Inequality in Older Volunteering: Association Between Volunteer Competency and Demographic Profiles.

The present survey research investigated older people's volunteering competency relating to social inequality by exploring the latent ability profile and demographic correlates of 1,000 older volunteers in 73 community care centersin southern Taiwan. Older volunteers were classified into advanced (n = 509), basic (n = 214), and novice (n = 277) groups. Demographics examined included: individualistic characteristics (religious beliefs), resources (education; number of chronic diseases), andsocial factors (serving area and spoken language, volunteering duration, marital status, and gender). Apparent inequality issues were revealed. The advanced group was better educated, Mandarin-speaking, and in urban areas. while the novice group featured the opposite (lower education Taiwanese-speaking suburban areas).

HC-HA/PTX3 from amniotic membrane reverts senescent limbal niche cells to Pax6+ neural crest progenitors to support limbal epithelial progenitors.

Quiescence and self-renewal of human corneal epithelial progenitor/stem cells (LEPC) are regulated by the limbal niche, presumably through close interaction with limbal (stromal) niche cells (LNC). Paired box homeotic gene 6 (Pax6), a conserved transcription factor essential for eye development, is essential for proper differentiation of limbal and corneal epithelial stem cells. Pax6 haploinsufficiency causes limbal stem cell deficiency, which leads to subsequent corneal blindness. We previously reported that serial passage of nuclear Pax6+ LNC resulted in the gradual loss of nuclear Pax6+ and neural crest progenitor status, the latter of which was reverted upon recovery of Pax6. These findings suggest Pax6 plays a pivotal role in supporting the self-renewal of LEPC in limbal niche. Herein, we show that HC-HA/PTX3, a unique matrix purified from amniotic membrane (AM) and consists of heavy chain 1of inter-α-trypsin inhibitor covalently linked to hyaluronic acid and complexed with pentraxin 3, is capable of reverting senescent LNC to nuclear Pax6+ neural crest progenitors that support self-renewal of LEPC. Such reversion is causally linked to early cell aggregation mediated by activation of C-X-C chemokine receptor type 4 (CXCR4)-mediated signaling followed by activation of bone morphogenetic protein (BMP) signaling. Furthermore, CXCR4-mediated signaling, but not BMP signaling, controls recovery of the nuclear Pax6+ neural crest progenitors. These findings not only explain why AM helps in vivo and ex vivo expansion of human LEPC, but they also illuminate the potential role of HC-HA/PTX3 as a surrogate matrix niche that complements stem cell-based therapies in regenerative medicine.

Converging experimental and computational views of the knotting mechanism of a small knotted protein.

MJ0366 from Methanocaldococcus jannaschii is the smallest topologically knotted protein known to date. 92 residues in length, MJ0366 ties a trefoil (31) knot by threading its C-terminal helix through a buttonhole formed by the remainder of the secondary structure elements. By generating a library of point mutations at positions pertinent to the knot formation, we systematically evaluated the contributions of individual residues to the folding stability and kinetics of MJ0366. The experimental Φ-values were used as restraints to computationally generate an ensemble of conformations that correspond to the transition state of MJ0366, which revealed several nonnative contacts. The importance of these nonnative contacts in stabilizing the transition state of MJ0366 was confirmed by a second round of mutagenesis, which also established the pivotal role of F15 in stapling the network of hydrophobic interactions around the threading C-terminal helix. Our converging experimental and computational results show that, despite the small size, the transition state of MJ0366 is formed at a very late stage of the folding reaction coordinate, following a polarized pathway. Eventually, the formation of extensive native contacts, as well as a number of nonnative ones, leads to the threading of the C-terminal helix that defines the topological knot.

Magnolol and Honokiol Inhibited the Function and Expression of BCRP with Mechanism Exploration.

