The role of phospholipase in beta-agonist-induced down regulation in guinea pig lungs

It has been observed that repeated and prolonged beta-agonist treatment causes the impairment of beta-adrenergic function, so-called "desensitization" or "down regulation". To clarify the mechanism of down regulation, the following experiment was performed using guinea pig lungs. Animals were divided into four groups: In the metaproterenol groups, guinea pigs were treated with metaproterenol (10 mg/kg/day) by intraperitoneal injection once a day for 1 day or for 7 successive days In the control groups, guinea pigs were treated with saline by the same procedure as in the metaproterenol groups. In the group treated with metaproterenol for 7 days, there was a 45% reduction in the number of beta-adrenoceptors and a 62% reduction in adenylate cyclase activity, compared with those of the control group. However, there were no significant changes in the dissociation constant (Kd) of the receptors. On the other hand, no reduction in the number of beta-adrenoceptors and adenylate cyclase activity was observed in the group treated with metaproterenol once a day for 1 day, compared with those of the control group. Phospholipase (PLase) activity in the lung microsomes of guinea pigs injected with metaproterenol for 1 day and for 7 days was elevated by 14.4 and 33.1%, respectively, compared with that of the control groups. Phospholipid contents of lung membranes prepared from the animals treated with metaproterenol for 7 days were significantly decreased compared with those of the control group, though in the group treated with metaproterenol once a day for 1 day, phospholipid contents did not differ from those of the control. Lung membranes treated with PLase A2 revealed decreases both in the number of beta-adrenoceptors and adenylate cyclase activity, dose dependently. These results and the fact that membrane phospholipids are involved in the beta-adrenoceptor system suggest that down regulation observed during beta-agonist administration is, at least in part, attributed to degradation of phospholipids of lung membranes by the persistent activation of PLase in the tissue.     

Effect of cytokines on Japanese encephalitis virus production by human monocytes

Japanese encephalitis (JE) virus was shown to grow in in vitro cultures of human monocytes. Interferon (IFN)-alpha and IFN-gamma inhibited JE virus production by the infected monocytes in the absence of anti-JE virus antibody, but interleukin (IL)-1 alpha, IL-2, IL-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte-CSF (G-CSF), and tumor necrosis factor (TNF)-alpha did not show a significant inhibition. Antibody against JE virus increased the JE virus production by the infected monocytes probably by enhanced uptake of virus-antibody complexes via Fc receptors. IFN-gamma and GM-CSF increased JE virus production by monocytes in the presence of anti-JE virus antibody, whereas IFN-alpha inhibited JE virus production even in the presence of the antibody. The other 5 cytokines (IL-1 alpha, IL-2, IL-3, G-CSF, and TNF-alpha) did not show a significant effect on JE virus production by monocytes in the presence or absence of the antibody.     

Lobeline production by hairy root culture of Lobelia inflata L.

Hairy roots were obtained following inoculation of the stems of Lobelia inflata L. with Agrobacterium rhizogenes strain ATCC 15834. These hairy roots contained agropine and mannopine. In addition, lobeline was detected by HPLC and confirmed by mass spectrometry. Various media were tested for the growth of hairy roots as well as for the content of lobeline in hairy roots. The growth rate of hairy roots cultured in Nitsch and Nitsch's medium was approximately one third of those cultured in other media. The lobeline content of hairy roots (18–54 g/g dry weight) cultured in these media was the same order of magnitude compared with that of roots of L. inflata (24 g/g dry weight) cultivated in pots. The hairy roots cultured in Nitsch and Nitsch's medium were morphologically different from those cultured in other media.Abbreviations MS medium Murashige and Skoog's medium - 1/2 MS medium one-half strength of the standard Murashige and Skoog's medium - B5 medium Gamborg's B5 medium - NN medium Nitsch and Nitsch's medium - FW fresh weight - DW dry weight     

Monitoring Primary Response to Chilling Stress in Etiolated Vigna radiata and V. mungo Seedlings Using Thermal Hysteresis of Water Proton NMR Relaxation Times

