Ketoconazole inhibits self-induced metabolism of 1,25-dihydroxyvitamin D3 and amplifies 1,25-dihydroxyvitamin D3 receptor up-regulation in rat osteosarcoma cells

Ketoconazole (an inhibitor of vitamin D-24 hydroxylase) was used to study the role of self-induced 1,25-dihydroxyvitamin D3 (1,25-D3) metabolism on cellular responsiveness to 1,25-D3. Eighteen hours of treatment with 1,25-dihydroxy-[26,27-methyl-3H]vitamin D3 (1,25-[3H]D3) increased total 1,25-D3 receptors (VDR) from 60 to 170 fmol mg/protein. In cells treated with both 1,25-[3H]D3 and ketoconazole, up-regulation of VDR was increased by 40% over that observed with cells receiving 1,25-[3H]D3 alone. Ketoconazole alone had no agonistic activity. Treatment of cells with 1 nM 1,25-[3H]D3 plus increasing doses of ketoconazole (0-30 microM) resulted in a dose-dependent increase in occupied VDR and total VDR. This up-regulation was associated with reduced 1,25-[3H]D3 catabolism. 1,25-[3H]D3-induced up-regulation of VDR typically peaked at 14 h and declined thereafter. Ketoconazole lengthened the time to reach peak VDR up-regulation to 20 h. The ability of ketoconazole to increase cell responsiveness (VDR up-regulation) was the result of both increased and prolonged occupancy of VDR by 1,25-[3H]D3. The t1/2 of occupied VDR was 2 h in the absence of ketoconazole and greater than 7 h when ketoconazole was present. Collectively, these results suggested that self-induced catabolism of 1,25-D3 is an important regulator of VDR occupancy and therefore cellular responsiveness to hormone. These data also demonstrate the usefulness of ketoconazole as an inhibitor of vitamin D hydroxylases in intact cells.     

Duplication of the streptokinase gene in the chromosome of Streptococcus equisimilis H46A

The erythromycin resistance plasmid pSM752 carrying the cloned streptokinase gene, skc, was introduced by protoplast transformation into Streptococcus equisimilis H46A from which skc was originally cloned. Cells transiently supporting the replication of pSM752 gave rise to an erythromycin-resistant clone designated H46SM which was plasmid free and produced streptokinase at levels approximately twice as high as the wild type. Southern hybridization of total cell DNA with an skc-containing probe provided evidence for the duplication of the skc gene in the H46SM chromosome. The results, which have some bearing on industrial streptokinase production, can be best explained by a single cross-over event between the chromosome and the plasmid in the region of shared homology leading to the integration of pSM752 in a Campbell-like manner.     

Seasonal variation of calcium,magnesium, potassium,and manganese contents in xylem sap of beech(Fagus sylvatica L.) in a 35-year-old limestone beech forest stand

Summary In a 35-year-old calcareous beech forest stand five beech trees (Fagus sylvatica L.) were felled every 2 weeks, and xylem sap was obtained by means of water displacement from the lowest trunk sections, each 100 cm in length. From mid-October 1988 to mid-October 1989 a total of 130 trees were investigated. The seasonal variations of the Ca, Mg, K and Mn contents, as well as those of pH, show four characteristic phases. Additionally, distribution of the mineral contents along the trunk was studied in four trees. The seasonal increase and decrease of xylem sap mineral contents along the trunk is shown for the characteristic phenophases. The Ca, Mg, K, and Mn contents of xylem saps were determined by means of atomicabsorptionspectrophotometry.     

Immunocytochemical localization of lipophorins in the flight muscles of the migratory locust (Locusts migratoria) at rest and during flight

Summary Locust lipoproteins (lipophorins) were localized by indirect immunofluorescence- and immunogold labelling in cryosections of dorsolongitudinal flight muscles. Immunolabelling was performed with monoclonal antibodies against apolipoprotein epitopes that are exposed at the surfaces of the lipophorin particles. Both at rest and during flight, lipophorins were located only in the wider spaces of the extracellular matrix, in the basement membranes of the individual muscle fibers and in the extracellular spaces that surround interfibrillar tracheoles. No internalization of lipophorins by the flight muscle cells was observed. Our results indicate that the unloading of lipophorins at the flight muscles is an extracellular event. Similarities with the vertebrate system of chylomicron and very-low-density lipoprotein degradation are discussed.     

Phenetic relationships inClerodendrum (Verbenaceae) and some phylogenetic considerations

Cluster analyses by different methods and a minimum spanning tree were used to study phenetic relationships in the genusChlerodendrum. 129 species were scored for 52 morphological characters corresponding to 119 character states. The phenetic results suggest a classification into 7 distinct groups, which may be grouped into two subgenera. This classification is supported by the iridoid distribution as well as by some phylogenetic considerations.     

