to the inflammation-related genes and pathways. CONCLUSIONS: This is the first study to identify miRNAs that might be associated with a more severe disease form of monogenic autoinflammatory diseases. All these miRNAs were associated with cytokine-mediated pathways and might be used for establishing diagnostic and therapeutic methods.

Microbiome is not linked to clinical disease severity of familial Mediterranean fever in an international cohort of children.

OBJECTIVES: The severity of familial Mediterranean fever (FMF) may vary in different areas, suggesting a role for environmental factors. We analysed the composition of gut microbiota among children with FMF and healthy controls from Turkey and the USA and determined its effect on disease severity. METHODS: Children with FMF with pathogenic MEFV mutations and healthy controls from Turkey and the USA were enrolled. FMF disease activity was evaluated with the Autoinflammatory Disease Activity Index (AIDAI). Gut bacterial diversity was assessed by sequencing 16S rRNA gene libraries. RESULTS: We included 36 children from Turkey (28 patients with FMF, 8 healthy controls), and 21 patients and 6 controls from the USA. In the Turkish group, 28.6% of patients had severe disease, while 13.3% of US group patients had severe disease. As expected, we observed substantial differences between the gut microbiota of children from the two geographic regions, with Turkish patients and controls exhibiting higher relative abundances of Bacteriodia, while US patients and controls exhibited higher relative abundances of Clostridia. Alpha- and betadiversity did not differ significantly between FMF patients and controls, and neither was predictive of disease severity within each geographic region. We observed differences between FMF patients and controls in the relative abundance of some bacterial taxa at the amplicon sequence variant (ASV) level, but these differences received mixed statistical support. CONCLUSIONS: Among an international cohort of children with FMF, we did not find a strong effect of gut microbiota composition on disease severity. Other environmental or epigenetic factors may be operative.

Comorbidities in familial Mediterranean fever: analysis of 2000 genetically confirmed patients.

OBJECTIVES: FMF is the most common periodic fever syndrome, characterized by recurrent episodes of fever and serosal inflammation accompanied with high acute phase reactants. The analysis of possible comorbidities is important to understand the impact of these conditions on clinical care and whether they share a common aetiological pathway. In this study, we aimed to evaluate the comorbidities associated with FMF patients in a large genetically diagnosed cohort. METHODS: We retrospectively evaluated the medical and genetic records of FMF patients who were followed up by rheumatologists in Hacettepe University for 15 years. The FMF patients who had homozygous or compound heterozygous mutations were included in the study. Comorbidities associated with FMF were divided into three groups: (i) comorbidities directly related to FMF, (ii) comorbidities due to increased innate inflammation, and (iii) comorbidities that were regarded as being incidental. RESULTS: A total of 2000 patients with a diagnosis of FMF were enrolled in the study. Among them 636 were children (31.8%) and M694V was the most common mutation in patients with associated inflammatory conditions. The frequency of AS, Iga Vasculitis (Henoch-Schönlein purpura), juvenile idiopathic arthritis, polyarteritis nodosa, multiple sclerosis and Behçet's disease were increased in patients with FMF when compared with those in the literature. CONCLUSION: This study represents the largest genetically confirmed cohort and compares the frequencies with existing national and international figures for each disease. The increased innate immune system inflammation seen in FMF may be considered as a susceptibility factor since it predisposes to certain inflammatory conditions.

Epigenetics for Clinicians from the Perspective of Pediatric Rheumatic Diseases.

