Greater Stimulatory Effect Research Articles

Cranberry phytochemicals act as non‐microbial antigens for different types of human peripheral blood mononuclear cells

To determine whether specific families of phytochemicals are able to interact with human peripheral blood mononuclear cell (PBMC) receptors and initiate an immune response, isolated human PBMC were incubated with three different extracts of cranberries representing anthocyanins, proanthocyanidins, or polyphenols. Proliferation of cells upon ex vivo stimulation and surface marker expression were measured by flow cytometry. The fraction containing mainly proanthocyanidins caused proliferation and activation of ãä T cells, as well as the activation of B cells. The fraction containing mainly anthocyanins slightly activated NK cells detected as an increased expression of NKG2D on NK cell surfaces. The polyphenol fraction increased B cell proliferation, and markedly increased expression of VLA‐1, about 10 fold, on monocytes. VLA‐1 is a cell adhesion molecule. PBMC secreted a number of cytokines in response to incubation with the cranberry fractions; the proanthocyanidin‐containing fraction having the greatest stimulatory effect. These results suggest that cranberry phytochemicals have the potential to immuno‐modulate in a variety of ways. Supported by Ocean Spray Cranberries, Inc and FAES

Extracellular Chloride Regulates the Epithelial Sodium Channel

The extracellular domain of the epithelial sodium channel ENaC is exposed to a wide range of Cl(-) concentrations in the kidney and in other epithelia. We tested whether Cl(-) alters ENaC activity. In Xenopus oocytes expressing human ENaC, replacement of Cl(-) with SO4(2-), H2PO4(-), or SCN(-) produced a large increase in ENaC current, indicating that extracellular Cl(-) inhibits ENaC. Extracellular Cl(-) also inhibited ENaC in Na+-transporting epithelia. The anion selectivity sequence was SCN(-) < SO4(2-) < H2PO4(-) < F(-) < I(-) < Cl(-) < Br(-). Crystallization of ASIC1a revealed a Cl(-) binding site in the extracellular domain. We found that mutation of corresponding residues in ENaC (alpha(H418A) and beta(R388A)) disrupted the response to Cl(-), suggesting that Cl(-) might regulate ENaC through an analogous binding site. Maneuvers that lock ENaC in an open state (a DEG mutation and trypsin) abolished ENaC regulation by Cl(-). The response to Cl(-) was also modulated by changes in extracellular pH; acidic pH increased and alkaline pH reduced ENaC inhibition by Cl(-). Cl(-) regulated ENaC activity in part through enhanced Na+ self-inhibition, a process by which extracellular Na+ inhibits ENaC. Together, the data indicate that extracellular Cl(-) regulates ENaC activity, providing a potential mechanism by which changes in extracellular Cl(-) might modulate epithelial Na+ absorption.

Effects of temperature, ultraviolet radiation and pectin methyl esterase on aerobic methane release from plant material

This study examines the effects of different irradiance types on aerobic methane (CH(4)) efflux rates from terrestrial plant material. Furthermore, the role of the enzyme pectin methyl esterase (PME) on CH(4) efflux potential was also examined. Different types of plant tissue and purified pectin were incubated in glass vials with different combinations of irradiation and/or temperature. Purified dry pectin was incubated in solution, and with or without PME. Before and after incubation, the concentration of CH(4) was measured with a gas chromatograph. Rates of CH(4) emission were found to depend exponentially on temperature and linearly on UV-B irradiance. UV-B had a greater stimulating effect than UV-A, while visible light had no effect on emission rates. PME was found to substantially reduce the potential for aerobic CH(4) emissions upon demethylation of pectin.

