Possible regulation of the growth of thyroid tissue by plasma iodide

Rats on iodine deficient diet for 6 months received propylthiouracil (PTU) (0.15%) during the last 2 months. At the end of this treatment, PTU was withdrawn and the rats were iodine refed. 48 hrs. before the iodine refeeding all rats were injected with 3H thymidine. The results showed that some prelabelled cells in the hyperplastic goitre preferentially disappeared during its involution and therefore are more sensitive to iodine.     

Spectrin αIIa variant in dominant and non-dominant spherocytosis

Several polymorphic mutations are located on the spectrin -chain; among these the variant termed IIa is characterized by an acid shift in the isoelectric point of the tryptic digest peptides 46 kDa and 35 kDa. In this variant a single amino acid substitution (alanine to aspartic acid) occured at position 972 of the spectrin -chain due to a point mutation (GCT to GAT) in the DNA. This variant, which seemed very rare in normal people, could be related to the recessive form of hereditary spherocytosis (HS) and could be absent in the dominant form of the disease. We have studied the IIa variant by denaturing electrophoresis of the spectrin tryptic digest peptides from 179 subjects: 46 controls, 78 patients with dominant (d) or non dominant (nd) HS and 55 relatives of the patients. The confirmation of the results was obtained at the DNA level in 41 subjects. The frequency of the chromosome bearing the IIa mutation was 7.6% in controls and higher (about 12–14%) in members of families with dHS as well ndHS. However, the family trees clearly showed that the mutation and the HS disease gene(s) were located on different chromosomes and inherited independently from each other. Furthermore, our study allows the conclusion that in most (if not all) cases of dHS, the IIa the variant is not the cause, is not a marker, and does not influence the phenotypic expression of the disease.     

Inability of the estrogen-induced uterine protein to serve as a substrate for the endogenous phosphokinase(s).

We have studied the phosphorylation of soluble proteins from uterine extracts by an endogenous protein kinase. The analysis of phosphorylation patterns by polyacrylamide gel electrophoresis did not reveal any significant difference in this respect between the soluble proteins from control or 17-beta-estradiol stimulated uteri. In both cases, three main components with mol. wt of about 120,000, 60,000 and 45,000 appear preferentially phosphorylated. Estrogen-induced protein did not coincide with any phosphorylated component, although some migrated very closely to it. This was observed whether phosphorylation was performed on uterine extract incubated with [gamma-3 2P]ATP or on intact organs incubated in the presence of 3 2Pi. We conclude that whatever the role of estrogen-induced protein, it is unlikely to be subjected to regulation through the phosphorylation process.     

Isolation of microvillus plasma membranes from suckling-rat intestine. The influence of premature induction of digestive enzymes by injection of cortisol acetate

1. Cortisone administration to suckling rats leads prematurely to induction of enzymes of the intestinal microvillus plasma membrane and lengthening of the intestinal microvilli. To investigate the membrane changes that might be involved, a method for the isolation of a fraction enriched with microvillus plasma membrane was developed in suckling rats. Plasma-membrane fractions were compared from 13-day-old control rats and from 13-day-old rats given cortisol acetate by subcutaneous injection for 3 days. 2. After cortisol injection, the activity of maltase, trehalase, sucrase and leucyl beta-naphthylamidase increased markedly, and to the same extent, in intestinal homogenates and plasma-membrane preparations. Purification, and recovery of five marker enzymes with respect to homogenate activity, and recovery of protein, were similar for both membrane preparations, particularly after correction for non-membrane activity, which was high in suckling rats and affected by cortisol. 3. In material released from the plasma membrane by digestion with papain, maltase protein was increased after cortisol injection at least as much as maltase activity. Sucrase activity increased at least 200-fold, and this increase was associated with the appearance of a new sucrase band on polyacrylamide-gel electrophoresis. 4. Sodium dodecyl sulphate electrophoresis of plasma-membrane proteins revealed at least four additional macromolecules after cortisol injection. Concurrently several proteins disappeared from the plasma membrane. The added proteins appeared in the main to be removed from the plasma membrane by papain, whereas the deleted proteins were in the papain-resistant fraction. 5. Enzymic stimulation induced by cortisol acetate in the suckling-rat plasma membrane therefore appears to involve the addition of new proteins, rather than activation of proteins in situ. Deletion of proteins from the membrane during induction of hydrolytic enzymes may reflect other phenomena such as protein reorganization associated with the change in microvillus shape.     

In vitro stimulation of prostaglandin synthesis in the rat pancreas by carbamylcholine, caerulein and secretin

Rat pancreas pieces spontaneously released PGE₂ (2.3 ng/100 mg × 45 min) and PGF_2α (7.6 ng/100 mg × 45 min). This release corresponds probably to a neo-synthesis since it was abolished by indomethacin. Carbamylcholine (≥ 10 μM), caerulein (≥ 10 nM) and secretin (≥ 10 nM) stimulated the release of PGE₂ and PGF_2α : the concentrations of stimulators required to increase PGs release were thus much higher than those which trigger enzyme secretion. Atropine specifically inhibited the cholinergic stimulation, whereas indomethacin blocked the stimulatory effects of all secretagogues. Stimulation of PGE₂ and PGF_2α release was reduced in a Ca⁺⁺-free medium, abolished by EGTA and mimicked by the ionophore A23187, underscoring the crucial role of Ca⁺⁺ in the regulation of PGs synthesis by the pancreas. Neither PGE₂ nor PGF_2α stimulated enzyme secretion in this system and indomethacin did not inhibit the secretory effect of carbamylcholine. Increased synthesis of prostaglandins in response to pancreatic secretagogues does not appear to be involved in the process of enzyme secretion.     

