A positive relationship between functional redundancy and temperature in Cenozoic marine ecosystems.

The long-term effects of climate change on biodiversity and biogeographic patterns are uncertain. There are known relationships between geographic area and both the number of species and the number of ecological functional groups-termed the species-area relationship and the functional diversity-area relationship, respectively. We show that there is a positive relationship between the number of species in an area, the number of ecological functional groups, and oceanic temperature in the shallow-marine fossil record of New Zealand over a time span of ~40 million years. One implication of this relationship is that functional redundancy increases with temperature. This reveals a long-lived and persistent association between the spatial structuring of biodiversity, the temperature-dependence of functional redundancy, and shallow-marine biodiversity in mid-latitudes.

Towards optimal use of phosphorus fertiliser.

Because phosphorus (P) is one of the most limiting nutrients in agricultural systems, P fertilisation is essential to feed the world. However, declining P reserves demand far more effective use of this crucial resource. Here, we use meta-analysis to synthesize yield responses to P fertilisation in grasslands, the most common type of agricultural land, to identify under which conditions P fertilisation is most effective. Yield responses to P fertilisation were 40-100% higher in (a) tropical vs temperate regions; (b) grass/legume mixtures vs grass monocultures; and (c) soil pH of 5-6 vs other pHs. The agronomic efficiency of P fertilisation decreased for greater P application rates. Moreover, soils with low P availability reacted disproportionately strong to fertilisation. Hence, low fertiliser application rates to P-deficient soils result in stronger absolute yield benefits than high rates applied to soils with a higher P status. Overall, our results suggest that optimising P fertiliser use is key to sustainable intensification of agricultural systems.

Oral corticosteroid prescribing habits for rhinosinusitis: The American Rhinologic Society membership.

BACKGROUND: In the field of otolaryngology, oral corticosteroids (OCS) are widely prescribed for rhinosinusitis. Although there is evidence in the literature regarding specific OCS dosing protocols, it is not known to what extent these recommendations are being followed. OBJECTIVE: To examine the current state of OCS prescribing habits for rhinosinusitis by American Rhinologic Society members. METHODS: An anonymous online survey was sent to all American Rhinologic Society members. Dosing, frequency, tapering, and overall prescribing habits for OCS were assessed in chronic rhinosinusitis with polyposis (CRSwP) and in chronic rhinosinusitis without polyposis and acute bacterial rhinosinusitis. The CRSwP group was subdivided into aspirin-exacerbated respiratory disease, allergic fungal sinusitis, and not otherwise specified. Results were compared with current guidelines. Descriptive statistics were used to analyze data. RESULTS: Ninety-three surveys were completed (response rate, 12.9%). Prednisone was the most common OCS prescribed. In the CRSwP-aspirin-exacerbated respiratory disease group (n = 86), the median starting dose was 60 mg (range, 4-80 mg) and the average duration was 8 days (range, 2-28 days). In the CRSwP-allergic fungal sinusitis group (n = 81), the median starting dose was 50 mg (range, 20-60 mg), and the average duration was 6 days (range, 2-35 days). In the CRSwP-not otherwise specified group (n = 84), the median starting dose was 50 mg (range, 20-80 mg) and the average duration was 5 days (range, 1-21 days). OCS were prescribed for chronic rhinosinusitis without polyposis and acute bacterial rhinosinusitis by 66.0 and 62.4% of respondents, respectively. CONCLUSION: Significant heterogeneity existed in OCS prescribing habits for rhinosinusitis. Discrepancies were observed between survey results and evidence-based recommendations. Developing standardized OCS treatment protocols for rhinosinusitis may improve the quality of care by optimizing clinical outcomes and reducing the risk of complications.

Earthworms increase plant production: a meta-analysis.

