Prognostic factors of Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis in children: report of the Japan Histiocytosis Study Group.

BACKGROUND: Despite several advances in the treatment of Epstein-Barr virus (EBV) in recent years, patients with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) do not always show satisfactory outcomes. We here conducted a nationwide survey in Japan to identify prognostic factors of EBV-HLH in children with this disease in an effort to improve the management and the outcomes of these patients. PROCEDURE: Between January 2003 and June 2008, we enrolled 98 children younger than 18 years of age who were diagnosed with EBV-HLH. We then studied the clinical characteristics and laboratory findings at the time of diagnosis with the aim to identify prognostic factors for EBV-HLH. RESULTS: The mean age of onset of EBV-HLH was 3.9 ± 2.8 years. Most of our patients presented with fever, hepatosplenomegaly, lymphadenopathy, and hemophagocytosis of bone marrow. Sixty-two percent of patients showed T cell clonality, and 97% had EBV infection in either T or natural killer cells. Most patients (60%) were treated with a multi-agent chemotherapeutic regimen, including corticosteroid, etoposide, and cyclosporine. After initial treatment, 90.3% of patients were in remission, and 7 patients (8.2%) experienced recurrence of EBV infection. Among several prognostic factors, patients with both hyperbilirubinemia (>1.8 mg/dl) and hyperferritinemia (>20,300 ng/ml) at the time of diagnosis had significantly poorer outcomes than those with low serum bilirubin and ferritin levels. CONCLUSIONS: These findings suggest that the therapeutic strategy for children with EBV-HLH could be tailored according to the laboratory findings at diagnosis.

Current research on chronic active Epstein-Barr virus infection in Japan.

Epstein-Barr virus (EBV) infection is usually asymptomatic and persists lifelong. Although EBV-infected B cells have the potential for unlimited proliferation, they are effectively removed by the virus-specific cytotoxic T cells, and EBV-associated lymphoproliferative disease develops only in immunocompromised hosts. Rarely, however, individuals without apparent immunodeficiency develop chronic EBV infection with persistent infectious mononucleosis-like symptoms. These patients have high EBV-DNA load in the peripheral blood and systemic clonal expansion of EBV-infected T cells or natural killer (NK) cells. Their prognosis is poor with life-threatening complications including hemophagocytic lymphohistiocytosis, organ failure, and malignant lymphomas. The term "chronic active EBV infection" (CAEBV) is now generally used for this disease. The geographical distribution of CAEBV is markedly uneven and most cases have been reported from Japan and other East Asian countries. Here we summarize the current understanding of CAEBV and describe the recent progress of CAEBV research in Japan.

Molecular diagnosis of mitochondrial respiratory chain disorders in Japan: focusing on mitochondrial DNA depletion syndrome.

BACKGROUND: Although mitochondrial respiratory chain disorders (MRCD) are one of the most common congenital metabolic diseases, there is no cumulative data on enzymatic diagnosis and clinical manifestation for MRCD in Japan and Asia. METHODS: We evaluated 675 Japanese patients having profound lactic acidemia, or patients having symptoms or signs of multiple-organ origin simultaneously without lactic acidemia on respiratory chain enzyme activity assay and blue native polyacrylamide gel electrophoresis. Quantitative polymerase chain reaction was used to diagnose mitochondrial DNA depletion syndrome (MTDPS). Mutation analysis of several genes responsible for MTDPS was also performed. RESULTS: A total of 232 patients were diagnosed with a probable or definite MRCD. MRCD are common, afflicting one in every several thousand people in Japan. More than one in 10 of the patients diagnosed lacked lactic acidemia. A subsequent analysis of the causative genes of MTDPS identified novel mutations in six of the patients. A 335 bp deletion in deoxyguanosine kinase (DGUOK; g.11692_12026del335 (p.A48fsX90)) was noted in two unrelated families, and may therefore be a common mutation in Japanese people. The proportion of all patients with MTDPS, and particularly those with recessive DNA polymerase γ (POLG) mutations, appears to be lower in Japan than in other studies. This is most likely due to the relatively high prevalence of ancient European POLG mutations in Caucasian populations. No other significant differences were identified in a comparison of the enzymatic diagnoses, disease classifications or prognoses in Japanese and Caucasian patients with MRCD. CONCLUSION: MTDPS and other MRCD are common, but serious, diseases that occur across all races.

