Anti-Photoaging Effects of Nanocomposites of Amphiphilic Chitosan/18ß-Glycyrrhetinic Acid.

Improving the transdermal absorption of weakly soluble drugs for topical use can help to prevent and treat skin photoaging. Nanocrystals of 18ß-glycyrrhetinic acid (i.e., NGAs) prepared by high-pressure homogenization and amphiphilic chitosan (ACS) were used to form ANGA composites by electrostatic adsorption, and the optimal ratio of NGA to ACS was 10:1. Dynamic light scattering analysis and zeta potential analysis were used to evaluate the nanocomposites' suspension, and the results showed that mean particle size was 318.8 ± 5.4 nm and the zeta potential was 30.88 ± 1.4 mV after autoclaving (121 °C, 30 min). The results of CCK-8 showed that the half-maximal inhibitory concentration (IC50) of ANGAs (71.9 µg/mL) was higher than that of NGAs (51.6 µg/mL), indicating that the cytotoxicity of ANGAs was weaker than that of NGAs at 24 h. After the composite had been prepared as a hydrogel, the vertical diffusion (Franz) cells were used to investigate skin permeability in vitro, and it was shown that the cumulative permeability of the ANGA hydrogel increased from 56.5 ± 1.4% to 75.3 ± 1.8%. The efficacy of the ANGA hydrogel against skin photoaging was studied by constructing a photoaging animal model under ultraviolet (UV) irradiation and staining. The ANGA hydrogel improved the photoaging characteristics of UV-induced mouse skin significantly, improved structural changes (e.g., breakage and clumping of collagen and elastic fibers in the dermis) significantly, and improved skin elasticity, while it inhibited the abnormal expression of matrix metalloproteinase (MMP)-1 and MMP-3 significantly, thereby reducing the damage caused by UV irradiation to the collagen-fiber structure. These results indicated that the NGAs could enhance the local penetration of GA into the skin and significantly improve the photoaging of mouse skin. The ANGA hydrogel could be used to counteract skin photoaging.

Grafting of 18ß-Glycyrrhetinic Acid and Sialic Acid onto Chitosan to Produce a New Amphipathic Chitosan Derivative: Synthesis, Characterization, and Cytotoxicity.

Chitosan is the only cationic polysaccharide found in nature. It has broad application prospects in biomaterials, but its application is limited due to its poor solubility in water. A novel chitosan derivative was synthesized by amidation of chitosan with 18ß-glycyrrhetinic acid and sialic acid. The chitosan derivatives were characterized by Fourier transform infrared spectroscopy, thermogravimetric analysis, and measurement of the zeta potential. We also investigated the solubility, cytotoxicity, and blood compatibility of chitosan derivatives. 18ß-glycyrrhetinic acid and sialic acid could be grafted onto chitosan molecular chains. The thermal stability of the synthesized chitosan derivatives was decreased and the surface was positively charged in water and phosphate-buffered saline. After chitosan had been modified by 18 ß-glycyrrhetinic acid and sialic acid, the solubility of chitosan was improved greatly in water and phosphate-buffered saline, and percent hemolysis was <5%. Novel amphiphilic chitosan derivatives could be suitable polymers for biomedical purposes.

Effects of Chitosan-Gentamicin Conjugate Supplement on Non-Specific Immunity, Aquaculture Water, Intestinal Histology and Microbiota of Pacific White Shrimp (Litopenaeus vannamei).