Menstrual-cycle phase and sexual behavior in semi-free-ranging stumptail macaques (Macaca arctoides)

The sexual behavior and female reproductive cycles of a group of island-dwelling stumptail macaques (Macaca arctoides)were monitored over a 6-month period, yielding 530 observation hr and 268 copulations. Compared to nondominant males, the dominant male copulated at a relatively high rate throughout the cycle, but largely with one high-ranking female. The non-dominant males copulated most frequently at midcycle. Female presenting was highest at midcycle, but only to the dominant male. Cross-study discrepancies may be due to different observation methods and restricted environmental conditions that mask female-initiated sexual behavior. The more naturalistic setting of this study allowed for a fuller expression of proceptivity. Contrary to some previous conclusions, present findings suggest that both hormonal and socioenvironmental factors influence the patterns of sexual behavior found in stumptail macaque colonies.     

Initiation of morphogenic cell-suspension and protoplast cultures of barley (Hordeum vulgare L.)

Cell-suspension cultures were initiated from embryogenic calli of various barley cultivars. Seven fast-growing suspension lines were obtained from four different cultivars (cvs. Dissa, Emir, Golden Promise and Igri). Two of these cell suspensions showed morphogenic capacity. From a cell suspension of cv. Dissa, albino plantlets were regenerated when aggregates were cultured on solid medium. Aggregates of cv. Igri usually stopped differentiation at the globular stage, but occasionally formed scutellum-like structures. Five suspension lines were used for protoplast isolation and culture. Dividing protoplasts were obtained from all lines, but with cv. Igri a few divisions only and no further development were observed. Protoplasts from the various lines differed in the time of first division (2–14 d), division frequency (0.09–70.9%) and efficiency of colony formation (0.09–7.3%). Protoplasts isolated from the morphogenic cell suspension of cv. Dissa developed compact calli which sporadically regenerated albino plantlets.Abbreviations CC, MS, N6, SH, Kao8p culture media; see Material and methods - cv chltivar - dicamba 3,6-dichloro-o-anisic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - picloram 4-amino-3,5,6-trichloropicolinic acid     

Acquisition of phosphorus and copper by VA-mycorrhizal hyphae and root-to-shoot transport in white clover

White clover (Trifolium repens L.) plants were grown in a calcareous soil in pots with three compartments, a central one for root growth and two outer ones for growth of vesicular-arbuscular (VA) mycorrhizal (Glomus mosseae [Nicol. & Gerd.] Gerdemann & Trappe) hyphae (hyphal compartments). Phosphorus (P) was applied at three levels (0, 20 and 50 mg kg⁻¹ soil) in the outer compartments in mycorrhizal treatments. Root and shoot dry weight were increased in mycorrhizal plants with hyphal access to outer compartments. Growth of the mycorrhizal hyphae in the outer compartments was not significantly affected by variation in P level in these compartments. However, both concentration and amount of P in roots and shoots sharply increased with increasing P supply in the outer (hyphal) compartments. With increasing P levels the calculated delivery of P by the hyphae from the outer compartments increased from 34% to 90% of total P uptake. Hyphal access to the outer compartments also significantly increased both concentration and quantity of Cu in the plants. The calculated delivery of Cu by the hyphae from the outer compartments ranged from 53% to 62% of total Cu uptake, irrespective of the P levels and the amounts of P taken up and transported by the hyphae. However, the distribution of Cu over roots and shoots was largely dependent on P levels. With increase in P level in the outer compartments the calculated hyphal contribution to the total amount of Cu in the shoots increased from 12% to 58%, but decreased in the roots from 75% to 46%. In conclusion, uptake and transport by VA-mycorrhizal hyphae may contribute substantially not only to P nutrition, but also to Cu nutrition of the host.     

Spatial and temporal localization of FGF receptors in Xenopus laevis

Summary Mesoderm formation is a result of cell-cell interactions between the vegetal and animal hemisphere and is thought to be mediated by inducing peptide growth factors including members of the FGF and TGF superfamilies. Our immunochemical study analyses the distribution of FGF receptors coded by the human flg gene during embryogenesis of Xenopus laevis. Immunostaining was detected in the dorsal and ventral ectoderm and also in the marginal zone of early cleavage, blastula and gastrula stages. Signals were very strong in the mid and late blastula (stage 8 and 9) and declined slightly in the early gastrula (stage 10). A dramatic decrease was observed up to the late gastrula (stage 11⁺). In stage 13 embryos, immunostaining was only found in cells around the blastopore. Isolated ectoderm cultured in vitro showed a similar temporal expression and decrease of the signal as the normal embryos. These results indicate that receptor expression is independent of the interaction of the animal cells with the vegetal part of the embryo. Of interest is the fact that the signal cannot only be found at or near the cell surface but also within the cell. This suggests the presence of an intracellular isoform of the receptor resulting from the endogenous expression of splice variants and the internalization of transmembrane receptor. Taken together our results suggest that the loss of competence (for bFGF around stage 10) is not directly correlated with the presence of receptors. The possible roles of heparan sulphate glucosaminoglycans (low affinity receptors) and control mechanisms in the intracellular signalling pathway downstream of the receptor level should be taken into consideration.