Astilbin Attenuates Cadmium-Induced Adipose Tissue Damage by Inhibiting NF-κB Pathways and Regulating the Expression of HSPs in Chicken.

Cadmium (Cd) can damage tissues by inducing oxidative stress, lymphocyte infiltration, and inflammation in these sites. Meanwhile, astilbin (Ast) is an antioxidant agent. At present, only a few mechanisms of Cd-induced adipose tissue damage have been described. Herein, we assessed the potential protective effects and the molecular mechanism underlying the antioxidant properly of Ast after Cd intake in chicken adipose tissue. In this study, a total of 160 7-day-old roosters were randomly divided into four groups. Roosters were fed with a basic diet (C group), Ast 40 mg/kg (Ast group), CdCl2 150 mg/kg + Ast 40 mg/kg (Cd/Ast group), and CdCl2 150 mg/kg (Cd group) for 60 days. We found that Cd intake changed the morphology and structure of adipose tissues and decreased the expression of several antioxidants, including total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), catalase (CAT), and total antioxidant capacity (T-AOC), but increased those of oxidative stress markers including malondialdehyde (MDA), inducible nitric oxide synthase (iNOS), NO, and H2O2. Cd further activated the nuclear factor kappa B (NF-κB) signaling pathway and increased the expression of the inflammation-related mediators, interleukin 1beta (IL-1ß), interleukin 6 (IL-6), interleukin 8 (IL-8), interleukin 10 (IL-10), cyclooxygenase-2 (COX-2), iNOS, prostaglandin E synthase (PTGES), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). Cd-induced oxidative stress upregulated the expression of three heat shock proteins (HSPs), including HSP27, HSP70, and HSP90. Summarily, Cd causes oxidative stress-mediated tissue damage by activating the NF-κB pathway, promoting inflammation and upregulating the expression of HSPs. However, Ast supplementation modulates oxidative stress in adipose tissue by inhibiting inflammation mediated by the NF-κB pathway and regulating the expression of HSPs.

Protective Effects of Astilbin Against Cadmium-Induced Apoptosis in Chicken Kidneys via Endoplasmic Reticulum Stress Signaling Pathway.

Cadmium (Cd) can cause endoplasmic reticulum stress (ERS) and apoptosis in animals. The kidney is an organ seriously affected by Cd because it can accumulate metal ions. Astilbin (ASB) is a dihydroflavonol rhamnoside, which has an anti-renal injury effect. This study aimed to evaluate the protective effect of ASB on Cd-induced ERS and apoptosis in the chicken kidney. In this study, a total of 120 1-day-old chickens were randomly divided into 4 groups. Chickens were fed with a basic diet (Con group), ASB 40 mg/kg (ASB group), CdCl2 150 mg/kg + ASB 40 mg/kg (ASB/Cd group), and CdCl2 150 mg/kg (Cd group) for 90 days. The results showed that Cd exposure induced pathological and ultrastructural damages and apoptosis in chicken kidneys. Compared with the Con group, metallothionein (MT1/MT2) level, nitric oxide (NO) content, inducible nitric oxide synthase (iNOS) activity, ERS-related genes 78-kDa glucose-regulated protein (Grp78), protein kinase PKR-like endoplasmic reticulum kinase (Perk), activating transcription factor 4 (Atf4) and CAAT/enhancer-binding protein (C/EBP) homologous protein (Chop), and pro-apoptotic gene B-cell lymphoma 2 (Bcl-2)-associated X (Bax), caspase-12, caspase-9, caspase-3 expression levels, and apoptotic rate were significantly increased in the Cd group. The expression level of Bcl-2 was significantly decreased in the Cd group. ASB/Cd combined treatment significantly improves the damage of chicken kidneys by ameliorating Cd-induced kidney ERS and apoptosis. Cd can cause the disorder of the GRP78 signal axis, activate the PERK-ATF4-CHOP pathway, aggravate the structural damage and dysfunction of ER, and promote the apoptosis of chicken kidneys, while the above changes were significantly alleviated in the ASB/Cd group. The results showed that ASB antagonizes the negative effects of Cd and against Cd-induced apoptosis in chicken kidneys via ERS signaling pathway.

Hyperoside Attenuates Zearalenone-induced spleen injury by suppressing oxidative stress and inhibiting apoptosis in mice.

