Photoresponsive Aqueous Dissolution of Poly(<i>N</i>‑Isopropylacrylamide) Functionalized with <i>o</i>‑Nitrobenzaldehyde through Phase Transition

We report a sharp photoinduced aqueous dissolution of the copolymer through phase transition based on the photochemical reaction of <i>o</i>-nitrobenzaldehyde (NBA) and the principle of polymer effect. We synthesized the copolymers having poly­(<i>N</i>-isopropylacrylamide) main chain and NBA side chain at 4, 7, and 10 mol % functionalizations and analyzed their photoresponsive characteristics. Light with 365 nm wavelength converted NBA groups at copolymer side chains to carboxylic acid efficiently at the rate of 7.3 cm²/J, and in the case of 10 mol % functionalization, the irradiation dosage no more than 56 mJ/cm² induced sharp aqueous dissolution of the copolymer thin layer in pH 7.4 at 25 °C. As example applications, we demonstrated on-demand release of polyethylene beads and fluorescent-labeled albumins, which had been immobilized on a substrate surface via the copolymers, by the precisely controlled light irradiation using a microprojection system. Also, we examined application of the copolymers to the selective recovery of living cells from culture substrate under microscopic observation. As a result, mild light irradiation at room temperature triggered immediate detachment of the cultured adherent cells only in the irradiated areas without critical influence on their viability

Photoresponsive Aqueous Dissolution of Poly(<i>N</i>‑Isopropylacrylamide) Functionalized with <i>o</i>‑Nitrobenzaldehyde through Phase Transition

We report a sharp photoinduced aqueous dissolution of the copolymer through phase transition based on the photochemical reaction of <i>o</i>-nitrobenzaldehyde (NBA) and the principle of polymer effect. We synthesized the copolymers having poly­(<i>N</i>-isopropylacrylamide) main chain and NBA side chain at 4, 7, and 10 mol % functionalizations and analyzed their photoresponsive characteristics. Light with 365 nm wavelength converted NBA groups at copolymer side chains to carboxylic acid efficiently at the rate of 7.3 cm²/J, and in the case of 10 mol % functionalization, the irradiation dosage no more than 56 mJ/cm² induced sharp aqueous dissolution of the copolymer thin layer in pH 7.4 at 25 °C. As example applications, we demonstrated on-demand release of polyethylene beads and fluorescent-labeled albumins, which had been immobilized on a substrate surface via the copolymers, by the precisely controlled light irradiation using a microprojection system. Also, we examined application of the copolymers to the selective recovery of living cells from culture substrate under microscopic observation. As a result, mild light irradiation at room temperature triggered immediate detachment of the cultured adherent cells only in the irradiated areas without critical influence on their viability

Photoresponsive Aqueous Dissolution of Poly(<i>N</i>‑Isopropylacrylamide) Functionalized with <i>o</i>‑Nitrobenzaldehyde through Phase Transition

We report a sharp photoinduced aqueous dissolution of the copolymer through phase transition based on the photochemical reaction of <i>o</i>-nitrobenzaldehyde (NBA) and the principle of polymer effect. We synthesized the copolymers having poly­(<i>N</i>-isopropylacrylamide) main chain and NBA side chain at 4, 7, and 10 mol % functionalizations and analyzed their photoresponsive characteristics. Light with 365 nm wavelength converted NBA groups at copolymer side chains to carboxylic acid efficiently at the rate of 7.3 cm²/J, and in the case of 10 mol % functionalization, the irradiation dosage no more than 56 mJ/cm² induced sharp aqueous dissolution of the copolymer thin layer in pH 7.4 at 25 °C. As example applications, we demonstrated on-demand release of polyethylene beads and fluorescent-labeled albumins, which had been immobilized on a substrate surface via the copolymers, by the precisely controlled light irradiation using a microprojection system. Also, we examined application of the copolymers to the selective recovery of living cells from culture substrate under microscopic observation. As a result, mild light irradiation at room temperature triggered immediate detachment of the cultured adherent cells only in the irradiated areas without critical influence on their viability

Study of Perfluoroalkyl Chain-Specific Band Shift in Infrared Spectra on the Chain Length

The CF₂ symmetric stretching vibration (ν_s(CF₂)) band of a perfluoroalkyl (Rf) group in an infrared (IR) spectrum exhibits a unique character, that is, an apparent high wavenumber shift with increasing the chain length, which is an opposite character to that of the CH stretching vibration band of a normal alkyl chain. To reveal the mechanism of the unusual IR band shift, two vibrational characters of an Rf chain are focused: (1) a helical conformation of an Rf chain, (2) the carbon (C) atoms having a smaller mass than the fluorine (F) atom dominantly vibrate as a coupled oscillator leaving F atoms stay relatively unmoved. These indicate that a “coupled oscillation of the skeletal C atoms” of an Rf chain should be investigated considering the helical structure. In the present study, therefore, the coupled oscillation of the Rf chain dependent on the chain length is investigated by Raman spectroscopy, which is suitable for investigating a skeletal vibration. The Raman-active ν_s(CF₂) band is found to be split into two bands, the splitting is readily explained by considering the helical structure and length with respect to group theory, and the unusual peak shift is concluded to be explained by the helical length

