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Endocytosis-mediated redistribution of antibiotics sheds light on the development of novel delivery platforms and alternative strategies to combat intracellular bacterial pathogens.

Biopharmaceutics Classification System (BCS) class II and IV drugs exhibit low solubility and suffer a limitation in oral administration. Exosomes have attracted intensive attention in the efficient delivery of such compounds. However, low gastrointestinal stability and high production cost of exosomes hinder their development as drug carriers. Here, milk exosomes are functionalized with phosphatidylserine and are capable of improving the solubility of BCS class II and IV drugs, resulting in facilitating the oral delivery of the drugs. A natural flavonoid, α-mangostin, is loaded into exosomes (AExo) to enhance the antibacterial efficiency, demonstrated by clearing 99% of bacteria in macrophages. Furthermore, AExo exhibits high mucus penetrability and shows a significant therapeutic efficacy in two animal infection models. Collectively, this work expands the application of exosomes from bovine milk with simple operation and low cost, shedding light on the potential of milk exosomes in improving the solubility of drugs to enhance the efficacy of oral administration.

Vancomycin (VCM) is an effective chemotherapeutic agent commonly used against gram-positive microorganisms but has serious nephrotoxic side effects that limit its effectiveness. New therapeutics and strategies are urgently needed to combat VCM associated nephrotoxicity. In this study, we determined the protective effect of chlorogenic acid (CA) in a rat model of VCM-induced nephrotoxicity. VCM administration led to markedly elevated blood urea nitrogen and serum creatinine levels that could be prevented with CA co-administration. VCM-mediated oxidative stress was also significantly attenuated by CA as reflected by decreased malondialdehyde and nitric oxide in VCM-treated kidneys. CA administration also prevented the VCM-mediated decrease in the renal antioxidative enzyme activities of glutathione reductase, glutathione peroxidase, and catalase and led to increased levels of reduced glutathione that had been depleted by VCM. Moreover, CA administration clearly inhibited VCM-induced expression of nuclear factor-kappa B, inducible nitric oxide synthase and the downstream pro-inflammatory mediators tumor necrosis factor-α and interleukins 1β and 6. Apoptotic markers were also markedly down-regulated with CA. Overall, CA treatment mitigated VCM-induced oxidative and nitrosative stresses and countered the apoptotic and inflammatory effects of VCM. Notably, CA did not affect the antibacterial activity of VCM in vitro.

The rapid dissemination of life-threatening multidrug-resistant bacterial pathogens calls for the development of new antibacterial agents and alternative strategies. The virulence factor secreted by bacteria plays a crucial role in the sophisticated processes during infections. Inspired by the unique capacity of many bacteria inducing clotting of plasma to initiate colonization, we propose a programmable antibiotic delivery system for precision therapy using methicillin-resistant S. aureus (MRSA) as a model. Coagulase utilized by MRSA to directly cleave fibrinogen into fibrin, is an ideal target not only for tracking bacterial status but for triggering the collapse of fibrinogen functionalized porous microspheres. Subsequently, staphylokinase, another virulence factor of MRSA, catalyzed hydrolysis of fibrin to further release the encapsulated antibiotics from microspheres. Our sequential triggered-release system exhibits high selectivity to distinguish live or dead MRSA from other pathogenic bacteria. Furthermore, such programmable microspheres clear 99% MRSA in 4 h, and show increased efficiency in a wound healing model in rats. Our study provides a programmable drug delivery system to precisely target bacterial pathogens using their intrinsic enzymatic cascades. This programmable platform with reduced selective stress of antibiotics on microbiota sheds light on the potential therapy for future clinical applications.

