Effective treatment by glycolic acid peeling for cutaneous manifestation of familial generalized acanthosis nigricans caused by FGFR3 mutation.

BACKGROUND: Acanthosis nigricans (AN) can occur as a cutaneous manifestation of genetic diseases, one of which is associated with activating mutations of the fibroblast growth factor receptor 3 gene (FGFR3). OBJECTIVE: We explored familial AN patients with FGFR3 mutations and examined the effectiveness of glycolic acid (GA) peeling in improving their skin manifestations. METHODS: Sanger sequencing was performed for the genomic DNA extracted from leucocytes of the family members involving familial AN. GA peeling was carried out for the two patients of familial AN once every 2 weeks. RESULTS: Heterozygous c.1949A>C (p.K650T) mutation in FGFR3 was identified for the affected family members examined, whereas the wild-type sequence was found for two unaffected individuals. Hyperpigmentation and coarseness of the skin were improved by GA peeling at regular intervals with few adverse effects. CONCLUSION: We diagnosed our cases as familial generalized AN caused by heterozygous c.1949A>C (p.K650T) mutation of FGFR3. We propose that GA peeling is a useful and safe therapeutic option to treat familial AN.

Control of an outbreak of carbapenem-resistant Pseudomonas aeruginosa in a haemato-oncology unit.

An outbreak of a multidrug-resistant Pseudomonas aeruginosa producing metallo-ß-lactamase (MBLPA) in a haemato-oncology unit was controlled using multidisciplinary interventions. The present study assesses the effects of these interventions by active surveillance of the incidence of MBLPA infection at the 1,240-bed tertiary care Kyoto University Hospital in Kyoto, Japan. Infection control strategies in 2004 included strengthening contact precautions, analysis of risk factors for MBLPA infection and cessation of urine collection. However, new MBLPA infections were identified in 2006, which prompted enhanced environmental cleaning, routine active surveillance, and restricting carbapenem usage. Between 2004 and 2010, 17 patients in the unit became infected with indistinguishable MBLPA strains. The final five infected patients were found by routine active surveillance, but horizontal transmission was undetectable. The MBLPA outbreak in the haemato-oncology unit was finally contained in 2008.

Pretransplant serum ferritin and C-reactive protein as predictive factors for early bacterial infection after allogeneic hematopoietic cell transplantation.

Although fluoroquinolones or other antibiotics are commonly used to prevent bacterial infections after hematopoietic cell transplantation (HCT), because of the growing presence of multidrug-resistant microorganisms, it is important to identify patients who are more likely to benefit from antibacterial prophylaxis. To evaluate risk factors for early bacterial infection after allogeneic HCT, we retrospectively analyzed clinical data for 112 consecutive adult patients with hematological malignancies who received transplants without any antibacterial prophylaxis. The cumulative incidence of bacterial infection at 30 days after transplantation was 16%. Among various pre-transplant factors, only high serum ferritin (>700 ng/mL, 47 patients) and high C-reactive protein (CRP) (>0.3 mg/dL, 28 patients) levels were significantly associated with the development of bacterial infection in a multivariate analysis (hazard ratio (95% confidence interval): ferritin, 4.00 (1.32-12.17); CRP, 3.64 (1.44-9.20)). In addition, septic shock and sepsis with organ failure were exclusively observed in patients who had high ferritin and/or high CRP levels. These results suggest that pretransplant serum ferritin and CRP levels can be useful markers for predicting the risk of early bacterial infection after allogeneic HCT. It may be prudent to limit antibacterial prophylaxis to patients with predefined risk factors to ensure the safety of HCT with the use of fewer antibiotics.

Close cooperation between infectious disease physicians and attending physicians can result in better management and outcome for patients with Staphylococcus aureus bacteraemia.

Staphylococcus aureus bacteraemia (SAB) is a serious infection that demands prompt clinical attention for good outcome. To assess the impact of intervention by infectious diseases physicians (IDPs) in cases with SAB, a retrospective cohort study of patients with SAB was performed in a 1240-bed, university hospital in Japan, with the aim of comparing the management and outcome of patients during the initial and the latter half of the intervention period,. Three hundred and forty-six patients with SAB during the 7-year period, from 2002 to 2008, were included, and 194 patients in the initial half of the period (from 2002 to 2005) were compared with 152 patients in the later period (from 2006 to 2008). There was no significant difference between the two groups with respect to patient's clinical background, although more patients in the later period were receiving immunosuppressive treatment. The proportion of methicillin resistant S. aureus was lower during the later period (56.2% vs. 43.3%; p 0.02). Echocardiography was used more frequently (37.1% vs. 64.5%; p < 0.001). Infective endocarditis and metastatic infections were diagnosed more frequently (10.8% vs. 20.4%; p 0.01). Follow-up blood cultures were obtained more regularly (52.1% vs. 73.7%; p <0.001) and therapy was more frequently administered for at least 14 days (47.4% vs. 82.2%; p <0.001). The 30-day mortality improved during the intervention period (25.8% vs. 16.4%; p 0.04). The total number of blood cultures received by the laboratory increased annually and the total number of consultations increased by approximately 1.6-fold compared to 2002. Proactive intervention by IDPs raised awareness of optimal management of bacteraemia and improved the adherence to the standards of care, which subsequently resulted in an improvement in the outcome.

