Rhodnius prolixus, a blood-sucking bug, is a unique insect that is known to produce nitric oxide (NO) in the salivary glands to use as a vasodilator for blood sucking. We report here the cloning of the NO synthase (NOS) cDNA from these salivary glands and its expression in a baculovirus system. This cDNA encodes a protein of 1174 amino acids with a calculated molecular mass of 132331 Da. The primary structures of mammalian NOS, including the putative cofactor-recognition sites for heme, tetrahydrobiopterin (BH,), calmodulin, FMN, FAD and NADPH are all conserved in salivary-gland NOS. Recombinant salivary-gland NOS differed from nerve NOS and endothelial NOS in that it lacked a large N-terminal domain and an N-terminal myristylation sequence, respectively. Salivary-gland NOS produced in a baculovirus system showed NOS activity and demonstrated that salivary-gland NOS was soluble and was Ca2+ and calmodulin dependent, similarly to mammalian constitutive NOS isoforms. Recombinant salivary-gland NOS was purified to near homogeneity and migrated at 130 kDa on SDS/PAGE.Keywords : insect; salivary gland ; nitric-oxide synthase; CaZ+ and calmodulin dependence; baculovirus.NO is produced in various mammalian tissues where it plays physiologically important roles [I]. NO acts as neurotransmitter in the peripheral and the central nervous systems and is involved in brain development, memory formation and learning. In the circulatory system, NO is produced in endothelial cells, and acts as a regulator of vascular tone and platelet aggregation. NO is induced in macrophages, neutrophils and liver cells, and acts as an important effector of the cell-mediated immune system. The enzyme critical for NO production, NO synthase (NOS), is found in these different tissues, and various isofoms have been purified and characterized, and some have been cloned [2, 31. Mammalian NOS have been grouped into two general categories: constitutive (Ca*' kalmodulin dependent, e.g. endothelial NOS and neuronal NOS) and inducible (Ca"/calmodulin independent, e.g. macrophage) types.By contrast, in invertebrates, the production and utilization of NO is not as well understood, although similar physiological functions are probably present. In some insects and molluscs, the existence of NOS in brain has been demonstrated, and the function of NO as a neurotransmitter was suggested [4, 51. In the silkworm, Bombyx mori, NOS activity in the fat body was induced by lipopolysaccharide, an agent that mimics bacterial assault, which suggested the involvement of NO in the insect immune defense system [ 3 ] . This activity required Ca2+, calmodulin, NADPH, FAD, FMN and tetrahydrobiopterin (BH,), similarly to mammalian NOS. Correspondence to M . Yuda, Department of Medical Zoology,Fux: f 8 l 592 31 5215. Abhreviufions. NO, nitric oxide; NOS, nitric-oxide synthase; RACE, rapid amplification of cDNA ends ; BH,, tetrahydrobiopterin ; MOI, multiplicity of infection.Note. The nucleotide sequence data published here have been submitted to the GenBank database and ar...

A novel complex of calphostin (UCN-1028), which specifically inhibits protein kinase C (PKC) has been isolated from the culture broth of a fungi Cladosporium cladosporioides. Purification of individual components was carried out by silica gel, non-porus resin Diaion HP-20SSand Sephadex LH-20 chromatography, leading to isolation of five closely related components, A, B, C, D and I. Calphostinsshowedcytotoxic activities against various tumorcells, andthese cytotoxicities were proportional to their inhibitory activities against PKC. Specific inhibitors of protein kinase C (PKC) can be excellent tools to study the role of PKC in cellular proliferation and differentiation, and are expected to be potential chemotherapeutic agents. In the course of screening new selective inhibitors of PKC, a fungi Cladosporium cladosporioides was found to produce a complex of novel and specific inhibitors of PKC, designated calphostin (originally called UCN-1028). The most hydrophobic component, calphostin A (UCN-1028A) which was the first purified compound, has already been reported1*. Further study of the calphostin fermentation showed the presence of four components in addition to calphostin A. This paper describes the fermentation, isolation, physico-chemical properties and biological activities of the calphostin complex. Materials and Methods Microorganism The producing organism, C. cladosporioides FERMBP-1285 was isolated from a block fence around a house in Osaka, Japan.

RP-1776, a novel cyclic peptide, was isolated from the culture broth of Streptomyces sp. KYI 1784. RP-1776 selectively inhibited the binding of PDGFBB to the extracellular domain of the PDGF /^receptor with an IC50 value of 11±6/im. Detailed binding experiments suggested that RP-1776 directly interacts with PDGF BB. RP-1776 inhibited the phosphorylation of the PDGFj3-receptor induced by PDGFBB. These results suggested that RP-1776 antagonizes the signaling of PDGFBB probably through the inhibition of PDGFBB binding to the PDGF^-receptor. Platelet-derived growth factor (PDGF) is a potent mitogen and chemotactic molecule for various cells, released from activated platelets. h2) PDGFis also expressed in embryonal tissues, and has neurotrophic activity for neuronal cells in rat brain, suggesting involvement of

