Molecular cloning and expression analysis of a macrophage-colony stimulating factor receptor-like gene from rainbow trout, Oncorhynchus mykiss

The M-CSF and its receptor (M-CSFR, CSF-1R or c-fms proto-oncogene) system were initially implicated as essential in mammals for normal monocyte development as well as for pregnancy. To allow a comparison with the M-CSF and M-CSFR system of an oviparous animal, we cloned a M-CSFR-like gene from rainbow trout (Oncorhynchus mykiss). The gene was cloned from a cDNA library of head kidney. It contained an open reading frame encoding 967 amino acids with a predicted size of 109 kDa. The putative amino acid sequence of rainbow trout M-CSFR showed 54% amino acid identity to fugu (Takifugu rubripes) M-CSFR, 52% to zebrafish (Danio rerio) M-CSFR and 40% to mouse (Mus musculus) and human (Homo sapiens) M-CSFR. The M-CSFR-like gene was constitutively expressed in head kidney, kidney, intestine, spleen and blood. The gene was detected especially in the ovary of immature female rainbow trout. These results suggest that a M-CSFR-like receptor may be involved in female reproductive tracts even in an oviparous animal like fish.     

Molecular and immunological approach to hematological disease: detection and analysis of intracellular modified nucleosides by flow cytometry

Modified nucleosides are components of ribosomal RNA (rRNA), transfer RNA (tRNA) and messenger RNA (mRNA). 1-methyladenosine and pseudouridine are members of those modified nucleosides. The urinary concentration of 1-methyladenosine and pseudouridine of cancer patients are higher than that of healthy controls, and those compounds were reduced after effective chemotherapy. Thus those compounds might be expected to use as tumor markers. In this study cellular origin of 1-methyladenosine and pseudouridine were analysed about two tumor cell lines (HUT-102, THP-1), peripheral blood lymphocytes (PBL) from healthy adult and PBL under the phytohemagglutinin stimulation, by flow cytometric analysis and immunofluorescent staining of cellular RNA using monoclonal antibodies specific for 1-methyladenosine (AMA) and pseudouridine (APU). Both 1-methyladenosine and pseudouridine were detected in more than 90% of tumor cells above the thresholds of flow cytometric detection (Spectrum III, Ortho). The PBL under the PHA stimulation also tended to take the same way of the tumor cell lines, whereas few of the PBL contained 1-methyladenosine above the thresholds. According to the DNA analysis of those cell lines, high contents of the modified nucleosides in the cell might follow DNA synthesis, this leads to one reason for high levels of the urinary excretion of the modified nucleosides in cancer patient.     

Detection of the putative E2 protein of hepatitis C virus in human liver

The question was asked whether a predicted envelope protein, considered to be processed from the polyprotein precursor encoded by the putative E2/NS1 region of the hepatitis C virus (HCV) genome, may be observed in HCV-infected humans. Two polyclonal antibodies against recombinant E2/NS1 proteins were prepared and their reactivity tested against liver extracts from HCV-infected patients by immunoblotting analysis. A band corresponding to a size of 44 kDa was detected in liver extracts from patients who were positive for the HCV-specific antibody anti-C100-3 but not in liver extracts from patients who did not have anti-C100-3 antibody. Additionally, no band was detected using preimmune sera or antisera which had been preabsorbed with recombinant E2/NS1 proteins. Deglycosylation studies demonstrated that the 44 kDa protein was a glycosylated form of a 38 kDa protein which corresponds to the predicted molecular weight of the putative E2/NS1 protein. These results suggest that the 44 kDa protein is a product of the E2/NS1 region. Frequent observation of the 44 kDa band in cases of chronic active hepatitis C suggests a correlation between the expression of this protein and the progression of hepatitis. © 1994 Wiley-Liss, Inc.     

