Development of the multiplex imaging chamber at PAL-XFEL.

Various X-ray techniques are employed to investigate specimens in diverse fields. Generally, scattering and absorption/emission processes occur due to the interaction of X-rays with matter. The output signals from these processes contain structural information and the electronic structure of specimens, respectively. The combination of complementary X-ray techniques improves the understanding of complex systems holistically. In this context, we introduce a multiplex imaging instrument that can collect small-/wide-angle X-ray diffraction and X-ray emission spectra simultaneously to investigate morphological information with nanoscale resolution, crystal arrangement at the atomic scale and the electronic structure of specimens.

Resonant X-ray emission spectroscopy using self-seeded hard X-ray pulses at PAL-XFEL.

Self-seeded hard X-ray pulses at PAL-XFEL were used to commission a resonant X-ray emission spectroscopy experiment with a von Hamos spectrometer. The self-seeded beam, generated through forward Bragg diffraction of the [202] peak in a 100 µm-thick diamond crystal, exhibited an average bandwidth of 0.54 eV at 11.223 keV. A coordinated scanning scheme of electron bunch energy, diamond crystal angle and silicon monochromator allowed us to map the Ir Lß2 X-ray emission lines of IrO2 powder across the Ir L3-absorption edge, from 11.212 to 11.242 keV with an energy step of 0.3 eV. This work provides a reference for hard X-ray emission spectroscopy experiments utilizing self-seeded pulses with a narrow bandwidth, eventually applicable for pump-probe studies in solid-state and diluted systems.

Ultrafast Carrier-Lattice Interactions and Interlayer Modulations of Bi2Se3 by X-ray Free-Electron Laser Diffraction.

As a 3D topological insulator, bismuth selenide (Bi2Se3) has potential applications for electrically and optically controllable magnetic and optoelectronic devices. Understanding the coupling with its topological phase requires studying the interactions of carriers with the lattice on time scales down to the subpicosecond regime. Here, we investigate the ultrafast carrier-induced lattice contractions and interlayer modulations in Bi2Se3 thin films by time-resolved diffraction using an X-ray free-electron laser. The lattice contraction depends on the carrier concentration and is followed by an interlayer expansion accompanied by oscillations. Using density functional theory and the Lifshitz model, the initial contraction can be explained by van der Waals force modulation of the confined free carrier layers. Our theoretical calculations suggest that the band inversion, related to a topological phase transition, is modulated by the expansion of the interlayer distance. These results provide insights into the topological phase control by light-induced structural change on ultrafast time scales.

Electric fields regulate cellular elasticity through intracellular Ca2+ concentrations.

Cellular elasticity is a key factor related to a broad range of physiological and pathological processes. The elasticity of a single cell has thus emerged as a potential biomarker to characterize the cellular state. Both internal and external stimuli affect cellular elasticity, and changes in elasticity can cause alterations in cellular characteristics or function. The application of electric fields (EFs) is a promising method that can be used to change cellular elasticity; however, the mechanisms underlying its effect remain unknown. Here, we demonstrate EFs-induced elasticity changes in human dermal fibroblasts and discuss the underlying mechanism related to actin polymerization. Cellular elasticity increases after EF (50 mV/mm) stimulation, reaching a maximum at 30 min before decreasing between 30 and 120 min. The cellular elasticity under EF stimulation, regardless of stimulation time, is higher than that of the control. F-actin regulates the elasticity of cells through gelsolin activation. We show changes in intracellular Ca2+ caused by EFs, which induced gelsolin activation and F-actin content changes. This result demonstrates a series of processes in which external electrical stimulation conditions regulate cellular elasticity.

Challenges of applying multicellular tumor spheroids in preclinical phase.

The three-dimensional (3D) multicellular tumor spheroids (MCTs) model is becoming an essential tool in cancer research as it expresses an intermediate complexity between 2D monolayer models and in vivo solid tumors. MCTs closely resemble in vivo solid tumors in many aspects, such as the heterogeneous architecture, internal gradients of signaling factors, nutrients, and oxygenation. MCTs have growth kinetics similar to those of in vivo tumors, and the cells in spheroid mimic the physical interaction of the tumors, such as cell-to-cell and cell-to-extracellular matrix interactions. These similarities provide great potential for studying the biological properties of tumors and a promising platform for drug screening and therapeutic efficacy evaluation. However, MCTs are not well adopted as preclinical tools for studying tumor behavior and therapeutic efficacy up to now. In this review, we addressed the challenges with MCTs application and discussed various efforts to overcome the challenges.

