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The characteristics of Lu2O3-doped ZrO2 as a solid electrolyte material were investigated in terms of its oxygen ion conductivity and flexural strength to realize its electrolytic function at intermediate and high temperatures. The effect of doping Lu3+, which has a high nuclear charge electric field strength, was examined through impedance spectroscopy, open-circuit potential measurements, and bending tests. The results with Lu2O3 dopant were compared with those obtained with a widely used dopant, Y3+, having a similar ionic radius with Lu3+, as well as a dopant that provides high ionic conduction, Sc3+, having a smaller ionic radius with Zr4+. The results revealed that, at the same dopant concentration, both the ionic conductivity and the flexural strength of Lu2O3-doped ZrO2 are higher than those of the widely used Y2O3-doped ZrO2. The conductivity of 8 mol% Lu2O3-doped ZrO2 surpassed that of 8 mol% Sc2O3-doped ZrO2 in the range of 800–950 °C (0.153 S/cm vs. 0.121 S/cm at 900 °C). These results indicate the potential of Lu3+ as a dopant for enhancing the performance of ZrO2 solid electrolytes.

The degradation behavior of yttria-stabilized zirconia by thermal aging was investigated in terms of phase transformation, local atomic structure, and electrical conductivity. The average grain size of 8YSZ was increased from 20.83 μm to 25.81 μm with increasing aging temperature. All 8YSZ samples degraded at different temperatures had a predominantly cubic structure. The (400) peak of 8YSZ deteriorated at 1300 and 1400 °C shifted to a high angle, and the peak of tetragonal was not indexed. For 8YSZ degraded at 1500 °C, the (400) peak shifted to a lower angle, and the peak of tetragonal was identified. Analysis of the local microstructure of aged 8YSZ using extended X-ray absorption fine structure showed that the intensity of the Zr-O peak gradually increased and that the intensity of the peak of cationic Zr decreased as the aging temperature increased. The changes in the peaks indicate that the oxygen vacancies were reduced and Y3+ ions escaped from the lattice, leading to the destabilization of 8YSZ. The activation energies of 8YSZ at 1300 °C and 1400 °C were derived to be 0.86 and 0.87 eV, respectively, and the activation energy of 8YSZ at 1500 °C increased significantly to 0.92 eV. With the thermal deterioration of 8YSZ, the cation (Y3+) escaped from the lattice and the number of oxygen vacancies decreased, resulting in the formation of a tetragonal structure and high activation energy at 1500 °C.

Legg-Calvé-Perthes disease is a juvenile ischemic osteonecrosis which produces extensive necrotic cell debris and release of damage associated molecular patterns (DAMPs) in the femoral head. The necrotic bone environment induces a chronic inflammatory repair response with excessive bone resorption leading to deformity and early osteoarthritis. Currently there is no minimally invasive method to clear the necrotic materials from the bone to decrease the inflammatory burden of the necrotic environment and to improve the healing process.We hypothesized that a novel minimally invasive two-needle saline washing technique would be effective to remove cell debris, proteins, and fat from the marrow space of porcine cadaveric humeral heads (HHs).Twenty-two HHs were subjected to three freeze-thaw cycles to simulate osteonecrosis prior to the wash procedure which consisted of placement of two 15-gauge intraosseous needles followed by incremental saline wash. After the washout procedure, the solutions were collected for measurements of turbidity, protein concentration, and cell count. The HHs were analyzed by optical scanning and histology.The solution collected after each wash showed a significant decrease in the turbidity, cell count, and protein concentration (p0.05). Histologic assessment showed significantly decreased cell debris and adipocytes in the washed group compared to the unwashed group (p0.001).The two-needle intraosseous wash technique effectively removed cell debris and proteins from the marrow space. The technique may be used to reduce the necrotic cell debris and DAMPs present in the necrotic bone.III, in vitro comparative study.

