Isolation of the 3ß-HSD promoter from Digitalis ferruginea subsp. ferruginea and its functional characterization in Arabidopsis thaliana.

BACKGROUND: Although members of the SDR gene family (short chain dehydrogenase) are distributed in kingdom of life, they have diverse roles in stress tolerance mechanism or secondary metabolite biosynthesis. Nevertheless, their precise roles in gene expression or regulation under stress are yet to be understood. METHODS: As a case study, we isolated, sequenced and functionally characterized the 3ß-HSD promoter from Digitalis ferruginea subsp. ferruginea in Arabidopsis thaliana. RESULTS: The promoter fragment contained light and stress response elements such as Box-4, G-Box, TCT-motif, LAMP element, ABRE, ARE, WUN-motif, MYB, MYC, W box, STRE and Box S. The functional analysis of the 3ß-HSD promoter in transgenic Arabidopsis seedlings showed that the promoter was expressed in cotyledon and root elongation zone in 2 days' seedlings. However, this expression was extended to hypocotyl and complete root in 6 days' seedlings. In 20 days-old seedlings, promoter expression was distributed to the whole seedling including hydathodes aperture, vascular bundle, shoot apical meristem, trichomes, midrib, leaf primordia, hypocotyl and xylem tissues. Further, expression of the promoter was enhanced or remained stable under the different abiotic stress conditions like osmotic, heat, cold, cadmium or low pH. In addition, the promoter also showed response to methyl jasmonate (MeJA) application. The expression could not be induced in wounded cotyledon most likely due to lack of interacting elements in the promoter fragment. CONCLUSIONS: Taken together, the 3ß-HSD promoter could be a candidate for the development of transgenic plants especially under changing environmental conditions.

Plastidial Expression of 3ß-Hydroxysteroid Dehydrogenase and Progesterone 5ß-Reductase Genes Confer Enhanced Salt Tolerance in Tobacco.

The short-chain dehydrogenase/reductase (SDR) gene family is widely distributed in all kingdoms of life. The SDR genes, 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and progesterone 5-ß-reductases (P5ßR1, P5ßR2) play a crucial role in cardenolide biosynthesis pathway in the Digitalis species. However, their role in plant stress, especially in salinity stress management, remains unexplored. In the present study, transplastomic tobacco plants were developed by inserting the 3ß-HSD, P5ßR1 and P5ßR2 genes. The integration of transgenes in plastomes, copy number and transgene expression at transcript and protein level in transplastomic plants were confirmed by PCR, end-to-end PCR, qRT-PCR and Western blot analysis, respectively. Subcellular localization analysis showed that 3ß-HSD and P5ßR1 are cytoplasmic, and P5ßR2 is tonoplast-localized. Transplastomic lines showed enhanced growth in terms of biomass and chlorophyll content compared to wild type (WT) under 300 mM salt stress. Under salt stress, transplastomic lines remained greener without negative impact on shoot or root growth compared to the WT. The salt-tolerant transplastomic lines exhibited enhanced levels of a series of metabolites (sucrose, glutamate, glutamine and proline) under control and NaCl stress. Furthermore, a lower Na+/K+ ratio in transplastomic lines was also observed. The salt tolerance, mediated by plastidial expression of the 3ß-HSD, P5ßR1 and P5ßR2 genes, could be due to the involvement in the upregulation of nitrogen assimilation, osmolytes as well as lower Na+/K+ ratio. Taken together, the plastid-based expression of the SDR genes leading to enhanced salt tolerance, which opens a window for developing saline-tolerant plants via plastid genetic engineering.

Synthesis, molecular docking studies, and biological evaluation of novel alkyl bis(4-amino-5-cyanopyrimidine) derivatives.

A series of bis(4-amino-5-cyano-pyrimidines) was synthesized and evaluated as dual inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). To further explore the multifunctional properties of the new derivatives, their antioxidant and antibacterial activities were also tested. The results showed that most of these compounds could effectively inhibit AChE and BChE. Particularly, compound 7c exhibited the best AChE inhibitory activity (IC50 = 5.72 ± 1.53 µM), whereas compound 7h was identified as the most potent BChE inhibitor (IC50 = 12.19 ± 0.57 µM). Molecular modeling study revealed that compounds 7c, 7f, and 7b showed a higher inhibitory activity than that of galantamine against both AChE and BChE. Anticholinesterase activity of compounds 7h, 7b, and 7c was significant in vitro and in silico for both enzymes, since these compounds have hydrophobic rings (Br-phenyl, dimethyl, and methoxyphenyl), which bind very well in both sites. In addition to cholinesterase inhibitory activities, these compounds showed different levels of antioxidant activities. Indeed, in the superoxide-dimethyl sulfoxide alkaline assay, compound 7j showed very high inhibition (IC50 = 0.37 ± 0.28 µM). Also, compound 7l exhibited strong and good antibacterial activity against Staphylococcus epidermidis and Staphylococcus aureus, respectively. Taking into account the results of biological evaluation, further modifications will be designed to increase potency on different targets. In this study, the obtained results can be a new starting point for further development of multifunctional agents for the treatment of Alzheimer's disease.

