Nasal Absorption Enhancement of 17β-Estradiol by Dimethyl-β-Cyclodextrin in Rabbits and Rats

A new formulation for nasal administration containing 17-estradiol (E₂) with dimethyl--cyclodextrin (DMC) as a solubilizer and absorption enhancer is described. Nasal administration of this E₂-DMC formulation gave a significantly higher E₂ absorption than an E₂ suspension in both rabbits and rats. Relative to an intravenous injection of the E₂-DMC formulation, absolute bioavailabilities of 94.6 and 67.2% were calculated for the nasal E₂-DMC formulation in rabbits and rats, respectively. Differences in bioavailability may have resulted from differences in experimental animal conditions. The effects on human nasal ciliary activity of the E₂-DMC formulation were studied with an in vitro method. The formulation was found to exert only a minor effect on ciliary beat frequency. Thus, nasal delivery of E₂, using a cyclodextrin inclusion formulation, may have potential for clinical application, e.g., in the therapy of postmenopausal disorders.     

α- and β-Adrenergic Receptors Differentially Modulate the Emission of Spontaneous and Amphetamine-Induced 50-kHz Ultrasonic Vocalizations in Adult Rats

Amphetamine (AMPH) increases adult rat 50-kHz ultrasonic vocalizations, preferentially promoting frequency-modulated (FM) calls that have been proposed to reflect positive affect. The main objective of this study was to investigate a possible noradrenergic contribution to AMPH-induced calling. Adult male Long-Evans rats were tested with AMPH (1 mg/kg intraperitoneal) or saline combined with various systemic pretreatments: clonidine (α2 adrenergic agonist), prazosin (α1 antagonist), atipamezole (α2 antagonist), propranolol, betaxolol, and/or ICI 118,551 (β1/β2, β1, and β2 antagonists, respectively), nadolol (β1/β2 antagonist, peripheral only), or NAD-299 (5HT_1A antagonist). In addition, effects of cirazoline (α1 adrenergic agonist) and cocaine (0.25–1.5 mg/kg intravenous) were studied alone. AMPH-induced calling was suppressed by low-dose clonidine and prazosin. Cirazoline and atipamezole did not significantly affect calling rate. Propranolol, without affecting the call rate, dose dependently promoted ‘flat'' calls under AMPH while suppressing ‘trills,'' thus reversing the effects of AMPH on the ‘call subtype profile.'' This effect of propranolol seemed to be mediated by simultaneous inhibition of CNS β1 and β2 rather than by 5HT_1A receptors. Finally, cocaine elicited fewer calls than did AMPH, but produced the same shift in the call subtype profile. Taken together, these results reveal differential drug effects on flat vs trill vs other FM 50-kHz calls. These findings highlight the value of detailed call subtype analyses, and show that 50-kHz calls are associated with adrenergic α1- and β-receptor mechanisms. These preclinical findings suggest that noradrenergic contributions to psychostimulant subjective effects may warrant further investigation.     

Preparation and characterization of D,L-PLA loaded 17-β-Estradiol valerate by emulsion/evaporation methods

PLA microparticles containing 17-β-estradiol valerate were prepared by an emulsion/evaporation method in order to sustain drug release. This system was characterized concerning particle size, particle morphology and the influence of formulation and processing parameters on drug encapsulation and in vitro drug release. The biodegradation of the microparticles was observed by tissue histological analysis. Scanning electron microscopy and particle size analysis showed that the microparticles were spherical, presenting non-aggregated homogeneous surface and had diameters in the range of 718–880 nm (inert micro-particles) and 3–4 µm (drug loaded microparticles). The encapsulation efficiency was ～80%. Hormone released from microparticles was sustained. An in vivo degradation experiment confirmed that microparticles are biodegradable. The preparation method was shown to be suitable, since the morphological characteristics and efficiency yield were satisfactory. Thus, the method of developed microparticles seems to be a promising system for sustained release of 17-β-estradiol.     

