808 nm Near-Infrared Light-Triggered Payload Release from Green Light-Responsive Donor-Acceptor Stenhouse Adducts Polymer-Coated Upconversion Nanoparticles.

Owing to deep activation in biotissues and enhanced targeting efficiency, developing photoresponsive polymer-upconversion nanoparticles (PP-UCNPs) nanovectors has witnessed rapid growth in the past decade. However, up to date, all developed nanovectors require high-order photon processes to initiate the release of cargos. The photodamage caused by high-power near-infrared laser light may be a critical obstacle to their clinical application. Here, for the first time, by leveraging absorption-emission spectral matching between donor-acceptor Stenhouse adducts (DASA) PP and UCNPs (λex , 808 nm) in the green region (≈530 nm), the designed nanovector is capable of releasing cargos at a low-power 808 nm excitation (0.2 W). Considering the high molar absorptivity, biobenign, and synthetic tunability of DASA, DASA PP can be utilized as an up-and-coming candidate to design and synthesize the next generation of upconversion nanovectors without photodamage.

Low-Power Near-Infrared-Responsive Upconversion Nanovectors.

Activating upconversion nanoparticle-based photoresponsive nanovectors (UCPNVs) by upconversion visible light at low-power near-infrared (NIR) excitation can realize deeper biotissue stimulation with a minimized overheating effect and photodamage. Here, we demonstrate a facile strategy to construct new surface-decorated UCPNVs based on Passerini three-component reaction (P-3CR) in highly convenient and effective manners. Such UCPNVs materials have a tailored deprotecting wavelength that overlaps upconversion blue light. By using 3-perylenecarboxaldehyde, Tm3+/Yb3+ ion-doped UCNP-containing isocyanides, and antitumor agent chlorambucil as the three components, the resulting monodisperse UCPNV exhibits an efficient release of caged chlorambucil under a very low 976 nm power. This approach expands the synthetic toolbox to enable quick development of UCPNVs for UCNP-assisted low-power NIR photochemistry.

Serum CCL27 predicts the response to Bacillus Calmette-Guerin immunotherapy in non-muscle-invasive bladder cancer.

The prediction of the response to Bacillus Calmette-Guerin (BCG) can help identify non-muscle-invasive bladder cancer (NMIBC) patients that may be better served with alternative therapy. Several cytokine profiles present promising results, but they are difficult to use in clinical practice. In this prospective, longitudinal study, we tried to identify reliable serum cytokines/chemokines to predict the response to BCG using samples collected before and during BCG induction therapy. We used the Bio-plex multiplex assays to identify potential BCG failure-related serum cytokines/chemokines in the discovery set (n = 13). After screening, we identified CCL27 as the top candidate biomarker for predicting the response to BCG (P = .003). In the validation set, we found that the AUC of the baseline CCL27 was 0.730 (95% CI 0.515-0.945, P = .040) along with 67% sensitivity, 78% specificity. The changes from baseline to last timepoint can also distinguish BCG responders from non-responders (AUC: 0.726, 95% CI 0.474-0.979, P = .044). Moreover, the combination score of serum CCL27 (CSCCL27), based on the baseline and changes of CCL27, could further improve the predictive accuracy with an AUC of 0.897 (95% CI 0.790-1.000, P < .001). The correlations between CCL27 and local/systemic immunologic parameters were further analyzed. The level of serum CCL27 was strongly correlated with regulatory T cells (Tregs) in the tumor microenvironment (P = .002), indicating that CCL27 may promote the recruitment of Tregs into the tumor microenvironment. Our results show that serum CCL27 may represent a practical and reliable marker for the prediction of the response to BCG in NMIBC.

Sensitive electrogenerated chemiluminescence biosensing method for the determination of DNA hydroxymethylation based on Ru(bpy)32+-doped silica nanoparticles labeling and MoS2-poly(acrylic acid) nanosheets modified electrode.