When the aquaculture water environment deteriorates or the temperature rises, shrimp are susceptible to viral or bacterial infections, causing a large number of deaths. This study comprehensively evaluated the effects of the oral administration of a chitosan-gentamicin conjugate (CS-GT) after Litopenaeus vannamei were infected with Vibrio parahaemolyticus, through nonspecific immunity parameter detection, intestinal morphology observation, and the assessment of microbial flora diversification by 16S rRNA gene sequencing. The results showed that the oral administration of CS-GT significantly increased total hemocyte counts and reduced hemocyte apoptosis in shrimp (p < 0.05). The parameters (including superoxide dismutase, glutathione peroxidase, glutathione, lysozyme, acid phosphatase, alkaline phosphatase, and phenoloxidase) were significantly increased (p < 0.05). The integrity of the intestinal epithelial cells and basement membrane were enhanced, which correspondingly alleviated intestinal injury. In terms of the microbiome, the abundances of Vibrio (Gram-negative bacteria and food-borne pathogens) in the water and gut were significantly reduced. The canonical correspondence analysis (CCA) showed that the abundances of Vibrio both in the water and gut were negatively correlated with CS-GT dosage. In conclusion, the oral administration of CS-GT can improve the immunity of shrimp against pathogenic bacteria and significantly reduce the relative abundances of Vibrio in aquaculture water and the gut of Litopenaeus vannamei.

Chitosan-Gentamicin Conjugate Hydrogel Promoting Skin Scald Repair.

Our earlier research indicated that chitosan-gentamicin conjugate (CS-GT) possesses superior antimicrobial activity and good water solubility. To develop CS-GT-based scald dressings, the antibacterial properties of CS-GT were further studied, and the biosafety of CS-GT and the healing mechanism of CS-GT hydrogel was systematically explored in this article. It was found that cell viability shows a declined inclination with the prolonged culture time and the increased concentration of CS-GT. After three day's culture, the cell viability could still remain at 79.72% when CS-GT concentration was as high as 1000 µg/mL. On the other hand, the hemolysis rate of CS-GT was lower than 5% when its concentration is 800 µg/mL. Therefore CS-GT has good cytocompatibility and hemocompatibility. A wound-healing experiment has shown that the skin healing rate of CS-GT hydrogel was the highest at 99.61%, followed by the positive control (wet burn ointment) 94.98%, GT hydrogel 87.50%, and matrix 77.39%. The blank control group, however, possessed the lowest healing rate of 75.45%. Further analysis indicated that CS-GT hydrogel could promote the synthesis of total protein (TP) in skin granulation tissue, resulting in the enhanced hydroxyproline (HYP) content, which facilitated collagen fibrogenesis, reduced cytokine expression in an inflammatory response, and, ultimately, accelerated wound healing. To sum up, CS-GT hydrogel is a promising scald dressing.

Anti-Aging Effect of Chitosan Oligosaccharide on d-Galactose-Induced Subacute Aging in Mice.

Chitosan oligosaccharide (COS), a natural polysaccharide with good antioxidant and anti-inflammatory properties, is the depolymerized product of chitosan possessing various biological activities. The present study was designed to investigate the possible anti-aging effect of COS on the aging model mouse induced by d-galactose (d-gal) and explore the underlying mechanism. In the experiment, 48 male Kunming mice (KM mice) were randomly divided into the normal group, model group, positive group, and low-medium-high dose polysaccharide groups (300, 600, 1200 mg/kg/day). The results showed that COS, by intragastric gavage after subcutaneous injection of d-gal (250 mg/kg/day) into the neck of mice consecutively for eight weeks, gradually recovered the body weight, the activity of daily living, and organ indices of mice, as well as effectively ameliorated the histological deterioration of the liver and kidney in mice triggered by d-gal. To be specific, COS obviously improved the activities of antioxidant enzymes in liver and kidney of KM mice, including catalase (CAT), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD), as well as decreased malondialdehyde (MDA) levels when compared with those in model group mice. Furthermore, COS not only elevated the diminished levels of serum immunoglobulin G (IgG) and IgM induced by d-gal, but also significantly inhibited the d-gal-caused upregulation of serum alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), uric acid (UA) and creatinine (CREA) levels as compared with those of mice in the model group. These results demonstrate that COS has an obvious anti-aging activity in d-gal-induced subacute aging mice, the mechanism of which, to some extent, is associated with enhancing the antioxidant defenses, reducing oxidative stress, and improving the immune function of aging model mice.