Pituitary adenylate cyclase-activating polypeptide directly stimulates LH and FSH secretion by human pituitary gonadotrophinomas

The effect of pituitary adenylate cyclase-activating polypeptide (PACAP) on LH and FSH secretion by human pituitary gonadotrophinomas in cell culture was studied. PACAP (1-38 peptide, 0.2-20 nmol/L) dose-dependently stimulated both LH and FSH secretion after 24 hours incubation. Of 11 tumours studied, PACAP (20 nmol/L) stimulated LH and/or FSH secretion by 1.7-4 fold in 9 cases. Two tumours did not respond to PACAP, although LHRH was stimulatory in these. None of the 11 tumours contained gsp mutations, excluding the possibility that these were the cause of the occassionally observed non-responsiveness to PACAP. A combination of PACAP (20 nmol/L) together with TRH (25 nmol/L) resulted in greater stimulatory effects on LH and FSH secretion than exerted by either peptide alone, but this was not observed with LHRH. In 3 tumours tested, PACAP stimulated cAMP production 2-3 fold by cultured human pituitary gonadotrophinomas but had no effect on rate of phosphatidylinositol (PI) turnover. These results indicate that PACAP can directly stimulate LH and FSH secretion by human pituitary gonadotrophs and that PACAP-receptors in gonadotrophin-secreting tumours are coupled with adenylate cyclase but not the PI second messenger system. We conclude that PACAP may play a role in controlling gonadotroph function in the human pituitary gland.

Growth promotion and protection against damping-off of wheat by two rock phosphate solubilizing actinomycetes in a P-deficient soil under greenhouse conditions

Micromonospora aurantiaca- and Streptomyces griseus-related strains isolated from Moroccan phosphate mines (MA MPM and SG MPM) were previously selected for their rock phosphate (RP) solubilizing abilities and their multiple plant growth promoting properties demonstrated in laboratory conditions. In order to assess whether these interesting properties could have a direct effect on plant growth and fitness, seeds of the wheat plant ( Triticum durum L. cv. Vitron) coated or not with mycelium of these strains and of the reference strain S. griseus M1323, were grown in a sterile soil deficient in soluble phosphate supplemented or not with soluble phosphate or with the insoluble RP, under greenhouse conditions. These studies revealed that the presence of the actinomycete strains in the soil supplemented with RP significantly promoted the growth of the wheat plants. MA MPM and SG MPM had the greatest stimulatory effect on plant growth with 50–47% and 80–78% weight increase of shoots and roots, respectively, in comparison with the sterile control. This increase correlated with a significant increase in the N and P content of plant tissues. The MA MPM- and SG MPM-dependent growth promotion in the RP supplemented soil was on average 10–13% lower than that achieved by the soluble phosphate supplement. Furthermore, in a soil infested with Pythium ultimum, the mediator of damping-off disease, the coating of wheat seeds with the mycelium of MA MPM strain resulted in a clear protection of the plant. The level of protection achieved by MA MPM was 14% lower than that conferred by the commercial bio-fungicide agent (Mycostop ®). This study demonstrated that MA MPM in association with pulverized RP could constitute a novel and non-polluting bio-fertilizer/biocontrol product useful for the development of sustainable agriculture.

Expression of ethylene-forming enzyme (EFE) of Pseudomonas syringae pv. glycinea in Trichoderma viride

The efe gene encoding an ethylene-forming enzyme from Pseudomonas syringae pv. glycinea has been expressed for the first time under the control of Trichoderma reesei cbh1 promoter in Trichoderma viride. Reverse transcription polymerase chain reaction analysis showed that transformant Y2 produced mRNA of the efe gene. Southern blot analysis showed that there was one copy of efe gene which was integrated into the chromosomal DNA of T. viride. Ethylene production by transformant Y2 was efficiently induced by cellulose, while very low level of ethylene was produced when sodium carboxymethyl cellulose or lactose was used as carbon source. Peptone exerted a much greater stimulatory effect on ethylene production. A high level of ethylene was produced when transformant Y2 was cultured in solid fermentation medium containing wheat straw, indicating that plant wastes could be directly converted to ethylene by the recombinant filamentous fungus.