Breast cancer resistance protein (BCRP), one of the ATP-binding cassette (ABC) transporters, was associated with the multidrug resistance (MDR) of chemotherapy. Magnolol (MN) and honokiol (HK) are major bioactive polyphenols of Magnolia officinalis. This study investigated the effects of MN and HK on the function and expression of BCRP for the purpose of developing BCRP inhibitor to overcome MDR. Cell lines including MDCKII-BCRP and MDCKII-WT were used for evaluating the function and expression of BCRP. The results showed that MN (100-12.5 µM) and HK (100-12.5 µM) significantly decreased the function of BCRP by 80~12% and 67~14%, respectively. In addition, MN and HK were verified as substrates of BCRP. Furthermore, MN and HK reduced the protein expression of BCRP, and inhibited the phosphorylation of epidermal growth factor receptor (EGFR) and phosphatidylinositol 3-kinase (PI3K). In conclusion, both MN and HK decreased the function and expression of BCRP via EGFR/PI3K signaling pathway. Therefore, both compounds were promising candidates for reversing the MDR of chemotherapy.

Materials Engineering of Violin Soundboards by Stradivari and Guarneri.

We investigated the material properties of Cremonese soundboards using a wide range of spectroscopic, microscopic, and chemical techniques. We found similar types of spruce in Cremonese soundboards as in modern instruments, but Cremonese spruces exhibit unnatural elemental compositions and oxidation patterns that suggest artificial manipulation. Combining analytical data and historical information, we may deduce the minerals being added and their potential functions-borax and metal sulfates for fungal suppression, table salt for moisture control, alum for molecular crosslinking, and potash or quicklime for alkaline treatment. The overall purpose may have been wood preservation or acoustic tuning. Hemicellulose fragmentation and altered cellulose nanostructures are observed in heavily treated Stradivari specimens, which show diminished second-harmonic generation signals. Guarneri's practice of crosslinking wood fibers via aluminum coordination may also affect mechanical and acoustic properties. Our data suggest that old masters undertook materials engineering experiments to produce soundboards with unique properties.

Two-photon fluorescence and second harmonic generation hyperspectral imaging of old and modern spruce woods.

Spruce is the commonly-used tonewood for the top plate of violin-family instruments, such as violins and cellos. The wood properties can critically determine the acoustic quality. It's been shown the wood of famous old instruments differ from modern ones due to chemical treatment and aging. To reveal the differences microscopically in both spatial and spectral domains, a two-photon hyperspectral system has been applied to investigate the autofluorescence and second harmonic generation within wood samples. Not only the cellular structures were observed through optical sectioning, but the spectral variations were revealed among different age wood samples and different cellular structures.

Development of a highly efficient, strongly coupled organic light-emitting diode based on intracavity pumping architecture.

We report a highly efficient polariton organic light-emitting diode (POLED) based on an intracavity pumping architecture, where an absorbing J-aggregate dye film is used to generate polariton modes and a red fluorescent OLED is used for radiative pumping of emission from the lower polariton (LP) branch. To realize the device with large-area uniformity and adjustable coupling strength, we develop a spin-coating method to achieve high-quality J-aggregate thin films with controlled thickness and absorption. From systematic studies of the devices with different J-aggregate film thicknesses and OLED injection layers, we show that the J-aggregate film and the pump OLED play separate roles in determining the coupling strength and electroluminescence efficiency, and can be simultaneously optimized under a cavity design with a good LP-OLED emission overlap for effective radiative pumping. By increasing the absorption with thick J-aggregate film and improving the electron injection of pump OLED with Li2CO3 interlayer, we demonstrate the POLED with a large Rabi splitting energy of 192 meV and a maximum external quantum efficiency of 1.2%, a record efficiency of POLEDs reported so far. This POLED architecture can be generally applied for exploration of various organic materials to realize novel polariton devices and electrically pumped lasers.

Niche regulation of limbal epithelial stem cells: HC-HA/PTX3 as surrogate matrix niche.