Investigation of the Effects of Molecular Parameters on the Hemostatic Properties of Chitosan.

Hemorrhea is one of the major problems in war, trauma care, and surgical operation that threaten the life of the injured and patients. As a novel polymeric hemostatic agent, biodegradable chitosan can stop bleeding through a variety of approaches. In this paper, chitosan with various molecular parameters was prepared from chitin as raw material through deacetylation, oxidative degradation, hydrophilic modification, and salt formation reactions. The influence of different polymer parameters on the hemostatic effects of chitosan was investigated by in vitro coagulation time and dynamic coagulation assay. The results showed that when the molecular weights were high (105⁻106) and approximate, the coagulation effect of chitosan improved with a decrease of the deacetylation degree and achieved a prominent level in a moderate degree of deacetylation (68.36%). With the same degree of deacetylation, the higher the molecular weight of chitosan, the better the procoagulant effect. The substituent derivatives and acid salts of chitosan showed significant procoagulant effects, especially the acid salts of chitosan. In addition, the hemostasis mechanism of chitosan with various parameters was preliminarily explored by analyzing the plasma recalcification time (PRT). The efforts in this paper laid a basis for further study of the structure⁻activity relationship and the mechanism of chitosan hemostasis.

Apigenin mediated protection of OGD-evoked neuron-like injury in differentiated PC12 cells.

Ischemic stroke has been confirmed to cause neuronal injury due to its insufficient supply of glucose and oxygen to brain tissue. Previous research has shown that oxidative stress, a result of excessive accumulation of reactive oxygen species (ROS), relates to pathophysiology of ischemic stroke, and causes oxidative damage to biomolecules, eventually leading to programmed cell death. Meanwhile, apigenin has been shown to exhibit antioxidant, anti-inflammatory, anti-cancer properties and neuroprotective action. Hence, this study was to investigate the potential mechanisms underlying the neural protection of apigenin on oxygen and glucose deprivation/reperfusion (OGD/R) induced neuronal injury in differentiated PC12 cells. Cells were pretreated with apigenin for 6 h, and then subjected to OGD for 12 h followed by reperfusion for 24 h. The results showed that OGD/R significantly decreased cell viability, mitochondrial membrane potential, mRNA levels of antioxidant and detoxifying enzymes and Nrf2 protein expression, while elevated the release of LDH, cell apoptosis, intracellular ROS level, P53 protein expression and upregulated its downstream genes in PC12 cells. However, apigenin effectively inhibited these undesirable changes induced by OGD/R. Our findings demonstrate that this compound attenuates OGD/R induced neuronal injury mainly by virtue of its anti-apoptosis and antioxidative properties via affecting the expression of Nrf2 and P53, and their downstream target gene transcription.

Protective effects of hydroxysafflor yellow A on ß-amyloid-induced neurotoxicity in PC12 cells.

The accumulation of extracellular amyloid-ß peptide (Aß) has been considered as one of the important causes of Alzheimer's disease (AD), the most prevalent form of dementia. Hydroxysafflor yellow A (HSYA), a major active chemical component isolated from Carthamus tinctorius L., has been shown to possess neuroprotective actions in various ischemic models in vivo. The present study aimed to investigate the potential protective effect of HSYA against Aß-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. The PC12 cells were pretreated with different concentrations (20, 40 and 80 µM) of HSYA for 2 h and then further treated with Aß (20 µM) for 24 h. The results showed that Aß could significantly decrease cell viability, glutathione level, mitochondrial membrane potential and the ratio of Bcl-2/Bax protein expression, while elevate the release of lactate dehydrogenase, the formation of DNA fragmentation, the levels of malondialdehyde and intracellular reactive oxygen species in PC12 cells. However, pretreatment with HSYA could effectively reverse these changes induced by Aß in PC12 cells. Our experimental results demonstrate that HSYA may be a potential neuroprotective agent warranting further development for treatment of AD.

