Fabrication of Perfluoropolyether Microfluidic Devices Using Laser Engraving for Uniform Droplet Production.

A perfluoropolyether (PFPE)-based microfluidic device with cross-junction microchannels was fabricated with the purpose of producing uniform droplets. The microchannels were developed using CO2 laser engraving. PFPE was chosen as the main material because of its excellent solvent resistance. Polyethylene glycol diacrylate (PEGDA) was mixed with PFPE to improve the hydrophilic properties of the inner surface of the microchannels. The microchannels of the polydimethylsiloxane microfluidic device had a blackened and rough surface after laser engraving. By contrast, the inner surface of the microchannels of the PFPE-PEGDA microfluidic device exhibited a smooth surface. The lower power and faster speed of the laser engraving resulted in the development of microchannels with smaller dimensions, less than 30 µm in depth. The PFPE and PFPE-PEGDA microfluidic devices were used to produce uniform water and oil droplets, respectively. We believe that such a PFPE-based microfluidic device with CO2-laser-engraved microchannels can be used as a microfluidic platform for applications in various fields, such as biological and chemical analysis, extraction, and synthesis.

Microfluidic Bioreactor with Fibrous Micromixers for In Vitro mRNA Transcription.

A new type of microfluidic bioreactor with fibrous micromixers for the ingredient mixing and a long macrochannel for the in vitro transcription reaction was fabricated for the continuous production of mRNA. The diameter of the fibrous microchannels in the micromixers was tuned by using an electrospun microfibrous disc with different microfiber diameters. The micromixer with a larger diameter of fibrous microchannels exhibited a better mixing performance than the others. The mixing efficiency was increased to 0.95 while the mixture was passed through the micromixers, suggesting complete mixing. To demonstrate the continuous production of mRNA, the ingredients for in vitro transcription were introduced into the perfluoropolyether microfluidic bioreactor. The mRNA synthesized by the microfluidic bioreactor had the same sequence and in vitro/in vivo performances as those prepared by the bulk reaction. The continuous reaction in the microfluidic bioreactor with efficient mixing performance can be used as a powerful platform for various microfluidic reactions.

Continuous production of lipid nanoparticles by multiple-splitting in microfluidic devices with chaotic microfibrous channels.

Polydimethylsiloxane (PDMS) microfluidic devices with chaotic microfibrous channels were fabricated for the continuous production of lipid nanoparticles (LNPs). Electrospun poly(ε-caprolactone) (PCL) microfibrous matrices with different diameters (3.6 ± 0.3, 6.3 ± 0.4, and 12.2 ± 0.8 µm) were used as a template to develop microfibrous channels. The lipid solution (in ethanol) and water phase were introduced into the microfluidic device as the discontinuous and continuous phases, respectively. The smaller diameter of microfibrous channels and the higher flow rate of the continuous phase resulted in the smaller LNPs with a narrower size distribution. The multiple-splitting of the discontinuous phase and the microscale contact between the two phases in the microfibrous channels were the key features of the LNP production in our approach. The LNPs containing doxorubicin with different average sizes (89.7 ± 35.1 and 190.4 ± 66.4 nm) were prepared using the microfluidic devices for the potential application in tumor therapy. In vitro study revealed higher cellular uptake efficiency and cytotoxicity of the smaller LNPs, especially in the HepG2 cells. The microfluidic devices with microfibrous channels can be widely used as a continuous and high-throughput platform for the production of LNPs containing various active agents.

In Situ Spontaneous Fabrication of Tough and Stretchable Polyurethane-Polyethyleneimine Hydrogels Selectively Triggered by CO2.