BACKGROUNDIn order to reveal the influence of cellular characteristics and media properties on membrane microalgal harvesting, changes in the cellular characteristics and media properties of Scenedesmus acuminatus culture contaminated by microzooplankton were characterized, and their influences on membrane harvesting were compared with the control culture. Cells and algogenic organic matter (AOM) in the contaminated and control cultures were fractionated to evaluate the membrane fouling potential. RESULTSThe contaminated culture had a 32% lower average flux than control culture. Both the fractionated cells and AOM in the contaminated culture with or without concentration adjustment had higher membrane fouling potentials. Microzooplankton contamination caused more cell aggregates and transparent exopolymer particles, which formed less permeable cake layer. Increased AOM in contaminated culture contained more carbohydrates and humic/fulvic acid-like substances, which caused a 15.5-fold higher adhesion force between AOM and the membrane surface, and thus resulted in higher flux declining rates and lower backwashing recovery rates. CONCLUSIONMicrozooplankton contamination caused the formation of cell aggregates and high concentration of AOM in S.acuminatus culture. AOM played a more important role than microalgal cells in flux declining and backwashing recovery. It is necessary to develop cultivation practice and culture pretreatment techniques to improve membrane harvesting efficiency. (c) 2019 Society of Chemical Industry

The placental tissue, due to its angiogenic, anti-inflammatory, antioxidative, antimicrobial, and anti-fibrotic properties, has become a compelling source towards a solution for several indications in regenerative medicine. However, methods to enhance and capture the therapeutic properties with formulations that can further the applications of viable placental tissue have not been explored. In this study, we investigated the regenerative effects of a hypoxia primed flowable placental formulation (FPF), composed of amnion/chorion and umbilical tissue, in two in vivo injury models. Laser Doppler data from rodent ischemia hindlimbs treated with FPF revealed significant tissue perfusion improvements compared to control ischemic hindlimbs. To further corroborate FPF’s effects, we used a rodent ischemic bipedicle skin flap wound model. FPF treatment significantly increased the rate of wound closure and the quality of wound healing. FPF-treated wounds displayed reduced inflammation and an increase in angiogenesis. Furthermore, quantitative PCR and next-generation sequencing analysis confirmed these changes in the FPF-treated group at both the gene and transcriptional level. The observed modulation in miRNAs was associated with angiogenesis, regulation of inflammatory microenvironment, cell migration and apoptosis, reactive oxygen species generation, and restoring epithelial barrier function, all processes involved in impaired tissue healing. Taken together, these data validate the tissue regenerative properties of the flowable placental formulation configuration tested.

The degradation behavior of yttria-stabilized zirconia under thermal stress was investigated in terms of phase transformation, local atomic structure, and electrical conductivity. The average grain size of 8YSZ were increased from 20.83 µm to 25.81 µm with increasing aging temperature. All 8YSZ samples degraded at different temperatures had a predominantly cubic structure. The (400) peak of 8YSZ deteriorated at 1300 and 1400°C shifted to high angle, and the peak of tetragonal was not indexed. For 8YSZ degraded at 1500°C, the (400) peak shifted to a lower angle, and the peak of tetragonal was identified. In the local atomic structure of the aged 8YSZ with extended X-ray absorption fine structure, the intensity of the Zr-O peak gradually increased, and the intensity of the Zr-Cation peak decreased as the aging temperature increased. The changes in the peaks indicate that the oxygen vacancies were reduced and Y3+ ions escaped from the lattice, leading to destabilization of 8YSZ. The activation energies of 8YSZ at 1300°C and 1400°C were derived to be 0.86 and 0.87 eV, respectively, and the activation energy of 8YSZ at 1500°C increased significantly to 0.92 eV. With the thermal deterioration of 8YSZ, the cation (Y3+) escaped from the lattice and the number of oxygen vacancies decrease, resulting in the formation of a tetragonal structure and high activation energy at 1500°C.

BACKGROUND Ischemic osteonecrosis of the femoral head produces necrotic cell debris and inflammatory molecules in the marrow space, which elicit a chronic inflammatory repair response. The purpose of this study was to determine the effects of flushing out the necrotic cell debris and inflammatory proteins on bone repair in a piglet model of ischemic osteonecrosis. METHODS Osteonecrosis of the femoral head of the right hindlimb was induced in 12 piglets by tying a ligature tightly around the femoral neck. One week after the surgery, 6 animals were treated with a percutaneous 3-needle bone washing procedure and non-weight-bearing (NWB) of the right hindlimb (wash group). The total saline solution wash volume was 450 mL per femoral head. Serial wash solutions were collected and analyzed. The remaining 6 animals were treated with NWB only (NWB group). At 8 weeks after the surgery, the femoral heads were assessed using radiography, micro-computed tomography (micro-CT), and histological analysis. In addition, we compared the results for these piglets with our published results for 6 piglets treated with multiple epiphyseal drilling (MED) plus NWB without bone washing (MED group). RESULTS Necrotic cells and inflammatory proteins were present in the bone wash solution collected 1 week after ischemia induction. The protein and triglyceride concentrations decreased significantly with subsequent washing (p < 0.005). At 8 weeks after ischemia induction, the wash group had a significantly higher bone volume than the MED or NWB group (p < 0.0001). Histological bone-formation measures were also significantly increased in the wash group compared with the MED group (p = 0.002) or NWB group (p < 0.0001) while macrophage numbers were significantly decreased in the wash group. CONCLUSIONS The percutaneous 3-needle procedure flushed out cell debris and inflammatory proteins from the necrotic femoral heads, decreased osteoclasts and macrophages, and increased bone formation following induction of ischemic osteonecrosis. CLINICAL RELEVANCE We believe that this is the first study to investigate the concept of washing out the necrotic femoral head to improve bone healing. The minimally invasive procedure may be useful to improve the necrotic bone environment and bone repair following ischemic osteonecrosis.