5-Hydroxymethylcytosine (5-hmC), an oxidation product of 5-mC (5-methylcytosine), is presented in DNA of most mammalian cells and play an important role in the alteration of cancer-related genes. Herein, a sensitive electrogenerated chemiluminescence (ECL) biosensing method for the determination of 5-hmC in DNA (5-hmC DNA) was established on the basis of chemical modification and nanomaterial amplification. First, electrochemically reduced molybdenum disulfide-poly(acrylic acid) (rMoS2-PAA) nanosheets were used to modify glassy carbon electrode (GCE) to form an ECL biosensing electrode (rMoS2-PAA/GCE) which has large accessible surface area to immobilize more DNA. Then, a capture probe with amino group was hybridized with the target 5-hmC DNA and immobilized on the surface of rMoS2-PAA/GCE via amido bond. When cysteamine was introduced, the M.HhaI methyltransferase (M.HhaI) was used as specific recognition element to replace the hydroxyl group of 5-hmC by thiol and generated the amine-derivated DNA. Finally, surface chemically activated Ru(bpy)32+-doped silica (Ru@SiO2) nanoparticles, carriers of ECL reagents, were employed as signal amplification unit which covalently bonded to the amine-derivated DNA resulting in an increased ECL intensity. The increased ECL intensity was linearity to the 5-hmC DNA concentration in a range from 5.0 × 10-14 M to 1.0 × 10-11 M, with a lower detection limit of 1.2 × 10-14 M. Besides, the proposed method also displayed a good selectivity to 5-hmC in the presence of 5-C and 5-mC. Moreover, the developed biosensing method was successfully employed to monitor human urine sample.

AP4 modulated by the PI3K/AKT pathway promotes prostate cancer proliferation and metastasis of prostate cancer via upregulating L-plastin.

The transition from androgen-dependent to metastatic castration-resistant prostate cancer (PCa) is a lethal event of uncertain molecular aetiology. Our previous studies demonstrated that L-plastin is involved in PCa invasion and metastasis and is upregulated by androgen and oestrogen in the hormone-dependent PCa cell line LNCaP. We recently found that L-plastin expression is consistently activated even after androgen deprivation, suggesting that androgen-independent transcription factors may regulate its expression. Herein, we performed sequential deletion and luciferase analysis of the L-plastin promoter and found that an androgen-independent regulatory factor prominently located in the region close to the transcription initiation site (-216 to +118) may facilitate L-plastin upregulation. AP4 was then identified as the relevant transcription activator that directly binds to the L-plastin promoter, as confirmed by EMSAs, supershift assays and CHIP-qPCR experiments. Moreover, we determined that the AP4/L-plastin axis is regulated by the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, contributing to PCa metastasis and castration resistance. Furthermore, we found that AP4 promotes PCa metastasis by upregulating L-plastin expression in vitro and in vivo. We collected a total of 136 PCa tissues and corresponding adjacent normal tissues from patients who underwent prostatectomy at Sun Yat-Sen Memorial Hospital from 2005 to 2015 and measured AP4 and L-plastin protein levels by immunohistochemistry. The results showed that AP4 levels strongly correlated with those of its downstream target gene L-plastin, were significantly upregulated in PCa tissues, were positively correlated with lymph node metastasis and Gleason scores over 7, and were an independent prognostic factor for patient survival. In summary, these findings support a plausible mechanism by which the AP4/L-plastin axis is regulated by the PI3K/AKT pathway in human PCa and may represent a novel therapeutic target in PCa treatment.

Measurement of DNA Translocation Dynamics in a Solid-State Nanopore at 100 ns Temporal Resolution.

Despite the potential for nanopores to be a platform for high-bandwidth study of single-molecule systems, ionic current measurements through nanopores have been limited in their temporal resolution by noise arising from poorly optimized measurement electronics and large parasitic capacitances in the nanopore membranes. Here, we present a complementary metal-oxide-semiconductor (CMOS) nanopore (CNP) amplifier capable of low noise recordings at an unprecedented 10 MHz bandwidth. When integrated with state-of-the-art solid-state nanopores in silicon nitride membranes, we achieve an SNR of greater than 10 for ssDNA translocations at a measurement bandwidth of 5 MHz, which represents the fastest ion current recordings through nanopores reported to date. We observe transient features in ssDNA translocation events that are as short as 200 ns, which are hidden even at bandwidths as high as 1 MHz. These features offer further insights into the translocation kinetics of molecules entering and exiting the pore. This platform highlights the advantages of high-bandwidth translocation measurements made possible by integrating nanopores and custom-designed electronics.

Hybrid integrated biological-solid-state system powered with adenosine triphosphate.