Thermal hysteresis of longitudinal relaxation times (T₁) ofwater protons in hypocotyls of etiolated Vigna radiata and V.mungo seedlings was investigated by pulse nuclear magnetic resonance(NMR) spectroscopy. Various lengths of chilling exposures duringa cool-warm cycle between 20 and 0?C (below 10?C, about 4 h)for the T₁ hysteresis measurement did not cause any visibleinjury symptoms in hypocotyls. However, the profiles of T₁ hysteresisvaried as a result of different chilling exposures. The sumsof the T₁ ratio (for detail see Introduction) reflecting T₁prolongation or shortening upon the warming process were a goodquantitative index for the extent of T₁ hysteresis, and thewide dispersion of this value ranging on the "minus" side (T₁prolongation upon warming) suggested the occurrence of a primaryresponse of cells to chilling stress before obvious visiblesymptoms occur while the T₁ ratio sums on the "plus" side (T₁shortening upon warming) corresponded to a response of seriousvisible injury. Therefore, the sums of the T₁ ratio can be usedas a non-destructive diagnostic tool for monitoring the primaryevent of chilling injury when lacking any visible injury symptoms.The data indicate that the critical temperature for the occurrenceof primary response for chilling stress was around 7.5?C forV. radiata and 12.5?C for V. mungo. (Received February 1, 1988; Accepted June 1, 1988)     

Immunochemical and histochemical studies on a phosphonoglycosphingolipid, SGL-II, isolated from the sea gastropod Aplysia kurodai

Antiserum was raised against 3-O-MeGal beta 1----3GalNAc alpha 1----3[6' -O-(2-aminoethylphosphonyl)Gal alpha 1----2 (2-aminoethylphosphonyl----6)Gal beta 1----4Glc beta 1----1Ceramide (SGL-II) isolated from the skin of a mollusc, Aplysia kurodai. This antiserum reacted with SGL-II and other phosphonoglycosphingolipids of Aplysia such as SGL-I', F-21, and some minor glycolipids on TLC plates, but it did not react with ganglioside or globoside. The sugars recognized were 3-O-methylgalactose at the non-reducing end and galactose at the branched chain of the glycolipids. One membrane glycoprotein (Mr 280,000) reacted strongly, and some other proteins reacted weakly with the antiserum. Immunohistochemical examination of the nervous tissues revealed distinct staining in the periganglionic tissue of the ganglia, and the perineural sheath of the proximal portion of the peripheral nerves. The neuropil and satellite cells were also stained. In the skin, subcutaneous connective tissues were moderately stained, and the cytoplasm of small mononuclear cells and foamy cells was also stained. The staining patterns were essentially the same in paraffin and cryostat sections. From the findings with sections pretreated with chloroform-methanol (2 : 1, v/v), it was suggested that the periganglionic and perineural stainings were due to glycoproteins, including an SDS-soluble glycoprotein of Mr 280,000, while those of the other regions were due to SGL-II and glycolipids immunologically related to SGL-II. The stainings in the skin sections were largely due to glycoproteins.     

Selective modification of myosin SH1 with 1,2,4-trinitrobenzene. VIII. Thiols of myosin

Myosin has 2 mol of the most reactive thiol, named SH1. 1,2,4-Trinitrobenzene (TNB), a novel dinitrophenyl(DNP)ating reagent [Takahashi et al. (1983) Chem. Lett. 1445-1448], was found to react only with SH1 without any other amino acid residues in myosin under the conditions used. Its reaction with myosin SH1 was about 30 times faster than that with N-acetylcysteine (NAC). The reaction rate of TNB with SH1 was about twice compared with that of NEM, the most reactive selective reagent for SH1 so far found, although its rate with NAC was only one sixtieth that of NEM. As to the lambda max of the absorption spectrum of SH1-DNP-myosin, a large red shift of as much as 20 nm was observed compared with low molecular S-DNP derivatives. This red shift disappeared in 8 M urea. This outstanding feature of SH1 modification with TNB was discussed in terms of affinity labeling by interaction with an aromatic amino acid near SH1.