Qualitative analysis of contribution of intracellular skeletal changes to cellular elasticity.

Cells are dynamic structures that continually generate and sustain mechanical forces within their environments. Cells respond to mechanical forces by changing their shape, moving, and differentiating. These reactions are caused by intracellular skeletal changes, which induce changes in cellular mechanical properties such as stiffness, elasticity, viscoelasticity, and adhesiveness. Interdisciplinary research combining molecular biology with physics and mechanical engineering has been conducted to characterize cellular mechanical properties and understand the fundamental mechanisms of mechanotransduction. In this review, we focus on the role of cytoskeletal proteins in cellular mechanics. The specific role of each cytoskeletal protein, including actin, intermediate filaments, and microtubules, on cellular elasticity is summarized along with the effects of interactions between the fibers.

High-Resolution XFEL Structure of the Soluble Methane Monooxygenase Hydroxylase Complex with its Regulatory Component at Ambient Temperature in Two Oxidation States.

Soluble methane monooxygenase (sMMO) is a multicomponent metalloenzyme that catalyzes the conversion of methane to methanol at ambient temperature using a nonheme, oxygen-bridged dinuclear iron cluster in the active site. Structural changes in the hydroxylase component (sMMOH) containing the diiron cluster caused by complex formation with a regulatory component (MMOB) and by iron reduction are important for the regulation of O2 activation and substrate hydroxylation. Structural studies of metalloenzymes using traditional synchrotron-based X-ray crystallography are often complicated by partial X-ray-induced photoreduction of the metal center, thereby obviating determination of the structure of the enzyme in pure oxidation states. Here, microcrystals of the sMMOH:MMOB complex from Methylosinus trichosporium OB3b were serially exposed to X-ray free electron laser (XFEL) pulses, where the ≤35 fs duration of exposure of an individual crystal yields diffraction data before photoreduction-induced structural changes can manifest. Merging diffraction patterns obtained from thousands of crystals generates radiation damage-free, 1.95 Å resolution crystal structures for the fully oxidized and fully reduced states of the sMMOH:MMOB complex for the first time. The results provide new insight into the manner by which the diiron cluster and the active site environment are reorganized by the regulatory protein component in order to enhance the steps of oxygen activation and methane oxidation. This study also emphasizes the value of XFEL and serial femtosecond crystallography (SFX) methods for investigating the structures of metalloenzymes with radiation sensitive metal active sites.

Characterizing the intrinsic properties of individual XFEL pulses via single-particle diffraction.

With each single X-ray pulse having its own characteristics, understanding the individual property of each X-ray free-electron laser (XFEL) pulse is essential for its applications in probing and manipulating specimens as well as in diagnosing the lasing performance. Intensive research using XFEL radiation over the last several years has introduced techniques to characterize the femtosecond XFEL pulses, but a simple characterization scheme, while not requiring ad hoc assumptions, to address multiple aspects of XFEL radiation via a single data collection process is scant. Here, it is shown that single-particle diffraction patterns collected using single XFEL pulses can provide information about the incident photon flux and coherence property simultaneously, and the X-ray beam profile is inferred. The proposed scheme is highly adaptable to most experimental configurations, and will become an essential approach to understanding single X-ray pulses.

Biomarkers to quantify cell migration characteristics.

BACKGROUND: Because cell movement is primarily driven by the connection between F-actin and integrin through a physical linkage, cellular elasticity and adhesion strength have been considered as biomarkers of cell motility. However, a consistent set of biomarkers that indicate the potential for cell motility is still lacking. METHODS: In this work, we characterize a phenotype of cell migration in terms of cellular elasticity and adhesion strength, which reveals the interdependence of subcellular systems that mediate optimal cell migration. RESULTS: Stiff cells weakly adhered to the substrate revealed superior motility, while soft cell migration with strong adhesion was relatively inhibited. The spatial distribution and amount of F-actin and integrin were highly variable depending on cell type, but their density exhibited linear correlations with cellular elasticity and adhesion strength, respectively. CONCLUSIONS: The densities of F-actin and integrin exhibited linear correlations with cellular elasticity and adhesion strength, respectively, therefore, they can be considered as biomarkers to quantify cell migration characteristics.

Electric field-induced changes in biomechanical properties in human dermal fibroblasts and a human skin equivalent.

