Case report: Rapidly progressive neurocognitive disorder with a fatal outcome in a patient with PU.1 mutated agammaglobulinemia.

Introduction: PU.1-mutated agammaglobulinemia (PU.MA) represents a recently described autosomal-dominant form of agammaglobulinemia caused by mutation of the SPI1 gene. This gene codes for PU.1 pioneer transcription factor important for the maturation of monocytes, B lymphocytes, and conventional dendritic cells. Only six cases with PU.MA, presenting with chronic sinopulmonary and systemic enteroviral infections, have been previously described. Accumulating literature evidence suggests a possible relationship between SPI1 mutation, microglial phagocytic dysfunction, and the development of Alzheimer's disease (AD). Case description: We present a Caucasian female patient born from a non-consanguineous marriage, who was diagnosed with agammaglobulinemia at the age of 15 years when the immunoglobulin replacement therapy was started. During the following seventeen years, she was treated for recurrent respiratory and intestinal infections. At the age of 33 years, the diagnosis of celiac-like disease was established. Five years later progressive cognitive deterioration, unstable gait, speech disturbances, and behavioral changes developed. Comprehensive microbiological investigations were negative, excluding possible infective etiology. Brain MRI, 18FDG-PET-CT, and neuropsychological testing were suggestive for a diagnosis of a frontal variant of AD. Clinical exome sequencing revealed the presence of a novel frameshift heterozygous variant c.441dup in exon 4 of the SPI1 gene. Despite intensive therapy, the patient passed away a few months after the onset of the first neurological symptoms. Conclusion: We describe the first case of PU.MA patient presenting with a rapidly progressive neurocognitive deterioration. The possible role of microglial dysfunction in patients with SPI1 mutation could explain their susceptibility to neurodegenerative diseases thus highlighting the importance of genetic testing in patients with inborn errors of immunity. Since PU.MA represents a newly described form of agammaglobulinemia, our case expands the spectrum of manifestations associated with SPI1 mutation.

Asthma Inflammatory Phenotypes: How Can We Distinguish Them?

BACKGROUND AND OBJECTIVES: induced sputum is used to assess different inflammatory phenotypes in asthma, but is not used routinely. We aimed to determine the proportion of inflammatory asthma phenotypes based on induced sputum, to find biomarkers that can discriminate between phenotypes, and to evaluate biomarkers in patients with and without biological therapy in different inflammatory asthma phenotypes. MATERIALS AND METHODS: this cross-sectional study investigated clinical characteristics, asthma control tests, skin prick test, impulse oscillometry (IOS), spirometry, induced sputum, biomarkers (IgE, eosinophils, fractional exhaled nitric oxide (FeNO), serum periostin, IL-5, IL-6, IL-8, IL-17A, IL-33) in 80 asthmatics. A total of 17/80 patients were treated with biologics (10 with omalizumab, 7 with benralizumab). RESULTS: a total of 31% of patients had eosinophilic asthma (EA), 30% had mixed granulocytic asthma (MGA), 24% had paucigranulocytic asthma (PGA), and 15% had neutrophilic asthma (NA). The difference was found in blood eosinophils (p = 0.002), the highest observed in EA. The cut-off ≥ 240/µL eosinophils, with 64% sensitivity and 72.7% specificity, identified EA (AUC = 0.743, p = 0.001). A higher IL-8 level was associated with NA (p = 0.025). In 63 non-biologic asthma group, eosinophils were higher in EA than in NA, MGA, and PGA (p = 0.012, p = 0.028, and p = 0.049, respectively). A higher IL-17A was associated with EA without biologics (p = 0.004). A significantly higher IL-5 was found in EA treated with biologics, in comparison with EA without biologics (p = 0.043). The number of leucocytes and neutrophils was higher in MGA without biologics (p = 0.049, p = 0.019), while IL-5, IL-6, and IL-8 levels were higher in MGA treated with biologics (p = 0.012, p = 0.032, p = 0.038, respectively). CONCLUSIONS: EA and MGA were the most prevalent asthma phenotypes. Blood eosinophils can identify EA, both in patients with and without biologics. Apart from the clinical profile, a broad spectrum of biomarkers for assessing inflammatory phenotypes is necessary for an adequate therapy approach to patients with asthma.