Homeostasis of the corneal epithelium is ultimately maintained by stem cells that reside in a specialized microenvironment within the corneal limbus termed palisades of Vogt. This limbal niche nourishes, protects, and regulates quiescence, self-renewal, and fate decision of limbal epithelial stem/progenitor cells (LEPCs) toward corneal epithelial differentiation. This review focuses on our current understanding of the mechanism by which limbal (stromal) niche cells (LNCs) regulate the aforementioned functions of LEPCs. Based on our discovery and characterization of a unique extracellular matrix termed HC-HA/PTX3 (Heavy chain (HC1)-hyaluronan (HA)/pentraxin 3 (PTX3) complex, "-" denotes covalent linkage; "/" denotes non-covalent binding) in the birth tissue, i.e., amniotic membrane and umbilical cord, we put forth a new paradigm that HC-HA/PTX3 serves as a surrogate matrix niche by maintaining the in vivo nuclear Pax6+ neural crest progenitor phenotype to support quiescence and self-renewal but prevent corneal fate decision of LEPCs. This new paradigm helps explain how limbal stem cell deficiency (LSCD) develops in aniridia due to Pax6-haplotype deficiency and further explains why transplantation of HC-HA/PTX3-containing amniotic membrane prevents LSCD in acute chemical burns and Stevens Johnson syndrome, augments the success of autologous LEPCs transplantation in patients suffering from partial or total LSCD, and assists ex vivo expansion (engineering) of a graft containing LEPCs. We thus envisage that this new paradigm based on regenerative matrix HC-HA/PTX3 as a surrogate niche can set a new standard for regenerative medicine in and beyond ophthalmology.

Invasion and Colonization Pattern of Fusarium fujikuroi in Rice.

Bakanae disease in rice can cause abnormal elongation of the stem and leaves, development of adventitious roots, a larger leaf angle, and even death. Little is known about the infection, colonization, and distribution of Fusarium fujikuroi in rice plants across different growth stages. In this study, microscopic observation and quantitative real-time PCR were combined to investigate the pathogenesis of bakanae, using artificially inoculated seedlings of a susceptible rice cultivar, Zerawchanica karatals (ZK), a resistant cultivar, Tainung 67 (TNG67), naturally infected adult field plants (cultivars Kaohsiung 139, Taikeng 2, and Tainan 11), and an F. fujikuroi isolate expressing green fluorescent protein. In rice seedlings, F. fujikuroi hyphae were found to directly penetrate the epidermis of basal stems and roots, then extend inter- and intracellularly to invade the vascular bundles. Occlusion of vascular bundles and radial hyphal expansion from vascular bundles to surrounding parenchyma were observed in adult plants. Analysis of consecutive 3-cm segments of the whole plant revealed that F. fujikuroi was largely confined to the embryo, basal stem, and basal roots in seedlings, and distributed unevenly in the lower aerial parts (including nodes and internodes) of adult plants. The elongation and development of adventitious roots did not necessarily correlate with the amount of F. fujikuroi in diseased plants. Treatment of rice seeds with gibberellic acid-3 (GA3) at 0.5 mg/liter resulted in significantly more elongation of ZK than TNG67 seedlings, suggesting that the susceptibility of ZK to bakanae is associated with its higher sensitivity to GA3.

Third-harmonic generation modulation achieved through ground-state depletion.

Without real-state transition, third-harmonic generation (THG) cannot be modulated simply by fluorescence-based methods. However, utilizing the absorption-enhancement property of THG, the modulation of THG intensity has been demonstrated through ground-state depletion (GSD) in this Letter. By suppressing the absorption of materials with GSD, the THG intensity can be reduced due to a decreasing of absorption enhancement. The ability to modulate THG intensity can benefit from applying super-resolution techniques to THG microscopy.

HC-HA/PTX3 Purified From Amniotic Membrane Promotes BMP Signaling in Limbal Niche Cells to Maintain Quiescence of Limbal Epithelial Progenitor/Stem Cells.

