Clinical and Molecular Characterization of SMAD4 Splicing Variants in Patients with Juvenile Polyposis Syndrome.

Juvenile polyposis syndrome (JPS) is an inherited autosomal dominant condition that predisposes to the development of juvenile polyps throughout the gastrointestinal (GI) tract, and it poses an increased risk of GI malignancy. Germline causative variants were identified in the SMAD4 gene in a subset (20%) of JPS cases. Most SMAD4 germline genetic variants published to date are missense, nonsense, and frameshift mutations. SMAD4 germline alterations predicted to result in aberrant splicing have rarely been reported. Here, we report two unrelated Italian families harboring two different SMAD4 intronic variants, c.424+5G>A and c.425-9A>G, which are clinically associated with colorectal cancer and/or juvenile GI polyps. In silico prediction analysis, in vitro minigene assays, and RT-PCR showed that the identified variants lead to aberrant SMAD4 splicing via the exonization of intronic nucleotides, resulting in a premature stop codon. This is expected to cause the production of a truncated protein. This study expands the landscape of SMAD4 germline genetic variants associated with GI polyposis and/or cancer. Moreover, it emphasizes the importance of the functional characterization of SMAD4 splicing variants through RNA analysis, which can provide new insights into genetic disease variant interpretation, enabling tailored genetic counseling, management, and surveillance of patients with GI polyposis and/or cancer.

Exploring the Relationship of rs2802292 with Diabetes and NAFLD in a Southern Italian Cohort-Nutrihep Study.

Background: The minor G-allele of FOXO3 rs2802292 is associated with human longevity. The aim of this study was to test the protective effect of the variant against the association with type 2 Diabetes and NAFLD. Methods: rs2802292 was genotyped in a large population of middle-aged subjects (n = 650) from a small city in Southern Italy. All participants were interviewed to collect information about lifestyle and dietary habits; clinical characteristics were recorded, and blood samples were collected from all subjects. The association between rs2802292 and NAFLD or diabetes was tested using a logistic model and mediation analysis adjusted for covariates. Results: Overall, the results indicated a statistical association between diabetes and rs2802292, especially for the TT genotype (OR = 2.14, 1.01 to 4.53 95% C.I., p = 0.05) or in any case for those who possess the G-allele (OR = 0.45, 0.25 to 0.81 95% C.I., p = 0.008). Furthermore, we found a mediation effect of rs2802292 on diabetes (as mediator) and NAFLD. There is no direct relationship between rs2802292 and NAFLD, but the effect is direct (ß = 0.10, -0.003 to 0.12 95% C.I., p = 0.04) on diabetes, but only in TT genotypes. Conclusions: The data on our cohort indicate that the longevity-associated FOXO3 variant may have protective effects against diabetes and NAFLD.

Lack of comparability of immunoassays for rheumatoid factor isotypes.

OBJECTIVES: Rheumatoid arthritis (RA) is a systemic autoimmune disease characterised by the presence of autoantibodies that are used for classification of the disease. Though routine diagnostics is commonly restricted to measuring rheumatoid factor (RF) and anti-citrullinated protein antibodies, detection of RF IgM, IgG and IgA isotypes, may increase the power of RA serodiagnosis by reducing the number of seronegative patients as well as provide prognostic information. The agglutination-based RF assays, such as nephelometry or turbidimetry, are unable to differentiate isotypes. We compared three different immunoassays used in current laboratory practice to detect RF isotypes. METHODS: We tested 117 consecutive serum samples that were positive for total RF at nephelometry, from 55 RA and 62 non-RA subjects. IgA, IgG, and IgM isotypes of RF were tested by immunoenzymatic (ELISA, Technogenetics), fluoroenzymatic (FEIA, ThermoFisher) and chemiluminescence (CLIA, YHLO Biotech Co.) immunoassays. RESULTS: Diagnostic performance differed considerably between the assays, especially with regard to RF IgG isotype. Agreement among methods by Cohen's kappa ranged from 0.05 (RF IgG CLIA vs. FEIA) to 0.846 (RF IgM CLIA vs. FEIA). CONCLUSIONS: The poor agreement observed in this study indicates substantial lack of comparability among assays for RF isotypes. Harmonization of these tests requires further efforts before their measurement can be used in clinical practice.

