BACKGROUNDGlucocorticoid is speculated to be able to have Aspergillus fumigatus (A. fumigatus) being more susceptible to reactive oxygen species (ROS) by inhibiting Afyap1, the transcription factor activating protein-1 (AP-1) homologue in A. fumigatus, which may provide a clue to expand the clinical use of glucocorticoid in patients with fungal infections. In this study, we used dexamethasone to determine the direct effect on oxidative killing susceptibility of A. fumigatus in vitro, as well as the expression level of Afyap1 gene and its target genes (catalase and superoxide dismutase (SOD) genes).

METHODSA. fumigatus spores were treated with different concentrations (0, 0.02, 0.2 mg/ml) of glucocorticoids and assigned to four groups (A: 0.5 hour, B: 2 hours, C: 7 hours, D: 16 hours) according to the time of treatment. The H2O2 oxidative killing assay was done, using the standard method-spot test, in each group of A. fumigatus. We measured the oxidative killing susceptibility as well as the expression level of the gene Afyap1, CATA, SOD1 and SOD2 in A. fumigatus at each group. The antifungal susceptibility to itraconazole and amphotericin B in each group of A. fumigatus was also measured with M38-A2 method.

RESULTSThe oxidative killing susceptibility of A. fumigatus was increased, consistent with the reduction of Afyap1, CATA, SOD1 and SOD2 gene expression level after being treated with dexamethasone for 0.5 hours. However, these observations were disappeared along with being treated for longer time. The antifungal susceptibility to itraconazole and amphotericin B in the A. fumigatus strains treated with dexamethasone indicated no change, compared with those without dexamethasone treatment.

CONCLUSIONDexamethasone can have A. fumigatus being more susceptible to ROS when treated for shorter period (0.5 to 2 hours) via the reduction of Afyap1 gene expression as well as the down-stream enzyme-coding gene expression.

Aspergillus fumigatus ; drug effects ; genetics ; metabolism ; Dexamethasone ; pharmacology ; Fungal Proteins ; genetics ; metabolism ; Hydrogen Peroxide ; pharmacology

The effect of dexamethasone (10(-5)M) and epinephrine (10(-6)M) on the biosynthesis of metallothionein (MT) in the perfused rat liver was investigated. MT synthesis was determined by measuring the incorporation of 14C-L-aspartic acid into liver MT fraction after the perfusion for five hours of isolated liver by artificial blood containing 14C-L-U-aspartic acid (0.2uci) with dexamethasone or epinephrine. MT was isolated by Sephadex G-75 column chromatography and DEAE Sephadex column chromatography. Incorporation of radioactive 14C into the MT fraction of perfused liver cytosol (9.0grams of liver) from dexamethasone treated, epinephrine treated and control groups were, respective1y, 0.72, 0.34 and 0.33% of total radioactivity infused. Total protein content in the MT fraction of liver perfused with dexamethasone and epinephrine were 0.80, 0.64mg/g liver compared to 0.52mg/g liver in the control. MT, a protein having a high content of cystein and metals is synthesized in the perfused rat liver and its induction is stimulated by dexamethasone, while epinephrine increased the accumulation of Zn in the MT fraction of the perfused rat liver. The present experiment confirms that MT synthesis and degradation are somewhat regulated by glucocorticoid hormone and epinephrine.
Animal ; Dexamethasone/pharmacology* ; Epinephrine/pharmacology* ; Female ; In Vitro ; Liver/drug effects ; Liver/metabolism* ; Metalloproteins/metabolism* ; Metallothionein/metabolism* ; Perfusion ; Rats ; Zinc/metabolism

The aim of this study was to explore the effect of IL-29 on the progression of airway allergic disease by detecting the level of IL-29 in airway allergic cell models stimulated by house dust mite (HDM) in the presence or absence of dexamethasone (DEX). The same batch of human bronchial epithelial cells in exponential growth phase was randomly divided into five groups: blank group (A), 300 ng/mL HDM group (B), 1000 ng/mL HDM group (C), 3000 ng/mL HDM group (D), and 300 ng/mL HDM+100 ng/mL DEX group (E). The IL-29 mRNA expression was detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The IL-29 protein expression in cell suspension was detected by ELISA. The results showed that after stimulation with HDM for 24 h, the expression of IL-29 was increased significantly, and after co-stimulation with HDM and DEX for 24 h, the expression of IL-29 in group E was significantly lower than that in the groups stimulated by HDM alone but higher than that in the group A. The differences between the different groups were significant (F=132.957, P<0.01). Additionally, the higher the concentration of HDM was, the more significant the increase in the IL-29 expression was. In conclusion, IL-29 may play a role in the progression of airway allergic disease including asthma.
Adult ; Animals ; Bronchi ; cytology ; drug effects ; metabolism ; Cells, Cultured ; Dexamethasone ; pharmacology ; Epithelial Cells ; drug effects ; metabolism ; Humans ; Interleukins ; metabolism ; Mites