Pancreatic cancer is the main cause of cancer-related deaths worldwide and is difficult to diagnose before the extensive local invasion and distant organ metastasis. Mesothelin is abnormally overexpressed in tumors such as ovary cancer and pancreatic cancer. Studies show a significant link between mesothelin overexpression and short survival in patients with pancreatic cancer. In this study, we screened a phage displayed-peptide library for peptides that selectively bind to mesothelin using mesothelin-overexpressing cells. After five rounds of screening, we selected a 9-mer peptide (named MSLN-Pep) that preferentially bound to mesothelin-high pancreatic cancer cells such as ASPC-1 and Panc-1 cells over mesothelin-low cells such as HEK 293 cells. MSLN-Pep was efficiently internalized into ASPC-1 cells and inhibited the cell migration and invasion while little affected the phosphorylation of Akt. Moreover, a MSLN-Pep-guided proapoptotic peptide (MSLN-Pep-KLA) exerted selective cytotoxicity against pancreatic cancer cells over mesothelin-low cells. MSLN-Pep-KLA when combined with gemcitabine, a chemotherapeutic agent against pancreatic cancer, sensitized ASPC-1 cells to the gemcitabine treatment. These results suggest that MSLN-Pep is a promising tool for a targeted therapy against pancreatic cancer expressing mesothelin at high levels. Citation Format: Min-Sung Park, Byungheon Lee. Identification of mesothelin binding peptide for targeted therapy against pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1743.

Fibrosis, the thickening and scarring of injured connective tissue, leads to a loss of organ function. Multiple cell types, including T-cells, macrophages, fibrocytes, and fibroblasts/myofibroblasts contribute to scar formation via secretion of inflammatory factors. This event results in an increase in oxidative stress and deposition of excessive extracellular matrix (ECM), characteristic of fibrosis. Further, aging is known to predispose connective tissue to fibrosis due to reduced tissue regeneration. In this study, we investigated the anti-fibrotic activity of a flowable placental formulation (FPF) using a bleomycin-induced dermal fibrosis model in aged mice. FPF consisted of placental amnion/chorion- and umbilical tissue-derived ECM and cells. The mice were injected with either FPF or PBS, followed by multiple doses of bleomycin. Histological assessment of FPF-treated skin samples revealed reduced dermal fibrosis, inflammation, and TGF-&beta, signaling compared to the control group. Quantitative RT-PCR and Next Generation Sequencing analysis of miRNAs further confirmed anti-fibrotic changes in the FPF-treated group at both the gene and transcriptional levels. The observed modulation in miRNAs was associated with inflammation, TGF-&beta, signaling, fibroblast proliferation, epithelial-mesenchymal transition and ECM deposition. These results demonstrate the potential of FPF in preventing fibrosis and may be of therapeutic benefit for those at higher risk of fibrosis due to wounds, aging, exposure to radiation and genetic predisposition.