Kinetics and mechanism study of competitive inhibition of jack-bean urease by baicalin.

Baicalin (BA) is the principal component of Radix Scutellariae responsible for its pharmacological activity. In this study, kinetics and mechanism of inhibition by BA against jack-bean urease were investigated for its therapeutic potential. It was revealed that the IC50 of BA against jack-bean urease was 2.74 ± 0.51 mM, which was proved to be a competitive and concentration-dependent inhibition with slow-binding progress curves. The rapid formation of initial BA-urease complex with an inhibition constant of K(i) = 3.89 × 10⁻³ mM was followed by a slow isomerization into the final complex with an overall inhibition constant of K(i)* = 1.47 × 10⁻4 mM. High effectiveness of thiol protectors against BA inhibition indicated that the strategic role of the active-site sulfhydryl group of the urease was involved in the blocking process. Moreover, the inhibition of BA was proved to be reversible due to the fact that urease could be reactivated by dithiothreitol but not reactant dilution. Molecular docking assay suggested that BA made contacts with the important activating sulfhydryl group Cys-592 residues and restricted the mobility of the active-site flap. Taken together, it could be deduced that BA was a competitive inhibitor targeting thiol groups of urease in a slow-binding manner both reversibly and concentration-dependently, serving as a promising urease inhibitor for treatments on urease-related diseases.

Preparation of Cod Skin Collagen Peptides/Chitosan-Based Temperature-Sensitive Gel and Its Anti-Photoaging Effect in Skin.

Background: Photoaging decreases quality of life and increases the risk of skin cancer, underscoring the urgent need to explore natural, high-efficacy, anti-skin photoaging (SP) active substances. Methods: In this study, a gel (CS/CSCPs/ß-GP gel) was prepared using chitosan (CS) and sodium ß-glycerophosphate (ß-GP) through crosslinking with small molecular CSCPs as the carried drug. We evaluated its structural characteristics and properties. The effect of CS/CSCPs/ß-GP gel on the degree of ultraviolet (UV)-induced skin aging of mice was investigated through comparative analysis of skin damage, the integrity of collagen tissues and elastic fibers, levels of reactive oxygen species (ROS) and key inflammatory factors (tumor necrosis factor [TNF]-α and interleukin [IL]-1ß, IL-6, and IL-10), and tissue expression of matrix metalloproteinase-3 (MMP-3) after repeated UV irradiation in a nude mice SP model. Results: The results showed that CS/CSCPs/ß-GP gel was successfully prepared and had the desired characteristics. Compared with CSCPs alone, the CS/CSCPs/ß-GP gel more evidently improved typical photoaging characteristics on mouse dorsal skin. It also increased the moisture content, causing the skin to become glossy and elastic. Pathological skin analysis revealed that this peptide-carrying gel can effectively inhibit epidermal thickening, reduce tissue inflammatory infiltration, suppress collagen fiber degradation, increase the collagen content, alleviate structural elastic fiber damage, and significantly inhibit abnormal MMP-3 expression. In addition, biochemical analysis showed that the CS/CSCPs/ß-GP gel can effectively inhibit the elevated expressions of ROS and key proinflammatory factors (TNF-α, IL-1ß, IL-6) in photoaging skin tissues and promote expression of the anti-inflammatory factor IL-10. Conclusion: SP can cause many clinical skin diseases, such as solar freckle-like nevus, solar keratosis, cutaneous melanoma, and squamous cell carcinoma. CSCPs are a high-efficacy anti-SP natural active substance and CS/CSCPs/ß-GP gel can synergistically enhance the CSCPs' anti-SP effect. The mechanism is likely related to the inhibited activation of ROS/nuclear transcription factor-κB signaling and the expression of downstream inflammatory factors.

Gastric acid-response chitosan/alginate/tilapia collagen peptide composite hydrogel: Protection effects on alcohol-induced gastric mucosal injury.