CO2 -triggered in situ hydrogels is developed from waterborne poly(ε-caprolactone)-based polyurethane (PU) dispersion and aqueous polyethyleneimine (PEI) solution without any other chemicals and apparatus (e.g., UV light). In the approach, nontoxic CO2 in air is used as a selective trigger for the hydrogel formation. CO2 adsorption onto PEI results in the formation of ammonium cations in PEI and the subsequent multiple ionic crosslinking between PU and PEI chains. Besides the amount of CO2 in air, the rate of hydrogel formation can be controlled by NaHCO3 in the PU-PEI mixture, which serves as a CO2 supplier. The PU hydrogels exhibit tough and stretchable properties with high tensile strength (2.05 MPa) and elongation at break (438.24%), as well as biocompatibility and biodegradability. In addition, the PU hydrogels exhibit high adhesion strength on skin and injectability due to the in situ formation. It is believed that these PU hydrogels have the ideal features for various future applications, such as tissue adhesion barriers, wound dressing, artificial skin, and injectable fillers.

Fabrication of Microfiber-Templated Microfluidic Chips with Microfibrous Channels for High Throughput and Continuous Production of Nanoscale Droplets.

A polydimethylsiloxane (PDMS) microfluidic chip with well-interconnected microfibrous channels was fabricated by using an electrospun poly(ε-caprolactone) (PCL) microfibrous matrix and 3D-printed pattern as templates. The microfiber-templated microfluidic chip (MTMC) was used to produce nanoscale emulsions and spheres through multiple emulsification at many small micro-orifice junctions among microfibrous channels. The emulsion formation mechanisms in the MTMC were the cross-junction dripping or Y-junction splitting at the micro-orifice junctions. We demonstrated the high throughput and continuous production of water-in-oil emulsions and polyethylene glycol-diacrylate (PEG-DA) spheres with controlled size ranges from 2.84 µm to 83.6 nm and 1.03 µm to 45.7 nm, respectively. The average size of the water droplets was tuned by changing the micro-orifice diameter of the MTMC and the flow rate of the continuous phase. The MTMC theoretically produced 58 trillion PEG-DA nanospheres per hour without high shear force. In addition, we demonstrated the higher encapsulation efficiency of the PEG-DA microspheres in the MTMC than that of the microspheres fabricated by ultrasonication. The MTMC can be used as a powerful platform for the large-scale and continuous productions of emulsions and spheres.

Enhanced osteogenic differentiation of alendronate-conjugated nanodiamonds for potential osteoporosis treatment.

BACKGROUND: Alendronate (Alen) is promising material used for bone-targeted drug delivery due to its high bone affinity and therapeutic effects on bone diseases. In addition, Alen can enhance the osteogenic differentiation of osteoblastic cell. Recently, nanodiamonds (NDs) with hardness, non-toxicity, and excellent biocompatibility are employed as promising materials for carrier systems and osteogenic differentiation. Therefore, we prepared Alen-conjugated NDs (Alen-NDs) and evaluated their osteogenic differentiation performances. METHODS: Alen-NDs were synthesized using DMTMM as a coupling reagent. Morphological change of Mouse calvaria-derived preosteoblast (MC3T3-E1) treated with Alen-NDs was observed using the confocal microscope. The osteogenic differentiation was confirmed by cell proliferation, alkaline phosphatase (ALP), calcium deposition, and real-time polymerase chain reaction assay. RESULTS: Alen-NDs were prepared to evaluate their effect on the proliferation and differentiation of osteoblastic MC3T3-E1 cells. The Alen-NDs had a size of about 100 nm, and no cytotoxicity at less than 100 µg/mL of concentration. The treatment of NDs and Alen-NDs reduced the proliferation rate of MC3T3-E1 cells without cell death. Confocal microscopy images confirmed that the treatment of NDs and Alen-NDs changed the cellular morphology from a fibroblastic shape to a cuboidal shape. Flow cytometry, alkaline phosphatase (ALP) activity, calcium deposition, and real-time polymerase chain reaction (RT-PCR) confirmed the higher differentiation of MC3T3-E1 cells treated by Alen-NDs, compared to the groups treated by osteogenic medium and NDs. The higher concentration of Alen-ND treated in MC3T3-E1 resulted in a higher differentiation level. CONCLUSIONS: Alen-NDs can be used as potential therapeutic agents for osteoporosis treatment by inducing osteogenic differentiation.

Organic matrix-free imaging mass spectrometry.