PURPOSE: An electric field (EF) can be used to change the mechanical properties of cells and skin tissues. We demonstrate EF-induced elasticity changes in human dermal fibroblasts (HDFs) and a human skin equivalent and identify the underlying principles related to the changes. METHODS: HDFs and human skin equivalent were stimulated with electric fields of 1.0 V/cm. Change in cellular elasticity was determined by using atomic force microscopy. Effects of EF on the biomechanical and chemical properties of a human skin equivalent were analyzed. In cells and tissues, the effects of EF on biomarkers of cellular elasticity were investigated at the gene and protein levels. RESULTS: In HDFs, the cellular elasticity was increased and the expression of biomarkers of cellular elasticity was regulated by the EF. Expression of the collagen protein in the human skin equivalent was changed by EF stimulation; however, changes in density and microstructure of the collagen fibrils were not significant. The viscoelasticity of the human skin equivalent increased in response to EF stimulation, but molecular changes were not observed in collagen. CONCLUSIONS: Elasticity of cells and human skin equivalent can be regulated by electrical stimulation. Especially, the change in cellular elasticity was dependent on cell age.

Surface Roughness on the Slots and Wings of Various Ceramic Self-Ligating Brackets and their Potential Concern on Biofilm Formation.

OBJECTIVE: The surface roughness of various orthodontic materials could affect biofilm formation and friction. The purpose of this study was to examine the surface roughness and chemical composition of the slots and wings of several ceramic self-ligating brackets. STUDY DESIGN: Four types of ceramic self-ligating brackets were separated into experimental groups (DC, EC, IC, and QK) while a metal self-ligating bracket (EM) was used as the control group. Atomic force microscopy and energy-dispersive x-ray spectroscope were used to examine the surface roughness and chemical composition of each bracket slot and wing. RESULTS: The control group was made of ferrum and chrome while all the experimental groups were comprised of aluminum and oxide. There was a statistically significant difference in the roughness average (Sa) among the various self-ligating brackets (p< 0.001 in slots and p<0.01 in the wing). The slots in the EC group had the lowest Sa, followed by the DC, IC, control, and QK groups. The wings in the IC group had the lowest Sa, followed by the EC, DC, control, and QK groups. CONCLUSIONS: There is a significant difference in the surface roughness of the slots and wings among several types of ceramic self-ligating brackets.

Biomechanical properties of red blood cells infected by Plasmodium berghei ANKA.

Malaria is a pathogenic disease in mammal species and typically causes destruction of red blood cells (RBCs). The malaria-infected RBCs undergoes alterations in morphology and its rheological properties, and the altered rheological properties of RBCs have a significant impact on disease pathophysiology. In this study, we investigated detailed topological and biomechanical properties of RBCs infected with malaria Plasmodium berghei ANKA using atomic force microscopy. Mouse (BALB/c) RBCs were obtained on Days 4, 10, and 14 after infection. We found that malaria-infected RBCs changed significantly in shape. The RBCs maintained a biconcave disk shape until Day 4 after infection and then became lopsided on Day 7 after infection. The central region of RBCs began to swell beginning on Day 10 after infection. More schizont stages were present on Days 10 and 14 compared with on Day 4. The malaria-infected RBCs also showed changes in mechanical properties and the cytoskeleton. The stiffness of infected RBCs increased 4.4-4.6-fold and their cytoskeletal F-actin level increased 18.99-67.85% compared with the control cells. The increase in F-actin depending on infection time was in good agreement with the increased stiffness of infected RBCs. Because more schizont stages were found at a late period of infection at Days 10 and 14, the significant changes in biomechanical properties might contribute to the destruction of RBCs, possibly resulting in the release of merozoites into the blood circulation.

Identification of a closed cutaneous injury after mechanical trauma caused by collision.

PURPOSE: Robotics has evolved rapidly in terms of mechanical design and control in the past few years. Collaborative robots that have direct contact with humans are being introduced in various fields, including industrial and medical services. Because collaborative robot systems are being introduced rapidly, the safety of the humans who work with them is becoming an important issue. In this study, we investigated skin injuries resulting from a collision between robots and humans using a freefall experiment system. METHODS: We particularly focused on closed skin injuries caused by a collision. To induce a closed injury, we struck mini-pigs with cubic-edge square and semi-sphere impactors at collision speeds of 1 and 3 m/s. We did not observe any open injuries with those conditions. Closed injuries were observed in the dermal layer of the skin after the collision test at both speeds and with both impactors. RESULTS: The collagen fiber in the dermal layer was separated and fragmented, and the subcutaneous fat layer became dense as a result of the collision. CONCLUSIONS: We closely observed and analyzed the histopathologic changes in the dermal and subcutaneous layers with intact epidermis after mechanical trauma to the inner skin layers.