Identifying pathogenic genetic background of simplex or multiplex retinitis pigmentosa patients: a large scale mutation screening study.

BACKGROUND AND PURPOSE: More than half of the retinitis pigmentosa (RP) cases are genetically simplex or multiplex. To date, 37 causative genes of RP have been identified; however, the elucidation of gene defects in simplex or multiplex RP patients/families remains problematic. The aim of our study was to identify the genetic causes of RP in patients with unknown or non-Mendelian inheritance. METHODS AND RESULTS: Since 2003, 52 simplex RP patients, 151 patients from 141 multiplex RP families, and six sporadic patients with retinal degeneration were studied. A total of 108 exons of 30 RP-causing genes that harboured the reported mutations were screened by an efficient denaturing high performance liquid chromatography (dHPLC) based assay. Aberrant fragments were subsequently analysed by automatic sequencing. Twenty-six mutations, including two frameshift mutations, one single amino acid deletion, and 23 missense mutations, were identified in 28 probands (14.07%). Eighteen mutations have not been reported to date. Three pairs of combined mutations in different genes were identified in two sporadic cases and one multiplex family, indicating the possibility of novel digenic patterns. Of the 23 missense mutations, 21 were predicted as deleterious mutations by computational methods using PolyPhen, SIFT, PANTHER, and PMut programs. CONCLUSION: We elucidated the mutation spectrum in Japanese RP patients and demonstrated the validity of the mutation detection system using dHPLC sequencing for genetic diagnosis in RP patients independent of familial incidence, which may provide a model strategy for identifying genetic causes in other diseases linked to a wide range of genes.

Effects on spectrum and susceptibility patterns of isolates causing bloodstream infection by restriction of fluoroquinolone prophylaxis in a hematology-oncology unit.

The emergence of fluoroquinolone-resistant gram-negative organisms has been demonstrated in patients given fluoroquinolone prophylaxis. To prevent increases in resistant bacteria, we restricted prophylactic use of fluoroquinolones. The spectrum and susceptibility patterns of isolates causing bloodstream infection (BSI) were assessed in patients receiving chemotherapy during periods of routine prophylaxis (period A: October 2001 to May 2003) and restricted prophylaxis (period B: June 2003 to January 2005). The total number of patients receiving chemotherapy was 442 during period A and 365 during period B. No significant differences were seen between periods with respect to patient characteristics. BSI was identified in 42 patients (44 episodes) during period A and 69 patients (74 episodes) during period B. Incidence of BSI increased significantly from 10.0% (44/442) during period A to 20.3% (74/365) during period B (P < 0.0001). Rate of Enterobacteriaceae BSI increased significantly, from 2.0% (9/442) during period A to 8.2% (30/365) during period B (P < 0.0001). For all BSI episodes, the proportion of BSI with gram-positive cocci decreased from 63.6% (28/44) during period A to 44.6% (33/74) during period B (P = 0.045), while the proportion of BSI with Enterobacteriaceae increased from 20.5% (9/44) to 40.5% (30/74) (P < 0.0001). The proportion of fluoroquinolone-resistant Enterobacteriaceae BSI for all Enterobacteriaceae BSI decreased from 75% (9/12) during period A to 17% (5/30) during period B (P = 0.0078). Restriction of fluoroquinolone prophylaxis affects the etiology of BSI and reduces the proportion of drug-resistant organisms.

Perioperative antimicrobial prophylaxis in urology: a multi-center prospective study.