Background: In the treatment of lung diseases, drug showed low bioavailability and efficacy by conventional administration methods. Passive lung-targeting microspheres provide a method to deliver drugs from the vascular side, there is still a paucity of systematic studies on the biocompatibility of biodegradable-polymer microspheres. Results: We used poly (lactic acid-glycolic acid) (PLGA) microspheres with different particle sizes (3, 10, 25, and 40 mm) as a model. The optimal lung-targeting particle size of the PLGA microspheres is 10 mm and the corresponding range of maximum tolerated dose is 125-150 mg/kg. We hypothesized that the decrease of blood oxygen saturation under treatment of microspheres was caused by pulmonary embolism. We found varying degrees of blood circulation loss of lungs by micro computed tomography test, which indicated severe pulmonary embolism subsequent to intravenous injection of microspheres with a particle size >25 mm. Furthermore, we found lung injury and microspheres leaked from the blood vessels into the alveoli by H&E staining. This process was likely induced by increased secretion of matrix metalloproteinases (MMPs), which destroyed the alveolar-capillary barrier and trigger an inflammatory cascade. Finally, we found that optimal particle size and dose conditions did not affect normal physiological activities after the microspheres degraded. The upregulation of vascular endothelial growth factor (VEGF) and platelet endothelial cell adhesion molecule-1 (PECAM-1 or CD31) induced vessel recanalization and reestablishment, which promoted the progress of lung repair. Conclusions: Collectively, our results highlight the importance of accurately designing the optimal size and dose of microspheres for passive lung-targeting delivery.

Vancomycin is a glycopeptide antibiotic widely used to treat infections caused by methicillin-resistant Staphylococcus aureus. However, nephrotoxicity is a major adverse side effect, and the development of effective nephroprotective agents remains a priority in antimicrobial chemotherapy. In this study, we investigated the cell protective effects of the flavonol glycoside rutin against vancomycin-induced toxicity. Vancomycin added to porcine renal tubular LLC-PK1 cells caused an increase of production of intracellular reactive oxygen species and subsequent apoptotic cell death. Pretreatment of LLC-PK1 cells with rutin at 5, 10, and 20 μM for 2 hr prior to 2-mM vancomycin exposure for 24 hr significantly decreased intracellular reactive oxygen species and increased superoxide dismutase and catalase activities. Rutin pretreatment also protected cells from vancomycin-induced caspase activation, mitochondrial membrane depolarization, and subsequent apoptosis. This study demonstrates a protective effect of rutin and suggests that rutin coadministration is an alternative therapy for treatment of vancomycin-induced nephrotoxicity.

The aim of this study was to prepare cefquinome-loaded polylactic acid microspheres and to evaluate their in vitro and in vivo characteristics and pharmacodynamics for the therapy of pneumonia in a rat model. Microspheres were prepared using a 0.7 mm two-fluid nozzle spray drier in one step resulting in spherical and smooth microspheres of uniform size (9.8 ± 3.6 μm). The encapsulation efficiency and drug loading of cefquinome were 91.6 ± 2.6% and 18.7 ± 1.2%, respectively. In vitro release of cefquinome from the microspheres was sustained for 36 h. Cefquinome-loaded polylactic acid microspheres as a drug delivery system was successful for clearing experimental Klebsiella pneumonia lung infections. A decrease in inflammatory cells and an inhibition of inflammatory cytokines TNF-α, IL-1β and IL-8 after microspheres treatment was found. Changes in cytokine levels and types are secondary manifestations of drug bactericidal effects. Rats were considered to be microbiologically cured because the bacterial load was less than 100 CFU/g. These results also indicated that the spray-drying method of loading therapeutic drug into polylactic acid microspheres is a straightforward and safe method for lung-targeting therapy in animals.

Oxidation of o-nitroaniline with sodium hypochlorite afforded benzofurazan oxide in 96 % yield, and treatment of benzofurazan oxide with acetylacetone in the presence of triethylamine gave 2-acetyl-3-methyl-quinoxaline- -1,4-dioxide in 94 % yield. Finally, condensation of 2-acetyl-3-methyl-quinoxaline- 1,4-dioxide with benzaldehyde using 4-(dimethylamino)pyridinium acetate as a catalyst led to quinocetone in 95 % yield. Subsequently, reduction of the synthesized quinocetone with sodium dithionite resulted in two deoxy derivatives, 1-(3-methyl-4-oxido-2-quinoxalinyl)-3-phenyl-2-propen-1-one and 1-(3-methyl-2-quinoxalinyl)-3-phenyl-2-propen-1-one in 88.5 and 92 % yield, respectively. Furthermore, the synthesized quinocetone, and its deoxy derivatives were characterized by 1H-NMR, 13C-NMR and elemental analysis.

