HPLC-studies on nonmercapt-mercapt conversion of human serum albumin

Human mercaptalbumin (HMA) and nonmercaptalbumin (HNA) could be separated by high-performance liquid chromatography (HPLC) at neutral pH. Using HPLC, the present authors found the nonmercapt-mercapt conversion (HNA----HMA) during hemodialysis and the mercapt-nonmercapt conversion (HMA----HNA) after hemodialysis in chronic renal failure, indicating HMA as the covalent carrier protein for sulfur-containing amino acids.     

Possible sensory receptor of nonadrenergic inhibitory nervous system

To determine the sensory receptor of the nonadrenergic inhibitory nervous system (NAIS), 22 cats were anesthetized and serotonin was continuously administered (50-250 micrograms.kg-1.min-1 iv) to increase pulmonary resistance (RL) to 377 +/- 57% (SE) of the control value. We then 1) mechanically irritated the trachea, 2) intravenously administered capsaicin (5 micrograms/kg), or 3) induced hypoxia (arterial PO2 30-40 Torr) to stimulate irritant and bronchial C-fiber receptors, pulmonary C-fiber receptors, or the carotid body (chemoreceptors), respectively. After treatment with atropine (3 mg/kg iv) and propranolol (2 mg/kg iv), the serotonin-induced change in RL was reduced by 58.6 +/- 14.3% by mechanical irritation and 63.3 +/- 12.1% by intravenous capsaicin. However, hypoxia produced no dilatation of the airways. In further experiments, we employed capsaicin inhalation to stimulate bronchial C-fiber receptors. Inhaled capsaicin (0.1%, for 5 breaths) also reduced RL by 79.2 +/- 9.2% of the elevated value, after atropine and propranolol. Treatment with a ganglionic blocking agent, hexamethonium (2 mg/kg iv), abolished bronchodilator responses, implying that a reflex pathway through vagal nerves is involved in this phenomenon. These results suggest that pulmonary and bronchial C-fiber receptors may be involved as sensory receptors in NAIS reflex bronchodilatation.     

Effect of 5-azacytidine on the differentiation of human leukemia K-562 cells

The treatment of K-562 cells with 10(-5) M to 10(-7) M 5-azacytidine induced a marked increase in benzidine-positive cells. Similarly, the exposure of K-562 cells to 2 X 10(-3) M butyric acid or 5 X 10(-7) M 1-beta-arabinofuranosylcytosine or 1 X 10(-3) M hydroxyurea induced an erythroid differentiation of K-562 cells. The activity of DNA-methyltransferase and the level of methylcytosine in newly synthesized DNA were significantly decreased when the cells were treated with 5-azacytidine or butyric acid, while 1-beta-arabinofuranosylcytosine or hydroxyurea had no inhibitory effect on DNA-methylation of K-562 cells. These results suggest that the inhibition of DNA-methylation is not necessarily a specific phenomenon for erythroid differentiation of K-562 cells.     

Distribution and biosynthesis of aminopeptidase N and dipeptidyl aminopeptidase IV in rat small intestine

The regional, cellular and subcellular distribution patterns of aminopeptidase N and dipeptidyl aminopeptidase IV were examined in rat small intestine. Aminopeptidase N of brush border membrane had maximal activity in the upper and middle intestine, while dipeptidyl aminopeptidase IV had a more uniform distribution profile with relatively high activity in the ileum. Along the villus and crypt cell gradient, the activity of both enzymes was maximally expressed in the mid-villus cells. However there was substantial dipeptidyl aminopeptidase IV activity in the crypt cells. Both enzymes were primarily associated with brush border membranes in all segments, however, in the proximal intestine, a significant amount of dipeptidyl aminopeptidase IV activity was associated with the cytosol fraction. The cytosol and brush border membrane forms of dipeptidyl aminopeptidase IV were immunologically identical and had the same electrophoretic mobility on disc gels. In contrast, the soluble and brush border membrane-bound forms of aminopeptidase N were immunologically distinct. When the total amount of aminopeptidase N and dipeptidyl aminopeptidase IV was determined by competitive radioimmunoassay, there were no regional or cellular differences in specific activity (enzyme activity/mg of enzyme protein) of either enzyme in brush border membrane and homogenate. The specific activity of both enzymes in a purified Golgi membrane fraction as measured by radioimmunoassay was about half that of the brush border membrane fraction. These results suggest that (1) aminopeptidase N and dipeptidyl aminopeptidase IV have different regional, cellular and subcellular distribution patterns; (2) there are enzymatically inactive forms of both enzymes present in a constant proportion to active molecules and that (3) a two-fold activation of precursor enzyme forms occurs during transfer from the Golgi membranes to the brush border membranes.     