In Legg-Calvé-Perthes disease (LCPD), a loss of blood supply to the juvenile femoral head leads to extensive cell death and release of damage-associated molecular patterns (DAMPs). Over time chronic inflammatory repair process is observed with impaired bone regeneration. Increased fibrous tissue and adipose tissue are seen in the marrow space with decreased osteogenesis in a piglet model of LCPD, suggesting inhibition of osteoblastic differentiation and stimulation of fibroblastic and adipogenic differentiation of mesenchymal stem cell (MSC) during the healing process. Little is known about the DAMPs present in the necrotic femoral head and their effects on MSC differentiation. The purpose of this study was to characterize the DAMPs present in the femoral head following ischemic osteonecrosis and to determine their effects on MSC differentiation. Necrotic femoral heads were flushed with saline at 48 h, 2 weeks and 4 weeks following the induction of ischemic osteonecrosis in piglets to obtain necrotic bone fluid (NBF). Western blot analysis of the NBF revealed the presence of prototypic DAMP, high mobility group box 1 (HMGB1), and other previously described DAMPs: biglycan, 4-hydroxynonenal (4-HNE), and receptor activator of NF-κB ligand (RANKL). ELISA of the NBF revealed increasing levels of inflammatory cytokines IL1β, IL6 and TNFα with the temporal progression of osteonecrosis. To determine the effects of NBF on MSC differentiation, we cultured primary porcine MSCs with NBF obtained by in vivo necrotic bone flushing method. NBF inhibited osteoblastic differentiation of MSCs with significantly decreased OSX expression (p = 0.008) and Von Kossa/Alizarin Red staining for mineralization. NBF also significantly increased the expression of proliferation markers Ki67 (p = 0.03) and PCNA (p 0.0001), and fibrogenic markers Vimentin (p = 0.02) and Fibronectin (p = 0.04). Additionally, NBF treated MSC cells showed significantly elevated RANKL/OPG secretion ratio (p = 0.003) and increased expression of inflammatory cytokines IL1β (p = 0.006) and IL6 (p 0.0001). To specifically assess the role of DAMPs in promoting the fibrogenesis, we treated porcine fibroblasts with artificial NBF produced by bone freeze-thaw method. We found increased fibroblastic cell proliferation in an NBF dose-dependent manner. Lastly, we studied the effect of HMGB1, a prototypic DAMP, and found that HMGB1 partially contributes to MSC proliferation and fibrogenesis. In summary, our findings show that DAMPs and the inflammatory cytokines present in the necrotic femoral head inhibit osteogenesis and promote fibrogenesis of MSCs, potentially contributing to impaired bone regeneration following ischemic osteonecrosis as observed in LCPD.

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Introduction Legg–Calve–Perthes disease is a juvenile ischemic osteonecrosis which produces extensive necrotic cell debris and release of damage associated molecular patterns (DAMPs) in the femoral head. The necrotic bone environment induces a chronic inflammatory repair response with excessive bone resorption leading to deformity and early osteoarthritis. Currently, there is no minimally invasive method to clear the necrotic materials from the bone to decrease the inflammatory burden of the necrotic environment and to improve the healing process. Hypothesis We hypothesized that a novel minimally invasive two-needle saline washing technique would be effective to remove cell debris, proteins, and fat from the marrow space of porcine cadaveric humeral heads (HHs). Materials and methods Twenty-two HHs were subjected to three freeze-thaw cycles to simulate osteonecrosis prior to the wash procedure which consisted of placement of two 15-gauge intraosseous needles followed by incremental saline wash. After the washout procedure, the solutions were collected for measurements of turbidity, protein concentration, and cell count. The HHs were analyzed by optical scanning and histology. Results The solution collected after each wash showed a significant decrease in the turbidity, cell count, and protein concentration (p Discussion/conclusion The two-needle intraosseous wash technique effectively removed cell debris and proteins from the marrow space. The technique may be used to reduce the necrotic cell debris and DAMPs present in the necrotic bone. Level of evidence III, in vitro comparative study.

Transcriptional coactivator PPAR γ coactivator (PGC)-1α and its splice variant N-terminal (NT)-PGC-1α mediate transcriptional regulation of brown adipose tissue (BAT) thermogenesis in response to changes in ambient temperature. PGC-1α is dispensable for cold-induced BAT thermogenesis as long as NT-PGC-1α is present. However, the functional significance of NT-PGC-1α in BAT has not been determined. In the present study, we generated NT-PGC-1α(−/−) mice to investigate the effect of NT-PGC-1α deficiency on adaptive BAT thermogenesis. At thermoneutrality, NT-PGC-1α(−/−) mice exhibited abnormal BAT phenotype with increased accumulation of large lipid droplets concomitant with marked downregulation of FA oxidation (FAO)-related genes. Consistent with transcriptional changes, mitochondrial FAO was significantly diminished in NT-PGC-1α(−/−) BAT. This alteration, in turn, enhanced glucose utilization within the NT-PGC-1α(−/−) BAT mitochondria. In line with this, NT-PGC-1α(−/−) mice had higher reliance on carbohydrates. In response to cold or β(3)-adrenergic receptor agonist, NT-PGC-1α(−/−) mice transiently exhibited lower thermogenesis but reached similar thermogenic capacities as their WT littermates. Collectively, these findings demonstrate that NT-PGC-1α is an important contributor to the maintenance of FAO capacity in BAT at thermoneutrality and provide deeper insights into the relative contributions of PGC-1α and NT-PGC-1α to temperature-regulated BAT remodeling.

