Light-driven liquid metal nanotransformers for biomedical theranostics.

Room temperature liquid metals (LMs) represent a class of emerging multifunctional materials with attractive novel properties. Here, we show that photopolymerized LMs present a unique nanoscale capsule structure characterized by high water dispersibility and low toxicity. We also demonstrate that the LM nanocapsule generates heat and reactive oxygen species under biologically neutral near-infrared (NIR) laser irradiation. Concomitantly, NIR laser exposure induces a transformation in LM shape, destruction of the nanocapsules, contactless controlled release of the loaded drugs, optical manipulations of a microfluidic blood vessel model and spatiotemporal targeted marking for X-ray-enhanced imaging in biological organs and a living mouse. By exploiting the physicochemical properties of LMs, we achieve effective cancer cell elimination and control of intercellular calcium ion flux. In addition, LMs display a photoacoustic effect in living animals during NIR laser treatment, making this system a powerful tool for bioimaging.

Photothermal and mechanical stimulation of cells via dualfunctional nanohybrids.

Stimulating cells by light is an attractive technology to investigate cellular function and deliver innovative cell-based therapy. However, current techniques generally use poorly biopermeable light, which prevents broad applicability. Here, we show that a new type of composite nanomaterial, synthesized from multi-walled carbon nanotubes, magnetic iron nanoparticles, and polyglycerol, enables photothermal and mechanical control of Ca2+ influx into cells overexpressing transient receptor potential vanilloid type-2. The nanohybrid is simply operated by application of highly biotransparent near-infrared light and a magnetic field. The technology may revolutionize remote control of cellular function.

Semiconducting Polymer Nanobioconjugates for Targeted Photothermal Activation of Neurons.

Optogenetics provides powerful means for precise control of neuronal activity; however, the requirement of transgenesis and the incapability to extend the neuron excitation window into the deep-tissue-penetrating near-infrared (NIR) region partially limit its application. We herein report a potential alternative approach to optogenetics using semiconducting polymer nanobioconjugates (SPNsbc) as the photothermal nanomodulator to control the thermosensitive ion channels in neurons. SPNsbc are designed to efficiently absorb the NIR light at 808 nm and have a photothermal conversion efficiency higher than that of gold nanorods. By virtue of the fast heating capability in conjunction with the precise targeting to the thermosensitive ion channel, SPNsbc can specifically and rapidly activate the intracellular Ca(2+) influx of neuronal cells in a reversible and safe manner. Our study provides an organic nanoparticle based strategy that eliminates the need for genetic transfection to remotely regulate cellular machinery.

Magnetically and Near-Infrared Light-Powered Supramolecular Nanotransporters for the Remote Control of Enzymatic Reactions.

Cancer is one of the primary causes of death worldwide. A high-precision analysis of biomolecular behaviors in cancer cells at the single-cell level and more effective cancer therapies are urgently required. Here, we describe the development of a magnetically- and near infrared light-triggered optical control method, based on nanorobotics, for the analyses of cellular functions. A new type of nanotransporters, composed of magnetic iron nanoparticles, carbon nanohorns, and liposomes, was synthesized for the spatiotemporal control of cellular functions in cells and mice. Our technology will help to create a new state-of-the-art tool for the comprehensive analysis of "real" biological molecular information at the single-cell level, and it may also help in the development of innovative cancer therapies.

Lipid Nanotube Tailored Fabrication of Uniquely Shaped Polydopamine Nanofibers as Photothermal Converters.

Helically coiled and linear polydopamine (PDA) nanofibers were selectively fabricated with two different types of lipid nanotubes (LNTs) that acted as templates. The obtained coiled PDA-LNT hybrid showed morphological advantages such as higher light absorbance and photothermal conversion effect compared to a linear counterpart. Laser irradiation of the coiled PDA-LNT hybrid induced a morphological change and subsequent release of the encapsulated guest molecule. In cellular experiments, the coiled PDA-LNT efficiently eliminated HeLa cells because of its strong affinity with the tumor cells. This work illustrates the first approach to construct characteristic morphologies of PDA nanofibers using LNTs as simple templates, and the coiled PDA-LNT hybrid exhibits attractive photothermal features derived from its unique coiled shape.

In Vivo Remote Control of Reactions in Caenorhabditis elegans by Using Supramolecular Nanohybrids of Carbon Nanotubes and Liposomes.

A supramolecular nanohybrid based on carbon nanotubes and liposomes that is highly biocompatible and capable of permeation through cells is described. The nanohybrid can be loaded with a variety of functional molecules and is structurally controlled by near-infrared laser irradiation for the release of molecules from the nanohybrids in a targeted manner via microscopy. We implemented the controlled release of molecules from the nanohybrids and demonstrated remote regulation of the photoinduced nanohybrid functions. As a proof of principle, nanohybrids loaded with amiloride were successfully used in the spatiotemporally targeted blocking of amiloride-sensitive mechanosensory neurons in living Caenorhabditis elegans. Our prototype could inspire new designs for biomimetic parasitism and symbiosis, and biologically active nanorobots for the higher-level manipulation of organisms.

Multifunctional carbon nanohorn complexes for cancer treatment.

