Role of ATP in the sensitivity to heat and the induction of apoptosis in mammalian cells.

Heat-induced cell death and apoptosis were studied with respect to intracellular ATP. Studies on the relationship between hyperthermic cell-killing at 44 degrees C and cellular ATP levels in four cell lines grown as monolayers and six cell lines grown in suspension showed good correlations between cellular ATP levels and the sensitivity to heat. D(0) values (the dose required to reduce survival in the linear portion of the response by 63%) linearly increased with an increase in cellular ATP levels. No such changes in sensitivity to heat were observed between the cells cultured at different cell densities, regardless of the change in the cellular ATP level. These results suggest that cellular intrinsic ability to supply ATP rather than the level of pooled ATP per se is responsible for the thermal response. Heat-induced apoptosis in L5178Y cells was observed following treatment at 42 degrees C for 70 min, 44 degrees C for 20 min or 47 degrees C for 3 min, which corresponded to surviving fractions of 25, 0.6 and 0.8%, respectively, but not at 47 degrees C for 20 min, indicating that mild heat shock induced apoptosis. 2-deoxyglucose (2DG) and 2,4-dinitrophenol (DNP) increased the sensitivity to heat and affected the mode of cell death. Cells treated with 2DG and DNP (2DG/DNP) were heated at 42 degrees C for 20 min, and then incubated at 37 degrees C for up to 2h in the presence or absence of 2DG/DNP. In the absence of 2DG/DNP, the cellular ATP level recovered to 76% of the control level and DNA ladder formation was observed, whereas in the presence of 2DG/DNP, the cellular ATP level was further decreased (3-7% of the control) and no DNA fragmentation was detected. These results suggest that the inhibition of ATP synthesis is closely associated with the enhancement of sensitivity to heat and that ATP is required for the induction of apoptosis.

[Evaluation of non-surgical therapy for the malignant lymphomas located mainly in the stomach and duodenum].

We evaluated the effects of the conservative treatment to 29 patients with non-Hodgkin lymphomas located mainly in stomach and duodenum. We could induce complete remission in all the cases of stage I, II, and III including MALT lymphomas by the combination therapy of Helicobacter pylori (H. pylori) eradication, chemotherapy, and radiation, and the complete remission persisted in all except for the three cases who died of other causes. Even in stage IV lymphomas mainly located in stomach and duodenum, we could induce complete remission in 50% of them. Considering the quality of life of patients with lymphomas located mainly in stomach and duodenum, the conservative treatment may be of benefit more than the surgical approach.

[Isoproterenol continuous nebulization of childhood status asthmaticus. II. Efficacy and side effects of low-dose method comparing with high-dose method].

We investigated the efficacy and side effects of "low-dose isoproterenol continuous nebulization" for childhood status asthmaticus, and compared them with those of "high-dose method". "Low-dose" is defined as 10 ml or less of 0.5% dl-isoproterenol solution diluted in 500 ml of normal saline. Subjects were 23 children who were hospitalized and underwent the nebulization therapy. The period of continuous nebulization was 26.3 +/- 12.5 hours. The Wood's clinical score clearly decreased in 22 cases, the average score changing from 7.3 +/- 1.2 to 2.8 +/- 1.5. Heart rate was not elevated significantly during the nebulization period, and decreased gradually (142 +/- 22/min at the start of the nebulization, 145 +/- 22/min at 1 hour, and 134 +/- 23/min at 3 hours, and 103 +/- 13/min at the cessation of the nebulization). Serum GOT, LDH, CPK, and potassium were decreased after the nebulization compared with the values before the treatment, the changes of the last 2 items being statistically significant. Two subjects who had vomited before the nebulization therapy complained nausea during the procedure, and one experienced transient finger tremor. We conclude that "low-dose isoproterenol continuous nebulization" is an effective and safe method for childhood status asthmaticus. We expect that this method will be familiar to all clinicians.

[Isoproterenol continuous nebulization for childhood status asthmaticus. I. Efficacy and side effects of high-dose method].

We investigated the efficacy and the side effects of "high-dose isoproterenol continuous nebulization" for childhood status asthmaticus. Subjects were 34 children who were hospitalized and underwent the nebulization therapy. The 50 ml solution of 0.5% dl-isoproterenol was diluted in 500 ml of normal saline and nebulized through an ultrasound nebulizer. The period of continuous nebulization was 25.5 +/- 16.0 hours. The Wood's clinical score clearly decreased in 32 cases, the average score changing from 7.7 +/- 0.8 to 2.9 +/- 1.3. Heart rate was elevated significantly during the first 3 hours (156 +/- 25/min at the start of the nebulization, 180 +/- 20/min at 1 hour, 171 +/- 23 at 3 hours), and then it decreased gradually to 122 +/- 25/min at the cessation of the nebulization. Serum GOT, LDH, CPK, and potassium were elevated after the nebulization compared with the values before the treatment, though the changes were not statistically significant. CPK-MB fraction after the nebulization was higher than normal range in 12 of 13 subjects. Of 34 subjects, 11 (32%) complained nausea or vomited, 2 showed arrhythmia on ECG (ventricular premature conduction), 1 developed myocardiac infarction, and 1 developed possible heart failure, some of which might be attributable to the pharmacological side effects of isoproterenol nebulization. We conclude that "high-dose isoproterenol continuous nebulization" is an effective method for childhood status asthmaticus, but there is some risk of serious side effects. This method was originally developed as a method indicated for the case of respiratory failure or threatened respiratory failure following status asthmaticus, and we should not extend the indication of this method thoughness.