Autophagy Receptor p62 Regulates SARS-CoV-2-Induced Inflammation in COVID-19.

As autophagy can promote or inhibit inflammation, we examined autophagy-inflammation interplay in COVID-19. Autophagy markers in the blood of 19 control subjects and 26 COVID-19 patients at hospital admission and one week later were measured by ELISA, while cytokine levels were examined by flow cytometric bead immunoassay. The antiviral IFN-α and proinflammatory TNF, IL-6, IL-8, IL-17, IL-33, and IFN-γ were elevated in COVID-19 patients at both time points, while IL-10 and IL-1ß were increased at admission and one week later, respectively. Autophagy markers LC3 and ATG5 were unaltered in COVID-19. In contrast, the concentration of autophagic cargo receptor p62 was significantly lower and positively correlated with TNF, IL-10, IL-17, and IL-33 at hospital admission, returning to normal levels after one week. The expression of SARS-CoV-2 proteins NSP5 or ORF3a in THP-1 monocytes caused an autophagy-independent decrease or autophagy-inhibition-dependent increase, respectively, of intracellular/secreted p62, as confirmed by immunoblot/ELISA. This was associated with an NSP5-mediated decrease in TNF/IL-10 mRNA and an ORF3a-mediated increase in TNF/IL-1ß/IL-6/IL-10/IL-33 mRNA levels. A genetic knockdown of p62 mimicked the immunosuppressive effect of NSP5, and a p62 increase in autophagy-deficient cells mirrored the immunostimulatory action of ORF3a. In conclusion, the proinflammatory autophagy receptor p62 is reduced inacute COVID-19, and the balance between autophagy-independent decrease and autophagy blockade-dependent increase of p62 levels could affect SARS-CoV-induced inflammation.

Molecular diagnostics and inhibition of cross-reactive carbohydrate determinants in Hymenoptera venom allergy.

BACKGROUND: The composition of venom extracts, cross-reactive carbohydrate determinants (CCD) and the component-resolved diagnostics (CRD) are important fields of investigation. IgE-reactivity to CCD complicates the interpretation of IgE to Hymenoptera venoms, especially in patients with multiple-positivity. We analyzed the clinical importance of CRD and CCD-inhibition for selection of allergens for venom immunotherapy (VIT). METHODS: In 71 patients, we measured specific IgE (sIgE) to honeybee venom (HBV), wasp venom (WV), hornet venom (HV), CCD, and recombinant allergens: phospholipase A2 (rApi m 1), hyaluronidase (rApi m 2), icarapin (rApi m 10), antigen 5 (rVes v 5), and phospholipase A1 (Immunoblot). In 29/71 HBV/WV/HV/CCD-positive patients CCD-inhibition was performed. According to CRD and CCD-inhibition, we identified true sensitization and defined groups of multiple-positive patients who needed CCD-inhibition before starting VIT. RESULTS: sIgE-rApi m 1, sIgE-rApi m 2, and sIgE-rApi m 10 were detected in 65.7%, 68.4%, and 58%, respectively. In HBV allergic patients, CRD sensitivity was 86.8%. In WV allergic patients, sensitivity of sIgE-rVes v 5 was 94%. True multiple-sensitization was found in 44.8% of HBV/WV/HV/CCD-positive patients after CCD-inhibition. Patients with multiple venom- and CCD-positivity had more frequent severe allergic reactions (p < 0.001). CCD-inhibition was helpful in HBV/WV/HV/CCD-positive patients who were negative to all tested recombinant honeybee allergens. Persistence of HBV-positivity after CCD-inhibition requires CRD to other honeybee recombinant allergens. CONCLUSION: CRD, using a profile of five most important recombinant allergens and CCD, has a high sensitivity for the diagnosis of venom allergy, especially in patients positive to several venom extracts. CRD and CCD-inhibition are helpful to reveal the clinically relevant, true sensitization and improve the selection of venoms for long-lasting VIT.

Mastocytosis in children: a single-center long-term follow-up study.