Long-term excessive alcohol intake can easily lead to gastritis, gastric ulcer, and gastric bleeding. In this paper, the gastric acid-responsive hydrogel of CS-NAC/alginate/tilapia collagen peptide (CS-NAC/ALG/TCP) was developed. Its structure and properties were determined. The alcohol-induced gastric mucosal injury models in mice were established to evaluate the protective effects of CS-NAC/ALG/TCP. The results showed that CS-NAC/ALG/TCP was successfully fabricated, and it showed a sustained release of TCP, strong mucoadhesion, and excellent biodegradability in vitro. In the animal experiments, CS-NAC/ALG/TCP improved the oxidative stress status of the gastric mucosa by increasing the levels of SOD, GSH, and CAT in tissues. It also down-regulated the expression of MPO, TNF-α, IL-1ß, and IL-6, and increased the production of gastric protective factors such as PGE2 and NO in mouse stomach, thereby reducing the alcohol-induced inflammation and protecting the gastric mucosal injury. Besides, CS-NAC/ALG/TCP can also increase the activities of alcohol metabolism enzymes to improve alcohol metabolism, thereby reducing alcoholic damage. In conclusion, CS-NAC/ALG/TCP is a promising candidate for the treatment of alcohol-induced gastric injury.

Optimized preparation of gastric acid-response sulfhydryl functionalized chitosan/alginate/tilapia peptide hydrogel and its protective effects on alcohol-induced liver and brain injury.

Long-term alcohol intake or drinking large quantities of alcohol at one time can cause organ damage, which in turn can lead to chronic diseases. It is of important clinical and social significance to find effective approaches for the prevention and treatment of alcohol-induced diseases. In this paper, sulfhydryl functionalized chitosan (chitosan-N-acetyl-l-cysteine, CS-NAC) and sodium alginate (SA) were used as the matrix materials to contain tilapia peptide (TP), and a gastric acid-response hydrogel (CS-NAC/SA/TP) was prepared. Taking the ethanol adsorption rate as the response index, based on the results of the single factor test, the preparation process of CS-NAC/SA/TP was optimized through the Box-Behnken design. The swelling and antioxidant properties of CS-NAC/SA/TP were tested in vitro, and the protective effects on alcohol-induced acute liver injury and chronic brain injury were assessed in vivo. Structural characterization showed that CS-NAC/SA/TP was successfully prepared. Under the optimal conditions (SA concentration of 1%, M CS-NAC/M CaCO3 of 1 : 1, M SA/M CS-NAC(CaCO3) of 15 : 1), the prepared CS-NAC/SA/TP had a porous structure, a swelling ratio of 2350%, an ethanol adsorption rate of 56.23% and strong antioxidant capacities in vitro. Animal experiment results demonstrated that CS-NAC/SA/TP effectively reduced liver and brain injuries in mice caused by alcoholism. Summarily, these findings indicate that CS-NAC/SA/TP has potential applications in preventing alcohol-induced liver and brain injuries.

Use of 18ß-glycyrrhetinic acid nanocrystals to enhance anti-inflammatory activity by improving topical delivery.

18ß-Glycyrrhetinic acid (GA) is often topically applied in clinical treatment of inflammatory skin diseases. However, GA has poor solubility in water, which results in poor skin permeability and low bioavailability. Nanocrystallization of drugs can enhance their permeability and improve bioavailability. We prepared GA nanocrystals (Nano GA) by high-pressure homogenization. These nanocrystals were characterized by photon correlation spectroscopy, scanning electron microscopy, thermogravimetric analysis, and X-ray diffractometry. The ability of Nano GA to improve dermal permeability was investigated ex vivo using Franz diffusion vertical cells and mouse skin. The topical anti-inflammatory activity of Nano GA was assessed in vivo by a 12-O-tetradecanoyl phorbol-13-acetate (TPA)-induced model in mouse ears. The average particle size of a GA nanocrystalline suspension was 288.6 ± 7.3 nm, with a narrow particle-size distribution (polydispersity index ∼0.13 ± 0.10), and the particle size of the lyophilized powder increased (552.0 ± 9.8 nm). After nanocrystallization, the thermal stability and crystallinity decreased but solubility increased significantly. Nano GA showed higher dermal permeability than Coarse GA. Macroscopic and staining-based observations of mouse ears and the levels of proinflammatory factors and myeloperoxidase revealed that the Nano GA hydrogel exhibited better anti-edema ability and more strongly inhibited inflammation development than the Coarse GA hydrogel and indomethacin hydrogel (positive drug). These results suggest that Nano GA could be an efficacious topical therapeutic agent for skin inflammation.