PURPOSE OF REVIEW: Epigenetics is the study of inherited phenotype changes that do not involve in alteration of DNA sequence. Epigenetic regulation can be examined under three main headings and study methodologies for all three will be discussed: DNA methylation is the addition of a methyl (CH3) group to DNA, histone modifications is a covalent post-translational modification of histones and non-coding RNAs are a group of functional RNA molecules that is copied from DNA but not converted into proteins. Epigenetic changes are being increasingly studied in the pathogenesis of most diseases including autoimmune and autoinflammatory diseases to shed light on the different phenotypes and disease courses. We have aimed to review the basic concepts in epigenetic studies and summarize the data for epigenetics in autoimmune and autoinflammatory rheumatic diseases. RECENT FINDINGS: Recent studies have assessed DNA hypomethylation in interferon-regulated genes in autoimmune diseases and in inflammatory pathways in systemic autoinflammatory diseases (SAIDs). Abnormal histone acetylation and methylation have been shown to be important in autoimmune diseases which was proven via effective targeted treatment trials against these pathways in mice models. miRNAs have an important role in the pathogenesis, and also, they can be used as diagnostic biomarkers in SAIDs (i.e., FMF, Behcet's disease) together with autoimmune diseases. Although the number of studies has increased over the years in parallel with the increase of interest in this field, we await further studies to improve the understanding and management of pediatric rheumatic diseases. Epigenetic studies in pediatric rheumatic diseases have enabled us to gain new information about disease pathogenesis, clinical heterogeneity, and prognosis. Further studies will help us define new diagnostic, prognostic, and therapeutic goals for rheumatic diseases.

Analysis of polymorphisms in the colchicine binding site of tubulin in colchicine-resistant familial Mediterranean fever patients.

Familial Mediterranean fever is a hereditary autoinflammatory syndrome. The typical treatment for the disease is colchicine. However, a subset of patients are not responsive to colchicine. In this study, polymorphisms in the colchicine-binding site of the TUBB1 gene, which encodes a tubulin isoform specific to leukocytes, were investigated in patients with colchicine-resistant disease. FMF patients who were followed in the Department of Pediatric Rheumatology at Hacettepe University were included in this study. Colchicine resistance was defined as ongoing disease activity (≥ 1 attack/month over 3 months or persistently elevated CRP) while taking the maximum tolerated dose of colchicine. A total of 62 Turkish FMF patients (42 colchicine-responsive and 20 colchicine-resistant) and a control group of healthy children were included in the study. DNA was extracted for analysis of TUBB1, and the colchicine binding site was sequenced. We did not observe A248T (rs148237574) or M257V (rs759579888), two variations that were previously associated with colchicine resistance in an in silico analysis. We did detect T274M (rs35565630), R306H (rs772479017), and R307H (rs6070697) variants in the FMF patients, but there was no statistically significant difference between the colchicine-responsive and colchicine-resistant groups. This is the first study to evaluate TUBB1 gene polymorphisms in the colchicine binding site in patients with FMF. Our data do not support the hypothesis that these polymorphisms are a possible cause of colchicine resistance in FMF patients.

Potential of miRNAs to predict and treat inflammation from the perspective of Familial Mediterranean Fever.

AIM: microRNAs (miRNAs) are small noncoding RNAs that play critical roles in physiological networks by regulating host genome expression at the post-transcriptional level. miRNAs are known to be key regulators of various biological processes in different types of immune cells, and they are known to regulate immunological functions. Differential expression of miRNAs has been documented in several diseases, including autoinflammatory and autoimmune diseases. This review aimed to focus on miRNAs and their association with autoimmune and autoinflammatory diseases. METHODS: All related literature was screened from PubMed, and we discussed the possible role of miRNAs in disease prediction and usage as therapeutic agents from the perspective of Familial Mediterranean Fever (FMF). CONCLUSIONS: FMF is an inherited autosomal recessive autoinflammatory disease caused by mutations in the MEditerranean FeVer (MEFV) gene, which encodes the protein pyrin. Recent studies have demonstrated that miRNAs may be effective in the pathogenesis of FMF and offer a potential explanation for phenotypic heterogeneity. Further understanding of the role of miRNAs in the pathogenesis of these diseases may help to identify molecular diagnostic markers, which may be important for the differential diagnosis of autoinflammatory diseases. Studies have shown that in the near future, traditional therapies in autoinflammatory diseases may be replaced with novel effective, miRNA-based therapies, such as the delivery of miRNA mimics or antagonists. These approaches may be important for predictive, preventive, and personalized medicine.