Effect of activin A and insulin-like growth factor-I on in vitro development of preantral follicles isolated from cryopreserved ovarian tissues in the mouse

Cryopreservation of ovarian tissue has been reported to delay the development of preantral follicles by temporary suppression of granulosa cell proliferation during in vitro culture. This delay might be overcome by treatment with activin A and/or IGF-I, known to stimulate granulosa cell proliferation. However, the effects of these growth factors, on delayed follicle development induced by ovarian tissue cryopreservation, have not been evaluated. Therefore, we studied the effects of activin A and/or IGF-I on granulosa cell proliferation and follicle development in preantral follicles isolated from mouse cryopreserved ovarian tissues. The preantral follicles isolated from fresh ovarian tissues were cultured with control medium (CM) for 10 days. The preantral follicles isolated from cryopreserved ovarian tissues were cultured with CM and with CM + activin A (100 ng/ml), IGF-I (50 ng/ml) or activin A + IGF-I added for 10, 12 and 14 days. The follicles were stimulated with hCG at the end of culture. The granulosa cell proliferation was evaluated by measuring the PCNA expression and the follicle development assessed by comparing the follicle diameter and oocyte maturation. The expressed level of PCNA was significantly decreased in the cryopreserved preantral follicles cultured with CM, compared to the fresh group ( p < 0.05), but increased to the level of the fresh group by the addition of activin A, IGF-I or activin A and IGF-I. The maximum follicle diameter and oocyte maturation rate were obtained in the fresh group after 10 days of culture, while the diameter and oocyte maturation rate of cryopreserved preantral follicles reached similar levels after 14 days. Under conditions of CM with added activin A or activin A + IGF-I, both the diameter and oocyte maturation rate of the cryopreserved preantral follicles improved to the levels of the fresh group after 12 days. However, the stimulatory effect was not different in comparisons between activin A and activin A + IGF-I. In addition, adding activin A significantly increased the survival rate of cryopreserved preantral follicles compared to IGF-I ( p < 0.05). On the other hand, the diameter of the cryopreserved preantral follicles did not increase to the level of the fresh group by addition of IGF-I, although the oocyte maturation rate was improved to the level of fresh group after 12 days of culture. These findings indicate that activin A has a greater stimulatory effect on the growth of preantral follicles than does IGF-I. In conclusion, the addition of activin A to culture media stimulated granulosa cell proliferation to overcome the delay in culture of the development of preantral follicles isolated from cryopreserved mouse ovaries.

Akt Mediates the Effect of Insulin on Epithelial Sodium Channels by Inhibiting Nedd4-2

The epithelial sodium channel (ENaC) plays an important role in transepithelial Na(+) absorption; hence its function is essential for maintaining Na(+) and fluid homeostasis and regulating blood pressure. Insulin is one of the hormones that regulates activity of ENaC. In this study, we investigated the contribution of two related protein kinases, Akt (also known as protein kinase B) and the serum- and glucocorticoid-dependent kinase (Sgk), on insulin-induced ENaC activity in Fisher rat thyroid cells expressing ENaC. Overexpression of Akt1 or Sgk1 significantly increased ENaC activity, whereas expression of a dominant-negative construct of Akt1, Akt1(K179M), decreased basal activity of ENaC. Inhibition of the endogenous expression of Akt1 and Sgk1 by short interfering RNA not only inhibited ENaC but also disrupted the stimulatory effect on ENaC of insulin and of the downstream effectors of insulin, phosphatidylinositol 3-kinase and PDK1. Conversely, overexpression of Akt1 or Sgk1 increased expression of ENaC at the cell membrane and overcame the inhibitory effect of Nedd4-2 on ENaC. Furthermore, mutation of consensus phosphorylation sites on Nedd4-2 for Akt1 and Sgk1, Ser(342) and Ser(428), completely abolished the inhibitory effect of Sgk1 and Akt1 on Nedd4-2 action. Together these data suggest that both Akt and Sgk are components of an insulin signaling pathway that increases Na(+) absorption by up-regulating membrane expression of ENaC via a regulatory system that involves inhibition of Nedd4-2.

