In Vitro and In Vivo Demonstration of Ultraefficient and Broad-Spectrum Antibacterial Agents for Photodynamic Antibacterial Chemotherapy.

Increasing threats from both pathogenic infections and antibiotic resistance highlight the pressing demand for nonantibiotic agents and alternative therapies. Herein, we report several new phenothiazinium-based derivatives, which could be readily synthesized via fragment-based assembly, which exhibited remarkable bactericidal activities both in vitro and in vivo. Importantly, in contrast to numerous clinically and preclinically used antibacterial photosensitizers, these compounds were able to eliminate various types of microorganisms, including Gram-(+) Staphylococcus aureus (S. aureus), Gram-(-) Escherichia coli, multidrug-resistant S. aureus, and their associated biofilms, at low drug and light dosages (e.g., 0.21 ng/mL in vitro and 1.63 ng/cm2 in vivo to eradicate S. aureus at 30 J/cm2). This study thus unveils the potential of these novel phenothiaziniums as potent antimicrobial agents for highly efficient photodynamic antibacterial chemotherapy.

Effects of Glycyrrhiza polysaccharide in diet on growth performance, serum antioxidant capacity, and biochemistry of broilers.

In the present study, we analyzed the effects of Glycyrrhiza polysaccharide (GCP) on growth performance, serum antioxidant capacity, and biochemistry of broilers. A total of 600, one-day-old AA broilers randomly divided into 5 treatment groups with 6 replicate pens of 20 birds per cage received dietary supplementation with GCP (0, 200, 500, 1,000, and 1,500 mg/kg) for 42 d. The supplementation of GCP linearly decreased (P < 0.05) feed conversion rate on day 22 to 42. Dietary supplementation with GCP reduced (P < 0.05) serum total cholesterol on day 21 and 42 and linearly improved (P < 0.05) albumin and high-density lipoprotein cholesterol. Dietary supplementation with 1,000 or 1,500 mg/kg GCP significantly increased (P < 0.05) serum total superoxide dismutase (T-SOD) activity on day 21 and 42 and reduced (P < 0.05) serum malondialdehyde content on 21 d. Dietary supplementation with 1,000 or 1,500 mg/kg GCP significantly improved (P < 0.05) interleukin-1ß (IL-1ß) and interferon-γ (IFN-γ) expressions in liver on day 21 and 42. At the end of the experiment, we randomly selected 20 broilers from 3 treatment groups (0, 1,000, and 1,500 mg/kg), respectively, to perform an lipopolysaccharide (LPS)-induced acute stress experiment. The 60 broilers were divided into 6 treatment groups with 10 birds per cage. The experiment was designed as a 3 × 2 factorial arrangement with GCP (0, 1,000, or 1,500 mg/kg) and LPS (injection of saline or 1 mg/kg body weight) levels as treatments. When the grouping was finished, the broilers were immediately intraperitoneally injected with LPS or normal saline. Six hours after challenged, serum antioxidant and liver immunity were analyzed. The results showed that dietary GCP prevented LPS-induced reductions in T-SOD activity and increases in malonaldehyde content (P < 0.05). Also, dietary GCP supplementation mitigated the LPS-induced increase in IL-1ß and IFN-γ in the liver. Supplementation with 1,500 mg/kg GCP showed the most optimal effect in broilers. GCP has the potential to be used as feed additive in broilers.

Smart Design of Nanomaterials for Mitochondria-Targeted Nanotherapeutics.

Mitochondria are the powerhouse of cells. They are vital organelles that maintain cellular function and metabolism. Dysfunction of mitochondria results in various diseases with a great diversity of clinical appearances. In the past, strategies have been developed for fabricating subcellular-targeting drug-delivery nanocarriers, enabling cellular internalization and subsequent organelle localization. Of late, innovative strategies have emerged for the smart design of multifunctional nanocarriers. Hierarchical targeting enables nanocarriers to evade and overcome various barriers encountered upon in vivo administration to reach the organelle with good bioavailability. Stimuli-responsive nanocarriers allow controlled release of therapeutics to occur at the desired target site. Synergistic therapy can be achieved using a combination of approaches such as chemotherapy, gene and phototherapy. In this Review, we survey the field for recent developments and strategies used in the smart design of nanocarriers for mitochondria-targeted therapeutics. Existing challenges and unexplored therapeutic opportunities are also highlighted and discussed to inspire the next generation of mitochondrial-targeting nanotherapeutics.