To explore how limbal niche cells (LNCs) may control quiescence, self-renewal, and corneal epithelial lineage commitment/differentiation of limbal epithelial progenitor/stem cells (LEPCs), we have established an in vitro sphere assay by reunion between the two cell types in three-dimensional Matrigel. The resultant sphere exhibits inhibition of corneal epithelial lineage commitment/differentiation and marked clonal growth of LEPCs, of which the latter is correlated with activation of canonical Wnt signaling. Herein, we have created a similar reunion assay in immobilized heavy chain-hyaluronic acid/pentraxin 3 (HC-HA/PTX3), which is purified from amniotic membrane (AM) and consists of a complex formed by hyaluronic covalently linked to heavy chain 1 of inter-α-inhibitor and noncovalently linked to pentraxin 3. The resultant spheres exhibited similar suppression of corneal epithelial lineage commitment/differentiation but upregulation of quiescence markers including nuclear translocation of Bmi-1, and negligible clonal growth of LEPCs. This outcome was correlated with the suppression of canonical Wnt but activation of noncanonical (Planar cell polarity) Wnt signaling as well as BMP signaling in both LEPCs and LNCs. The activation of BMP signaling in LNCs was pivotal because nuclear translocation of pSmad1/5/8 was prohibited in hLEPCs when reunioned with mLNCs of conditionally deleted Bmpr1a;Acvr1(DCKO) mice. Furthermore, ablation of BMP signaling in LEPCs led to upregulation of cell cycle genes, downregulation of Bmi-1, nuclear exclusion of phosphorylated Bmi-1, and marked promotion of the clonal growth of LEPCs. Hence, HC-HA/PTX3 uniquely upregulates BMP signaling in LNCs which leads to BMP signaling in LEPCs to achieve quiescence, helping explain how AM transplantation is clinically useful to be used as a matrix for ex vivo expansion of LEPCs and to treat corneal blindness caused by limbal stem cells deficiency.

Constitutive expression of pentraxin 3 (PTX3) protein by human amniotic membrane cells leads to formation of the heavy chain (HC)-hyaluronan (HA)-PTX3 complex.

Heavy chain (HC)-hyaluronan (HA), a complex formed by the covalent linkage between HC1 from the inter-α-trypsin inhibitor (IαI) and HA, purified from the human amniotic membrane (AM), is responsible for the anti-inflammatory, antiscarring, and antiangiogenic actions of the AM. This HC-HA complex is produced by constitutive expression of TNF-stimulated gene 6 and endogenous production of IαI by AM cells. Pentraxin 3 (PTX3), a prototypic long pentraxin that plays a non-redundant role in innate immunity against selected pathogens, also helps stabilize HC-HA to ensure female fertility. Here we noted strong positive PTX3 staining in the AM epithelium and compact stroma. PTX3 was constitutively expressed and secreted by cultured AM epithelial and stromal cells and, further, greatly up-regulated by TNF and IL-1ß. Using an agarose overlay to trap the HA-containing matrix, the HC-HA-PTX3 complex was formed, as analyzed by Western blot analysis, by AM cells but not human skin fibroblasts, despite being cultured in the presence of serum and TNF. However, exogenous PTX3 helps human skin fibroblasts form the HC-HA-PTX3 complex with an agarose overlay. Furthermore, PTX3 can be coimmunoprecipitated with the HC-HA complex from agarose-overlaid AM cell extracts by an anti-human IαI antibody. Such a HC-HA-PTX3 complex can be reconstituted in vitro and exhibit similar effects as those reported for AM HC-HA-PTX3 on polarization of M2 macrophages. The tight binding between PTX3 and AM HC-HA withstands four runs of CsCl ultracentrifugation in the presence of 4 m GnHCl. These results indicate that PTX3 is constitutively expressed and secreted by AM cells as an integral component of the AM HC-HA-PTX3 complex and contributes to the biological function of AM HC-HA-PTX3.

Study of temperature distributions in pc-WLEDs with different phosphor packages.

Phosphor converted white LEDs (pc-WLED) with different geometries, namely remote-dome, remote-plate, half-dome, and conformal-coating packages, were studied using simulations to obtain the thermal power and the temperature distribution. The results of an experiment carried out to measure the temperature distribution in these pc-WLEDs validate the simulation results. A hotspot always occurs in the phosphor region. The conformal-coating package is the most feasible choice to reduce the hotspot temperature comparing with the other geometries.

Resolution enhancement of two-photon microscopy via intensity-modulated laser scanning structured illumination.

Conventional structured illumination microscopy (SIM) with wide-field illumination is an applicable tool to provide resolution enhancement. And yet its applications in thick specimens are still full of challenges. By combing the structured illumination concept with two-photon excitation, a laser scanning two-photon structured illumination microscope (LSTP-SIM) was constructed to gain ∼1.42-fold lateral resolution enhancement in contrast to two-photon fluorescence microscopy. With a point-scanning geometry, an acoustic-optical modulator was used to modulate temporally the excitation intensity in order to produce the structured illumination pattern. The theoretical models of image formation and image reconstruction were clearly established. Simulation and experiments were both performed to show the capability of this system to enhance the lateral resolution. Combined with the inherent optical sectioning power of the two-photon excitation, LSTP-SIM would have the potential for applications in optically-thick specimens.