Colorectal Cancer Chemoprevention: A Dream Coming True?

Colorectal cancer (CRC) is one of the deadliest forms of cancer worldwide. CRC development occurs mainly through the adenoma-carcinoma sequence, which can last decades, giving the opportunity for primary prevention and early detection. CRC prevention involves different approaches, ranging from fecal occult blood testing and colonoscopy screening to chemoprevention. In this review, we discuss the main findings gathered in the field of CRC chemoprevention, focusing on different target populations and on various precancerous lesions that can be used as efficacy evaluation endpoints for chemoprevention. The ideal chemopreventive agent should be well tolerated and easy to administer, with low side effects. Moreover, it should be readily available at a low cost. These properties are crucial because these compounds are meant to be used for a long time in populations with different CRC risk profiles. Several agents have been investigated so far, some of which are currently used in clinical practice. However, further investigation is needed to devise a comprehensive and effective chemoprevention strategy for CRC.

Classic Galactosemia: Clinical and Computational Characterization of a Novel GALT Missense Variant (p.A303D) and a Literature Review.

Classic galactosemia is an autosomal recessive inherited liver disorder of carbohydrate metabolism caused by deficient activity of galactose-1-phosphate uridylyltransferase (GALT). While a galactose-restricted diet is lifesaving, most patients still develop long-term complications. In this study, we report on a two-week-old female patient who is a compound heterozygote for a known pathogenic variant (p.K285N) and a novel missense variant (p.A303D) in the GALT gene. Segregation analysis showed that the patient inherited the p.K285N pathogenic variant from her father and the p.A303D variant from her mother. A bioinformatics analysis to predict the impact of the p.A303D missense variant on the structure and stability of the GALT protein revealed that it may be pathogenic. Based on this finding, we performed a literature review of all GALT missense variants identified in homozygous and compound heterozygous galactosemia patients carrying the p.K285N pathogenic variant to explore their molecular effects on the clinical phenotype of the disease. Our analysis revealed that these missense variants are responsible for a wide range of molecular defects. This study expands the clinical and mutational spectrum in classic galactosemia and reinforces the importance of understanding the molecular consequences of genetic variants to incorporate genetic analysis into clinical care.

Soluble urokinase Plasminogen Activator Receptor (suPAR) levels are predictive of COVID-19 severity: an Italian experience.

BACKGROUND: The soluble urokinase Plasminogen Activator Receptor (suPAR) has been identified as a reliable marker of COVID-19 severity, helping in personalizing COVID-19 therapy. This study aims to evaluate the correlation between suPAR levels and COVID-19 severity, in relation to the traditional inflammatory markers. METHODS: Sera from 71 COVID-19 patients were tested for suPAR levels using Chorus suPAR assay (Diesse Diagnostica Senese SpA, Italy). suPAR levels were compared with other inflammatory markers: IL-1ß, IL-6, TNF-α, circulating calprotectin, neutrophil and lymphocyte counts, and Neutrophil/Lymphocytes Ratio (NLR). Respiratory failure, expressed as P/F ratio, and mortality rate were used as indicators of disease severity. RESULTS: A positive correlation of suPAR levels with IL-6 (r = 0.479, p = 0.000), TNF-α (r = 0.348, p = 0.003), circulating calprotectin (r = 0.369, p = 0.002), neutrophil counts (r = 0.447, p = 0.001), NLR (r = 0.492, p = 0.001) has been shown. Stratifying COVID-19 population by suPAR concentration above and below 6 ng/mL, we observed higher levels of circulating calprotectin (10.1 µg/mL, SD 7.9 versus 6.4 µg/mL, SD 7.5, p < 0.001), higher levels of P/F ratio (207.5 IQR 188.3 vs 312.0 IQR 127.8, p = 0.013) and higher mortality rate. Median levels of suPAR were increased in all COVID-19 patients requiring additional respiratory support (Nasal Cannula, Venturi Mask, BPAP and CPAP) (6.5 IQR = 4.9) compared to the group at room air (4.6 IQR = 4.2). CONCLUSION: suPAR levels correlate with disease severity and survival rate of COVID-19 patients, representing a promising prognostic biomarker for the risk assessment of the disease.