Quinocetone (QCT), an antimicrobial growth promoter, is widely used in food-producing animals. However, information about pharmacokinetics (PK) of QCT in ducks still remains unavailable up to now. In this study, QCT and its major metabolites (1-desoxyquinocetone, di-desoxyquinocetone and 3-methyl-quinoxaline-2-carboxylic) in ducks were studied using a simple and sensitive UHPLC-MS/MS assay. Twenty ducks were divided into two groups. (n = 10/group). One group received QCT by oral administration at dose of 40 mg/kg while another group received QCT intravenously at 10 mg/kg. Plasma samples were collected at various time points from 0 to 96 hr. QCT and its major metabolites in duck plasma samples were extracted by 1 ml acetonitrile and detected by UHPLC-MS/MS, with the gradient mobile phase that consisted of 0.1% formic acid in water (A) and acetonitrile (B). A noncompartment analysis was used to calculate the PK parameters. The results showed that following oral dosing, the peak plasma concentration (Cmax ) of QCT was 32.14 ng/ml and the area under the curve (AUCINF_obs) was 233.63 (h ng)/ ml. Following intravenous dosing, the Cmax , AUCINF_obs and Vss_obs were 96.70 ng/ml, 152.34 (h ng)/ ml and 807.00 L/kg, respectively. These data indicated that the QCT was less absorbed in vivo following oral administration, with low bioavailability (38.43%). QCT and its major metabolites such as 1-desoxyquinocetone and 3-methyl-quinoxaline-2-carboxylic were detected at individual time points in individual ducks, while the di-desoxyquinocetone was not detected in all time points in all ducks. This study enriches basic scientific data about pharmacokinetics of QCT in ducks after oral and intravenous administration and will be beneficial for clinical application in ducks.

The aims of this study were to prepare pidotimod (PDM) soluble powder and to investigate the immune enhancement properties of PDM in chickens vaccinated with Newcastle disease virus vaccine. In vivo experiment, 360 6-day-old chickens were averagely divided into 6 groups. The chickens, except blank control (BC) group, were vaccinated with Newcastle disease vaccine (NDV). At the same time of the vaccination, the chickens in three PDM groups were given water with PDM for 5days, respectively, with the PDM at low, medium and high concentrations (0.25g/L, 0.5g/L, 1g/L), in control drug group was treated with 0.2ml/PDM dose via drinking water, in vaccination control (VC) and BC group, with equal volume physiological saline, once a day for five successive days. On days 14, 21 and 28 after the vaccination, the growth performance, the lymphocyte proliferation, serum antibody titer, the CD4/CD8 cell ratios and interleukin-2 (IL-2) and interferon-gamma (IFN-γ) were measured. The results showed that PDM at suitable dose could significantly promote growth performance, lymphocyte proliferation, enhance serum antibody titer, CD4/CD8 cell ratios and improve serum IL-2 and IFN-γ concentrations. It indicated that PDM could significantly improve the immune efficacy of Newcastle disease vaccine using doses of 0.5g/L, these results are consistent with the drug acting as an immunopotentiator.

The polymers of poly (lactic-co-glycolic acid), gelatin, and ethyl cellulose were used to control the microscopic structure of microspheres prepared by spray drying. We investigated controlled release by varying material concentrations, sprayer inlet temperatures and air velocities, and optimized process parameters of yield, particle size, and distribution. Three types of polymeric microspheres with a mean particle size range of 13–25 µm were loaded with cefquinome and characterized. In vitro drug release studies determined that microspheres were the most suitable structures for controlling release due to their high entrapment capacities and release times greater than 24 h.