BACKGROUND: Mastocytosis is a heterogeneous group of rare disorders characterized by the accumulation of clonal mast cells in organs such as the skin and bone marrow. The diagnosis of cutaneous mastocytosis (CM) is based on clinical findings, positive Darier's sign, and histopathology, if necessary. METHODS: Medical records of 86 children with CM diagnosed during a 35-year long period were reviewed. Most patients (93%) developed CM during the first year of life (median age 3 months). Clinical features at presentation and during the follow-up period were analyzed. Baseline serum tryptase level was measured in 28 patients. RESULTS: A total of 85% of patients had maculopapular cutaneous mastocytosis/urticaria pigmentosa (MPCM/UP), 9% had mastocytoma, and 6% had diffuse cutaneous mastocytosis (DCM). Boy to girl ratio was 1.1:1. Fifty-four of 86 patients (63%) were followed from 2 to 37 years (median 13 years). Complete resolution was registered in 14% of mastocytoma cases, 14% of MCPM/UP, and in 25% of DCM patients. After the age of 18, skin lesion persisted in 14% mastocytoma, 7% MCPM/UP, and 25% children with DCM. Atopic dermatitis was diagnosed in 9.6% of patients with MPCM/UP. Three of 28 patients had elevated serum tryptase. Prognosis in all patients was good, and there were no signs of progression to systemic mastocytosis (SM). CONCLUSION: To the best of our knowledge, our results represent the longest single-center follow-up study of childhood-onset CM. We found no complications of massive mast cell degranulation or progression to SM.

Developing COVID-19 vaccine recommendations during the pandemic: The experience of Serbia's Expert Committee on Immunization.

A National Immunization Technical Advisory Group (NITAG) is a multi-disciplinary body of national experts that provide evidence-based recommendations to policy-makers to assist them in making informed immunization policy and programme decisions. During the COVID-19 pandemic, NITAGs faced many challenges in making evidence-based recommendations for COVID-19 vaccines due to the rapidly evolving situation with new vaccine products available in a short time period and limited data on vaccine effectiveness. The authors reviewed the process used by Serbia's NITAG, which is called the Serbian Expert Committee on Immunization, to develop COVID-19 vaccine recommendations during the pandemic. The article examines the challenges and successes faced by the committee. Serbia's expert committee used the best available evidence to develop over forty recommendations on all aspects of COVID-19 vaccination. These expert committee recommendations facilitated the early procurement and successful roll-out of COVID-19 vaccines, guidance for vaccination of individuals at the highest risk, and high COVID-19 vaccination coverage in the country. The availability of five COVID-19 vaccines in Serbia was an advantage for the successful roll-out but posed challenges for the expert committee. Serbia's expert committee plans to use the experience and best practices developed during the pandemic to improve and expand its work moving forward.

Autoimmune and immunoserological markers of COVID-19 pneumonia: Can they help in the assessment of disease severity.

Background: Immune dysregulation and associated inefficient anti-viral immunity during Coronavirus Disease 2019 (COVID-19) can cause tissue and organ damage which shares many similarities with pathogenetic processes in systemic autoimmune diseases. In this study, we investigate wide range autoimmune and immunoserological markers in hospitalized patients with COVID-19. Methods: Study included 51 patients with confirmed Severe Acute Respiratory Syndrome Coronavirus 2 infection and hospitalized due to COVID-19 pneumonia. Wide spectrum autoantibodies associated with different autoimmune inflammatory rheumatic diseases were analyzed and correlated with clinical and laboratory features and pneumonia severity. Results: Antinuclear antibodies (ANA) positivity was found in 19.6%, anti-cardiolipin IgG antibodies (aCL IgG) in 15.7%, and anti-cardiolipin IgM antibodies (aCL IgM) in 7.8% of patients. Positive atypical x anti-neutrophil cytoplasmic antibodies (xANCA) were detected in 10.0% (all negative for Proteinase 3 and Myeloperoxidase) and rheumatoid factor was found in 8.2% of patients. None of tested autoantibodies were associated with disease or pneumonia severity, except for aCL IgG being significantly associated with higher pneumonia severity index (p = 0.036). Patients with reduced total serum IgG were more likely to require non-invasive mechanical ventilation (NIMV) (p < 0.0001). Serum concentrations of IgG (p = 0.003) and IgA (p = 0.032) were significantly lower in this group of patients. Higher total serum IgA (p = 0.009) was associated with mortality, with no difference in serum IgG (p = 0.115) or IgM (p = 0.175). Lethal outcome was associated with lower complement C4 (p = 0.013), while there was no difference in complement C3 concentration (p = 0.135). Conclusion: Increased autoimmune responses are present in moderate and severe COVID-19. Severe pneumonia is associated with the presence of aCL IgG, suggesting their role in disease pathogenesis. Evaluation of serum immunoglobulins and complement concentration could help assess the risk of non-invasive mechanical ventilation NIMV and poor outcome.