To meet the challenge of feeding a growing world population with minimal environmental impact, we need comprehensive and quantitative knowledge of ecological factors affecting crop production. Earthworms are among the most important soil dwelling invertebrates. Their activity affects both biotic and abiotic soil properties, in turn affecting plant growth. Yet, studies on the effect of earthworm presence on crop yields have not been quantitatively synthesized. Here we show, using meta-analysis, that on average earthworm presence in agroecosystems leads to a 25% increase in crop yield and a 23% increase in aboveground biomass. The magnitude of these effects depends on presence of crop residue, earthworm density and type and rate of fertilization. The positive effects of earthworms become larger when more residue is returned to the soil, but disappear when soil nitrogen availability is high. This suggests that earthworms stimulate plant growth predominantly through releasing nitrogen locked away in residue and soil organic matter. Our results therefore imply that earthworms are of crucial importance to decrease the yield gap of farmers who can't -or won't- use nitrogen fertilizer.

Evaluation of two commercial, rapid, ELISA kits testing for scrapie in retro-pharyngeal lymph nodes in sheep.

AIMS: To estimate the number of cases of scrapie that would occur in sheep of different prion protein (PrP) genotypes if scrapie was to become established in New Zealand, and to compare the performance of two commercially available, rapid ELISA kits using ovine retro-pharyngeal lymph nodes (RLN) from non-infected and infected sheep of different PrP genotypes. METHODS: Using published data on the distribution of PrP genotypes within the New Zealand sheep flock and the prevalence of cases of scrapie in these genotypes in the United Kingdom, the annual expected number of cases of scrapie per genotype was estimated, should scrapie become established in New Zealand, assuming a total population of 28 million sheep. A non-infected panel of RLN was collected from 737 sheep from New Zealand that had been culled, found in extremis or died. Brain stem samples were also collected from 131 of these sheep. A second panel of infected samples comprised 218 and 117 RLN from confirmed scrapie cases that had originated in Europe and the United States of America, respectively. All samples were screened using two commercial, rapid, transmissible spongiform encephalopathy ELISA kits: Bio-Rad TeSeE ELISA (ELISA-BR), and IDEXX HerdChek BSE-Scrapie AG Test (ELISA-ID). RESULTS: If scrapie became established in New Zealand, an estimated 596 cases would occur per year; of these 234 (39%) and 271 (46%) would be in sheep carrying ARQ/ARQ and ARQ/VRQ PrP genotypes, respectively. For the non-infected samples from New Zealand the diagnostic specificity of both ELISA kits was 100%. When considering all infected samples, the diagnostic sensitivity was 70.4 (95% CI=65.3-75.3)% for ELISA-BR and 91.6 (95% CI=88.2-94.4)% for ELISA-ID. For the ARQ/ARQ genotype (n=195), sensitivity was 66.2% for ELISA-BR and 90.8% for ELISA-ID, and for the ARQ/VRQ genotype (n=107), sensitivity was 81.3% for ELISA-BR and 98.1% for ELISA-ID. CONCLUSIONS: In this study, the ELISA-ID kit demonstrated a higher diagnostic sensitivity for detecting scrapie in samples of RLN from sheep carrying scrapie-susceptible PrP genotypes than the ELISA-BR kit at comparable diagnostic specificity. CLINICAL RELEVANCE: The diagnostic performance of the ELISA-ID kit using ovine RLN merits the consideration of including this assay in the national scrapie surveillance programme in New Zealand.

Comparison of the Q-fever complement fixation test and two commercial enzyme-linked immunosorbent assays for the detection of serum antibodies against Coxiella burnetti (Q-fever) in ruminants : recommendations for use of serological tests on imported animals in New Zealand.