Nuclear p120 catenin unlocks mitotic block of contact-inhibited human corneal endothelial monolayers without disrupting adherent junctions.

Contact inhibition ubiquitously exists in non-transformed cells that are in contact with neighboring cells. This phenomenon explains the poor regenerative capacity of in vivo human corneal endothelial cells during aging, injury and surgery. This study demonstrated that the conventional approach of expanding human corneal endothelial cells by disrupting contact inhibition with EDTA followed by bFGF activated canonical Wnt signaling and lost the normal phenotype to endothelial-mesenchymal transition, especially if TGFß1 was added. By contrast, siRNA against p120 catenin (CTNND1) also uniquely promoted proliferation of the endothelial cells by activating trafficking of p120 catenin to the nucleus, thus relieving repression by nuclear Kaiso. This nuclear p120-catenin-Kaiso signaling is associated with activation of RhoA-ROCK signaling, destabilization of microtubules and inhibition of Hippo signaling, but not with activation of Wnt-ß-catenin signaling. Consequently, proliferating human corneal endothelial cells maintained a hexagonal shape, with junctional expression of N-cadherin, ZO-1 and Na(+)/K(+)-ATPase. Further expansion of human corneal endothelial monolayers with a normal phenotype and a higher density was possible by prolonging treatment with p120 catenin siRNA followed by its withdrawal. This new strategy of perturbing contact inhibition by selective activation of p120-catenin-Kaiso signaling without disrupting adherent junction could be used to engineer surgical grafts containing normal human corneal endothelial cells to meet a global corneal shortage and for endothelial keratoplasties.

Synthesis of uniform poly(d,l-lactide) and poly(d,l-lactide-co-glycolide) microspheres using a microfluidic chip for comparison.

Applications of poly(l-lactide) (PLA) and poly(d,l-lactide-co-glycolide) (PLGA) microspheres are widely used in the biomedical and pharmaceutical fields. The effects of PLA/PLGA on microsphere properties when using conventional particulate preparation methods are not easily defined due to the uncontrollable particle size and size distribution. This study was aimed to synthesize uniform PLA and PLGA microspheres using a phenol formaldehyde resin-based microfluidic chip, which has the advantage of being solvent-resistant, flexible, and is readily disassembled for cleaning. The proposed chip can rapidly fabricate reproducible PLA and PLGA microspheres. Uniform emulsion droplets can be achieved by hydrodynamic flow focusing. After solvent evaporation, the free-flowing PLA and PLGA microspheres have a high level of morphological uniformity and size, allowing for a clear comparison of material effects. The results indicate that the sizes of the PLA and PLGA microspheres for the various flow rates of dispersed/continuous phases are very similar. The PLA/PLGA materials do not have a significant effect on particle size, but the particle surface indicates a different morphology. The result of the cytotoxicity evaluation shows no difference between PLA and PLGA and ensures the biocompatibility of both prepared PLA and PLGA microspheres for biomedical and pharmaceutical applications in the future.

Social Skill Abilities and Demographic Correlates of Older Volunteers.

Older age is associated with reduced social networks while social skill abilities are important assets for older people to cope with these situations. To better understand older people's social skill ability and important demographic correlates, the present cross-sectional survey research interviewed 1000 Taiwanese older volunteers from 73 community care centers. Findings of a mixed model indicated that education and serving area outweighed other demographic factors significantly correlated with older volunteers' social skill ability. Latent class analyses further identified different latent ability groups for different education (high vs. low) or serving-area (urban vs. suburban) older volunteers. Specifically, low-education or suburban volunteers were correlated with disadvantageous social skill profiles. Notably, the suburban eclectic ability group exhibited a skill pattern signifying the risk of loneliness. Overall, the findings called for further investigation into the relationship between older people's socioeconomic factors (e.g., education and serving area) and their social skill ability.

An acute herb-drug interaction of Magnoliae Officinalis Cortex with methotrexate via inhibiting multidrug resistance-associated protein 2.