Clinical characteristics and outcomes of chédiak-Higashi syndrome: a nationwide survey of Japan.

BACKGROUND: Chédiak-Higashi syndrome (CHS) is a rare autosomal recessive disorder characterized by immunodeficiency, neurological dysfunction, and oculocutaneous albinism. Recently, several clinical CHS phenotypes have been reported. Here, we report results of a nationwide survey performed to clarify clinical characteristics and outcomes of CHS patients in Japan. METHODS: Questionnaires were sent to 287 institutions to collect data regarding CHS patients diagnosed between 2000 and 2010, including results of lysosomal trafficking regulator (LYST) gene analysis. Cytotoxicity and degranulation activity of cytotoxic T lymphocytes were analyzed in available patient samples. RESULTS: A total of 15 patients diagnosed with CHS were eligible for enrollment in this study. Of these, 10 (67%) had recurrent bacterial infections, five (33%) developed life-threatening hemophagocytic lymphohistiocytosis (HLH), and one patient had complicated malignant lymphoma. Hematopoietic stem cell transplantation (HSCT) was performed for six patients including three with HLH, and 10 of the enrolled patients have survived at the time of this writing. LYST analysis was performed for 10 patients; seven different mutations were detected in seven patients, whereas no mutation was identified in three patients. Cytotoxicity and degranulation activity were impaired in patients with and without LYST mutation. DISCUSSION: Results of this survey indicate that one or two patients with CHS were newly diagnosed each year in Japan. The incidence of HLH was not as high as expected. Mutations of genes other than LYST were suspected in some cases. We conclude that determining indication for HSCT for CHS patients should be based on genetic and cytotoxic analysis.

Characterization of Epstein-Barr virus (EBV) BZLF1 gene promoter variants and comparison of cellular gene expression profiles in Japanese patients with infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis.

Epstein-Barr virus (EBV) genotypes can be distinguished based on gene sequence differences in EBV nuclear antigens 2, 3A, 3B, and 3C, and the BZLF1 promoter zone (Zp). EBV subtypes and BZLF1 Zp variants were examined in Japanese patients with infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis. The results of EBV typing showed that samples of infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis all belonged to EBV type 1. However, sequencing analysis of BZLF1 Zp found three polymorphic Zp variants in the same samples. The Zp-P prototype and the Zp-V3 variant were both detected in infectious mononucleosis and chronic active EBV infection. Furthermore, a novel variant previously identified in Chinese children with infectious mononucleosis, Zp-V1, was also found in 3 of 18 samples of infectious mononucleosis, where it coexisted with the Zp-P prototype. This is the first evidence that the EBV variant distribution in Japanese patients resembles that found in other Asian patients. The expression levels of 29 chronic active EBV infection-associated cellular genes were also compared in the three EBV-related disorders, using quantitative real-time reverse transcription polymerase chain reaction analysis. Two upregulated genes, RIPK2 and CDH9, were identified as common specific markers for chronic active EBV infection in both in vitro and in vivo studies. RIPK2 activates apoptosis and autophagy, and could be responsible for the pathogenesis of chronic active EBV infection.

Hemophagocytic lymphohistiocytosis after hematopoietic stem cell transplantation in children: a nationwide survey in Japan.