A rapid access to novel and diverse 3-oxothiazolo[3,2-c]pyrimidine-8-carboxylates using multicomponent Mannich cyclisation reactions.

An efficient synthesis of novel 3-oxotetrahyrothiazolo[3,2-c]pyrimidine-8-carboxylate derivatives was developed by reacting 4-oxothiazolidines with formaldehyde and amines under very mild conditions. A variety of novel thiazolopyrimidine carboxylates were attained rapidly by double Mannich/intramolecular cyclisation reactions in very good yields.

Quercetin- and caffeic acid-functionalized chitosan-capped colloidal silver nanoparticles: one-pot synthesis, characterization, and anticancer and antibacterial activities.

The presented study comprises the one-pot synthesis and the characterization of quercetin- and caffeic acid-functionalized chitosan-capped colloidal silver nanoparticles (Ch/Q- and Ch/CA-Ag NPs), and their antibacterial and anticancer activities. The formation of Ch/Q- and Ch/CA-Ag NPs has been confirmed by ultraviolet-visible (UV-vis) spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, and transmission electron microscopy (TEM). The characteristic surface plasmon resonance (SPR) absorption band has been found at 417 and 424 nm for Ch/Q- and Ch/CA-Ag NPs, respectively. The formation of a chitosan shell comprising quercetin and caffeic acid, which surround the colloidal core Ag NPs, was confirmed by UV-vis, and FTIR analyses, and monitored by TEM microscopy. The size of nanoparticles has been determined as 11.2 and 10.3 nm for Ch/Q- and Ch/CA-Ag, respectively. The anticancer activity of Ch/Q- and Ch/CA-Ag NPs has been evaluated against U-118 MG (human glioblastoma) and ARPE-19 (human retinal pigment epithelium) cells. Both NPs showed anticancer activity, but Ch/Q-Ag NPs seemed to be more effective on cancer cell lines (U-118 MG) in comparison to healthy ones (ARPE-19). Furthermore, the antibacterial activity of Ch/Q- and Ch/CA-Ag NPs against Gram-negative (P. aeruginosa and E. coli) and Gram-positive (S. aureus and S. epidermidis) bacteria was determined, and dose-dependent antibacterial effects were found.

Trimetazidine has Promising Preventive Effects on Experimental Chronic Pancreatitis Rat Model.

BACKGROUND: It was aimed to evaluate the preventive efficacy of trimetazidine in an experimental chronic pancreatitis rat model. METHODS: Chronic pancreatitis model was accomplished with caerulein and alcohol administration. In the study, 40 female Sprague Dawley rats were randomized into 5 groups containing 8 animals in each. Group 1 (chronic pancreatitis); group 2 (chronic pancreati- tis+low-dose trimetazidine group); group 3 (chronic pancreatitis+high-dose trimetazidine group); group 4 (placebo group (chronic pancreatitis + saline)); group 5 (sham group). 24 hours after the last injection, all animals were sacrificed. Tumor necrosis factor-alpha, transforming growth factor-ß, malondialdehyde, and glutathione peroxidase levels were tested in blood samples. Histopathologic exam- inations were conducted by a senior pathologist who was unaware of the group allocations. RESULTS: Results of biochemical parameters of the trimetazidine groups (groups 2 and 3) were significantly favorable compared with the chronic pancreatitis group (group 1) (P < .05). The difference between the low-dose- and the high-dose trimetazidine group (group 3) was significant in terms of blood tests (P < .05). The difference between the low-dose trimetazidine group and the chronic pancreatitis group was not significant in terms of histopathologic scores (P > .05); however, the difference was significant between the high-dose trimetazidine group and the chronic pancreatitis group (P < .05). CONCLUSIONS: To the best of our knowledge, this current research is the first study that evaluates trimetazidine's efficacy in the chronic pancreatitis rat model. Trimetazidine has affirmative preventive properties in the chronic pancreatitis course.

Research note: modeling of the water quality of Büyüksu Stream, Bolu, Turkey.

The QUAL2E model was applied to the Büyüksu Stream, Bolu, Turkey, to predict the effect of conventional and industrial pollutant sources on stream quality. The model parameters were conditioned using data from eight sampling stations on a 24-km stretch of the stream during a steady-state period between May 2003 and June 2004. The higher biochemical oxygen demand (> 50 mg/L) and lower dissolved oxygen concentrations measured (< 4.0 mg/L), with their expected values, indicated that the water quality of the Büyüksu Stream was threatened by industrial and domestic pollution. The oxygen-sag curves obtained by the model calculations proved that discharges of the Bolu City wastewater and wood-processing plant effluent were the two primary pollutant sources affecting the stream. Results suggested that the conditioned model can be used as a tool to show the effects of pollutant sources on the Büyüksu Stream and to assess improvements expected by reducing the contribution of pollutant sources.