Background aims Previously, we showed that human mesenchymal stromal cells (hMSCs) were activated to express tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) upon TNF-α stimulation, induced cell death in triple-negative breast cancer (TNBC) MDA-MB-231 cells (MDA), and RNA released from apoptotic MDA further increased TRAIL expression in hMSCs. This feed-forward stimulation increased apoptosis in MDA cells. Here, we tested whether TRAIL-expressing hMSCs, in combination with a sub-toxic-dose of a chemotherapy drug doxorubicin, would overcome TRAIL resistance and create synergistic effects on targeting metastatic TNBC. Methods To optimize conditions for the combination treatment, we (i) selected an optimal condition to activate hMSCs for TRAIL expression, (ii) selected an optimal dose of doxorubicin treatment, (iii) examined underlying mechanisms in vitro and (iv) tested the efficacy of the optimized conditions in a xenograft mouse model of human breast cancer lung metastasis. Results The results showed that DNA fragments from apoptotic MDA triggered hMSCs to increase further TRAIL expression in an absent in melanoma 2 (AIM2)–dependent manner, and thus higher TRAIL-expressing hMSCs stimulated with synthetic DNA, poly(deoxyadenylic-deoxythymidylic) acid [poly(dA:dT)], more effectively suppressed tumor progression in vivo . Furthermore, activated hMSCs increased apoptosis in MDA cells when combined with a sub-toxic dose of doxorubicin, which was mediated by up-regulating TRAIL and Fas-related pathways. When we combined the optimized conditions, pre-activated hMSCs with poly (dA:dT) synergistically reduced tumor burden even with minimal doxorubicin treatment in a xenograft mouse model of human breast cancer lung metastasis. Conclusions These results suggest that the treatment of hMSCs with a sub-toxic dose of doxorubicin can overcome TRAIL resistance and be a potential novel therapy for TNBC metastasis treatment.

Bone marrow-derived mesenchymal stem cells (BMSCs) are a good cell source for regeneration of cartilage as they can migrate directly to the site of cartilage injury and differentiate into articular chondrocytes. Articular cartilage defects do not heal completely due to the lack of chondrocytes or BMSCs at the site of injury. In this study, the chemotaxis of BMSCs toward chemokines, which may give rise to a complete regeneration of the articular cartilage, was investigated. CCR2, CCR4, CCR6, CXCR1, and CXCR2 were expressed in normal BMSCs and were increased significantly upon treatment with proinflammatory cytokines. BMSC migration was increased by MIP-3α and IL-8 more than by MCP-1 or SDF-1α. IL-8 and MIP-3α significantly enhanced the chemotaxis of BMSCs compared with MCP-1, SDF-1α, or PBS. Human BMSC recruitment to transplanted scaffolds containing either IL-8 or MIP-3α significantly increased in vivo compared to scaffolds containing PBS. Furthermore, IL-8- and MIP-3α-containing scaffolds enhanced tissue regeneration of an osteochondral defect site in beagle knee articular cartilage. Therefore, this study suggests that IL-8 and MIP-3α are the candidates that induce the regeneration of damaged articular cartilage.

Mesenchymal stem or stromal cells (MSCs) have many potential therapeutic applications including therapies for cancers and tissue damages caused by cancers or radical cancer treatments. However, tissue-derived MSCs such as bone marrow MSCs (BM-MSCs) may promote cancer progression and have considerable donor variations and limited expandability. These issues hinder the potential applications of MSCs, especially those in cancer patients. To circumvent these issues, we derived MSCs from transgene-free human induced pluripotent stem cells (iPSCs) efficiently with a modified protocol that eliminated the need of flow cytometric sorting. Our iPSC-derived MSCs were readily expandable, but still underwent senescence after prolonged culture and did not form teratomas. These iPSC-derived MSCs homed to cancers with efficiencies similar to BM-MSCs but were much less prone than BM-MSCs to promote the epithelial-mesenchymal transition, invasion, stemness, and growth of cancer cells. The observations were probably explained by the much lower expression of receptors for interleukin-1 and TGFβ, downstream protumor factors, and hyaluronan and its cofactor TSG6, which all contribute to the protumor effects of BM-MSCs. The data suggest that iPSC-derived MSCs prepared with the modified protocol are a safer and better alternative to BM-MSCs for therapeutic applications in cancer patients. The protocol is scalable and can be used to prepare the large number of cells required for "off-the-shelf" therapies and bioengineering applications.