Zearalenone (ZEA) is a ubiquitous mycotoxin contaminant that causes immune toxicity, apoptosis, and oxidative stress in animals. Hyperoside (Hyp) is a flavonol glycoside compound with antioxidant and anti-apoptotic properties. However, the potential of Hyp to prevent ZEA-induced spleen injury remains unknown. To evaluate the chemoprotective effect of Hyp against ZEA-induced spleen injury, 60 male Kunming mice were randomly assigned into five groups. The first two groups were orally treated with ZEA (40 mg/kg) for 30 days, and combined with Hyp (0, 100 mg/kg) treatment. The other three groups are orally treated with normal saline, olive oil, or Hyp (100 mg/kg) for 30 days. Hyperoside had an inhibitory effect against ZEA-induced spleen lesions. In addition, Hyp significantly increased the activity of antioxidant enzymes [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT)], the total antioxidant capacity (T-AOC), and significantly reduced the malondialdehyde (MDA) content reducing ZEA-induced oxidative stress in the spleen. Moreover, the translation of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream target genes (CAT, NQO1, SOD1, GSS, GCLM, and GCLC) were ameliorated using co-therapy with Hyp before treatment with ZEA. Hyperoside also significantly inhibited the translation and expression of apoptotic genes (caspase3, casepase9, Bax, Bcl-2) and the production of apoptotic bodies induced by ZEA in the spleen. In conclusion, the findings revealed that Hyp inhibited ZEA-induced spleen injury through its antioxidant and anti-apoptotic effects. Thus, it provides a new treatment option for immune system diseases caused by ZEA.

Size-Dependent Biological Effects of Quercetin Nanocrystals.

Quercetin (QE) is an attractive natural compound for cancer prevention due to its beneficial anti-oxidative and anti-proliferative effects. However, QE is poorly soluble in water and slightly soluble in oil, which results in its low oral bioavailability and limits its application in the clinic. The aim of this study was to prepare QE nanocrystals (QE-NCs) with improved solubility and high drug loading, furthermore, the size-dependent anti-cancer effects of QE-NCs were studied. We prepared QE-NCs with three different particle sizes by wet milling, then, cell proliferation, migration and invasion were studied in A549 cells. The QE-NCs had antitumor effects in a dose- and size-dependent manner. Compared with the large particles, the small particles had a strong inhibitory impact on cell biological effects (p < 0.05 or p < 0.01). Moreover, Western blot assay indicated that QE-NCs may inhibit the migration and invasion of A549 cells by inhibiting the STAT3 signaling pathway, and the particle size may have an effect on this process. In this study, it was proven that NCs could dramatically enhance the anticancer efficacy of QE at the cellular level. In addition, particle size had a considerable influence on the dissolution behavior and antitumor effects of NCs.

Preparation and Evaluation of Midazolam Rectal Gel in vitro and in vivo.

Midazolam has been successfully used for sedation, which the tablets, injections and oral solutions were available in market. However, the oral bioavailability of midazolam is less due to the first effect, while injection formulation has a low patient compliance. The purpose of this study was to obtain midazolam rectal gel to meet the production and market requirements. We evaluated the in vitro release behavior of midazolam powder, a physical mixture of midazolam and HPMC, and midazolam gel. The results showed that the midazolam gel has superior release degree compared to the other. Correspondingly, midazolam gel produced relevant plasma concentrations result in the rats. At the same dose (1 mg/kg), the Cmax and AUC (0-t) of midazolam after administration of the midazolam rectal gel were 8-10 times higher than that of the oral solution. The midazolam rectal gel was evaluated for physico-chemical tests including pH, viscosity, drug content at 40°C /75%RH (relative humidity) for 180 days (6 months). It provides information about the shelf lives of medicine, as well as the conditions for their storage. Rectum irritation test demonstrated that frequent application of midazolam rectal gel did not induce rectal mucosal damage. In conclusion, midazolam rectal gel was a potential new dosage form, which could alleviate the feeling of alien, discomfort and refusal during application of the patients. It was likely to be a more convenient and effective sedative medicine form specifically for infants and children.

Solidification of Nanostructured Lipid Carriers Loaded Testosterone Undecanoate: In Vivo and In Vitro Study.

To enhance the bioavailability of testosterone undecanoate (TU) and overcome the current problem of soft capsules (Andriol Testocaps®), Nano-structured lipid carriers (NLC) for TU was developed. First, suspension of TU-loaded NLC (TU-NLC) was prepared by high pressure homogenization; then adsorbent or a protective agent ß-cyclodextrin was used to solidify the suspension through a vacuum system; finally, the solid powder of TU-loaded NLC (solid TU-NLC) was filled into hard capsules. The characteristics of solid TU-NLC, were investigated in vitro and vivo. The particle size of TU-NLC was about 273.3 nm, the potential was 0.156±0.04. Transmission electron microscope (TEM) revealed that the NLC was spherical and uniform. Differential scanning calorimetry (DSC) suggested the drug had been encapsulated into NLC lipid matrix. The drug release proved that solid TU-NLC showed a higher dissolution in vitro. The CaCO-2 cell permeability showed that solid TU-NLC could enhance trans-membrane absorption of the TU. Moreover, the AUC of solid TU-NLC formulations (4304±550.50 µg/L*min) was higher than commercial product Andriol Testocaps® (3075±372.50 µg/L*min). In conclusion, solid TU-NLC could enhance the rate of dissolution, and had a relatively higher bioavailability than Andriol Testocaps® in vivo GRAPHICAL ABSTRACT: .