Effect of rapidly resorbable bone substitute materials on the temporal expression of the osteoblastic phenotype in vitro

Ideally, bioactive ceramics for use in alveolar ridge augmentation should possess the ability to activate bone formation and, thus, cause the differentiation of osteoprogenitor cells into osteoblasts at their surfaces. Therefore, in order to evaluate the osteogenic potential of novel bone substitute materials, it is important to examine their effect on osteoblastic differentiation. This study examines the effect of rapidly resorbable calcium-alkali-orthophosphates on osteoblastic phenotype expression and compares this behavior to that of beta-tricalcium phosphate (TCP) and bioactive glass 45S5. Test materials were three materials (denominated GB14, GB9, GB9/25) with a crystalline phase Ca(2)KNa(PO(4))(2) and with a small amorphous portion containing either magnesium potassium phosphate (GB14) or silica phosphate (GB9 and GB9/25, which also contains Ca(2)P(2)O(7)); and a material with a novel crystalline phase Ca(10)[K/Na](PO(4))(7) (material denominated 352i). SaOS-2 human bone cells were grown on the substrata for 3, 7, 14, and 21 days, counted, and probed for an array of osteogenic markers. GB9 had the greatest stimulatory effect on osteoblastic proliferation and differentiation, suggesting that this material possesses the highest potency to enhance osteogenesis. GB14 and 352i supported osteoblast differentiation to the same or a higher degree than TCP, whereas, similar to bioactive glass 45S5, GB9/25 displayed a greater stimulatory effect on osteoblastic phenotype expression, indicating that GB9/25 is also an excellent material for promoting osteogenesis.

A comparison of the leaf shredding ability and influence on microbial films of surface and cave forms of Gammarus minus Say

Gammarus minus has both cave and surface forms that differ considerably in their morphology. We experimentally compared the ability of these two forms to consume leaves and influence the activity of the microbial films associated with leaves. Both forms of G. minus were effective leaf shredders, and were equally efficient at converting leaves to small particles. We found little relationship between amphipod size and the rate at which they shredded leaves. Both forms of G. minus stimulated the respiration rate of microbes associated with leaves by 32–52%. Cave forms of G. minus had a 15% greater stimulatory effect on microbial respiration. The stimulatory effect of G. minus on microbial respiration is evident in cave streams as well as in the laboratory. It appears that G. minus plays a particularly important role in cave systems by converting leaves to fine particles and stimulating microbial films which are important foods for many cave animals.

Translation initiation region sequence preferences in Escherichia coli

BackgroundThe mRNA translation initiation region (TIR) comprises the initiator codon, Shine-Dalgarno (SD) sequence and translational enhancers. Probably the most abundant class of enhancers contains A/U-rich sequences. We have tested the influence of SD sequence length and the presence of enhancers on the efficiency of translation initiation.ResultsWe found that during bacterial growth at 37°C, a six-nucleotide SD (AGGAGG) is more efficient than shorter or longer sequences. The A/U-rich enhancer contributes strongly to the efficiency of initiation, having the greatest stimulatory effect in the exponential growth phase of the bacteria. The SD sequences and the A/U-rich enhancer stimulate translation co-operatively: strong SDs are stimulated by the enhancer much more than weak SDs. The bacterial growth rate does not have a major influence on the TIR selection pattern. On the other hand, temperature affects the TIR preference pattern: shorter SD sequences are preferred at lower growth temperatures. We also performed an in silico analysis of the TIRs in all E. coli mRNAs. The base pairing potential of the SD sequences does not correlate with the codon adaptation index, which is used as an estimate of gene expression level.ConclusionIn E. coli the SD selection preferences are influenced by the growth temperature and not influenced by the growth rate. The A/U rich enhancers stimulate translation considerably by acting co-operatively with the SD sequences.

The selective recruitment of mRNA to the ER and an increase in initiation are important for glucose-stimulated proinsulin synthesis in pancreatic β-cells

Glucose acutely stimulates proinsulin synthesis in pancreatic beta-cells through a poorly understood post-transcriptional mechanism. In the present study, we demonstrate in pancreatic beta-cells that glucose stimulates the recruitment of ribosome-associated proinsulin mRNA, located in the cytoplasm, to the ER (endoplasmic reticulum), the site of proinsulin synthesis, and that this plays an important role in glucose-stimulated proinsulin synthesis. Interestingly, glucose has greater stimulatory effect on the recruitment of proinsulin mRNA to the ER compared with other mRNAs encoding secretory proteins. This, as far as we are aware, is the first example whereby mRNAs encoding secretory proteins are selectively recruited to the ER and provides a novel regulatory mechanism for secretory protein synthesis. Contrary to previous reports, and importantly in understanding the mechanism by which glucose stimulates proinsulin synthesis, we demonstrate that there is no large pool of 'free' proinsulin mRNA in the cytoplasm and that glucose does not increase the rate of de novo initiation on the proinsulin mRNA. However, we show that glucose does stimulate the rate of ribosome recruitment on to ribosome-associated proinsulin mRNA. In conclusion, our results provide evidence that the selective recruitment of proinsulin mRNA to the ER, together with increases in the rate of initiation are important mediators of glucose-stimulated proinsulin synthesis in pancreatic beta-cells.