Clinical utility of circulating calprotectin to assist prediction and monitoring of COVID-19 severity: An Italian study.

BACKGROUND: Calprotectin (S100A8/A9) has been identified as a biomarker that can aid in predicting the severity of disease in COVID-19 patients. This study aims to evaluate the correlation between levels of circulating calprotectin (cCP) and the severity of COVID-19. METHODS: Sera from 245 COVID-19 patients and 110 apparently healthy individuals were tested for calprotectin levels using a chemiluminescent immunoassay (Inova Diagnostics). Intensive care unit (ICU) admission and type of respiratory support administered were used as indicators of disease severity, and their correlation with calprotectin levels was assessed. RESULTS: Samples from patients in the ICU had a median calprotectin concentration of 11.6 µg/ml as compared to 3.5 µg/ml from COVID-19 patients who were not in the ICU. The median calprotectin concentration in a cohort of healthy individuals collected before the COVID-19 pandemic was 3.0 µg/ml (95% CI: 2.820-2.969 µg/ml). Patients requiring a Venturi mask, continuous positive airway pressure, or orotracheal intubation all had significantly higher values of calprotectin than controls, with the increase of cCP levels proportional to the increasing need of respiratory support. CONCLUSION: Calprotectin levels in serum correlate well with disease severity and represent a promising serological biomarker for the risk assessment of COVID-19 patients.

Cardiovascular safety, cancer and Jak-inhibitors: Differences to be highlighted.

Rheumatoid arthritis (RA) is a chronic inflammatory disease whose natural history leads to articular and extra-articular damage. Both cardiovascular risk and malignancy risk results higher in RA patients, compared to general population. Janus kinase inhibitors (JAKis) are oral targeted synthetic disease modifying antirheumatic drugs (tsDMARDs) that disrupt cytokine cascade and exert anti-inflammatory effects by interfering with signaling through the JAK-STAT intracellular pathways. A recent RCT comparing tofacitinib 5 mg twice daily, tofacitinib 10 mg twice daily and anti-TNF in rheumatoid arthritis demonstrated an increased risk of MACE HR 1.33 and cancer HR 1.49 at a follow-up of 4 years. This has led the FDA to class warnings for tofacitinib, baricitinib and upadacitinib. Cumulative RCT data, RCT extension data demonstrated a safety profile for Jak inhibitors. Conflicting data results from real life registries; the different selectivity for JAKs (JAK1, JAK2, JAK3 and Tyk2) probably determines differences in efficacy and safety profiles among the members of this group which should actually be evaluated. In order to better understand the cardiovascular and neoplastic risk linked to these class of drugs, we aim to provide a literature review on existing evidence of the safety of Jak-Inhibitors in rheumatoid arthritis.

The role of neutralizing antibodies by sVNT after two doses of BNT162b2 mRNA vaccine in a cohort of Italian healthcare workers.

OBJECTIVES: Evaluating anti-SARS-CoV-2 antibody levels is a current priority to drive immunization, as well as to predict when a vaccine booster dose may be required and for which priority groups. The aim of our study was to investigate the kinetics of anti-SARS-CoV-2 Spike S1 protein IgG (anti-S1 IgG) antibodies and neutralizing antibodies (NAbs) in an Italian cohort of healthcare workers (HCWs), following the Pfizer/BNT162b2 mRNA vaccine, over a period of up to six months after the second dose. METHODS: We enrolled 57 HCWs, without clinical history of COVID-19 infection. Fluoroenzyme-immunoassay was used for the quantitative anti-S1 IgG antibodies at different time points T1 (one month), T3 (three months) and T6 (six months) following the second vaccine shot. Simultaneously, a commercial surrogate virus neutralization test (sVNT) was used for the determination of NAbs, expressed as inhibition percentage (% IH). RESULTS: Median values of anti-S1 IgG antibodies decreased from T1 (1,452 BAU/mL) to T6 (104 BAU/mL) with a percent variation of 92.8% while the sVNT showed a percent variation of 34.3% for the same time frame. The decline in anti-S1 IgG antibodies from T1 to T6 was not accompanied by a loss of the neutralizing capacity of antibodies. In fact at T6 a neutralization percentage <20% IH was observed only in 3.51% of HCWs. CONCLUSIONS: Our findings reveal that the decrease of anti-S1 IgG levels do not correspond in parallel to a decrease of NAbs over time, which highlights the necessity of using both assays to assess vaccination effectiveness.

