In situ monitoring of toxic effects of algal toxin on cells by a novel microfluidic flow cytometry instrument.

Algal toxins produced by microalgae, such as domoic acid (DA)1, have toxic effects on humans. However, toxicity tests using mice only yield lethal doses of algal toxins without providing insights into the mechanism of action on cells. In this study, a fast segmentation of microfluidic flow cytometry cell images based on the bidirectional background subtraction (BBS)2 method was developed to get the visual evidence of apoptosis in both bright-field and fluorescence images. This approach enables mapping of changes in cell morphology and activity under algal toxins, allowing for fast (within 60 s) and automated biological detection. By combining microfluidics with flow cytometry, the intricate cellular-level reaction process can be observed in micro samples of 293 T cells and mouse spleen cells, offering potential for future in vitro experiments.

Controlling Supramolecular Fiber Formation of Nucleopeptide by Guanosine Triphosphate.

Cells use dynamic self-assembly to construct functional structures for maintaining cellular homeostasis. However, using a natural biological small molecule to mimic this phenomenon remains challenging. This work reports the dynamic microfiber formation of nucleopeptide driven by guanosine triphosphate, the small molecule that controls microtubule polymerization in living cells. Deactivation of GTP by enzyme dissociates the fibers, which could be reactivated by adding GTP. Molecular dynamic simulation unveils the mystery of microfiber formation of GBM-1 and GTP. Moreover, the microfiber formation can also be controlled by diffusion-driven GTP gradients across a semipermeable membrane in bulk conditions and the microfluidic method in the defined droplets. This study provides a new platform to construct dynamic self-assembly materials of molecular building blocks driven by GTP.

Controlling Intracellular Enzymatic Self-Assembly of Peptide by Host-Guest Complexation for Programming Cancer Cell Death.

Controlling the enzymatic reaction of macromolecules in living systems plays an essential role in determining the biological functions, which remains challenging in the synthetic system. This work shows that host-guest complexation could be an efficient strategy to tune the enzymatic self-assembly of the peptide. The formed host-guest complexation prevents the enzymatic kinetics of peptide assemblies on the cell surface and promotes cellular uptake of assemblies. For uptake inside cells, the host-guest complex undergoes dissociation in the acidic lysosome, and the released peptide further self-assembles inside the mitochondria. Accumulating assemblies at mitochondria induce the ferroptosis of cancer cells, resulting in cancer cell death in vitro and the tumor-bearing mice model. As the first example of using host-guest complexation to modulate the kinetics of enzymatic self-assembly, this work provides a general method to control enzymatic self-assembly in living cells for selective programming cancer cell death.

Design and Application of Thymol Electrochemical Sensor Based on the PtNPs-CPOFs-MWCNTs Composite.

In this study, the preparation of covalent polyoxometalate organic frameworks (CPOFs) is introduced using the idea of polyoxometalate and covalent organic frameworks. Firstly, the prepared polyoxometalate was functionalized with an amine group (NH2-POM-NH2), and then the CPOFs were prepared by a solvothermal Schiff base reaction with NH2-POM-NH2 and 2,4,6-trihydroxybenzene-1,3,5-tricarbaldehyde (Tp) as monomers. After the incorporation of PtNPs and MWCNTs into the CPOFs material, the PtNPs-CPOFs-MWCNTs nanocomposites, which possess excellent catalytic activity and electrical conductivity, were formed and utilized as new electrode materials for the electrochemical thymol sensors. The obtained PtNPs-CPOFs-MWCNTs composite exhibits excellent activity toward thymol, which is attributable to its large special surface area, good conductivity and the synergistic catalysis of each component. Under optimal experimental conditions, the sensor presented a good electrochemical response to thymol. The sensor shows two good linear relationships between the current and thymol concentration in the range of 2-65 µM (R2 = 0.996) and 65-810 µM (R2 = 0.997), with the corresponding sensitivity of 72.7 µA mM-1 and 30.5 µA mM-1, respectively. Additionally, the limit of detection (LOD) was calculated to be 0.2 µM (S/N = 3). At the same time, the prepared thymol electrochemical sensor revealed superior stability and selectivity. The constructed PtNPs-CPOFs-MWCNT electrochemical sensor is the first example of thymol detection.

