Associations between clinical features and therapy with macrophage subpopulations and T cells in inflammatory lesions in the aorta from patients with Takayasu arteritis.

Takayasu arteritis (TAK) is a large-vessel granulomatous vasculitis; the inflammatory infiltration in arteries comprises macrophages, multi-nucleated giant cells, CD4+ and CD8+ T cells, γδ T cells, natural killer (NK) cells and neutrophils. However, it is unknown which subtype of macrophages predominates. This study aims to evaluate macrophages subpopulations in the aorta in TAK. Immunohistochemistry was performed in the aorta from TAK patients (n = 22), patients with atherosclerotic disease (n = 9) and heart transplant donors (n = 8) using the markers CD68, CD86, CD206, CD3, CD20 and CD56. Active disease was observed in 54·5% of patients and active histological lesions were found in 40·9%. TAK patients presented atherosclerotic lesions in 27·3% of cases. The frequency of macrophages, M1 macrophages, T, B and NK cells was higher in the aorta from TAK and atherosclerotic patients compared to heart transplant donors. In TAK, macrophages and T cells were the most abundant cells in the aorta, and the expression of CD206 was higher than CD86 (P = 0·0007). No associations were found between the expression of cell markers and active disease or with atherosclerotic lesions. In TAK patients, histological disease activity led to higher T cell counts than chronic fibrotic lesions (P = 0.030), whereas prednisone use was associated with lower T cell counts (P = 0·035). In conclusion, M1 macrophages were more frequent in TAK and atherosclerotic patients compared to heart transplant donors, while M2 macrophages dominated M1 macrophages in TAK. T cells were associated with histological disease activity and with prednisone use in TAK.

Diagnostic accuracy of thromboelastometry and its correlation with the HPLC-MS/MS quantification test.

The aim of the study was to evaluate the diagnostic accuracy of thromboelastometry for assessing rivaroxaban concentrations. The accuracy of thromboelastometry was compared with the high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method, which is the gold standard for drug plasma monitoring (the reference standard). Forty-six clinically stable patients were treated with 10, 15, or 20 mg of rivaroxaban once daily (OD group) or 15 mg twice a day (BID group) (no particular indication for treatment). Patient samples were collected 2 h after the use of the medication (peak) and 2 h before the next dose (trough). The rivaroxaban plasma concentrations were determined via HPLC-MS/MS, and thromboelastometry was performed using a ROTEM® delta analyzer. There were significant prolongations in clotting time (CT) for the 10, 15, and 20 mg of rivaroxaban treatments in the OD groups. In the 15 mg BID group, the responses at the peak and trough times were similar. At the peak times, there was a positive correlation between the plasma concentration of rivaroxaban and CT (Spearman correlation rho=0.788, P<0.001) and clot formation time (rho=0.784, P<0.001), and a negative correlation for alpha angle (rho=-0.771, P<0.001), amplitude after 5 min (rho=-0.763, P<0.001), and amplitude after 10 min (rho=-0.680, P<0.001). The CT presented higher specificity and sensitivity using the cut-off determined by the receiver characteristics curve. ROTEM has potential as screening tool to measure possible bleeding risk associated with rivaroxaban plasma levels.

Pre-analytical parameters associated with unsuccessful karyotyping in myeloid neoplasm: a study of 421 samples.

Cytogenetics is essential in myeloid neoplasms (MN) and pre-analytical variables are important for karyotyping. We assessed the relationship between pre-analytical variables (time from collection to sample processing, material type, sample cellularity, and diagnosis) and failures of karyotyping. Bone marrow (BM, n=352) and peripheral blood (PB, n=69) samples were analyzed from acute myeloid leukemia (n=113), myelodysplastic syndromes (n=73), myelodysplastic syndromes/myeloproliferative neoplasms (n=17), myeloproliferative neoplasms (n=137), and other with conclusive diagnosis (n=6), and reactive disorders/no conclusive diagnosis (n=75). The rate of unsuccessful karyotyping was 18.5% and was associated with the use of PB and a low number of nucleated cells (≤7×103/µL) in the sample. High and low cellularity in BM and high and low cellularity in PB samples showed no metaphases in 3.9, 39.7, 41.9, and 84.6% of cases, respectively. Collecting a good BM sample is the key for the success of karyotyping in MN and avoids the use of expensive molecular techniques.

Diagnostic accuracy of thromboelastometry and its correlation with the HPLC-MS/MS quantification test

The aim of the study was to evaluate the diagnostic accuracy of thromboelastometry for assessing rivaroxaban concentrations. The accuracy of thromboelastometry was compared with the high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method, which is the gold standard for drug plasma monitoring (the reference standard). Forty-six clinically stable patients were treated with 10, 15, or 20 mg of rivaroxaban once daily (OD group) or 15 mg twice a day (BID group) (no particular indication for treatment). Patient samples were collected 2 h after the use of the medication (peak) and 2 h before the next dose (trough). The rivaroxaban plasma concentrations were determined via HPLC-MS/MS, and thromboelastometry was performed using a ROTEM® delta analyzer. There were significant prolongations in clotting time (CT) for the 10, 15, and 20 mg of rivaroxaban treatments in the OD groups. In the 15 mg BID group, the responses at the peak and trough times were similar. At the peak times, there was a positive correlation between the plasma concentration of rivaroxaban and CT (Spearman correlation rho=0.788, P<0.001) and clot formation time (rho=0.784, P<0.001), and a negative correlation for alpha angle (rho=−0.771, P<0.001), amplitude after 5 min (rho=−0.763, P<0.001), and amplitude after 10 min (rho=−0.680, P<0.001). The CT presented higher specificity and sensitivity using the cut-off determined by the receiver characteristics curve. ROTEM has potential as screening tool to measure possible bleeding risk associated with rivaroxaban plasma levels.