Experimental studies on vertical infection of mice with Japanese encephalitis virus. IV. Effect of virus strain on placental and fetal infection

An experiment was carried out to examine the effect of an inoculated strain of Japanese encephalitis virus on the establishment of experimental vertical infection of mice with this virus. In it, closed-colony mice of the CFW strain were inoculated intravenously with seven strains of the virus at 7 days of pregnancy. After that, an attempt was made to recover the virus from placenta and fetus, so that the infection rate of each strain might be determined. As a result, the infection rate was high for both placenta and fetus in the case of the AS-6 and Sagara strains both of which had undergone three passages in the mouse brain. The placental infection rate was high and the fetal infection rate relatively low in the case of the JaGAr01 and Fuji strains which had undergone 7 and 150 passages, respectively, in the mouse brain. The infection rate was very low for both placenta and fetus in the case of the Nakayama-Yakken strain which had undergone more than 100 passages in the mouse brain. There was no difference in the severity of viremia after inoculation between the AS-6 and Fuji strains. Both placental and fetal infection rates were low in the case of the JaTH160 strain which had undergone passages in mice by intraperitoneal inoculation and which presented a strong peripheral infectivity and induced a severe viremia after inoculation. Neither placental nor fetal infection occurred in the case of the S- strain used as live virus vaccine. These results indicated that placental and fetal infection rates varied from one virus strain to another.(ABSTRACT TRUNCATED AT 250 WORDS)     

A temperature-sensitive mutant of E. coli exhibiting slow processing of exported proteins

A temperature-sensitive E. coli mutant with a mutation in the spc ribosomal protein operon was found to have a conditional defect in the processing of precursor proteins destined for the periplasmic space or the outer membrane. At high temperatures, significant amounts of precursor proteins having unprocessed signal sequences are detected in the mutant cell by pulse-labeling. The precursors are processed at very slow rates during a subsequent chase. Genetic analysis indicates that the mutation impairs the function of a gene, termed secY, located at the promoter-distal part of the spc operon. The secY gene is distinct from those genes previously known to specify ribosomal proteins, yet it is within the spc operon. It is suggested that the product of the secY gene is a component of the cellular apparatus that is essential for protein secretion across the cytoplasmic membrane. The gene secY is probably identical with prlA, previously identified as a suppressor of signal sequence mutations.     

Role of lymphokines in regulation of macrophage differentiation

The regulatory mechanism of guinea pig lymphokines was investigated in regard to differentiation of myeloid cells to macrophages. The Ml-cell line, established from a myeloid leukemia of an SL-strain mouse, was induced to differentiate in vitro into mature macrophages possessing Fc receptors and the ability to phagocytize latex particles by treatment with crude lymphokines. Both concanavalin A- and antigen-induced lymphokines showed the differentiation-inducing factor (D factor) activity. However, macrophage migration inhibitory factor/ macrophage activation factor (MIF/MAF) purified by an immunoadsorbent column with anti-MIF antibody had no such an activity. The D-factor activity was detected in the lymphokine preparation that was not retained on the immunoadsorbent column. In contrast, colony-stimulating factor (CSF) was adsorbed to the immunoadsorbent column, and could be recovered in the purified MIF/MAF preparation. These findings suggest that the molecular entity of D factor is distinct from MIF/ MAF and CSF. A culture supernatant of guinea pig peritoneal macrophages activated with MIF/ MAF (CSF) exhibited strong D-factor activity. However, the supernatant possessed rather reduced CSF activity as compared to that of the original MIF/MAF (CSF) preparation. Thus, MIF/MAF may play an important role in macrophage differentiation by regulating the production of D factor or CSF from macrophages.     

The Effects of Direct Instillations of a Therapeutic Mixture Into the Tracheobronchial Tree

Twenty-two patients with chronic productive bronchitis or bronchiectasis were treated by direct instillations of normal saline and N-acetylcysteine into the trachea through a percutaneous catheter following a period of conventional routine therapy.The instillation, using either normal saline or varying concentrations of N-acetylcysteine did not produce significant change in alveolar gas exchange as reflected by measurement of arterial PaCO₂ and the alveolar arterial gradient for oxygen during and after the introduction of the medication into the bronchial tree. Studies were carried out after patients had been stabilized breathing pure oxygen on an IPPB machine for 30 minutes.Evaluation of the treatment by means of pulmonary function tests demonstrated significant improvement in overall function following therapy.The results indicate that the technique of tracheobronchial lavage is physiologically benign and that overall improvement in pulmonary function can be obtained by this means in cases of the type described in this study.