Production of sterigmatocystin by Aspergillus versicolor isolated from roughage

A total of 69 samples of hay and straw collected during the winter period of 1984/85 were surveyed for their contamination by Aspergillus versicolor. The percentage of A. versicolor-positive samples was 14.5%. Nineteen A. versicolor strains mainly isolated from roughage were tested for the production of sterigmatocystin. All of the isolates examined were capable of producing different levels of sterigmatocystin on a cracked corn substrate. The majority of these strains were highly toxigenic; 53% of the isolates produced more than 500 mg/kg of sterigmatocystin. These findings suggest that corn is a very suitable substrate for sterigmatocystin production and that particularly in the surface layers of feed stocks and corn silos such toxigenic strains of A. versicolor can produce considerable growth and possibly sterigmatocystin, too.     

Stereospecific microbial reduction of 4,5-dihydro-4-(4-methoxyphenyl)-6-(trifluoromethyl-1H-1)-benzazepin+ ++-2-o ne.

A key intermediate, (3R-cis)-1,3,4,5-tetrahydro-3-hydroxy-4-(4-methoxyphenyl)-6-(trifluorome thyl)- 2H-1-benzazepin-2-one (compound II or SQ32191), with high optical purity was made by the stereoselective microbial reduction of the parent ketone 1. Several strains of bacterial and yeast cultures were screened for the ability to catalyse the stereoselective reduction of 4,5-dihydro-4-(4-methoxyphenyl)-6-(trifluoromethyl)-1H-1-benzazepin++ +-2,3-dione [compound I or SQ32425]. Microorganisms from the genera Nocardia, Rhodococcus, Alkaligenes, Corynebacterium, Arthrobacter, Hansenula, and Candida reduced compound I to compound II with 60-70% conversion yield. In contrast, microorganisms from the genera Pseudomonas and Acinetobacter reduced compound I stereospecifically to (trans)-1,3,4,5-tetrahydro-3-hydroxy-4-(4-methoxyphenyl)-6-(trifluoromet hyl-2H- 1-benzazepin-2-one (compound III or SQ32408). Among various cultures evaluated, N. salmonicolor SC6310 effectively catalysed the transformation of compound I to compound II with 96% conversion yield at 1.5-2.0 gl-1 concentration. Compound II was isolated and identified by NMR analysis, mass spectrometry, and comparison to an authentic sample. Preparative scale fermentation process and transformation process were developed using cell suspensions of N. salmonicolor SC6310 to catalyse the transformation of compound I to compound II. The isolated compound II had a melting point of 222 degrees C (reference 221-223 degrees C), optical rotation of +130.4 (reference +128 degrees C), and optical purity of greater than 99.9% as analyzed by NMR and chiral HPLC.     

Sequence and structural organization of the human gene encoding ciliary neurotrophic factor.

Ciliary neurotrophic factor (CNTF) is a potent polypeptide hormone whose actions appear to be restricted to the nervous system where it promotes survival, neurotransmitter synthesis and neurite outgrowth in certain neuronal populations. We have cloned the gene encoding human CNTF (hCNTF) and have characterized its structure and organization. The hCNTF gene appears to be a unique-copy gene with a simple genetic organization, since only a single intron interrupts the coding domain. The hCNTF gene is located on chromosome 11, as determined using human-hamster somatic cell hybrids. The CNTF protein is highly conserved in evolution. The amino acid (aa) sequences of rat and rabbit CNTF translated from cDNAs display approx. 85% homology with the deduced aa sequence encoding hCNTF.     

Enrichment of diazotrophic bacteria from rice soil in continuous culture

Summary A chemostat was used as a model system to study competitive interactions of diazotrophic microorganisms. Enrichment experiments were carried out under microaerobic conditions (8.7 mol O₂/l) with malate as the sole carbon source. The starting material was a Korean rice soil including intact root pieces. The enrichment process was governed by the dilution rate. High dilution rates resulted in the enrichment ofAzospirillum lipoferum, whereas low dilution rates led to the predominance of an unidentified organism, named Isolate R. Dilution rates were set in the range from D=0.005 to D=0.1 h^–1. The growth kinetics of both organisms followed Monod's model in the enrichment culture. From the experiments, the maximum specific growth rate ofA. lipoferum and Isolate R were 0.069 h^–1 and 0.025 h^–1, respectively. The corresponding K_s-values were 8.4 and 0.9 (mg. 1^–1). The point of theoretical coexistence of both organisms was calculated to occur at a substrate concentration of s=3.0 (mg.l^–1) with a growth of rate =0.018 h^–1. Hence the preset nutritional niches occupied by at least two organisms.Azospirillum lipoferum seems to represent the copiotroph microflora and Isolate R is of the oligotroph type. In addition to its high substrate affinity Isolate R liberatedca. 75% of the fixed nitrogen into the medium, which indicates its potential role for mutualistic interactions in the rhizosphere.     