Preparation and Evaluation of Progesterone Nanocrystals to Decrease Muscle Irritation and Improve Bioavailability.

Progesterone (PG) is a crucial immunomodulatory agent during early pregnancy, and nowadays PG oil-based injection (PG/OI) has a huge market all over the world. However, PG/OI may accumulate the local muscle and further cause irritations after long-term administration. In this study, PG nanocrystals (PG/NCs) injection was developed to decrease muscle toxicity. PG/NCs injection containing 10% (w/v) PG was first prepared using a wet grinding method. Then, particle size, zeta potential, morphology powder, X-ray diffraction (PXRD), differential scanning calorimetry (DSC), and Fourier transform infrared (FTIR) studies were carried out to evaluate the characteristics of dosage form. The rabbit muscle irritation, hemolysis, and rat pharmacokinetics tests were used to estimate the in vivo characteristics of PG/NCs. The results showed that the mean particle size and the zeta potentials of NCs were 299.5 ± 9.0 nm and - 36.8 ± 1.5 mV, respectively. The crystalline state of PG/NCs was not altered during particle size reduction according to PXRD, DSC, and FTIR results. Muscle irritation presented that PG/NCs had lower irritation than that of PG/OI. Hemolysis test suggested that PG/NCs injection was functioned without hemolysis and red cell agglutination. The pharmacokinetics study showed that the AUC0-t and Cmax of PG/NCs was 3.2-fold (p < 0.05) and 3.1-fold higher than PG/OI, which demonstrated that PG/NCs injection had greater bioavailability than PG/OI. Therefore, it was obvious that PG/NCs injection exhibited a lower muscle irritation, hemolysis rate, and higher bioavailability, which was a better dosage form than OI.

Preparation and evaluation of carfentanil nasal spray employing cyclodextrin inclusion technology.

Carfentanil (CFTN), a derivative of fentanyl, is highly effective as an analgesic, but its relatively poor solubility in water has limited its nasal application. The objective of this study was to develop the new CFTN-CD inclusion technology to increase the solubility of CFTN. The inclusion compound CFTN-DM-ß-CD was prepared by the ultrasonic method and characterized using X-ray powder diffraction and morphological shapes analysis (the scanning electron microscopy). The in vitro dissolution profiles of CFTN-DM-ß-CD were assessed in hydrochloric acid and phosphate buffer. Nasal ciliotoxicity studies were carried out using isolated toad palate. Rats were treated with CFTN-DM-ß-CD (250 µg/kg) by intravenous, intramuscular injection, oral, or nasal drops. The results showed that CFTN was successfully enveloped by DM-ß-CD. The in vitro cumulative dissolution of CFTN-DM-ß-CD was obviously enhanced compared to free CFTN in two buffers. Nasal ciliotoxicity studies have shown that the CFTN-DM-ß-CD does not exhibit higher nasal ciliotoxicity than that of free CFTN. Pharmacokinetic studies demonstrated that CFTN-DM-ß-CD by nasal administration was absorbed more rapidly and has higher Cmax and bioavailability than that of either intramuscular injection or oral administration. In conclusion, the CFTN-DM-ß-CD nasal spray was shown to be a relatively safe dosage form for the rapid and effective intranasal delivery of CFTN.

Metoclopramide nasal spray in vitro evaluation and in vivo pharmacokinetic studies in dogs.

Metoclopramide (MCP) can effectively alleviate motion sickness-caused nausea and vomiting. Nasal administration offers the greatest patient compliance. It is suitable for self-administration and offers rapid and complete absorption, no first-pass effects and high bioavailability. In the present study, a MCP nasal spray was prepared and evaluated in vitro and in vivo. Nasal cilia toxicity of Bufo toads was used to screen the preservative types and concentrations. Rabbit nasal mucosa was used to evaluate the mucosa permeability of different MCP nasal sprays with different penetration enhancers and preservative. A three-period crossover trial was then carried out in beagle dogs with three different MCP dosage forms: nasal sprays, oral tablets and intramuscular (IM) solution. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) was performed to measure dog plasma MCP, and pharmacokinetic parameters were calculated. The results of ciliatoxicity and permeation study showed that 0.03% methyl paraben lacking penetration enhancers was optimal. Compared to control IM, the bioavailability of oral tablets of MCP was 24.9%, while that of nasal spray was 62.3%. Meanwhile time-to-maximal plasma concentration (Tmax) of nasal spray was significantly shorter than that of oral tablets. In conclusion, MCP nasal spray prepared here is safe with minimal ciliatoxicity, rapid onset and high relative bioavailability.

