Leumorphin is a novel endogenous opioid peptide in man

Porcine leumorphin, a putative opioid peptide corresponding to amino acid residues 228-256 of preproenkephalin B has been demonstrated to exist in the porcine neurointermediate pituitary. A recent study on the sequence analysis of genomic DNA of human preproenkephalin B has shown that human leumorphin differs in 3 amino acid residues from porcine leumorphin. In order to clarify whether leumorphin is an endogenous opioid peptide in man, we have studied its existence in the human brain using a radioimmunoassay and its opioid activity by a bioassay with the guinea-pig ileum myenteric plexus-longitudinal muscle preparation. High performance gel permeation chromatography and reverse-phase high performance liquid chromatography coupled with the radioimmunoassay for leumorphin have revealed that human leumorphin exists in water extracts of the human striatum. In the guinea-pig ileum assay, synthetic human leumorphin exhibited potent opioid activity, with the concentration of 3nM to give 50 per cent inhibition. These results indicate that leumorphin is a novel endogenous opioid peptide in man.     

The presence of l-2,4-diaminobutyric acid decarboxylase activity in Vibrio species: A new biosynthetic pathway for 1,3-diaminopropane

Abstract A new enzyme activity, which catalyzes decarboxylation of l -2,4-diaminobutyric acid (DABA) to yield 1,3-diaminopropane (DAP), has been found in dialyzed crude extracts prepared from Vibrio alginolyticus . The pH optimum for the activity was 8.0–8.5, and the enzyme showed a pyridoxal 5'-phosphate (PLP) requirement. Mg²⁺ caused about 30% stimulation in activity. The enzyme was active to only l -DABA among the diamino acids examined, and the K _m value for l -DABA was 0.13 mM. Ammonium sulfate fractionation of a dialyzed crude extract followed by HPLC separation allowed us to conclude that this enzyme differed from the decarboxylase which occurs in Vibrio spp. to produce norspermidine (Nspd) for carboxynorspermidine (C-Nspd) having a moiety similar in structure to DABA. The same enzyme activity was detected in several other Vibrio species.     

Induction and inhibition of cathepsin B and hemoglobin-hydrolase activity in murine B16 melanoma by thiol protease inhibitors

The effects of potent thiol protease inhibitors in vitro (leupeptin, antipain, chymostatin and E-64 (N-[N-(L-3-trans-carboxyoxirane-2-carbonyl)-L-leucyl]agmatine) on intracellular cathepsin B and hemoglobin (Hb)-hydrolase from cultured B16 melanoma cells were studied. E-64 induced cultured B16 melanoma cells to decrease the activities of intracellular cathepsin B (EC 3.4.22.1.) but did not have this effect with Hb-hydrolase or acid phosphatase (EC 3.1.3.2). Leupeptin, antipain and chymostatin induced B16 melanoma cells to increase the activities of intracellular cathepsin B and Hb-hydrolase but not that of acid phosphatase. These results indicate that there are two kinds of thiol protease inhibitors, each with a varying reaction to cultured B16 melanoma--inhibition of intracellular cathepsin B, and conversely, inducement of both cathepsin B and Hb-hydrolase.     

Evidence for cryptic northern refugia in the last glacial period in Cryptomeria japonica

MethodsGenetic diversity and structure were examined using 20 microsatellite markers in 37 populations of C. japonica. The locations of glacial refugia were assessed using STRUCTURE analysis, and potential habitats under current and past climate conditions were predicted using SDM. The process of genetic divergence was also examined using the approximate Bayesian computation procedure (ABC) in DIY ABC to test the divergence time between the gene pools detected by the STRUCTURE analysis.ConclusionsThe combined evidence from microsatellites and SDM clearly indicates that climatic changes have shaped the genetic structure of C. japonica. The gene pool detected in northern Tohoku district is likely to have been established by cryptic northern refugia on the coast of the Japan Sea to the west of the Archipelago. The gene pool in Yakushima Island can probably be explained simply by long-term isolation from the other gene pools since the LGM. These results are supported by those of SDM and the predicted divergence time determined using ABC analysis.     

Expression Level of Quiescin Sulfhydryl Oxidase 1 (Qsox1) in Neuroblastomas

Neuroblastoma is the most common extracranial solid malignant tumor observed during childhood. Although these tumors can sometimes regress spontaneously or respond well to treatment in infants, genetic alterations that influence apoptosis can, in some cases, confer resistance to chemotherapy or result in relapses and adversely affect prognosis for these patients. The aim of this study was to correlate immunohistochemical expression of the protein quiescin sulfhydryl oxidase 1 (QSOX1) in samples obtained from untreated neuroblastomas with the patients’ clinical and pathological prognostic factors and clinical course. Neuroblastoma samples (n=23) obtained from histology blocks were arrayed into tissue microarrays and analysed by immunohistochemistry. The cases were classified according to the following clinical and pathological prognostic factors: age at diagnosis greater or less than/equal to 18 months; location of the lesion at diagnosis (abdominal or extra-abdominal); presence or absence of bone-marrow infiltration; tumor differentiation (well or poorly differentiated); Shimada histopathologic classification (favourable or unfavourable); state of the tumor extracellular matrix (Schwannian-stroma rich or poor); amplification of the MYCN oncogene; and clinical course (dead or alive with or without relapses/residual lesions). Twelve of the cases were female, 9 children were over 18 months old, 9 cases presented with extra-abdominal tumors and 9 cases exhibited tumors with unfavourable histologies. Fifteen patients underwent bone-marrow biopsy, and 4 of these were positive for metastasis. Nine patients died. The higher immunohistochemical expression of QSOX1 was more common in well-differentiated samples (P=0.029), in stroma-rich samples (P=0.029) and in samples from patients with a high prevalence of relapses/residual disease. The functions of QSOX1 include extracellular matrix maturation and the induction of apoptosis. Therefore, QSOX1 may be involved in neuroblastoma differentiation and regression and may thus function as a biomarker for identifying risk groups for this neoplasm.Key words: neuroblastoma, QSOX1, proliferation, apoptosis, prognostic factors     

