PDA modified NIR-II NaEr0.8Yb0.2F4nanoparticles with high photothermal effect.

Polydopamine (PDA)-modified NaEr0.8Yb0.2 F4nanoparticles were synthesized, with strong NIR-II emission, quantum yield of 29.63%, and excellent photothermal performance. Crystal phases and microstructures are characterized. Optical properties such as absorption, NIR-II emission, and light stability are studied, and the luminescence mechanism is discussed in detail. Key factors in NIR-II imaging were evaluated in fresh pork tissue, including penetration depth, spatial resolution, and signal-to-noise ratio (SNR). A high penetration depth of 5 mm and a high spatial resolution of 1 mm were detected. Mice are imaged in vivo afterintravenousinjection. Due to the accumulation of nanoparticles in the liver, high image quality with an SNR of 5.2 was detected in the abdomen of KM mice with hair. The photothermal conversion effect of PDA-modified NPs was twice that of the reported material. These NIR-II nanoparticles have superior optical properties, high photothermal efficiency and low cytotoxicity, and are potential fluorescent probes for further disease diagnosis and treatment.

Effects of astaxanthin on plasma membrane function and fertility of boar sperm during cryopreservation.

Mammalian sperm is highly susceptible to reactive oxygen species (ROS) during the cryopreservation process. Astaxanthin (AST), a red pigment of the carotenoid family, is recognized as having a variety of beneficial biological activities and effects, including antioxidant, anticancer, anti-diabetic, and anti-inflammatory. The present study aimed to investigate whether the presence of AST protected boar sperm from ROS stress during cryopreservation. Boar sperm was diluted with a freezing medium supplemented with different concentrations of AST (0, 0.5, 1, 2, or 5 µM). The addition of AST, especially at a concentration of 2 µM, exerted positive effects on post-thaw sperm motility parameters. Meanwhile, sperm plasma membrane integrity and acrosome integrity of post-thaw sperm were significantly increased, while lipid peroxidation was inhibited in response to 2 µM AST treatment. Interestingly, compared to the control, supplementation with 2 µM AST increased unsaturated fatty acids (UFAs) levels and decreased saturated fatty acids (SFAs) content in post-thaw sperm, leading to a decreased ratio of SFAs/UFAs in the AST group. In conclusion, the addition of AST to freezing extenders inhibited lipid peroxidation and regulated fatty acid composition of the sperm membrane, improved post-thaw sperm quality, and had no adverse effect on boar sperm in vitro fertilization (IVF) capacity and potential for embryonic development. Our data provide a novel insight into understanding the mechanisms of AST concerning protecting boar sperm quality against ROS damage during cryopreservation.

Effect of L-carnitine on sperm quality during liquid storage of boar semen.

OBJECTIVE: This study was conducted to investigate the effect of L-carnitine on the pig semen characteristics during storage. METHODS: Spermatozoa samples were examined for spermatozoa quality and then randomly divided into 5 groups: 0 (control), 12.5, 25, 50, and 100 mM L-carnitine. Sperm motility, plasma membrane integrity and antioxidant parameters (total reactive oxygen species, total antioxidant capacity, and malondialdehyde) were evaluated after 0, 3, 5, and 10 day cooledstorage at 17°C. Moreover, ATP content, mitochondria activity as well as sperm-binding and in vitro fertilizing ability of preserved boar sperm were also investigated. RESULTS: Supplementation with 50 mM L-carnitine could effectively maintain boar sperm quality parameters such as sperm motility and membrane integrity. Besides, we found that L-carnitine had positive effects on boar sperm quality mainly through improving antioxidant capacities and enhancing ATP content and mitochondria activity. Interestingly, by assessing the effect of L-carnitine on sperm fertility and developmental potential, we discovered that the extender containing L-carnitine could improve sperm quality and increase the number of sperms bounding to zona pellucida, without improving in vitro fertility and development potential. CONCLUSION: These findings suggested that the proper addition of L-carnitine to the semen extender improved boar sperm quality during liquid storage at 17°C.

Simultaneous separation and determination of seven isoflavones in Radix Puerariae by capillary electrophoresis with a dual cyclodextrin system.