Multifunctional carbon nanohorn (CNH) complexes were synthesized using oxidized CNH, magnetite (MAG) nanoparticles, and polyethyleneimine (PEI). The ferromagnetic MAG nanoparticles were loaded onto CNH (MAG-CNH) using iron(II) acetate and subsequent heat treatment. Chemical functionalization of the MAG-CNH complexes with PEI improved their water-dispersibility and allowed further conjugation with a fluorophore. The application of an external magnetic field significantly intensified the targeted vectorization of CNH complexes into human cervical cancer (HeLa) cells. Following cell uptake, laser irradiation of the cells showed a significant enhancement in the photothermal effects of CNHs leading to cell death. We have confirmed that the photothermal properties and ferromagnetic characteristics of CNH complexes show efficient cell elimination. The present study is an essential step toward the development of an innovative cancer therapy and a highly sensitive detection of cancer cells at the single-cell level.

Transcriptome profiling of Lotus japonicus roots during arbuscular mycorrhiza development and comparison with that of nodulation.

To better understand the molecular responses of plants to arbuscular mycorrhizal (AM) fungi, we analyzed the differential gene expression patterns of Lotus japonicus, a model legume, with the aid of a large-scale cDNA macroarray. Experiments were carried out considering the effects of contaminating microorganisms in the soil inoculants. When the colonization by AM fungi, i.e. Glomus mosseae and Gigaspora margarita, was well established, four cysteine protease genes were induced. In situ hybridization revealed that these cysteine protease genes were specifically expressed in arbuscule-containing inner cortical cells of AM roots. On the other hand, phenylpropanoid biosynthesis-related genes for phenylalanine ammonia-lyase (PAL), chalcone synthase, etc. were repressed in the later stage, although they were moderately up-regulated on the initial association with the AM fungus. Real-time RT-PCR experiments supported the array experiments. To further confirm the characteristic expression, a PAL promoter was fused with a reporter gene and introduced into L. japonicus, and then the transformants were grown with a commercial inoculum of G. mosseae. The reporter activity was augmented throughout the roots due to the presence of contaminating microorganisms in the inoculum. Interestingly, G. mosseae only colonized where the reporter activity was low. Comparison of the transcriptome profiles of AM roots and nitrogen-fixing root nodules formed with Mesorhizobium loti indicated that the PAL genes and other phenylpropanoid biosynthesis-related genes were similarly repressed in the two organs.

Knockdown of an arbuscular mycorrhiza-inducible phosphate transporter gene of Lotus japonicus suppresses mutualistic symbiosis.

cDNA for a major arbuscular mycorrhiza (AM)-inducible phosphate (Pi) transporter of Lotus japonicus, LjPT3, was isolated from Glomus mosseae-colonized roots. The LjPT3 transcript was expressed in arbuscule-containing cells of the inner cortex. The transport activity of the gene product was confirmed by the complementation of a yeast mutant that lacks high-affinity Pi transporters. In contrast to most AM-inducible Pi transporters thus far reported, LjPT3 has an amino acid sequence that has much in common with those of other members of the Pht1 family of plant Pi transporters, such as StPT3 of potato. To understand better the physiological role of this AM-inducible Pi transporter, knockdown transformants of the gene were prepared through hairy root transformation and RNA interference. Under Pi-limiting conditions, the transformants showed a reduction of Pi uptake via AM and growth retardation. The transformants also exhibited a decrease in G. mosseae arbuscules. Additionally, when Mesorhizobium loti was inoculated into the knockdown transformants in combination with G. mosseae, necrotic root nodules were observed. Based on these findings, we consider that the genetically engineered host plants had monitored insufficient Pi uptake via AM or low expression of LjPT3, excluding the existing fungi and rhizobia and/or preventing further development of the fungal and nodule structures.

Cloning and expression analysis of a MAPKKK gene and a novel nodulin gene of Lotus japonicus.

We isolated a cDNA encoding mitogen-activated protein kinase kinase kinase alpha, designated LjM3Kalpha, from Lotus japonicus, a model legume. The gene was expressed constitutively in roots, root nodules, and shoots. We also identified a novel nodulin gene, LjNUF, that shows specific expression in nodules. LjNUF resembles the C-terminal half of a hypothetical protein (pir//D85436), the N-terminal half of which is similar to a portion of mitogen-activated protein kinase kinase kinase gamma. Although LjNUF was predicted to be a secreted protein, its function remains to be clarified.

cDNA cloning, mRNA expression, and mutational analysis of the squalene synthase gene of Lotus japonicus.

A full-length cDNA for squalene synthase was isolated from Lotus japonicus, a model leguminous plant. The transcript was abundant in roots, symbiotic root nodules, and shoots, in that order. In situ hybridization revealed that the mRNA level is high in expanding root cells but low in dividing root tip ones. The transcript is also abundant in vascular bundles and the basal portions of mature nodules. L. japonicus squalene synthase has an unusual Asp residue near the active site, where mammalian enzymes have Gln, and replacement of the Gln by Glu has been reported to cause severe inactivation. Site-directed mutagenesis of the L. japonicus enzyme and assaying in vitro showed that this Asp residue can be substituted by not only Gln but also Glu, suggesting that the local structure of plant squalene synthases is different from that of mammalian enzymes.