Biomass is considered an alternative energy which can solve an greenhouse gas problem like CO2 which is a major contributor to global warming. The biomass can be converted to various energy sources through thermochemical conversion. In this study, a continuous gasifier was engineered for a wood biomass gasification. The biomass was used a waste wood. The experiments of CO2 gasification were achieved as the gasification temperature, moisture content and input CO2 concentration. The results showed that the yield of producer gas increased with an increasing the gasification temperature. The amount of the light tar increased due to the decomposition of gravimetric tar by the thermal cracking, and the char was confirmed pore development through the SEM analysis. The CO concentration was increased with an increased input CO2 concentration from Boudouard reaction. Through the parametric screening studies, the hydrogen and carbon monoxide concentration were 32.91% and 48.33% at the optimal conditions of this test rig.

Background: Tendinopathy is a clinical problem for which treatment shows mixed results and treatment options are limited. Gene expression signatures early in the mechanotransduction pathway can accurately predict risk and correlate with different clinical outcomes. Studies aimed at elucidating the molecular mechanisms of tendinopathy have focused on small cohorts of genes that show an incomplete picture of the degeneration process. This study compared the effect of cyclic strain on gene expression in tendon cells from normal tendon and chronically painful areas of tendinopathy in 3 patients. Methods: We measured a panel of mechanotransduction genes and cytoskeletal tensional balance with and without cyclic strain, which disrupts connective tissue synthetic-degradative equilibrium. Normal and degenerative tendons were obtained from patients undergoing surgery to treat chronic painful tendinopathy. A cyclic strain model was established to measure cytoskeletal tensional homeostasis. Results: Prior to cyclic strain, the normal tendon cells exhibited varying patterns of elevated expression of 7 genes compared with degenerative tendon cells. In response to cyclic strain, gene expression of COL1A1, ITGA6, CTNNA1, and CLEC3B was up-regulated in normal tendon cells. Cyclic strain had no effect on degenerative tendon cells. Cyclic strain exacerbated the inhibition of protein synthesis in both cell types, especially in the degenerative tendon cells. Conclusion: Alterations in the pattern of gene expression are suggestive of a dynamic equilibrium between synthesis and degradation, whereby cell adhesion molecules are predominantly up-regulated to facilitate cellular reorientation in response to their altered functional environment. Clinical Relevance: These data might have future applications, including the identification of markers for early diagnosis, targets for drug design, and indicators for treatment responsiveness and prognosis.

microRNAs are small molecules, about 17-23 nucleotides in length, that act as translational regulators of their target gene. By binding to a target, microRNAs are known to either inhibit translation or induce degradation of the target. Despite the great interest in microRNAs, however, the exact targets of each individual microRNA in different processes remain largely unknown. In this study, we determined that the lymphoid enhancer-binding factor-1 (LEF-1) was expressed during the chondrogenesis of human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and sought to identify a novel microRNA targeting this gene. Through subsequent studies, we have identified, for the first time, one particular microRNA, miR-449a, that recognizes and regulates the expression of LEF-1 in a dose-dependent and sequence-specific manner. In addition, we observed that the inhibition of LEF-1 via miR-449a led to the subsequent repression of Sox 9, which is a well-established regulator of chondrogenesis. Collectively, this study demonstrated that miR-449a directly targets LEF-1, which in turn affects the expression of Sox 9, ultimately leading to the proper regulation of the differentiation and chondrogenesis of human MSCs (hBM-MSCs).