Gypenosides ameliorate memory deficits in MPTP-lesioned mouse model of Parkinson's disease treated with L-DOPA.

BACKGROUND: Previous studies have revealed that gypenosides (GPS) improve the symptoms of anxiety disorders in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned rat model of Parkinson's disease (PD). The present study aimed to investigate the effects of GPS on memory deficits in an MPTP-lesioned mouse model of PD treated with L-3,4-dihydroxyphenylalanine (L-DOPA). METHODS: MPTP (30 mg/kg/day, 5 days)-lesioned mice were treated with GPS (50 mg/kg) and/or L-DOPA (10 and 25 mg/kg) for 21 days. After the final treatments, behavioral changes were assessed in all mice using passive avoidance and elevated plus-maze tests. We then evaluated the biochemical influences of GPS treatment on levels of tyrosine hydroxylase (TH), dopamine, N-methyl-D-aspartate (NMDA) receptors, extracellular signal-regulated kinase (ERK1/2), and cyclic AMP-response element binding protein (CREB) phosphorylation. RESULTS: MPTP-lesioned mice exhibited deficits associated with habit learning and spatial memory, which were further aggravated by treatment with L-DOPA (25 mg/kg). However, treatment with GPS (50 mg/kg) ameliorated memory deficits. Treatment with GPS (50 mg/kg) also improved L-DOPA (25 mg/kg)-treated MPTP lesion-induced decreases in retention latency on the passive avoidance test, as well as levels of TH-immunopositive cells and dopamine in the substantia nigra and striatum. GPS treatment also attenuated increases in retention transfer latency on the elevated plus-maze test and in NMDA receptor expression, as well as decreases in the phosphorylation of ERK1/2 and CREB in the hippocampus. Treatment with L-DOPA (10 mg/kg) also ameliorated deficits in habit learning and spatial memory in MPTP-lesioned mice, and this effect was further enhanced by treatment with GPS (50 mg/kg). CONCLUSION: GPS ameliorate deficits in habit learning and spatial memory by modulating the dopaminergic neuronal and N-methyl-D-aspartate receptor-mediated signaling systems in MPTP-lesioned mice treated with L-DOPA. GPS may serve as an adjuvant therapeutic agent for memory deficits in patients with PD receiving L-DOPA.

Intrinsic and extrinsic mechanical properties related to the differentiation of mesenchymal stem cells.

Diverse intrinsic and extrinsic mechanical factors have a strong influence on the regulation of stem cell fate. In this work, we examined recent literature on the effects of mechanical environments on stem cells, especially on differentiation of mesenchymal stem cells (MSCs). We provide a brief review of intrinsic mechanical properties of single MSC and examined the correlation between the intrinsic mechanical property of MSC and the differentiation ability. The effects of extrinsic mechanical factors relevant to the differentiation of MSCs were considered separately. The effect of nanostructure and elasticity of the matrix on the differentiation of MSCs were summarized. Finally, we consider how the extrinsic mechanical properties transfer to MSCs and then how the effects on the intrinsic mechanical properties affect stem cell differentiation.

Selective cytotoxic effect of non-thermal micro-DBD plasma.

Non-thermal plasma has been extensively researched as a new cancer treatment technology. We investigated the selective cytotoxic effects of non-thermal micro-dielectric barrier discharge (micro-DBD) plasma in cervical cancer cells. Two human cervical cancer cell lines (HeLa and SiHa) and one human fibroblast (HFB) cell line were treated with micro-DBD plasma. All cells underwent apoptotic death induced by plasma in a dose-dependent manner. The plasma showed selective inhibition of cell proliferation in cervical cancer cells compared to HFBs. The selective effects of the plasma were also observed between the different cervical cancer cell lines. Plasma treatment significantly inhibited the proliferation of SiHa cells in comparison to HeLa cells. The changes in gene expression were significant in the cervical cancer cells in comparison to HFBs. Among the cancer cells, apoptosis-related genes were significantly enriched in SiHa cells. These changes were consistent with the differential cytotoxic effects observed in different cell lines.

