Role of the Abcg2 transporter in plasma, milk, and tissue levels of the anthelmintic monepantel in mice.

Breast cancer resistance protein/ATP-binding cassette subfamily G2 (BCRP/ABCG2) is an ATP-binding cassette efflux (ABC) transporter expressed in the apical membrane of cells in tissues, such as the liver, intestine, kidney, testis, brain, and mammary gland. It is involved in xenobiotic pharmacokinetics, potentially affecting the efficacy and toxicity of many drugs. In this study, the role of ABCG2 in parasiticide monepantel (MNP) and its primary metabolite, monepantel sulfone (MNPSO2)'s systemic distribution and excretion in milk, was tested using female and male wild-type and Abcg2-/- mice. Liquid chromatography coupled with a tandem mass spectrometer (LC-MS/MS) was used for the analysis in a 10-min run time using positive-mode atmospheric pressure electrospray ionization (ESI+) and multiple reaction monitoring (MRM) scanning. For the primary metabolite tested, milk concentrations were 1.8-fold higher in wild-type mice than Abcg2-/- female lactating mice (P = 0.042) after intravenous administration of MNP. Finally, despite the lack of a difference between groups, we investigated potential differences in MNP and MNPSO2's plasma and tissue accumulation levels between wild-type and Abcg2-/- male mice. In this study, we demonstrated that MNPSO2 milk levels were affected by Abcg2, with potential pharmacological and toxicological consequences, contributing to the undesirable xenobiotic residues in milk.

Preparation, characterization, and evaluation of antibacterial and cytotoxic activity of chitosan-polyethylene glycol nanoparticles loaded with amoxicillin as a novel drug delivery system.

In this study, nanoparticles of amoxicillin (AMX) were prepared using chitosan (CHI) and polyethylene glycol (PEG). The physicochemical properties of the particles were investigated by FT-IR, DSC, SEM, and zeta potential analyses. The nanoparticles showed a spherical shape, and the average size of formulations was within the range of 696.20 ± 24.86 - 359.53 ± 7.41 nm. Zeta potential data demonstrated that the formulations had positive surface charges with a zeta potential range of 21.38 ± 2.28 - 7.73 ± 1.66 mV. FTIR analysis showed that the drug was successfully entrapped in the nanoparticles. DSC results suggested that the drug was present in amorphous form in the polymer matrix. In vitro release studies demonstrated that the release pattern consisted of two phases, with an initial burst release followed by a controlled and sustained release. The MTT assay results on mouse fibroblast cell line indicated that the prepared formulations did not affect the viability of the cells. In the in vitro antibacterial activity test, it was found that the drug-loaded nanoparticles have AMX-equivalent antibacterial activity against E. coli, and S. aureus. These findings revealed that the obtained nanoparticles might be a promising and safe nanocarrier system for efficient delivery of AMX.

Amoxicillin has been encapsulated in PEG-CHI nanoparticles.The structure of nanoparticles was investigated by SEM, FTIR, and DSC studies.The nanoparticles showed an initial fast release followed by a slow release.PEG-CHI nanoparticles displayed equivalent antibacterial activity to amoxicillin, and a non-cytotoxic profile in healthy cells.

Effects of feed intake and water hardness on fluralaner pharmacokinetics in layer chickens.