Dynamic Control of Cyclic Peptide Assembly to Form Higher-Order Assemblies.

Chirality correction, asymmetry, ring-chain tautomerism and hierarchical assemblies are fundamental phenomena in nature. They are geometrically related and may impact the biological roles of a protein or other supermolecules. It is challenging to study those behaviors within an artificial system due to the complexity of displaying these features. Herein, we design an alternating D,L peptide to recreate and validate the naturally occurring chirality inversion prior to cyclization in water. The resulting asymmetrical cyclic peptide containing a 4-imidazolidinone ring provides an excellent platform to study the ring-chain tautomerism, thermostability and dynamic assembly of the nanostructures. Different from traditional cyclic D,L peptides, the formation of 4-imidazolidinone promotes the formation of intertwined nanostructures. Analysis of the nanostructures confirmed the left-handedness, representing chirality induced self-assembly. This proves that a rationally designed peptide can mimic multiple natural phenomena and could promote the development of functional biomaterials, catalysts, antibiotics, and supermolecules.

Controlling supramolecular filament chirality of hydrogel by co-assembly of enantiomeric aromatic peptides.

Supramolecular chirality plays an indispensable role in living and synthetic systems. However, the generation and control of filament chirality in the supramolecular hydrogel of short peptides remains challenging. In this work, as the first example, we report that the heterodimerization of the enantiomeric mixture controls the alignment, chirality, and stiffness of fibrous hydrogels formed by aromatic building blocks. The properties of the resulting racemic hydrogel could not be achieved by either pure enantiomer. Cryo-EM images indicate that the mixture of L and D enantiomers forms chiral nanofibers, the percentage of which can be readily controlled through stoichiometric co-assembly of heterochiral enantiomers. 2D NOESY NMR and diffusion-ordered NMR spectroscopy reveal that heterodimerization of enantiomers plays a crucial role in the formation of chiral nanofibers. Further mechanistic studies unravel the mechanism of supramolecular chirality formation in this two-component system. Molecular dynamics simulations confirm that the intermolecular hydrogen bond and π-π interaction of heterodimers play important roles in forming a chiral hydrogel. Furthermore, regulation of the adhesion and morphology of mammalian cells is achieved by tuning the relative ratio of L and D enantiomers at the same concentration. This work illustrates a novel strategy to control the supramolecular chirality of aromatic peptide hydrogels for materials science.

Carbon Dots-in-EuAPO-5 Zeolite: Triple-Emission for Multilevel Luminescence Anti-Counterfeiting.

Multilevel luminescence materials have aroused wide attention for their advanced anti-counterfeiting abilities. However, various complicated stimuli factors involved in multilevel luminescence anti-counterfeiting (MlLA) limit the practical applications of such materials. Herein, carbon dots (CDs) are in situ introduced into Eu-substituted AlPO4 -5 zeolite (named CDs@EuAPO-5) via a solvent-free thermal crystallization method, which exhibits triple emissions including pink fluorescence mainly associated with Eu3+ in the zeolite framework, blue fluorescence and green room temperature phosphorescence (RTP) associated with CDs. CDs are uniformly embedded in the EuAPO-5 zeolite matrix. Such composite displays excellent photo-, thermo-, and solvent resistance, as well as long-term storage-stability. Moreover, the triple emissions of the composite only need two kinds of common excitation lights to trigger, without involving other complicated stimuli. A triple-level luminescence anti-counterfeiting (TlLA) label has been built, realizing facile, quick, and advanced luminescence anti-counterfeiting that is hard to copy.

Matrix Metalloproteinases (MMP-2) and Tissue Inhibitors of Metalloproteinases (TIMP-2) in Male Inguinal Hernia Patients at Different Ages.

