Intelligent Hybrid Hydrogels for Rapid In Situ Detection and Photothermal Therapy of Bacterial Infection.

Diseases induced by bacterial infections increasingly threaten the health of people all over the world; thus, it is urgent and significant to early diagnose and effectively eliminate infections to save people's lives. To this end, we synthesized an intelligent hydrogel that integrated in situ visualized diagnosis and photothermal therapy of bacterial infections. By simply and subtly incorporating pH-sensitive bromothymol blue (BTB) and near-infrared (NIR)-absorbing conjugated polymer (termed as PTDBD) into thermosensitive chitosan (CS)-based hydrogel, the synthesized BTB/PTDBD/CS hydrogel can diagnose the acidic microenvironment of Staphylococcus aureus (S. aureus) biofilm and infected wounds by showing visualized color change. After rapid diagnosis, the hydrogel can immediately treat the infection site by local hyperthermia under irradiation of NIR laser (808 nm) and even the stubborn biofilm that is difficult to eradicate. Since the dominating antibacterial mechanism is hyperthermia, the hybrid hydrogel shows broad-spectrum antibacterial activity against Gram-positive, Gram-negative, and drug-resistant bacteria. In addition, it has low cytotoxicity to normal cells and no effect on the main organs of mice. It paves a brand new avenue to develop smart and facile diagnosis and a treatment platform for bacterial infections.

Organic Polymer Nanoparticles with Primary Ammonium Salt as Potent Antibacterial Nanomaterials.

Bacterial infections induced by drug-resistant strains have become a global crisis. A membrane-disrupted mechanism is considered as an effective way to kill bacteria with little chance to trigger drug resistance. It is necessary to explore and develop new materials based on the membrane-disrupted mechanism to combat bacterial resistance. Here we report the design of organic nanoparticles based on a polymer (PDCP) as highly effective inhibition and bactericidal reagents. The PDCP is devised to have a hydrophobic skeleton and hydrophilic side chain modified with protonated primary amines, which could self-assemble to form organic nanoparticles (PDCP-NPs). By taking advantage of the large surface to volume ratio of nanoparticles, the synthesized PDCP-NPs have enriched positive charges and multiple membrane-binding sites. Research results display that PDCP-NPs have highly potent antibacterial activity in vitro and vivo, especially for Gram-negative bacteria with low toxicity against mammalian cells. This work design will inspire researchers to develop more membrane-disrupted bactericide and advance the applications of organic nanoparticles in the antibacterial area.

Flower extracts from Paeonia decomposita and Paeonia ostii inhibit melanin synthesis via cAMP-CREB-associated melanogenesis signaling pathways in murine B16 melanoma cells.

This investigation seeks to identify the effects of the EtOAc fractions of different flower parts of Paeonia decomposita (Pd) and Paeonia ostii (Po) on melanogenesis and their mechanisms of action in B16 melanoma cells. Cell viability assay showed that Pd-1, Po-1 (the petals of Pd or Po), Pd-3 and Po-3 (the stamens of Pd or Po) at 25 µg/ml produced lower toxic activities in B16 cells. Pd-1 and Po-1 extracts considerably reduced the melanin content and inhibited tyrosinase and DOPA oxidase activity. Moreover, Pd-1 and Po-1 down-regulated the expressions of MC1R, MITF, TRP-1, TRP-2, and tyrosinase. These extracts also reduce cAMP levels and inhibited the phosphorylation of CREB, which might be due to the presence of high concentrations of phenolic compounds and flavonoids. Our results suggested that Pd-1 and Po-1 are able to modulate the cAMP-CREB signaling pathway and down-regulate the melanogenesis-related proteins resulting in the observed anti-melanogenic effects. PRACTICAL APPLICATIONS: In China, the flower of Paeonia is often consumed as a dietary supplement and as an additive in skin whitening products. In November 2013, the National Health and Family Planning Commission of the People's Republic of China has approved the flower of Paeonia ostii as a novel food resource. The current study firstly demonstrated that the effects of EtOAc fractions of the petals of Paeonia decomposita (Pd) and Paeonia ostii (Po) considerably reduced the melanin content in B16 cells, which is due to the modulation of the cAMP-CREB signaling pathway followed by the down-regulation of melanogenesis-related proteins. Pd and Po extracts, as natural tyrosinase inhibitors, may serve as good candidates in food additives, cosmetic materials, or even in treating hyperpigmentation diseases.

