NLRP3 augmented resistance to gemcitabine in triple-negative breast cancer cells via EMT/IL-1ß/Wnt/ß-catenin signaling pathway.

BACKGROUND: Gemcitabine is widely used in the treatment of breast cancer (BC). However, the resistance to drugs remains a tough concern. The study explored the potential mechanism concerning gemcitabine resistance in triple-negative BC (TNBC) in vitro. METHODS: TNBC cells (TNBCC) and gemcitabine-resistance cell lines (GRC) were used. We investigated the sensitivity to gemcitabine responsive to regulation of Nod-like receptor protein 3 (NLRP3) expression in TNBCC in different gemcitabine concentrations. RT-PCR checked NLRP3 mRNA expression and MTT assessed the cell cytotoxicity. Gemcitabine resistance was studied in GRC exposed to 0, 1, 3, 5 nm gemcitabine after GRC were treated with NLRP3 agonist Nigericin sodium salt (NSS) or antagonist CY-09. Epithelial-to-mesenchymal transition (EMT) biomarkers were evaluated via RT-PCR and inflammasome IL-1ß, ß-catenin content and GSK-3ß activity were measured by ELISA methods. Last, we inactivated the signaling and examined the NLRP3, EMT mRNA expression by RT-PCR, IL-1ß, ß-catenin content and GSK-3ß activity by ELISA and cell cytotoxicity through MTT. RESULTS: NLRP3 up-regulation improved cell survival and reduced sensitivity to gemcitabine (P<0.05). NLRP3 had higher expression in GRC than TNBCC. GRC cell viability dropped as the gemcitabine concentration increased. NLRP3 up-regulation added to resistance to gemcitabine in GRC (P<0.05). NLRP3 agonist might induce EMT process, activate wnt/ß-catenin signaling and IL-1ß, while inactivation of wnt/ß-catenin signaling could result in the inhibition of NLRP3, IL-1ß and EMT as well as cell viability in GRC (P<0.05). CONCLUSION: NLRP3 could enhance resistance to gemcitabine via IL-1ß/EMT/Wnt/ß-catenin signaling, which suggested that NLRP3 antagonist CY-09 might be incorporated into gemcitabine treatment for TNBC.

PageRank as a collective variable to study complex chemical transformations and their energy landscapes.

A reduced set of reaction coordinates is often employed in chemistry to describe the collective change between reactants and products within the context of rare event theories and the exploration of energy landscapes. Yet selecting the proper collective variable becomes increasingly challenging as the systems under study become more complex. Recent advancement of new descriptions of collective molecular coordinates has included graph-theoretical metrics, including social permutation invariant and PageRank (PR) coordinates, based upon the network of interactions about molecules and atoms within a system. Herein we continue the development of PR by (1) presenting a new formulation that is continuous along a reaction path, (2) illustrating that the fluctuations in PR are demonstrative of the fundamental motions of the atoms/molecules, and (3) providing the analytical derivatives with respect to atomic coordinates. The latter is subsequently combined with a harmonic bias to create the potential of mean force (PMF). As an example, we first consider the transformation of tetrahedral [Al(OH)4](aq) - to octahedral [Al(OH)4(H2O)2](aq) - using the PR PMF. Second, we explore the interchange of contact ion pair and solvent separated ion pairs of aqueous Na⋯OH, where the distance-biased PMF is projected onto PR space. In turn, this reveals where solvent rearrangement has the most impact upon the reaction pathway.

Structural and Dynamic Heterogeneity of Capillary Wave Fronts at Aqueous Interfaces.

Using a unique combination of slab-layering analyses and identification of truly interfacial molecules, this work examines water/vapor and water/n-hexane interfaces, specifically the structural and dynamic perturbations of the interfacial water molecules at different locations within the surface capillary waves. From both the structural and dynamic properties analyzed, it is found that these interfacial water molecules dominate the perturbations within the interfacial region, which can extend deep into the water phase relative to the Gibbs dividing surface. Of more importance is the demonstration of structural and dynamic heterogeneity of the interfacial water molecules at the capillary wave front, as indicated by the dipole orientation and the structural and dynamic behavior of hydrogen bonds and their networks.

Towards a unified description of the hydrogen bond network of liquid water: a dynamics based approach.

The definition of a hydrogen bond (H-bond) is intimately related to the topological and dynamic properties of the hydrogen bond network within liquid water. The development of a universal H-bond definition for water is an active area of research as it would remove many ambiguities in the network properties that derive from the fixed definition employed to assign whether a water dimer is hydrogen bonded. This work investigates the impact that an electronic-structure based definition, an energetic, and a geometric definition of the H-bond has upon both topological and dynamic network behavior of simulated water. In each definition, the use of a cutoff (either geometric or energetic) to assign the presence of a H-bond leads to the formation of transiently bonded or broken dimers, which have been quantified within the simulation data. The relative concentration of transient species, and their duration, results in two of the three definitions sharing similarities in either topological or dynamic features (H-bond distribution, H-bond lifetime, etc.), however no two definitions exhibit similar behavior for both classes of network properties. In fact, two networks with similar local network topology (as indicated by similar average H-bonds) can have dramatically different global network topology (as indicated by the defect state distributions) and altered H-bond lifetimes. A dynamics based correction scheme is then used to remove artificially transient H-bonds and to repair artificially broken bonds within the network such that the corrected network exhibits the same structural and dynamic properties for two H-bond definitions (the properties of the third definition being significantly improved). The algorithm described represents a significant step forward in the development of a unified hydrogen bond network whose properties are independent of the original hydrogen bond definition that is employed.