Level and avidity of antineutrophil cytoplasmic antibodies specific to lactoferrin are useful biomarkers in systemic lupus erythematosus.

OBJECTIVES: To evaluate specificity, level, and avidity of antineutrophil cytoplasmic antibodies (ANCA) in systemic lupus erythematosus (SLE). There are no studies of ANCA avidity in SLE. METHODS: Level (ELISA) and avidity (ELISA) of myeloperoxidase (MPO-), proteinase 3 (PR3-), lactoferrin (LF-), cathepsin G, elastase (EL-), and bactericidal/permeability increasing protein (BPI)-ANCA in 142 SLE patients were studied. SLE activity was measured by SLEDAI-2 K. 25/40 ANCA-positive patients were immunoserologically followed (12 ± 2 months). RESULTS: 40/142 (28.2%) SLE patients were ANCA-positive: LF- (21/40), MPO- (19/40), EL- (6/40), PR3- (3/40), and BPI-ANCA (1/40). Only LF-ANCA were associated with renal manifestations (p < 0.05), and positive predictive value for renal involvement in ANCA-positive SLE was 76.2%. LF-ANCA-positive patients had higher SLEDAI-2 K (p < 0.05) and more frequently had anti-dsDNA (p < 0.05), low C3 (p < 0.001), and low C4 (p < 0.05) than LF-ANCA-negative patients. LF-ANCA level was in a positive correlation with SLEDAI-2 K, anti-dsDNA, and anti-C1q (p < 0.01) and in a negative correlation with C3 and C4 (p < 0.05). LF-ANCA avidity was higher than MPO-, EL-, PR3-, and BPI-ANCA avidity (p < 0.01). In LF-ANCA-positive patients, renal manifestations were associated with higher LF-ANCA level (p < 0.01) and avidity (p < 0.05). Based on LF-ANCA level and avidity, the receiver operating characteristic curves for discriminating patients with and without renal involvement had areas under the curves of 0.988 (95% CI: 0.949-1.00) and 0.813 (95% CI: 0.607-1.00), respectively. After the follow-up period, number of LF-ANCA-positive patients decreased (p < 0.01). CONCLUSIONS: In contrast to other ANCAs, only LF-ANCA level correlated with activity and standard serological SLE markers. LF-ANCA level and avidity might be biomarkers of renal involvement in SLE. LF-ANCA are promising serological marker in SLE. Key Points • LF- and MPO-ANCA were most frequently found, while EL-, PR3-, and BPI-ANCA were rarely detected in SLE. • In contrast to other ANCAs, only LF-ANCA were associated with renal involvement, and their level correlated with the activity and standard serological markers of SLE. • LF-ANCA avidity was higher than other ANCAs' avidity; LF-ANCA level and avidity might be useful biomarkers of renal manifestations in SLE. • Detection of ANCA specificity, level, and avidity may help in the diagnosis of particular clinical SLE phenotypes.

The Diagnostic Importance of Recombinant Allergen IgE Testing in Patients with Hymenoptera Venom Allergy: Comparison of Two Methods.