AIM: To make valid recommendations on the use of serological test methods for the detection of serum antibodies in ruminants against Coxiella burnetii (Q-fever), by comparing the performance of the complement fixation test (CFT) and two ELISA, and by identifying reasons for discrepancies between the test methods. METHODS: A total of 73 serum samples from infected cattle, 69 from infected goats, and 100 samples from non-infected cattle and 57 samples from non-infected sheep, as well as 95 samples from infected cattle herds (mix of seropositive and seronegative samples), were tested using the CFT, the IDEXX ELISA (I-ELISA) and the Pourquier ELISA (P-ELISA). A mixed panel of 12 serum samples from sheep from inter-laboratory proficiency testing (proficiency panel) was also tested using the CFT and both ELISA, and further investigated using IgG- and IgM-specific ELISA. RESULTS: Generally, the two commercial ELISA were more sensitive than the CFT for the detection of infected ruminants. Good agreement between ELISA for positive and negative results was found for samples from the infected herd, while results for the positive panels varied between the two ELISA. For the total of the positive serum panels, the I-ELISA detected 95% of samples as positive or suspicious, while the P-ELISA detected only 81%. In the P-ELISA, more samples were considered suspicious (18%) than in the I-ELISA (14%). All sera from non-infected sheep and cattle tested negative in the serological test methods employed, except for one positive sample from a sheep in the P-ELISA. Further investigation revealed that a CFT-positive but ELISA-negative result was due to high IgM and low IgG reactivity. CONCLUSIONS: The two commercial ELISA were more sensitive than the CFT in all panels from infected ruminants. However, they could only detect IgG. The I-ELISA should be the serological test method of choice for cattle, sheep and goats for import testing of animals into New Zealand because it was more sensitive than the P-ELISA and was equally specific to the PELISA and the CFT. For other animal species, such as deer and camelids, the CFT should still be used since none of the ELISA has been evaluated for these species. This study has shown that the two commercial ELISA will detect the majority of infected ruminants but may miss animals that have not developed an IgG response.

Effect of conformity and contact stress on wear in fixed-bearing total knee prostheses.

Ultra high molecular weight polyethylene (PE) remains the primary bearing surface of choice in total knee replacements (TKR). Wear is controlled by levels of cross-shear motion and contact stress. The aim of this study was to compare the wear of fixed-bearing total knee replacements with curved and flat inserts and to test the hypothesis that the flat inserts which give higher contact stresses and smaller contact areas would lead to lower levels of surface wear. A low-conforming, high contact stress knee with a low-medium level of cross shear resulted in significantly lower wear rates in comparison to a standard cruciate sacrificing fixed-bearing knee. The low wear solution found in the knee simulator was supported by fundamental studies of wear as a function of pressure and cross shear in the pin on plate system. Current designs of fixed-bearing knees do not offer this low wear solution due to their medium cross shear, moderate conformity and medium contact stress.

Rate, extent and concentration dependence of histamine-evoked Weibel-Palade body exocytosis determined from individual fusion events in human endothelial cells.

The rate, concentration dependence and extent of histamine-evoked Weibel-Palade body (WPB) exocytosis were investigated with time-resolved fluorescence microscopy in cultured human umbilical vein endothelial cells expressing WPB-targeted chimeras of enhanced green fluorescent protein (EGFP). Exocytosis of single WPBs was characterized by an increase in EGFP fluorescence, morphological changes and release of WPB contents. The fluorescence increase was due to a rise of intra-WPB pH from resting levels, estimated as pH 5.45+/-0.26 (s.d., n=144), to pH 7.40. It coincided with uptake of extracellular Alexa-647, indicating the formation of a fusion pore, prior to loss of fluorescent contents. Delays between the increase in intracellular free calcium ion concentration evoked by histamine and the first fusion event were 10.0+/-4.42 s (n=9 cells) at 0.3 microM histamine and 1.57+/-0.21 s (n=15 cells) at 100 microM histamine, indicating the existence of a slow process or processes in histamine-evoked WPB exocytosis. The maximum rates of exocytosis were 1.20+/-0.16 WPB s(-1) (n=9) at 0.3 microM and 3.66+/-0.45 WPB s(-1) at 100 microM histamine (n=15). These occurred 2-5 s after histamine addition and declined to lower rates with continued stimulation. The initial delays and maximal rate of exocytosis were unaffected by removal of external Ca2+ indicating that the initial burst of secretion is driven by Ca2+ release from internal stores, but sustained exocytosis required external Ca2+. Data were compared to exocytosis evoked by a maximal concentration of the strong secretagogue ionomycin (1 microM), for which there was a delay between calcium elevation and secretion of 1.67+/-0.24 s (n=6), and a peak fusion rate of approximately 10 WPB s(-1).