Anodized tubular TiO 2 electrodes (ATTEs) are prepared using an organic additive consisting of either (i) ethylene glycol (EG) or (ii) glycerol (Gly) to make various photoanodes with different length of TiO 2 tubes and thereby to investigate the effect of their length on the photo-driven activity for hydrogen evolution and Cr(VI) reduction, as well as on the photocurrent. The ATTEs with EG have longer TiO 2 tubes (3.42–15.6 μm) than those with Gly (0.26–1.95, 6.82 μm). The former samples exhibit higher photocurrent densities (22.8–32.8 mA cm −2 ) than the latter (8.0–19.4, 20.3 mA cm −2 ). The latter samples (tube length of less than 7 μm) clearly exhibit a change of the rate-determining step from electron migration to photohole capture as the scanned applied bias increases, since the photocurrent shows a plateau for tube lengths above 2 μm. Meanwhile, the samples with EG remain in the electron migration step up to a tube length of 16 μm and is due to the difference of the morphology, crystal phase and crystallinity. This favourable characteristic is also applied to and well matched with the results from the reactions of Cr(VI) reduction and hydrogen evolution (up to ca. 250 μmol h −1 ).

The properties of regenerated cartilage using bone marrow-derived mesenchymal stem cells (MSCs) and poly lactic-co-glycolic acid (PLGA) scaffold composites pretreated with TGF-beta3 were investigated and compared to the non-TGF-beta3 treated MSCs/PLGA composites in a rabbit model. We prepared MSCs/PLGA scaffold composites and pretreated it with TGF-beta3 for 3 weeks prior to transplantation. Then, composites were transplanted to the osteochondral defect in the rabbit knee. After 12 weeks of transplantation, 10 of the 12 rabbits in which TGF-beta3 pretreated MSCs/PLGA scaffold composites were transplanted showed cartilaginous regeneration. In gross morphology, regenerated cartilage showed smooth, flush, and transparent features. In indentation test, this had about 80% of Young's modulus of normal articular cartilage. Histological examination demonstrated hyaline like cartilage structures with glycosaminoglycan and type II collagen expression. Histological scores were not statistically different to the normal articular cartilage. These results showed improvement of cartilage regeneration compared to the non-TGF-beta3 pretreated MSCs/PLGA scaffold composite transplanted group. Thus, we have successfully regenerated improved hyaline-like cartilage and determined the feasibility of treating damaged articular cartilage using MSCs/PLGA scaffold composite pretreated with TGF-beta3. Also, we suggest this treatment modality as another concept of cartilage tissue engineering.

Bone marrow derived-stromal cells (BMSCs) are pluriplotent progenitors for a variety of cell types, including osteoblast, chondrocyte, adipocyte, and so on. Stem cell research has enormous potential in the future clinical treatment of a wide range of diseases. Tracking stem cell localization, survival, differentiation, and proliferation after transplantation in living subjects is essential for understanding stem cell biology and physiology. However, we don’t have exact evaluation methods and safeguards for clinical application. In this study, we investigated tracking BMSCs differentiation, localization, toxicity, and migration in vivo. The fluorescent vector used in our studies did not affect BMSCs viability or their ability to undergo osteogenic and adipogenic differentiation in vitro. During differentiation, EGFP-BMSCs by Oil Red O and Alizarin Red S were stained. EGFP-BMSCs were transplanted into the femoral region in autologous rabbit. After one month, these cells were detectable by confocal microscopy and RTPCR. Transplanted EGFP-BMSCs were not detected another organs (spleen, kidney, liver, and muscle) in immunohistochemistry and RT-PCR. In organ-function test and cell-toxicity examination, there is no difference between the before and after EGFP-BMSCs transplantation. we observed that transplanted EGFP-BMSCs were not affected cell-toxicity and migration. This results offer some evaluation methods and safeguards for clinical application using BMSCs. In further study, it will be needed to test reproductive and developmental toxicity after transplantation and observe migration of EGFP-BMSCs after transplantation.

Poly-L-lactic acid (PLLA) is a desirable and very attractive polymer for fabricating porous scaffolds. As of now, a solvent casting method with organic solvents has been used in scaffold fabrication process. However, residual organic solvents in the scaffolds have the problems of decreasing the effect of osteogenic induction due to the hindrance of bioceramic by polymer solution and it’s harmfulness in vivo. To avoid these disadvantages of scaffolds by organic solvent casting method, we developed a new method fabricating polymer (PLLA)/ceramic (β -TCP) composite scaffolds by baking method without using solvent, and then we tested properties of scaffolds on animals. As the result, non-toxicity has been proved through animal experiment and newly fabricated polymer/ceramic composites by a novel sintering method were induced rapid bone regeneration through enhancing the interaction of cells and a bone induction factor without any host immune response.