Water, sanitation, and hygiene services in health care facilities in the Autonomous Province of Vojvodina, Serbia.

Provision of safe water, sanitation, and hygiene (WASH) services in health care facilities is a priority at the global, national, and local levels. To inform improvements planning, conditions of WASH, waste management, and environmental cleaning were assessed in 81 facilities in the Autonomous Province of Vojvodina, Serbia, as part of a nationally representative survey in 2019. The survey included on-site checks, structured interviews, and drinking-water quality analysis. WHO/UNICEF indicators for WASH service levels and an advanced service level defined at the national level were applied. The results showed that all investigated facilities provided basic water services; 94% of facilities provided basic hygiene and waste management services; 58 and 2%, respectively, provided basic cleaning and sanitation services. Only 1% of investigated facilities met the basic level for all five WASH dimensions. Advanced service levels were only met for hygiene, waste management, and/or cleaning in 15-38% of facilities. In 33% of health care facilities, drinking-water quality was not in compliance with the national standards. The results revealed that there is a need for increased awareness and efforts to ensure basic provisions for sanitation, environmental cleaning, and drinking-water safety.

Assessing T-Cell Immunity in Kidney Transplant Recipients with Absent Antibody Production after a 3rd Dose of the mRNA-1273 Vaccine.

The vulnerable population of kidney transplant recipients (KTRs) are low responders to COVID-19 vaccines, so specific immune surveillance is needed. The interferon-gamma (IFN-γ) release assay (IGRA) is effective in assessing T cell-mediated immunity. We assessed SARS-CoV-2-directed T cell responses in KTRs with absent antibody production after a third dose of the mRNA-1273 vaccine, using two different IGRAs. A cohort of 57 KTRs, who were actively followed up, received a third dose of the mRNA-1273 vaccine. After the evaluation of humoral immunity to SARS-CoV-2, 14 seronegative patients were tested with two commercial IGRAs (SD Biosensor and Euroimmun). Out of 14 patients, one and three samples were positive by IGRAs with Euroimmun and SD Biosensor, respectively. The overall agreement between the two assays was 85.7% (κ = 0.444). In addition, multivariate linear regression analysis showed no statistically significant association between the IFN-γ concentration, and the independent variables analyzed (age, gender, years since transplant, total lymphocytes cells/mcl, CD3+ cells/mcl, CD3+ CD4+ cells/mcl, CD3+ CD8+ cells/mcl, CD19+ cells/mcl, CD3-CD16+CD56+ cells/mcl) (p > 0.01). In a vulnerable setting, assessing cellular immune response to complement the humoral response may be advantageous. Since the two commercial IGRAs showed a good agreement on negative samples, the three discordant samples highlight the need for further investigations.

Closing the serological gap in the diagnostic testing for COVID-19: The value of anti-SARS-CoV-2 IgA antibodies.

