T-cadherin (CDH13, H-cadherin) expression downregulated surfactant protein D in bronchioloalveolar cells

T cadherin is a unique cadherin cell adhesion molecule that is anchored to the surface membrane through a glycosyl phosphatidyl inositol (GPI) moiety. In the present study, we postulated that T cadherin could regulate surfactant protein (SP)-D gene expression in human bronchioloalveolar type-II cells. We transfected A549 cells (human lung cancer cell line with alveolar type-II cell characteristics) with the T-cadherin expression vector. Both original and control plasmid-transfected A549 cells expressed SP-D; however, neither human nor murine T-cadherin-transfected A549 cells expressed SP-D mRNA. The downregulation of SP-D production in human T-cadherin-expressed A549 cells was also demonstrated using Western immunoblotting techniques. Control vector-transfected A549 cells showed a positive band of SP-D but not of T cadherin. In contrast, T-cadherin-transfected A549 cells, which expressed T-cadherin protein, did not produce SP-D. We further examined the relationship of T cadherin and SP-D expression in secondary pulmonary alveolar proteinosis associated with hematolymphoid malignancies. SP-D was detected in bronchioloalveolar type-II cells in alveolar proteinosis. However, little or no T-cadherin expression was detected in alveolar type-II cells in these patients. To our knowledge, this is the first report describing an effect of cadherin on SP production in bronchioloalveolar cells.     

The adrenergic modulation of firings of respiratory rhythm-generating neurons in medulla-spinal cord preparation from newborn rat

We analysed the modulation of respiratory neurons by adrenaline or noradrenaline (NA) in a newborn rat brainstem-spinal cord preparation. Adrenaline or NA caused a dose-dependent depression of the respiratory rhythm and induced C4 spinal tonic discharges. The inhibitory effect of adrenaline (ED₅₀=0.5 μM) on the respiratory rhythm was stronger than NA (ED₅₀=5 μM). The adrenaline respiratory rhythm depression was partially blocked by the α₁-antagonist prazosin or by the α₂-antagonist yohimbine. The C4 tonic discharge elicited by adrenaline was blocked by the α₁-antagonist prazosin. The direct effects of adrenaline on pre-inspiratory (Pre-I) neurons were examined in a synaptic blockade solution (low Ca), and fifty-six percent of Pre-I neurons were found to continue firing. In low-Ca solution, Pre-I neurons were excited (n=29 of 39) or depressed (n=5 of 39) by adrenaline, and excited by α₁-agonist phenylephrine or depressed by α₂-agonist clonidine. These results suggest that the respiratory rhythm depression under intact network conditions is mediated by some other inhibitory system. The inhibitory effect of adrenaline on the respiratory rhythm was partially blocked by the GABA_A-antagonists bicuculline or picrotoxin, but not by the GABA_B-antagonist phaclofen. The present results suggest that: (1) respiratory rhythm generation is more sensitive to adrenaline than NA through α-adrenergic action of adrenaline; (2) the activity of Pre-I neurons could be directly regulated by excitation via α₁-receptors and inhibition via α₂-receptors; and (3) the depression of the respiratory rhythm by adrenaline is partly mediated by GABA_Aergic neurons. Received: 8 April 1997 / Accepted: 6 October 1997     

Analysis of proteins in urinary tract stones and urine of urolithic patients

In order to investigate the mechanism of urinary tract stone formation, we analyzed protein components in urine and the stone. Urinary proteins of healthy subjects and urolithic patients as well as protein components urinary tract stone of the urolithic patients were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Electrophoretic patterns of urinary proteins of the patients differed from those of healthy subjects after separating protein patterns into those larger than 66kDa or smaller than 30kDa. Protein constituents of urinary tract stone were mainly separated into 18 bands ranging from 26.8 to 143 kDa. Major bands among these 18 bands differed among stones from different patients. On western blotting, the developed intensities of Tamm-Horsfall protein (THP) were fainter than those of healthy subjects. Whereas intensities of albumin (ALB) were stronger than those of healthy subjects. Moreover, blotting patterns of THP of the patients on non-reducing SDS-PAGE were obviously broad. Thus, we suggest that analysis of fractionated urinary proteins or protein components of urinary tract stone may provide a tool for monitoring the prognosis or relapse in the patients.     

The expression of neuronal nitric oxide synthase and dystrophin in rat regenerating muscles

We investigated the expression of neuronal nitric oxide synthase (nNOS) and dystrophin in the regenerating skeletal muscles of rats after cardiotoxin-induced myonecrosis by immunohistochemical studies and western blot analysis. In normal muscles, nNOS was moderately immunostained on type 2B fibers, but was faintly immunostained on type 2A or type 1 fibers. In immunohistochemical studies of regenerating muscles, nNOS was first observed at the sarcolemma of type 2B fibers on day 10, when the type discrimination between types 2A and 2B was first detected by ATP reactions. Subsequently, the immunostaining of nNOS grew progressively stronger in type 2B fibers, with faint staining in type 2A and type 1 fibers until day 28. Meanwhile, the immunostaining of dystrophin grew stronger equally in all three fibers until day 21. In western blot analysis of regenerating muscles, nNOS regenerated more slowly than dystrophin. The present data suggest that the expression of nNOS is related to the muscle fiber type differentiation, and that the role of nNOS is related to the function of the type 2B fibers of the muscle. This revised version was published online in July 2006 with corrections to the Cover Date.     

