Effects of progesterone and estradiol on the proliferation of phytohemagglutinin-stimulated human lymphocytes

In this paper we report on a study to elucidate whether the response of human lymphocytes to mitogenic stimulation was modified by physiological changes which occur during the menstrual cycle. Experiments with untreated cultures showed intra-individual variation to mitogen stimulation in female lymphocyte cultures, but a significant correlation between the menstrual cycle and the proliferation kinetics of lymphocytes was not found. Consequently, we performed experiments in which two of the hormones that regulate the menstrual cycle in women, estradiol and progesterone, were added to cultured human lymphocytes obtained from both men and women. The results indicate that both hormones at physiological concentrations have the capacity to modify the proliferation of PHA-stimulated human lymphocytes. Therefore, both hormones could play a role in the induction of the intra-individual variation observed in the untreated female cultures. However, in vivo other factors could also modify the proliferation kinetics of human lymphocytes preventing the demonstration of the effects of a single factor, such as the hormonal changes occurring during the menstrual cycle.     

Living immobilized Acetobacter in Ca-alginate in vinegar production: Perliminary study on optimum conditions for immobilization

Summary The optimum conditions forAcetobacter immobilization were investigated. The results show that: 1) the maximum oxygen uptake rate (OURm) and cell release are related to alginate and cell concentration in the gel; 2)different alginate concentration does not affect cell viability, but long storage in CaCl₂ reduces the number of living cells; 3)the double alginate gel layers had no influence on cell viability and on the OURm and prevented cell leakage from the gel matrix.     

Insulin and progesterone increase 32PO4-labeling of phospholipids and inositol 1,4,5-trisphosphate mass in Xenopus oocytes.

After a 4-6 h induction period, insulin or progesterone induces Xenopus oocytes to enter prophase of meiosis. During the period of induction, both insulin and progesterone induced an increase in 32PO4 labeling of phosphatidylcholine and phosphatidylinositol. Through a mass assay, we found that insulin and progesterone increase inositol 1,4,5-trisphosphate (IP3) at about 15-30 s, 15 min and at about 2-3 h (0.5 GVBD50) after hormone addition. Since IP3 increases were small (from a basal of 66 to 104 nM), the results agree with prior conclusions that progesterone does not induce a large, cytosolic calcium elevation. Insulin is probably acting through the insulin-like growth factor-1 receptor as insulin concentrations greater than about 50 nM are required to increase IP3.     

Trehalase activity and its regulation during growth of Saccharomyces cerevisiae.

Trehalase activity decreased in 95% at the onset of the transition phase of growth of S. cerevisiae. The question which we raised was whether this phenomenon was due to proteolysis or to conversion of the enzyme to a less active form (dephosphorylation). Immunological methods allowed to identify the presence of the trehalase protein during cell growth. At the same stage of growth, an increase in the non-phosphorylated enzyme was detected "in vitro". Results utilizing mutant strains also indicated that regulation occurred by interconversion of forms. The same mechanism also seems to control trehalase activity in non proliferating conditions.     

Influence of dilution rate and induction of cloned gene expression in continuous fermentations of recombinant yeast

The effects of growth rate on cloned gene product synthesis in recombinant Saccharomyces cerevisiae have been studied in continuous culture. The plasmid employed contains a yeast GAL10-CYC1 hybrid promoter directing expression of the E. coli lacZ gene. beta-Galactosidase production was therefore controlled by the yeast galactose regulatory circuit, and the induction process and its effects were studied at the various dilution rates. At all dilution rates plasmid stability decreased with induction of lacZ gene expression. In some instances, two induced "steady states" were observed, the first 10-15 residence times after induction and the second after 40-50 residence times. The second induced steady state was characterized by greater biomass concentration and lower beta-galactosidase specific activity relative to the first induced "steady-state." beta-Galactosidase specific activity and biomass concentration increased as dilution rate was reduced, and despite lower flow rate and plasmid stability, overall productivity (activity/L/hr) was substantially higher at low dilution rate. Important factors influencing all of the trends were the glucose and galactose (inducer) concentrations in the vessel and inducer metabolism.     

Role of tissue hypoxia as the mechanism of lactic acidosis during E. coli endotoxemia.

We compared the hemodynamic and metabolic alterations produced in rabbits by similar decreases in cardiac output created by inflating a balloon placed in the right ventricle (n = 6) with those produced by an intravenous bolus of Escherichia coli lipopolysaccharide (LPS; SEP group; n = 6). We measured O2 consumption (VO2), O2 transport (TO2), and O2 extraction ratio (ERO2) for the whole animal and also for the left hindlimb. Both groups experienced similar decreases in cardiac output, systemic TO2, and VO2 and similar increases in ERO2. For the hindlimb, TO2 was similar, but VO2 and ERO2 were lower for the SEP group 30 min after LPS administration (P less than 0.05); however, this difference disappeared during the remainder of the experiment. Arterial lactate concentration was greater (P less than 0.05) for the SEP group. There were no differences in skeletal muscle PO2, measured with a multiwire surface electrode, or in cardiac and skeletal muscle concentrations of high-energy phosphates. We hypothesize that a direct effect of LPS on cellular metabolism may have resulted in greater arterial lactate concentration for the SEP group.     