Molecular Mechanisms of Modified Sensitivity of the Adenylate Cyclase Signaling System to Biogenic Amines during Streptozotocin-Induced Diabetes

We demonstrated changes in the sensitivity of the adenylate cyclase signaling system to biogenic amines (adrenoceptor agonists and serotonin) underwent a change in skeletal muscles of rats with 30-day streptozotocin-induced diabetes. Isoproterenol had a less significant stimulatory effect on adenylate cyclase in diabetic rats. Hormonal signals via Gi proteins were suppressed in animals with diabetes, which determined a greater stimulatory effect of norepinephrine and serotonin on adenylate cyclase. Hormones less significantly increased guanosine triphosphate-binding activity of G proteins in diabetic rats, which reflects the impairment of their functional coupling with receptors.

Interactive effects of functionally different earthworm species on aggregation and incorporation and decomposition of newly added residue carbon

The interactive effects of two functionally different earthworm species ( Aporrectodea caliginosa (endogeic species) and Lumbricus rubellus (epigeic species)) on the incorporation of fresh residue into large macroaggregates and formation of microaggregates within these large macroaggregates were investigated during a short-term laboratory experiment using 13C-labelled sorghum ( Sorghum bicolor (L.) Moench) residues. Soil was collected from a long-term no-tillage agricultural field, crushed through a 250-μm sieve and incubated under laboratory conditions. The following earthworm treatments were applied: (i) soil+ 13C-labelled residue+ A. caliginosa; (ii) soil+ 13C-labelled residue+ L. rubellus; (iii) soil+ 13C-labelled residue+ A. caliginosa+ L. rubellus and; (iv) soil+ 13C-labelled residue. Two residue placement treatments (i.e. surface and incorporated) were superimposed on the earthworm treatments. Earthworms were added after 8 days of incubation. Aggregate size distribution and total C and 13C were measured after 22 days. Microaggregates, fine inter-microaggregate particulate organic matter (inter-POM) and intra-microaggregate POM (intra-POM) were isolated from macroaggregates. Earthworms had a greater stimulating effect on the formation of large macroaggregates (>2000 μm) and microaggregates within large macroaggregates when residue was incorporated in the soil, especially in the presence of A. caliginosa. When residue was placed on the surface, residue-derived intra-POM C was highest when L. rubellus was present and significantly lower in the presence of A. caliginosa. Residue-derived inter-POM C was highest when a mix of both species was present. These results indicate that earthworm species differentially affect incorporation of fresh organic matter into stable microaggregates within macroaggregates, and that interactive effects of earthworm species might have important consequences for the incorporation and protection of C inside of microaggregates within macroaggregates especially when residues are placed on the soil surface.

Differential Growth Properties of Normal and Malignant Esophageal Epithelial Cells: A Possible Cross Talk between Transforming Growth Factor-.BETA.1 and Epidermal Growth Factor Signaling

Comparative cultures of normal and malignant cells are important for understanding the growth properties of tumors. Although many cell lines have been established from esophageal cancers, the growth properties of normal and cancer-derived esophageal epithelial cells have not been compared extensively. We succeeded in establishing an assay system in serum-free conditions for normal human esophageal epithelial cells (HEE cells) and 14 cancer-derived esophageal epithelial cell lines (TE-cell lines). The growth properties of these cells were characterized upon stimulation with various growth-related factors. Among these factors, acidic fibroblast growth factor (aFGF) was the most effective stimulant for both the HEE cells and all the TE-cell lines. Most TE-cell lines required a higher concentration of calcium for their growth than did the HEE cells. Transforming growth factor-beta1 (TGF-beta1) inhibited the growth of HEE cells and 7 TE-cell lines; however, the other 7 TE-cell lines were resistant to the inhibitory effect of TGF-beta1. Interestingly, epidermal growth factor (EGF) had a much greater stimulatory effect on the TGF-beta1-resistant cells than the TGF-beta1-sensitive cells. Although ethanolamine enhanced the growth-promoting ability of EGF or aFGF in the TGF-beta1-sensitive cells, it had no effect on the TGF-beta1-resistant cells. These findings suggested a possible cross talk between TGF-beta1 and EGF signaling, and an important role of ethanolamine in the signaling pathways of growth factors. This serum-free culture system will contribute to clarify the altered signaling pathways of esophageal cancer.