Wear, debris, and biologic activity of cross-linked polyethylene in the knee: benefits and potential concerns.

Cross-linked polyethylene currently is being introduced in knee prostheses. The wear rates, wear debris, and biologic reactivity of non cross-linked, moderately cross-linked, and highly cross-linked polyethylene have been compared in multidirectional wear tests and knee simulators. Multidirectional pin-on-plate wear studies of noncross-linked, moderately cross-linked (5 Mrad), and highly cross-linked (10 Mrad) polyethylene showed a 75% reduction in wear with the highly cross-linked material under kinematics found in the hip, but only a 33% reduction under wear in kinematics representative of the knee. In knee simulator studies, with the fixed-bearing press-fit, condylar Sigma cruciate-retaining knee under high kinematic input conditions, the wear of 5 Mrad moderately cross-linked polyethylene was 13 +/- 4 mm per 1 million cycles, which was lower (p < 0.05) than the wear of clinically used, gamma vacuum foil GUR 1020 polyethylene (23 +/- 6 mm/1 million cycles). For the low-contact stress mobile-bearing knee, the wear of moderately cross-linked polyethylene was 2 +/- 1 mm per 1 million cycles, which was lower (p < 0.05) than GVF GUR 1020 polyethylene (5 +/- 2 mm/1 million cycles). The wear debris isolated from the fixed-bearing knees showed the moderately cross-linked material had a larger percentage volume of particles smaller than 1 mum in size, compared with GVF GUR 1020 polyethylene. Direct cell culture studies of wear debris generated in sterile wear simulators using multidirectional motion showed a increase (p < 0.05) in tumor necrosis factor-alpha levels and reactivity for GUR 1050 cross-linked polyethylene debris compared with an equivalent volume of noncross-linked GUR 1050 polyethylene. The use of cross-linked polyethylene in the knee reduces the volumetric wear rate. However, the clinical significance of reduced fracture toughness, elevated wear in abrasive conditions, and the elevated tumor necrosis factor-alpha release from smaller more reactive particles warrant further investigation.

Orbitally induced oscillations in the East Antarctic ice sheet at the Oligocene/Miocene boundary.

Between 34 and 15 million years (Myr) ago, when planetary temperatures were 3-4 degrees C warmer than at present and atmospheric CO2 concentrations were twice as high as today, the Antarctic ice sheets may have been unstable. Oxygen isotope records from deep-sea sediment cores suggest that during this time fluctuations in global temperatures and high-latitude continental ice volumes were influenced by orbital cycles. But it has hitherto not been possible to calibrate the inferred changes in ice volume with direct evidence for oscillations of the Antarctic ice sheets. Here we present sediment data from shallow marine cores in the western Ross Sea that exhibit well dated cyclic variations, and which link the extent of the East Antarctic ice sheet directly to orbital cycles during the Oligocene/Miocene transition (24.1-23.7 Myr ago). Three rapidly deposited glacimarine sequences are constrained to a period of less than 450 kyr by our age model, suggesting that orbital influences at the frequencies of obliquity (40 kyr) and eccentricity (125 kyr) controlled the oscillations of the ice margin at that time. An erosional hiatus covering 250 kyr provides direct evidence for a major episode of global cooling and ice-sheet expansion about 23.7 Myr ago, which had previously been inferred from oxygen isotope data (Mi1 event).

Cholesterol binds to synaptophysin and is required for biogenesis of synaptic vesicles.

Here, to study lipid-protein interactions that contribute to the biogenesis of regulated secretory vesicles, we have developed new approaches by which to label proteins in vivo, using photoactivatable cholesterol and glycerophospholipids. We identify synaptophysin as a major specifically cholesterol-binding protein in PC12 cells and brain synaptic vesicles. Limited cholesterol depletion, which has little effect on total endocytic activity, blocks the biogenesis of synaptic-like microvesicles (SLMVs) from the plasma membrane. We propose that specific interactions between cholesterol and SLMV membrane proteins, such as synaptophysin, contribute to both the segregation of SLMV membrane constituents from plasma-membrane constituents, and the induction of synaptic-vesicle curvature.