BACKGROUND: Fluralaner is a novel drug belonging to the isoxazoline class that acts on external parasites of domestic animals. It is used systemically via drinking water, especially against red poultry mite in layer chickens. Fluralaner is frequently used in layers infected with D. gallinae. However, no study to date has investigated the effects of feed intake and water hardness. OBJECTIVES: This study aimed to investigate the effects of variable water hardness and feed intake on the pharmacokinetic profile of fluralaner. METHODS: Layer chickens were divided into four groups (n = 8): fed + purified water (Group 1), feed restricted + purified water (Group 2), feed restricted + hard water (Group 3), and feed restricted + soft water (Group 4). After administering a single dose of the drug with drinking water, the blood samples were collected for 21 days. Fluralaner concentrations in plasma samples were determined by liquid chromatography/tandem mass spectrometry. The maximum plasma concentration (Cmax), time to reach maximum plasma concentration (tmax), area under the concentration-time curve values (AUC0-21d), half-life (t1/2), and other pharmacokinetic parameters were calculated. RESULTS: Although the highest maximum plasma concentration (Cmax) was determined in Group 1 (fed + purified water), no statistically significant difference was found in the Cmax, tmax, t1/2, MRT0-inf_obs, Vz/Fobs, and Cl/F_obs parameters between the experimental groups. CONCLUSIONS: It was concluded that the feed intake or water hardness did not change the pharmacokinetic profile of fluralaner in layer chickens. Therefore, fluralaner could be used before or after feeding with the varying water hardness in poultry industry.

Effects of Tyrosine Kinase Inhibitor-masitinib Mesylate on Canine Mammary Tumour Cell Lines.

INTRODUCTION: Masitinib mesylate, a selective tyrosine kinase inhibitor of the c-KIT receptor, is used for the treatment of mast cell tumours in dogs. Masitinib has previously been investigated in various cancers; however, its potential anticancer effect in canine mammary tumours (CMTs) is unknown. In the present paper, we investigated the antiproliferative effect of masitinib in CMT cells and its possible mechanisms of action. MATERIAL AND METHODS: The effect of masitinib on the proliferation of CMT-U27 and CMT-U309 cells was assessed by MTT assay and DNA fragmentation. Flow cytometric analysis was used to measure the effect of masitinib on apoptosis and the cell cycle. Additionally, vascular endothelial growth factor levels (VEGF) were measured, and the proliferation marker Ki-67 was visualised in immunocytochemical stainings in CMT cells. RESULTS: Treatment with masitinib inhibited the proliferation of CMT cells in a concentration-dependent manner. Maximal apoptotic activity and DNA fragmentation were observed at approximately IC50 of masitinib in both cell lines. In addition, cell cycle distribution was altered and VEGF levels and Ki-67 proliferation indices were decreased in masitinib-treated cells in comparison with control cells. CONCLUSION: In this study, masitinib suppressed cell proliferation concomitantly via induction of apoptosis and cell cycle arrest by decreasing VEGF levels and the Ki-67 proliferation index in CMT-U27 and CMT-U309 cells in vitro, suggesting its potential as a therapeutic tool in the clinical setting of mammary cancer treatment in dogs.

The screening of the safety profile of polymeric based amoxicillin nanoparticles in various test systems.

Nanotechnology-based drugs show superiority over conventional medicines because of increased bioavailability, lower accumulation in non-target tissues, and improved therapeutic index with increased accumulation at target sites. However, it is important to be aware of possible problems related to the toxicity of these products, which have therapeutically superior properties. Accordingly, the present study was designed to investigate the safety profile of amoxicillin nanoparticles (AmxNPs) that we developed to increase the oral bioavailability of amoxicillin (Amx) in poultry. In the first part of the study, the genotoxicity potential of AmxNPs was evaluated using the Ames test and the in vitro comet assay. The results of Ames test showed that none of the tested concentrations of Amx and AmxNPs cause a significant increase in the revertant number of Salmonella typhimurium strains TA98, and TA100, either with or without metabolic activation. Similarly, the comet assay revealed that AmxNPs did not induce DNA damage at any of the concentrations used, whereas high-dose (200 µg/mL) of Amx caused a significant increase in the percentage of DNA in the tail. In the second part of the study, the toxicity potential of AmxNPs on broilers was investigated by measuring biochemical parameters. In vivo results demonstrated that AmxNps did not cause a significant change in biochemical parameters, whereas Amx increased ALT, glucose, and cholesterol levels at certain sampling times. The obtained findings suggest that AmxNPs could be a safe promising potential drug in drug delivery systems.