Minimal Erythema Dose (MED) Testing

Ultraviolet radiation (UV) therapy is sometimes used as a treatment for various common skin conditions, including psoriasis, acne, and eczema. The dosage of UV light is prescribed according to an individual''s skin sensitivity. Thus, to establish the proper dosage of UV light to administer to a patient, the patient is sometimes screened to determine a minimal erythema dose (MED), which is the amount of UV radiation that will produce minimal erythema (sunburn or redness caused by engorgement of capillaries) of an individual''s skin within a few hours following exposure. This article describes how to conduct minimal erythema dose (MED) testing. There is currently no easy way to determine an appropriate UV dose for clinical or research purposes without conducting formal MED testing, requiring observation hours after testing, or informal trial and error testing with the risks of under- or over-dosing. However, some alternative methods are discussed.     

Analysis of creatinine in mouse and rat serum by ion exchange high performance liquid chromatography for in vivo studies of renal function

An ion exchange high performance liquid chromatography method was developed for determining creatinine levels in both mouse and rat serum samples. Separation of creatinine from other serum components was achieved in 10 min using a 100 x 4.1-mm, 10 microm strong cation exchange column following acetonitrile precipitation of serum proteins. Incorporation of a guard cartridge placed in-line prior to the analytical column was employed to prevent interference from compounds used in renal disease animal trials. Creatinine levels in normal and diseased animals were accurately determined in the 0.01-10 mg/dL range, and average recovery of the method was approximately 85% for both mouse and rat serum. Addition of 0.5-1.0% acetic acid to the acetonitrile used for protein precipitation significantly improved creatinine recovery to above 97% in mouse serum. The method was used for routine preclinical diagnosis of rat and mouse model renal function, and for the evaluation of renal disease treatment efficacy.     

Modulation of gastrin-releasing peptide (GRP) receptors in insulin secreting cells

Gastrin-releasing peptide (GRP) receptors are present in pancreatic islets, though their regulation is unknown except for homologous desensitization. The modulation of binding of GRP to mouse pancreatic islets and INS-1 cells was studied. At 60 min (steady-state), total binding of [(125)I-Tyr(15)] GRP was 1.62 per cent of total radioactivity per 50 islets; non-specific binding (presence of 1 mM unlabelled GRP(1-27)) was 0.05 to 0.61 per cent of total radioactivity. A preincubation with 1000 nM cholecystokinin (CCK(8)) or with 1000 nM glucose-dependent insulinotropic peptide (GIP) augmented the number of GRP binding sites but not their affinity. [(125)I-Tyr(15)]GRP binding to INS-1 cells was saturable (90 min) and specific with respect to compounds that are not chemically related to GRP (e.g. calcitonin gene-regulated peptide-CGRP and atrial natriuretic peptide-ANP). Displacement studies showed one binding site with a K(d) of 0.39 nM and a B(max) of 13.2 fmoles mg(-1) protein. When the cells were pretreated for 24 h with 10 nM GIP or CCK(8), only GIP but not CCK(8) increased the B(max) of the GRP binding site. The affinity (K(d)) was not changed by either compound. This effect of GIP pretreatment was not affected by downregulating PKC by TPA (phorbol ester; long-term pretreatment). These data indicate that: (1) specific binding sites for GRP are present in mouse pancreatic islets and INS-1 cells; (2) the GRP binding is upregulated by GIP in both islets and INS-1 cells and additionally by CCK(8 ), albeit only in islets; and (3) PKC does not seem to be involved in the up-regulation process. Thus a positive interplay between both the incretins GIP and CCK(8) and the neurotransmitter GRP is obvious.     

Differential role of proline-rich tyrosine kinase 2 and focal adhesion kinase in determining glioblastoma migration and proliferation

The propensity of malignant gliomas to invade surrounding brain tissue contributes to poor clinical outcome. Integrin-mediated adhesion to extracellular matrix regulates the migration and proliferation of many cell types, but its role in glioma progression is undefined. We investigated the role of the cytoplasmic tyrosine kinases FAK and Pyk2, potential integrin effectors, in the phenotypic determination of four different human glioblastoma cell lines. While FAK expression was similar between the four cell lines, increased FAK activity correlated with high proliferation and low migratory rates. In contrast, Pyk2 activity was significantly increased in migratory cell lines and depressed in proliferative cell lines. Overexpression of Pyk2 stimulated migration, whereas FAK overexpression inhibited cell migration and stimulated cellular proliferation. These data suggest that FAK and Pyk2 function as important signaling effectors in gliomas and indicate that their differential regulation may be determining factors in the temporal development of proliferative or migrational phenotypes.