Ionic liquid foam floatation coupled with ionic liquid dispersive liquid-liquid microextraction for the separation and determination of estrogens in water samples by high-performance liquid chromatography with fluorescence detection.

An ionic liquid foam floatation coupled with ionic liquid dispersive liquid-liquid microextraction method was proposed for the extraction and concentration of 17-α-estradiol, 17-ß-estradiol-benzoate, and quinestrol in environmental water samples by high-performance liquid chromatography with fluorescence detection. 1-Hexyl-3-methylimidazolium tetrafluoroborate was applied as foaming agent in the foam flotation process and dispersive solvent in microextraction. The introduction of the ion-pairing and salting-out agent NH4 PF6 was beneficial to the improvement of recoveries for the hydrophobic ionic liquid phase and analytes. Parameters of the proposed method including concentration of 1-hexyl-3-methylimidazolium tetrafluoroborate, flow rate of carrier gas, floatation time, types and concentration of ionic liquids, salt concentration in samples, extraction time, and centrifugation time were evaluated. The recoveries were between 98 and 105% with relative standard deviations lower than 7% for lake water and well water samples. The isolation of the target compounds from the water was found to be efficient, and the enrichment factors ranged from 4445 to 4632. This developing method is free of volatile organic solvents compared with regular extraction. Based on the unique properties of ionic liquids, the application of foam floatation, and dispersive liquid-liquid microextraction was widened.

Inhibition of prostate cancer by suicide gene targeting the FCY1 and HSV-TK genes.

Prostate cancer is one of the most prevalent tumors. The switch of androgen signal dependence makes therapy more complex. Although reports on introduction of a single suicide gene exist, double suicide gene therapy has not been reported yet. In the current study, two suicide genes were constructed in the pIRES plasmid driven by PSMA promoter. 5-FC and GCV combination in vitro led to a higher growth inhibition on prostate cancer compared to a single pro-drug. Retarded xenograft tumor growth was observed in castrated nude mice after double suicide gene activation. Furthermore, decreased metastasis was observed with double suicide gene treatment. These findings suggest that specific double suicide gene strategy could be a potential option for the therapy of prostate cancer.

Microfluidic Reactor Array Device for Massively Parallel In-situ Synthesis of Oligonucleotides.

We have designed and fabricated a microfluidic reactor array device for massively parallel in-situ synthesis of oligonucleotides (oDNA). The device is made of glass anodically bonded to silicon consisting of three level features: microreactors, microchannels and through inlet/outlet holes. Main challenges in the design of this device include preventing diffusion of photogenerated reagents upon activation and achieving uniform reagent flow through thousands of parallel reactors. The device embodies a simple and effective dynamic isolation mechanism which prevents the intermixing of active reagents between discrete microreactors. Depending on the design parameters, it is possible to achieve uniform flow and synthesis reaction in all of the reactors by proper design of the microreactors and the microchannels. We demonstrated the use of this device on a solution-based, light-directed parallel in-situ oDNA synthesis. We were able to synthesize long oDNA, up to 120 mers at stepwise yield of 98 %. The quality of our microfluidic oDNA microarray including sensitivity, signal noise, specificity, spot variation and accuracy was characterized. Our microfluidic reactor array devices show a great potential for genomics and proteomics researches.

[Sodium nitroprusside facilitates human sperm capacitation and acrosome reaction].

OBJECTIVE: To explore the effect of nitric oxide (NO) on human sperm capacitation and acrosome reaction (AR). METHODS: Different concentrations of sodium nitroprusside (SNP) were added to the sperm suspension from 48 healthy fertile men, and the suspension was incubated in 1 x Earle at 37 degrees C for 1 hour. Progesterone was used to induce AR for 15, 30, 45 and 60 min, and then acid phosphatase (ACP) activity in the suspension before and after capacitation and at different time of AR was measured by p-nitrophenyl sodium phosphate assay. In the meantime, sperm motile parameters were assayed by CASA to observe sperm capacitation and AR. RESULTS: ACP activity and sperm motile parameters increased in the 50 approximately 100 nmol/L NO concentration group, showed no significant variation in the 150 approximately 200 nmol/L group, and decreased in the 250 approximately 300 nmol/L group. CONCLUSION: NO can facilitate sperm capacitation, AR and sperm motile parameters in low concentration and suppress them in high concentration. ACP activity assay of sperm is an objective and reliable method to evaluate sperm capacitation and AR in whole sperm population.

Individually addressable parallel peptide synthesis on microchips.

Miniaturized, spatially addressable microchips of peptides and peptidomimetics are powerful tools for high-throughput biomedical and pharmaceutical research and the advancement of proteomics. Here we report an efficient and flexible method for the parallel synthesis of peptides on individually addressable microchips, using digital photolithography and photogenerated acid in the deprotection step. We demonstrate that we are able to synthesize thousands of peptides in a 1 cm(2) area on a microchip using 20 natural amino acids as well as synthetic amino acid analogs, with high stepwise yields and short reaction-cycle times. Epitope screening experiments using a p53 antibody (PAb240) produced clearly defined binding patterns. The peptidomimetic sequences on the microchip show specific antibody binding and provide insights into the molecular details responsible for specificity of epitope binding. Our approach requires just a conventional synthesizer and a computer-controllable optical module, thereby allowing potential development of peptide microchips for various pharmaceutical and proteomic applications in routine research laboratories.