The feasible maintenance dose of corticosteroid in Takayasu arteritis in the era of biologic therapy.

Objective: Although biologic agents are used in Takayasu arteritis (TAK), corticosteroids are still the mainstay of treatment. This study aimed to investigate the feasible maintenance dose of prednisolone (PSL) in the biologic therapy era.Method: We enrolled 93 patients with TAK who satisfied the criteria of the American College of Rheumatology and visited our department from 2008 to 2018. The clinical characteristics and PSL dose of the patients were retrospectively evaluated.Results: The mean ± sd maintenance dose of PSL was 5.0 ± 3.0 mg/day. In patients having TAK for > 20 years, PSL discontinuation and drug-free status were achieved in 27.2% and 18%, respectively. Although tapering the PSL dose to 10 mg/day was achieved within 12 months, tapering to 5 mg/day required 10 years. Relapse significantly interfered with the PSL dose reduction. The clinical characteristics of patients with relapse included a lower rate of combination therapy using immunosuppressants. Moreover, biologics were used in > 60% of patients with relapse. Tapering of PSL was significantly possible in patients receiving biologics and additional relapse was observed in 6.3% and 50% of patients with and without biologics, respectively. Such PSL-sparing effect enabled the reduction of the median PSL dose from 10 to 5 mg/day. Steroid discontinuation was achieved in some patients.Conclusions: The use of biologics significantly reduced the PSL dose in relapsed patients. A PSL dose of ≤ 5 mg/day is a feasible target for TAK, especially when biologic agents are used. Nevertheless, corticosteroid discontinuation may also be the target in some patients.

Four distinct clinical phenotypes of vasculitis affecting medium-sized arteries.

Objective: Within the spectrum of polyarteritis nodosa (PAN), cutaneous PAN (cPAN) is further classified into mild cPAN and severe cPAN which presents with ulcers, necrosis, or neuritis. As distinguishing between severe cPAN and systemic PAN can be difficult, this study evaluated the clinical characteristics of patients with necrotizing arteritis of medium-sized arteries. Methods: Forty-one patients diagnosed with necrotizing arteritis of medium-sized arteries between 2008 and 2017 at our institution were enrolled in this study. Clinical background, laboratory findings, treatments, and rates of relapse and death were evaluated. Results: Thirty-six patients were classified as having cPAN (mild, 15; ulcer, nine; neuritis, eight; both, four), and five cases manifested systemic vasculitis. Clinical characteristics of mild cPAN included female predominance (84.6%) and younger age (median 31 years); those of systemic PAN included older age (median 71 years) and higher levels of inflammatory markers. Severe cPAN manifested with intermediate phenotypes. The median doses of prednisolone used to treat mild cPAN, severe cPAN, and systemic PAN were 20.0, 40.0, and 40.0 mg/day, respectively. Immunosuppressants were used in 20.0% of mild cPAN, 90.5% of severe cPAN, and 80.0% of systemic PAN patients. Although the mortality rates were indistinguishable, the relapse rates of severe cPAN (ulcer type) were significantly higher than those of other types (88.9%). Conclusion: The clinical characteristics of mild cPAN, severe cPAN (ulcer type), severe cPAN (neuritis type), and systemic PAN were distinct from each other. In particular, patients with severe cPAN (ulcer type) had higher relapse rates, indicating the importance of combination therapy.

A single-stranded DNA-cross-reactive immunogenic epitope of human homocysteine-inducible endoplasmic reticulum protein.