Protective effects of collagen polypeptide from tilapia skin against injuries to the liver and kidneys of mice induced by d-galactose.

We wished to investigate the role of a tilapia skin collagen polypeptide (TSCP; molecular weight <3 kDa) in alleviating liver and kidney injuries in aging mice induced by d-galactose (d-gal) and its underlying mechanism of action. First, we characterized TSCP. TSCP was passed through a 3-kDa ultrafiltration membrane, desalted in water by a solid-phase extraction column, purified further by reverse phase-high performance liquid chromatography, and analyzed by electrospray ionization mass spectrometry and tandem mass spectrometry. TSCP contained 17 types of amino acids (AAs) and 41 peptide chains of length 7 AAs to 22 AAs. The content of free AAs and total AAs of TSCP was 13.5% and 93.79%, respectively. Next, we undertook animal experiments. Mice were injected once-daily with D-gal (300 mg/kg body weight, s.c.) for 8 weeks, and TSCP was administered simultaneously once-daily by intragastric gavage. TSCP could visibly improve the decreased body weight, depressed appetite, and mental deterioration of mice triggered by d-gal. TSCP could also alleviate d-gal-induced damage to the liver and kidneys according to histopathology (especially high-dose TSCP). Consistent with these macroscopic and pathologic changes, TSCP could also prevent d-gal-induced increases in serum levels of alanine aminotransferase, aspartate transaminase, alkaline phosphatase, lipid peroxidation, creatinine and uric acid, as well as decreases in serum levels of immunoglobulin (Ig)G and IgM. Moreover, TSCP improved the activities of superoxide dismutase, catalase, and glutathione peroxidase, but also inhibited the increases in the levels of malondialdehyde and inducible nitric oxide synthase expression in the liver and kidneys of d-gal-treated mice. These results suggest that TSCP can alleviate the injuries to the liver and kidneys in aging mice induced by d-gal, and that its mechanism of action might be, at least partially, associated with attenuation of oxidative stress and enhancement of immune function.

Preparation and evaluation of squid ink polysaccharide-chitosan as a wound-healing sponge.

A new type of wound healing agent was developed using two marine biomaterials (squid ink polysaccharide and chitosan) as carriers and calcium chloride as an initiator for coagulation. Based on central composite design-response surface methodology, comprehensive evaluation of appearance quality for composite sponges and water absorbency were used as evaluation indices to identify the optimized preparation conditions and further evaluate the performance of the squid ink polysaccharide-chitosan sponge (SIP-CS). The optimized formulation of SIP-CS was as follows: chitosan concentration, 2.29%; squid ink polysaccharide concentration, 0.55%; and calcium chloride concentration, 2.82%, at a volume ratio of 15:5:2. SIP-CS was conducive to sticking on the wound, characterized by the spongy property, strong absorptivity, and tackiness. Rabbit ear arterial, hepatic, and femoral artery hemorrhage experiments indicated that, compared with chitosan dressings and absorbable gelatin, the hemostatic times were shorter and the bleeding volume was smaller. Furthermore, SIP-CS absorbed a large amount of hemocytes, leading to rapid hemostasis. The healing areas and wound pathological sections in scalded New Zealand rabbits indicated that SIP-CS promoted wound healing more rapidly than chitosan and better than commercially available burn cream. Thus, SIP-CS is a good wound healing agent for rapid hemostasis, promoting burn/scalded skin healing, and protecting from wound infection.