Synaptic vesicle biogenesis.

Synaptic vesicles, which have been a paradigm for the fusion of a vesicle with its target membrane, also serve as a model for understanding the formation of a vesicle from its donor membrane. Synaptic vesicles, which are formed and recycled at the periphery of the neuron, contain a highly restricted set of neuronal proteins. Insight into the trafficking of synaptic vesicle proteins has come from studying not only neurons but also neuroendocrine cells, which form synaptic-like microvesicles (SLMVs). Formation and recycling of synaptic vesicles/SLMVs takes place from the early endosome and the plasma membrane. The cytoplasmic machinery of synaptic vesicle/SLMV formation and recycling has been studied by a variety of experimental approaches, in particular using cell-free systems. This has revealed distinct machineries for membrane budding and fission. Budding is mediated by clathrin and clathrin adaptors, whereas fission is mediated by dynamin and its interacting protein SH3p4, a lysophosphatidic acid acyl transferase.

Selective localization of the polytopic membrane protein prominin in microvilli of epithelial cells - a combination of apical sorting and retention in plasma membrane protrusions.

Prominin is a recently identified polytopic membrane protein expressed in various epithelial cells, where it is selectively associated with microvilli. When expressed in non-epithelial cells, prominin is enriched in plasma membrane protrusions. This raises the question of whether the selective association of prominin with microvilli in epithelial cells is solely due to its preference for, and stabilization in, plasma membrane protrusions, or is due to both sorting to the apical plasma membrane domain and subsequent enrichment in plasma membrane protrusions. To investigate this question, we have generated stably transfected MDCK cells expressing either full-length or C-terminally truncated forms of mouse prominin. Confocal immunofluorescence and domain-selective cell surface biotinylation experiments on transfected MDCK cells grown on permeable supports demonstrated the virtually exclusive apical localization of prominin at steady state. Pulse-chase experiments in combination with domain-selective cell surface biotinylation showed that newly synthesized prominin was directly targeted to the apical plasma membrane domain. Immunoelectron microscopy revealed that prominin was confined to microvilli rather than the planar region of the apical plasma membrane. Truncation of the cytoplasmic C-terminal tail of prominin impaired neither its apical cell surface expression nor its selective retention in microvilli. Both the apical-specific localization of prominin and its selective retention in microvilli were maintained when MDCK cells were cultured in low-calcium medium, i.e. in the absence of tight junctions. Taken together, our results show that: (i) prominin contains dual targeting information, for direct delivery to the apical plasma membrane domain and for the enrichment in the microvillar subdomain; and (ii) this dual targeting does not require the cytoplasmic C-terminal tail of prominin and still occurs in the absence of tight junctions. The latter observation suggests that entry into, and retention in, plasma membrane protrusions may play an important role in the establishment and maintenance of the apical-basal polarity of epithelial cells.

p24 and p23, the major transmembrane proteins of COPI-coated transport vesicles, form hetero-oligomeric complexes and cycle between the organelles of the early secretory pathway.

COPI-coated vesicles that bud off the Golgi complex contain two major transmembrane proteins, p23 and p24. We have localized the protein at the Golgi complex and at COPI-coated vesicles. Transport from the intermediate compartment (IC) to the Golgi can be blocked at 15 degrees C, and under these conditions p24 accumulates in peripheral punctated structures identified as IC. Release from the temperature block leads to a redistribution of p24 to the Golgi, showing that p24, similar to p23, cycles between the IC and Golgi complex. Immunoprecipitations of p24 from cell lysates and from detergent-solubilized Golgi membranes and COPI-coated vesicles show that p24 and p23 interact with each other to form a complex. Transient transfection of p23 in HeLa cells shows that p23 and p24 colocalize in structures induced by the overexpression of p23. Taken together p24 interacts with p23 and constitutively cycles between the organelles of the early secretory pathway.

Differential extraction of proteins from paraformaldehyde-fixed cells: lessons from synaptophysin and other membrane proteins.