New workflow for classification of genetic variants' pathogenicity applied to hereditary recurrent fevers by the International Study Group for Systemic Autoinflammatory Diseases (INSAID).

BACKGROUND: Hereditary recurrent fevers (HRFs) are rare inflammatory diseases sharing similar clinical symptoms and effectively treated with anti-inflammatory biological drugs. Accurate diagnosis of HRF relies heavily on genetic testing. OBJECTIVES: This study aimed to obtain an experts' consensus on the clinical significance of gene variants in four well-known HRF genes: MEFV, TNFRSF1A, NLRP3 and MVK. METHODS: We configured a MOLGENIS web platform to share and analyse pathogenicity classifications of the variants and to manage a consensus-based classification process. Four experts in HRF genetics submitted independent classifications of 858 variants. Classifications were driven to consensus by recruiting four more expert opinions and by targeting discordant classifications in five iterative rounds. RESULTS: Consensus classification was reached for 804/858 variants (94%). None of the unsolved variants (6%) remained with opposite classifications (eg, pathogenic vs benign). New mutational hotspots were found in all genes. We noted a lower pathogenic variant load and a higher fraction of variants with unknown or unsolved clinical significance in the MEFV gene. CONCLUSION: Applying a consensus-driven process on the pathogenicity assessment of experts yielded rapid classification of almost all variants of four HRF genes. The high-throughput database will profoundly assist clinicians and geneticists in the diagnosis of HRFs. The configured MOLGENIS platform and consensus evolution protocol are usable for assembly of other variant pathogenicity databases. The MOLGENIS software is available for reuse at http://github.com/molgenis/molgenis; the specific HRF configuration is available at http://molgenis.org/said/. The HRF pathogenicity classifications will be published on the INFEVERS database at https://fmf.igh.cnrs.fr/ISSAID/infevers/.

Potential role of pyrin, the protein mutated in familial Mediterranean fever, during inflammatory cell migration.

Familial Mediterranean fever (FMF), the most common of the systemic autoinflammatory disorders, is caused by mutations in the MEFV (Mediterranean Fever) gene, which encodes the protein pyrin. Neutrophils, one of the major components during inflammation, are the main cell type that expresses pyrin. In response to an inflammatory stimulus, neutrophils migration to their main active site. To date, several pyrin-interacting proteins have been demonstrated to co-localise with the cytoskeletal protein actin, which is important in the process of neutrophil migration and raises the question of whether pyrin plays a role in the actin cytoskeletal network during inflammatory cell migration. In this study, we examined the possible role of pyrin during inflammatory cell migration in neutrophils. We generated a cell migration assay with neutrophils and primary neutrophils from patients. We also knocked down pyrin expression using siRNA and then performed cell migration assay. We showed co-localisation of pyrin and F-actin at the leading edge during inflammatory cell migration. In pyrin knocked down cells, we identified a significant decrease in neutrophil migration. In addition, we demonstrated a dramatic increase in migration in the neutrophils of FMF patients compared with a healthy control group. These data together provide new insight into the cellular function of pyrin and demonstrate an important link between pyrin and polymerising actin in the process of inflammatory cell migration.

Alteration of the microRNA expression profile in familial Mediterranean fever patients.