Bactericidal activity-tunable conjugated polymers as a human-friendly bactericide for the treatment of wound infections.

Photodynamic therapy (PDT) has been reported to be an effective alternative to combat bacterial infections even those triggered by drug-resistant strains as there is little chance to develop resistance to this therapy. Therefore, it is imperative to design and synthesize a superior photo-active bactericide for the treatment of bacterial infections. Herein, we synthesized three bactericidal activity-tunable conjugated polymers (P1-P3) with various photoactive capabilities and employed them for the treatment of wound infections with little damage to cells; by altering the construction unit of π-conjugated backbone structures with electron-rich and electron-deficient aromatic heterocycles, the optical properties and ability of reactive oxygen species (ROS) generation could be regulated; this resulted in a tunable killing ability. The cationic quaternary ammonium (QA) groups on the side chains endowed the CPs with not only good dispersibility but also a better interaction with the negatively charged membrane of bacteria. The antibacterial experiments towards ampicillin-resistant Escherichia coli TOP10 (E. coli) and the treatment of wound infections in mice indicate that the P1-P3 have varied bactericidal activities; moreover, P3 has been demonstrated to be a human-friendly bactericide with excellent antibacterial capability. It not only acts as a potential bactericide for the practical treatment of infectious wounds, but also offers guidance for the design and structure control of photo-active bactericides.

Geniposide Suppresses Hepatic Glucose Production via AMPK in HepG2 Cells.

Geniposide is one of the main compounds in Gardenia jasminoides ELLIS and has many pharmacological activities, but its anti-hyperglycemic activity has not yet been fully explored. This study was designed to determine, for the first time, how geniposide from G. jasminoides regulates hepatic glucose production, and the underlying mechanisms. During in vitro study, we found the inhibitory effect of geniposide on the hepatic glucose production is partly through AMP-activated protein kinase (AMPK) activation in HepG2 cells. Geniposide significantly inhibited hepatic glucose production in a dose-dependent manner. AMPK, acetyl coenzyme A synthetase (ACC) and forkhead box class O1 (FoxO1) phosphorylation were stimulated by different concentrations of geniposide. In addition, the enzyme activities of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) were all significantly suppressed. What is important is that these effects were partly reversed by (1) inhibition of AMPK activity by compound C, a selective AMPK inhibitor, and by (2) suppression of AMPKα expression by small interfering RNA (siRNA). In summary, geniposide potentially ameliorates hyperglycemia through inhibition of hepatic gluconeogenesis by modulation of the AMPK-FoxO1 signaling pathway. Geniposide or geniposide-containing medicinal plants could represent a promising therapeutic agent to prevent type 2 diabetes on gluconeogenesis.

Preclinical pharmacokinetics and disposition of a novel selective VEGFR inhibitor fruquintinib (HMPL-013) and the prediction of its human pharmacokinetics.