A simple, comprehensive and efficient capillary electrophoresis method using a dual cyclodextrin system was developed for the simultaneous determination of seven isoflavones (3'-methoxypuerarin, puerarin, 3'-hydroxypuerarin, ononin, daidzin, daidzein and genistin). Baseline separations of the seven isoflavones were achieved within 11 min with the running buffer consisting of 35 mm sodium tetraborate, 9.0 mm sulfobutylether-ß-cyclodextrin and 30 mm α-cyclodextrin at pH 9.34, and peaks were detected at 254 nm. Other separation parameters included the separation voltage for 15 kV and the working temperature for 25°C. Under the optimum conditions, good linearities were obtained with linear correlation coefficients of seven isoflavones of 0.9978-0.9992. The limits of detection and the limits of quantification were 0.7-2.9 and 2.5-9.5 µg/mL, respectively. Excellent precision and accuracy were obtained. The intraday and interday precision ranged from 0.7 to 2.0% and from 0.8 to 1.9%, respectively. The recoveries of seven analytes were from 97.7 to 103.1%. This method was successfully applied to determine the seven analytes in Radix Puerariae and its preparations.

Separation and simultaneous determination of seven bioactive components in Tripterygium wilfordii Hook. F. and Tripterygium preparations by micellar electrokinetic capillary chromatography.

A simple, comprehensive, and highly selective MEKC method has been developed for simultaneous analysis of seven bioactive components (triptolide, wilfortrine, wilfordine, wilforgine, wilforine, triptophenolide, and triptonide) in the root extracts of Tripterygium wilfordii Hook. F. (TWHF) and Tripterygium preparations (TPs). Optimal BGE consisted of 10 mM sodium tetraborate, 30 mM SDS, and 30% v/v methanol. The separation voltage was 20 kV and the temperature was 25°C. A DAD was used and the detection wavelength was at 218 nm. Under the optimum conditions, the baseline separation of seven components was achieved in less than 26 min. Excellent precision, good stability, and accuracy were obtained. For all analytes, linear calibrations were established within 10-100 µg/mL. The LOD and LOQ were within 1.2-4.2 µg/mL and 4.0-14 µg/mL, respectively. The developed method was suitable for the determination of key components in TWHF and TPs.

Effects of L-glutamine on boar sperm quality during liquid storage at 17°C.

The quality of boar spermatozoa is affected by oxidative stress during preservation in vitro. It has been demonstrated that L-Glutamine (Gln) can effectively protect cells from oxidative stress-induced injury. There are, however, no reports to date evaluating the effects of Gln on boar semen liquid preservation at 17 °C. The aims of the present study were to elucidate whether the addition of Gln to the extender BTS could improve the quality of boar spermatozoa when stored at 17 °C and to determine the mechanism underlying Gln protection of spermatozoa against preservation-induced damage. Boar semen samples were collected and diluted with Beltsville Thawing Solution (BTS) containing different concentrations (0, 10, 20, 40 or 80 mM) of Gln. The results indicated the addition of 20 mM Gln to the BTS improved (P < 0.05) the motility, acrosome integrity and membrane integrity of boar sperm during liquid preservation. Interestingly, treatment of spermatozoa with Gln addition to the extender resulted in ROS quenching, while enhancing γ-glutamyl cysteine synthetase (γ-GCS) activity, and glutathione (GSH) content of spermatozoa. These results suggest that BTS supplemented with Gln can provide greater protective capacity to boar sperm against oxidative stress by enhancing GSH synthesis during liquid preservation.

Identification of hydrolyzable tannins (punicalagin, punicalin and geraniin) as novel inhibitors of hepatitis B virus covalently closed circular DNA.

The development of new agents to target HBV cccDNA is urgently needed because of the limitations of current available drugs for treatment of hepatitis B. By using a cell-based assay in which the production of HBeAg is in a cccDNA-dependent manner, we screened a compound library derived from Chinese herbal remedies for inhibitors against HBV cccDNA. Three hydrolyzable tannins, specifically punicalagin, punicalin and geraniin, emerged as novel anti-HBV agents. These compounds significantly reduced the production of secreted HBeAg and cccDNA in a dose-dependent manner in our assay, without dramatic alteration of viral DNA replication. Furthermore, punicalagin did not affect precore/core promoter activity, pgRNA transcription, core protein expression, or HBsAg secretion. By employing the cell-based cccDNA accumulation and stability assay, we found that these tannins significantly inhibited the establishment of cccDNA and modestly facilitated the degradation of preexisting cccDNA. Collectively, our results suggest that hydrolyzable tannins inhibit HBV cccDNA production via a dual mechanism through preventing the formation of cccDNA and promoting cccDNA decay, although the latter effect is rather minor. These hydrolyzable tannins may serve as lead compounds for the development of new agents to cure HBV infection.