In Vitro Evaluation of Antioxidant, Anti-Inflammatory, Antimicrobial and Wound Healing Potential of Thymus Sipyleus Boiss. Subsp. Rosulans (Borbas) Jalas.

Thymus sipyleus Boiss. subsp. rosulans (Borbas) Jalas (TS) is a commonly used plant in the treatment of various complaints, including skin wounds in Turkish folk medicine. Despite the widespread traditional use of TS, there is not any scientific report confirming the effectiveness of this plant on the healing process. This research aimed to investigate the effects of different extracts obtained from TS on biological events during wound healing, on a cellular basis. In this context, proliferative activities of the extracts, as well as the effects on wound closure and hydroxyproline synthesis, were determined. In addition to wound healing properties, the antioxidant, antibacterial and anti-inflammatory activities of the extracts were evaluated. Decoction (D) and infusion (I) extracts contained the highest amount of phenolic content and showed the most potent activity against DPPH radical. All extracts exhibited complete protection against the damage induced by hydrogen peroxide (H2O2) by increasing cell viability compared to only H2O2-treated groups, both in co-treatment and pre-treatment protocols. None of the extracts exhibited cytotoxic activity, and most of the extracts from the TS stimulated fibroblast proliferation and migration. All TS extracts exert anti-inflammatory activity by suppressing the overproduction of tumor necrosis factor-alpha (TNF-α) and nitric oxide (NO). The most pronounced activity on hydroxyproline synthesis was observed in D extract. In summary, it was observed that TS extracts can promote the healing process by enhancing fibroblast migration, proliferation and collagen synthesis as well as suppressing pro-inflammatory cytokines. The obtained data in this work support the traditional use of TS as a valuable plant-based compound for the treatment of wounds.

Development and characterization of polymeric-based nanoparticles for sustained release of amoxicillin - an antimicrobial drug.

In this study, amoxicillin (AMO)-loaded poly(vinyl alcohol)/sodium alginate (PVA/NaAlg) nanoparticles were prepared as a polymer-based controlled release system. The physicochemical properties of the obtained nanoparticles were investigated by XRD, DSC/TGA, particle size analyses and zeta potential measurements. The average particle sizes were in the range from 336.3 ± 25.66 to 558.3 ± 31.39 nm with negative zeta potential values from -41.86 ± 0.55 to -47.3 ± 2.76 mV. The influences of PVA/NaAlg ratio, span 80 concentration, exposure time to glutaraldehyde (GA) and the drug/polymer ratio on AMO release profiles were evaluated. In vitro drug release studies showed a controlled and pH dependent AMO release with an initial burst effect. XRD patterns and DSC thermograms of AMO-loaded nanoparticles revealed that the drug in the nanoparticles was in amorphous form, which was more stable than the crystalline form. The antibacterial activity of the optimal formulation was also investigated. The minimum inhibitory concentration (MIC) values of this formulation had the comparable antibacterial activity with that of pure AMO. These results indicate that the developed nanoparticles could be a promising candidate drug delivery system for AMO.

Response of cultured normal canine mammary epithelial cells to deracoxib-doxorubicin combination.

Currently, there is a growing interest in combining anticancer drugs with the aim to improve outcome in patients suffering from tumours and reduce the long-term toxicity associated with the current standard of treatment. In this study, we evaluated the possible role of deracoxib against the toxicity of doxorubicin on normal canine mammary epithelial cells. The effect of deracoxib and doxorubicin combination on cell viability was determined by MTT assay. Apoptosis was characterised by flow cytometry. Cell nitrite concentrations were measured with the Griess reaction. Deracoxib (50 and 100 µM) treatment decreased the cytotoxic action of doxorubicin at 0.9 µM in the cells, from 33.63% to 13.4% and 25.82%, respectively. Our results also showed that the reverse effect of deracoxib on doxorubicin-induced cytotoxic activity in the cells was associated with a marked (3.04- to 3.57-fold) decrease in apoptosis. In additional studies identifying the mechanism of the observed effect, deracoxib exhibited an activity to prevent doxorubicin-mediated overproduction of nitric oxide in the cells. Our in vitro study results indicate that deracoxib (50 and 100 µM) can be beneficial in protecting normal cells from the toxic effect of doxorubicin in conjunction with apoptosis by the modulation of nitric oxide production.

