Isolation of inhibin like peptides from human placenta

Two moieties of inhibin could be obtained by chromatography of partially purified preparations of inhibin from human placenta on Sephadex G-100, G-25 and ion exchange chromatography on diethylaminoethyl Sephadex A-50. The higher molecular weight moiety (14,000) designated as HPI-H appears to be similar to inhibin from human seminal plasma. While the lower molecular weight moiety (1500) designated as HPI-L appears to be similar to that of sheep testicular inhibin. The preparations from both human term placenta and human seminal plasma inhibited the binding of [¹²⁵I] human follicle stimulating hormone to rat testicular receptors. This effect of inhibins could be neutralized by antisera raised against corresponding polypeptide. Further these antibodies could neutralize endogenous inhibin resulting in 2 to 3 fold increase in serum follicle stimulating harmone levels, which could then be reversed by exogenous administration of the isolated inhibin preparations.     

Albumin stabilizes 14,15-leukotriene A4

14,15-Leukotriene A4 is a pivotal biosynthetic intermediate in 15-lipoxygenase initiated leukotriene biosynthesis. This compound hydrolyzes instantaneously in phosphate buffer at pH 7.4. However, addition of human or bovine albumin to otherwise identical buffer solutions increases its stability. Intact 14,15-leukotriene A4 then decomposes by first-order kinetics with rate constants inversely proportional to the albumin concentration. Stabilization of 14,15-leukotriene A4 under certain conditions may influence its proportionate transformation by enzymatic vs non-enzymatic processes.     

GRF-induced feeding: Evidence for protein selectivity and opiate involvement

Growth hormone-releasing factor (GRF) is a hypothalamic peptide named for its ability to induce release of growth hormone from the anterior pituitary. GRF also acts as a neurotransmitter in the suprachiasmatic nucleus/medial preoptic area (SCN/MPOA) to stimulate food intake. The purpose of this series of experiments was to explore the nature of GRF-induced feeding, with a particular emphasis on macronutrient selectivity, and to examine the role of opiate activity in the paraventricular nucleus of the hypothalamus (PVN). Chow intake stimulated by GRF microinjection (1 pmol/0.5 μl) into the SCN/MPOA was blocked by injection of methyl-naltrexone (3 μg/0.5 μl) into the PVN. In animals habituated to macronutrient diets (Teklad, WI), GRF preferentially stimulated intake of protein at 2 and 4 h postinjection, whereas it had no effect on carbohydrate intake. Further, this effect was blocked by injection of naloxone (40 nmol/0.5 μl) into the PVN. Microinjection of morphine (0, 1, 10, and 17 μg/0.5 μl) into the PVN also specifically stimulated protein intake at 2 and 4 h postinjection. These results suggest that feeding derived from GRF actions in the SCN/MPOA is macronutrient selective, and is dependent on PVN opiate activity for expression.     

The effects of monocyte-conditioned medium and interleukin 1 on the synthesis of collagenous and non-collagenous proteins by mouse bone and human bone cells in vitro

Cultural adherent human mononuclear cells produce factor(s) which stimulate the release of calcium from new-born mouse calvaria in organ culture. This stimulation of bone resorption is accompanied by an inhibition of the incorporation of [³H]proline into collagen which is independent of increased prostaglandin production by the bone. When human osteoblast-like cells are treated with conditioned medium from human mononuclear cells, collagen accounts for a decreased proportion of the protein synthesised. This effect on matrix synthesis is not accompanied by an inhibitory action of the monocyte-conditioned medium preparations on net cell proliferation. In human osteoblast-like cell cultures, partially purified human interleukin 1 also inhibits the production of the bone-specific protein osteocalcin in a dose-dependent fashion. These observations are consistent with the hypothesis that products of human monocytes similar to, or identical with, human interleukin 1 may be important regulators of bone metabolism and may contribute to the bone loss seen in diseases such as chronic rheumatoid arthritis.     

Reliability of the ICP-AES for trace elements studies of biological materials

In order to investigate the reliability of the Inductively Coupled Plasma Atomic Emission Spectrometer (ICP-AES) for trace element analysis of biological materials, we have carried out extensive investigations on human plasma, using an Applied Research Laboratory’s ICP-AES. When we aspirated the untreated plasma into the spectrometer, we obtained unreliable and nonreproducible results. However, when we pretreated the plasma by wet digestion (to destroy all the organic material), we achieved reproducible and consistent results. It is, therefore, suggested that biological samples should be pretreated, preferably by wet digestion, before being aspirated into the ICP-AES for analysis.     

Interleukin-1-β and Tumor Necrosis Factor-α Increase Peripheral-Type Benzodiazepine Binding Sites in Cultured Polygonal Astrocytes

Peripheral-type benzodiazepine binding sites (PTBBS) are markedly increased in the injured CNS. Astrocytes appear to be the primary cell type which express increased PTBBS. Because certain cytokines within the injured CNS are potent mitogens for astrocytes, we examined the effects of two such cytokines, interleukin (IL)-1 beta and tumor necrosis factor (TNF), on PTBBS in cultured astrocytes using [3H]Ro 5-4864 as the specific ligand. Purified cultures of either polygonal or process-bearing astrocytes were prepared from neonatal rat cerebral hemispheres. At a concentration of 1.8 nM, specific binding of the radioactive ligand to polygonal astrocytes reached equilibrium within 60 min and was half-maximal by 5-10 min. By contrast, specific binding to process-bearing astrocytes barely exceeded background levels. IL-1 and TNF increased PTBBS within polygonal astrocytes in both dose- and time-dependent manners. At 10-50 ng/ml, IL-1 beta and TNF-alpha elevated [3H]Ro 5-4864 binding in polygonal astrocyte cultures 65 and 87%, respectively, above the level in control cultures. However, no changes in PTBBS were seen within polygonal astrocytes after IL-2 treatment. Scatchard analysis of saturation binding experiments suggested that the increase in PTBBS promoted by TNF was due to an increased number of binding sites present in polygonal astrocytes and not due to an increase in receptor affinity. Binding data suggested that PTBBS within cultures of process-bearing astrocytes were virtually absent irrespective of the treatment. These in vitro data suggest that certain cytokines found in the injured brain may be involved in up-regulating PTBBS within a particular subtype of astrocyte.     

Spreading of non-transformed and transformed cells

Mechanisms of cellular reactions responsible for the spreading non-transformed cultured tissue cells on the surface of various substrata and relationships of these reactions to the control of cell proliferation are reviewed; the special role of the membrane-cystoskeleton interactions leading to extension and attachment of psuedopods is stressed.Transition of cells from non-transformed to transformed phenotype is characterized by decreased spreading and by decreased dependence of proliferation on spreading. Manifestations of both of these spreading-associated changes are reviewed and their possible mechanisms are discussed. It is suggested that cell transition to transformed phenotype involves shift of an equilibrium between the reactions induced by the two groups of membrane-bound ligands: those attached and those not attached to the substratum.