Preparation, physical characterization and pharmacokinetic study of paclitaxel nanocrystals.

Paclitaxel (PTX) is a natural broad-spectrum anticancer drug with poor aqueous solubility. PTX nanocrystals were formulated to improve the water solubility, and PTX nanosuspensions were prepared using anti-solvent precipitation, and then organic solvent and surfactants were removed by filtering through a vacuum system. The physical characterization of PTX nanocrystals were measured by transmission electron microscope, X-ray diffraction and differential scanning calorimetry. In addition, saturation solubility, in vitro release, stability and pharmacokinetic characteristics were examined. The average particle size of PTX nanocrystals was ∼200 nm, and they had a stable potential and a uniform distribution. Paclitaxel nanocrystals can effectively improve drug solubility and in vitro release. PTX pharmacokinetic and tissue distribution studies were compared after intravenous administration of nanocrystals versus a commercial injection formulation. PTX nanocrystals were rapidly distributed with a longer elimination phase. Moreover, tissue distribution indicated that PTX nanocrystals are mainly absorbed by the liver and spleen and may offer reduced renal and cardiovascular toxicity which may reduce side effects.

Progress in the study of drug nanocrystals.

The poor water solubility of many candidate drugs remains a major obstacle to their development and clinical use, especially for oral drug delivery. Nanocrystal technology can improve the solubility and dissolution rates of many poorly water-soluble drugs very effectively, significantly improving their oral bioavailability and decreasing the food effect. For this reason, this technology is becoming a key area of drug delivery research. This review presents much of the recent progress in nanocrystal drug pharmaceuticals, including the characteristics, composition, preparation technology, and clinical applications of these drugs. Finally, the effect of nanocrystal technology on insoluble drugs is quantified and described.

Formulation and evaluation of thienorphine hydrochloride sublingual delivery system.

Thienorphine hydrochloride (ThH) is a highly insoluble and readily metabolized partial-opioid agonist. It is used for the treatment of pain and heroin addiction. This study aimed to formulate and evaluate sublingual delivery systems containing ThH. Dimethyl-ß-cyclodextrin (DM-ß-CD) can enhance the solubility and permeability of hydrophobic drugs. In this paper, ThH cyclodextrin inclusion complexes were prepared and administrated sublingually with the objective of improving the drug's aqueous solubility, in vitro permeation rate, and in vivo absorption rate. The formulation was prepared with DM-ß-CD using the freeze-dried method and characterized using phase solubility, differential scanning calorimetry (DSC), X-ray and NMR analyses. The results of each test indicated the formation of dynamic inclusion complexes between ThH and DM-ß-CD. The inclusion complexes also showed significant increases in in vitro aqueous solubility and mucosal permeability. According to the pharmacokinetic study of the complex in rats, the AUC and C(max) values of the sublingual delivery group were 40 and 46 times higher than those of the gastrointestinal group, whereas t(max) was shorter, which proved that in vivo absorption and metabolism had been improved. It can therefore be concluded that the inclusion technology and sublingual delivery system were suitable for ThH development.

Preparation and characterization of insulin-loaded bioadhesive PLGA nanoparticles for oral administration.

Poly(D,L-lactide-co-glycolide) nanoparticles (PLGA-NP) have been extensively used as a drug delivery system for proteins and peptides. However, their negative surface charge decreases bioavailability under oral administration. Recently, cationically modified PLGA-NP has been introduced as novel carriers for oral delivery. The characteristics of the nanoparticles, such as particle size, surface charge, and bioadhesion are considered the most significant determinants of the effect of these nanoparticles both in vitro and in vivo. Our aim was to introduce and evaluate the physiochemical characteristics, bioadhesion, and biological activity of positively charged chitosan-coated PLGA-NP (CS-PLGA-NP), using insulin as a model drug. Results were compared to those of common negatively charged PLGA-NP and the in vitro cytotoxicity of the two types of nanoparticles was examined. These results indicate that both CS-PLGA-NP and PLGA-NP had a narrow size distribution, averaging less than 150 nm. CS-PLGA-NP was positively charged (+43.1 ± 0.3 mV), exhibiting the cationic nature of chitosan, whereas PLGA-NP showed a negative surface charge (-1.72 ± 0.2 mV). CS-PLGA-NP exhibited stronger bioadhesive potency than PLGA-NP and much greater relative pharmacological availability with regard to orally delivered insulin. In addition, an evaluation of cytotoxicity by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed no increase in toxicity in either kind of nanoparticle during the formulation process. The study proves that CS-PLGA-NP can be used as a vector in oral drug delivery systems for proteins and peptides due to its positive surface charge and bioadhesive properties.