Preparation and characterization of D, L-PLA loaded 17-β-Estradiol valerate by emulsion/evaporation methods

PLA microparticles containing 17-β-estradiol valerate were prepared by an emulsion/evaporation method in order to sustain drug release. This system was characterized concerning particle size, particle morphology and the influence of formulation and processing parameters on drug encapsulation and in vitro drug release. The biodegradation of the microparticles was observed by tissue histological analysis. Scanning electron microscopy and particle size analysis showed that the microparticles were spherical, presenting non-aggregated homogeneous surface and had diameters in the range of 718-880 nm (inert micro-particles) and 3-4 μm (drug loaded microparticles). The encapsulation efficiency was ～80%. Hormone released from microparticles was sustained. An in vivo degradation experiment confirmed that microparticles are biodegradable. The preparation method was shown to be suitable, since the morphological characteristics and efficiency yield were satisfactory. Thus, the method of developed microparticles seems to be a promising system for sustained release of 17-β-estradiol.     

Analysis of Gene Expression Profiles in Largemouth Bass Exposed to 17-β-Estradiol and to Anthropogenic Contaminants that Behave as Estrogens

Novel molecular based methods are being developed to study changes in gene expression in wildlife exposed to anthropogenic chemicals. Gene arrays, in particular, are useful tools that can be used to simultaneously monitor hundreds to thousands of genes within a single experiment, giving an investigator the ability to determine how exposure affects multiple metabolic pathways. These methods are thought to be both sensitive and able to reveal biochemical mechanisms of action. A largemouth bass (LMB) array containing 132 genes has been designed to study the impact of gene expression in male fish exposed to 17-beta estradiol or to the compounds 4-nonylphenol (4-NP) or 1,1-dichloro-2, 2-bis (p-chlorophenyl) ethylene (p,p'-DDE). The results of these experiments demonstrate distinct gene expression patterns in LMB exposed to these compounds.     

Design, synthesis and in vitro antibacterial and antifungal activities of some novel spiro[azetidine-2,3��-indole]-2,4(1��H)-dione

The present study deals with the synthesis of novel spiro[azetidine-2,3′-indole]-2′,4(1′H)-dione derivative from the reactions of 3-(phenylimino)-1,3-dihydro-2H-indol-2-one derivatives with chloracetyl chloride in the presence of triethylamine (TEA). All the compounds were characterized using IR, ¹H-NMR, MS, and elemental analysis. They were screened for their antibacterial and antifungal activities. The bacterial strains used were Gram-positive Staphylococcus aureus (MTCC-96) and Gram-negative Escherichia coli (MTCC-521) and Pseudomonas aeruginosa (MTCC-647). The antifungal screening was done on Candida albicans (MTCC-183) and Asperigillus niger (MTCC-343) fungal strains. Results revealed that, compounds (7a), (7b), (7c), (7d), and (7e) showed very good activity with MIC value of 6.25–12.5 μg/ml against three evaluated bacterial strains and the remaining compounds showed good to moderate activity comparable to standard drugs as antibacterial agents. Compounds (7c) and (7h) displayed equipotent antifungal activity in comparison to standard drugs. Amoxicillin, gentamycin, and streptomycin were used as standard drugs for antibacterial activity while fluconazole and itraconazole were used as standard drugs for antifungal activity. Structure–activity relationship study of the compounds showed that the presence of electron withdrawing group substitution at 5′ and 7′ positions of indoline ring and on ortho or para position of phenyl ring increases both antibacterial and antifungal activity of the compound. Henceforth, our findings will have a good impact on chemists and biochemists for further investigations in search of spiro-fused antimicrobial agents.     

Determination of oestrone, 17α- and 17β-oestradiol in bovine aqueous humor using gas chromatography-negative ion chemical ionization mass spectrometry