Identification of c-fos related genes and their induction by neural activation in rainbow trout brain.

A number of studies have shown that the induction of c-fos gene is an indicator of the responses of cells and tissues to the environmental stimuli. In the present study, using RT-PCR-based strategy, we isolated, from the brain of the rainbow trout, two partial cDNA clones (RT-fos1 and RT-fos2) that code proteins homologous to c-Fos proteins of higher vertebrates. Sequence analysis of the two clones indicated that the two rainbow trout clones are very similar to each other over the entire cloned region (88% amino acid identity) and showed moderate similarity to c-Fos proteins of higher vertebrates (40% amino acid identity with human c-Fos). Two functionally important domains (i.e. the leucine zipper and zinc finger) are highly conserved among all the vertebrate species analyzed, although the region between the two domains was highly variable between different species. Intraperitoneal administration of kainic acid, a stable agonist of glutamate receptors, transiently induced the mRNAs for both RT-fos1 and RT-fos2 in the rainbow trout brain. These results indicate that the expression pattern of the two clones can be utilized as a suitable anatomical marker for the increased neural activities in salmonid fish brain to investigate the higher order behavior such as the learning and imprinting of odors of the home river.

Cladinose analogues of sixteen-membered macrolide antibiotics. IV. Improved therapeutic effects of 4-O-acyl-L-cladinose analogues of sixteen-membered macrolide antibiotics.

Six derivatives of sixteen-membered macrolides possessing 4-O-acyl-alpha-L-cladinose as a neutral sugar were synthesized via 3"-methylthiomethyl ether intermediates in reasonable yield. Introduction of a methyl group on the 3"-hydroxyl group of midecamycin A1 was effective for enhancing its antibacterial activity. All these derivatives exhibited excellent therapeutic effects in mice, and some of them showed improved pharmacokinetics compared with the natural antibiotics (mycarose type) in mice. Facile synthesis of 9-O-acylated analogues are also described.

[Three cases of typical cryptomeria pollen asthma].

We treated three asthmatic children diagnosed as typical cryptomeria pollen asthma (Cryptomeria japonica). Wheezing started from one to four years after onset of pollinosis. Asthmatic attacks occurred one or several days after pollen counts increased. We could find asthmatic attacks in the years when the pollen counts were higher than in average years. In the laboratory findings, only specific IgE antibody to cryptomeria with its high titer of over 17.5 PRU/ml was detected in these three cases. Allergen inhalation tests were performed with positive results in the two cases, suggesting that these three cases were typical child cryptomeria pollen asthma.

Taste transduction mechanism: similar effects of various modifications of gustatory receptors on neural responses to chemical and electrical stimulation in the frog.

Responses in the frog glossopharyngeal nerve induced by electrical stimulation of the tongue were compared with those induced by chemical stimuli under various conditions. (a) Anodal stimulation induced much larger responses than cathodal stimulation, and anodal stimulation of the tongue adapted to 5 mM MgCl2 produced much larger responses than stimulation with the tongue adapted to 10 mM NaCl at equal current intensities, as chemical stimulation with MgCl2 produced much larger responses than stimulation with NaCl at equal concentration. (b) The enhansive and suppressive effects of 8-anilino-1-naphthalenesulfonate, NiCl2, and uranyl acetate on the responses to anodal current were similar to those on the responses to chemical stimulation. (c) Anodal stimulation of the tongue adapted to 50 mM CaCl2 resulted in a large response, whereas application of 1 M CaCl2 to the tongue adapted to 50 mM CaCl2 produced only a small response. This, together with theoretical considerations, suggested that the accumulation of salts on the tongue surface is not the cause of the generation of the response to anodal current. (d) Cathodal current suppressed the responses induced by 1 mM CaCl2, 0.3 M ethanol, and distilled water. (e) The addition of EGTA or Ca-channel blockers (CdCl2 and verapamil) to the perfusing solution of the lingual artery reversibly suppressed both the responses to chemical stimulus (NaCl) and to anodal current with 10 mM NaCl. (f) We assume from the results obtained that electrical current from the microvillus membrane of a taste cell to the synaptic area supplied by anodal stimulation or induced by chemical stimulation activates the voltage-dependent Ca channel at the synaptic area.

Selective modification of positive chemotaxis in the true slime mold Physarum polycephalum by ethylenediaminetetraacetic acid treatment.

The plasmodium of the true slime mold Physarum polycephalum was treated with EDTA or EGTA and the effect of the treatment on the chemotactic response was examined by measuring the chemotactic motive force with the double-chamber method. The results obtained were as follows: (1) The treatment of the plasmodium with 5 mM EDTA (pH 7.0, 20 min) did not give any significant effect on the protoplasmic streaming or motility. (2) The plasmodium treated with EDTA exhibited no chemotactic response to non-electrolyte attractants (D-glucose, D-galactose, D-mannose, and maltose) and negative chemotaxis to electrolyte attractants (cyclic AMP and NaH2PO4). (3) The EDTA treatment gave no effect on the chemotactic response to repellents (D-fructose, NaCl, CaCl2, MgCl2). (4) The EDTA-treated plasmodium exhibited changes in the membrane potential in response to both attractants and repellents as similar to the untreated plasmodium. (5) The treatment of the plasmodium with 5 mM EGTA (pH 7.0, 20 min) gave results similar to those obtained with the EDTA treatment. The results obtained suggested that EDTA (or EGTA) treatment did not affect the receptor sites but modified the transduction mechanism from reception into tactic movement.