Since there are few published reports regarding the impact of urologic surgery on perioperative infections, an epidemiologic analysis was performed on data from 1,156 open or laparoscopic operations in urology collected by the 21 hospitals participating in this study between September 2002 and August 2003. Prophylactic antibiotics were administered intravenously according to our protocol designed on the basis of the invasiveness and contamination levels. The surgical site infection (SSI) rates following clean, clean-contaminated and contaminated surgery were 1.2%. 5.8% and 23.4%, respectively, while the remote infection (RI) rates were 3.5%. 7.1% and 35.9%, respectively. Methicillin-resistant Staphylococcus aureus (MRSA) was most frequently isolated from SSIs as well as RIs, whereas Enterococcus faecalis and Pseudomonas aeruginosa were more frequently discovered in RIs than in SSIs. Several risk factors for SSI and/or RI, such as older age, high ASA score, obesity, diabetes, preoperative chemotherapy, long operation time and much blood loss, were identified by univariate analysis.

Clinical factors associated with fluconazole resistance and short-term survival in patients with Candida bloodstream infection.

In a 1-year national surveillance program of Candida bloodstream infections in Japan, clinical factors predicting fluconazole resistance and survival of the patients were analyzed. Blood isolates and complete clinical histories were obtained from 326 patients. Fluconazole-resistant isolates were found in 15 (4.6%) of the cases. Univariate analysis of the demographic and clinical factors associated with fluconazole resistance revealed that age, hematologic malignancy, neutropenia, and immunosuppression were of statistical significance. A multiple logistic regression model showed that only hematologic malignancy as the underlying disease (odds ratio, 6.6; 95% confidence interval, 1.6-26.9; P=0.009) was independently associated with resistance. In 242 cases in which data regarding management and prognosis were available, the 30-day survival rate was 68.4%. In the univariate analysis of factors predicting survival, a significant association was found for Candida species, age of the patient, neutropenia, recent abdominal surgery, removal of the central venous catheter, and use of appropriate antifungal therapy. In the multivariate analysis, removal of the central venous catheter (odds ratio, 6.0; 95% confidence interval, 2.2-16.1; P<0.001) and the use of appropriate therapy (odds ratio, 2.1; 95% confidence interval, 1.1-4.1; P=0.03) were independent factors significantly associated with survival after the diagnosis of Candida bloodstream infection.

Diagnostic testing in Epstein-Barr virus infection.

Laboratory diagnosis of Epstein-Barr virus (EBV) infection is improving with the development of new technologies. Quantification of the virus by real-time polymerase chain reaction (PCR) and evaluation of EBV-specific T cells, especially by tetrameric human leukocyte antigens, are noteworthy candidates for monitoring procedures in clinical laboratories involved in the management of transplant recipients. Standardization of PCR is essential for improving the quality of these monitoring procedures.

Adhesion via CD43 induces Syk activation and cell proliferation in TF-1 cells.

The effect of adhesion via CD43 (leukosialin, sialophorin) on cell proliferation and phosphorylation signaling were examined in a growth factor-dependent hematopoietic progenitor cell line, TF-1. TF-1 cells promptly resulted in death after withdrawal of growth factors. However, the viable cell number increased when TF-1 cells were cultured on anti-CD43 monoclonal antibody-coated plates. In this case, sustained activation of protein tyrosine kinase Syk and extracellular signal-regulated kinase (Erk) 1/2 were detected. Overexpression of exogenous Syk on TF-1 cells by the adenovirus vector system induced enhancement of the cell proliferation accompanied with enhancement of the Erk activation by a dominant-positive effect. The signal transducer and activator of transcription (STAT) 5 seemed not to be associated with the CD43-mediated cell proliferation. These results indicated that adhesion via CD43 induces the proliferation of TF-1 cells in the absence of growth factors in part by Syk-dependent Erk 1/2 signaling.

[Characterization of cryoglobulin, M protein, low molecular weight IgM in a patient with chronic hepatitis C and type II mixed cryoglobulinemia].

The common extrahepatic manifestation of hepatitis C virus(HCV) infection is mixed cryoglobulinemia. We have analyzed serum cryoglobulin, IgM and various antibody activities from a patient with chronic hepatitis C and type II cryoglobulinemia. Cryoprecipitates were consisted of polyclonal IgG and monoclonal IgM-kappa with rheumatoid factor activity. About 20% of total IgM was found to be low molecular weight IgM by gel-filtration and SDS-PAGE. The anti-streptolysin O(ASO) activity measured by Latex agglutination method was found to be markedly elevated despite normal activity by Rantz-Randell method. The patient's serum has revealed to react against bovine gamma globulin, which crosslinked streptolysin O to Latex particles, in a nonspecific manner. Phenotypic analysis of the surface markers on abnormal lymphoid cells from peripheral blood and bone marrow showed monoclonal expansion of B-cell lineage by flow cytometry. The patient was treated with interferon-alpha, which resulted in an improvement of liver dysfunction, decreased amounts of cryoglobulin and IgM. It was concluded that the patient has suffered from lymphoproliferative disorder, namely HCV infection-associated primary macroglobulinemia.