Pre-analytical parameters associated with unsuccessful karyotyping in myeloid neoplasm: a study of 421 samples

Cytogenetics is essential in myeloid neoplasms (MN) and pre-analytical variables are important for karyotyping. We assessed the relationship between pre-analytical variables (time from collection to sample processing, material type, sample cellularity, and diagnosis) and failures of karyotyping. Bone marrow (BM, n=352) and peripheral blood (PB, n=69) samples were analyzed from acute myeloid leukemia (n=113), myelodysplastic syndromes (n=73), myelodysplastic syndromes/myeloproliferative neoplasms (n=17), myeloproliferative neoplasms (n=137), and other with conclusive diagnosis (n=6), and reactive disorders/no conclusive diagnosis (n=75). The rate of unsuccessful karyotyping was 18.5% and was associated with the use of PB and a low number of nucleated cells (≤7×103/µL) in the sample. High and low cellularity in BM and high and low cellularity in PB samples showed no metaphases in 3.9, 39.7, 41.9, and 84.6% of cases, respectively. Collecting a good BM sample is the key for the success of karyotyping in MN and avoids the use of expensive molecular techniques.

The PTPN22 1858T allele but not variants in the proximal promoter region of IL-21 gene is associated with the susceptibility to type 1 diabetes and the presence of autoantibodies in a Brazilian cohort.

Interleukin (IL)-21 and protein tyrosine phosphatase non-receptor 22 (PTPN22) regulate lymphocyte function and have been implicated in the pathogenesis of autoimmune diabetes. We sequenced the proximal promoter of the IL-21 gene for the first time and analysed the PTPN22 1858T polymorphism in type 1A diabetes (T1AD) patients and healthy controls (HC). We correlated the frequencies of islet and extra-pancreatic autoantibodies with genotypes from both loci. The case series comprised 612 T1AD patients and 792 HC. Genotyping of PTPN22 C1858T was performed on 434 T1AD patients and 689 HC. The -448 to +83 base pairs (bp) region of the IL-21 gene was sequenced in 309 Brazilian T1AD and 189 HC subjects. We also evaluated human leucocyte antigen (HLA) DR3/DR4 alleles. The frequencies of glutamic acid decarboxylase (GAD65), tyrosine phosphatase-like protein (IA)-2, anti-nuclear antibody (ANA), thyroid peroxidase (TPO), thyroglobulin (TG), thyrotrophin receptor autoantibody (TRAb), anti-smooth muscle (ASM) and 21-hydroxylase (21-OH) autoantibodies were higher in T1AD patients than in HC. The PTPN22 1858T allele was associated with an increased risk for developing T1AD [odds ratio (OR) = 1·94; P < 0·001], particularly in patients of European ancestry, and with a higher frequency of GAD65 and TG autoantibodies. HLA-DR3/DR4 alleles predominated in T1AD patients. A heterozygous allelic IL-21 gene variant (g.-241 T > A) was found in only one patient. In conclusion, only PTPN22 C1858T polymorphism and HLA-DR3 and/or DR4 alleles, but not allelic variants in the 5'-proximal region of the IL-21 gene were associated with T1AD risk. Patients with T1AD had increased frequencies of anti-islet-cell, anti-thyroid, anti-nuclear, anti-smooth muscle and anti-21-OH autoantibodies. The C1858T PTPN22 polymorphism was also associated with a higher frequency of GAD65 and TG autoantibodies.

Antinucleosome antibodies in patients with juvenile systemic lupus erythematosus.

Our objective was to evaluate the frequency of antinucleosome antibodies (anti-Ncs) in juvenile systemic lupus erythematosus (JSLE) comparing it to that observed for anti-DNA and to correlate the presence of these antibodies with clinical manifestations and disease activity. Anti-Ncs and anti-DNA were detected by ELISA in 74 patients with JSLE and 64 normal controls. Clinical records were reviewed. Disease activity was assessed by SLEDAI score. Anti-Ncs and anti-DNA showed sensitivity of 52.7% and 54% and specificity of 98.4% and 95.3%, respectively. Disagreement between the two assays was found in 25.7% of the cases: isolated positive Anti-Ncs in nine cases (12.2%) and isolated positive anti-DNA in 10 cases (13.5%). Agreement was found in 74.3%: both positive antibodies in 30 cases and both negative in 25. The presence of anti-Ncs was significantly associated with malar erythema, hemolytic anemia, anti-DNA and low complement levels, but not with renal manifestations. The presence of anti-Ncs was associated with a higher SLEDAI median (P < 0.001) and its titers correlated with the SLEDAI score (r = 0.504; P < 0.001). The frequency, sensitivity and specificity values were similar between anti-Ncs and anti-DNA antibodies in patients with JSLE. Nevertheless, the discordance of 25.7% between the two assays suggests that both antibodies may have a complementary diagnostic role. The association and correlation between anti-Ncs and several disease activity parameters demonstrated its usufulness in the follow-up of these patients.