Gypenosides exert cardioprotective effects by promoting mitophagy and activating PI3K/Akt/GSK-3ß/Mcl-1 signaling.

Background: Gynostemma pentaphyllum (Thunb.) Makino, a well-known edible and medicinal plant, has anti-aging properties and is used to treataging-associated conditions such as diabetes, metabolic syndrome, and cardiovascular diseases. Gypenosides (GYPs) are the primary constituents of G. pentaphyllum. Increasing evidence indicates that GYPs are effective at preserving mitochondrial homeostasis and preventing heart failure (HF). This study aimed to uncover the cardioprotective mechanisms of GYPs related to mitochondrial regulation. Methods: The bioactive components in GYPs and the potential targets in treating HF were obtained and screened using the network pharmacology approach, followed by drug-disease target prediction and enrichment analyses. The pharmacological effects of GYPs in cardioprotection, mitochondrial function, mitochondrial quality control, and underlying mechanisms were further investigated in Doxorubicin (Dox)-stimulated H9c2 cardiomyocytes. Results: A total of 88 bioactive compounds of GYPs and their respective 71 drug-disease targets were identified. The hub targets covered MAPK, EGFR, PI3KCA, and Mcl-1. Enrichment analysis revealed that the pathways primarily contained PI3K/Akt, MAPK, and FoxO signalings, as well as calcium regulation, protein phosphorylation, apoptosis, and mitophagy process. In Dox-stimulated H9c2 rat cardiomyocytes, pretreatment with GYPs increased cell viability, enhanced cellular ATP content, restored basal oxygen consumption rate (OCR), and improved mitochondrial membrane potential (MMP). Furthermore, GYPs improved PINK1/parkin-mediated mitophagy without influencing mitochondrial fission/fusion proteins and the autophagic LC3 levels. Mechanistically, the phosphorylation of PI3K, Akt, GSK-3ß, and the protein level of Mcl-1 was upregulated by GYP treatment. Conclusion: Our findings reveal that GYPs exert cardioprotective effects by rescuing the defective mitophagy, and PI3K/Akt/GSK-3ß/Mcl-1 signaling is potentially involved in this process.

Extracellular Matrix-Mimetic Peptide Scaffolds Prolonged the Hypothermic Preservation of Stem Cells for Storage and Transportation.

Encouraging advances in both regenerative medicine and tissue engineering with stem cells require a short-term preservation protocol to provide enough time for quality control or the transportation of cell products from manufacturing facilities to clinical destinations. The hypothermic preservation of stem cells under refrigerated conditions (2-8 °C) in their specific culture medium provides an alternative and low-cost method for cryopreservation or commercial preservation fluid for short-term storage. However, most stem cells are vulnerable to hypothermia, which might result in cell damage from the cooling process and the lack of extracellular matrix (ECM). Herein, we report a peptide scaffold cell-culture-medium additive for mimicking in vivo ECM to enhance the storage efficiency of mesenchymal stem cells (MSCs) under hypothermic preservation. Peptide scaffolds exhibit protective effects against hypothermic injury by maintaining the viability, proliferation, migration, and differentiation capabilities of cells. The mechanistic study showed that the peptide scaffold was conducive to maintain mitochondrial function by retaining mitochondrial respiration, mitochondrial membrane potential (ΔΨm), and mass to alleviate intracellular and mitochondrial reactive oxygen species (ROS) production. Moreover, the peptide scaffold also prolonged the survival and retained the multipotency of hematopoietic stem and progenitor cells (HSPCs) under hypothermic conditions. In conclusion, these results demonstrate a feasible and convenient preservation system for stem cells that has the potential to promote the clinical application of hematopoietic stem cell therapy.

[Factors Affecting the Preservation of Human Peripheral Blood Mononuclear Cells at 4 ℃].

OBJECTIVE: To analyze the preservation effect and related influencing factors of human peripheral blood mononuclear cells under serum-free condition at 4 ℃. METHODS: Human peripheral blood mononuclear cells were isolated by density gradient centrifugation, and stored at 4 ℃ under different cell concentrations, supplemented with human serum albumin, and glucose. The cell viability, total cell number, viable cell number and cell phenotype were detected during preservation of 72 h. RESULTS: With the prolongation of storage time, the number of human peripheral blood mononuclear cells gradually decreased(r=0.982). Compared with the cell concentration of (5-6)×106 cells/ml, the cell number decreased more slowly when the cell storage concentration was (1-2)×106 cells/ml; Adding human serum albumin and glucose can effectively improve the survival rate of human peripheral blood mononuclear cells, among which 2% human serum albumin has a better preservation effect; Compared with the blank control group, the analysis results of cell subsets showed that the downward trends of NK cells and T cells were significantly slowed after adding albumin and glucose. CONCLUSION: The cell density of (1-2)×106/ml and 2% human serum albumin are more suitable for the preservation of PBMC, and 5% glucose can improve the preservation effect of human peripheral blood mononuclear cells at 4 ℃.