Cross-sectional surveys of parasitic infection were performed using the agar plate culture technique (APCT) and modified formalin-ethyl acetate concentration technique (MFECT) to assess the true prevalence of Strongyloides stercoralis relative to other parasites in north-east Thailand. The enzyme-linked immunosorbent assay (ELISA) for diagnosis of S. stercoralis infection was used to estimate the seroprevalence for comparison with coproprevalence. Faecal and serum samples were collected from study participants during October-November 2000. Within the sample population of 332 rural northeast Thais from 3 communities, S. stercoralis was the most common parasitic infection (average 28.9%, range 27.7-30.3%) as determined by APCT; by MFECT the average was 5.4% (range 1.8-8.6%). Other intestinal parasites by order of prevalence were Opisthorchis viverrini (average 14.2%, range 8.6-19.4%), hookworm (average 12.3%, range 4-20.2%), Echinostoma sp. (7.5%), Giardia intestinalis (0.9%), Trichuris trichiura (0.6%), and Taenia sp., Hymenolepis nana and Entamoeba coli (all 0.3%). In an analysis of a subset of the sample population for which serum samples were available (n = 120), coproprevalence by APCT was 33.3% (range 27-53.8%) and seroprevalence was 47.5% (range 29.7-57.9%) by modified unit-based ELISA and 34.2% (range 21.6-42.1%) by conventional optical density (OD)-based ELISA. Taking APCT as the reference method for diagnosis of strongyloidiasis, the sensitivity and specificity of the OD-based ELISA were 65% and 81.3%, respectively, and of the unit-based ELISA were 77.5% and 71.3%, respectively. Our results indicate that S. stercoralis is the predominant parasite in rural north-east Thailand, and that APCT and ELISA should be used as complementary diagnostic methods for community-based parasite surveys, at least among those in high-risk groups.

Key Points• ATL patients who relapsed after allogeneic HSCT have a very high mortality rate and present a serious therapeutic challenge.• No large study exists that assesses the role of salvage therapies for relapsed ATL after HSCT; this is the first report summarizing the outcome.Adult T-cell leukemia/lymphoma (ATL) relapse is a serious therapeutic challenge after allogeneic hematopoietic stem cell transplantation (allo-SCT). In the present study, we retrospectively analyzed 35 patients who experienced progression of or relapsed persistent ATL after a first allo-SCT at 3 institutions in Nagasaki prefecture (Japan) between 1997 and 2010. Twenty-nine patients were treated by the withdrawal of immune suppressants as the initial intervention, which resulted in complete remission (CR) in 2 patients. As the second intervention, 9 patients went on to receive a combination of donor lymphocyte infusion and cytoreductive therapy and CR was achieved in 4 patients. Of 6 patients who had already had their immune suppressants discontinued before the relapse, 3 patients with local recurrence received local cytoreductive therapy as the initial treatment, which resulted in CR for more than 19 months. Donor lymphocyte infusion-induced remissions of ATL were durable, with 3 cases of long-term remission of more than 3 years and, interestingly, the emergence or progression of chronic GVHD was observed in all of these cases. For all 35 patients, overall survival after relapse was 19.3% at 3 years. The results of the present study suggest that induction of a graft-versus-ATL effect may be crucial to obtaining durable remission for ATL patients with relapse or progression after allo-SCT. (Blood. 2013;121(1):219-225)

No abstract

No abstract

ObjectivesTo clarify the association between glucose intolerance and high altitudes (2900–4800 m) in a hypoxic environment in Tibetan highlanders and to verify the hypothesis that high altitude dwelling increases vulnerability to diabetes mellitus (DM) accelerated by lifestyle change or ageing.DesignCross-sectional epidemiological study on Tibetan highlanders.ParticipantsWe enrolled 1258 participants aged 40–87 years. The rural population comprised farmers in Domkhar (altitude 2900–3800 m) and nomads in Haiyan (3000–3100 m), Ryuho (4400 m) and Changthang (4300–4800 m). Urban area participants were from Leh (3300 m) and Jiegu (3700 m).Main outcome measureParticipants were classified into six glucose tolerance-based groups: DM, intermediate hyperglycaemia (IHG), normoglycaemia (NG), fasting DM, fasting IHG and fasting NG. Prevalence of glucose intolerance was compared in farmers, nomads and urban dwellers. Effects of dwelling at high altitude or hypoxia on glucose intolerance were analysed with the confounding factors of age, sex, obesity, lipids, haemoglobin, hypertension and lifestyle, using multiple logistic regression.ResultsThe prevalence of DM (fasting DM)/IHG (fasting IHG) was 8.9% (6.5%)/25.1% (12.7%), respectively, in all participants. This prevalence was higher in urban dwellers (9.5% (7.1%)/28.5% (11.7%)) and in farmers (8.5% (6.1%)/28.5% (18.3%)) compared with nomads (8.2% (5.7%)/15.7% (9.7%)) (p=0.0140/0.0001). Dwelling at high altitude was significantly associated with fasting IHG+fasting DM/fasting DM (ORs for >4500 and 3500–4499 m were 3.59/4.36 and 2.07/1.76 vs <3500 m, respectively). After adjusting for lifestyle change, hypoxaemia and polycythaemia were closely associated with glucose intolerance.ConclusionsSocioeconomic factors, hypoxaemia and the effects of altitudes >3500 m play a major role in the high prevalence of glucose intolerance in highlanders. Tibetan highlanders may be vulnerable to glucose intolerance, with polycythaemia as a sign of poor hypoxic adaptation, accelerated by lifestyle change and ageing.