OBJECTIVE: To study the dexamethasone pharmacokinetics of human inner ear perilymph under different drug administration using computer simulations. METHOD: The dexamethasone pharmacokinetics in guinea pigs inner ear perilymph under an intratympanic application with high-performance liquid chromatography. Dexamethasone pharmacokinetics in the guinea pigs cochlear fluid were simulated with a computer model, the Washington University Cochlear Fluids Simulator, version 1.6 and the best Simulations parameters to the experimental data could be obtain. With best Simulations parameters based on the experimental data, seven kinds application protocols were designed for human inner ear perilymph. RESULT: After an intratympanic application dose of 0.5% dexamethasone 150 ml in guinea pigs, the T(1/2K) was (2.918 +/- 0.089) h, and Cmax was (231.25 +/- 6.89) microg/L. The best Simulations parameters were that concentration of the dexamethasone 21-Phosphate disodium salt was 0.5% and the formula weight was 516, as well as drug diffusion coefficient was 0.6939 x 10(-5) cm2/s and round window permeability was 2.2 x 10(-11) cm/s while drug clearance half time was 175 minutes and scala tympaniscala vestibuli communication was 45 minutes. After an intratympanic application dose of 0.5% dexamethasone 500 mL, which the applied drug stayed in contact with the round window membrane for 15, 30, 60 and 120 minutes, the Cmax was 32.8, 64.3, 122.6 and 203.3 microg/L and the AUC was 116.5, 229.1, 423.6 and 759.2 microg/(h x L), respectively. After an intratympanic application dose of 0.5%, 1%, 2% and 4% dexamethasone 500 ml, which the applied drug stayed in contact with the round window membrane for 30 minutes respectively, the Cmax was 64.3, 127.3, 255.4 and 575.6 microg/L respectively and the AUC was 229.1, 462.8, 920.59 and 1525.2 microg/(h x L), respectively. CONCLUSION The dexamethasone pharmacokinetics in human inner ear perilymph by computer simulations was reported. As the time contact with the round window membrane increased, the inner ear perilymph concentration of dexamethasone increased. As the concentration of dexamethasone increased, the inner ear perilymph concentration of drug increased.
Animals ; Computer Simulation ; Dexamethasone ; pharmacokinetics ; Ear, Inner ; metabolism ; Guinea Pigs ; Humans ; Perilymph ; metabolism ; Round Window, Ear ; metabolism

Animals ; Dexamethasone ; administration & dosage ; metabolism ; pharmacokinetics ; Drug Administration Routes ; Ear, Middle ; Guinea Pigs ; Nanoparticles ; administration & dosage

BACKGROUNDThe efficacies of current treatments for invasive aspergillus (IA) are unsatisfactory and new therapeutic targets or regimens to treat IA are urgently needed. Previous studies have indicated that the ability of conidia to invade host cells is critical in IA development and fibronectin has a hand in the conidia adherence process. In the clinical setting, many patients who receive glucocorticoid for extended periods are susceptible to Aspergillus fumigatus (A. fumigatus) infection, for this reason we investigated the effect of glucocorticoid on conidia invasiveness by comparing the invasiveness of A. fumigatus conidia in the type II human alveolar cell line (A549) cultured with different concentrations of dexamethasone. We also explored the relationships between dexamethasone and fibronectin expression.

METHODSFollowing culture with anti-fibronectin antibodies and/or dexamethasone, type II human alveolar A549 cells were infected with conidia of A. fumigatus. After 4 hours, the extracellular free conidia were washed away and the remaining immobilized conidia were released using Triton-X 100 and quantified by counting the colony-forming units. The invasiveness of conidia was measured by calculating the invasion rate (%). The transcription of the fibronectin gene in cells cultured with different concentrations of dexamethasone for 24 hours was tested by fluorogenic quantitative RT-PCR while the expression of fibronectinin cells cultured for 48 hours was tested by Western blotting and immunocytochemistry.