Analysis of human innate immune responses to PRINT fabricated nanoparticles with cross validation using a humanized mouse model.

Ideal nanoparticle (NP)-based drug and vaccine delivery vectors should be free of inherent cytotoxic or immunostimulatory properties. Therefore, determining baseline immune responses to nanomaterials is of utmost importance when designing human therapeutics. We characterized the response of human immune cells to hydrogel NPs fabricated using Particle Replication in Non-wetting Templates (PRINT) technology. We found preferential NP uptake by primary CD14(+) monocytes, which was significantly reduced upon PEGylation of the NP surface. Multiplex cytokine analysis of NP treated primary human peripheral blood mononuclear cells suggests that PRINT based hydrogel NPs do not evoke significant inflammatory responses nor induce cytotoxicity or complement activation. We furthered these studies using an in vivo humanized mouse model and similarly found preferential NP uptake by human CD14(+) monocytes without systemic inflammatory cytokine responses. These studies suggest that PRINT hydrogel particles form a desirable platform for vaccine and drug delivery as they neither induce inflammation nor toxicity. From the clinical editor: The authors here fabricated hydrogel nanorods using the PRINT (Particle Replication In Nonwetting Templates) fabrication process. They tested the interaction of human immune cells with these particles and found no immunoreactivity. This finding would suggest that monodisperse PRINT particles of identical shape and size could serve a variety of clinical applications.

Mutant Enpp1asj mice as a model for generalized arterial calcification of infancy.

Generalized arterial calcification of infancy (GACI), an autosomal recessive disorder, is characterized by early mineralization of blood vessels, often diagnosed by prenatal ultrasound and usually resulting in demise during the first year of life. It is caused in most cases by mutations in the ENPP1 gene, encoding an enzyme that hydrolyzes ATP to AMP and inorganic pyrophosphate, the latter being a powerful anti-mineralization factor. Recently, a novel mouse phenotype was recognized as a result of ENU mutagenesis - those mice developed stiffening of the joints, hence the mutant mouse was named 'ages with stiffened joints' (asj). These mice harbor a missense mutation, p.V246D, in the Enpp1 gene. Here we demonstrate that the mutant ENPP1 protein is largely absent in the liver of asj mice, and the lack of enzymatic activity results in reduced inorganic pyrophosphate (PPi) levels in the plasma, accompanied by extensive mineralization of a number of tissues, including arterial blood vessels. The progress of mineralization is highly dependent on the mineral composition of the diet, with significant shortening of the lifespan on a diet enriched in phosphorus and low in magnesium. These results suggest that the asj mouse can serve as an animal model for GACI.

Screening and identification of compounds with antiviral activity against hepatitis B virus using a safe compound library and novel real-time immune-absorbance PCR-based high throughput system.

There are now seven nucleoside/tide analogues, along with interferon-α, that are approved by the FDA for the management of chronic hepatitis B virus (HBV) infection, a disease affecting hundreds of millions of people worldwide. These medications, however, are limited in usefulness, and significant side effects and the emergence of viral escape mutants make the development of novel and updated therapeutics a pressing need in the treatment of HBV. With this in mind, a library containing 2000 compounds already known to be safe in both humans and mice with known mechanisms of action in mammalian cells were tested for the possibility of either antiviral activity against HBV or selective toxicity in HBV producing cell lines. A modified real-time immune-absorbance-polymerase chain reaction (IA-PCR) assay was developed for this screen, utilizing cells that produce and secrete intact HBV virions. In this procedure, viral particles are first captured by an anti-HBs antibody immobilized on a plate. The viral load is subsequently assessed by real-time PCR directly on captured particles. Using this assay, eight compounds were shown to consistently reduce the amount of secreted HBV viral particles in the culture medium under conditions that had no detectable impact on cell viability. Two compounds, proparacaine and chlorophyllide, were shown to reduce HBV levels 4- to 6-fold with an IC50 of 1 and 1.5 µM, respectively, and were selected for further study. The identification of these compounds as promising antiviral drug candidates against HBV, despite a lack of previous recognition of HBV antiviral activity, supports the validity and utility of testing known compounds for "off-pathogen target" activity against HBV, and also validates this IA-PCR assay as an important tool for the detection of anti-viral activity against enveloped viruses.