Magnoliae Officinalis Cortex (MOC), an herbal drug, contains polyphenolic lignans mainly magnolol (MN) and honokiol (HK). Methotrexate (MTX), a critical drug for cancers and autoimmune deseases, is a substrate of multidrug resistance-associated protein 2 (MRP2) and breast cancer resistance protein (BCRP). This study investigated the effect of coadministration of MOC on the pharmacokinetics of MTX and relevant mechanisms. Sprague-Dawley rats were orally administered MTX alone and with single dose (2.0 and 4.0 g/kg) and repeated seven doses of MOC (2.0 g/kg thrice daily for 2 days, the 7th dose given at 0.5 h before MTX). The serum concentrations of MTX were determined by a fluorescence polarization immunoassay. The results showed that a single dose of MOC at 2.0 g/kg significantly increased the AUC0-t and MRT of MTX by 352% and 308%, and a single dose at 4.0 g/kg significantly enhanced the AUC0-t and MRT by 362% and 291%, respectively. Likewise, repeated seven doses of MOC at 2.0 g/kg significantly increased the AUC0-t and MRT of MTX by 461% and 334%, respectively. Mechanism studies indicated that the function of MRP2 was significantly inhibited by MN, HK and the serum metabolites of MOC (MOCM), whereas BCRP was not inhibited by MOCM. In conclusion, coadministration of MOC markedly enhanced the systemic exposure and mean residence time of MTX through inhibiting the MRP2-mediated excretion of MTX.

A microfluidic chip using phenol formaldehyde resin for uniform-sized polycaprolactone and chitosan microparticle generation.

This study develops a new solvent-compatible microfluidic chip based on phenol formaldehyde resin (PFR). In addition to its solvent-resistant characteristics, this microfluidic platform also features easy fabrication, organization, decomposition for cleaning, and reusability compared with conventional chips. Both solvent-dependent (e.g., polycaprolactone) and nonsolvent-dependent (e.g., chitosan) microparticles were successfully prepared. The size of emulsion droplets could be easily adjusted by tuning the flow rates of the dispersed/continuous phases. After evaporation, polycaprolactone microparticles ranging from 29.3 to 62.7 µm and chitosan microparticles ranging from 215.5 to 566.3 µm were obtained with a 10% relative standard deviation in size. The proposed PFR microfluidic platform has the advantages of active control of the particle size with a narrow size distribution as well as a simple and low cost process with a high throughput.

Manufacturing of human corneal endothelial grafts.

PURPOSE: Human corneal endothelial cells (HCECs) play a significant role in maintaining visual function. However, these cells are notorious for their limited proliferative capacity in vivo. Current treatment of corneal endothelial dysfunction resorts to corneal transplantation. Herein we describe an ex vivo engineering method to manufacture HCEC grafts suitable for transplantation through reprogramming into neural crest progenitors. METHODS: HCECs were isolated by collagenase A from stripped Descemet membrane of cadaveric corneoscleral rims, and induced reprogramming via knockdown with p120 and Kaiso siRNAs on collagen IV-coated atelocollagen. Engineered HCEC grafts were released after assessing their identity, potency, viability, purity and sterility. Phase contrast was used for monitoring cell shape, graft size, and cell density. Immunostaining was used to determine the normal HCEC phenotype with expression of N-cadherin, ZO-1, ATPase, acetyl-α-tubulin, γ-tubulin, p75NTR, α-catenin, ß-catenin, and F-actin. Stability of manufactured HCEC graft was evaluated after transit and storage for up to 3 weeks. The pump function of HCEC grafts was measured by lactate efflux. RESULTS: One HCEC graft suitable for corneal transplantation was generated from 1/8th of the donor corneoscleral rim with normal hexagonal cell shape, density, and phenotype. The manufactured grafts were stable for up to 3 weeks at 37 °C or up to 1 week at 22 °C in MESCM medium and after transcontinental shipping at room temperature by retaining normal morphology (hexagonal, >2000 cells/mm2, >8 mm diameter), phenotype, and pump function. CONCLUSIONS: This regenerative strategy through knockdown with p120 and Kaiso siRNAs can be used to manufacture HCEC grafts with normal phenotype, morphology and pump function following prolonged storage and shipping.