OBJECTIVES: Phenotypic heterogeneity in familial Mediterranean fever (FMF) disease indicated that FMF is not a simple monogenic disease. Therefore it has been suggested that epigenetic factors can be one of the reason for the variations. We undertook this study to test potential involvement of miRNAs in the pathogenesis of FMF. METHODS: miRNA array was performed on whole blood RNA samples from 6 healthy controls (-/-), 6 FMF patients (M694V/M694V), 6 carriers who displayed the disease phenotype (M694V/-) and 6 healthy carriers (M694V/-). The raw data was analysed by Multi Experiment Viewer (MeV) and candidate miRNAs were determined according to fold change (more than 2.0 or less than -2.0). The validation of differentially expressed miRNAs was done by qRT-PCR. Then we performed pathway analyses with using bioinformatics tools. RESULTS: 14 miRNAs were found to be significant among groups through the analysis with MeV. miR-20a-5p, miR-197-3p, let-7d-3p and miR-574-3p were found to be associated with inflammatory pathway related genes according to DAVID analysis. MiR-20a-5p (FDR: 0,00, FCH: 5.55) was significantly up regulated whereas miR-197-3p (FDR: 0,00, FCH: -2.27) was down regulated in homozygotes patients. Both let-7d-3p (FDR: 0.00, FCH: 28.75) and miR-574-3p (FDR: 0.00, FCH: 3.95) were up regulated in heterozygote patients group. CONCLUSIONS: We showed that there are several differentially expressed miRNAs both in homozygote and heterozygote FMF patients compared to controls and healthy carriers. Thus we suggest that these miRNAs, related with inflammatory pathways may be responsible for the expression of the disease in FMF.

Pyrin-PSTPIP1 colocalises at the leading edge during cell migration.

A set of mutations in the MEditerranean FeVer (MEFV) gene causes familial Mediterranean fever (FMF), the most common auto-inflammatory disease. The gene encodes a protein named pyrin, which appears to play an important role in inflammatory pathways. Furthermore, pyrin, which is expressed in neutrophils, has been reported to interact with proline-serine-threonine phosphatase-interacting protein 1 (PSTPIP1) and actin proteins. However, the relations between pyrin and PSTPIP1 during the cell migration have not yet been elucidated. In the present study, we constructed a cell migration assay method using HL-60 cells. Pyrin-PSTPIP1 interactions were analysed by immunofluorescence staining in control, differentiated and differentiated-stimulated HL-60 cells. In stimulated cells, pyrin-polymerised actin, PSTPIP1-polymerised actin and pyrin-PSTPIP1 were found to be colocalised. Pyrin has been shown to be colocalised with actin and PSTPIP1 at the leading edge of the migrating cell. For the first time, PSTPIP1 was found to interact with dynamic actin and pyrin at the site of polarisation.

Cellular models in autoinflammatory disease research.

Systemic autoinflammatory diseases are a heterogeneous group of rare genetic disorders caused by dysregulation of the innate immune system. Understanding the complex mechanisms underlying these conditions is critical for developing effective treatments. Cellular models are essential for identifying new conditions and studying their pathogenesis. Traditionally, these studies have used primary cells and cell lines of disease-relevant cell types, although newer induced pluripotent stem cell (iPSC)-based models might have unique advantages. In this review, we discuss the three cellular models used in autoinflammatory disease research, their strengths and weaknesses, and their applications to inform future research in the field.

Behçet's syndrome: recent advances to aid diagnosis.

Behçet's syndrome is a recurring inflammatory multiorgan disorder affecting the skin, mucosa, eyes, joints, stomach, and central nervous system. Behçet's syndrome epidemiology varies greatly among populations (0.64-420/100,000), and Behçet's syndrome has gained increasing international acclaim in the recent 50 years due to raising awareness of the syndrome, although it is rare in most population. In addition to the unclear etiology of the syndrome, the diagnosis of Behçet's syndrome is complicated by a vague clinical presentation, phenotypic heterogeneity and/or incomplete representation, and the lack of any specific laboratory, radiographic, or histological findings. There exists a dire need to elucidate factors that contribute to disease pathogenesis and/or are associated with clinical features of Behçet's syndrome and the classification of different forms of the syndrome. The identification of such molecular, cellular, and/or clinical factors are crucial for timely diagnosis and efficacious management of Behçet's syndrome. We discuss recent advances in the clinical diagnosis of Behçet's syndrome and related contributions of genetics, epigenetics, microbiome, inflammasomes, and autoantibodies to the improved diagnosis, management, and understanding of Behçet's syndrome.

