Preferred ambient temperature for old and young men in summer and winter

To investigate the effects of age on thermal sensitivity, preferred ambient temperature (T _pref) was compared between old (71–76 years) and young (21–30 years) groups, each consisting of six male subjects in summer and winter. The air temperature (T _a) was set at either 20° C or 40° C at commencement. The subject was directed to adjust theT _a for 45 min by manipulating a remote control switch to the level at which he felt most comfortable. In the older group, theT _pref was significantly lower in trials starting at 20° C than that starting at 40° C in summer. The fluctuation ofT _pref (temperature difference between maximum and minimumT _a during the last 10 min) was significantly wider in the older group in both summer and winter. Repetition of the same experiment on each subject showed a poorer reproducibility ofT _pref in the older group than in the younger group in summer. Tympanic and esophageal temperatures of the older group kept falling throughout the trial starting at 20° C in summer. These results suggest that thermal sensitivity is decreased with advancing age and that thermal perception in the elderly, especially to cold, is less sensitive in summer.     

Electrotransformation of Thiobacillus ferrooxidans with plasmids containing a mer determinant.

The mer operon from a strain of Thiobacillus ferrooxidans (C. Inoue, K. Sugawara, and T. Kusano, Mol. Microbiol. 5:2707-2718, 1991) consists of the regulatory gene merR and an operator-promoter region followed by merC and merA structural genes and differs from other known gram-negative mer operons. We have constructed four potential shuttle plasmids composed of a T. ferrooxidans-borne cryptic plasmid, a pUC18 plasmid, and the above-mentioned mer determinant as a selectable marker. Mercury ion-sensitive T. ferrooxidans strains were electroporated with constructed plasmids, and one strain, Y4-3 (of 30 independent strains tested), was found to have a transformation efficiency of 120 to 200 mercury-resistant colonies per microgram of plasmid DNA. This recipient strain was confirmed to be T. ferrooxidans by physiological, morphological, and chemotaxonomical data. The transformants carried a plasmid with no physical rearrangements through 25 passages under no selective pressure. Cell extracts showed mercury ion-dependent NADPH oxidation activity.     

Functional involvement of P-glycoprotein in blood-brain barrier.

P-glycoprotein, an active efflux pump of antitumor agents in multidrug-resistant tumor cells, exists in various normal tissues, including brain capillaries. To study the physiological function of P-glycoprotein expressed in brain capillary endothelium, we established nine mouse brain capillary endothelial cell (MBEC) lines and examined the transport of antitumor agents across the monolayer of MBEC epithelia. In the MBECs, the activities of alkaline phosphatase and gamma-glutamyl transpeptidase, specific markers for brain capillary endothelial cells, were about three times higher than those in other cells including human umbilical vein endothelial cells. By immunoblot analysis, P-glycoprotein was detected in all of the nine MBEC clones. The P-glycoprotein expressed in MBECs specifically bound [125I]iodoaryl azidoprazosin as that in multidrug-resistant cells, and efflux of vincristine was observed in the MBECs. When MBECs were grown on a porous filter membrane, they formed a monolayer of epithelium. By immunoelectron microscopic analysis, P-glycoprotein in MBEC epithelia was shown to be localized to the apical surface of the cells. Moreover, the unidirectional transepithelial transport of vincristine from basal side to apical side was demonstrated in vitro. These observations indicate that P-glycoprotein in brain capillary endothelium prevents vincristine from entering the central nervous system and thus may be one of the functional components of the blood-brain barrier.     

Differential effects of protein synthesis inhibition on CTL and targets in cell-mediated cytotoxicity

The reactions that lead to target cell lysis by cytotoxic T cells (CTL) are despite intensive investigations poorly understood. To examine the relative roles effectors and targets play in the lytic reaction, protein synthesis in either CTL or targets was inhibited before assay of lysis. We show, in agreement with previous results, that de novo protein synthesis is not necessary in either effectors or targets during the cytolytic reaction. However, activation of CTL requires protein synthesis. Activated CTL respond to protein synthesis inhibitors with a cycling of activity, a result that is interpreted to be consistent with a stimulus secretion mechanism. Treatment of targets with protein synthesis inhibitors prior to incubation with CTL leads to a very rapid and irreversible loss of lytic susceptibility. It is shown that the decrease in lysability is not due to lack of proper CTL target interaction: MHC class I antigens are expressed on drug-treated targets and these cells serve as cold targets in competitive inhibition experiments. Moreover, drug-treated targets trigger transient Ca2+ mobilization and generation of inositol phosphates in CTL. It is therefore concluded that drug-treated targets are able to trigger CTL function but lack a component that is required for their successful lysis.     