Synergistic growth inhibitory effect of deracoxib with doxorubicin against a canine mammary tumor cell line, CMT-U27.

Cyclooxygenase (COX) inhibitors have been shown to exert anti-angiogenic and anti-tumor activities on many types of malignant tumors. These anticancer properties make it worthwhile to examine the possible benefit of combining COX inhibitors with other anti-cancer agents. In the present study, we evaluated the potential of deracoxib (DER) in potentiating antitumor activity of doxorubicin (DOX) in canine mammary carcinoma cells (CMT-U27). DER (50-250 µM) enhanced the antiproliferative activity of DOX by reducing the IC50 (approximately 3- to 3.5 fold). Interaction analysis of the data showed that combinations of DOX at 0.9 µM with DER (100-250 µM) produced synergism in the CMT-U27 cell line, with a ratio index ranging from 1.98 to 2.33. In additional studies identifying the mechanism of observed synergistic effect, we found that DER strongly potentiated DOX-caused G0/G1 arrest in cell cycle progression. Also, DER (100-250 µM) augmented apoptosis induction with approximately 1.35- and 1.37- fold increases in apoptotic response caused by DOX in the cells. DER enhanced the antiproliferative effect of DOX in conjunction with induction of apoptosis by modulation of Bcl-2 expression and changes in the cell cycle of the CMT-U27 cell line. Although the exact molecular mechanism of the alterations in the cell cycle and apoptosis observed with DER and DOX combinations require further investigations, the results suggest that the synergistic effect of DOX and DER combinations in CMT therapy may be achieved at relatively lower doses of DOX with lesser side effects. Therefore, combining DER with DOX may prove beneficial in the clinical treatment of canine mammary cancer.

In vitro effects of doxorubicin and deracoxib on oxidative-stress-related parameters in canine mammary carcinoma cells.

The present study evaluated the effects of doxorubicin (DOX) and deracoxib (DER), as single agents and in combination treatments, on antioxidant parameters in the canine mammary carcinoma cell line CMT-U27. The cells were exposed to DOX and DER for 24, 48 and 72 h. The viability and malondialdehyde (MDA), nitric oxide (NO), catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSHPx) and total glutathione (GSH) activities of CMT-U27 cells were determined. The half inhibition concentration (IC50) of DOX was found to be ∼0.9 µM in the 72-h period. IC50 and 1/10 IC50 concentrations of DOX were combined with all concentrations of DER (50-1000 µM) in the combination experiments. The results showed increased oxidative status associated with significant decreases of CAT and GSH levels in CMT-U27 cells exposed to 10-µM and higher concentrations of DOX compared to control cells. In contrast, there were no significant changes in the groups tested with any of the concentrations of DER (50-1000 µM). In combination treatments, DER attenuated DOX-induced oxidative damage by modulating the enzymatic and non-enzymatic components in CMT-U27 cells. We suggest that the combination of DOX and DER can be beneficial in the treatment of cancer cells by increasing cellular responses to oxidative stress. In conclusion, the use of COX inhibitor in conjunction with a chemotherapeutic agent may provide a basis for new concepts of cancer treatment through systematic modulation of the antioxidant defence systems in mammary cancers of animals.

The effects of piroxicam and deracoxib on canine mammary tumour cell line.