PURPOSE: This study evaluated the preclinical pharmacokinetics (PK) and disposition of fruquintinib (HMPL-013), a small molecule vascular endothelial growth factor receptors inhibitor. METHODS: In vitro and in vivo PK/ADME assays were conducted. Allometry and PK modeling/simulation were conducted to predict human PK parameters and the time course profiles. RESULTS: HMPL-013 has high permeability without efflux. It shows moderate oral bioavailability of 42-53 % and Tmax < 4 h in mouse, rat, dog and monkey, with exposure-dose linearity proved in rats and dogs. No significant food effect is on dog PK. HMPL-013 has moderately high tissue distribution. It majorly distributes in gastrointestinal tract, liver, kidney, adrenal and adipose. The plasma protein binding fraction is 88-95 % in mouse, rat, dog and human, invariable up to 10 µM. The in vivo clearance of HMPL-013 is low, consistent with the in vitro scaling. Three major oxidative metabolites were identified in liver microsomes of mouse, rat, dog, monkey and human. Dog is mostly similar to human regarding in vitro metabolism. Demethylation, hydroxylation and sequential glucuronidation are the major in vivo metabolic reactions. Direct urinary and biliary excretion of HMPL-013 is negligible. Metabolizing to M1 (demethylation), sequentially glucuronidating, followed by biliary excretion, and to a less extent, by urinary excretion, are important elimination pathways for HMPL-013 in rats. HMPL-013 has low risk of drug-drug interaction. It is predicted to have favorable human PK properties and low efficacious dose. CONCLUSION: HMPL-013 demonstrates good preclinical PK and enables successful human PK and dose projection. It is valuable for further clinical development.

Glucagon-like peptide 1 receptor plays a critical role in geniposide-regulated insulin secretion in INS-1 cells.

AIM: To explore the role of the glucagon-like peptide 1 receptor (GLP-1R) in geniposide regulated insulin secretion in rat INS-1 insulinoma cells. METHODS: Rat INS-1 insulinoma cells were cultured. The content of insulin in the culture medium was measured with ELISA assay. GLP-1R gene in INS-1 cells was knocked down with shRNA interference. The level of GLP-1R protein in INS-1 cells was measured with Western blotting. RESULTS: Geniposide (0.01-100 µmol/L) increased insulin secretion from INS-1 cells in a concentration-dependent manner. Geniposide (10 µmol/L) enhanced acute insulin secretion in response to both the low (5.5 mmol/L) and moderately high levels (11 mmol/L) of glucose. Blockade of GLP-1R with the GLP-1R antagonist exendin (9-39) (200 nmol/L) or knock-down of GLP-1R with shRNA interference in INS-1 cells decreased the effect of geniposide (10 µmol/L) on insulin secretion stimulated by glucose (5.5 mmol/L). CONCLUSION: Geniposide increases insulin secretion through glucagon-like peptide 1 receptors in rat INS-1 insulinoma cells.

[Activating effect of citrus flavonoids on neuromedin U2 receptor and analysis on siRNA interference].

OBJECTIVE: To screen out active substances on Neuromedin U2 receptor (NMU2R) by using stable NMU2R cell lines and negative cell lines and analyzing siRNA interference. METHOD: NMU2R cells were used to observe the activating effect of nine nine citrus flavonoids on NMU2R cell. Afterwards, false-positive interference of citrus flavonoids that showed higher activating effect was eliminated by using negative cells and analyzing the efficiency of siRNA interference. RESULT: Hesperidin and nobiletin contained in citrus flavonoids were found to effectively activate NMU2R. The efficacy, EC50 and potency values of hesperidin were 4.688, 318.970 micromol x L(-1) and 200.933 micromol x L(-1), while the efficacy, EC50 and potency values of nobiletin were 4.758, 5.832 micromol x L(-1) and 3.124 micromol x L(-). CONCLUSION: Hesperidin and nobiletin contained in citrus flavonoids can activate NMU2R. Nobiletin shows such a low EC50 that it has medicinal value.

Geniposide induces the expression of heme oxygenase-1 via PI3K/Nrf2-signaling to enhance the antioxidant capacity in primary hippocampal neurons.

Oxidative stress in brain is emerging as a potential causal factor in aging and age-related neurodegenerative disorders. A large body of evidence shows that induction of endogenous antioxidative proteins seems to be a reasonable strategy for delaying the progression of cell injury. In this study, geniposide upregulates the expression of heme oxygenase-1 (HO-1) to attenuate the cell apoptosis induced by 3-morpholinosydnonimine hydrochloride (SIN-1) in primary cultured hippocampal neurons. Furthermore, geniposide induces the nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and activation of phosphatidylinositol 3'-kinase (PI3K) in the presence of oxidative stress, and both LY294002 (a specific inhibitor of PI3K) and Zinc protoporphyrin (ZnPP, an inhibitor of HO-1) decrease the cytoprotective action of geniposide in hippocampal neurons. Taken together, the novel cytoprotective mechanism of geniposide to antagonize oxidative stress may be involved in PI3K- and Nrf2-mediated upregulation of the antioxidative enzyme HO-1.