MSN-on-a-Chip: Cell-Based Screenings Made Possible on a Small-Molecule Microarray of Native Natural Products.

Standard small-molecule microarrays (SMMs) are not well-suited for cell-based screening assays. Of the few attempts made thus far to render SMMs cell-compatible, all encountered major limitations. Here we report the first mesoporous silica nanoparticle (MSN)-on-a-chip platform capable of allowing high-throughput cell-based screening to be conducted on SMMs. By making use of a glass surface on which hundreds of MSNs, each encapsulated with a different native natural product, were immobilized in spatially defined manner, followed by on-chip mammalian cell growth and on-demand compound release, high-content screening was successfully carried out with readily available phenotypic detection methods. By combining this new MSN-on-a-chip system with small interfering RNA technology for the first time, we discovered that (+)-usniacin possesses synergistic inhibitory properties similar to those of olaparib (an FDA-approved drug) in BRCA1-knockdown cancer cells.

A method for small molecule microarray-based screening for the rapid discovery of affinity-based probes.

We describe herein a new method for the high-throughput identification of affinity-based probes (AfBPs) using a small molecule microarray (SMM) approach. A hydroxylethylene-based small molecule library was first generated by solid-phase combinatorial synthesis. The library was tagged with biotin to facilitate immobilization on avidin-coated slides. Preliminary screening with γ-secretase (both the recombinantly purified protein as well as cellular lysates overexpressing the enzyme) was carried out, in order to identify potential small molecule binders, which were subsequently redesigned into AfBPs. Several specific and potent probes for γ-secretase were thus identified through the binding profiles observed on the SMMs. The SMM platform was able to sensitively and conveniently report activity-based binding interactions between aspartic proteases and their small molecule inhibitors. This new approach thus provides a potentially more rapid and efficient method for developing AfBPs using SMMs.

Rapid synthesis of Abelson tyrosine kinase inhibitors using click chemistry.

Protein kinases catalyze the phosphorylation of serine, threonine, tyrosine and histidine residues in proteins. Aberrant regulation of kinase activity has been implicated in many diseases including cancer. Thus development of new strategies for kinase inhibitor design remains an active area of research with direct relevance to drug development. Abelson (Abl) tyrosine kinase is one of the Src-family of tyrosine kinases and is directly implicated in Chronic Myelogenous Leukemia (CML). In this article, we have, for the first time, developed an efficient method for the construction of small molecule-based bisubstrate inhibitors of Abl kinase using click chemistry. Subsequent biochemical screenings revealed a set of moderately potent inhibitors, a few of which have comparable potency to Imatinib (an FDA-approved drug for treatment of chronic myeloid leukemia) against Abl.

High-throughput synthesis of azide libraries suitable for direct "click" chemistry and in situ screening.

A key challenge in current drug discovery is the development of high-throughput (HT) amenable chemical reactions that allow rapid synthesis of diverse chemical libraries of enzyme inhibitors. The Cu(I)-catalyzed, 1,3-dipolar cycloaddition between an azide and an alkyne, better known as "click chemistry", is one such method that has received the most attention in recent years. Despite its popularity, there is still a lack of robust and efficient chemical strategies that give access to diverse libraries of azide-containing building blocks (key components in click chemistry). We report herein a highly robust and efficient strategy for high-throughput synthesis of a 325-member azide library. The method is highlighted by its simplicity and product purity. The utility of the library is demonstrated with the subsequent "click" synthesis of the corresponding bidentate inhibitors against PTP1B.

Solid-phase assembly and in situ screening of protein tyrosine phosphatase inhibitors.