BACKGROUND: Collagen metabolism, controlled by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), might be related to inguinal hernia formation. It was reported that the incidence of inguinal hernia and the recurrence rate after inguinal hernia repair were higher in the elderly. The objective of the research was to assess the amounts of MMP-2 and TIMP-2 in patients at different ages in order to examine the relationship between age and inguinal hernia occurrence. METHODS: The research included 40 primary inguinal hernia male patients, and four groups were created: 50-59 years old (A group); 60-69 years old (B group); 70-79 years old (C group); 80-89 years old (D group). We got the samples from anterior rectus sheath fascia. Real-time fluorescence quantitative polymerase chain reaction (RT-PCR) and immunohistochemistry were applied to estimate the levels of MMP-2 and TIMP-2. RESULTS: The MMP-2 amounts in C and D group were statistically higher than control group (P < 0.05), and the TIMP-2 levels in C and D group were statistically lower than control group (P < 0.05). We found a positive correlation between age and expression levels of MMP-2 (r = 0.537, P < 0.001; r = 0.569, P < 0.001) and a negative correlation between age and TIMP-2 in inguinal hernia patients (r = - 0.759, P < 0.001; r = - 0.759, P < 0.001). CONCLUSIONS: Increased MMP-2 and reduced TIMP-2 may have some relationships with higher inguinal hernia incidence of the elderly.

Structure-Based Programming of Supramolecular Assemblies in Living Cells for Selective Cancer Cell Inhibition.

Here we report on the design, synthesis, and assembly of an enzymatic programmable peptide system inspired by endocytic processes to induce molecular assemblies formation spatiotemporally in living cancer cells, resulting in glioblastoma cell death mainly in necroptosis. Our results indicate the stability and glycosylation of molecules play an essential role in determining the final bioactivity. Detailed mechanistic studies by CLSM, Flow cytometry, western blot, and Bio-EM suggest the site-specific formation of assemblies, which could induce the LMP and activate the downstream cell death pathway. Moreover, we also demonstrate that our strategy can boost the activity of commercial chemotherapy drug by escaping lysosome sequestration. We expected this work would be expanded towards artificial intelligent biomaterials for cancer therapy and imaging precisely.

Programmed death-ligand 1 triggers PASMCs pyroptosis and pulmonary vascular fibrosis in pulmonary hypertension.

Pyroptosis is a pro-inflammatory form of programmed cell death, whose genesis directly depended on caspase-1 activation. Pulmonary hypertension (PH) is a disease characterized, in part, by vascular fibrosis. Up to now, there is no report on the relationship between pyroptosis and vascular fibrosis in PH. Here, we confirmed that pyroptosis had occurred in the media of pulmonary arteries in two PH rat models and hypoxic human pulmonary arterial smooth muscle cells (hPASMCs). Caspase-1 inhibition attenuated the pathogenesis of PH, as assessed by vascular remodeling, right ventricular systolic pressure, right ventricle hypertrophy and hemodynamic parameters of pulmonary vasculature. Moreover, caspase-1 inhibition suppressed pulmonary vascular fibrosis as demonstrated by Masson staining, as well as immunohistochemistry and Western blot analysis of fibrillar collagen. In addition, Programmed death-ligand 1 (PD-L1) was markedly increased in PH, which was regulated by the transcription factor STAT1. Furthermore, PD-L1 knockdown in hPASMCs repressed the onset of hypoxia-induced pyroptosis and fibrosis. Overall, these data identify a critical STAT1-dependent posttranscriptional modification that promotes PD-L1 expression in the pyroptosis of PASMCs to modulate pulmonary vascular fibrosis and accelerate the progression of PH.

Carbon Dots in Porous Materials: Host-Guest Synergy for Enhanced Performance.

Carbon dots (CDs) are emerging as a new class of carbon nanomaterials, which have inspired growing interest for their widespread applications in anti-counterfeiting, sensing, bioimaging, optoelectronic and energy-related fields. In terms of the concept of host-guest assembly, immobilizing CDs into porous materials (PMs) has proven to be an effective strategy to avoid the aggregation of bare CDs in solid state, in particular, the host-guest synergy with both merits of CDs and PMs affords composites promising properties in afterglow and tunable emissions, as well as optimizes their performance in optics, catalysis, and energy storage. This Minireview summarizes the recent progress in the research of CDs@PMs, and highlights synthetic strategies of constructing composites and roles of porous matrices in boosting the applications of CDs in diverse areas. The prospect of future exploration and challenges are proposed for designing advanced CDs-based functional nanocomposite materials.