BACKGROUND: Hemophagocytic lymphohistiocytosis (HLH) is associated with hypercytokinemia in children. Although HLH can be also observed after hematopoietic stem cell transplantation (HSCT), the incidence and clinical features of HLH after HSCT remain obscure. PROCEDURE: The clinical features of HLH after HSCT (post-HSCT HLH) were investigated in children with malignancies, immune deficiencies, or aplastic anemia. The HLH/Langerhans Cell Histiocytosis (LCH) Committee of the Japanese Society of Pediatric Hematology (JSPH) sent questionnaires to hospitals with JPSH members asking for details of cases in which HLH occurred after HSCT between 1998 and 2008. RESULTS: Among 42 children who were diagnosed with post-HSCT HLH between 1998 and 2008 in Japan, 37 fulfilled our inclusion criteria; of these, 26 were classified as early-onset (onset <30 days after HSCT) and 11 were classified as late-onset (onset >30 days after HSCT). In the early-onset group, the presence of respiratory symptoms, high levels of total bilirubin, and triglycerides at onset and the lack of control of GVHD with tacrolimus were significantly associated with non-resolution of HLH (P < 0.05). The survival rate was significantly higher in patients with resolution of HLH than in those without resolution (59% vs. 14%, P < 0.05). CONCLUSIONS: These findings suggest that early-onset post-HSCT HLH is a specific entity of HLH, and appropriate diagnosis and prompt management need to be established.

A novel anti-peroxiredoxin autoantibody in patients with Kawasaki disease.

Antibodies to the anti-oxidative peroxiredoxin (Prx) enzymes occur in both systemic autoimmune disease and vasculitis in adulthood. Because increased oxidative stress induces vasculitis in Kawasaki disease (KD), autoimmunity to Prxs in patients with KD was investigated. The presence of antibodies to Prx 1, 2 and 4 was analyzed by ELISA and Western blot. Of 30 patients with KD, 13 (43.3%) possessed antibodies to Prx 2, whereas these antibodies were present in only 1 of 10 patients (10.0%) with sepsis (4 with purulent meningitis and 6 with septicemia). In contrast, antibodies to Prx 1 and 4 were not detected in either group. There was no significant correlation among the titers of the three antibodies. Clinical parameters were compared between anti-Prx 2-positive and -negative patients. The presence of anti-Prx 2 antibodies correlated with a longer period of fever and poor response to high-dose γ-globulin therapy in patients with KD. Anti-Prx 2-positive patients had significantly greater excretion of urinary 8-isoprostaglandin than did anti-Prx 2-negative patients. These results provide the first evidence for an antibody to Prx 2 in patients with KD. They also suggest that this antibody might serve as a marker of disease severity and be involved in the pathophysiology of vasculitis in some patients with KD.

Effect of single-dose oral mizoribine pulse therapy twice per week for frequently relapsing steroid-dependent nephrotic syndrome.

AIM: To evaluate the efficacy of single-dose oral mizoribine (MZB) pulse therapy given twice weekly for frequently relapsing steroid-dependent nephrotic syndrome (FR-SDNS). METHODS: The subjects were 8 patients with FR-SDNS with a median age of 6.9 years old (range 3.1 - 18.0 y). The study was performed as a Phase II trial. The MZB dose was adjusted to achieve a peak blood level of 3 - 5 µg/ml (3.9 - 15.9 mg/kg/d, maximum dose: 750 mg) using a single dose given twice weekly before a meal. The therapeutic benefits of MZB pulse therapy were assessed based on a comparison of the incidence of relapse and the required daily dosage of prednisolone (PSL) in the 12 months prior to and following therapy. RESULTS: The incidence of relapse after therapy was significantly lower than that before therapy (2.5 ± 1.4 vs. 4.3 ± 0.5, p < 0.01) and the required daily dosage of prednisolone (PSL) after therapy was lower than that before therapy (0.48 ± 0.23 vs. 0.52 ± 0.32 mg/kg/d, not significant). However, this therapy was not effective for 3 out of 4 patients treated with cyclosporine. During follow-up, discontinuation of PSL was possible in 4 of 5 patients who showed a decreased rate of relapse after therapy. The peak blood concentration of MZB in these patients was significantly higher than that in 3 patients who did not show a decreased rate of relapse (3.95 ± 0.11 vs. 3.05 ± 0.21 µg/ml, p < 0.01). No adverse effects were observed in any patients. CONCLUSION: Our results show that single-dose oral MZB pulse therapy is effective in decreasing the frequency of relapse in some pediatric patients with FR-SDNS. A peak concentration of MZB of ~3.8 - 4.0 µg/ ml may be required for FR-SDNS therapy.