A cycloaddition product of a chiral maleimide: 4-{(3aS*,6aS*)-4,6-dioxo-1-phenyl-5-[(1R)-1-phenyl-ethyl]-1,3a,4,5,6,6a-hexa-hydro-pyrrolo[3,4-c]pyrazol-3-yl}phenyl acetate.

In the title mol-ecule, C(27)H(23)N(3)O(4), the two central five-membered rings form a dihedral angle of 63.66 (4)°. The absolute configuration was determined by analysis of Bijvoet pairs based on resonant scattering of light atoms, yielding a Hooft parameter y = -0.10 (7).

(3aR,6aR)-1-Phenyl-5-[(R)-1-phenyl-ethyl]-3-[4-(trifluoro-meth-yl)phen-yl]-1,6a-dihydro-pyrrolo[3,4-c]pyrazole-4,6(3aH,5H)-dione.

In the title mol-ecule, C(26)H(20)F(3)N(3)O(2), the two central five-membered rings form a dihedral angle of 62.94 (8)°. The absolute configuration was determined by analysis of Bijvoet pairs based on resonant scattering of light atoms, yielding a Hooft parameter y = -0.05 (11). Notable intra- and inter-molecular contacts include C-H⋯O and C-H⋯π(arene) hydrogen bonds.

Chitosan/poly(ethylene glycol)/hyaluronic acid biocompatible patches obtained by electrospraying.

Electrospray is a promising technique to scale-up production of microparticles and nanoparticles. In this study, electrospraying was used in order to produce candidate biopatches (CPH) by using chitosan, poly(ethylene glycol) (PEG) and hyaluronic acid (HA). Four different ratios of polymer blend compositions (CPH1, CPH2, CPH3 and CPH4) were tested by dissolving in 2% acetic acid solution (Ac.A.). The HA amount in each blend was kept the same to designate the optimum surface with different chitosan/PEG ratios for electrospray process. Fourier-transform infrared (FTIR) microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM) studies showed that obtained patches had highly adhesive surfaces with the aid of heterogeneously distributed micro- and nano-particles. Additionally, video images of FTIR microscopy and AFM images proved that all surfaces have similar heterogeneity except CPH2. The most homogenous surface was obtained by CPH3. Patches were directly subjected to antibacterial tests against ten different types of gram positive and gram negative bacteria using disc diffusion assay (Kirby-Bauer method). Extraordinarily there was no antibacterial property of patches coated with microparticles. Finally, biocompatibility studies were performed by using mouse fibroblast L929 cell lines (ATTC number CCL-1) to test cell adhesion and proliferation properties of the patches. Results of 72 h viability tests proved the electrospray of ternary blends had displayed good biocompatibility; in particular, CPH3 had the highest cell viability.

Efficacy of tocilizumab treatment in cerulein-induced experimental acute pancreatitis model in rats.

BACKGROUND/AIMS: Acute pancreatitis (AP) is a disease that can cause local and systemic complications that may have high morbidity and mortality. Currently, there is not any specific treatment for AP. In this study, we created an experimental model of AP in rats, and we aimed to demonstrate the histological effectiveness of tocilizumab treatment that antagonizes interleukin-6 (IL-6), one of the key cytokines in the development of AP. MATERIALS AND METHODS: Forty-eight rats were divided into six groups for this study. AP model was created by subcutaneous injections of cerulein (20 µg/kg) four times at 1-h intervals. Tocilizumab 4 mg/kg was administered to one of the treatment groups and 8 mg/kg to the other treatment group intraperitoneally. The effects of tocilizumab were revealed by examining pancreatic tissue of the rats histopathologically according to the Schonberg scoring system. RESULTS: A comparison between tocilizumab treatment group and AP control group provides statistically significant improvement in AP (p<0.0001). Furthermore, the dose of 8 mg/kg is shown to be more effective than 4 mg/kg (p=0.004). CONCLUSION: Our study points out that tocilizumab may be an effective agent for pancreatitis treatment.

New Lyotropic Mixtures with Non-Chiral N-Acylamino Acid Surfactants Presenting the Biaxial Nematic Phase Investigated by Laser Conoscopy, Polarized Optical Microscopy and X-ray Diffraction.

Amino acid-based surfactants were used as the main surfactants to prepare new lyotropic mixtures presenting three nematic phases. One of them is biaxial (NB), and the two others are uniaxial, discotic (ND) and calamitic (NC). These surfactants were the non-chiral molecules, potassium N-dodecanoyl-DL-alaninate (DL-KDDA), potassium N-dodecanoyl-DL-serinate (DL-KDDS), disodium N-dodecanoyl-DL-aspartate (DL-NaDDAs) and potassium N-dodecanoyl-glycinate (KDDGly). Measurements of the optical birefringences and X-ray diffraction analysis were used to characterize the nematic phases and phase transitions. Mixtures with DL-KDDS exhibited the largest biaxial phase domain (~9 °C) with respect to the other mixtures in this study. The results obtained with the KDDGly mixture showed that the existence of hydrogen bonding between the head groups of the surfactant molecules seems to hinder the orientation of the micelles under the action of an external magnetic field.