Characterization of serum extracellular vesicles and their differential level of miR-197-3p in familial Mediterranean fever patients.

OBJECTIVES: The aim of this study was to analyze the existence of miRNAs derived from serum extracellular vesicles (EVs) in familial Mediterranean fever (FMF) patients. Our group has previously shown the association of certain miRNAs with FMF. METHODS: Serum samples of adult and pediatric FMF patients and their age matched controls were used in the study. Serum EVs were characterized by transmission electron microscopy (TEM) and flow cytometry. RNAs were isolated from EVs and levels of miR-197-3p and miR-20a-5p were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). RESULTS: EV characterization using TEM demonstrated fraction of 30-120 nm-sized particles with cup-shaped morphology. Flow cytometry results revealed the CD63 and CD81 positive populations as 53.3% in serum EVs. We showed that miR-197-3p and miR-20a-5p were "circulating miRNAs" and carried in EVs of FMF patients and controls. In FMF patients, level of miR-197-3p was significantly decreased. There was no significant alteration in the level for miR-20a-5p between patients and controls. CONCLUSION: We showed the differential level of miR-197-3p in serum EVs of the FMF patients. miR-197-3p's potential as a biomarker and therapeutic target in FMF pathogenesis warrants further investigation.

EVs and EV-miRNAs can be identified in FMF patients' sera.Serum EV-miR-197-3p is dysregulated in FMF patients.Serum EV-miR-197-3p might have both diagnostic and therapeutic potentials.

Drug metabolism and inflammation related distinct miRNA signature of colchicine resistant familial Mediterranean fever patients.

OBJECTIVE: Colchicine is the primary treatment for familial Mediterranean fever (FMF). Although colchicine is safe and effective in FMF patients, around 5-10% of patients show resistance to the drug. This study investigates the possibility of a link between colchicine resistance and the distinct miRNA profiles in colchicine resistant FMF patients. METHODS: Differentially expressed miRNAs in colchicine resistant FMF patients were detected by Affymetrix 4.0 miRNA array analysis. These miRNAs were then categorized based on the role of their target genes in drug metabolism and inflammation related pathways. qRT-PCR was used to validate candidate miRNAs selected by Enrichr, a gene enrichment analysis system based on the relevance of possible target genes in drug metabolism pathways. Expression levels of these miRNAs' potential target genes were investigated by qRT-PCR. Then, a colchicine resistant hepatoblastoma cell line (HEPG2) was established, and the differentially expressed miRNAs and genes identified in patients were also analyzed in this colchicine-resistant cell line. RESULTS: 25 differentially expressed miRNAs were detected in colchicine resistant FMF patients. miR-183-5p, miR-15b-5p, miR-505-5p, and miR-125a-5p were identified to be associated with drug resistance and inflammatory pathways and thus chosen for further validation. miR-183-5p, miR-15b-5p, miR-505-5p miRNAs showed significantly differential expression in qRT-PCR. NFKB1, NR3C1, PPARα - drug absorption, distribution, metabolism, and excretion (ADME) genes were predicted to be targeted by these miRNAs. Among these targets, NFKB1 and NR3C1 were differentially over expressed in colchicine resistant FMF patients. These findings were validated in the colchicine resistant hepatoblastoma cell line (HEPG2). CONCLUSION: This is the first study evaluating the role of miRNAs in colchicine resistant patients with FMF. Their differential expression may result in resistance to standard colchicine treatment by affecting the expression of genes that take place in drug absorption, distribution, metabolism, and excretion (ADME) or nuclear receptors that regulate ADME genes, thus potentially playing a role in both drug metabolism and inflammation.