Mass spectrometry (MS) is an ideal tool for analyzing multiple types of (bio)molecular information simultaneously in complex biological systems. In addition, MS provides structural information on targets, and can easily discriminate between true analytes and background. Therefore, imaging mass spectrometry (IMS) enables not only visualization of tissues to give positional information on targets but also allows for molecular analysis of targets by affording the molecular weights. Matrixassisted laser desorption/ionization-time of flight (MALDI-TOF) MS is particularly effective and is generally used for IMS. However, the requirement for an organic matrix raises several limitations that get in the way of accurate and reliable images and hampers imaging of small molecules such as drugs and their metabolites. To overcome these problems, various organic matrix-free LDI IMS systems have been developed, mostly utilizing nanostructured surfaces and inorganic nanoparticles as an alternative to the organic matrix. This minireview highlights and focuses on the progress in organic matrix-free LDI IMS and briefly discusses the use of other IMS techniques such as desorption electrospray ionization, laser ablation electrospray ionization, and secondary ion mass spectrometry. [BMB Reports 2020; 53(7): 349-356].

Polyaniline-grafted nanodiamonds for efficient photothermal tumor therapy.

Polyaniline-grafted nanodiamond (PAN-ND) nanoparticles were fabricated by polymerizing aniline at the surface of amine-modified NDs for efficient photothermal therapy (PTT). A series of PAN from different aniline concentrations were also prepared to compare the properties and the efficiency of PTT. The polymerization rate of aniline was faster in the presence of NDs than that of aniline alone. Compared to PAN nanoparticles, PAN-ND has a spherical shape, smaller size, and ultimately higher cellular uptake efficiency. The temperature of aqueous PAN-ND dispersion increased to 44.4 °C after laser irradiation for 5 min. In addition, the UV absorbance intensity of PAN-ND increased at the lower pH at the near infrared (NIR) region, resulting in an enhanced photothermal effect at a tumor site. Notably, the viability of HeLa cells treated with PAN-ND decreased by less than 20%, suggesting the high efficiency of PTT. The PAN-ND can be a potential candidate for efficient photothermal tumor therapy.

Immobilization of phenol-containing molecules on self-assembled monolayers on gold via surface chemistry.

Various phenol-containing molecules such as flavonoids have a wide range of biological effects including anticancer, antimicrobial, and anti-inflammatory properties, and, therefore, they have become subjects of active research for various medicinal and biological applications. To construct applicable materials incorporated with phenol-containing molecules, strategies for immobilization of phenol-containing molecules on solid substrates are required. Although several immobilization methods have been devised and reported, mostly harnessing phenol functionality, however, development of a general immobilization method has been hampered due to its complicated chemical reactions and low reaction yields on surfaces. Furthermore, the use of phenol as a reaction center may compromise the biological activity of phenol-containing molecules. Here, we describe a simple, fast, and reliable method for the surface immobilization of phenol-containing molecules by introducing chemical functional groups, carboxylic acid, thiol, and azide, while maintaining phenol functionality by way of the Mannich-type condensation reaction. We examined the chemical functionalization of naphthol, tyrosine, and flavanone and their immobilization to the self-assembled monolayers on gold via various surface chemistries: the carbodiimide coupling reaction, Michael addition, and the 'click' reaction. We strongly believe our method can be a general and practical platform for immobilization of various phenol-containing molecules on surfaces of various materials.

On-Demand Modulation of Bacterial Cell Fates on Multifunctional Dynamic Substrates.

This paper reports unprecedented dynamic surfaces based on zwitterionic low-density self-assembled monolayers (LDSAMs) of alkanethiolates on gold, which integrate three interconvertible states-bacteria-adherable, bactericidal, and nonfouling states-through electrical modulations. The conformations of alkanethiolates were electrically modulated to generate zwitterionic, anionic, and cationic surfaces, which responded differently to bacteria and determined the fate of bacteria. Furthermore, the reversible switching of multifunctions of the surface was realized for killing bacteria and subsequently releasing dead bacteria from the surface. For practical application of our strategy, we examined the selective antibacterial effect of our surface for eradication of mycoplasma contaminants in contaminated mammalian cell cultures.