Medical applications of the intrinsic mechanical properties of single cells.

The mechanical properties of single cells have been recently identified as the basis of an emerging approach in medical applications because they are closely related to the biological processes of cells and, ultimately, human health conditions. In this article, we provide a brief review of the intrinsic mechanical properties of single cells related to cancer and aging. The mechanical properties can be used as biomarkers for early cancer diagnosis because cancer cells have a lower Young's modulus, indicating higher elasticity or softness than their counterpart normal cells. The metastatic potential of cancer cells is inversely correlated with their elastic properties. Aging induces stiffness through an increased amount of cytoskeletal fiber. Changes in the mechanical properties also show potential for drug screening. Although there are several challenges to be met before clinical applications can be made, such mechanical properties of single cells may provide new approaches to human diseases.

Medical applications of atomic force microscopy and Raman spectroscopy.

This paper reviews the recent research and application of atomic force microscopy (AFM) and Raman spectroscopy techniques, which are considered the multi-functional and powerful toolkits for probing the nanostructural, biomechanical and physicochemical properties of biomedical samples in medical science. We introduce briefly the basic principles of AFM and Raman spectroscopy, followed by diagnostic assessments of some selected diseases in biomedical applications using them, including mitochondria isolated from normal and ischemic hearts, hair fibers, individual cells, and human cortical bone. Finally, AFM and Raman spectroscopy applications to investigate the effects of pharmacotherapy, surgery, and medical device therapy in various medicines from cells to soft and hard tissues are discussed, including pharmacotherapy--paclitaxel on Ishikawa and HeLa cells, telmisartan on angiotensin II, mitomycin C on strabismus surgery and eye whitening surgery, and fluoride on primary teeth--and medical device therapy--collagen cross-linking treatment for the management of progressive keratoconus, radiofrequency treatment for skin rejuvenation, physical extracorporeal shockwave therapy for healing of Achilles tendinitis, orthodontic treatment, and toothbrushing time to minimize the loss of teeth after exposure to acidic drinks.

Hierarchical channel morphology in O-rings after two cycling exposures to 70 MPa hydrogen gas: a case study of sealing failure.

This study investigates the impact of high-pressure hydrogen gas exposure on the structural and morphological characteristics of O-ring materials. O-ring specimens undergo two cycles of sealing under 70 MPa hydrogen gas, and their resulting variations are examined using advanced characterization techniques, including powder X-ray diffraction (PXRD), small-angle X-ray scattering (SAXS), scanning electron microscopy (SEM) and atomic force microscopy (AFM). Our findings reveal that the lattice parameters of the O-ring material show no significant changes when exposed to 70 MPa hydrogen gas. However, in the micrometre range, the formation of a hierarchical channel morphology becomes evident. This morphology is accompanied by the separation of carbon black filler from the rubber matrix, contributing to mechanical weakening of the O-ring. These observations can be attributed to the pressure gradient that develops between the inner and outer radii of the O-ring, resulting from compression forces acting perpendicularly to the radial direction due to clamp locking.

Analysis of aging effects on chemical property of human hair by Fourier transform infrared spectroscopy.

BACKGROUND: In the previous work, we investigated the aging effect on morphology and mechanical property of the hair by using atomic force microscopy. OBJECTIVES: The effects of aging on chemical properties of human hair were investigated by Fourier transform infrared (FT-IR) spectroscopy. METHODS: Healthy hair samples with no diseases were collected from 60 Koreans (30 males and 30 females) and they were grouped by age: 1-10, 11-20, 21-30, 31-40, 41-50, and 51-60 years. RESULTS: The characteristic parameters of FT-IR absorbance bands including center frequency, half width, height, and area were analyzed using the Gaussian model. To quantitatively analyze the chemical composition of hair, the height and area of all bands in the spectra were normalized to the amide I centered at 1652-1659 and 1654-1658 cm(-1), for male and female hairs, respectively. In all male and female hairs, the spectra of specific components of the hair keratin showed to have the same dependence on aging. The center positions of the bands arising from amide A, CH(3) mode, and amide I were altered by aging. The female hair contains more cystein than the male hair. Changes in the amount of amide II and amide A by aging were more significant in male hair than in female hair. CONCLUSIONS: The changes in chemical components of the hair according to the ages were shown at the inflection point at 30 s.