Tunable narrow-linewidth high-peak-power sub-nanosecond optical parametric generator by injection seeding.

We present an efficient tunable optical parametric generator (OPG) with its linewidth close to the Fourier transform limit by injection seeding a tunable diode laser. Benefitting from high-peak-power sub-nanosecond (426 ps) laser pumping and a high-gain MgO:PPLN (PPMgLN) crystal, the OPG produced signal peak power up to 0.343 MW at 1638 nm and the total conversion efficiency reached 47.9% at 1-kHz pulse repetition rate. Considering the linewidth limit of short signal pulses (∼ 350 ps), a tunable seeder with the linewidth at hundred-MHz level was applicable. The achieved OPG signal tuning range was 1510-1638 nm with linewidth at GHz level, which is two orders of magnitude narrower than the unseeded OPG. Injection seeding a non-resonant OPG device does not introduce extra cavity feedback electronics that are essential for an optical parametric oscillator (OPO), greatly improving robustness and reducing cost. It is believed such a compact, tunable and costless PPMgLN OPG with high peak power, high repetition rate and relatively narrow linewidth has great significance in lidar, spectroscopy, etc.

Spectroscopic Technique-Based Comparative Investigation on the Interaction of Theaflavins with Native and Glycated Human Serum Albumin.

Theaflavin is a kind of multi-pharmacological and health beneficial black tea factor. The aim of this study is to investigate the mechanisms by which theaflavin interacts with glycosylated and non-glycosylated serum albumins and compares their binding properties. Fluorescence and ultraviolet spectra indicated that theaflavin interacted with native and glycated human serum albumin through a static quenching mechanism and had a higher degree of quenching of human serum albumin. The thermodynamic parameters revealed that the combinations of theaflavin with native and glycated human serum albumin were a spontaneous endothermic reaction, and the hydrophobic force was a major driving force in the interaction process. Zeta potential, particle size, synchronous fluorescence, three-dimensional fluorescence spectroscopy and circular dichroism further clarified the effect of theaflavin on the conformation of human serum albumin structure were more pronounced. In addition, site competition experiments and molecular docking technique confirmed that the binding sites of theaflavin on both native and glycated human serum albumin were bound at site II. This study had investigated the effects of glycation on the binding of HSA with polyphenols and the potential nutriology significance of these effects.

Mapping ApoE/Aß binding regions to guide inhibitor discovery.

Blocking the interaction between the E4 isoform of apolipoprotein E (ApoE) and amyloid beta-peptide (Aß) may be an avenue for pharmacological intervention in Alzheimer's disease (AD). The main regions of interaction of the two proteins are, respectively, ApoE244-272 and Aß12-28. These protein segments are too large to facilitate the design of small molecule inhibitors. We mapped the primary components of ApoE/Aß interaction to smaller peptide segments. Within the three motifs that are primarily responsible for ApoE/Aß interaction, we identified four peptides that substantially block ApoE/Aß interaction and further improved their inhibitory activity by rational hydrophobic amino acid substitution. Moreover, the mapping results provide the clue that the Aß residues which interact with ApoE appear to be in the same region where Aß self-interacts. According to this information, we found that Congo Red and X-34 could strongly inhibit ApoE/Aß interaction. Our findings extend our understanding of ApoE/Aß interaction and may guide the discovery of inhibitors that treat AD by antagonizing ApoE/Aß interaction.

Fabrication of freestanding, microperforated membranes and their applications in microfluidics.

This manuscript describes a convenient method for the fabrication of freestanding, microperforated membranes in photocurable polymers using only one step of photolithography. We used photosensitive prepolymers to make the membranes and photolithography to define the micropatterns. We demonstrated the fabrication of single- and multilayer microperforated membranes in SU-8 photoresist and Norland Optical Adhesive prepolymer. These membranes can be used to pattern surfaces in various materials and to fabricate complex three-dimensional microfluidic channel structures.

