Naturally occurring CAR bacillus infection in a laboratory rat colony and epizootiological observations

An epizootic of chronic respiratory disease was found in a rat colony. Lungs of the symptomatic rats showed histopathologically severe peribronchial lymphoid cuffing. Filamentous bacteria were detected on the border of the tracheal and bronchial epithelium by light and electron microscopy. These bacteria did not grown on artificial media but propagated in embryonated chicken eggs. The disease was thus diagnosed as cilia-associated respiratory (CAR) bacillus infection. Epizootiological observations of the natural and experimentally induced cases revealed that the disease was highly contagious, slowly progressive and intractable. Contact infection may play a major role in the transmission of this disease.     

Production of bovine identical twins via transfer of demi-embryos without zonae pellucidae

Embryos were collected nonsurgically from n?turally-cycling or superovulated donors 7 d after estrus. Forty-four morulae, early blastocysts and blastocysts classified as good to excellent were bisected using a fine glass needle to produce forty-four identical demi-embryos. The bisected demi-embryos, without zonae pellucidae, were nonsurgically transferred, either by twin or single transfer. An additional forty-eight embryos were collected from the same donors and transferred as a control. Among the twin transfers, 8 of the 13 recipients became pregnant (61.5%). Seven of them conceived twin fetuses (87%) and one a single fetus. However, only two sets of normal identical twin calves were obtained. Among the single transfers, 72.6% (45/62) of bisected embryos without zonae pellucidae resulted in pregnancy, of which 48.4% (30/62) were identical twins, and 24.2% (15/62) were singletons. Another 27.4% (17/62) of the recipients did not became pregnant. The pregnancy rate for whole embryos with zonae pellucidae was 72.9% (35/48). These data show that there was no significant difference between the pregnancy rates of bisected embryos without zonae pellucidae and whole embryos with zonae pellucidae transferred 7 d after estrus. Bisection of bovine embryos was simplified and even morula stage embryos were transferred without zonae pellucidae.     

Normotensive glucocorticoid-suppressible hyperaldosteronism in adult

A 40 year-old man was admitted to our hospital for detailed examination of hypokalemia (2.7 mEq/l). His blood pressure was normal. Metabolic alkalosis, ACTH dependent hyperaldosteronism (18 ng/dl) and over-response to synthetic ACTH were observed. Plasma renin activity, on the other hand, was within the normal range (1.7 ng/ml/hr). Serum potassium was normalized to 4.1 mEq/l and the responsiveness of the renin-angiotensin-aldosterone system was recovered after the administration of dexamethasone. These results led us to suggest that this case might be normotensive glucocorticoid-suppressible hyperaldosteronism. The etiology which was not associated with hypertension and low plasma renin activity has not been clarified but may be related to the shortness of duration of this disease. Our case was also afflicted with mild hypercortisolemia and excessive excretion of urinary 17-hydroxycorticosteroid and 17-ketosteroid which was suppressed by the administration of dexamethasone (2 mg/day). These findings may be related to hypersensitivity of the fascicular zone of the adrenal gland to ACTH.     

Enhancement of ATPase Activity by a Lipid Peroxide of Arachidonic Acid in Rat Brain Microvessels

The effects of 15-hydroperoxyarachidonic acid (15-HPAA) on Na+, K+- and Mg+-ATPase activities in the blood-brain barrier (BBB) were examined using rat brain microvessels (MV). 15-HPAA markedly stimulated these ATPase activities in MV at low concentrations whereas the synaptosomal Na+, K+-ATPase activity was inhibited in a dose-dependent manner. Further neurochemical analysis revealed that this stimulatory effect of 15-HPAA in MV was not due to a simple detergent-like action of the compound on the membranes but rather to stimulation of the phospholipase A2 and lipoxygenase activity within MV. In addition, it was shown that free radical reactions were involved in the mechanism. Since such anti-edema drugs as 1,2-bis(nicotinamido)propane were proved to be potent suppressors of the enhanced ATPase activity, further speculations on the role of this effect for ischemic brain edema are offered.     

Microheterogeneity of PSI-E Subunit of Photosystem I in Nicotiana sylvestris

Microheterogeneity of a photosystem I (PSI) subunit encodedby a nuclear gene psaE was examined in Nicotiana sylvestris,with the aid of cDNA cloning, peptide mapping analysis and proteinsequencing. The psaE product of this plant has four isoformswhose mobilities in PAGE are slightly different from each other.We isolated two types of psaE cDNAs from a N. sylvestris cDNAlibrary, and designated the corresponding genes as psaEa andpsaEb, respectively. The psaEa and psaEb genes are 77% homologousat DNA level, and their translation products share 80.4% homologyfor the precursor proteins and 89.1% for the mature forms. Comparativeanalysis of the four isoproteins and the putative products ofthe two psaE genes revealed that two isoproteins out of fourare derived from psaEa gene, and the difference between thesetwo isoproteins lies in the respective presence or absence ofN-terminal alanine. Likewise, the other two proteins are derivedfrom psaEb with similar N-terminal heterogeneity. These resultsindicate that multi-gene organization and heterogeneous N-terminalformation at post-translational level are two possible causesfor PSI subunit polymorphism in isogenic plant lines. (Received October 8, 1993; Accepted November 30, 1993)     

Human endometrial stromal cells and decidual cells express cluster of differentiation (CD) 13 antigen/aminopeptidase N and CD10 antigen/neutral endopeptidase.