Influence of the parameters of several pulse sequences on the image contrast: comparison between the two major contrast media

We compared the influences of the parameters of several pulse sequences using two major commercially available gadolinium (Gd) contrast media for MR imaging. The phantom of Gd solutions of various concentration (0.1 - 10mmol/L) was prepared, and was scanned with a 1.5T clinical MR unit, using a spin-echo T1-weighted sequence, 2DFLASH, 3DFLASH, and 3DVIBE. The signal intensity was measured and the contrast enhancement ratio (CER) was calculated and plotted as a function of Gd concentration. The results were compared between the pulse sequences, and between the contrast media as well. Both 3DFLASH and 3DVIBE showed higher CER than other two sequences, showing similar CER curve configuration. There was no significant difference both in CER value and CER curve configuration between the two contrast media for each pulse sequence.     

Development of enzyme-labeled oligonucleotide probe for detection of mecA gene in methicillin-resistant Staphylococcus aureus.

A DNA hybridization method with an enzyme-labeled oligonucleotide probe (mecA-ELONP) was developed to detect the methicillin-resistant gene (mecA) in methicillin-resistant Staphylococcus aureus. For rapid identification, bacterial colonies were transferred from agar plates directly onto nylon membranes. Lysis of cells, denaturation of DNA, and hybridization were performed on the membranes. These procedures required only 3 h for completion. The results obtained by this test closely corresponded with those obtained by determining the MICs of oxacillin against S. aureus. The results of the mecA-ELONP also correlated well with those of a commercially available PCR test. Thus, mecA-ELONP proved to be a reliable and convenient method for the rapid identification of methicillin-resistant S. aureus, which could be useful in clinical microbiology laboratories.     

Purification and characterization of a hemolysin produced by Vibrio cholerae biotype El Tor: another toxic substance produced by cholera vibrios.

A thermolabile direct hemolysin from an El Tor cholera vibrio strain has been isolated and partially characterized as a simple protein of ca. 20,000 molecular weight. In addition to its hemolytic activity, the hemolysin is cytotoxic, cardiotoxic, and rapidly lethal. In these respects it resembles the thermostable direct hemolysin/cytotoxin/cardiotoxin/lethal toxin of Vibrio parahaemolyticus and certain other bacterial hemolysins, although there are other significant differences. Because identical diseases are produced by both hemolytic and nonhemolytic cholera vibrios, the El Tor hemolysin may be presumed to be pathogenetically irrelevant. These observations raise the question of "When is a toxic substance also a toxin?"     

Effect of Azelastine Hydrochloride on Macrophage Chemotaxis and Phagocytosis in Vitro

Azelastine, a newly synthesized anti-allergic agent, was tested for its effects on guinea pig macrophage chemotaxis and phagocytosis. As specific macrophage chemo-attractants, we used macrophage chemotactic factors a and c; separated and highly purified from inflamed skin sites. Macrophage chemotaxis induced by skin extract or chemotactic factors was significantly suppressed by a low concentration of the agent (1 microgram/ml); the effect was dose-dependent. The inhibition of chemotaxis was reversible, because chemotactic activity was restored when the agents was removed by washing cells before chemotactic assay. Inactivation of chemotactic factors was not detected by mixing azelastine and factors a and c. Azelastine may directly interact with macrophages to decrease their chemotactic responsiveness. beta-Glucuronidase activity in the medium and macrophages after phagocytosis of polystyrene latex particles was not affected by this agent at concentrations ranging from 1 to 10 micrograms/ml. The phagocytosis of latex particles or sheep red blood cells opsonized with IgG antibodies (EA) and anchoring of macrophages to substrate were not inhibited and azelastine did not damage the macrophages as determined by lactate dehydrogenase (LDH) release assay.     

Evaluation of "M-point" in hepatic artery to identify left medial segment of liver. Angiographic study

The hepatic arteries of 122 patients were analysed on angiography to identify the left medial segment of the liver. Left medial arterial branches were classified into three types: type I arising from the left hepatic artery on the umbilical portion of the portal vein; type II arising from proximal portion of left hepatic artery before reaching the umbilical portion of the portal vein; type III arising from right hepatic artery. Incidence of each type is 37.2%, 35.8% and 27.0%, respectively. The artery frequently kinks at the right side of the umbilical portion of the portal vein and a total of incidence is 68% and that of each type is 23.5%, 89.8% and 100%, respectively. We call this characteristic kinking point of the left medial arterial branches, the "M-point".