During coronavirus disease 2019 (COVID-19) pandemic, the early diagnosis of patients is a priority. Serological assays, in particular immunoglobulin (Ig)M and IgG anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have today several applications but the interpretation of their results remains an open challenge. Given the emerging role of the IgA isotype in the COVID-19 diagnostics, we aimed to identify the SARS-CoV-2 IgA antibodies in a COVID-19 population seronegative for IgM. A total of 30 patients hospitalized in San Giovanni di Dio Hospital (Florence, Italy) for COVID-19, seronegative for IgM antibodies, have been studied for anti-SARS-CoV-2 antibodies. They all had a positive oro/nasopharyngeal swab reverse transcription-polymerase chain reaction result. Assays used were a chemiluminescent assay measuring SARS-CoV-2 specific IgM and IgG (S + N) and an ELISA, measuring specific IgG (S1) and IgA antibodies against SARS-CoV-2. Among the 30 patients, eight were positive for IgA, seven were positive for IgG (N + S), and two for IgG (S1), at the first point (5-7 days from the onset of symptoms). The IgA antibodies mean values at the second (9-13 days) and third (21-25 days) time points were even more than twice as high as IgG assays. The agreement between the two IgG assays was moderate (Cohen's K = 0.59; SE = 0.13). The inclusion of the IgA antibodies determination among serological tests of the COVID-19 diagnostic is recommended. IgA antibodies may help to close the serological gap of the COVID-19. Variations among anti-SARS-CoV-2 IgG assays should be considered in the interpretation of results.

Functional evidence of mTORß splice variant involvement in the pathogenesis of congenital heart defects.

mTOR dysregulation has been described in pathological conditions, such as cardiovascular and overgrowth disorders. Here we report on the first case of a patient with a complex congenital heart disease and an interstitial duplication in the short arm of chromosome 1, encompassing part of the mTOR gene. Our results suggest that an intragenic mTOR microduplication might play a role in the pathogenesis of non-syndromic congenital heart defects (CHDs) due to an upregulation of mTOR/Rictor and consequently an increased phosphorylation of PI3K/AKT and MEK/ERK signaling pathways in patient-derived amniocytes. This is the first report which shows a causative role of intragenic mTOR microduplication in the etiology of an isolated complex CHD.

Do new and old biomarkers of early undifferentiated arthritis correlate with Arthritis Impact Measurement Scales?

OBJECTIVES: In early undifferentiated arthritis (EUA), the relationship between inflammatory biomarkers and disability is still unclear. The aim of this study was to correlate inflammatory biomarkers with the Arthritis Impact Measurement Scales (AIMS) in EUA. METHODS: Seventy patients with EUA were compared with 20 patients with established rheumatoid arthritis (RA). The association of AIMS [mobility, physical impairment (PI), dexterity, household activities, activities of daily living (ADL), social activity, pain, anxiety, depression] with serum laboratory [phase acute reactants, calprotectin, interleukin-6, tumour necrosis factor (TNF)-α, rheumatoid factor, anti-nuclear and anti-citrullinated peptide antibodies, HLA-DRB], clinical [Clinical Disease Activity Index (CDAI), fatigue, pain and stiffness NRS], x-ray and ultrasound biomarkers was analysed with non-parametric Spearman's rank correlation and Mann-Whitney U tests. RESULTS: No differences in AIMS were found between EUA and established RA patients, or between EUA patients that evolved into early RA (n=17) and those that remained EUA (n=53) at six months of follow-up. In EUA, erythrocyte sedimentation rate correlated with mobility impairment, PI and depression (p=0.04, p=0.03 and p=0.022, respectively), TNF-α correlated with PI (p=0.01) and calprotectin with anxiety (p=0.02). HLA-DRB1*11-positive EUA patients had lower ADL deficiency (p=0.006), depression (p=0.0004) and anxiety (p=0.01). CDAI correlated with PI (p=0.01) and pain (p=0.01), fatigue with PI (p=0.0001) and ADL (p=0.009), stiffness with PI (p=0.01), and Power Doppler ultrasound synovitis with PI (p=0.02) and pain (p=0.007). CONCLUSIONS: In EUA, physical and mood disorders are associated with new and old inflammatory serological, clinical and imaging biomarkers. HLA-DRB1*11-positivity may be protective against these disease-related features.

Integrating quality assurance in autoimmunity: the changing face of the automated ANA IIF test.