Conditional VHL gene deletion activates a local NO-VEGF axis in a balanced manner reinforcing resistance to endothelium-targeted glomerulonephropathy.

BACKGROUND/AIMS: We have reported that tubular epithelial cell injury caused by renal ischemia-reperfusion is attenuated in conditional VHL knockout (VHL-KO) mice and also that induction of hypoxia-inducible factor (HIF) suppresses angiotensin II-accelerated Habu snake venom (HV) glomerulonephropathy in rats. However, it remains unknown whether VHL knockdown protects glomerular endothelial cells from endothelium-targeted glomerulonephritis. METHODS AND RESULTS: VHL-KO mice with HV glomerulonephropathy (HV GN) had fewer injured glomeruli, a lower mesangiolysis score and reduced blood urea nitrogen levels. Immunoreactivity of vascular endothelial growth factor (VEGF) in the glomerular capillaries was enhanced by VHL knockdown and was conserved even in VHL-KO mice with HV GN, despite HV-attenuating endothelial VEGF expression in vitro. VHL-KO mice showed enhanced nitric oxide (NO) production in glomerular endothelial cells and tubular cells, associated with activated VEGF expression in the kidney (i.e. an activated NO-VEGF axis). The levels of NO in glomeruli and tubules were conserved even in mice with HV GN. In contrast, suppressing NO production in glomerular endothelial cells by an NO synthase inhibitor, N(ϖ)-nitro-L-arginase, completely blunted the protection of VHL-KO from HV GN. The activated NO-VEGF axis in the kidney of VHL-KO mice was also associated with an elevation in Flk-1 phosphorylation and increased levels of IL-10 and IP-10. CONCLUSION: Conditional VHL knockdown may enhance the NO-VEGF axis and protect glomerular endothelial cells from HV GN, thereby providing resistance to injury of tubular epithelial cells and glomerular endothelial cells.

Asymptomatic high Epstein-Barr viral load carriage in pediatric renal transplant recipients.

High viral load carriage of EBV is one of the risks for PTLD in transplant recipients. We reviewed retrospectively in pediatric renal transplant recipients with EBV seronegative. EBV loads in peripheral blood and EBV-CTLs were measured every 1-3 months in 13 patients after grafting. Immunosuppressants were reduced when the patients were considered to have persistent high EBV loads (>1000 copies/µgDNA for over six months). All showed primary EBV infection: six with asymptomatic persistent high EBV loads (group A) and seven with neither EBV-associated symptoms nor persistent high EBV loads (group B). No patient developed PTLD in either group. Chronic rejection occurred in one patient in group A after immunosuppressants' reduction. There was no difference in renal dysfunction rates between the two groups. The maximum and increase rates in EBV loads were significantly higher in group A. The CTLs' percentage was significantly lower in group A when EBV loads first rose above 100 copies/µg DNA. This study suggests the possibility that EBV loads and CTLs' monitoring may be useful for avoidance of PTLD, as patients with asymptomatic persistent high EBV loads had higher EBV loads and lower percentages of CTLs.

Neonatal lupus erythematosus in identical twins, showing transient bullous lesions.

One of identical twin girls was born with ulcers on her leg, and shortly after birth developed a flaccid blister on the leg. Subepidermal blister with vacuolar degeneration of basal cell layer and the heavy infiltration of mononuclear cells in the upper dermis were observed in the blister lesion. She also had generalized livedo. Her identical twin sister did not exhibit ulcers or blisters, but was born with milia on her limbs. Their mother was found to have lupus erythematosus with positive anti-Ro/SSA antibodies and developed Sjögren syndrome. We emphasize neonatal blistering and congenital milia unique manifestations of neonatal lupus erythematosus.