The influence of hydrocortisone on the synthesis and turnover of microvillous membrane glycoproteins in suckling rat intestine.

Synthesis and degradation of intestinal mucosal and microvillous membrane glycoproteins were studied in control suckling rats, and suckling rats given cortisol acetate by intraperitoneal injection for 3 days. Cortisol acetate had no effect on total uptake of radioactive glucosamine by the protein free compartment of rat intestine. Early incorporation of [1(-14)C]glucosamine by intestinal glycoproteins was enhanced by cortisol, but stimulation was the same in membrane and homogenate fractions. Polyacrylamide gel electrophoresis of membrane proteins solubilized with 2% sodium dodecyl sulphate demonstrated a cortisol dependent change, characterized by loss of faster travelling glycoproteins, and a corresponding shift in maximum labelling at 3 h from these glycoproteins to more slowly migrating glycoproteins. Degradation was studied qualitatively with a double isotope technique. Glycoprotein degradative rates appeared to be stimulated by cortisol, but similarly in membrane and total homogenate fractions. On polyacrylamide gels, the areas occupied by glycoproteins with the highest apparent degradative rates, corresponded closely with the areas of most active labelling at 3 h. The rate of degradation in the most actively labelled zone appeared to be higher after cortisol than in the controls. The results indicate that cortisol does not alter membrane composition by inhibiting degradation of selected glycoproteins, and are consistent with a model in which cortisol stimulates the synthesis of specific membrane glycoproteins in suckling rats, while inhibiting synthesis of other glycoproteins.     

Distribution of greenhouse gases in hyper-arid and arid areas of northern Chile and the contribution of the high altitude wetland microbiome (Salar de Huasco,Chile)

Northern Chile harbors different bioclimatic zones including hyper-arid and arid ecosystems and hotspots of microbial life, such as high altitude wetlands, which may contribute differentially to greenhouse gases (GHG) such as carbon dioxide (CO₂), methane (CH₄) and nitrous oxide (N₂O). In this study, we explored ground level GHG distribution and the potential role of a wetland situated at 3800 m.a.s.l, and characterized by high solar radiation <?1600 W m^?2, extreme temperature ranges (?12 to 24 °C) and wind stress (<?17 m s^?1). The water source of the wetland is mainly groundwater springs, which generates streams and ponds surrounded by peatlands. These sites support a rich microbial aquatic life including diverse bacteria and archaea communities, which transiently form more complex structures, such as microbial mats. In this study, GHG were measured in the water and above ground level air at the wetland site and along an elevation gradient in different bioclimatic areas from arid to hyper-arid zones. The microbiome from the water and sediments was described by high-throughput sequencing 16S rRNA and rDNA genes. The results indicate that GHG at ground level were variable along the elevation gradient potentially associated with different bioclimatic zones, reaching high values at the high Andean steppe and variable but lower values in the Atacama Desert and at the wetland. The water areas of the wetland presented high concentrations of CH₄ and CO₂, particularly at the spring areas and in air bubbles below microbial mats. The microbial community was rich (>?40 phyla), including archaea and bacteria potentially active in the different matrices studied (water, sediments and mats). Functional microbial groups associated with GHG recycling were detected at low frequency, i.e., <?2.5% of total sequences. Our results indicate that hyper-arid and arid areas of northern Chile are sites of GHG exchange associated with various bioclimatic zones and particularly in aquatic areas of the wetland where this ecosystem could represent a net sink of N₂O and a source for CH₄ and CO₂.     

Comparison of methods based on annexin-V binding, DNA content or TUNEL for evaluating cell death in HL-60 and adherent MCF-7 cells

HL-60 and MCF-7 cells were treated with 0.15 μ M camptothecin (CPT) or with the solvent dimethylsulfoxide (DMSO) for the controls, for 2, 3 and 4 h or for 24, 48 and 72 h, respectively. The apoptotic index (AI) was then evaluated in parallel by the following flow cytometric methods: (1) double staining of unfixed cells with fluoresceinated annexin V and propidium iodide (PI), this after detachment by trypsinization in the case of MCF-7 cultures; (2) prefixation in 70% ethanol, extraction of degraded, low molecular weight DNA with 0.2 M phosphatecitrate buffer and analysis of the DNA content stained with PI; (3) TUNEL, i.e. labelling of DNA strand breaks with biotin-dUTP, followed by staining with streptavidin-fluorescein and counterstaining with PI. In HL-60 cells, the three methods gave similar results for the AI (3–4% in the controls and at 2 h of CPT treatment, and 35–43% at 3 and 4 h after CPT). This indicates that CPT-induced membrane alteration and DNA fragmentation occurred concomitantly in those cells. For MCF-7 cells, CPT-induced apoptosis developed more slowly, the AI, whether based on annexin V or on DNA content, remained unchanged at 24 h, then was increasing to 8% at 48 h and to 25% at 72 h of treatment. In these cells, the TUNEL index did not increase prior to 72 h, and the increase was minor (up to 9% vs. 2–3% in the controls) at 72 h of the treatment. This indicates that in MCF-7 cells DNA strand breaks cannot be effectively labelled, which may be due to inaccessibility of 3-OH ends in the breaks to exogenous terminal deoxynucleotidyl transferase. The mechanism of endonucleolytic DNA fragmentation thus may be different, depending on the cell type.