While investigating the localization of synaptophysin in PC12 cells using immunofluorescence microscopy, we noticed a striking difference in its apparent subcellular distribution depending on whether digitonin or Triton X-100 was used as permeabilization agent of paraformaldehyde (PFA)-fixed cells. We found that this difference was due to epitope inaccessibility in the digitonin-treated cells combined with an almost quantitative extraction of the antigen on Triton X-100 permeabilization. Both phenomena were differential with respect to the various synaptophysin-containing compartments. The extraction of antigen from PFA-fixed cells was also seen with other membrane proteins but not with cytosolic proteins and proteins in the lumen of the secretory pathway. Significantly, some of the membrane proteins were extracted from the PFA-fixed cells in higher-molecular-weight forms which we believe represent their in vivo oligomeric states. The implications of our observations are discussed with respect to the method of immunofluorescence microscopy and also to the possible use of paraformaldehyde as an in vivo crosslinker for the study of membrane protein quaternary structure.

Synaptic-like microvesicles of neuroendocrine cells originate from a novel compartment that is continuous with the plasma membrane and devoid of transferrin receptor.

We have characterized the compartment from which synaptic-like microvesicles (SLMVs), the neuroendocrine counterpart of neuronal synaptic vesicles, originate. For this purpose we have exploited the previous observation that newly synthesized synaptophysin, a membrane marker of synaptic vesicles and SLMVs, is delivered to the latter organelles via the plasma membrane and an internal compartment. Specifically, synaptophysin was labeled by cell surface biotinylation of unstimulated PC12 cells at 18 degrees C, a condition which blocked the appearance of biotinylated synaptophysin in SLMVs and in which there appeared to be no significant exocytosis of SLMVs. The majority of synaptophysin labeled at 18 degrees C with the membrane-impermeant, cleavable sulfo-NHS-SS-biotin was still accessible to extracellularly added MesNa, a 150-D membrane-impermeant thiol-reducing agent, but not to the 68,000-D protein avidin. The SLMVs generated upon reversal of the temperature to 37 degrees C originated exclusively from the membranes containing the MesNa-accessible rather than the MesNa-protected population of synaptophysin molecules. Biogenesis of SLMVs from MesNa-accessible membranes was also observed after a short (2 min) biotinylation of synaptophysin at 37 degrees C followed by chase. In contrast to synaptophysin, transferrin receptor biotinylated at 18 degrees or 37 degrees C became rapidly inaccessible to MesNa. Immunofluorescence and immunogold electron microscopy of PC12 cells revealed, in addition to the previously described perinuclear endosome in which synaptophysin and transferrin receptor are colocalized, a sub-plasmalemmal tubulocisternal membrane system distinct from caveolin-positive caveolae that contained synaptophysin but little, if any, transferrin receptor. The latter synaptophysin was selectively visualized upon digitonin permeabilization and quantitatively extracted, despite paraformaldehyde fixation, by Triton X-100. Synaptophysin biotinylated at 18 degrees C was present in these subplasmalemmal membranes. We conclude that SLMVs originate from a novel compartment that is connected to the plasma membrane via a narrow membrane continuity and lacks transferrin receptor.

Corticotropin-releasing (CRH) activity of thymic peptides on CRH-insensitive corticotropic tumor cells

Se sabe que ciertas preparaciones tímicas son capaces de estimular la liberación de corticotrofina (ACTH) en células hipofisarias. No resulta claro, sin embargo, si este efecto es mediado por el receptor para la hormona liberadora de ACTH (CRH). En el presente estudio se observó que la timosina fracción cinco (TF5), el péptido tímico MB-35 y posiblemente ciertas histonas de timo de ternera son capaces de estimular la liberación de ACTH en una sublínea insensible al CRH derivada de la línea tumoral coricotropa AtT20 y por lo tanto denominada AtT20(Cl). El rango de concentraciones efectivas en el cual TF5 y MB-35 mostraron un efecto dosis-dependiente sobre la liberación de ACTH fue de 100 a 2000 µg/ml y de 10 a 100 mg/ml para TF5 y MB-35, respectivamente. Aunque ninguna de estas preparaciones indujo cambios significativos en el contenido intracelular de ACTH en las células AtT20(Cl), se observó una tendencia hacia la depleción a las dosis más altas de RF5. Los resultados obtenidos sugieren que ciertos péptidos tímicos son capaces de estimular la liberación de ACTH en células corticotropas por un mecanismo independiente del receptor par CRH