Fabrication of a microfluidic Ag/AgCl reference electrode and its application for portable and disposable electrochemical microchips.

This report describes a convenient method for the fabrication of a miniaturized, reliable Ag/AgCl reference electrode with nanofluidic channels acting as a salt bridge that can be easily integrated into microfluidic chips. The Ag/AgCl reference electrode shows high stability with millivolt variations. We demonstrated the application of this reference electrode in a portable microfluidic chip that is connected to a USB-port microelectrochemical station and to a computer for data collection and analysis. The low fabrication cost of the chip with the potential for mass production makes it disposable and an excellent candidate for real-world analysis and measurement. We used the chip to quantitatively analyze the concentrations of heavy metal ions (Cd(2+) and Pb(2+)) in sea water. We believe that the Ag/AgCl reference microelectrode and the portable electrochemical system will be of interest to people in microfluidics, environmental science, clinical diagnostics, and food research.

A prototypic microfluidic platform generating stepwise concentration gradients for real-time study of cell apoptosis.

This work describes the development of a prototypic microfluidic platform for the generation of stepwise concentration gradients of drugs. A sensitive apoptotic analysis method is integrated into this microfluidic system for studying apoptosis of HeLa cells under the influence of anticancer drug, etoposide, with various concentrations in parallel; it measures the yellow fluorescent proteincyan fluorescent protein fluorescence resonance energy transfer (FRET) signal that responds to the activation of caspase-3, an indicator of cell apoptosis. Sets of microfluidic valves on the chip generate stepwise concentration gradient of etoposide in various cell-culture microchambers. The FRET signals from multiple chambers are simultaneously monitored under a fluorescent microscope for long-time observation and the on-chip results are compared with those from 96-well plate study and the methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. The microfluidic platform shows several advantages including high-throughput capacity, low drug consumption, and high sensitivity.

Generation of alginate microfibers with a roller-assisted microfluidic system.

This work describes a convenient microfluidic strategy with a simple and minimal-requirement design for the generation of uniformly-sized alginate gel fibers with diameters one order of magnitude smaller than those formed in conventional methods. Different from common microfluidic methods, our system contains a single microchannel through which alginate solutions are transported and squeezed into a calcium chloride solution. Ca2+ ions immediately cross link the alginate solution into a gel state that is collected with a rotor. We use microchannels to define the initial size of the alginate fibers and a roller to further reduce the fiber size (diameter) by one order of magnitude down to approximately 1 microm. The size of the formed fibers can be well controlled by adjusting the corresponding parameters, and be predicted with a simple equation. In addition, various functional materials (colloidal particles, bacteria and nanoparticles) have been encapsulated into the gel fibers with this technique.

A screw-actuated pneumatic valve for portable, disposable microfluidics.

This work describes a simple and inexpensive approach for controlling the pneumatic valves that were invented in Quake's group to miniaturize the whole system for portable and disposable microfluidic devices. The valves are assembled from two parts. One is the polydimethylsiloxane (PDMS) channels formed by multilayer soft lithography. The other is a polymethylmethacrylate (PMMA) frame with machine screws for pressure control. Turning the screws into the control channel inlet (filled with water and covered with a thin PDMS membrane) actuates the valve by creating pressure in the control channel. This method avoids the bulky and expensive external pressure-control facilities and can be easily integrated into portable and disposable devices.

Supercritical fluid extraction of ecdysterone from the roots of Achyranthes bidentata BL.

Ecdysterone has been found in a great many plants and animals and has some valuable pharmaceutical properties. The present study was conducted to investigate optimal conditions for the extraction of the compound by supercritical fluid extraction from the roots of Achyranthes bidentata BL. An orthogonal array design (OAD), OA(9)(3(4)), was employed as a chemometric method for optimization of the extraction of ecdysterone from the herbal medicine. Four parameters, namely, pressure and temperature of the supercritical fluid, the dynamic extraction time, and the flow rate of dimethyl sulfoxide, were studied and optimized by a three-level OAD. Determinations of the extracts were performed by high-performance liquid chromatography. The effects of the parameters were studied using analysis of variance. The results shown that the yield of ecdysterone could be influenced by the four parameters to a similar degree. The yield for DMSO-modified supercritical CO(2) was in the range from 0.65 to 1.03 mg/g under the selected conditions. In comparison with methanol-modified supercritical CO(2 )and Soxhlet extraction, a higher yield was obtained when DMSO-modified supercritical CO(2) was used.