OBJECTIVES: Currently available computer-aided diagnosis (CAD) systems for the detection of anti-nuclear antibodies (ANA) by indirect immunofluorescence (IIF) assay enable a standardized measurement of system-specific fluorescent intensity (FI) measures. We aimed to evaluate an internal quality control (iQC) program that controls the total ANA IIF process in routine practice. METHODS: In addition to the kit iQC materials, supplemental quality indicators were integrated in a total quality assurance (QA) program: patient-derived iQC's samples (negative, 1/160 fine speckled and 1/160 homogeneous), median sample FI per run and percentage of ANA IIF positive samples per run. Analytical rejection criteria were based on the imprecision of the positivity index (PI) measure of the Zenit PRO system (Menarini). Clinical rejection criteria were based on changes in FI that correspond to a change in ANA IIF titer of ≥2. To evaluate the QA program, different artificial errors were introduced during the ANA IIF process. After every run, quality indicators were evaluated and compared to the pre-set target values. RESULTS: Rescanning the ANA IIF slides five times, using an old conjugate and a needle obstruction resulted in analytically and even clinically relevant errors in ANA IIF results. All errors were correctly detected by the different defined quality indicators. Traditional Westgard rules, including analytically (and clinically) defined rejection limits were useful in monitoring quality indicators. CONCLUSIONS: The integration of a total process iQC program in CAD systems, based on the specific FI measurands and performance criteria of the system, adds value to QA.

Gastric polyposis and desmoid tumours as a new familial adenomatous polyposis clinical variant associated with APC mutation at the extreme 3'-end.

Germline mutations of the APC gene, which encodes a multidomain protein of 2843 amino acid residues, cause familial adenomatous polyposis (FAP). Three FAP clinical variants are correlated with the location of APC mutations: (1) classic FAP with profuse polyposis (>1000 adenomas), associated with mutations from codon 1250 to 1424; (2) attenuated FAP (<100 adenomas), associated with mutations at APC extremities (before codon 157 and after codon 1595); (3) classic FAP with intermediate colonic polyposis (100-1000 adenomas), associated with mutations located in the remaining part of APC In an effort to decipher the clinical phenotype associated with APC C-terminal germline truncating mutations in patients with FAP, after screening APC mutations in one family whose members (n=4) developed gastric polyposis, colon oligo-polyposis and desmoid tumours, we performed a literature meta-analysis of clinically characterised patients (n=97) harbouring truncating mutations in APC C-terminus. The APC distal mutations identified in this study cluster with a phenotype characterised by colon oligo-polyposis, diffuse gastric polyposis and desmoid tumours. In conclusion, we describe a novel FAP clinical variant, which we propose to refer to as Gastric Polyposis and Desmoid FAP, that may require tailored management.

Diagnostic accuracy of an automated chemiluminescent immunoassay for anti-SARS-CoV-2 IgM and IgG antibodies: an Italian experience.

A pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been spreading throughout the world. Though molecular diagnostic tests are the gold standard for COVID-19, serological testing is emerging as a potential surveillance tool, in addition to its complementary role in COVID-19 diagnostics. Indubitably quantitative serological testing provides greater advantages than qualitative tests but today there is still little known about serological diagnostics and what the most appropriate role quantitative tests might play. Sixty-one COVID-19 patients and 64 patients from a control group were tested by iFlash1800 CLIA analyzer for anti-SARS CoV-2 antibodies IgM and IgG. All COVID-19 patients were hospitalized in San Giovanni di Dio Hospital (Florence, Italy) and had a positive oro/nasopharyngeal swab reverse-transcription polymerase chain reaction result. The highest sensitivity with a very good specificity performance was reached at a cutoff value of 10.0 AU/mL for IgM and of 7.1 for IgG antibodies, hence near to the manufacturer's cutoff values of 10 AU/mL for both isotypes. The receiver operating characteristic curves showed area under the curve values of 0.918 and 0.980 for anti-SARS CoV-2 antibodies IgM and IgG, respectively. iFlash1800 CLIA analyzer has shown highly accurate results for the anti-SARS-CoV-2 antibodies profile and can be considered an excellent tool for COVID-19 diagnostics.

