Studies on the resistance of a congenic resistant strain of mice (DKIR) against infection with Salmonella enteritidis (author's transl)]

A congenic strain of mice (DKIR) having a relatively resistant gene for mouse typhoid was established by the beckcross mating between C3H/He and DKI strains. DKI strain is highly and uniformly susceptible to the infection with Salmonella enteritidis and C3H/He is relatively resistant to that infection. The present paper reported the consistency of resistance to Salmonella infection of DKIR strain throughout generations after 10 backcross matings. Difference of the number of infected organisms in the peritoneal fluid and organs between DKIR and DKI or C3H/He strains was also described. The newly established DKIR strain seems to be a suitable experimental animal for the study of genetical resistance for mouse typhoid when compared with its original DKI strain.     

Effect of expectorants on relaxation behavior of sputum viscoelasticity in vivo

We studied the effects of expectorants (mucolytic agents) in vivo on the relaxation behavior of sputum viscoelasticity. Seven female and thirty-three male patients (56.8 +/- 19.3 yrs, range: 21 to 82 years old) with a chronic pulmonary disease except bronchial asthma were studied. They were randomly put into the control group or a group which would be given oral treatments with an expectorant for a week after a one week washout period. The groups were as follows: Group I (n = 8), control; Group II (n = 7), Bromhexine hydrochloride 24 mg per day; Group III (n = 10), Ambroxol 90 mg per day; Group IV (n = 9) alpha - Chymotrypsin buccle 100 ch.u. per day; Group V (n = 6), Serratiopeptidase 30 mg per day. In Groups IV & V, frequency dependence of sputum viscoelasticity at the range of omega = 10(-3) to 10(0) rad.sec-1 were clearly changed after the treatments, and the magnitude of the relaxation and its main relaxation time were significantly increased. On the other hand, in Groups I, II & III, no significant changes of the frequency dependences were observed. These findings suggest that proteolytic enzymes administered orally work on the molecular structure of sputum, and break down their linkages between subunits of the structure.     

Spodiobacter cordis gen. nov. sp. nov., a member of the family Flavobacteriaceae isolated from patients with infective endocarditis

Two gram‐negative, catalase‐negative, oxidase‐positive strains (PAGU 1467^T and PAGU 1468) isolated from patients with infective endocarditis were investigated to determine their taxonomic status. 16S rRNA gene sequence analysis indicated that the two strains were members of the Bergeyella‐Chryseobacterium‐Riemerella branch of the family Flavobacteriaceae. Strains PAGU 1467^T and PAGU 1468 were highly related to each other (98.8% 16S rRNA gene sequence similarity). Phylogenetically closely‐related species to PAGU 1467^T comprised Bergeyella zoohelcum (95.0% 16S rRNA gene sequence similarity), Riemerella anatipestifer (94.3%) and Cloacibacterium normanense (94.3%). The major fatty acids of the two isolates were iso‐C_15:0, iso‐C_17:0 3‐OH and iso‐C_15:0 3‐OH. The presence of C_16:0 3‐OH and iso‐C_15:0 2‐OH allowed these isolates to be distinguished from B. zoohelcum. Menaquinone MK‐6 was the only respiratory quinone in these organisms; this is a consistent characteristic of the family Flavobacteriaceae. The guanine‐plus‐cytosine content of the genomic DNA was 42.0%, which is higher than that of other close phylogenetic relatives. On the basis of their phenotypic properties and genetic distinctiveness, isolates PAGU 1467^T and PAGU 1468 were classified within the novel genus Spodiobacter, as Spodiobacter cordis gen. nov., sp. nov., which is also the type species. The type strain of S. cordis is PAGU 1467^T ( = CCUG 65564^T = NBRC 109998^T).     

Production and characterization of monoclonal antibodies to 17 alpha-hydroxyprogesterone

Hybridoma clones producing antibodies to 17 alpha-hydroxyprogesterone (17-OHP) were established by using a 17-OHP-bovine serum albumin conjugate as an immunogen. Six representative IgG-class monoclonal antibodies of high affinity (10(8)-10(9) M-1) showed differential reactivities with several structurally related steroids. Two enzyme immunoassay (EIA) systems (fluorescence EIA and micro-EIA) for 17-OHP using OHP 4B2.2.3, which showed the lowest cross-reactivity with other steroids, were established. The micro-EIA system was shown to be applicable to the mass-screening of congenital adrenal hyperplasia.     