Perfluorotolyl (PFT)-ether and perfluorotoly-trimethylsilyl (PFT-TMS) ether derivatives of oestrone, 17α- and 17β-oestradiol were prepared under phase transfer conditions. The former derivatives under negative ion chemical ionization conditions gave significant ions in the mass spectrometer but 17α- and 17α-oestradiol gave poor resolution. However, the PFT-TMS derivatives of 17α- and 17β-oestradiol showed good resolution. These derivatives were used for the analysis of oestrogens in bovine aqueous humour, vitreous humour and retina. The mean (±SEM) concentrations of oestrone in bovine aqueous humour (n=18), vitreous humour (n=18) and bovine retina (n=4) were 0.47±0.11, 0.46±0.14 and 1.10±0.24 ng.ml⁻¹, respectively. 17α-Oestradiol was detected in 16 out of 18 samples of bovine aqueous humour and vitreous humour and the mean (±SEM) concentrations were 0.30±0.10 and 0.08±0.02 ng.ml⁻¹, respectively. The mean (±SEM) concentration of 17β-oestradiol in aqueous humour (n=7) and vitreous homour (n=11) 0.83±0.26 ng ml⁻¹ and 0.39±0.09 ng ml⁻¹, respectively. In retina the concentrations of both steroids were below the detection limit.     

Inhibition of microRNA-17-5p reduces the inflammation and lipid accumulation,and up-regulates ATP-binding cassette transporterA1 in atherosclerosis

Atherosclerosis (AS) is a chronic inflammatory disease of the arterial wall. Macrophages are considered to be closely associated with the development and progression of AS. However, the precise mechanism of miR-17-5p in the macrophages under AS remains incompletely clarified. This study investigated the regulatory effect of miR-17-5p on the inflammation and lipid accumulation in mouse macrophages both in vivo and in vitro. It was found that miR-17-5p was highly expressed with lowered ATP-binding cassette transporterA1 (ABCA1) level in the peripheral blood leucocytes (PBLs) of AS patients. Moreover, the level of miR-17-5p was up-regulated in the macrophages of ApoE^?/? mice fed with a high-cholesterol diet. Furthermore, we injected miR-17-5p antagomir into AS mice or transfected miR-17-5p inhibitors into mouse macrophage RAW264.7 cells. Results showed that downregulation of miR-17-5p significantly reduced the production of inflammatory cytokines, inhibited the lipid accumulation and up-regulated ABCA1, and activated peroxisome proliferator-activated receptor (PPAR) γ/Liver X receptor (LXR) α signaling pathway. Additionally, ABCA1 was found to be a target of miR-17-5p by directly binding to 3′-untranslated region (3′-UTR) of its mRNA. Our study indicates a novel regulatory mechanism for miR-17-5p by interacting with ABCA1, which could be a therapy-target for the treatment of AS.     

BAER-101, a selective potentiator of α2- and α3-containing GABAA receptors,fully suppresses spontaneous cortical spike-wave discharges in Genetic Absence Epilepsy Rats from Strasbourg (GAERS)

BAER-101 (formerly AZD7325) is a selective partial potentiator of α2/3-containing γ-amino-butyric acid A receptors (GABAARs) and produces minimal sedation and dizziness. Antiseizure effects in models of Dravet and Fragile X Syndromes have been published. BAER-101 has been administered to over 700 healthy human volunteers and patients where it was found to be safe and well tolerated. To test the extent of the antiseizure activity of BAER-1010, we tested BAER-101 in the Genetic Absence Epilepsy Rats from Strasbourg (GAERS) model, a widely used and translationally relevant model. GAERS rats with recording electrodes bilaterally located over the frontal and parietal cortices were used. Electroencepholographic (EEG) signals in freely moving awake rats were analyzed for spike-wave discharges (SWDs). BAER-101 was administered orally at doses of 0.3–100 mg/kg and diazepam was used as a positive control using a cross-over protocol with a wash-out period between treatments. The number of SWDs was dose-dependently reduced by BAER-101 with 0.3 mg/kg being the minimally effective dose (MED). The duration of and total time in SWDs were also reduced by BAER-101. Concentrations of drug in plasma achieved an MED of 10.1 nM, exceeding the K_i for α2 or α3, but 23 times lower than the K_i for α5-GABAARs. No adverse events were observed up to a dose 300× MED. The data support the possibility of antiseizure efficacy without the side effects associated with other GABAAR subtypes. This is the first report of an α2/3-selective GABA PAM suppressing seizures in the GAERS model. The data encourage proceeding to test BAER-101 in patients with epilepsy.