A highly efficient solid-phase strategy for assembly of small molecule inhibitors against protein tyrosine phosphatase 1B (PTP1B) is described. The method is highlighted by its simplicity and product purity. A 70-member combinatorial library of analogues of a known PTP1B inhibitor has been synthesized, which upon direct in situ screening revealed a potent inhibitor ( Ki = 7.0 microM) against PTP1B.

Solid-phase synthesis of azidomethylene inhibitors targeting cysteine proteases.

An efficient strategy for the solid-phase synthesis of azidomethylene inhibitors targeting cysteine proteases is described. The method is highlighted by its compatibility with readily available building blocks, as well as its ability to accommodate different functional groups. A 249-member library has thus far been successfully synthesized, characterized, and screened against Caspase-1, -3 and -7.

[Study on anticancer activity of Syngnathus in vitro].

OBJECTIVE: To study the anticancer activity of Syngnathus in vitro. METHOD: Observing the influence of different extracts from Syngnathus on growth of different cancer cell strains by MTT method. RESULT: It has been found out that the fat-soluble nonsaponified extract from Temminck et Schlegel and the alcoholic extract from Syngnathus acus have cytotoxic activities. The nonsaponified extract from Temminck et Schlegel can inhibit the growth of cancer cell strains KB, Hela, PAA, K562, and Bcap37, and the alcoholic extract from Syngnathus acus can inhibit the growth of cancer cell strin KB, But Bloch shows no apparent anticancer activity. CONCLUSION: Syngnathus has promising prospects as an anticancer Chinese medicine.

[Study on proteins of allele by using the microisoelectric focusing].

Clear protein electrophoresis pattern of single pollen grain from Cucurbita pepo L. was obtained with isoelectric focusing and supersensitive stain technique. The results showed that there were two special distributing protein bands near acidity pole. In species I, the two protein bands were a little near to acidity pole and in species II, that was a little far from acidity pole. In species III, the two protein bands distributed either like species I or like species II and the distributed ratio was 1:1. Moreover, we discuss how to identify relative purity of gene with the sorts of the special proteins.

Genetic transformation of Lycium barbarum L. via A. tumefaciens.

A system for transformation and regeneration of Lycium barbarum L., an important Chinese medical plant, has been established. Young stem segments from Lycium barbarum L. were infected with Agrobacterium tumefaciens C58cl(pGV3850::neo1103), and the transformed calli selected from the callus induction medium containing 50 micrograms/ml kanamycin could regenerate buds on differentiation medium containing 25 micrograms/ml kanamycin. 30% of the regenerated buds were normal in morphology. The normal buds could develop into whole plantlets after they were transferred to the rooting medium to induce roots. Nopaline detection, NPT-II enzyme activity assay and Southern blotting hybridization indicated that the foreign genes had been integrated into the genome of Lycium barbarum L. and expressed in the plant. In the processes of experiments, it was found that (i) after the pre-processes, the explants which formed callus quickly were easy to transform; (ii) the rate of normal regenerated plants from transgenic calli was higher than that from the untransgenic ones.

Hypervitaminosis D associated with drinking milk.

BACKGROUND: Vitamin D has been added to milk in the United States since the 1930s to prevent rickets. We report the unusual occurrence of eight cases of vitamin D intoxication that appear to have been caused by excessive vitamin D fortification of dairy milk. METHODS: Medical records were reviewed and a dietary questionnaire was sent to eight patients who had unexplained hypervitaminosis D. Vitamin D analyses with high-performance liquid chromatography were performed on samples of the patients' serum, the dairy milk they drank, and the vitamin D concentrate added to the milk. RESULTS: All eight patients drank milk produced by a local dairy in amounts ranging from 1/2 to 3 cups (118 to 710 ml) daily. All had elevated serum 25-hydroxyvitamin D concentrations (mean [+/- SD], 731 +/- 434 nmol per liter [293 +/- 174 ng per milliliter]). Six of the eight patients had elevated serum vitamin D3 concentrations. Of the eight patients, seven had hypercalcemia and one had hypercalciuria but normocalcemia (mean serum calcium, 3.14 +/- 0.51 mmol per liter [12.6 +/- 2.1 mg per deciliter]). Analysis of the dairy's vitamin D-fortified milk revealed concentrations of vitamin D3 (cholecalciferol) that ranged from undetectable to as high as 232,565 IU per quart (245,840 IU per liter). An analysis of the concentrate that was used to fortify the milk, labeled as containing vitamin D2 (ergocalciferol), revealed that it contained vitamin D3. CONCLUSIONS: Hypervitaminosis D may result from drinking milk that is incorrectly and excessively fortified with vitamin D. Milk that is fortified with vitamin D must be carefully monitored.