Adults with systemic anaphylactic reactions (SAR) to insect sting show often multiple-positivity of serum-specific IgE (sIgE) to Hymenoptera venoms. Unnecessary long-lasting venom-specific immunotherapies (VIT) in false-positive patients increase the risk of recurrent SAR. This report aims to analyze the diagnostic importance of recombinant allergen IgE testing in patients with SAR to Hymenoptera sting. In 82 patients we measured sIgE to honeybee venom (HBV), wasp venom (WV) and hornet venom (HV) extracts, recombinant phospholipase A2 from HBV (sIgE-rApi m1), recombinant antigen 5 from WV (sIgE-rVes v5), and cross-reactive carbohydrate determinants-CCD-bromelain by ImmunoCAP. We analyzed the correlation of ImmunoCAP and Immunoblot for HBV and WV extracts, rApi m1, and rVes v5 in 39/82 patients. According to the history of the culprit insect, we compared sensitivity and specificity between the two methods. The severity of the SAR does not depend on the sIgE level to venom extracts and recombinant allergens. Fifty-one percent of the patients had a multiple-positivity to HBV/WV or HBV/WV/HV extracts. Severe SAR and CCD-sIgE were more frequent in multiple-positive than single-positive patients. CCD-sIgE were more frequent in HBV allergic patients than WV and HV allergic patients. There was a significant correlation between levels of sIgE to venom extracts and recombinant allergens measured by ImmunoCAP and Immunoblot. ImmunoCAP has higher sensitivity and specificity than Immunoblot for diagnosis of SAR to Hymenoptera venoms. IgE testing to recombinant CCD-free allergens is necessary for the adequate selection of long-lasting VIT, especially in patients with multiple sensitivities to venom extracts.

Combined intravenous pulse and topical corticosteroid therapy for severe alopecia areata in children: Comparison of two regimens.

There is no universally accepted treatment for severe pediatric alopecia areata (AA). This prospective study comprised 73 patients (aged 1-18 years) with severe AA (>30% of scalp surface area): 37 received 1-day intravenous dexamethasone pulses (1-DP) and 36 received 3-day pulses (3-DP), monthly, for 6-12 months. Also, all patients applied topical clobetasol propionate under plastic wrap occlusion. Patients achieving >50% regrowth were considered good responders (GR). All patients reached short term, while 65/73 were available for the long-term follow-up (mean 33.3 ± 15.3 vs. 27.7 ± 14.3 months, 1-DP and 3-DP, respectively). Relapses during therapy were more frequent in 1-DP group. 3-DP patients were more frequently GR in comparison with 1-DP. 3-DP patients with disease duration <6 months had better outcomes. Patients without Hashimoto thyroiditis (HT) had 9.8-fold higher chance of being GR in comparison with HT patients. The best results were achieved in AA plurifocalis (AAP). No patient had severe short-term side-effects. At the long-term follow-up, 67% of 3-DP patients had stable results. Only 14.2% AAP patients experienced relapses. Patients had no long-term side-effects. 3-DP were more efficacious than 1-DP. Short disease duration and no HT were good prognostic factors. 3-DP protocol is well-tolerated, with beneficial effects and long-lasting results in severe pediatric AA.

Neutrophil extracellular traps-associated markers are elevated in patients with systemic lupus erythematosus.

Neutrophil extracellular traps (NETs) are the main source of autoantigens in systemic lupus erythematosus (SLE). The aim of this study was to evaluate the clinical importance of NETs-associated markers in SLE. We compared NETs-associated markers in SLE patients (n = 111) with healthy controls (n = 50). Moreover, in 35 patients with drug-naïve SLE (n = 35), we investigated correlation between NETs-associated markers [DNase I concentration, myeloperoxidase (MPO) activity, anti-MPO antibodies, cell-free DNA (cfDNA), NETolytic activity] with serological parameters [anti-dsDNA antibodies, C3, C4 and B-cell activating factor (BAFF) levels] and disease activity measured by modified SLE Disease Activity Index (M-SLEDAI-2K). In comparison with healthy controls, SLE patients had higher cfDNA, MPO activity, anti-MPO antibodies (p < 0.001), BAFF and DNase I concentration (p < 0.01). Contrary, NETolytic activity was lower in SLE patients (p < 0.05), despite higher concentration of DNase I. MPO activity and cfDNA levels showed correlation with DNase I concentration (p < 0.001, p < 0.01, respectively). BAFF levels correlated with cfDNA, DNase I concentration and MPO activity (p < 0.05). Anti-dsDNA antibodies showed correlation with MPO activity (p < 0.01), cfDNA and BAFF levels (p < 0.001). Anti-dsDNA and C3 levels were independent predictors of M-SLEDAI-2K in multivariate analysis (p < 0.01). We demonstrated that sera of SLE patients have decreased NETolytic activity, leading to increased levels of various NETs-associated markers, which correlate with anti-dsDNA antibodies in drug-naïve SLE. We showed that BAFF participates in a complex relationship between NETosis and anti-dsDNA antibodies production. These findings have important implications for a better understanding of SLE pathogenesis and development of therapy that inhibits NETs persistence and disease progression.