Corticotropin-releasing (CRH) activity of thymic peptides on CRH-insensitive corticotropic tumor cells

Se sabe que ciertas preparaciones tímicas son capaces de estimular la liberación de corticotrofina (ACTH) en células hipofisarias. No resulta claro, sin embargo, si este efecto es mediado por el receptor para la hormona liberadora de ACTH (CRH). En el presente estudio se observó que la timosina fracción cinco (TF5), el péptido tímico MB-35 y posiblemente ciertas histonas de timo de ternera son capaces de estimular la liberación de ACTH en una sublínea insensible al CRH derivada de la línea tumoral coricotropa AtT20 y por lo tanto denominada AtT20(Cl). El rango de concentraciones efectivas en el cual TF5 y MB-35 mostraron un efecto dosis-dependiente sobre la liberación de ACTH fue de 100 a 2000 Ag/ml y de 10 a 100 mg/ml para TF5 y MB-35, respectivamente. Aunque ninguna de estas preparaciones indujo cambios significativos en el contenido intracelular de ACTH en las células AtT20(Cl), se observó una tendencia hacia la depleción a las dosis más altas de RF5. Los resultados obtenidos sugieren que ciertos péptidos tímicos son capaces de estimular la liberación de ACTH en células corticotropas por un mecanismo independiente del receptor par CRH (AU)

Thymosin peptides stimulate corticotropin release by a calcium-dependent mechanism.

Thymosin fraction five (TF5), a well-characterized immunoregulatory thymic preparation, has been reported to stimulate corticotropin (ACTH) release from rat pituitary cells. Since a previous study in our laboratory had shown that TF5 was able to stimulate ACTH release from corticotropin-releasing hormone (CRH)-insensitive corticotropic tumor cells, it was of interest to assess the role of calcium in the mechanism of action of TF5 on corticotropic cells. A CRH-insensitive variant, denoted AtT-20(CI), of the wild-type corticotropic tumor cell line AtT-20 was used. Synthetic h/rCRH within a dose range of 0.1-100 nM was completely ineffective to stimulate basal ACTH release from AtT-20(CI) cells, although the same batch of neuropeptide displayed the expected ACTH-releasing activity on dispersed rat pituitary cells (for instance, 0.1 nM CRH induced a 3.7-fold increase in ACTH release in this cell system). Median eminence extracts (1/10) induced only a 12% increase in ACTH release from AtT-20(CI) cells as compared to the 395% stimulation induced in normal pituitary cells. As expected, TF5 induced a dose-dependent increase in ACTH release from AtT-20(CI) cells. However, this ACTH-releasing activity of TF5 was completely abolished when cells were incubated in Ca-free medium or Ca-free medium containing 0.5 mM EGTA. On the other hand, the presence of the Ca ionophore A23187 (5 microM) in medium containing normal Ca levels (2.5 mM) did not affect the ACTH-releasing activity of TF5 on AtT-20(CI) cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Corticotropin-releasing (CRH) activity of thymic peptides on CRH-insensitive corticotropic tumor cells.

A number of thymic preparations are known to stimulate corticotropin (ACTH) release from pituitary cells but it remains unclear whether this effect is mediated by the corticotropin-releasing hormone (CRH) receptor-associated pathway. We report here that thymosin fraction five (TF5), peptide MB-35 and possibly calf thymus histones can stimulate the release of ACTH from a CRH-insensitive variant of the mouse corticotropic cell line AtT20. The effective concentration range at which TF5 and MB-35 displayed their ACTH-releasing activity in a dose-dependent manner was 100 to 2,000 micrograms/ml and 10 to 100 ng/ml, respectively, whereas neither preparation induced a significant depletion of intracellular ACTH stores. Our data suggest that thymosin peptides can stimulate ACTH release from corticotrophs by a CRH receptor-independent mechanism.