Reactive astrocytes protect melanoma cells from chemotherapy by sequestering intracellular calcium through gap junction communication channels.

Brain metastases are highly resistant to chemotherapy. Metastatic tumor cells are known to exploit the host microenvironment for their growth and survival. We report here that melanoma brain metastases are surrounded and infiltrated by activated astrocytes, and we hypothesized that these astrocytes can play a role similar to their established ability to protect neurons from apoptosis. In coculture experiments, astrocytes, but not fibroblasts, reduced apoptosis in human melanoma cells treated with various chemotherapeutic drugs. This chemoprotective effect was dependent on physical contact and gap junctional communication between astrocytes and tumor cells. Moreover, the protective effect of astrocytes resulted from their sequestering calcium from the cytoplasm of tumor cells. These data suggest that brain tumors can, in principle, harness the neuroprotective effects of reactive astrocytes for their own survival and implicate a heretofore unrecognized mechanism for resistance in brain metastasis that might be of relevance in the clinic.

[High throughput screening method of identifying D5 receptor agonist from Chinese herbs].

D5 receptor is a subtype of dopamine D1-like receptor, and it plays a functional role in many neurological disorders. Some natural compounds from Chinese herbs, which were shown to have the property as that of receptor agonist, might provide a rich source in search of new candidates for therapeutic use. For exploring this possibility, we developed a cell-based high throughput (HTS) D5 receptor assay to screen the herb-based natural compound library established in our centre. The D5 receptor plasmid (hD5R/pcDNA3.1) and reporter gene plasmid (4 x CRE/TK/Luci/pGL3) were co-transfected into HEK293 cell line. After G418 being selected, the monoclonal cell lines bearing hD5R and the reporter gene were established and used for agonist screening. To optimize the assay condition, the effects of some factors such as cell number per well, incubation time, and the doses of SKF38393 (a potent selective partial D1-like agonist) were examined by using forskolin, a positive compound for cAMP response element. The best condition for this HTS assay included: the cell number at 5 x 10(4)/mL, the dose of forskolin at 5-20 micromol/L, the dose of SKF38393 at 100 nmol/L-100 micromol/L, and the agonist incubation time at 6 -8 h. Thereafter, water extracts from more than 200 Chinese herbs in our library were screened and three of these water extracts showed positive activity, with higher or similar activity as SKF38393. In conclusion, we have established a cell-based HTS assay for D5 receptor agonist screening, and by use of this HTS assay, 3 Chinese herbs maybe contain components exhibiting D5 receptor agonist property. This work provides an alternative vision of how to use herb medicines and a way to develop potential drugs for treatment of neurological disorders.

[Effect of total glucosides of paeony on nuclear factor-kappaB activation in rat peritoneal macrophages].

OBJECTIVE: To study the effect of total glucosides of paeony (TGP) on lipopolysaccharides (LPS)-induced nuclear factor-kappaB (NF-kappaB) activation in macrophages. METHOD: Rat peritoneal macrophages were pre-treated with TGP for 2 h and stimulated with LPS for 20 min or 0.5 h. Inhibitory kappaBalpha (IkappaBalpha) protein in the cytoplasm and NF-kappaB p65 protein in the nuclear were analyzed by western blot. Further, DNA binding activity of NF-kappaB complex was detected. RESULT: TGP enhanced the amounts of IkappaBalpha protein in the cytoplasm and decreased the amounts of NF-kappaB p65 protein in the nuclear of LPS-induced macrophages. TGP also inhibited the LPS-mediated DNA binding activity of NF-kappaB complex in macrophages. CONCLUSION: TGP can inhibit LPS-induced NF-kappaB activation in macrophages through arresting IKBalpha protein degradation, NF-kappaB p65 protein nuclear translocation and DNA binding activity of NF-kappaB complex.