Penicillin-induced Cutaneous Necrotizing Eosinophilic Vasculitis with Cryofibrinogenemia.

Cutaneous necrotizing eosinophilic vasculitis (CNEV) is a rare type of vasculitis. Eosinophilic vasculitis is a necrotizing vasculitis with eosinophilic vascular infiltration, in which eosinophils mediate vascular damage in the disease process. We present a case of an 18-year-old girl who developed palpable purpura and hemorrhagic bullae over the lower extremities associated with itching, 7 days after the commencement of penicillin therapy. Plasma cryofibrinogen was positive. Histopathology showed an infiltration of eosinophils within and around the vessel walls and a complete absence of nuclear dust and neutrophils. Oral prednisone at 1 mg/kg induced remission in 2 weeks; the prednisone dose was tapered and discontinued after 2.5 months. There was no evidence of recurrence after 37 months of follow-up. Our patient represents a rare case of drug/penicillin-induced CNEV associated with cryofibrinogenemia, without systemic organ involvement.

HCV related severe cryoglobulinemic vasculitis treated with plasma exchange and rituximab: case report and literature review.

Mixed cryoglobulinemia is the most prevalent extrahepatic manifestation of chronic HCV infection. It is usually a benign lymphoproliferative disorder which presents as vasculitis affecting different organs. Although life-threatening cryoglobulinemic vasculitis (CryoVas) is rare, it is sometimes the first and possibly lethal complication. Its treatment depends on the severity of vasculitis and can be challenging. High dose of corticosteroids, immunosuppressive agents and plasma exchange represent the first-line treatment, which should be followed by antiviral therapy. Rituximab is an effective and safe treatment option. However, the data about its use in life-threatening conditions are scarce. We report the case of a patient with severe, relapsing and life-threatening HCV-related CryoVas  resistant to standard therapy who had had an initial beneficial response to rituximab added to plasma exchange that was later compromised by the development of sepsis. We also review the literature and discuss manifestations and therapy of life-threatening Cryovas with focus on rituximab use.

Evaluation of cytokine genetic polymorphisms in adult patients with common variable immunodeficiency: A single-center study.

Common variable immunodeficiency (CVID) is a heterogeneous disease characterized by impaired B-cell differentiation and maturation accompanied with the defective antibody production. Several investigators addressed the possibility that disturbed cytokine production of TNF, IL-6, IFN-γ and IL-10, among a variety of others, may be implicated in CVID. The aim of this study was to test the hypothesis that genetic polymorphisms involving TNF (-308G/A), IFNG (+874 T/A), IL10 (-1082G/A, -819T/C and -592A/C), and IL6 (-174G/C) cytokine genes might contribute to susceptibility to CVID. Thirty five patients with CVID and 250 healthy controls were genotyped for indicated single nucleotide polymorphisms (SNP) in TNF, IL6, IFNG and IL10 using Taqman-based assays. CVID patients had significantly higher frequency of TNF A allele and AA genotype than in healthy subjects (p=0.006; OR=2.27; 95%CI=1.24-4.17 and p=0.038, OR=15.64; 95%CI=1.38-177.20, respectively). In addition, the frequency of GG genotype was significantly higher in healthy controls than in patient group (p=0.019, OR=0.43, 95%CI=0.21-0.89). Genetic analysis of IL6 SNP showed that allele G confers increased risk for CVID (p=0.037, OR=1.78, 95% CI=1.03-3.08) while IFNG allele T was associated with splenomegaly in CVID (p=0.032; OR=2.86; 95% CI=1.08-7.56). We observed no association between genotypes, alleles and haplotypes of IL-10 gene and CVID or its clinical complications. In conclusion, our results indicated association between CVID and cytokine gene polymorphisms -308G/A TNF and -174G/C IL6. In addition, we demonstrated that splenomegaly, one of the most common complications in this disease, is associated with +874T/A IFNG polymorphism. These findings add further support to the notion that cytokines may play significant role in pathogenesis of this primary antibody deficiency. However, further investigation that would involve a larger study group of CVID patients is warranted to confirm our findings.

Assessment of Deoxyribonuclease Activity in Serum Samples of Patients With Systemic Lupus Erythematosus: Fluorescence-Based Method Versus ELISA.