Two Novel Technologies for the Detection of Anti-cardiolipin and Anti ß2-Glycoprotein Antibodies in the Real Life: Chemiluminescent in Comparison to the Addressable Laser Bead Immunoassays.

In the present study, we evaluated two novel technologies, the chemiluminescent immunoassay (CIA) QUANTA Flash on BIO-FLASH (Inova Diagnostics, San Diego, CA, USA) and the addressable laser bead immunoassay (ALBIA) on BioPlex™ 2200 (Bio-Rad, Hercules, CA, USA) for the detection of anti-cardiolipin IgG/IgM (aCL) and anti-ß2-glycoprotein IgG/IgM (aß2GPI) antibodies. The study was performed on 134 samples from consecutive patients (59 males and 75 females, mean age 54 ± 10 years) who consulted a rheumatologist because thrombosis and/or pregnancy complications were present or another immunological disease (Sjogren's syndrome, inflammatory arthritis). Fourteen patients of the total fulfilled 25the Sydney criteria for APS and for these patients previous results of aPLs were available. Sera were tested for aCL and aß2GPI of IgG and IgM isotypes using CIA (BIO-FLASH) and ALBIA (BioPlex™ 2200). Overall agreement between CIA and ALBIA ranged from 88.1% (aCL IgG) to 97.8% (aß2GPI IgG). Cohen's kappa coefficient ranged from 0.53 to 0.91, implying moderate to almost perfect agreement. Almost perfect agreement was found between BioPlex™ 2200 and BIO-FLASH aß2GPI IgG and aCL IgM with Cohen's kappa of 0.91 and 0.88, respectively. On the other hand, moderate agreement was found between BioPlex™ 2200 and BIO-FLASH aCL IgG and ß2GPI IgM assays with Cohen's kappa of 0.57 and 0.53, respectively. The two novel technologies look promising and comparable but further studies with larger cohorts are needed to contribute to the better understanding of the new aPLs antibodies assays performance.

The Laboratory Role in anti-TNF Biological Therapy Era.

Along years, the advent of biological therapy widely modified treatment of rheumatic diseases and other disorders. However, many agents may elicit in anti-drug antibodies (ADAbs) upon consecutive infusions, with a loss of response. For the right strategy of a personalized medicine, the therapeutic monitoring of TNF-α inhibitors and ADAbs represents an important effort in diagnostic-therapeutic pathway, to improve overall patient management and favoring an appropriate clinical approach. A raising number of diagnostic tests have been designed to elucidate the efficacy and/or safety of a specific drug or class of drugs for a targeted patient's group. Our paper reviewed the current understanding of the immunogenicity of biological drugs employed in the treatment of inflammatory diseases underlying the laboratory role.

The longevity SNP rs2802292 uncovered: HSF1 activates stress-dependent expression of FOXO3 through an intronic enhancer.

The HSF and FOXO families of transcription factors play evolutionarily conserved roles in stress resistance and lifespan. In humans, the rs2802292 G-allele at FOXO3 locus has been associated with longevity in all human populations tested; moreover, its copy number correlated with reduced frequency of age-related diseases in centenarians. At the molecular level, the intronic rs2802292 G-allele correlated with increased expression of FOXO3, suggesting that FOXO3 intron 2 may represent a regulatory region. Here we show that the 90-bp sequence around the intronic single nucleotide polymorphism rs2802292 has enhancer functions, and that the rs2802292 G-allele creates a novel HSE binding site for HSF1, which induces FOXO3 expression in response to diverse stress stimuli. At the molecular level, HSF1 mediates the occurrence of a promoter-enhancer interaction at FOXO3 locus involving the 5'UTR and the rs2802292 region. These data were confirmed in various cellular models including human HAP1 isogenic cell lines (G/T). Our functional studies highlighted the importance of the HSF1-FOXO3-SOD2/CAT/GADD45A cascade in cellular stress response and survival by promoting ROS detoxification, redox balance and DNA repair. Our findings suggest the existence of an HSF1-FOXO3 axis in human cells that could be involved in stress response pathways functionally regulating lifespan and disease susceptibility.