Identification of the Functional Domains of the Telomere Protein Rap1 in Schizosaccharomyces pombe

The telomere at the end of a linear chromosome plays crucial roles in genome stability. In the fission yeast Schizosaccharomyces pombe, the Rap1 protein, one of the central players at the telomeres, associates with multiple proteins to regulate various telomere functions, such as the maintenance of telomere DNA length, telomere end protection, maintenance of telomere heterochromatin, and telomere clustering in meiosis. The molecular bases of the interactions between Rap1 and its partners, however, remain largely unknown. Here, we describe the identification of the interaction domains of Rap1 with its partners. The Bqt1/Bqt2 complex, which is required for normal meiotic progression, Poz1, which is required for telomere length control, and Taz1, which is required for the recruitment of Rap1 to telomeres, bind to distinct domains in the C-terminal half of Rap1. Intriguingly, analyses of a series of deletion mutants for rap1 ⁺ have revealed that the long N-terminal region (1–456 a.a. [amino acids]) of Rap1 (full length: 693 a.a.) is not required for telomere DNA length control, telomere end protection, and telomere gene silencing, whereas the C-terminal region (457–693 a.a.) containing Poz1- and Taz1-binding domains plays important roles in those functions. Furthermore, the Bqt1/Bqt2- and Taz1-binding domains are essential for normal spore formation after meiosis. Our results suggest that the C-terminal half of Rap1 is critical for the primary telomere functions, whereas the N-terminal region containing the BRCT (BRCA1 C-terminus) and Myb domains, which are evolutionally conserved among the Rap1 family proteins, does not play a major role at the telomeres.     

Human Immunodeficiency Virus Type 1 Enhancer-binding Protein 3 Is Essential for the Expression of Asparagine-linked Glycosylation 2 in the Regulation of Osteoblast and Chondrocyte Differentiation

Human immunodeficiency virus type 1 enhancer-binding protein 3 (Hivep3) suppresses osteoblast differentiation by inducing proteasomal degradation of the osteogenesis master regulator Runx2. In this study, we tested the possibility of cooperation of Hivep1, Hivep2, and Hivep3 in osteoblast and/or chondrocyte differentiation. Microarray analyses with ST-2 bone stroma cells demonstrated that expression of any known osteochondrogenesis-related genes was not commonly affected by the three Hivep siRNAs. Only Hivep3 siRNA promoted osteoblast differentiation in ST-2 cells, whereas all three siRNAs cooperatively suppressed differentiation in ATDC5 chondrocytes. We further used microarray analysis to identify genes commonly down-regulated in both MC3T3-E1 osteoblasts and ST-2 cells upon knockdown of Hivep3 and identified asparagine-linked glycosylation 2 (Alg2), which encodes a mannosyltransferase residing on the endoplasmic reticulum. The Hivep3 siRNA-mediated promotion of osteoblast differentiation was negated by forced Alg2 expression. Alg2 suppressed osteoblast differentiation and bone formation in cultured calvarial bone. Alg2 was immunoprecipitated with Runx2, whereas the combined transfection of Runx2 and Alg2 interfered with Runx2 nuclear localization, which resulted in suppression of Runx2 activity. Chondrocyte differentiation was promoted by Hivep3 overexpression, in concert with increased expression of Creb3l2, whose gene product is the endoplasmic reticulum stress transducer crucial for chondrogenesis. Alg2 silencing suppressed Creb3l2 expression and chondrogenesis of ATDC5 cells, whereas infection of Alg2-expressing virus promoted chondrocyte maturation in cultured cartilage rudiments. Thus, Alg2, as a downstream mediator of Hivep3, suppresses osteogenesis, whereas it promotes chondrogenesis. To our knowledge, this study is the first to link a mannosyltransferase gene to osteochondrogenesis.     

Induction of osteoblastic cell differentiation by forskolin. Stimulation of cyclic AMP production and alkaline phosphatase activity

The effects of forskolin on differentiation of osteoblastic cells (clone MC3T3-E1) cultured in alpha-minimum essential medium containing 0.1% bovine serum albumin were investigated by assays of intracellular cyclic AMP level and alkaline phosphatase activity in the cells. Forskolin increased cyclic AMP production in the cells in a dose-related manner, the maximum increase being 250-fold above that of the controls. Alkaline phosphatase activity in the cells was also elevated as early as 24 h and rose to nearly its maximum at 48 h. The elevation was dose-dependent, with a maximum increase at 5 X 10(-6) M forskolin. Forskolin and prostaglandin E2 showed a supraadditive effect on cyclic AMP production in the cells and had an additive effect on alkaline phosphatase activity, whereas forskolin and dibutyryl cyclic AMP had little additive effect on either cyclic AMP production or enzyme activity. These results suggest that cyclic AMP is closely linked to the differentiation of osteoblastic cells in vivo.