Flow cytometric method for enumeration and classification of reactive immature granulocyte populations.

We developed a flow cytometric method for the enumeration and classification of nonmalignant immature granulocytes (IG). In this study, IG are defined as most immature (IG stage 1: promyelocytes and myelocytes) and as more mature (IG stage 2: metamyelocytes). Blood specimens from 46 patients with documented infectious or inflammatory disease and known presence of IG (by routine manual microscopy) were analyzed. For a reference manual differential count, we used a 400 white blood cell (WBC) differential and separated granulocytes into promyelocytes and myelocytes combined, metamyelocytes, and included band cells in the mature, segmented neutrophil population. The flow cytometric method is based on three-color staining of whole, anticoagulated blood with CD45-PerCP, CD16-FITC, and CD11b-PE-labeled monoclonal antibodies and a three-step gating procedure. The flow cytometric results were confirmed by cell sorting and microscopic evaluation of the sorted cells. A total of 10,000 events, excluding debris, were recorded per specimen and IG stage 1 (CD16-/CD11b-), IG stage 2 (CD16-/CD11b+), and mature neutrophils (CD16+/CD11b+) were categorized. Regression and correlation between flow cytometric IG and the manual differential showed y = 1.34x + 0.95, r(2) = 0.86 for IG stages 1 and 2 combined versus promyelocytes, myelocytes, and metamyelocytes. For IG stage 1 versus microscopic counts of promyelocytes and myelocytes, the results were y = 1.53x + 1.24, r(2) = 0.76; for IG stage 2 versus manual metamyelocyte count, y = 0.77x + 0.21, r(2) = 0.58. Reproducibility of the flow cytometric method showed a coefficient of variation (CV) of 6.8% for all IG combined compared with a CV of 50.2% for manual differential IG count (based on a routine 100 WBC count). Samples were found stable at least 12 h at 25 degrees C and at least 48 h at 4 degrees C for flow cytometry. After staining and lysing, the sample was stable for at least 120 min at room temperature. We analyzed samples from patients with myelodysplastic and myeloproliferative disease separately. We found that CD16- mature neutrophils falsely elevated the flow cytometric IG count. Similar results were obtained in blood from patients treated with granulocyte-colony stimulating factor (G-CSF). Although this restricts the use of the method somewhat, we believe that this flow cytometric method is useful for enumerating reactive IG, as well as for evaluating automated methods for IG identification by hematology analyzers.

IL-15 is elevated in the patients of postoperative enterocolitis.

Serum interleukin 15 (IL-15) levels were measured in 77 patients who were consecutively admitted to our intensive care unit. Postoperative enterocolitis occurred in four patients and Methicillin-resistant Staphylococcus aureus (MRSA), but not Clostridium difficile, was identified in the faecal specimens from these patients. The IL-15 levels in the patients with MRSA enterocolitis were significantly elevated compared with those of other MRSA infections without enterocolitis including pneumonia (n=6) and cholangitis (n=1), and other MRSA non-colonized patients (n=66) (21.2+/-5.2 pg/ml vs 4.3+/-0.2, 4.3+/-0.5). Notably, an increase in serum IL-15 was observed just before clinical manifestation of severe diarrhoea. Our findings suggest that IL-15 may be associated in the pathogenesis of postoperative enterocolitis and its serum level may be a severity indicator of the disease.

[Evaluation of COBAS CORE anti-HIV-1/HIV-2 EIA DAGS for the detection of antibodies to HIV by COBAS CORE].

COBAS CORE Anti-HIV-1/HIV-2 EIA DAGS (CORE HIV-1/2) for the detection of antibodies to HIV-1 and HIV-2 was evaluated by a fully automated immunoassay system, COBAS CORE II (Roche Diagnostics K.K.). The sensitivity of CORE HIV-1/2 was assessed in comparison with PA1/2, Enzygnost, VIDAS and AxSYM by testing serial two-fold dilutions of HIV-1 and HIV-2 sera. CORE HIV-1/2 showed the highest sensitivity than other methods. The agreements of the results of anti-HIV antibodies determinations between the CORE HIV-1/2 and other methods, PA1/2, Enzygnost, VIDAS, and AxSYM, were 99.4%, 100%, 100%, and 99.7%, respectively in the tests for anti-HIV-1 antibody. In the tests for anti-HIV-2, all results were agreed at 100%. Discrepant samples which showed a negative by CORE HIV-1/2 and a positive by PA1/2 and AxSYM were clearly negative of anti-HIV-1 antibody by p24 ACA, PCR, Western blot, and continuous clinical studies. This assay showed excellent coefficients of variation for both within-run and day to day precision. The determination of anti-HIV antibodies was not interfered with hemoglobin, bilirubin, and chylemia. We conclude that the excellent specificity and sensitivity of the CORE HIV-1/2 make the CORE HIV-1/2 a suitable method for diagnosis of HIV-1 and HIV-2 infections in clinical laboratories.

