Calcium mediates the interconversion between two states of the liver inositol 1,4,5-trisphosphate receptor

D-myo-Inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) regulates intracellular Ca2+ by mobilizing Ca2+ from a non-mitochondrial store. We have investigated the effects of Ca2+ on the binding of [32P]Ins (1,4,5)P3 to permeabilized rat hepatocytes and a liver plasma membrane-enriched fraction. Increasing the free Ca2+ concentration in the medium from 0.1 nM to 0.7 microM increased the capacity of a high affinity binding component (KD = 2-3 nM) in permeabilized cells by a factor of 10. If the membrane fraction was preincubated at 37 degrees C before binding was measured at 4 degrees C, all of the Ins(1,4,5)P3 receptors were transformed to a low affinity state (KD = 65 +/- 12 nM, Bmax = 3.1 +/- 0.1 fmol/mg, n = 4). When 0.7 microM of Ca2+ was added, the receptors were totally transformed to a high affinity state (KD = 2.8 +/- 0.4 nM, Bmax = 2.7 +/- 0.4 fmol/mg, n = 4). The EC50 of the Ca2(+)-induced interconversion of the Ins(1,4,5)P3 receptor was 140 nM. This Ca2(+)-induced transformation of the Ins(1,4,5)P3 receptor from a low affinity to a high affinity state was associated with an inhibition of the Ins(1,4,5)P3-induced Ca2+ release in permeabilized hepatocytes. These data suggest that the Ins(1,4,5)P3-dependent hormones, by increasing the intracellular Ca2+ concentration, induce a reversible transformation of the receptor from its low affinity state, coupled to the Ca2+ release, to a desensitized high affinity state. Transformation of the receptor may play a role in the oscillatory release of Ca2+ observed in single isolated hepatocytes.     

Chemo- and karyotaxonomic studies on some rhizomatous species of the genusSymphytum (Boraginaceae)

InS. tuberosum subspp.tuberosum andnodosum, S. grandiflorum andS. ibericum the presence of the pyrrolizidine alkaloids lycopsamine, echimidine and symphytine could be demonstrated. The taxonS. tuberosum contains an unknown compound that seems to be specific for this taxon. This compound is not the pyrrolizidine alkaloid anadoline which has previously been reported for this species. It is possibly represented by a peak on GC/MS with a molecular ion peak at m/z 623 (as TMS derivative) and can be used as a chemotaxonomic marker for the speciesS. tuberosum. The pyrrolizidine alkaloid pattern of the two subspecies ofS. tuberosum reinforces the close relationship. Fresh material ofS. tuberosum contained the triterpene isobauerenol, but in herbarium material isobauerenol was lacking. InS. grandiflorum, neither fresh nor dried material contains isobauerenol. In herbarium material ofS. ibericum also no isobauerenol could be found. More extensive chemotaxonomical research is necessary to support the view thatS. abchasicum is more closely related toS. ibericum than toS. grandiflorum.     

Purification and partial characterization of an iron regulated outer membrane protein of B. fragilis under non-denaturing conditions

The CHAPS-PAGE gelsystem we applied gave a good separation of the proteins of Bacteroides fragilis under non-denaturing conditions. We succeeded with preparative CHAPS-PAGE in purifying an iron regulated outer membrane protein (a 44 kDa polypeptide on SDS-PAGE) of B. fragilis. This integral membrane protein proved to be a lipopolysaccharide binding protein with an isoelectric point of approximately pH 5.5. This method of purifying membrane proteins could be an important step in research into the function of membrane proteins.     

The indices of antipneumococcal immunity in children with chronic pneumonia

In 76 children with chronic pneumonia the levels of serum antibodies to pneumococcal capsular polysaccharide antigens (serotypes 1, 3, 6B, 8, 9N, 15F, 23F), O-polysaccharide and pneumococcal protein somatic antigens were determined by ELISA techniques. The study showed that in sick children the content of antipneumococcal antibodies in the blood increased with age. No correlation between the content of total immunoglobulins and that of antipneumococcal antibodies in the blood of the patients was established, but a sharp decrease in the concentration of antibodies was registered in a child with hypoglobulinemia. No increase in the level of antibodies to pneumococcal antigens was observed in cases of the exacerbation of Pneumococcus-induced inflammatory process in the lungs.     

The quantitative analysis of lipopolysaccharide antigen in the blood serum of patients with salmonellosis by using complement-bound liposome lysis]

Titration of group B Salmonella O-antigen in the blood sera of patients and donors was carried out by means of the complement-dependent lysis of liposomes sensitized with S. typhimurium LPS. Good correlation (r = 0.95) of the levels of S. typhimurium somatic O-antigen in the patients' sera determined by liposomal immunoassay and aggregate hemagglutination test was established. The concentration of the antigens in the tested samples was within 0.5-50 micrograms/ml. Statistical analysis of the results obtained by liposomal immunoassay techniques demonstrated differences in the distribution functions for the blood sera of patients with different diseases and of donors.     

Effect of mastectomy versus mastectomy plus adjuvant ovariectomy on prolactin response to TRH in breast cancer

The endocrine effects of ovariectomy need to be further investigated. The present study was carried out to evaluate the influence of the adjuvant ovariectomy on the mastectomy-induced changes in PRL response to TRH in breast cancer. The study included 34 patients with locally limited breast carcinoma, 18 of whom were treated with radical mastectomy, whereas the other 16 underwent mastectomy plus adjuvant ovariectomy. PRL secretion in response to TRH (200 mcg I.V. as bolus) was evaluated one day before and 7 days after surgery. In patients treated with mastectomy only, PRL increase after TRH was significantly higher after surgery than before. On the contrary, no difference was seen in patients treated with mastectomy plus ovariectomy. This study shows that the adjuvant ovariectomy may block the increase in PRL response to TRH induced by mastectomy in breast cancer.     

Determination of the amounts of the protein synthesis initiation and elongation factors in wheat germ

Previous work by Browning et al. (Browning, K. S., Lax, S. R., Humphreys, J., Ravel, J. M., Jobling, S. A., and Gehrke, L. (1988) J. Biol. Chem. 263, 9630-9634) indicated that wheat germ extracts do not contain sufficient amounts of some of the protein synthesis initiation factors to obtain optimal translation of all mRNAs. In this investigation, a quantitative enzyme-linked immunosorbent assay was used to determine the amounts of eukaryotic initiation factors (eIF) 2, 3, 4A, 4F, and (iso)4F as well as the amounts of 40 S ribosomal subunits and elongation factors (EF) 1 alpha and 2 present in wheat germ extracts. EF-1 alpha is present in the highest amount (approximately 5% of the total protein), and eIF-4F is present in the lowest amount (approximately 0.03% of the total protein). The micromolar amounts of the factors and ribosomes are as follows: EF-1 alpha, 34; EF-2, 5.2; eIF-2, 1.5; eIF-3, 0.7; eIF-4A, 3.0, eIF-4F, 0.09; eIF-(iso)4F, 0.8; and 40 S ribosomal subunits, 3.2. The molar ratios of the factors to 40 S ribosomal subunits are approximately 11:1 for EF-1 alpha, 1.6:1 for EF-2, 0.45:1 for eIF-2, 0.2:1 for eIF-3, 0.9:1 for eIF-4A, 0.03:1 for eIF-4F, and 0.25:1 for eIF-(iso)4F. These findings strongly suggest that the concentrations of the initiation factors, particularly those factors required for the binding of mRNA to ribosomes, may play a major role in regulating the translation of mRNAs within the cell.