The vitamin D content of fortified milk and infant formula.

BACKGROUND: The fortification of milk and infant formula with vitamin D has had an important role in eliminating rickets in children and osteomalacia in adults. A recent outbreak of vitamin D intoxication caused by drinking milk fortified with excess vitamin D has led to questions about the level of vitamin D in milk from other producers. METHODS: We used high-performance liquid chromatography to measure vitamin D in samples of 13 brands of milk with various fat contents and 5 brands of infant formula purchased at random from local supermarkets in five Eastern states. RESULTS: Only 12 (29 percent) of the 42 samples of the 13 brands of milk and none of the 10 samples of the 5 brands of infant formula contained 80 to 120 percent of the amount of vitamin D stated on the label. Twenty-six of the 42 milk samples (62 percent) contained less than 80 percent of the amount claimed on the label. No vitamin D was detected in 3 of the 14 samples of skim milk tested (lower limit of assay, 4.7 IU per quart [5.0 IU per liter]). One milk sample labeled as containing vitamin D2 (ergocalciferol) contained vitamin D3 (cholecalciferol). Seven of the 10 samples of infant formula contained more than 200 percent of the amount stated on the label; the sample with the highest concentration contained 419 percent of the stated amount. None of the samples of infant formula contained less than the amount stated. CONCLUSIONS: Milk and infant-formula preparations rarely contain the amount of vitamin D stated on the label and may be either underfortified or overfortified. Since both underfortification and overfortification are hazardous, better monitoring of the fortification process is needed.

Regeneration of transgenic Lycium barbarum L.

A simple and effective system for the transformation and regeneration of Lycium barbarum L. has been developed. Young stem segments from Lycium barbarum L. were infected by Agrobacterium tumefaciens harboring a vector containing neomycin phosphotransferase II (npt-II) gene derived from non-oncogenic Ti plasmid. Calli originating from young stem segments on selective induction medium could differentiate into buds on selective differentiation medium rapidly and finally developed into whole plants. NPT-II enzyme activity assay and DNA hybridization indicated that the foreign gene had been integrated into the genome of Lycium barbarum L. and could be expressed in plants.

In vitro pollen plant induction and embryoid clone establishment of Panax ginseng.

In anther culture of Panax ginseng, its callus formation showed a wide adaptability to culture media. Large numbers of calli were induced on media exhibiting better effects of induction. Supplements of 5 mg 2,4-D/1 and 1 mg KT/1 to the media proved to be much effective. Regeneration of the whole plantlets from anther culture of Panax ginseng is usually quite difficult. During the past three years, however, sixteen of the 100 medium formulae tested were proved to be suitable. The formulae of MS + 0.5 mg BA/1 + 2 mg GA/1 + 1000 mg LH/1 + 3% sucrose were considered good and effective. A visibly differentiated body, which was light-milky white and later turned into a light green spot, was formed 40 days after the callus was transferred to the differentiation media. This body differentiated subsequently into buds, roots and, eventually, seedlings. The embryoid clones have been established in order to maintain its ability of continual differentiation into plantlets through successive culturings of many generations. The test-tube ginseng thus formed were transferred to regular flowerpots and grew well. Based upon chromosome examination of the callus cells and the root tips, we tentatively affirmed that the majority of these regenerated from anther of Panax ginseng were originated from pollen cells.