BACKGROUND: We report the improvement of previously described method for determining deoxyribonuclease (DNase) activity in serum samples that uses a fluorescently labeled DNA fragment as a substrate METHODS: Activity of serum DNase was analyzed in 31 patients with systemic lupus erythematosus (SLE) and 13 healthy individuals by fluoresence-based method and ELISA test RESULTS: We found a mean decrease in DNase activity between cases and controls of 12.46% measured by the fluoresence-based method and of 12.21% measured by ELISA method. High level of positive correlation between two methods for DNase activity was observed: P < 0.001 and Pearson correlation coefficient 0.740. Decreased DNase activity was found in 25 of 31 SLE patients (81%) by fluoresence-based method and in 24 of 31 SLE patients (77%) by ELISA test. We also observed the significant positive correlation between titer of anti-dsDNA antibodies and DNase activity measured by both methods (P < 0.05). CONCLUSIONS: The key improvement is the use of internal control in the fluorescence-based method, which diminishes the influence of technical errors on the obtained results and increases reliability of the assay. This improved fluorescence-based method, with additional validation, may provide an alternative to more expensive and time-consuming conventional methods, such as ELISA.

Coexistence of Autoantibodies against the Golgi Complex and Ro52 Antigen in a Patient with Nonspecific Interstitial Pneumonia.

Nonspecific interstitial pneumonia (NSIP) is often associated with connective tissue diseases (CTD). The diagnosis of NSIP was confirmed in a 63-year-old man by high-resolution computed tomography and an open lung biopsy. Anti-Golgi complex autoantibodies (AGA) and anti-Ro52 antibodies were simultaneously detected at high concentrations. Autoantibodies to aminoacyl-tRNA synthetases (ARS) were negative. The patient was treated with corticosteroids for six months. During the seven-year follow-up, NSIP had a slow progression and patient had not developed the clinical features of CTD. The present study potentially demonstrates that the autoimmune process elicited by AGA and/or Ro/SSA may play a role in promoting idiopathic NSIP independently of the typical ARS routes, which has not been reported thus far.

Hereditary Angioedema Due to C1 Inhibitor Deficiency in Serbia: Two Novel Mutations and Evidence of Genotype-Phenotype Association.

Hereditary angioedema due to C1 inhibitor deficiency (C1-INH-HAE) is a rare autosomal dominant disease characterized by recurrent life-threatening oedemas and/or abdominal pain and caused by mutations affecting the C1 inhibitor gene, SERPING1. We sought to investigate the spectrum of SERPING1 mutations in Serbia and the possible genotype-phenotype association. C1-INH-HAE was diagnosed on the basis of clinical and laboratory criteria in 40 patients from 27 families; four were asymptomatic. Mutational analysis of the SERPING1 gene was performed by sequencing and multiplex ligation-dependent probe amplification. Disease-causing mutations in SERPING1 were identified in all patients. In C1-INH-HAE type I, we identified 19 different mutations, including 6 missense mutations, 6 nonsense mutations, 2 small deletions, 1 small insertion, 2 splicing defects and 2 large deletions. Two of the mutations (c.300C>T and c.1184_1185insTA) are reported here for the first time. All C1-INH-HAE type II patients from three families harboured the same substitution (c.1396C>T). Based on the type of mutation identified in the SERPING1 gene, patients were divided into two groups: group 1 (nonsense, frameshift, large deletions/insertions, splicing defect, and mutations at Arg444) or group 2 (missense, excluding mutations at Arg444). Significant differences were found in the clinical severity score (P = 0.005), prevalence of laryngeal (P = 0.040) and facial (P = 0.013) oedema, and long-term prophylaxis (P = 0.023) between the groups with different types of mutations. Because our population consisted of related subjects, differences in the severity score between mutation groups were further confirmed using the generalized estimating equation (P = 0.038). Our study identified 20 different disease-causing mutations, including two novel mutations, in all C1-INH-HAE patients, highlighting the heterogeneity of mutations in the SERPING1 gene. Furthermore, it appears that mutations with a clear effect on C1-INH function might be responsible for a more severe disease phenotype.

Combined oral pulse and topical corticosteroid therapy for severe alopecia areata in children: a long-term follow-up study.

