M-CSF potently augments RANKL-induced resorption activation in mature human osteoclasts

Macrophage-CSF (M-CSF) is critical for osteoclast (OC) differentiation and is reported to enhance mature OC survival and motility. However, its role in the regulation of bone resorption, the main function of OCs, has not been well characterised. To address this we analysed short-term cultures of fully differentiated OCs derived from human colony forming unit-granulocyte macrophages (CFU-GM). When cultured on dentine, OC survival was enhanced by M-CSF but more effectively by receptor activator of NFκB ligand (RANKL). Resorption was entirely dependent on the presence of RANKL. Co-treatment with M-CSF augmented RANKL-induced resorption in a concentration-dependent manner with a (200-300%) stimulation at 25 ng/mL, an effect observed within 4-6 h. M-CSF co-treatment also increased number of resorption pits and F-actin sealing zones, but not the number of OCs or pit size, indicating stimulation of the proportion of OCs activated. M-CSF facilitated RANKL-induced activation of c-fos and extracellular signal-regulated kinase (ERK) 1/2 phosphorylation, but not NFκB nor nuclear factor of activated T-cells, cytoplasmic-1 (NFATc1). The mitogen-activated protein kinase kinase (MEK) 1 inhibitor PD98059 partially blocked augmentation of resorption by M-CSF. Our results reveal a previously unidentified role of M-CSF as a potent stimulator of mature OC resorbing activity, possibly mediated via ERK upstream of c-fos.     

Rem2-targeted shRNAs reduce frequency of miniature excitatory postsynaptic currents without altering voltage-gated Ca²⁺ currents

Ca²⁺ influx through voltage-gated Ca²⁺ channels (VGCCs) plays important roles in neuronal cell development and function. Rem2 is a member of the RGK (Rad, Rem, Rem2, Gem/Kir) subfamily of small GTPases that confers potent inhibition upon VGCCs. The physiologic roles of RGK proteins, particularly in the brain, are poorly understood. Rem2 was implicated in synaptogenesis through an RNAi screen and proposed to regulate Ca²⁺ homeostasis in neurons. To test this hypothesis and uncover physiological roles for Rem2 in the brain, we investigated the molecular mechanisms by which Rem2 knockdown affected synaptogenesis and Ca²⁺ homeostasis in cultured rat hippocampal neurons. Expression of a cocktail of shRNAs targeting rat Rem2 (rRem2) reduced the frequency of miniature excitatory postsynaptic currents (mEPSCs) measured 10 d after transfection (14 d in vitro), but did not affect mEPSC amplitude. VGCC current amplitude after rRem2-targeted knockdown was not different from that in control cells, however, at either 4 or 10 d post transfection. Co-expression of a human Rem2 that was insensitive to the shRNAs targeting rRem2 was unable to prevent the reduction in mEPSC frequency after rRem2-targeted knockdown. Over-expression of rRem2 resulted in 50% reduction in VGCC current, but neither the mEPSC frequency nor amplitude was affected. Taken together, the observed effects upon synaptogenesis after shRNA treatment are more likely due to mechanisms other than modulation of VGCCs and Ca²⁺ homeostasis, and may be independent of Rem2. In addition, our results reveal a surprising lack of contribution of VGCCs to synaptogenesis during early development in cultured hippocampal neurons.     

Archaeology on the Great Plains

Archaeology on the Great Plains. W. Raymond Wood. ed. Lawrence: University Press of Kansas, 1998. 522 pp.     

Repeated grazing of a salt marsh grass by moulting greylag geese Anser anser– does sequential harvesting optimise biomass or protein gain?