With specific monoclonal antibodies, we found that human endometrial stromal cells and decidual cells express two function-related surface antigens. Indirect immunofluorescence staining revealed that both endometrial stromal cells and decidual cells during the first trimester of pregnancy expressed cluster of differentiation (CD) 13 antigen and CD10 antigen, which are identical to aminopeptidase N and neutral endopeptidase, respectively. By flow cytometric analysis, CD13 antigen was detected on 82-93% of the examined cells, and CD10 antigen was detected on 75-93% of the examined cells in endometrial stromal cell-enriched preparations. Furthermore, peptidase activity was detected in these cell preparations by an assay based on the hydrolysis of alanine-p-nitroanilide into p-nitroaniline and alanine.     

Inhibitory effects of interleukin-1 on follicle-stimulating hormone induction of aromatase activity, progesterone secretion, and functional luteinizing hormone receptors in cultures of porcine granulosa cells

Effects of interleukin-1 (IL-1) on FSH-induced differentiation of immature porcine granulosa cells in vitro were examined in short-term (48-h) cultures. IL-1 inhibited FSH induction of aromatase activity and of LH-stimulated cAMP accumulation by granulosa cells. Both these inhibitory actions of IL-1 were concentration-dependent. Significant inhibitory effects were observed with as low as 0.05-0.25 ng/ml of IL-1, with maximal effects at 25 ng/ml. IL-1 also significantly inhibited increases in [125I]iodo-LH binding and progesterone secretion induced by FSH, as well as reducing basal levels of aromatase activity and LH-stimulated cAMP accumulation. Studies on the mechanisms of IL-1 actions on FSH-induced differentiation of immature porcine granulosa cells revealed that IL-1 reduced cAMP accumulation by the cells in response to FSH in a time- and concentration-dependent manner. IL-1 also inhibited induction of aromatase activity and LH-stimulated cAMP accumulation induced by dibutyryl cAMP, suggesting that IL-1 also affects the steps distal to cAMP generation. In contrast, IL-1 had no effect on progesterone secretion induced by dibutyryl cAMP, suggesting that post-cAMP steps of progesterone secretion were unaffected by IL-1.     

Identification and characterization of a resident vascular stem/progenitor cell population in preexisting blood vessels

Vasculogenesis, the in-situ assembly of angioblast or endothelial progenitor cells (EPCs), may persist into adult life, contributing to new blood vessel formation. However, EPCs are scattered throughout newly developed blood vessels and cannot be solely responsible for vascularization. Here, we identify an endothelial progenitor/stem-like population located at the inner surface of preexisting blood vessels using the Hoechst method in which stem cell populations are identified as side populations. This population is dormant in the steady state but possesses colony-forming ability, produces large numbers of endothelial cells (ECs) and when transplanted into ischaemic lesions, restores blood flow completely and reconstitutes de-novo long-term surviving blood vessels. Moreover, although surface markers of this population are very similar to conventional ECs, and they reside in the capillary endothelium sub-population, the gene expression profile is completely different. Our results suggest that this heterogeneity of stem-like ECs will lead to the identification of new targets for vascular regeneration therapy.     

Membrane glycoproteomics of fetal lung fibroblasts using LC/MS

Some aberrant N‐glycosylations are being used as tumor markers, and glycoproteomics is expected to provide novel diagnosis markers and targets of drug developments. However, one has trouble in mass spectrometric glycoproteomics of membrane fraction because of lower intensity of glycopeptides in the existence of surfactants. Previously, we developed a glycopeptide enrichment method by acetone precipitation, and it was successfully applied to human serum glycoproteomics. In this study, we confirmed that this method is useful to remove the surfactants and applicable to membrane glycoproteomics. The glycoproteomic approach to the human fetal lung fibroblasts membrane fraction resulted in the identification of over 272 glycoforms on 63 sites of the 44 glycoproteins. According to the existing databases, the structural features on 41 sites are previously unreported. The most frequently occurring forms at N‐glycosylation site were high‐mannose type containing nine mannose residues (M9) and monosialo‐fucosylated biantennary oligosaccharides. Several unexpected N‐glycans, such as fucosylated complex‐type and fucosylated high‐mannose and/or fucosylated pauci‐mannose types were found in ER and lysosome proteins. Our method provides new insights into transport, biosynthesis, and degradation of glycoproteins.     

Chemical identification and functional analysis of apocarotenoids involved in the development of arbuscular mycorrhizal symbiosis

Arbuscular mycorrhizae formed between more than 80% of land plants and arbuscular mycorrhizal (AM) fungi represent the most widespread symbiosis on the earth. AM fungi facilitate the uptake of soil nutrients, especially phosphate, by plants, and in return obtain carbohydrates from hosts. Apocarotenoids, oxidative cleavage products of carotenoids, have been found to play a critical role in the establishment of AM symbiosis. Strigolactones previously isolated as seed-germination stimulants for root parasitic weeds act as a chemical signal for AM fungi during presymbiotic stages. Stimulation of carotenoid metabolism, leading to massive accumulation of mycorradicin and cyclohexenone derivatives, occurs during root colonization by AM fungi. This review highlights research into the chemical identification of arbuscular mycorrhiza-related apocarotenoids and their role in the regulation and establishment of AM symbiosis conducted in the past 10 years.