Analysis of the expression of chicken and rat gene products in myoblast x myoblast cell hybrids

Hybrid cells were isolated by fusing primary chicken myoblasts to HPRT-deficient rat L6 myoblasts and incubating the cells in medium containing HAT and ouabain. All hybrid clones contained both rat and chicken chromosomes and expressed a number of gene products characteristic of both species. Although all clones were capable of fusing spontaneously to form myofibers, immunofluorescence and isoenzyme analysis revealed only the rat forms of skeletal muscle myosin and MM-creatine kinase. No differentiated gene products of chicken origin were detected. Analysis of the expression of chicken HPRT revealed that some hybrid clones were capable of modulating this enzyme activity when switched from HAT medium into thioguanine medium and back into HAT, even though HPRT is normally a constitutively expressed enzyme. Parental control cells were incapable of this modulation phenomenon.     

Characterisation of stroma-dependent blast colony-forming cells in human marrow

Human bone marrow contains a population of haemopoietic progenitor cells that can be distinguished by their ability to adhere to preformed stromal layers (cultured in the presence of methylprednisolone [MP+] and form blast cell colonies. The stromal layers function in the colony assay after they have been heavily irradiated but not after they have been passaged. The binding of the progenitor cells to the stromal cells is complete after 2 hours of coincubation, and stromal layers of 9.6 cm2 can provide adhesion sites for at least 2,000 blast colony-forming cells. The blast colony-forming cells were shown by micromanipulation to self-renew as well as to give rise to multipotential and lineage-committed colony-forming progenitor cells.     

Immunocytochemical localization of somatostatin in the brain of the lizard, Ctenosauria pectinata

The brain of the lizard, Ctenosauria pectinata, was studied light microscopically using an immunocytochemical staining method that is specific for neurohypophysial hormones and somatostatin. It was shown that the telencephalon and particularly the diencephalon contain somatostatin-producing perikarya, while somatostatinergic fibers occur in the entire brain. Similar to the situation in other vertebrates, somatostatin neurons in Ctenosauria pectinata form a population distinct from the neurohypophysial hormone-producing neurons. The small-sized somatostatin neurons were found in the cortex and the diencephalon: (1) ventral from, and partially overlapping with, the classical neurosecretory paraventricular nucleus; and (2) in the region of the infundibular (tuberal) nucleus. Somatostatin fibers were found among the classical neurosecretory fibers of the supraoptico-paraventricular system (tract, median eminence, neural lobe), near to and within the epiphysis, in the septum, in the vicinity of the tectum opticum and the cerebellum, and in the tegmentum.     

Control in congenital adrenal hyperplasia monitored by frequent saliva 17OH-progesterone measurements

Treatment in a group of 19 patients with congenital adrenal hyperplasia (CAH) has been monitored by frequent, serial measurements of saliva 17OH-progesterone (17OHP) concentrations. Detailed 17OHP profiles were obtained during consecutive weekend days and every 1-2 h over a separate 24-hour period. Patients showed a marked diurnal rhythm in 17OHP levels, particularly when treated with hydrocortisone. In some patients, 10 mg/m2/day of hydrocortisone was sufficient glucocorticoid replacement to produce adequate control, although there was considerable individual variation. Saliva 17OHP profiles provided valuable information on the efficacy of hydrocortisone, cortisone acetate, prednisolone and dexamethasone as glucocorticoid suppressive regimes in the treatment of CAH. Preliminary results suggest that hydrocortisone given in two divided doses during the day, supplemented by a small dose of prednisolone at bedtime, is suitable treatment for CAH patients who are still growing. In the patient who has completed statural growth, single daily dose dexamethasone therapy ensures adequate adrenal suppression and is convenient in the longterm.     

Requirement for extracellular calcium or magnesium in mitogen-induced activation of human peripheral blood lymphocytes

The importance of calcium in lymphocyte activation is well recognized, but the levels of extracellular ionized free calcium (Ca++) necessary for lymphocyte proliferation via various pathways have not been investigated in detail. We studied the ability of a lectin mitogen (PHA) and a calcium ionophore (ionomycin) to induce interleukin 2 receptors, interleukin 2 (IL2) production, and proliferation over various concentrations of extracellular Ca++. Reducing the Ca++ levels from the normal 200 microM to 10 microM in PHA-stimulated cultures partially inhibited IL2 receptor expression, IL2 production, and subsequent proliferation. At 1 microM Ca++, both IL2 activity and proliferation were eliminated, but partial IL2 receptor expression was still observed. Ionomycin did not induce any of these events in cultures where the extracellular Ca++ concentration was below 100 microM. Restoring calcium in the medium resulted in normal levels of IL2 receptor expression, IL2 activity, and proliferation when PBL were stimulated with either mitogen. Exogenous magnesium partially restored these events in PHA-stimulated cultures, but had no effect when ionomycin was used as the mitogen. These data indicate that stimulation by ionomycin is much more dependent upon the levels of extracellular Ca++ than is PHA. Extracellular calcium also appears to be necessary subsequent to IL2 receptor acquisition, since the latter was seen without IL2 activity or proliferation at very low extracellular Ca++, and IL2 failed to restore the proliferative response under these conditions. The data also suggest that PHA, but not ionomycin, can activate lymphocytes via a magnesium-dependent pathway, or that PHA has a lower specificity for divalent cation cofactors.     

Diethyl phthalate, a chemotactic factor secreted by Helicobacter pylori.

The structure of a small-molecule, non-peptide chemotactic factor has been determined from activity purified to apparent homogeneity from Helicobacter pylori supernatants. H. pylori was grown in brucella broth media until one liter of solution had 0.9 absorbance units. The culture was centrifuged, and the bacteria re-suspended in physiological saline and incubated at 37 degrees C for 4 h. A monocyte migration bioassay revealed the presence of a single active chemotactic factor in the supernatant from this incubation. The chemotactic factor was concentrated by solid phase chromatography and purified by reverse phase high pressure liquid chromatography. The factor was shown to be indistinguishable from diethyl phthalate (DEP) on the basis of multiple criteria including nuclear magnetic resonance spectroscopy, electron impact mass spectroscopy, UV visible absorption spectrometry, GC and high pressure liquid chromatography retention times, and chemotactic activity toward monocytes. Control experiments with incubated culture media without detectable bacteria did not yield detectable DEP, suggesting it is bacterially derived. It is not known if the bacteria produce diethyl phthalate de novo or if it is a metabolic product of a precursor molecule present in culture media. DEP produced by H. pylori in addition to DEP present in man-made products may contribute to the high levels of DEP metabolites observed in human urine. DEP represents a new class of chemotactic factor.     

Characterization of the ribosomal DNA units in two related Prunus species (P. Cerasifera and P. Spinosa)

Summary The genetic relationships between two Prunus species, involved in rootstock breeding, were examined at the level of the ribosomal RNA genes. Twenty clones of P. cerasifera, a diploid species, and 12 clones of P. spinosa, a tetraploid wild species, were studied. The use of three heterologous ribosomal DNA probes covering different regions of the ribosomal tandem repeats enabled us to construct restriction maps for EcoRI and BamHI. We identified two unit types (unit I and unit II) in P. cerasifera. In P. spinosa, P. cerasifera units were present in addition to a third ribosomal unit type (unit III). These results appeared to confirm previous cytological studies (Salesses 1973) indicating that one of the genomes in P.spinosa has homology with the one from P. cerasifera.