RESULTSA significant reduction in the invasiveness of conidia was seen in the cells cultured with anti-fibronectin antibody ((14.42 ± 1.68)% vs. (19.17 ± 2.53)%, P < 0.05), but no significant difference was observed in cells cultured with a combination of anti-fibronectin antibody and dexamethasone (6.37 ± 10(-5) mol/L). There was no correlation between the dexamethasone concentration and the invasiveness of conidia after dexamethasone pretreatment of cells for 4 hours. In contrast, after pretreated for 24 hours, the invasiveness of conidia in the presence of 6.37×10(-5) mol/L dexamethasone ((24.66 ± 2.41)%) was higher than for the control ((19.17 ± 2.53)%) and the 0.25×10(-5) mol/L group ((19.93 ± 3.06)%), and the invasiveness in the 1.27×10(-5) mol/L group ((22.47 ± 2.46)%) was also higher than in the control, P < 0.05. The relative transcripts of the fibronectin gene after exposure to 6.37×10(-5) mol/L dexamethasone (9.19×10(-3)±1.2×10(-3)) was higher than for the control (4.61×10(-3)± 1.54×10(-3)) and the 0.25×10(-5) mol/L group (6.20×10(-3)± 1.93×10(-3)), and expression in the 1.27×10(-5) mol/L group (7.94×10(-3)± 2.24×10(-3)) was also higher than for the control, P < 0.05. High concentrations of dexamethasone promoted fibronectin production after culture for 48 hours.

CONCLUSIONSDexamethasone can increase invasiveness of A. fumigatus conidia by promoting fibronectin expression. This may partially explain why patients who are given large doses of glucocorticoids for extended periods are more susceptible to A. fumigatus infection.

Aspergillus fumigatus ; pathogenicity ; Cell Line, Tumor ; Dexamethasone ; pharmacology ; Fibronectins ; genetics ; metabolism ; Gene Expression ; drug effects ; Humans

OBJECTIVETo investigate the effect of bortezomib (Bor) alone or in combination with As(2)O(3) (ATO) and/or dexamethasone (DXM) on proliferation and apoptosis in KM3 human multiple myeloma cell line KM3.

METHODSKM3 cells were cultured with different concentrations of Bor and ATO and/or DXM in combination or Bor, ATO, DXM alone for different times. Cell proliferation was assayed by MTT assay, and IC(50) was calculated. Cell morphology was observed with light and electric microscopy. The agarose gel electrophoresis was used to evaluate DNA content, and the flow cytometry was used to exam Annexin V-FITC/PI stain.

RESULTSBor, ATO and DXM inhibited KM3 cell proliferation in a time-and dose-dependent manner with the IC(50) of 0.27, 3.10 and 8.01 micromol/L, respectively. The inhibition rate of KM3 cells by Bor plus ATO and DXM was significantly higher than Bor plus ATO or DXM \[(34.51 +/- 0.51)% vs (25.39 +/- 0.90)% and (34.51 +/- 0.51)% vs (23.80 +/- 0.78)% respectively\]. Typical morphology for apoptosis and DNA ladder were observed in KM3 cell treated with 0.25 micromol/L Bor for 48 h, by Annexin V positivity. The apoptosis rate induced by Bor plus both ATO and DXM was higher than that induced by Bor plus DXM.

CONCLUSIONBor can inhibit the proliferation and induce apoptosis of KM3 cells. Bor enhances the inhibitory effect of ATO and DXM on the growth of KM3 cell. ATO enhances the apoptosis effects of Bor and DXM on KM3 cells.

Apoptosis ; drug effects ; Boronic Acids ; pharmacology ; Bortezomib ; Cell Line, Tumor ; Dexamethasone ; pharmacology ; Humans ; Multiple Myeloma ; metabolism