Reconfigurable Assembly of Active Liquid Metal Colloidal Cluster.

We report the reconfigurable assembly of rod-shaped eutectic gallium-indium alloy (EGaIn) liquid metal colloidal motors by mimicking the growth behavior of a dandelion. EGaIn nanorods with a diameter of 210 nm and a length of 850 nm were synthesized via an ultrasound-assisted physical dispersion method. The nanorods possess a core-shell structure with a 30 nm GaOOH shell and zero-valent liquid core. The EGaIn motors move autonomously at a speed of 41.2 µm s-1 under an acoustic field. By modulating the frequency of the applied acoustic field, the EGaIn colloidal motors self-organize into various striped and circular patterns, followed by a flower-like cluster. The dandelion-like EGaIn colloidal motor clusters move collectively and redisperse when the applied acoustic frequency is changed. Numerical simulations reveal that the flower-like clusters are created by the acoustic propulsion in combination with steric repulsion and hydrodynamics.

Iron overload threatens the growth of osteoblast cells via inhibiting the PI3K/AKT/FOXO3a/DUSP14 signaling pathway.

Iron overload is a common stress in the development of cells. Growing evidence has indicated that iron overload is associated with osteoporosis. Therefore, enhancing the understanding of iron overload would benefit the development of novel approaches to the treatment of osteoporosis. The purpose of the present study was to analyze the effect of iron overload on osteoblast cells, via the MC3T3-E1 cell line, and to explore its possible underlying molecular mechanisms. Ferric ammonium citrate (FAC) was utilized to simulate iron overload conditions in vitro. FAC-induced iron overload strongly suppressed proliferation of osteoblast cells and induced apoptosis. Moreover, iron overload strongly suppressed the expression of dual-specificity phosphatase 14 (DUSP14). Additionally, overexpression of DUSP14 protected osteoblast cells from the deleterious effects of iron overload, and this protective effect was mediated by FOXO3a. Additionally, matrine rescued the function of DUSP14 in osteoblast cells. Most importantly, our analysis demonstrated the essential role of the PI3K/AKT/FOXO3a/DUSP14 signaling pathway in the defense against iron overload in osteoblast cells. Overall, our results not only elucidate deleterious effects of iron overload, but also unveil its possible signaling pathway in osteoblast cells.

Carbon Dots-in-Matrix Boosting Intriguing Luminescence Properties and Applications.

As a new class of luminescent nanomaterials, carbon dots (CDs) have aroused significant interest because of their fascinating photoluminescence properties and potential applications in biological, optoelectronic, and energy-related fields. Strikingly, embedding CDs in host matrices endow them with intriguing luminescent properties, in particular, room temperature phosphorescence and thermally activated delayed fluorescence, due to the confinement effect of the host matrix and the H-bonding interactions between CDs and the matrix. Here, the state-of-the-art strategies for introducing CDs in various host matrices are summarized, such as nanoporous materials, polyvinyl alcohol, polyurethane, potash alum, layered double hydroxides, amorphous silica, etc. The resultant luminescent properties of the composites and their emission mechanisms are discussed. Their applications in bioimaging, drug delivery/release, sensing, and anticounterfeiting are also presented. Finally, current problems and challenges of CDs-based composites are noted for future development of such luminescent materials.

Novel laser positioning navigation to aid puncture during percutaneous nephrolithotomy: a preliminary report.

PURPOSE: Accurate puncture of the renal collecting system is crucial to the success of percutaneous nephrolithotomy and presents a technical challenge for urologists. Here, we introduced the Surgical Approach Visualization and Navigation (SAVN) system, a novel navigation system to assist puncture and reduce intraoperative radiation. MATERIALS AND METHODS: Twenty kidneys of 10 cadavers were randomly divided into two groups for renal calyx puncture. In the control group, traditional fluoroscopy was used for guidance, while SAVN system was used in the experimental group. Puncture duration, number of puncture attempts, total number of intraoperative fluoroscopies, and number of fluoroscopies during the puncture procedure were recorded. RESULTS: The puncture duration was 14.2 ± 2.5 s in SAVN group and 48.3 ± 7.1 s in conventional group (P < 0.05). One puncture attempt was needed for successful puncture in SAVN group, while more than one in conventional group (P = 0.28). The total number of intraoperative fluoroscopies was 3.3 ± 1.0 in SAVN group and 14.5 ± 3.1 in control group (P < 0.05),while the number of fluoroscopies during the puncture procedure was 0 and 11.2 ± 2.4, respectively (P < 0.05). CONCLUSIONS: The novel SAVN system has a simplified structure and is easy to use. It can be used to successfully assist with puncture of the renal calyx, thus reducing puncture duration and radiation dose.