A novel insertional mutation at exon VII of the myelin proteolipid protein gene in Pelizaeus -- Merzbacher disease

Pelizaeus — Merzbacher disease (PMD) is an X-linked neurologicaldisorder characterized by dysmyelination in the central nervoussystem (CNS). Recently mutations of the myelln proteollpid protein(PLP) gene which encodes both PLP and Its Isoform, DM-20 generatedby alternative spllcing, have been demonstrated In PMD patients.We analyzed the seven exons of the PLP gene of a Japanese boyaffected with PMD by direct sequencing and identified an Insertionevent In exon Vll of the PLP gene. This mutation was also presentIn his carrier mother, but was absent In ninety-five X chromosomesof normal Japanese. The frame-shift mutation leads to the productionof truncated PLP with altered carboxyl terminal amlno acid sequences,resulting In conslderable change of the structure of PLP andDM-20 necessary for functional purposes. This is the first reportof a mutation In exon Vll of the PLP gene associated with PMD.     

Development of early neutropenic fever, with or without bacterial infection, is still a significant complication after reduced-intensity stem cell transplantation.

Little information is available on the clinical characteristics of infectious complications that occur in the early period after reduced-intensity stem cell transplantation (RIST). We retrospectively investigated the clinical features of neutropenic fever and infectious episodes within 30 days after RIST in 76 patients who had received fluoroquinolones as part of their antibacterial prophylaxis. Preparative regimens included cladribine 0.66 mg/kg or fludarabine 180 mg/m2 plus busulfan 8 mg/kg. All but 1 patient survived 30 days after transplantation, and 75 patients (99%) became neutropenic within a median duration of 9 days. Neutropenic fever was observed in 29 patients (38%), and bacterial infection was confirmed in 15 (20%) of these, including bacteremia (n = 13), bacteremia plus pneumonia (n = 1), and urinary tract infection (n = 1). The causative organisms were gram-positive (n = 9) and gram-negative organisms (n = 7), with a mortality rate of 6%. Neither viral nor fungal infection was documented. Multivariate analysis showed that the presence of neutropenia at the initiation of preparative regimens was an independent risk factor for subsequent documented bacterial infections (P =.026; 95% confidence interval, 1.25-35.1). We conclude that neutropenic fever and bacteremia remain common complications in RIST.     

A pore-forming toxin produced by Aeromonas sobria activates Ca2+ dependent Cl- secretion

Bacteria produce many types of hemolysin that induce diarrhea by mechanisms that are not completely understood. Aeromonas sobria hemolysin (ASH) is a major virulence factor produced by A. sobria, a human pathogen that causes diarrhea. Since epithelial cells in the intestine are the primary targets of hemolysin, we investigated the effects of ASH on ion transport in human colonic epithelial (Caco-2) cells. ASH increased short-circuit currents (Isc) in a dose-dependent manner, and it also activated a 125I efflux from Caco-2 cells. ASH-induced Isc increases and 125I efflux activations were both suppressed by low Ca2+ levels in the extracellular solution or by pretreatment with the Ca2+ chlelator BAPTA-AM. Intracellular Ca2+ levels were increased by ASH in a biphasic fashion characterized by a rapid sharp increase (peak 1) followed by a sustained low plateau (peak 2). ASH-induced peak 1 was inhibited by pretreatment with pertussis toxin, indicating that Ca2+ was mobilized from intracellular stores, and peak 2 was induced by an influx of extracellular Ca2+. Peak 2 but not peak 1 was related to Cl- secretion. These results indicate that ASH activates Ca2+-dependent Cl- secretion.     

Long‐term observation of a Japanese mucolipidosis IV patient with a novel homozygous p.F313del variant of MCOLN1

Mucolipidosis type IV (MLIV) is an autosomal recessively inherited lysosomal storage disorder characterized by progressive psychomotor delay and retinal degeneration that is associated with biallelic variants in the MCOLN1 gene. The gene, which is expressed in late endosomes and lysosomes of various tissue cells, encodes the transient receptor potential channel mucolipin 1 consisting of six transmembrane domains. Here, we described 14‐year follow‐up observation of a 4‐year‐old Japanese male MLIV patient with a novel homozygous in‐frame deletion variant p.(F313del), which was identified by whole‐exome sequencing analysis. Neurological examination revealed progressive psychomotor delay, and atrophy of the corpus callosum and cerebellum was observed on brain magnetic resonance images. Ophthalmologically, corneal clouding has remained unchanged during the follow‐up period, whereas optic nerve pallor and retinal degenerative changes exhibited progressive disease courses. Light‐adapted electroretinography was non‐recordable. Transmission electron microscopy of granulocytes revealed characteristic concentric multiple lamellar structures and an electron‐dense inclusion in lysosomes. The in‐frame deletion variant was located within the second transmembrane domain, which is of putative functional importance for channel properties.     

Isolation,characterization and chromosomal mapping of an actin gene from the primitive red alga Cyanidioschyzon merolae

Based on the results of cytological studies, it has been assumed that Cyanidioschyzon merolae does not contain actin genes. However, Southern hybridization of C. merolae cell-nuclear DNA with a yeast actin-gene probe has suggested the presence of an actin gene in the C. merolae genome. In the present study, an actin gene was isolated from a C. merolae genomic library using a yeast actin-gene probe. The C. merolae actin gene has no intron. The predicted actin is composed of 377 amino acids and has an estimated molecular mass of 42003 Da. Southern hybridization indicated that the C. merolae genome contains only one actin gene. This gene is transcribed at a size of 2.4 kb. When Southern hybridization was performed with C. merolae chromosomes separated by pulsed-field gel electrophoresis, a band appeared on unseparated chromosomes XI and XII. A phylogenetic tree based on known eucaryote actin-gene sequences revealed that C. merolae diverged after the division of Protozoa, but before the division of Fungi, Animalia and Chlorophyta.