The interaction of ethanol with reconstituted synaptosomal plasma membrane Ca 2+-ATPase

The primary effect of ethanol is on the central nervous system. However, the molecular mechanisms responsible for the physiological symptoms of ethanol intoxication are still unknown. Low concentrations of ethanol were observed to stimulate the activity of the calcium pump from reconstituted synaptosomal plasma membrane Ca 2+-ATPase (PMCA), and ethanol inhibited Ca 2+-ATPase activity at concentrations above 5%. The greatest stimulating effect was obtained with 5% (v/v) ethanol and was lipid-dependent, being 74% when the protein had been reconstituted in phosphatidylcholine (PC) and less when the reconstituted protein had previously been activated by calmodulin or after removal of a 9-kDa autoinhibitory site by controlled trypsinization. Stimulation of the pump by ethanol was lower for the native or trypsin-digested protein in the presence of phosphatidylserine than in PC. These results suggest a direct ethanol–protein interaction, because the activating effect depended on the state of Ca 2+-ATPase (native or truncated, or in presence of calmodulin). The activating mechanism of ethanol may involve opening an autoinhibitory domain located close to the calmodulin binding domain.

Modulation of Gardos channel activity by oxidants and oxygen tension: effects of 1-chloro-2,4-dinitrobenzene and phenazine methosulphate

We compare the effects of 1-chloro-2,4-dinitrobenzene (CDNB) and phenazine methosulphate (PMS) on Gardos channel activity in normal human red cells. Both stimulate channel activity, both are dependent on the presence of extracellular Ca 2+, and neither is affected by inhibitors of protein (de)phosphorylation. Of the two, PMS has a considerably greater effect. In addition, a major difference is that whilst CDNB has a greater stimulatory effect in oxygenated cells, by contrast, PMS is more effective in deoxygenated cells. These actions are correlated with ca. 30% inhibition of the plasma membrane Ca 2+ pump (PMCA) and an increased sensitivity of the Gardos channel to Ca 2+ (EC 50 falling to about 150 nM). These findings are important in understanding how oxidants alter red cell cation permeability and may be relevant to the abnormal permeability phenotype shown by deoxygenated sickle cells.

Differential GH-releasing hormone regulation of GHRH receptor mRNA expression in the rat pituitary.

To understand the capacity of growth hormone-releasing hormone (GHRH) to regulate expression of the GHRH receptor, we studied the effects of GHRH on GHRH receptor mRNA expression in immature and adult rats by use of pituitary cell culture and immunoneutralization approaches. Pituitary cell cultures from neonatal (2-day-old) and adult (70-day-old) rats were treated with GHRH for 4, 24, or 72 h. The effect of GHRH on GHRH receptor mRNA expression depended on the duration of GHRH exposure in both age groups; short-term (4 h) GHRH treatment significantly reduced GHRH receptor mRNA expression (P < 0.05), whereas intermediate treatment (24 h) restored GHRH receptor mRNA to basal levels, and long-term treatment (72 h) stimulated GHRH receptor mRNA expression (P < 0.02). The long-term stimulatory effect of GHRH on GHRH receptor mRNA expression required the presence of serum in the culture medium, and, in the absence of serum, the stimulatory effect was completely abolished. Moreover, the capacity of the pituitary to increase GHRH receptor mRNA expression in response to 72-h GHRH treatment was age dependent, with neonatal pituitaries exhibiting a much greater stimulatory effect than adult pituitaries (P < 0.025). Immunoneutralization of endogenous GHRH significantly reduced GHRH receptor mRNA expression in neonatal (P < 0.004), juvenile (P < 0.003), and mature (P < 0.004) pituitaries compared with age-matched controls. Taken together, these results indicate that GHRH is a potent regulator of GHRH receptor gene expression in immature and mature pituitaries; however, the nature and direction of GHRH regulation of its receptor depend significantly on several variables, including the duration of GHRH exposure, the presence of permissive components in serum, and the developmental stage of the pituitary.