Thermoresponsive Polymer Brush Modulation on the Direction of Motion of Phoretically Driven Janus Micromotors.

We report a thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) brush functionalized Janus Au-Pt bimetallic micromotor capable of modulating the direction of motion with the change of the ambient temperature. The PNIPAM@Au-Pt micromotor moved along the Au-Pt direction with a speed of 8.5 µm s-1 in 1.5 % H2 O2 at 25 °C (below the lower critical solution temperature (LCST) of PNIPAM), whereas it changed the direction of motion (i.e., along the Pt-Au direction) and the speed decreased to 2.3 µm s-1 at 35 °C (above LCST). Below LCST, PNIPAM brushes grafted on the Au side were hydrophilic and swelled, which permitted the electron transfer and proton diffusion on the Au side, and thus the motion is regarded as a self-electrophoretic mechanism. However, PNIPAM brushes above LCST became hydrophobic and collapsed, and thus the driving mechanism switched to the self-diffusiophoresis like that of Pt-modified Janus silica motors. These motors could reversibly change the direction of motion with the transition of the hydrophobic and hydrophilic states of the grafted PNIPAM brushes. Such a thermoresponsive polymer brush functionalization method provides a new strategy for engineering the kinematic behavior of phoretically driven micro/nanomotors.

Carbon Dots in a Matrix: Energy-Transfer-Enhanced Room-Temperature Red Phosphorescence.

High-efficiency red room-temperature phosphorescence (RTP) emissions have been achieved by embedding carbon dots (CDs) in crystalline Mn-containing open-framework matrices. The rationale of this strategy relies on two factors: 1) the carbon source, which affects the triplet energy levels of the resulting CDs and thus the spectral overlap and 2) the coordination geometry of the Mn atoms in the crystalline frameworks, which determines the crystal-field splitting and thus the emission spectra. Embedding the carbon dots into a matrix with 6-coordinate Mn centers resulted in a strong red RTP with a phosphorescence efficiency of up to 9.6 %, which is higher than that of most reported red RTP materials. The composite material has an ultrahigh optical stability in the presence of strong oxidants, various organic solvents, and strong ultraviolet radiation. A green-yellow RTP composite was also prepared by using a matrix with 4-coordinate Mn centers and different carbon precursors.

MMP-2 and MMP-9 contribute to the angiogenic effect produced by hypoxia/15-HETE in pulmonary endothelial cells.

Matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) are the predominant gelatinases in the developing lung. Studies have shown that the expression of MMP-2 and MMP-9 is upregulated in hypoxic fibroblasts, 15-hydroxyeicosatetraenoic acid (15-HETE) regulated fibroblasts migration via modulating MMP-2 or MMP-9, and that hypoxia/15-HETE is a predominant contributor to the development of pulmonary arterial hypertension (PAH) through increased angiogenesis. However, the roles of MMP-2 and MMP-9 in pulmonary arterial endothelial cells (PAECs) angiogenesis as well as the molecular mechanism of hypoxia-regulated MMP-2 and MMP-9 expression have not been identified. The aim of this study was to investigate the role of MMP-2 and MMP-9 in PAEC proliferation and vascular angiogenesis and to determine the effects of hypoxia-induced 15-HETE on the expression of MMP-2 and MMP-9. Western blot, immunofluorescence, and real-time PCR were used to measure the expression of MMP-2 and MMP-9 in hypoxic PAECs. Immunohistochemical staining, flow cytometry, and tube formation as well as cell proliferation, viability, scratch-wound, and Boyden chamber migration assays were used to identify the roles and relationships between MMP-2, MMP-9, and 15-HETE in hypoxic PAECs. We found that hypoxia increased MMP-2 and MMP-9 expression in pulmonary artery endothelium both in vivo and in vitro in a time-dependent pattern. Moreover, administration of the MMP-2 and MMP-9 inhibitor MMI-166 significantly reversed hypoxia-induced increases in right ventricular systemic pressure (RVSP), right ventricular function, and thickening of the tunica media. Furthermore, up-regulation of MMP-2 and MMP-9 expression was induced by 15-HETE, which regulates PAEC proliferation, migration, and cell cycle transition that eventually leads to angiogenesis. Our study demonstrated that hypoxia increases the expression of MMP-2 and MMP-9 through the 15-lipoxygenase/15-HETE pathway, and that MMP-2 and MMP-9 promote PAEC angiogenesis. These findings suggest that MMP-2 and MMP-9 may serve as new potential therapeutic targets for the treatment of PAH.