There are no widely accepted therapy protocols for severe alopecia areata (AA). We treated 65 children/adolescents with AA affecting >30% of scalp. Fourty-three percent of patients had AA plurifocalis (AAP). Fifty-seven percent had AA subtotalis (AAS), AAP+ophiasis (AAP+OPH), and alopecia totalis/universalis (AT/AU). Long-term follow-up (median 96 months) data were available for 69% of patients. Oral dexamethasone (prednisolone 5 mg/kg equivalent) was given once in 4 weeks. Patients received 6, 9, or 12 pulses. Clobetasol propionate 0.05% ointment under plastic wrap occlusion was applied 6 days a week. Hair growth was assessed on a scale ranging 0-100% of regrowth in individual AA lesions. Regrowth >50% was considered good response. Six to twelve months months after the therapy, 56.9% of patients had >75% of hair regrowth. In AAP, 65.5% had complete regrowth. 61.5% of all patients were considered good responders. Significantly, higher percentage of good responders was found in AA lasting ≤12 months. No patients had serious side effects. There was no change in stability of the hair status at the long-term follow-up. Most AA patients had beneficial effects with this protocol. Best results were in AAP and AAP+OPH. Combined topical and oral pulse corticosteroid therapy of AA in children shows long-lasting results, without serious side effects.

Importance of low serum DNase I activity and polyspecific anti-neutrophil cytoplasmic antibodies in propylthiouracil-induced lupus-like syndrome.

OBJECTIVE: To study the role of deoxyribonuclease (DNase) I activity and ANCA in propylthiouracil (PTU)-induced lupus-like syndrome (LLS). METHODS: We compared 36 SLE patients with 17 PTU-induced LLS patients diagnosed from 2008 to 2014. We studied ANCA profile (MPO, PR3, lactoferrin, CTG, elastase, bactericidal/permeability-increasing protein), anti-dsDNA, anti-ENA, anti-nucleosome, anti-histone, anti-C1q, anti-aCL, complement components, cryoglobulins and serum DNase I activity. Healthy persons and patients without LLS treated with PTU comprised the control groups. Twelve LLS patients were serologically and clinically followed for 4.1 (S.D. 2.0) years. RESULTS: PTU-induced LLS patients less frequently had arthritis, renal and neurological manifestations, but more frequently had fever, purpura, urticarial-like vasculitis and ulceration (P < 0.01). PTU-induced LLS patients more frequently had polyspecific ANCA (anti-MPO, anti-elastase and anti-PR3 were most commonly detected) (P < 0.01). SLE patients more frequently had anti-dsDNA, anti-ENA, anti-nucleosome, anti-C1q (P < 0.01) and anti-histone antibodies (P < 0.05). PTU-induced LLS patients had lower DNase I activity than SLE patients and controls (P < 0.01). Discontinuation of PTU increased DNase I activity, although it did not reach the levels of controls (P < 0.01). After remission, MPO-ANCA decreased (P < 0.01), but persisted for a long time. CONCLUSION: PTU, as a trigger, and low DNase I activity, as a predisposing factor, may lead to LLS. Polyspecific ANCAs are useful markers for differentiating SLE from PTU-induced LLS. Low DNase I activity might be an important prognostic biomarker for PTU-induced LLS. Monitoring of ANCA and DNase I activity may prevent long-lasting exposure to causal drugs, unnecessary immunosuppressive therapy and severe complications of LLS.

Fatal sulfasalazine-induced eosinophilic myocarditis in a patient with periodic fever syndrome.

OBJECTIVE: The aim of this paper is to report the first case of drug-induced eosinophilic myocarditis (EM) in a patient with hereditary periodic fever syndrome (PFS). CASE: A 28-year-old man with hyper-IgD syndrome, one of the PFS, developed a sulfasalazine-induced systemic hypersensitivity reaction complicated by EM. Thirteen days after sulfasalazine introduction, which had been given for arthritis, the patient developed fever, facial/neck edema, rash and cardiogenic shock, and died within 8 h. The autopsy revealed hemophagocytosis, while acute heart failure caused by necrotizing EM was established as the cause of death. CONCLUSION: This was a case of drug-induced EM in a patient with PFS that had an atypical presentation, rapid evolution and poor outcome.