IGF2 modulates the microenvironment for osteoclastogenesis

We previously reported that hypoxic stress enhanced osteoclast differentiation via increasing insulin-like growth factor 2 (IGF2) production. However, the mechanisms underlying IGF2 stimulation remains unknown. In this study, we investigated the molecular mechanisms of osteoclastogenesis by IGF2 treatment. Primary mouse bone marrow cells were cultured with IGF2. Total RNAs were applied to a DNA microarray analysis, and quantitative RT-PCR was then used to confirm the microarray data and clarify which cells expressed the relative genes. The most interesting findings were the upregulations of CXC chemokine ligand 7 (CXCL7) expression in stromal cells and stromal cell-derived factor 1 (SDF1) expression in osteoblastic cells with IGF2 treatment. The addition of exogenous SDF1 to CXCL7 increased the number of osteoclasts and promoted the formation of giant osteoclasts. These results suggest that IGF2 modulates the microenvironment around osteoclast precursor cells. SDF1 together with CXCL7 may promote the formation of giant osteoclasts.     

Genome Sequence of the Lager Brewing Yeast, an Interspecies Hybrid

This work presents the genome sequencing of the lager brewing yeast (Saccharomyces pastorianus) Weihenstephan 34/70, a strain widely used in lager beer brewing. The 25 Mb genome comprises two nuclear sub-genomes originating from Saccharomyces cerevisiae and Saccharomyces bayanus and one circular mitochondrial genome originating from S. bayanus. Thirty-six different types of chromosomes were found including eight chromosomes with translocations between the two sub-genomes, whose breakpoints are within the orthologous open reading frames. Several gene loci responsible for typical lager brewing yeast characteristics such as maltotriose uptake and sulfite production have been increased in number by chromosomal rearrangements. Despite an overall high degree of conservation of the synteny with S. cerevisiae and S. bayanus, the syntenies were not well conserved in the sub-telomeric regions that contain lager brewing yeast characteristic and specific genes. Deletion of larger chromosomal regions, a massive unilateral decrease of the ribosomal DNA cluster and bilateral truncations of over 60 genes reflect a post-hybridization evolution process. Truncations and deletions of less efficient maltose and maltotriose uptake genes may indicate the result of adaptation to brewing. The genome sequence of this interspecies hybrid yeast provides a new tool for better understanding of lager brewing yeast behavior in industrial beer production.Key words: Saccharomyces pastorianus, beer, genome, interspecies hybrid, larger yeast     

Serological Studies of Neurologic Helminthic Infections in Rural Areas of Southwest Cameroon: Toxocariasis,Cysticercosis and Paragonimiasis

Background

Both epilepsy and paragonimiasis had been known to be endemic in Southwest Cameroon. A total of 188 people (168 and 20 with and without symptoms confirmed by clinicians, respectively, 84.6% under 20 years old) were selected on a voluntary basis. Among 14 people (8.3%) with history of epilepsy, only one suffered from paragonimiasis. Therefore, we challenged to check antibody responses to highly specific diagnostic recombinant antigens for two other helminthic diseases, cysticercosis and toxocariasis, expected to be involved in neurological diseases. Soil-transmitted helminthic infections were also examined.

Methodology/Principal Findings

Fecal samples were collected exclusively from the 168 people. Eggs of Ascaris lumbricoides, Trichuris trichiura and hookworms were found from 56 (33.3%), 72 (42.8%), and 19 (11.3%) persons, respectively. Serology revealed that 61 (36.3%), 25 (14.9%) and 2 (1.2%) of 168 persons showed specific antibody responses to toxocariasis, paragonimiasis and cysticercosis, respectively. By contrast, 20 people without any symptoms as well as additional 20 people from Japan showed no antibody responses. Among the 14 persons with epilepsy, 5 persons were seropositive to the antigen specific to Toxocara, and one of them was simultaneously positive to the antigens of Paragonimus. The fact that 2 children with no history of epilepsy were serologically confirmed to have cysticercosis strongly suggests that serological survey for cysticercosis in children is expected to be useful for early detection of asymptomatic cysticercosis in endemic areas.

Conclusions/Significance

Among persons surveyed, toxocariasis was more common than paragonimiasis, but cysticercosis was very rare. However, the fact that 2 children were serologically confirmed to have cysticercosis was very important, since it strongly suggests that serology for cysticercosis is useful and feasible for detection of asymptomatic cysticercotic children in endemic areas for the early treatment.