Treatment of multi-drug resistant (MDR) Escherichia coli intestinal infections are being hampered by the presence of the mcr-1 (colistin) and tet (tetracycline) resistance genes in these strains. We screened seven traditional Chinese medicines for their ability to synergize with tetracycline to provide an effective new drug for the treatment of animal intestinal diseases caused by MDR E. coli. Our primary screen identified quercetin as a compound that reduced the minimum inhibitory concentration (MIC) of tetracycline against the E. coli standard test strain American Type Culture Collection (ATCC) 25922 and clinical isolates fourfold from 4 and 256 μg/mL to 1 and 64 μg/mL, respectively. Low levels of quercetin in combination with tetracycline were bactericidal for clinical E. coli isolates and after 24 h, the differences between this combination and each drug singly were 108 CFU/mL. We used this combination therapy in a mouse infection model and found 100% survival after 48 h compared with

Staphylococcus aureus is a bacterial pathogen that causes food poisoning, various infections, and sepsis. Effective strategies and new drugs are needed to control S. aureus associated infections due to the emergence and rapid dissemination of antibiotic resistance. In the present study, the antibacterial activity, potential mode of action, and applications of flavonoids from licorice were investigated. Here, we showed that glabrol, licochalcone A, licochalcone C, and licochalcone E displayed high efficiency against methicillin-resistant Staphylococcus aureus (MRSA). Glabrol, licochalcone A, licochalcone C, and licochalcone E exhibited low cytotoxicity without hemolytic activity based on safety evaluation. Glabrol displayed rapid bactericidal activity with low levels of resistance development in vitro. Meanwhile, glabrol rapidly increased bacterial membrane permeability and dissipated the proton move force. Furthermore, we found that peptidoglycan, phosphatidylglycerol, and cardiolipin inhibited the antibacterial activity of glabrol. Molecular docking showed that glabrol binds to phosphatidylglycerol and cardiolipin through the formation of hydrogen bonds. Lastly, glabrol showed antibacterial activity against MRSA in both in vivo and in vitro models. Altogether, these results suggest that glabrol is a promising lead compound for the design of membrane-active antibacterial agents against MRSA and can be used as a disinfectant candidate as well.

This study aimed to compare in vivo activity between cefquinome (CEQ)-loaded poly lactic-co-glycolic acid (PLGA) microspheres (CEQ-PLGA-MS) and CEQ injection (CEQ-INJ) against Klebsiella pneumonia in a rat lung infection model. Forty-eight rats were divided into control group (sham operated without infection and drug treatment), Klebsiella pneumonia model group (KPD + Saline), CEQ-PLGA-MS and CEQ-INJ therapy groups (KPD + CEQ-PLGA-MS and KPD + INJ, respectively). In the KPD + Saline group, rats were infected with Klebsiella pneumonia ATCC 10031. In the KPD + CEQ-PLGA-MS and KPD + INJ groups, infected rats were intravenously injected with 12.5 mg/kg body weight CEQ-PLGA-MS and CEQ-INJ, respectively. Compared to CEQ-INJ treatment group, CEQ-PLGA-MS treatment further decreased the number of bacteria colonies (decreased to 1.94 lg CFU/g) in lung tissues and the levels of inflammatory cytokine including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-4 (p

Nephrotoxicity is the major limiting factor for the clinical use of vancomycin (VCM) for treatment of serious infections caused by multiresistant Gram-positive bacteria. This study investigated the renal protective activity of rutin in a rat model of VCM-induced kidney injury in male Wistar rats. VCM administered intraperitoneally at 200 mg/kg twice daily for 7 successive days resulted in significant elevation of blood urea nitrogen and creatinine, as well as urinary N-acetyl-β-D-glucosaminidase. Coadministration of VCM with oral rutin at 150 mg/kg significantly reduced these markers of kidney damage. Rutin also significantly attenuated VCM-induced oxidative stress, inflammatory cell infiltration, apoptosis, and decreased interleukin-1β and tumor necrosis factor alpha levels (all P