Morphology-based optical separation of subpopulations from a heterogeneous murine breast cancer cell line

<div><p>Understanding tumor heterogeneity is an urgent and unmet need in cancer research. In this study, we used a morphology-based optical cell separation process to classify a heterogeneous cancer cell population into characteristic subpopulations. To classify the cell subpopulations, we assessed their morphology in hydrogel, a three-dimensional culture environment that induces morphological changes according to the characteristics of the cells (i.e., growth, migration, and invasion). We encapsulated the murine breast cancer cell line 4T1E, as a heterogeneous population that includes highly metastatic cells, in click-crosslinkable and photodegradable gelatin hydrogels, which we developed previously. We observed morphological changes within 3 days of encapsulating the cells in the hydrogel. We separated the 4T1E cell population into colony- and granular-type cells by optical separation, in which local UV-induced degradation of the photodegradable hydrogel around the target cells enabled us to collect those cells. The obtained colony- and granular-type cells were evaluated <i>in vitro</i> by using a spheroid assay and <i>in vivo</i> by means of a tumor growth and metastasis assay. The spheroid assay showed that the colony-type cells formed compact spheroids in 2 days, whereas the granular-type cells did not form spheroids. The tumor growth assay in mice revealed that the granular-type cells exhibited lower tumor growth and a different metastasis behavior compared with the colony-type cells. These results suggest that morphology-based optical cell separation is a useful technique to classify a heterogeneous cancer cell population according to its cellular characteristics.</p></div

Tumor growth in mice transplanted with 4T1E cells, and colony- (C1 and C2) and granular-type (G1 and G2) cells separated from 4T1E cells.

<p>(A) Tumor growth. (B) Body weight of mice. The table below the figure shows the statistical profile as analyzed by using Scheffe’s test for tumor growth. Asterisks in the table indicate statistical significance. One asterisk indicates <i>p</i>-value less than 0.05 and double asterisks indicate <i>p</i>-value less than 0.01. Error bars indicate the standard deviation (n = 3–4).</p

Lateral Diffusion and Molecular Interaction in a Bilayer Membrane Consisting of Partially Fluorinated Phospholipids

Fluorinated lipids and surfactants are attractive biomimetic materials for the extraction and reorganization of membrane proteins because of the biological inertness of fluorocarbons. We investigated the fundamental physical properties of a partially fluorinated phospholipid (F4-DMPC), such as phase transition, area thermal expansion, and lateral lipid diffusion, to evaluate the intermolecular interaction of F4-DMPC in the hydrophobic region quantitatively on the basis of free-volume theory. Fluorescence microscope observation of the supported lipid bilayer (SLB) of F4-DMPC showed that the phase transition between the liquid crystalline and gel phases occurred at 5 °C and that the area thermal expansion coefficient was independent of the temperature near the phase transition temperature. We performed a single particle tracking of the F4-DMPC-SLB on a SiO₂/Si substrate, to measure the diffusion coefficient and its temperature dependence. The apparent activation energy (<i>E</i>′_a) of lateral lipid diffusion, which is an indicator of intermolecular interaction, was 39.1 kJ/mol for F4-DMPC, and 48.2 kJ/mol for a nonfluorinated 1,2-dioleoyl-<i>sn</i>-glycero-3-phosphocholine as a control. The difference of 9 kJ/mol in <i>E</i>′_a was significant compared with the difference due to the acyl chain species among nonfluorinated phosphatidylcholine and also that caused by the addition of cholesterol and alcohol in the bilayer membranes. We quantitatively evaluated the attenuation of intermolecular interaction, which results from the competition between the dipole-induced packing effect and steric effect at the fluorocarbon segment in F4-DMPC

An overview of the optical cell separation procedure.

<p>(A) Cell encapsulation in the PD-gelatin hydrogel. 4T1E cells were encapsulated in photodegradable hydrogels prepared by mixing azide-gelatin solution and DBCO-PC-4armPEG solution. (B) Optical cell separation from the PD-gelatin hydrogel. The target cells are irradiated with UV light and subsequently retrieved after hydrogel degradation. (C) Microscopic images of 4T1E cells, and colony- and granular-type cells to be separated by use of the optical cell separation process.</p

Frequency of metastasis in tissues obtained 38 days post-transplantation of 4T1E cells, and colony- (C1 and C2) and granular-type (G1 and G2) cells separated from 4T1E cells.

<p>Arrows indicate metastatic tumors on the surface of the tissues. The table shows the frequency of metastases confirmed by observation of the dissected tissues. The number of tumors detected in lung, liver, colon, kidney, and intestine is indicated as none (-), one (+), one to five (++), and more than five (+++). The upward-pointing arrows in the table indicate spleen enlargement.</p

Microscopic images of 4T1E cells, and colony- and granular-type cells separated from 4T1E cells.

<p>The separated cells were established as colony- (C1 and C2) and granular-type (G1 and G2) cells by optical separation based on morphology in PD-gelatin hydrogels. 2D, cell culture on a culture dish; 3D, cell culture in the PD-gelatin hydrogels. The images were taken after 3 days of culture under 2D and 3D conditions. (A) Raw images. The scale bar indicates 500 μm. (B) Digitally magnified images. The scale bar indicates 125 μm.</p