[A case of persistent cerebellar ataxia complicated by conversion disorder--confirmed by positive cerebrospinal fluid glutamate receptor delta2 and epsilon2 antibodies].

We recently encountered a 13-year-old girl who developed persistent cerebellar symptoms one month after mixed measles/rubella vaccination, making it difficult to distinguish this condition from conversion disorders. Severe truncal ataxia was the initial manifestation in this case. The patient had no abnormalities in objective tests but began to show extraordinary circadian variations in certain parameters. Her cerebellar symptoms were thus considered to possibly be associated with conversion disorders. Later, she tested positive for cerebrospinal fluid anti-glutamic acid receptor (GluR) delta2 antibody. The lymphocyte stimulation test yielded a positive reaction to GluRdelta2 antigen. In addition, in the chronic stage SPECT revealed reduced cerebellar blood flow. She was thus diagnosed as having persistent cerebellar ataxia due to autoimmune mechanisms and modification of cerebellar symptoms due to secondary conversion disorders. Our experience with this case suggests that checking cerebrospinal fluid for anti-GluRdelta2 antibody is possibly useful for distinguishing between conversion disorders and cerebellar ataxia due to autoimmune mechanisms.

Isolation and characterization of a novel Staphylococcus aureus bacteriophage, phiMR25, and its therapeutic potential.

A novel bacteriophage, phiMR25, was isolated from a lysogenic Staphylococcus aureus strain by mitomycin C induction. Its biological features were analyzed in comparison with phiMR11, which was described previously as a prototype therapeutic phage. phiMR25 is morphologically similar to phiMR11 (morphotype B1 of family Myoviridae) but has a broader host range than phiMR11 on S. aureus strains. phiMR25 can also multiply on S. aureus lysogens of phiMR11. Its DNA is 44,342 bp in size, is predicted to include 70 open reading frames, and does not contain genes related to toxin or drug resistance. The lysogenic module and most of the putative virion protein genes are completely different from those of phiMR11. In spite of their genetic diversity, intraperitoneal administration of phiMR25 rescued mice inoculated with a lethal dose of S. aureus, as was the case for phiMR11. These results suggest that phiMR25 could be another candidate phage to treat S. aureus infection.

Diagnosis of infectious mononucleosis caused by Epstein-Barr virus in infants.

BACKGROUND: The diagnosis of infectious mononucleosis (IM) is usually on serologic tests. The responses of anti-Epstein-Barr virus (anti-EBV) antibodies are weak in infants. The authors encountered some IM infants in whom anti-EBV antibodies were undetectable during early stage, although EBV genome was found in their blood. The aim of the present study was therefore to clarify the frequency of anti-EBV-antibody negative IM cases. METHODS: The EBV serostatus of 104 IM children diagnosed on Sumaya criteria was retrospectively studied. The EBV genome in peripheral blood mononuclear cells was measured. RESULTS: The anti-viral capsid antigen-IgM (anti-VCA-IgM)-positive rate in the acute phase was only 25% in infants but 80% in patients ≥ 4 years of age. Twenty percent of the infants were negative for all anti-EBV antibodies and required repeated serologic tests. For infants, the significant rise in anti-VCA-IgG was the most sensitive marker. Three seronegative infants with IM symptoms, with circulating EBV genome during acute phase, were eventually considered as having IM on anti-VCA-IgG seroconversion thereafter. CONCLUSIONS: To diagnose IM in infants the serologic test alone in the acute phase is not sensitive enough. It is proposed that the EBV genome be evaluated in peripheral blood mononuclear cells when infants presenting with IM symptoms are negative for anti-EBV antibodies during the acute phase.

Characteristics of a novel Pseudomonas aeruginosa bacteriophage, PAJU2, which is genetically related to bacteriophage D3.

Pseudomonas aeruginosa bacteriophage (phage) is one of the most taxonomically and genetically diverse phages. Although phage D3 is one of well-studied P. aeruginosa phages, no D3-related P. aeruginosa phage has been reported. We report a novel P. aeruginosa siphovirus, PAJU2, which is genetically related to but morphology distinct (highly elongated head) from phage D3. A PAJU2 capsid protein, Orf3, is thought to be synthesized as a protein fused to a prohead protease and is autocatalytically cleaved, which may form the head chain mail. Despite such morphological differences, PAJU2 is expected to be a useful genetic reference for phage D3.

