Antiserum Against Neurite Outgrowth Factor in Chick Gizzard Extract and Its Inhibitory Effect on Neuritic Response in Cultured Ciliary Neurons

Abstract: Antiserum against a neurite outgrowth factor (NOF) of gizzard extract that promotes neurite outgrowth from dissociated ciliary ganglionic neurons (CG neurons) of 8-day-old chick embryo was prepared to determine whether or not the antiserum inhibits neurite outgrowth from cultured neurons or explants of chick and murine tissues. When CG neurons were cultured on a polyornithine-coated well exposed to NOF (NOF-bound POR well), marked neurite outgrowth was observed. When NOF-bound POR wells were exposed to antiserum, neurite outgrowth from CG neurons was gradually inhibited with increasing amounts of antiserum, while exposure to preimmune serum did not prevent neurite outgrowth. Antiserum had no effect on neuronal survival during a 48-h incubation. The diluted antiserum, which produced nearly 100% inhibition of the NOF activity, was almost equally active in suppressing the activity of NOFs in conditioned media (CM) of various chick embryo tissues, but showed much less inhibitory effects on NOFs in CM of murine tissues. The appearance of neurites from explants of spinal cord, dorsal root ganglion, or retina of chick embryo was also inhibited by the antiserum. These results indicate that antiserum against NOF from gizzard extract suppressed the activity of NOFs from various sources, and that there are species differences in NOFs, at least between chick and murine.     

Measurement of Pseudomonas aeruginosa multidrug efflux pumps by quantitative real-time polymerase chain reaction

Multidrug efflux pumps contribute to multiple antibiotic resistance in Pseudomonas aeruginosa. Pump expression usually has been quantified by Western blotting. Quantitative real-time polymerase chain reaction has been developed to measure mRNA expression for genes of interest. Whether this method correlates with pump protein quantities is unclear. We devised a real-time PCR for mRNA expression of MexAB-OprM and MexXY-OprM multidrug efflux pumps. In laboratory strains differing in MexB and MexY expression and in several clinical isolates, protein and mRNA expression correlated well. Quantitative real-time PCR should be a useful alternative in quantitating expression of multidrug efflux pumps by P. aeruginosa isolates in clinical laboratories.     

Cooperation between alteration of DNA gyrase genes and over-expression of MexB and MexX confers high-level fluoroquinolone resistance in Pseudomonas aeruginosa strains isolated from a patient who received a liver transplant followed by treatment with fluoroquinolones

Clinical isolates of highly fluoroquinolone-resistant Pseudomonas aeruginosa had a mutation in either A or B subunit of DNA gyrase and over-expressed MexB and MexX, the efflux system proteins. Introduction of wild-type gyrase genes of Escherichia coli into the isolates made them as fluoroquinolonesusceptible as the moderately fluoroquinolone-resistant strains that only over-expressed efflux system proteins. These findings demonstrate that high fluoroquinolone-resistance in P. aeruginosa is attributed to cooperation between alteration in DNA gyrase genes and over-expression of efflux systems proteins.     

Immunocytochemical detection of enhanced expression of c-myc protein in the heart of cardiomyopathic hamster

An immunocytochemical study was performed to examine the expression of cellular c-myc protein in the heart of 30-, 120- and 180-day-old cardiomyopathic Syrian UM-X7.1 hamsters. The heart of age- and sex-matched BIO-RB hamster was used as normal control. In paraffin sections, an immunostaining for c-myc was markedly increased in cytoplasm of cells from the UM-X7.1 heart as compared with that of the BIO-RB heart which showed a weak staining. However, c-myc was localized in nuclei of cells in frozen sections of the heart. Specific cell types of the heart were differentiated with anti-vimentin, and we found that the increased expression of c-myc was present in nuclei of muscle cells of the UM-X7.1 myocardium. A quantitative study of c-myc-positive nuclei of muscle and nonmuscle cells was carried out by a video micrometer. The mean number of c-myc-positive nuclei of muscle cells was significantly higher in the cardiomyopathic heart than in the control heart from hamsters of all ages studied. These results suggest that the increase of c-myc protein may relate to the pathological state or pathogenesis of the hereditary cardiomyopathy.     