The mechanism involved in generating anti-DNA antibodies (Abs) remains unclear, as DNA is poorly immunogenic. Molecular mimicry between DNA and non-DNA substances has been implicated as a possible mechanism. We previously reported that homocysteine-inducible endoplasmic reticulum protein (Herp), which is induced by endoplasmic reticulum stress, is recognized by anti-double-stranded DNA (dsDNA) IgG from patients with systemic lupus erythematosus and that immunization with Herp elicits anti-dsDNA Abs in BALB/c mice. In this study, we observed that anti-single-stranded DNA (ssDNA) Abs were also generated in Herp-immunized BALB/c mice and established an anti-Herp monoclonal antibody (mAb), HT4, which specifically cross-reacted with ssDNA. The epitope of the HT4 mAb on Herp, 'EPAGSNR', was identified by screening a synthetic peptide library. The binding of the HT4 mAb to the peptide was competitively inhibited by ssDNA. Immunization of the epitope peptide elicited anti-ssDNA Abs in BALB/c mice. These results indicate that the epitope exists in a human self-protein, mimics ssDNA and shows antigenicity for anti-ssDNA Abs in normal mice. Anti-ssDNA Abs are often found in patients with drug-induced lupus erythematosus. Treatment with representative drugs that cause drug-induced lupus (chlorpromazine, procainamide and hydralazine) induced Herp expression and apoptosis in HeLa cells. These findings suggest that molecular mimicry between Herp and ssDNA is involved in anti-ssDNA Ab production in drug-induced lupus.

Myeloablative cord blood transplantation for adults with hematological malignancies using tacrolimus and short-term methotrexate for graft-versus-host disease prophylaxis: single-institution analysis.

We studied clinical outcomes of 25 adult patients with hematological malignancies who underwent cord blood transplantation (CBT) after a myeloablative conditioning regimen, including high-dose cytosine arabinoside (CA) (8 g/m(2)), cyclophosphamide (CY) (120 mg/kg), and total-body irradiation (TBI) (12 Gy). For graft-versus-host disease (GVHD) prophylaxis, all patients received a combination of tacrolimus and short-term methotrexate (sMTX). Neutrophil engraftment was achieved in 20 of 25 patients. Of the 22 evaluable patients, 2 and 7 had grades I and II acute GVHD, respectively, and only 1 developed grade III acute GVHD after discontinuation of tacrolimus due to encephalopathy. Chronic GVHD developed in 13 of 19 evaluable patients, including 4 with the extensive type. However, the Karnofsky scores of survivors at 1 year after CBT were 90% or 100%. Eight of 25 patients died of nonrelapse causes (n = 4) and relapse/progressive disease (n = 4); 17 patients are currently alive with 15 free of disease at the present time (median follow-up, 24 months). The probability of disease-free survival at 2 years among patients with standard risk was 89% and that of high-risk patients was 30%. Transplantation-related mortality within 100 days was 12%. These results suggested that the CA/CY/TBI combination is a promising conditioning regimen for myeloablative CBT. Furthermore, tacrolimus and sMTX seemed to have suppressed severe acute GVHD and chronic GVHD, which may also contribute to the favorable results.

Evaluation of a fully automated system (RAISUS) for rapid identification and antimicrobial susceptibility testing of Staphylococci.

RAISUS is a system for rapid bacterial identification and antimicrobial susceptibility testing. RAISUS and VITEK showed 97.8% and 75.9% agreement in identification of 45 Staphylococcus aureus strains and 58 coagulase-negative staphylococci (CoNS), respectively, and RAISUS and CLSI (formerly NCCLS) methods showed 87.2% and 87.9% agreement in the MICs for S. aureus and CoNS, respectively. RAISUS provided these data within 3.75 h, suggesting its utility for clinical bacteriological laboratories.

Residual formaldehyde on plastic materials and medical equipment following low-temperature steam and formaldehyde sterilization.

We measured the amount of residual formaldehyde on 16 plastic materials and five medical devices following low-temperature steam and formaldehyde (LTSF) sterilization, based on the European Standard EN14180. The amounts of formaldehyde residue on the plastic materials were compared with that on a filter paper of similar dimensions. The amount of residual formaldehyde on polyamide 6, polyurethane, natural rubber and polyacetal was higher (21.9, 15.2, 3.0 and 2.1 times, respectively) than that on the filter paper. The amount of formaldehyde recovered from a breathing circuit, anaesthesia circuit, oxygen tubing, airway tube and tweezers was 260, 240, 594, 56 and 0 microg, respectively, following LTSF sterilization. Our results emphasize the need to verify the main material composing the medical equipment before LTSF sterilization, as the amount of formaldehyde retrieved following sterilization varies according to the material used for construction.

Evaluation of a low-temperature steam and formaldehyde sterilizer.