Lactobacillus rhamnosus GG and Lactobacillus casei suppress Escherichia coli-induced chemokine expression in intestinal epithelial cells.

BACKGROUND: Recently, some strains of lactic acid bacteria (LAB) have been reported to prevent the development of atopic dermatitis and to improve allergic symptoms, especially in young children. However, the mechanisms involved in these effects are not fully understood. Intestinal microbiota play critical roles in the development of host immune development and are recognized and regulated by the host through intestinal epithelial cells (IECs). We thus hypothesized that LAB influence the host immune system through the activation of IECs. To begin testing this hypothesis, chemokine expression in IECs exposed to intestinal bacteria was investigated. METHODS: Caco-2 cell monolayers were stimulated with different concentrations of various live or heat-killed intestinal bacteria or bacterial components for up to 3 h. Changes in the gene expressions of various chemokines were measured using quantitative real-time PCR. RESULTS: The expressions of CCL20, CXCL8, CXCL10 and CX3CL1 were strongly induced by nonpathogenic Escherichia coli in a dose-dependent manner and were partially induced by some commensal LAB. In contrast, Lactobacillus rhamnosus GG (LGG) and Lactobacillus casei did not induce these chemokine expressions. In addition, LGG significantly suppressed the expressions of CCL20 and CXCL10 induced by E. coli, peptidoglycan or flagellin when cultured simultaneously. CONCLUSIONS: LGG and L. casei markedly suppressed E. coli-induced chemokine expression, presumably through the suppression of the Toll-like receptor-mediated signal transduction pathway, at least in part. The clinical importance of this suppressive effect and the mechanisms involved require further investigation; however, such effects can be used as a marker to identify clinically useful LAB.

In silico and in vivo evaluation of bacteriophage phiEF24C, a candidate for treatment of Enterococcus faecalis infections.

Along with the increasing threat of nosocomial infections by vancomycin-resistant Enterococcus faecalis, bacteriophage (phage) therapy has been expected as an alternative therapy against infectious disease. Although genome information and proof of applicability are prerequisites for a modern therapeutic phage, E. faecalis phage has not been analyzed in terms of these aspects. Previously, we reported a novel virulent phage, phiEF24C, and its biology indicated its therapeutic potential against E. faecalis infection. In this study, the phiEF24C genome was analyzed and the in vivo therapeutic applicability of phiEF24C was also briefly assessed. Its complete genome (142,072 bp) was predicted to have 221 open reading frames (ORFs) and five tRNA genes. In our functional analysis of the ORFs by use of a public database, no proteins undesirable in phage therapy, such as pathogenic and integration-related proteins, were predicted. The noncompetitive directions of replication and transcription and the host-adapted translation of the phage were deduced bioinformatically. Its genomic features indicated that phiEF24C is a member of the SPO1-like phage genus and especially that it has a close relationship to the Listeria phage P100, which is authorized for prophylactic use. Thus, these bioinformatics analyses rationalized the therapeutic eligibility of phiEF24C. Moreover, the in vivo therapeutic potential of phiEF24C, which was effective at a low concentration and was not affected by host sensitivity to the phage, was proven by use of sepsis BALB/c mouse models. Furthermore, no change in mouse lethality was observed under either single or repeated phage exposures. Although further study is required, phiEF24C can be a promising therapeutic phage against E. faecalis infections.

Induction of apoptosis in human basophils by anti-Fas antibody treatment in vitro.