Potentiation by BL191 of differentiation of neuroblastoma cells induced by dibutyryl cAMP and prostaglandin E1

BL191, a newly developed phosphodiesterase inhibitor, markedly potentiated a differentiation of neuroblastoma cell clones (Neuro2a, NS-20Y, and N1E115) induced by dibutyryl cyclic adensoine 3′:5′-monophosphate(dibutyryl cAMP) and prostaglandin E₁ (PGE₁). BL191 (1 mM) inhibited DNA synthesis more strongly when used together with PGE₁ (0.5 μg/ml) and dibutyryl cAMP (0.5 mM) than papaverine (1.6 μg/ml) alone did. The inhibition rates of DNA synthesis were 72.5% for N1E-115, 75.3% for Neuro2a, and 82.5% for NS-20Y. After the treatment with BL191. PGE₁, and dibutyryl cAMP for 48 h all of three cell lines became enlarged and flattened, and extended long processes. The specific activities of choline acetyl transferase (EC 2.3.1.9) of NS-20Y and dopamine β-hydroxylase (EC 1.14.17.1) of N1E-115 increased about 3-fold as compared to the controls. The tumorigenicities of Neuro2a and N1E-115 cells were decreased, but not of NS-20Y. These data suggest the heterogenous responsiveness in neuroblastoma cells to drug treatment.     

Purification and some properties of beta-lactamases from Proteus rettgeri and Proteus inconstans

Two beta-lactamases were isolated from strains of Proteus species and purified, one from a strain of P. rettgeri and the other from a strain of P. inconstans. Each enzyme preparation gave a single protein band on polyacrylamide gel electrophoresis. Molecular weights of P. rettgeri and P. inconstans enzymes were found to be 42,000 and 43,000, and their isoelectric points pH 8.7 and 8.6, respectively. The two enzymes presented typical cephalosporinase profiles. Cefmetazole (CS-1170) and cefoxitin, both cephamycin antibiotics, not only resisted hydrolysis by both of the enzymes, but also inhibited their activities competitively. Rabbit antiserum against purified P. rettgeri enzyme inhibited the activity of both purified and crude enzyme preparations from other strains of P. rettgeri so far tested. None of the beta-lactamases produced by other species of Proteus including P. inconstans was inhibited by the antiserum, thus showing that the purified cephalosporinase was of the species-specific types. The enzymological properties of the preparations were compared with those of beta-lactamases derived from other gram-negative enteric bacteria.     

Cloning and nucleotide sequence of the mono- and diacylglycerol lipase gene (mdlB) of Aspergillus oryzae

Abstract Aspergillus oryzae IFO4202 produces at least two extracellular lipolytic enzymes L1 and L2 (cutinase, and mono- and diacylglycerol lipase, respectively). Southern hybridization of restriction enzyme-digested genomic DNA fragments with 23mer oligonucleotides synthesized according to the amino acid sequence of the L2 as probe suggested the presence of the L2 gene (tentatively designated as mdlB ) and an additional weakly hybridizing region. A fragment containing the genomic mdlB gene was cloned in Escherichia coli . Nucleotide sequencing of the fragment revealed an open reading frame, comprising 1021 nucleotides, which contains two introns (51 and 52 nucleotides). Putative polyadenylation signals were found 182 and 287 bp downstream of the stop codon. The deduced amino acid sequence of the mdlB gene corresponds to 306 amino acid residues including a leader sequence of 28 amino acids and is highly similar to that of the mdlA gene of Penicillium camembertii . Three residues presumed to form the catalytic triad (serine, aspartic acid and histidine) of lipases were also conserved.