Transplantation of elastin-secreting myoblast sheets improves cardiac function in infarcted rat heart

Myoblast sheet transplantation for cardiac failure is a promising therapy to enhance cardiac function via paracrine mechanism. However, their efficacies of treatment showed a gradual decline. The gene modification of the implanted myoblast is important in improving the long-term results of the treatment. Elastin fiber enhances the extensibility of the infarcted wall and can prevent left ventricular dilation. We therefore hypothesized that the elastin gene modification of the implanted myoblast could strengthen and maintain the long-term improvement effects of cardiac function. In this study, we evaluated long-term follow-up benefits of functional myoblast sheets that secrete elastin in an infarcted model. The animal models were divided into three groups: a group transplanted with nontransfected, wild-type, skeletal myoblast-type sheets (WT-rSkM); group transplanted with myoblast sheets that secreted elastin fragments (ELN-rSkM); and a control group (ligation only). Cardiac function was examined by echocardiography, and cardiac remodeling after infarction was evaluated by histological examination. The cardiac function was significantly improved and the left ventricle end-diastolic dimensions were significantly reduced in the ELN-rSkM group. Histological analysis showed that left ventricular remodeling was attenuated in the ELN-rSkM group and that elastic fiber was formed in the epicardial area of ELN-rSkM group. The functionalization of myoblast sheet by elastin gene transfer showed the long-term improvement of cardiac function. Expressed recombinant elastin fiber prevented the dilation of the left ventricular chamber after myocardial infarction. The functional myoblast sheet transplantation maintained the treatment effect by the paracrine effect of myoblast and the formed recombinant elastin.     

Gicerin,a cell adhesion molecule,participates in the histogenesis of retina

Gicerin is a novel cell adhesion molecule that belongs to the immunoglobulin superfamily. Gicerin protein adheres to neurite outgrowth factor (NOF), an extracellular matrix protein in the laminin family, and also exhibits homophilic adhesion. Heterophilic adhesion of gicerin to NOF is thought to play an active role in neurite outgrowth of developing retinal cells in vitro. In this study, we examined the adhesion activity of gicerin during the retinal development of Japanese quail using an antibody directed against gicerin, to elucidate the biological importance of gicerin in retinal histogenesis. Immunohistochemical and Western blot analysis showed that gicerin was highly expressed in the developing retina but suppressed in the mature retina. The aggregation of neural retinal cells from 5-day embryonic quail retina was significantly inhibited when incubated with a polyclonal antibody to gicerin, suggesting that gicerin protein participates in the adhesion of neural retinal cells of the developing retina. Furthermore, histogenesis of retina both in the organ cultures and in ovo embryos was severely disrupted by incubation with a gicerin antibody. These findings provide evidence that gicerin plays an important role in retinal histogenesis. © 1997 John Wiley & Sons, Inc. J Neurobiol 33: 769–780, 1997     

Ethylene-induced putrescine accumulation modulates K+ partitioning between roots and shoots in barley seedlings

We investigated the cause and effect relationships among ethylene, polyamines, and K⁺ in barley ( Hordeum vulgare L. cv. Amagi) seedlings. Application of 1-aminocyclopropane-1-carboxylic acid (ACC), a precursor of ethylene, to the growth medium caused a decrease in K⁺ concentration in roots and an increase in shoots. Addition of ACC induced putrescine accumulation in roots, while spermidine and spermine levels remained unchanged. Exogenous supply of putrescine led to putrescine accumulation and reduced K⁺ concentration. Application of Co²⁺, an inhibitor of ethylene biosynthesis, together with ACC, inhibited putrescine accumulation with a decrease in K⁺ concentration in roots. ACC-treated roots showed K⁺ uptake capacity equivalent to that of control roots, implying that the majority of K⁺ is translocated to shoots. These results suggest that ethylene regulates K⁺ partitioning between roots and shoots through the level of accumulation of putrescine in barley seedlings.     

Isolation and characteristics of a short rod-shaped mutant of Pseudomonas aeruginosa

Abstract Among 40 short rod-shaped mutants of Pseudomonas aeruginosa PAO isolated after nitrosoguanidine mutagenesis, a stable clone designated KG1034 was obtained and studied for its cellular and genetical features. Cells of the parent strain, PAO2302 had a mean cell length of 1.9 μm, whereas that of KG1034 was 1.3 μm. The doubling time of KG1034 was less than that of the parent strain, although both strains elongated at the same rate and exhibited the same relative amounts and pattern of penicillin-binding proteins. These results suggested that the phenotype of KG1034 was due to the initiation of septation at an earlier stage. The new gene responsible for this phenotype was named srs (abbreviation for short r od shape) and mapped by conjugation between cys -54 and puuC .