Cyclooxygenase (COX) inhibitors, already widely used for the treatment of pain and inflammation, are considered as promising compounds for the prevention and treatment of neoplasia. The aim of our study was to determine the direct antiproliferative effects of nonsteroidal anti-inflammatory drugs (NSAIDs), piroxicam and deracoxib, at a variety of concentrations as both single and combined treatments on canine mammary carcinoma cell line CMT-U27 and to understand the mechanisms of cell death. MTT assay was performed to determine cell viability, and flow cytometric analyses were performed to evaluate apoptosis and cell cycle alterations. Significant decrease in cell viability was observed at high concentrations of piroxicam and deracoxib in both single and combined treatments after 72 h incubation. Combined treatment produced a significantly greater inhibition than that caused by either agent alone. Also apoptotic cell number was increased by both drugs at the cytotoxic concentrations. However, concomitant treatment of cells with piroxicam and deracoxib resulted in significant induction of apoptosis at lower concentrations and accumulation of cells in the G0/G1 phase. Significant cytotoxic effects exhibited by the combination of piroxicam and deracoxib against canine mammary carcinoma cells in vitro suggest an attractive approach for the treatment of canine mammary carcinoma.

In vivo assessment of antidiabetic and antioxidant activities of rosemary (Rosmarinus officinalis) in alloxan-diabetic rabbits.

Rosemary (Rosmarinus officinalis), used in traditional Turkish folk medicine for the treatment of hyperglycaemia, is widely accepted as one of the medicinal herb with the highest antioxidant activity. Accordingly, the present study was designed to investigate the possible actions of ethanolic extract of the leaves of Rosmarinus officinalis on glucose homeostasis and antioxidant defense in rabbits. In the first set of experiments, hypoglycaemic effects of oral administration of various doses (50, 100 and 200 mg/kg) of the extract were examined in normoglycaemic and glucose-hyperglycaemic rabbits. Optimal effect was observed in both of the animal groups with a dose of 200 mg/kg of the extract and this activity was independent from the effects of insulin. In another part of experiments, acute effect of various doses of the Rosmarinus officinalis extract on blood glucose and serum insulin levels was studied in alloxan-induced diabetic rabbits. Of the three doses of extract, the highest dose (200 mg/kg) significantly lowered blood glucose level and increased serum insulin concentration in alloxan-diabetic rabbits. The last set of experiments designed to investigate the subacute effect of the Rosmarinus officinalis extract on repeated administration in alloxan-diabetic rabbits. At the doses of 100 and 200 mg/kg, antihyperglycaemic effect of extract was accompanied by a significant increase in serum insulin levels in diabetic rabbits. Furthermore, during 1 week of treatment of diabetic rabbits with a dose of 200 mg/kg of the extract showed that the extract possessed a capability to inhibit the lipid peroxidation and activate the antioxidant enzymes. It was concluded that probably, due to its potent antioxidant properties, the Rosmarinus officinalis extract exerts remarkable antidiabetogenic effect.

Effect of linear alkylbenzene sulphonate (LAS) on non-specific defence mechanisms in rainbow trout (Oncorhynchus mykiss).

Linear alkylbenzene sulphonate (LAS) is among the most widely disseminated xenobiotics to enter waste streams and the aquatic environment. In the present investigation, we present a novel approach to evaluate in toxicity of LAS. The effects of sublethal levels (0.2 and 0.4 mg/l) of LAS on non-specific immune system, phagocytosis, respiratory burst and lyzosyme activity, and specific growth rate in the rainbow trout, Oncorhynchus mykiss, during a 54-day exposure were examined by a static bioassay test procedure. The phagocytic activity of leukocytes from fish exposed to 0.4 mg/l LAS statistically decreased compared with the control fish values. No significant reductions were observed in the extra-intracellular respiratory burst and lysozyme activities after exposure to LAS at any of the concentrations tested. The final body weight in fish groups exposed to the LAS were found to be significantly lower than in the control. The specific growth rate results also supported the result above. The results of this study showed sublethal doses (0.2-0.4 mg/l) of LAS caused to statistically insignificant suppression of non-specific immune system mechanisms excluding phagocytosis in fish at laboratory conditions. These doses of LAS may produce potential synergism on immune system when presented with other environmental pollutants.