The objective of this study was to prepare a new compound fenbendazole tablet containing 29.7 % fenbendazole, 1.50 % praziquantel and 0.059 % ivermectin for oral administration. The tablets were successfully prepared using mannitol as filler agent, polyvinyl polypyrrolidone as disintegrant, 5 % povidone (PVAK30) as a binder agent and magnesium stearate as lubricant. The appearance, hardness, fragility, time limit of disintegration and fenbendazole dissolution at 45 min all met the technical standards of the Ministry of Agriculture for the People's Republic of China. We used high performance liquid chromatography and electrospray-mass spectrometry for drug detection. Oral administration of 100 mg/kg fenbendazole, 5 mg/kg praziquantel and 0.2 mg/kg ivermectin using a non-compartmental model defined peak plasma concentrations (Cmax) of 495, 826, 73 ng/mL, and 218 ng/mL for the metabolite oxfendazole, respectively. The area under the curve (AUClast) values for these drugs were 4653, 1045, 1971 and 5525 h×ng/mL, respectively. This study enriches the pharmacokinetic data of compound fenbendazole tablets using dogs as a model system. The new tablet formulation was assimilated quickly and systemically and this study will be beneficial for the clinical application of parasite treatments in dogs.

The aim of this study was to prepare CEQ-loaded gelatin microspheres and compare two preparation methods, evaluate targeting to the lungs. Gelatin microspheres containing CEQ were prepared by an emulsion cross-linking method (ECLM) and a spray-drying method (SDM) and were characterized in terms of morphology, size, drug-loading coefficient, encapsulation ratio and in vitro release. The microspheres prepared by ECLM gave a drug loading (DL) of 19.4 ± 2.4% and an entrapment efficiency (EE) of 80.8 ± 3.2%. The microspheres prepared by SDM resulted in a DL value of 20.8 ± 2.7% and an EE of 95.3 ± 3.8%. The average particle size of microspheres was 7-30 μm by both methods and both preparations sustained CEQ release for 36 h in the target tissue (lungs). The in vitro release profile of the microspheres matched the Korsmeyer-Peppas release pattern. In vivo studies identified the lung as the target tissue and the region of maximum CEQ release. Histopathological examination showed a partial lung inflammation that disappeared spontaneously as the microspheres were biodegraded. In general, the formulations were safe. The well-sustained CEQ release from the microspheres revealed its suitability as a drug delivery vehicle that minimized injury to healthy tissues while achieving the accumulation of therapeutic drug for lung targeting. The intravenous administration of CEQ gelatin microspheres prepared by SDM is of potential value in treating lung diseases in animals.

The aim of this study was to prepare cefquinome-loaded poly lactic-co-glycolic acid (PLGA) microspheres and to evaluate their in vitro and in vivo characteristics. Microspheres were prepared using a spry drier and were characterized in terms of morphology, size, drug-loading coefficient, encapsulation ratio and in vitro release. The prepared microspheres were spherical with smooth surfaces and uniform size (12.4 ± 1.2 μm). The encapsulation efficiency and drug loading of cefquinome was 91.6 ± 2.6 and 18.3 ± 1.3%, respectively. In vitro release of cefquinome from the microspheres was sustained for 36 h. In vivo studies identified the lung as the target tissue and the region of maximum cefquinome release. A partial lung inflammation was observed but disappeared spontaneously as the microspheres were removed through in vivo decay. The sustained cefquinome release from the microspheres revealed its applicability as a drug delivery system that minimized exposure to healthy tissues while increasing the accumulation of therapeutic drug at the target site. These results indicated that the spray-drying method of loading cefquinome into PLGA microspheres is a straightforward method for lung targeting in animals.

Additional file 1: Table S1. Correlations of ADG, FGR and Diarrhea.