Warfarin accelerates ectopic mineralization in Abcc6(-/-) mice: clinical relevance to pseudoxanthoma elasticum.

Pseudoxanthoma elasticum (PXE) is a multisystem ectopic mineralization disorder caused by mutations in the ABCC6 gene. Warfarin, a commonly used anticoagulant, is associated with increased mineralization of the arterial blood vessels and cardiac valves. We hypothesized that warfarin may accelerate ectopic tissue mineralization in PXE, with clinical consequences. To test this hypothesis, we developed a model in which Abcc6(-/-) mice, which recapitulate features of PXE, were fed a diet supplemented with warfarin and vitamin K1. Warfarin action was confirmed by significantly increased serum levels of oxidized vitamin K. For mice placed on a warfarin-containing diet, quantitative chemical and morphometric analyses revealed massive accumulation of mineral deposits in a number of tissues. Mice fed a warfarin-containing diet were also shown to have abundant uncarboxylated form of matrix Gla protein, which allowed progressive tissue mineralization to ensue. To explore the clinical relevance of these findings, 1747 patients with PXE from the approximately 4000 patients in the PXE International database were surveyed about the use of warfarin. Of the 539 respondents, 2.6% reported past or present use of warfarin. Based on the prevalence of PXE (approximately 1:50,000), thousands of patients with PXE worldwide may be at risk for worsening of PXE as a result of warfarin therapy.

[Seasonal variation characteristics of phosphorus speciation in the sediment of Changshou Lake].

To further know the seasonal variation characteristics of phosphorus species in lake sediment, the contents of different phosphorus species in the surface sediment sampling from Changshou Lake were determined by using the SMT sequential extraction method in July and November, 2009, and March, 2010. The contents of total phosphorus (TP) basically show an order of the low-water period, high-water period, and normal-water period from high to low, and the maximum appeared in Shoudao (2960.29 mg/kg) while the minimum in Guanjiakou (586.05 mg/kg). The mass fractions of phosphorus associated with hydrous ferric/aluminum oxides (Fe/Al-P) and organic phosphorus (OP) in TP at all sampling sits are in an order of the normal-water period, high-water period, and low-water period from high to low, and the maximum of Fe/Al-P appeared in Lewen (40.01%) while that of OP in the dam mouth (72.44%). The mass fraction of phosphorus bound to calcium salt (Ca-P) in TP is in an order of the low-water period, normal-water period, and high-water period, and the maximum appeared in Baibudang (15.52%). These results suggested that the phosphorus speciation in the sediment of Changshou Lake shows obviously temporal and spatial distribution characteristics, and Fe/Al-P is mainly influenced by pH of the overlying water, while OP is not apparently influenced by dissolved oxygen (DO).

Hepatitis B virus e antigen production is dependent upon covalently closed circular (ccc) DNA in HepAD38 cell cultures and may serve as a cccDNA surrogate in antiviral screening assays.

Currently available antiviral nucleoside analogs for the treatment of chronic hepatitis B virus (HBV) infections profoundly reduce virus load, but rarely cure the virus infection. This is due, at least in part, to their failure to eliminate viral covalently closed circular (ccc) DNA from the nuclei of infected hepatocytes. To screen compound libraries for antiviral drugs targeting cccDNA, we set out to develop a cell-based assay suitable for high throughput screening. Since cccDNA is time-consuming to assay, it was desirable to use a viral gene product that could serve as a reporter for intracellular cccDNA level. We predicted that the secretion of HBV e antigen (HBeAg) by HepAD38 cells, a tetracycline inducible HBV expression cell line, would be cccDNA-dependent. This is because a large portion of pre-core mRNA leader sequence in the 5' terminus of integrated viral genome was deleted, preventing HBeAg expression from transgene, but could be restored from the 3' terminal redundancy of pre-genomic RNA during viral DNA replication and subsequent cccDNA formation. Our experimental results showed that following induction, HepAD38 produced and accumulated cccDNA, which became detectable between 7 and 8 days. HBeAg synthesis and secretion into culture fluid were dependent upon and proportional to the level of cccDNA detected. Therefore, the secretion of HBeAg by HepAD38 cells could potentially serve as a convenient reporter for the high throughput screening of novel antiviral drugs targeting HBV cccDNA.