Anti-photoaging effects of chitosan oligosaccharide in ultraviolet-irradiated hairless mouse skin.

Skin photoaging (SP) is a premature skin-aging damage after repeated exposure to ultraviolet (UV) radiation, mainly characterized by oxidative stress and inflammatory disequilibrium, which makes skin show the typical symptoms of photoaging such as coarse wrinkling, dryness, irregular pigmentation and laxity. Chitosan oligosaccharide (COS), a natural polysaccharide with good humectant property, is the depolymerized product of chitosan with various biological activities, among which the antioxidant and anti-inflammatory effects have been frequently reported in recent years. However, no existing invivo study indicates whether COS has direct protective effect on UV-induced SP. In the current research, we investigated the potential preventive effect of COS against UV-caused damage in hairless mouse dorsal skin. The data showed that COS, by topical application after each UV-radiation for 10weeks, effectively inhibited the undesirable changes on the skin induced by UV. To be specific, COS obviously alleviated the macroscopic and histopathological damages of mice skin, via mitigating the disrupted collagenous fibers, as well as improving the relative content of type I collagen and the amount of total collagen. Furthermore, COS effectively inhibited the levels of pro-inflammatory cytokines such as TNF-α, IL-1ß and IL-6, and markedly improved the activities of antioxidant enzymes (SOD, GSH-Px, CAT), as well as the content of skin hydroxyproline and moisture. These findings demonstrated that this natural polysaccharide attenuated UV-induced SP, at least in part, by virtue of favorable regulation of antioxidant and anti-inflammatory status, which presumably worked in concert to maintain the morphology and level of dermal collagen.

Preparation and Characterization of Chitosan-Agarose Composite Films.

Nowadays, there is a growing interest to develop biodegradable functional composite materials for food packaging and biomedicine applications from renewable sources. Some composite films were prepared by the casting method using chitosan (CS) and agarose (AG) in different mass ratios. The composite films were analyzed for physical-chemical-mechanical properties including tensile strength (TS), elongation-at-break (EB), water vapor transmission rate (WVTR), swelling ratio, Fourier-transform infrared spectroscopy, and morphology observations. The antibacterial properties of the composite films were also evaluated. The obtained results reveal that an addition of AG in varied proportions to a CS solution leads to an enhancement of the composite film's tensile strength, elongation-at-break, and water vapor transmission rate. The composite film with an agarose mass concentration of 60% was of the highest water uptake capacity. These improvements can be explained by the chemical structures of the new composite films, which contain hydrogen bonding interactions between the chitosan and agarose as shown by Fourier-transform infrared spectroscopy (FTIR) analysis and the micro-pore structures as observed with optical microscopes and scanning electron microscopy (SEM). The antibacterial results demonstrated that the films with agarose mass concentrations ranging from 0% to 60% possessed antibacterial properties. These results indicate that these composite films, especially the composite film with an agarose mass concentration of 60%, exhibit excellent potential to be used in food packaging and biomedical materials.

(-)-Patchouli alcohol protects against Helicobacter pylori urease-induced apoptosis, oxidative stress and inflammatory response in human gastric epithelial cells.