We evaluated a low-temperature steam and formaldehyde (LTSF) sterilizer based on the draft European Standard prEN 14180. Microbiological tests were conducted on small and full loads using process challenge devices in five programs (P1-P5). With small loads all tests showed no growth of Bacillus stearothermophilus (ATCC7953) spores. However, positive cultures were observed with full-load tests using P5 (sterilization temperature, 50 degrees C). Our data indicated that the load influenced the efficacy of the LTSF sterilizer. Desorption tests were conducted to determine residual formaldehyde in indicator strips. The mean concentrations of formaldehyde in P1-P5 were 31.9, 56.3, 54.9, 82.2 and 180.6 microg, respectively, which are below the limits allowed by the draft Standard. Our results indicate that the LTSF sterilizer is useful for sterilization because of its excellent efficacy, short handling time, and safety.

Autoimmune neutropenia in pregnant women causing neonatal neutropenia.

Autoimmune neutropenia (AIN) can occur during pregnancy. However, neonatal neutropenia occurring in an infant born to a mother with AIN has only rarely been documented. Recently, we have experienced two cases of AIN during pregnancy, both of which caused severe yet transient neonatal neutropenia (< 0.3 x 10(9)/l), probably as a result of transplacental maternal anti-neutrophil autoantibodies. The anti-neutrophil antibodies seemed to be against antigens other than NA1/NA2 because the autoantibodies did not bind to neutrophils of specific NA types selectively in the granulocyte indirect immunofluorescence test. Although AIN is a relatively uncommon disease, neonatal neutropenia caused by maternal AIN may not be quite as rare.

Childhood T-cell acute lymphoblastic leukemia with four distinct immunophenotypes representing different stages of T-cell development.

The authors report on a 14-year-old boy who developed T-cell acute lymphoblastic leukemia (FAB:L1) displaying 4 immunophenotypically distinct leukemic cell populations by 3-color immunofluorescence staining. Cytogenetic analysis at diagnosis showed 46,XY,add(4)(p16)[12]/46,XY[2]. A single rearrangement of the T-cell antigen receptor beta- and gamma-chain genes in these cells indicated monoclonality of the leukemic cells. These findings suggest that leukemic blast cells of monoclonal origin in this case were divided into 4 immunophenotypic populations, representing various stages of differentiation.

Decreased expression of transcription factor GATA-2 in haematopoietic stem cells in patients with aplastic anaemia.

Aplastic anaemia is characterized by reduced haematopoiesis resulting in pancytopenia. It has been speculated that there is an injury in haematopoietic stem cells in the bone marrow; however, the precise nature of the injury has not been elucidated. In this study, the levels of expression of mRNAs for three transcription factors, GATA-2, SCL and AML1, which function in the early stages of haematopoiesis, were examined by quantitative polymerase chain reaction in patients with aplastic anaemia, idiopathic thrombocytopenic purpura (ITP) and normal subjects. Among these factors, expression of GATA-2 mRNA in purified CD34-positive cells was markedly decreased in aplastic anaemia compared with that in ITP and in normal subjects. The expression levels of SCL and AML1 mRNA in CD34-positive cells in aplastic anaemia were not different from those in normal subjects. When the expression of GATA-2 protein in CD34-positive cells was examined by immunocytochemical analysis, the percentage of GATA-2-positive cells in aplastic anaemia was lower than that in normal subjects. These findings strongly suggest that there is an aberrant expression of transcription factors in stem cells in aplastic anaemia, which may be responsible for the development of the disease.

Clonal evolution from trisomy into tetrasomy of chromosome 8 associated with the development of acute myeloid leukemia from myelodysplastic syndrome.