Enhancement of tumoricidal activity of alveolar macrophages via CD40-CD40 ligand interaction.

CD40-CD40 ligand (CD40L) interaction was originally defined as important molecules for the development of humoral immunity. Thereafter, some investigations have focused on its essential roles for the induction of cell-mediated immunity in host defenses. Here we investigated the antitumor activity of murine alveolar macrophages through CD40-CD40L interaction. The CD40L gene was transfected into murine lung cancer cells (3LLSA), and CD40L-expressing clones (3LLSA-CD40L) were established. Stimulation of CD40 molecules on the surface of alveolar macrophages with 3LLSA-CD40L cells induced the production of nitric oxide, tumor necrosis factor-alpha, and interleukin-12 and the tumoricidal activity of alveolar macrophages in the presence of interferon-gamma, which increased the surface expression of CD40 molecules on alveolar macrophages. These findings were not observed when alveolar macrophages were obtained from CD40-deficient mice. On the other hand, interleukin-6 production by alveolar macrophages did not depend on CD40-CD40L interaction. We also established a murine melanoma cell line expressing CD40L (B16 4A5-CD40L) that could induce tumoricidal activity of alveolar macrophages. Furthermore, when spleen cells were cocultivated with 3LLSA-CD40L cells, specific cytotoxic T lymphocytes for wild-type 3LLSA cells could be induced. These results suggest that CD40L gene transfer into tumor cells may induce antitumor immunity in a tumor-bearing host and may offer a new strategy for cancer gene therapy.

Constrictive bronchiolitis obliterans and paraneoplastic pemphigus.

Constrictive bronchiolitis obliterans is rare, and the pathogenesis of the disease often remains unknown. This study reports on the case of a 38 yr-old female with constrictive bronchiolitis obliterans and paraneoplastic pemphigus associated with malignant lymphoma. The patient developed progressive obstructive lung disease. The chest radiograph showed almost normal lungs. Paraneoplastic pemphigus is a newly described syndrome in which patients have autoantibodies binding to some epithelia, including in the respiratory tract. The disease develops in association with non-Hodgkin's lymphomas or other malignant neoplasms. The case presented here suggests that constrictive bronchiolitis obliterans associated with paraneoplastic pemphigus may be one of the facets of autoimmune responses in this context.

Detection and sequence diversity of Kaposi's sarcoma-associated herpesvirus (KSHV)/human herpesvirus 8 (HHV-8) DNA.

Detection of Kaposi's sarcoma (KS)-associated herpesvirus (KSHV)/human herpesvirus (HHV-8) has been reported frequently in patients with KS associated with acquired immunodeficiency syndrome (AIDS). We examined the presence of the KSHV sequence in 8 HIV-positive patients comprising 5 with KS, 2 with syphilis, 1 with prurigo, and 2 HIV-negative patients with angiosarcoma. Using the polymerase chain reaction, we observed amplification of a DNA fragment of the expected size in 4 patients with KS. Sequencing analysis of the amplified fragments revealed several base substitutions upon comparison with the originally reported sequence. Our results support the hypothesis of a pathogenic role of KSHV in the development of skin lesions in HIV-positive patients with KS, and the sequences of KSHV DNA fragments isolated in this study also demonstrated strain diversity similar to that reported previously.

Immunization with the "immunogenic peptide" of TSH receptor induces oligoclonal antibodies with various biological activities.

Balb/c mice were immunized with a synthetic peptide (P354-14) corresponding to "the immunogenic peptide" of human thyrotropin receptor (hTSH-R). Through screening for binding to the peptide, we obtained several monoclonal antibodies with various biological activities: thyroid stimulation (SAb), inhibition of TSH stimulation (BAb) and no significant effect on cAMP production. One of the stimulatory clones was further studied. This clone enhanced cAMP production in Cos-7 cells transformed with the truncated TSH-R cDNA deleting the immunogenic peptide. These results indicated that the immunogenic peptide of the TSH-R induces oligoclonal anti-TSH-R antibodies, although the region is not essential for the functional epitope.