The effects of P2Y receptor agonists and adenosine on prostaglandin production by the guinea-pig uterus.

1. This study has investigated the effects adenosine 5'-triphosphate (ATP), analogues of ATP, uridine 5'-triphosphate (UTP) and adenosine on prostaglandin output from the guinea-pig uterus superfused in vitro, and from guinea-pig endometrium and myometrium cultured for 24 h. 2. ATP, 2-methylthio ATP and adenosine increased the outputs of prostaglandin F(2 alpha) (PGF(2 alpha)) and 6-keto-PGF(1 alpha) (reflecting PGI(2) production), and UTP increased the output of PGF(2 alpha) from the superfused guinea-pig uterus. These findings support the hypothesis that the contractile effects of ATP, 2-methylthio ATP, UTP and adenosine are mediated by prostaglandins. 3. Suramin (a P2 receptor antagonist) and 8-sulphophenyltheophylline (an A receptor antagonist) blocked the stimulatory actions of ATP and adenosine, respectively, on PGF(2 alpha) output, suggesting that ATP acts on P2 receptors (probably of the P2Y type) and adenosine acts on A receptors in the guinea-pig uterus to increase PGF(2 alpha) production. 4. ATP, 2-methylthio ATP, alpha,beta-methylene ATP, beta,gamma-methylene ATP, UTP and adenosine increased the output of PGF(2 alpha) from guinea-pig endometrium and myometrium after 24 h of culture, with a greater stimulatory effect being exerted on the endometrium than on the myometrium. Little or no stimulatory effect was seen after 2 and 8 h of culture. In addition the effects of ATP, ATP analogues, UTP and adenosine on the outputs of PGE(2) and 6-keto-PGF(1 alpha) from cultured endometrium and myometrium were more variable, with both stimulation and inhibition being observed. 5. The stimulatory effects of ATP and adenosine on PGF(2 alpha) output from the endometrium and myometrium were associated with an increase in the prostaglandin synthesizing capacity of both tissues, due probably to an increase in the amount of prostaglandin H synthase present.

Deflazacort increases osteoclast formation in mouse bone marrow culture and the ratio of RANKL/OPG mRNA expression in marrow stromal cells.

Information on precise effects of deflazacort on bone cell function, especially osteoclasts, is quite limited. Therefore, the present study was undertaken to test effects of deflazacort on osteoclast-like cell formation in mouse bone marrow cultures and on the regulation of osteoprotegerin (OPG) and its ligand (RANKL) mRNA expressions by RT-PCR in the ST2 marrow stromal cells. TRAP-positive mononuclear cells increased after the treatment of deflazacort at 10(-9) to 10(-7) M alone for 6 days in a dose-dependent manner. Number of TRAP-positive multi-nucleated cells (MNCs) increased significantly with combined treatment of deflazacort at 10(-7) M and 1,25-(OH)2D3 at 10(-9) M compared to that of cultures treated with 1,25-(OH)2D3 alone (p<0.05). Exposure to deflazacort at 10(-7) M in the presence of 1,25-(OH)2D3 at 10(-9) M in the last 3-day culture had greater stimulatory effect on osteoclast-like cell formation than that of the first 3-day culture did. Deflazacort at 10(-10) -10(-6) M downregulated OPG and upregulated RANKL in mRNA levels in a dose-dependent manner. These observations suggest that deflazacort stimulate osteoclast precursor in the absence of 1,25-(OH)2D3 and enhance differentiation of osteoclasts in the presence of 1,25-(OH)2D3. These effects are, in part, thought to be mediated by the regulation of the expression of OPG and RANKL mRNA in marrow stromal cells.