(-)-Patchouli alcohol (PA), the major active principle of Pogostemonis Herba, has been reported to have anti-Helicobacter pylori and gastroprotective effects. In the present work, we aimed to investigate the possible protective effect of PA on H. pylori urease (HPU)-injured human gastric epithelial cells (GES-1) and to elucidate the underlying mechanisms of action. Results showed that pre-treatment with PA (5.0, 10.0, 20.0µM) was able to remarkably ameliorate the cytotoxicity induced by 17.0U/mg HPU in GES-1 cells. Flow cytometric analysis on cellular apoptosis showed that pre-treatment with PA effectively attenuated GES-1 cells from the HPU-induced apoptosis. Moreover, the cytoprotective effect of PA was found to be associated with amelioration of the HPU-induced disruption of MMP, attenuating oxidative stress by decreasing contents of intracellular ROS and MDA, and increasing superoxide dismutase (SOD) and catalase (CAT) enzymatic activities. In addition, pre-treatment with PA markedly attenuated the secretion of nitric oxide (NO) and pro-inflammatory cytokines such as interleukin-2 (IL-2), interleukin-4 (IL-4) and tumor necrosis factor-α (TNF-α), whereas elevated the anti-inflammatory cytokine interleukin-13 (IL-13) in the HPU-stimulated GES-1 cells. Molecular docking assay suggested that PA engaged in the active site of urease bearing nickel ions and interacted with important residues via covalent binding, thereby restricting the active urease catalysis conformation. Our experimental findings suggest that PA could inhibit the cellular processes critically involved in the pathogenesis of H. pylori infection, and its protective effects against the HPU-induced cytotoxicity in GES-1 cells are believed to be associated with its anti-apoptotic, antioxidative, anti-inflammatory and HPU inhibitory actions.

Meta-analysis of estradiol for luteal phase support in in vitro fertilization/intracytoplasmic sperm injection.

OBJECTIVE: To evaluate whether the addition of E(2) for luteal phase support (LPS) in IVF/intracytoplasmic sperm injection (ICSI) could improve the outcome of clinical pregnancy. DESIGN: Meta-analysis. SETTING: University hospital center. PATIENT(S): Women underwent IVF or ICSI using the GnRH agonist or GnRH antagonist protocol. INTERVENTION(S): Progesterone alone or combined with E(2) for LPS. MAIN OUTCOME MEASURE(S): Clinical pregnancy rate per patient (CPR/PA), clinical pregnancy rate per ET, implantation rate, ongoing pregnancy rate per patient, clinical abortion rate, and ectopic pregnancy rate. RESULT(S): Fifteen relevant randomized controlled trials (RCTs) were identified that included a total of 2,406 patients. There was no statistical difference between E(2) + P group and P-only group regarding the primary outcome of CPR/PA for different routes of administration of E(2) (oral, vaginal, and transdermal) or other relevant outcome measures. No significant effect was observed for different daily doses of E(2) (6, 4, and 2 mg), even through oral medication in CPR/PA. CONCLUSION(S): The best available evidence suggests that E(2) addition during the luteal phase does not improve IVF/ICSI outcomes through oral medication, even with different daily doses. Furthermore, RCTs that study other administration routes are needed.

Protective effects of apigenin against 1-methyl-4-phenylpyridinium ion­induced neurotoxicity in PC12 cells.

Parkinson's disease is recognized as the second most common neurodegenerative disorder after Alzheimer's disease, characterized by the loss of dopominergic neurons in the substantia nigra pars compacta and can be experimentally mimicked by the use of the neurotoxin, 1­methyl­4­phenylpyridinium ion (MPP(+)), in in vitro models. In this study, we investigated the potential protective effects of apigenin (AP), galangin and genkwanin, naturally occurring plant flavonoids, on the MPP(+)­induced cytotoxicity in cultured rat adrenal pheochromocytoma cells (PC12 cells). The PC12 cells were pre-treated with various concentrations of the test compounds for 4 h, followed by the challenge with 1,000 µM MPP(+) for 48 h. We found that only pre-treatment with AP (3, 6 and 12 µM) before injury significantly increased cell viability, decreased the release of lactate dehydrogenase, reduced the level of intracellular reactive oxygen species and elevated mitochondrial membrane potential in the MPP(+)­treated PC12 cells. In addition, AP markedly suppressed the increased rate of apoptosis and the reduced Bcl­2/Bax ratio induced by MPP(+) in the PC12 cells. Taken together, the findings of this study demonstrate that AP exerts neuroprotective effects against MPP(+)­induced neurotoxicity in PC12 cells, at least in part, through the inhibition of oxidative damage and the suppression of apoptosis through the mitochondrial pathway.