Tetrasomy 8, though rare, is usually associated with trisomy 8, a far more common chromosomal abnormality in acute myeloid leukemia (AML). Yet the clonal relationship between trisomy 8 and tetrasomy 8 in the cases with these chromosomal abnormalities has been unclear. Here, we report a case of a 17-year-old male, diagnosed as having a myelodysplastic syndrome (MDS). Chromosome analysis showed the presence of trisomy 8. Five years later, he developed overt AML exhibiting tetrasomy 8 only. After chemotherapy, the blast cells in the bone marrow decreased to 3.4%, and the karyotype showed trisomy 8 alone. Fluorescence in situ hybridization using a probe specific for chromosome 8 showed that the percentages of cells exhibiting 2/ 3 /4 signals were 7.8/89.2/2.0 at the MDS stage, 20.5/36.1/41.0 when overt AML developed and 24.0/72.1/2.4 after chemotherapy. These results suggested that tetrasomy 8 is derived from the AML clone, possibly evolved from the MDS clone with trisomy 8. To our knowledge, this is the first detailed case report of clonal evolution from trisomy 8 into tetrasomy 8 associated with the development of AML from MDS.

Genomic structure and regulation of a novel human gene, Klp1.

Klp1 (K562 cells-derived leucine zipper-like protein 1) is a transcription factor which binds to the coproporphyrinogen oxidase promoter regulatory element (GGACTACAG). In order to clarify the function of Klp1, we determined the complete human Klp1 genomic structure and regulatory element in the promoter region. The gene spans about 2.4 kb and has three exons. Its promoter region has multiple GC boxes, E2F binding site, one cAMP response element (CRE), and no TATA box with multiple transcription initiation sites, which is characteristic of housekeeping and growth regulating genes. Promoter analysis showed that the promoter was more active in K562 cells entered into the cell cycle by serum stimulation than quiescent cells. Further promoter analysis revealed that CRE at -42 is essential for full promoter activity, and c-Jun and activation transcription factor 1/cAMP response element binding protein 1 proteins bind to this element. These structural characteristics and the promoter function suggest that Klp1 may play a role in cell cycle regulation.

Molecular analysis of delta-aminolevulinate dehydratase deficiency in a patient with an unusual late-onset porphyria.

Cloning, expression, and genotype studies of the defective gene for delta-aminolevulinate dehydratase (ALAD) in a patient with an unusual late onset of ALAD deficiency porphyria (ADP) were carried out. This patient was unique in that he developed the inherited disease, together with polycythemia, at the age of 63. ALAD activity in erythrocytes of the patient was less than 1% of the normal control level. ALAD complementary DNA (cDNA) isolated from the patient's Epstein-Barr virus (EBV)-transformed lymphoblastoid cells had 2 base transitions in the same allele, G(177) to C and G(397) to A, resulting in amino acid substitutions K59N and G133R, respectively. It has been verified that the patient had no other ALAD mutations in this and in the other allele. By restriction fragment length polymorphism (RFLP) analysis, all family members of the proband who had one-half ALAD activity compared with the ALAD activity of the healthy control were shown to have the same set of base transitions. Expression of ALAD cDNA in CHO cells revealed that K59N cDNA produced a protein with normal ALAD activity, while G133R and K59N/G133R cDNA produced proteins with 8% and 16% ALAD activity, respectively, compared with that expressed by the wild type cDNA. These findings indicate that while the proband was heterozygous for ALAD deficiency, the G(397) to A transition resulting in the G133R substitution is responsible for ADP, and the clinical porphyria developed presumably due to an expansion of the polycythemic clone in erythrocytes that carried the mutant alad allele.

Cloning of a coproporphyrinogen oxidase promoter regulatory element binding protein.

Coproporphyrinogen oxidase [CPO] gene promoter regulatory element (CPRE) plays an important role in CPO gene regulation. To isolate a CPRE binding protein, we performed Southwestern screening of K562 cDNA expression library using CPRE as a probe and isolated a cDNA clone which encoded a novel protein, Klp1 (K562 cell-derived leucine-zipper-like protein 1). Klp1 mRNA was highly expressed in K562 cells, HeLa cells, and brain as a single transcript (1.4 kb). Gel mobility shift assays revealed that Klp1 specifically binds to CPRE. Computational analysis revealed that Klp1 has a leucine-zipper-like structure, a Leu-X-X-Leu-Leu motif, and a putative nuclear localization signal in the basic amino acid rich region. Transfection of the Klp1 expression vector into THP-1 cells resulted in transcriptional activation of a reporter construct containing CPRE. These results indicate that Klp1 is a DNA sequence-specific transcription factor that regulates gene expression of genes that contain CPRE in their regulatory region.