Effect of miR-182 on hepatic fibrosis induced by Schistosomiasis japonica by targeting FOXO1 through PI3K/AKT signaling pathway.

The study aimed to investigate the impact of miR-182 and FOXO1 on S. japonica-induced hepatic fibrosis. Microarray analysis was performed to screen out differential expressed miRNAs and mRNAs. Rat hepatic fibrosis model and human hepatocellular cell line LX-2 were used to study the effect of miR-182 and FOXO1. qRT-PCR and Western blot were used to detect the expression of miR-182, FOXO1 or other fibrosis markers. The targeting relationship between FOXO1 and miR-182 was verified by luciferase reporter assay. Immunohistochemistry or immunofluorescence staining was conducted to detect FOXO1 or α-SMA in rat hepatic tissues. Cell viability and apoptosis were detected by MTT assay and flow cytometry. The expression of PI3K/AKT pathway-related proteins was detected by Western blot. miR-182 was highly expressed in liver fibrosis samples, and FOXO1 expression was negatively correlated with miR-182 expression. After transfection of miR-182, FOXO1 expression was down-regulated, with the results of LX-2 cells proliferation inhibition and apoptosis induction, as well as the aggravation of rat hepatic fibrosis. The expression of p-AKT/AKT and p-S6/S6 was increased, meaning that the PI3K/AKT signal pathway was activated. The results were reversed when treated with Wortmannin (PI3K inhibitor). After transfection of miR-182 inhibitor, FOXO1 expression was up-regulated, LX-2 cell proliferation was inhibited, and apoptosis rate was increased. High-expressed miR-182 and low-expressed FOXO1 promoted proliferation and inhibiting apoptosis on liver fibrosis cells, stimulating the development of S. japonica-induced hepatic fibrosis through feeding back to PI3K/AKT signaling pathway.

Bone morphogenetic protein-7 inhibits endothelial-mesenchymal transition in pulmonary artery endothelial cell under hypoxia.

Pulmonary artery hypertension (PAH) is characterized by structural changes in pulmonary arteries. Increased numbers of cells expressing α-smooth muscle actin (α-SMA) is a nearly universal finding in the remodeled artery. It has been confirmed endothelial-to-mesenchymal transition (EndoMT) may be a source of those α-SMA-expressing cells. In addition, the EndoMT is reversible. Here, we show that under hypoxia, the expression of bone morphogenetic protein 7 (BMP-7) was decreased both in vivo and in vitro. We also found that under normoxia, BMP-7 deficiency induced spontaneous EndoMT and cell migration. The hypoxia-induced EndoMT and cell migration were markedly attenuated after pretreatment with rh-BMP-7. Moreover, m-TOR phosphorylation was involved in EndoMT and BMP-7 suppressed hypoxia-induced m-TORC1 phosphorylation in pulmonary artery endothelial cells. Our results demonstrate that BMP-7 attenuates the hypoxia-induced EndoMT and cell migration by suppressing the m-TORC1 signaling pathway. Our study revealed a novel mechanism underlying the hypoxia-induced EndoMT in pulmonary artery endothelial cells and suggested a new therapeutic strategy targeting EndoMT for the treatment of pulmonary arterial hypertension.