The effects of simulated goose grazing on common saltmarsh-grass Puccinellia maritima plants were tested on a Danish salt marsh during the flightless moulting period of greylag geese Anser anser (3–21 June 1998). Plants in an area exclosed from the influence of grazing and the nutrient effects of goose faeces were subject to removal of youngest lamina at 3-, 6-, 9- and 18-day intervals during this period. Average biomass and protein accumulation between harvests was highest at defoliation intervals of 9 days or more. Field observations from two separate study areas demonstrated geese returned to regraze the Puccinellia sward after 6–8 days and oesophageal contents from feeding geese showed selection for lamina lengths consistent with the results of clipping every 6 days. Geese therefore regrazed Puccinellia patches at shorter intervals than expected were they to maximise their intake of biomass or protein at each visit. However, total cumulative lamina elongation, equivalent to the long term gain during the entire moult period, showed no significant difference between the three most intensive defoliation treatments, which were significantly greater than those of plants defoliated at 18 day intervals. Highest overall lamina protein levels were maintained at 6- and 9-day defoliation intervals. This suggests geese regrazed Puccinellia patches at a rate that maximised their number of harvests during the flightless period, but maintained highest protein levels and overall biomass in the sward. This suggests, in line with earlier studies, that moulting greylag geese combine dietary selection, reduced nitrogen excretion and regrazing patterns to meet protein demands during regrowth of flight feathers.     

C20orf9-003 (ACI-1), a gene localized on chromosome 20q13.12 encoding for a 49 kD cytoplasmic protein with a putative nucleotide binding site.

Murine NGD5 is a gene identified from NG108-15 cells which is postulated to be involved in opioid receptor function. Here we report the cloning and characterization of a cDNA C20orf9-003 (ACI-1) encoding the human orthologue of the mouse NGD5. Analysis of the genomic structure revealed that C20orf9-003 (ACI-1) contains 13 exons and 12 introns, spanning 52.5kb of genomic DNA and is a variant of C20orf9. Chromosomal localization of human C20orf9-003 (ACI-1) assigned this gene to chromosome 20q13.12. Genes at this locus have been associated with the progression and possibly the development of various cancers. In addition several linkage studies support the possibility that one or more genes affecting obesity are located in 20q13. No function can be clearly assigned to C20orf9-003 (ACI-1), however, the protein has a cytoplasmic subcellular location and the secondary structure contains a Rossman fold like feature which is found in many nucleotide binding proteins.     

Amino acids and peptides: Part XLIII. Structure-activity studies in the bradykinin series: Synthesis of analogs modified in position

β-Alanyl-, acetimidoyl-, and carbamoyl-bradykinin [(IV), (V), and (VI), respectively] have been synthesized by the picolyl ester method. The first two analogs have very low smooth muscle contracting activity, but the carbamoyl derivative is as fully active as bradykinin itself.     

Coming of age: steroid hormones of wild immature baboons (Papio cynocephalus)

Large gaps exist in our knowledge about common patterns and variability in the endocrinology of immature nonhuman primates, and even normal hormonal profiles during that life stage are lacking for wild populations. In the present study we present steroid profiles for a wild population of baboons (Papio cynocephalus) from infancy through reproductive maturation, obtained by noninvasive fecal analyses. Fecal concentrations of glucocorticoid (fGC) and testosterone (fT) metabolites for males, and of fGC, estrogen (fE), and progestin (fP) metabolites for females were measured by radioimmunoassay (RIA). In males, infancy was characterized by high and declining levels of fGC and fT, whereas steroid concentrations were low during the juvenile years. During the months immediately prior to testicular enlargement, fT (but not fGC) concentration tended to increase. Males that matured early consistently had higher fT and fGC concentrations than those that matured late, but not significantly so at any age. Individual differences in fT concentrations were stable across ages, and average individual fT and fGC concentrations were positively correlated. For females, high and declining levels of fE characterized infancy, and values increased again after 3.5 years of age, as some females reached menarche by that age. Both fP and fGC were relatively low and constant throughout infancy and the juvenile period. During the months immediately prior to menarche, fGC concentration significantly decreased, while no changes were observed for fE levels. fP exhibited a complicated pattern of decrease that was subsequently followed by a more modest and nonsignificant increase as menarche approached. Early- (EM) and late-maturing (LM) females differed only in fP concentration; the higher fP concentrations in EM females reached significance at 4-4.5 years of age. Maternal rank at offspring conception did not predict concentrations of any hormone for either sex. Our results demonstrate the presence of individual endocrine variability, which could have important consequences for the timing of sexual maturation and subsequently for individual reproductive success. Further evaluation of the factors that affect hormone concentrations during the juvenile and adolescent periods should lead to a better understanding of mechanisms of life-history variability.