Enterobacter kobei sp. nov., a new species of the family Enterobacteriaceae resembling Enterobacter cloacae

The name Enterobacter kobei sp. nov. is proposed for a group of organisms referred to as NIH Group 21 at the National Institute of Health, Tokyo. The members of this species are Gram-negative, motile rods conforming to the definition of the family Enterobacteriaceae. The DNA relatedness of 23 strains of NIH Group 21 to the representative proposed as the type strain of this species averaged 82% at 70°C, whereas the relatedness to other species within the family Enterobacteriaceae was less than 42%. Because the phenotypic resemblance to Enterobacter cloacae is very close and the DNA relatedness (12–42%) is closer to species of the genus Enterobacter than to other species of the family, the members of NIH Group 21 were placed in the genus Enterobacter. Close phenotypic and genetic relationships were also found between NIH Group 21 and a member of a group of organisms referred to as Enteric Group 69 at the Centers for Disease Control and Prevention (CDC), Atlanta, Georgia, USA. It is suggested that the latter could be regarded as a subspecific rank of E. kobei, though this is subject to study of further strains. The majority of strains of E. kobei were isolated from clinical specimens. A culture of the type strain (NIH 1485-79) has been deposited in the Japan Collection of Microorganisms as JCM 8580. Received: 22 March 1996 / Accepted: 19 April 1996     

Expression of host genes in influenza virus infected cells

The NS1 protein of influenza virus shuts off host gene expression by inhibiting the polyadenylation-site cleavage of host pre-mRNAs, resulting in a general decline in cellular protein synthesis. On the other hand, an activation of several host genes related to host antiviral defense such as interferon- alpha/beta, MxA, 2',5'-oligoadenylate synthetase, and Fas occures upon infection. Therefore, balance of the shut-off and the activation of cellular genes during virus growth may be crucial in determining the outcome of infection. To obtain a comprehensive view of the global effects of influenza virus infection on human respiratory epithelial cells at the cytoplasmic mRNA level, we performed oligo DNA microarray analysis using GeneChip arrays (Affymetrix). In NCl-H292 cells infected with A/Udorn/72 virus, more than 4-fold increase of expression level was observed for 164 genes at 12 h pi. Approximately 60% of the virus-stimulated genes (VSGs) were also stimulated with interferon-beta treatment and contained the genes known to possess antiviral activity. Interestingly, majority of the VSGs were stimulated before induction of interferons, suggesting that the stimulation of the VSGs during early phase of infection is not mediated by interferons, but it is triggered from within by the virus infection.     

Utility of NucleoCounter for the chondrocyte count in the collagenase digest of human native cartilage

In cartilage tissue engineering, viable cell numbers should be correctly counted in the collagenase digest of the biopsied cartilage. However, this is a difficult task due to the presence of matrix debris, cell ghosts and their aggregates. To search for the correct cell counting method in this situation, we evaluated the utility of an automatic cell counting device, the NucleoCounter, and compared it with conventional staining using the LIVE/DEAD® kit. We first measured the cell numbers of a standard chondrocyte sample by the NucleoCounter, which showed a high accuracy (R² = 0.9999) and reproducibility (%CV: 2.00–8.66). We then calculated the cell numbers and viability in some collagenase digests of native cartilage using either the NucleoCounter or LIVE/DEAD® kit, revealing that the total cell numbers, viable ones and viability were highly correlated between them (R² = 0.9601, 0.9638 and 0.917, respectively). However, both the intrapersonal and interpersonal variabilities in the NucleoCounter was significantly decreased to about 1/20–1/5, compared to that of the LIVE/DEAD® kit. The NucleoCounter was regarded as a useful tool for simple, rapid, and highly reproducible cell counts, which may not only provide constant experimental data in a certain laboratory, but also contribute to the high reproducibility of the clinical results of cartilage tissue engineering among multiple institutions.     

Expression of novel organic cation/carnitine transporter (OCTN2) in the mouse pancreas

Among the organic cation transporters, OCTN2 is identified as the most important carnitine transporter owing to the ability to transport carnitine. Although the OCTN2 is previously found in various tissues, there have been no reports showing the OCTN2 in the pancreas. In this study, we examined the expression and localization of OCTN2 in the mouse pancreas by the aid of an in situ hybridization technique and immunohistochemistry with anti-OCTN2 antibody. As a result, the OCTN2 expression was found in the A-cells for the first time. OCTN2 was not expressed in B-cells, notwithstanding that the metabolism of long-chain fatty acids, which are transported into the mitochondria with the help of carnitine, was expected for fatty acid-stimulated insulin secretion. Thus, this study suggests the possibility of carnitine uptake in the pancreatic A-cells through OCTN2 and implies the presence of carnitine transporter(s) other than OCTN2 in the B-cell.     

Dynamic changes in expression of heme oxygenases in mouse heart and liver during hypoxia

Heme oxygenase cleaves heme to form biliverdin, carbon monoxide (CO), and iron, and consists of two structurally related isozymes, HO-1 and HO-2. HO-2 is also known as a potential oxygen sensor. Here we show that the relative CO content in arterial blood, which reflects the total amount of endogenous heme degradation, dynamically changes in mice during acclimatization to normobaric hypoxia (10% O2), with the two peaks at 1 day and 21 days of hypoxia. The expression levels of HO-1 and HO-2 proteins were decreased by 20% and 40%, respectively, in the mouse liver at 7 days of hypoxia, which returned to the basal levels at 14 days. On the other hand, HO-1 and HO-2 proteins were increased 2-fold and 1.3-fold, respectively, in the heart at 28 days of hypoxia. Thus, hypoxia induces or represses the expression of HO-1 and HO-2 in vivo, depending on cellular microenvironments.