There is enormous potential in combining the capabilities of the biological and the solid state to create hybrid engineered systems. While there have been recent efforts to harness power from naturally occurring potentials in living systems in plants and animals to power complementary metal-oxide-semiconductor integrated circuits, here we report the first successful effort to isolate the energetics of an electrogenic ion pump in an engineered in vitro environment to power such an artificial system. An integrated circuit is powered by adenosine triphosphate through the action of Na(+)/K(+) adenosine triphosphatases in an integrated in vitro lipid bilayer membrane. The ion pumps (active in the membrane at numbers exceeding 2 × 10(6) mm(-2)) are able to sustain a short-circuit current of 32.6 pA mm(-2) and an open-circuit voltage of 78 mV, providing for a maximum power transfer of 1.27 pW mm(-2) from a single bilayer. Two series-stacked bilayers provide a voltage sufficient to operate an integrated circuit with a conversion efficiency of chemical to electrical energy of 14.9%.

Polymyxin E Induces Rapid Paenibacillus polymyxa Death by Damaging Cell Membrane while Ca2+ Can Protect Cells from Damage.

Polymyxin E, produced by Paenibacillus polymyxa, is an important antibiotic normally against Gram-negative pathogens. In this study, we found that polymyxin E can kill its producer P. polymyxa, a Gram-positive bacterium, by disrupting its cell membrane. Membrane damage was clearly revealed by detecting the leakage of intracellular molecules. The observation using scanning electron microscopy also supported that polymyxin E can destroy the cell membrane and cause an extensive cell surface alteration. On the other hand, divalent cations can give protection against polymyxin E. Compared with Mg2+, Ca2+ can more effectively alleviate polymyxin E-induced damage to the cell membrane, thus remarkably increasing the P. polymyxa survival. Our findings would shed light on a not yet described bactericidal mechanism of polymyxin E against Gram-positive bacteria and more importantly the nature of limited fermentation output of polymyxin E from P. polymyxa.

Exploring regulation genes involved in the expression of L-amino acid oxidase in Pseudoalteromonas sp. Rf-1.

Bacterial L-amino acid oxidase (LAAO) is believed to play important biological and ecological roles in marine niches, thus attracting increasing attention to understand the regulation mechanisms underlying its production. In this study, we investigated genes involved in LAAO production in marine bacterium Pseudoalteromonas sp. Rf-1 using transposon mutagenesis. Of more than 4,000 mutants screened, 15 mutants showed significant changes in LAAO activity. Desired transposon insertion was confirmed in 12 mutants, in which disrupted genes and corresponding functionswere identified. Analysis of LAAO activity and lao gene expression revealed that GntR family transcriptional regulator, methylase, non-ribosomal peptide synthetase, TonB-dependent heme-receptor family, Na+/H+ antiporter and related arsenite permease, N-acetyltransferase GCN5, Ketol-acid reductoisomerase and SAM-dependent methytransferase, and their coding genes may be involved in either upregulation or downregulation pathway at transcriptional, posttranscriptional, translational and/or posttranslational level. The nhaD and sdmT genes were separately complemented into the corresponding mutants with abolished LAAO-activity. The complementation of either gene can restore LAAO activity and lao gene expression, demonstrating their regulatory role in LAAO biosynthesis. This study provides, for the first time, insights into the molecular mechanisms regulating LAAO production in Pseudoalteromonas sp. Rf-1, which is important to better understand biological and ecological roles of LAAO.

Antibacterial mechanisms of polymyxin and bacterial resistance.

Multidrug resistance in pathogens is an increasingly significant threat for human health. Indeed, some strains are resistant to almost all currently available antibiotics, leaving very limited choices for antimicrobial clinical therapy. In many such cases, polymyxins are the last option available, although their use increases the risk of developing resistant strains. This review mainly aims to discuss advances in unraveling the mechanisms of antibacterial activity of polymyxins and bacterial tolerance together with the description of polymyxin structure, synthesis, and structural modification. These are expected to help researchers not only develop a series of new polymyxin derivatives necessary for future medical care, but also optimize the clinical use of polymyxins with minimal resistance development.

Improving signal-to-noise performance for DNA translocation in solid-state nanopores at MHz bandwidths.

DNA sequencing using solid-state nanopores is, in part, impeded by the relatively high noise and low bandwidth of the current state-of-the-art translocation measurements. In this Letter, we measure the ion current noise through sub 10 nm thick Si3N4 nanopores at bandwidths up to 1 MHz. At these bandwidths, the input-referred current noise is dominated by the amplifier's voltage noise acting across the total capacitance at the amplifier input. By reducing the nanopore chip capacitance to the 1-5 pF range by adding thick insulating layers to the chip surface, we are able to transition to a regime in which input-referred current noise (∼ 117-150 pArms at 1 MHz in 1 M KCl solution) is dominated by the effects of the input capacitance of the amplifier itself. The signal-to-noise ratios (SNRs) reported here range from 15 to 20 at 1 MHz for dsDNA translocations through nanopores with diameters from 4 to 8 nm with applied voltages from 200 to 800 mV. Further advances in bandwidth and SNR will require new amplifier designs that reduce both input capacitance and input-referred amplifier noise.