Association of neuronal pp60c-src with growth cone glycoproteins of rat brain

Tyrosine phosphorylation and protein tyrosine kinase (PTK) activity in the growth cone membrane-associated glycoprotein (GCGP) fraction of 1-day-old rat brain were examined. Using immunoblotting and immunoprecipitation techniques, pp60^c-src was identified as one of the major PTKs associated with GCGPs. Furthermore, only GCGP-associated src that was also tyrosine phosphorylated was active. Immunoprecipitation experiments using various src antibodies revealed that pp60^c-src contributed partially to the PTK activity detected in GCGPs, and that it is associated with several proteins of Mr 140 K, 120 K, 85 K and 50 K. This association of src protein with GCGPs was specific, and another src family member p59^fyn, which is also abundant in the brain, did not exhibit such an association. In addition to pp60^c-src, the GCGP fraction contained several major phosphotryosine-containing proteins of Mr 140 K, and a 97/90 K doublet that corresponded to the beta subunits of IGF-I/ insulin receptors. These studies show that pp60^c-src associated with GCGPs is an active PTK that could be involved in neuronal growth and development, transmembrane signalling, and in recognition and/or adhesive events. © 1992 John Wiley & Sons, Inc.     

Proteinases and amylases of larval midgut of Zabrotes subfasciatus reared on cowpea (Vigna unguiculata) seeds

Proteinase and amylase activities in larval midguts of the bruchid beetle Zabrotes subfasciatus (Boh.) (Coleoptera: Bruchidae) reared on cowpea (Vigna unguiculata (L.) Walp.) seeds were investigated. We could detect and isolate a proteolytic activity with a pH optimum of 5.5 (on azo-casein as substrate) which was activated by thiol reagents and inhibited by several compounds reactive against-SH groups. None of the plant protein inhibitors of serine proteinases utilized were effective inhibitors of this activity. This activity has characteristics of a cysteine class proteinase. We could also detect and isolate a proteolytic activity with a pH optimum of 3.5 (on hemoglobin as substrate) which was not influenced by activators or inhibitors of cysteine, serine, or metalloproteinases. This activity was totally inhibited by pepstatin, a specific inhibitor of aspartic proteinases. We conclude that this activity is due to an aspartic class proteinase. We found also that the aspartic class proteolytic activity is higher than the cysteine class proteinase activity in the midguts of Z. subfasciatus. This seems to be contrary to what is found in Callosobruchus maculatus (F.) larvae midguts. An amylolytic activity with the charateristics of an -amylase was also detected and isolated.

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Résumé Les activités protéinase et amylase ont été étudiées sur l'intestin moyen de larves de Zabrotes subfasciatus Boh. (Coléo, Bruchidae), élevées sur graines de Vigna unguiculata Walp. Nous avons pu déceler et isoler une activité protéolytique optimale à pH 5,5 (sur substrat d'azo-caséine) activée par des réactifs thiol et inhibée par plusieurs composés réagissant aux groupements SH. Aucun inhibiteur végétal des sérine-protéases utilisé n'a inibé efficacement cette activité qui présente les caractéristiques des protéines de la famille des cystéines. Nous avons pu déceler et isoler aussi une activité protéolytique optimale au pH 3,5 (sur hémoglobine comme substrat) qui n'était pas modifiée par les activateurs ou les inhibiteurs de cystéine, de sérine ou de métalloprotéinases. Cette activité était totalement inhibée par la pepstatine, inhibiteur spécifique des protéinases aspartiques. Nous en concluons que l'activité est due à une protéinase de la famille aspartique. Nous avons trouvé aussi que l'activité protéolytique de la famille aspartique était supérieure à l'activité protéinase de la famille cystéine dans l'intestin moyen de Z. subfasciatus. Ceci semble l'inverse de ce qui a été observé dans l'intestin moyen des larves de Callosobruchus maculatus F. (C.P. Silva & al, in litt.). Une activité amylolytique ayant les caractéristiques d'une -amylase a aussi été décelée.

     

Metabolism of retinoic acid in vivo in the vitamin A-deficient rat.

Sample preparation and high-pressure liquid-chromatography separation methods useful for the study of retinoic acid metabolism are reported. The sample preparation procedure does not cause significant degradation of retinoic acid, and the gradient high-pressure liquid-chromatography separation method gives excellent separation of the major metabolites of retinoic acid. These methods were used to examine the metabolites of retinoic acid in blood, trachea and lung, testes, kidneys and small intestine of vitamin A-deficient rats dosed subcutaneously with 2 micrograms of [11,12-3H] retinoic acid. At 6h after dosing, a total of eight metabolites of retinoic acid produced in vivo were found in the tissues examined. Of these, four were found in most of the epithelial tissues examined, and therefore may be of interest as possible active metabolites in the epithelial functions of vitamin A.