BACKGROUND: Basophils are thought to play an important role in the pathogenesis of allergic inflammation; however, the factors associated with basophil death are not fully understood. Fas (CD95) is a member of the TNF receptor superfamily and is known to induce apoptosis in activated T cells, neutrophils and eosinophils. In the present study, the expression and function of Fas in human basophils were investigated in vitro. METHODS: Human cultured basophils were obtained by culturing cord blood-derived CD34+ cells in the presence of 2.5 ng/ml of IL-3 for 5-6 weeks. The expression of Fas was measured using flow cytometry. Cell viability and morphological changes after the incubation of basophils with anti-Fas mAb (clone CH11, IgM) in the presence of 1 ng/ml of IL-3 were measured using the trypan blue dye exclusion test and light microscopy, respectively. RESULTS: Human cultured basophils constitutively and significantly expressed Fas on their cell surfaces. Treatment with anti-Fas monoclonal antibody (mAb) significantly reduced basophil viability in a time- and dose-dependent manner. When basophils were incubated with 10 ng/ml of anti-Fas mAb or control for 72 h, the basophil viability was 27.3 +/- 8.8% and 89.3 +/- 5.2%, respectively (p < 0.01). Anti-Fas mAb-treated basophils were shrunken and exhibited condensed nuclei, consistent with apoptosis. CONCLUSIONS: Our findings indicate that human basophils express functional Fas on their cell surfaces, and signaling via Fas may regulate basophil survival in vivo.

Isolation and characterization of a novel Enterococcus faecalis bacteriophage phiEF24C as a therapeutic candidate.

Vancomycin-resistant Enterococcus faecalis (VRE) has become a significant threat in nosocomial settings. Bacteriophage (phage) therapy is frequently proposed as a potential alternative therapy for infections caused by this bacterium. To search for candidate therapeutic phages against Enterococcus faecalis infections, 30 Enterococcus faecalis phages were isolated from the environment. One of these, virulent phage phiEF24C, which has a broad host range, was selected for analysis. The plaque-forming ability of phiEF24C was virtually unaffected by differences in the clinical host strains. Furthermore, the phage had a shorter latent period and a larger burst size than ordinary tailed phages, indicating that phiEF24C has effective lytic activity against many Enterococcus faecalis strains, including VRE. Morphological and genomic analyses revealed that phiEF24C is a large myovirus (classified as family Myoviridae morphotype A1) with a linear double-stranded DNA genome of c. 143 kbp. Analyses of the N-terminal amino acid sequences of the virion proteins, together with the morphology and the genome size, speculated that phiEF24C is closely related to other myoviruses of Gram-positive bacteria that have been used experimentally or practically for therapy or prophylaxis. Considering these results, phiEF24C may be a potential candidate therapeutic phage against Enterococcus faecalis infections.

Centrally administered neuromedin U elevates plasma adrenaline by brain prostanoid TP receptor-mediated mechanisms in rats.

Neuromedin U is a hypothalamic peptide involved in energy homeostasis and stress responses. The peptide, when administered intracerebroventricularly (i.c.v.), decreases food intake and body weight while increasing body temperature and heat production. We examined the effect of i.c.v. administered neuromedin U on plasma catecholamines with regard to the brain prostanoid using anesthetized rats. Neuromedin U (0.1, 0.5 and 1 nmol/animal, i.c.v.) effectively elevated plasma adrenaline (a maximal response was obtained at 0.5 nmol/animal), but had little effect on plasma noradrenaline. However, intravenously administered neuromedin U (0.5 nmol/animal) had no effect on plasma catecholamines. Neuromedin U (0.5 nmol/animal, i.c.v.)-induced elevation of plasma adrenaline was effectively reduced by intracerebroventricular pretreatments with indomethacin (an inhibitor of cyclooxygenase) (0.6 and 1.2 micromol/animal), furegrelate (an inhibitor of thromboxane A2 synthase) (0.9 and 1.8 micromol/animal) and (+)-S-145 (a blocker of prostanoid TP receptors) (250 and 625 nmol/animal), respectively. The neuromedin U-induced adrenaline response was also abolished by acute bilateral adrenalectomy. These results suggest that centrally administered neuromedin U evokes the secretion of adrenaline from the adrenal medulla by brain prostanoid TP receptor-mediated mechanisms in rats.