Advances in detection methods of L-amino acid oxidase activity.

L-amino acid oxidase (LAAO) is widely distributed in many different organisms and found to play important biological roles, thus attracting a great deal of attention for characterization of its activity. Diverse detection methods with their own properties have been established. This review advanced different LAAO activity assays based on substrate consumption, cofactor amount, and product accumulation. The description of benefits and drawbacks of each method is expected to help researchers find appropriate detection method of LAAO activity for their own purpose.

Effects of magnolol on impairment of learning and memory abilities induced by scopolamine in mice.

Alzheimer's disease (AD), one of the most common forms of dementia, is primarily ascribed to the cholinergic deficits and neuronal dysfunction. Magnolol (Mag), a bioactivator extracted from Magnolia officinalis, has protective effects on cholinergic neurons, but the specific mechanism remains unknown. To further evaluate the therapeutic effects of Mag on the learning and memory impairment in a scopolamine (Scop)-induced mouse model, the passive avoidance and the Morris water maze tests, the measurement of the ratio of brain/hippocampus to body weight, activities of acetyl cholinesterase (AChE), superoxide dismutase (SOD), total nitric oxide synthase (total NOS) and the content of methane dicarboxylic aldehyde (MDA) in hippocampus homogenate as well as the immunefluorescence staining of the AChE positive nerve fibers were performed. Therapeutically treated with Mag, the impaired abilities of learning and memory of the Scop-induced mice were almost restored to the native levels. The restored AChE, total NOS and SOD activities and the MDA level were observed, with a relatively normal density of AChE positive nerve fibers in hippocampus CA3 molecular layer. The improving efficacy of Mag on learning and memory impairment induced by Scop is dose-dependent, indicating that Mag has potential neuroprotective effects against neuronal impairment and memory dysfunction induced by Scop in mice. The underlying mechanisms may be associated with the anti-oxidative effects of Mag and its protective effects on hippocampus cholinergic neurons.

Nanopore detachment kinetics of poly(A) binding proteins from RNA molecules reveals the critical role of C-terminus interactions.

The ubiquitous and abundant cytoplasmic poly(A) binding protein (PABP) is a highly conserved multifunctional protein, many copies of which bind to the poly(A) tail of eukaryotic mRNAs to promote translation initiation. The N-terminus of PABP is responsible for the high binding specificity and affinity to poly(A), whereas the C-terminus is known to stimulate PABP multimerization on poly(A). Here, we use single-molecule nanopore force spectroscopy to directly measure interactions between poly(A) and PABPs. Both electrical and biochemical results show that the C-C domain interaction between two consecutive PABPs promotes cooperative binding. Up to now, investigators have not been able to probe the detailed polarity configuration (i.e., the internal arrangement of two PABPs on a poly(A) streak in which the C-termini face toward or away from each other). Our nanopore force spectroscopy system is able to distinguish the cooperative binding conformation from the noncooperative one. The ∼50% cooperative binding conformation of wild-type PABPs indicates that the C-C domain interaction doubles the cooperative binding probability. Moreover, the longer dissociation time of a cooperatively bound poly(A)/PABP complex as compared with a noncooperatively bound one indicates that the cooperative mode is the most stable conformation for PABPs binding onto the poly(A). However, ∼50% of the poly(A)/PABP complexes exhibit a noncooperative binding conformation, which is in line with previous studies showing that the PABP C-terminal domain also interacts with additional protein cofactors.

Helix-coil kinetics of individual polyadenylic acid molecules in a protein channel.

Helix-coil transition kinetics of polyadenylic acid [poly(A)] inside a small protein channel is investigated for the first time, at the single molecule level. The confinement of a RNA molecule inside the channel slows its kinetics by nearly 3 orders of magnitude as compared to bulk measurements of free poly(A). These findings are related to the interaction energy of the RNA structure with the interior of the pore, explained by a simple two-state model. These results shed light on the way intermolecular interactions alter nucleic acid kinetics.