Bone is a dynamic tissue which is constantly remodelled by subsequent cycles of bone resorption and formation. Glucocorticoid and vitamine D3 are known as regulating substances in bone metabolism. In vitro experiments using bone tissue, it was suggested that glucoccorticoid inhibits bone resorption, whereas the effect of glucocorticoid on bone formation are complex- increasing or decreasing effect. The active form of vitamin D3, 1,25-dihydroxycholecalciferol [1.25-(OH)2D3], has been reported to stimulate osteoblastic activities including the production of ALP, type I collagen, and osteoclacin. The purpose of this study was to evaluate the effect of admixture of vitamin D3 and dexamethasone, one of glucocorticoids, on osteblastic cell line(MC3T#-E1). Alkaline phosphatase(ALP) and MTT assay were conducted in the cultivated cells with 1, 10, 100nm/ml of 1,25-(OH)2D3 and/or 10nM/ml, 100nM/ml, 1micrometer/ml of dexamethasone. The observed results were as follows. 1. The activity of osteoblastic cells with 1micrometer/ml of dexamethasone was significantly increased at 1-day cultivation with comparison to control group, but was decreased afterwards. But the activity of ALP was greatest in 1micrometer/ml of dexamethasone and increased with time lapsed. 2. The activity of osteoblastic cells with vitamin D3 was significantly increased dose-dependently at 1-day cultivation, but was significantly decreased in 10nM/ml or 100nM/ml at 2-day or 3-day cultivation, and was greatest in 100nM/ml at 3-day cultivation. 3. In case of admixture of dexamethasone and vitamin D3 at 2-day cultivation, but was increased again at 3-day cultivation, which was greater than that in control or dexamethasone only group. The activity of ALP was decreased at 1-day cultivation, but was increased in the admixture of 10nM/ml or 100nM/ml of dexamethasone with 100nM/ml of vitamin D3 at 2-day cultivation, and was again decreased at 3-day cultivation.
Bone and Bones ; Bone Resorption ; Calcitriol ; Cholecalciferol ; Collagen Type I ; Dexamethasone* ; Glucocorticoids ; Metabolism ; Osteoblasts* ; Osteogenesis ; Vitamins*

Chronic psychological stress can promote vascular diseases, such as hypertension and atherosclerosis. This study aims to explore the effects and mechanism of chronic psychological stress on aortic medial calcification (AMC). Rat arterial calcification model was established by nicotine gavage in combination with vitamin D₃ (VitD₃) intramuscular injection, and rat model of chronic psychological stress was induced by humid environment. Aortic calcification in rats was evaluated by using Alizarin red staining, aortic calcium content detection, and alkaline phosphatase (ALP) activity assay. The expression levels of the related proteins, including vascular smooth muscle cells (VSMCs) contractile phenotype marker SM22α, osteoblast-like phenotype marker RUNX2, and endoplasmic reticulum stress (ERS) markers (GRP78 and CHOP), were determined by Western blot. The results showed that chronic psychological stress alone induced AMC in rats, further aggravated AMC induced by nicotine in combination with VitD₃, promoted the osteoblast-like phenotype transformation of VSMCs and aortic ERS activation, and significantly increased the plasma cortisol levels. The 11β-hydroxylase inhibitor metyrapone effectively reduced chronic psychological stress-induced plasma cortisol levels and ameliorated AMC and aortic ERS in chronic psychological stress model rats. Conversely, the glucocorticoid receptor agonist dexamethasone induced AMC, promoted AMC induced by nicotine combined with VitD₃, and further activated aortic ERS. The above effects of dexamethasone could be inhibited by ERS inhibitor 4-phenylbutyrate. These results suggest that chronic psychological stress can lead to the occurrence and development of AMC by promoting glucocorticoid synthesis, which may provide new strategies and targets for the prevention and control of AMC.
Rats ; Animals ; Glucocorticoids/metabolism* ; Rats, Sprague-Dawley ; Nicotine/metabolism* ; Hydrocortisone/metabolism* ; Muscle, Smooth, Vascular ; Dexamethasone/metabolism* ; Vascular Calcification/metabolism* ; Myocytes, Smooth Muscle/metabolism* ; Cells, Cultured

OBJECTIVETo investigate the potential of synovial mesenchymal stem cells (SMSC) in osteogenic differentiation.

METHODSSMSC were obtained by limited dilution method and expanded to culture in 25-milliliter flasks. The attached cells were treated with inductive medium containing dexamethasone, glycerophosphate and vitamin C at 3rd passage SMSC. The mineralized nodule was stained by Von Kossa method. The expression of ALP and osteopontin were detected by histochemical, immunohistological staining technique, respectively, while the expression of cbfa1 mRNA by RT-PCR.

RESULTSPure SMSC which were of spindle shape and star shape, uniform in size, could be induced to pleomorphism osteoblast in vitro, which were intensive positive in ALP and osteopontin. The expression of cbfa1 mRNA were also verified by RT-PCR and the polygonal cells formed nodular structure at 4 weeks. All these were coincident with the characters of osteoblast.

CONCLUSIONSMSC can be purified and induced into osteoblast in vitro.

Ascorbic Acid ; Cell Differentiation ; Dexamethasone ; Glycerophosphates ; Humans ; Mesenchymal Stromal Cells ; Osteoblasts ; Osteogenesis ; Osteopontin ; metabolism ; RNA, Messenger ; metabolism