In this paper, we propose a novel LuxUI based on an ambient light sensor as an alternative input method for smartwatches. We present a concept of LuxUI with design and implementation details of LuxUI interactions and events followed by possible applications and discussion issues. The proposed LuxUI is simple in design yet effective which works well for 200 ms interval interaction with a modest battery usage. For contact-based explicit interaction on smartwatches, the proposed LuxUI can be used as an alternative UI for otherwise expressivity-limited smartwatches.

The aim of this study was to identify the signal transduction pathways and mechano-transducers that play critical roles in the processes induced by changes in cyclic hydrostatic pressure and fluid shear in 3-dimensional (3D) culture systems. Mesenchymal stem cells were loaded into a polymeric scaffold and divided into three groups according to the stress treatment: static, fluid shear, and hydrostatic pressure with fluid shear. Cells were exposed daily to a hydrostatic pressure of 0.2 MPa for 1 min followed by 14 min rest with fluid flow at 30 rpm. Protein extracts were analyzed by Western blot for extracellular signal-regulated kinase 1/2 (ERK1/2). The complexes were cultured under the mechanical stimuli for 21 days with or without phospho-ERK1/2 inhibitor (U0126) and evaluated by RT-PCR, calcium contents, and immunohistochemistry. Under conditions of mechanical stimulation, the activation of ERK1/2 was sustained or increased with time. U0126 suppressed mechanical stimuli-induced expression of osteocalcin. In addition, calcium contents and the degrees of osteocalcin and osteopontin staining were decreased by this inhibitor. These results demonstrate that mechanical stimuli, particularly hydrostatic pressure with fluid shear, enhance osteogenesis in 3D culture systems via ERK1/2 activation.

The transformation of acyclic allylic benzamides 4 and homoallylic benzamides 12 to vinyl oxazolines 3 is achieved in the presence of base by the catalysis of Pd(0) in high yield and with high diastereoselectivity. Especially, in the case of homoallylic benzamides 12, trans-oxazolines 3 are formed exclusively or predominantly over cis-oxazolines 8, irrespective of the composition of their stereoisomers. The reaction is believed to proceed via the same π-allylpalladium complex that arises from either primary or secondary allylic acetates. We applied this method to the syntheses of β-amino-α-hydroxy acids 1 and γ-amino-β-hydroxy acids 2, conveniently protected as oxazoline.

Our recent study showed that human mesenchymal stem/stromal cells (hMSCs) are activated to express tumor necrosis factor (TNF)-α-related apoptosis-inducing ligand (TRAIL) by exposure to TNF-α and these activated hMSCs effectively induce apoptosis in triple-negative breast cancer MDA-MB-231 (MDA) cells in vitro and in vivo. Here, we further demonstrated that activated hMSCs not only induced apoptosis of MDA cells but also reduced metastatic features in MDA cells. These activated hMSC-exposed MDA cells showed reduced tumorigenicity and suppressed formation of lung metastasis when implanted in the mammary fat pad. Surprisingly, the activated hMSC-exposed MDA cells increased TRAIL expression, resulting in apoptosis in MDA cells. Interestingly, upregulation of TRAIL in MDA cells was mediated by interferon-beta (IFN-β) secreted from activated hMSCs. Furthermore, IFN-β in activated hMSCs was induced by RNA and DNA released from apoptotic MDA cells in absent in melanoma 2 (AIM2) and IFN induced with helicase C domain 1 (IFIH1)-dependent manners. These observations were only seen in the TRAIL-sensitive breast cancer cell lines but not in the TRAIL-resistant breast cancer cell lines. Consistent with these results, Kaplan–Meier survival analysis also showed that lack of innate sensors detecting DNA or RNA is strongly associated with poor survival in estrogen receptor-negative breast cancer patients. In addition, cancer-associated fibroblasts (CAF) isolated from a breast cancer patient were also able to express TRAIL and IFN-β upon DNA and RNA stimulation. Therefore, our results suggest that the crosstalk between TRAIL-sensitive cancer cells and stromal cells creates a tumor-suppressive microenvironment and further provide a novel therapeutic approach to target stromal cells within cancer microenvironment for TRAIL sensitive cancer treatment.

The purpose of this study was to evaluate the mechanism of crosstalk between the type II collagen and TGF-beta1 signaling pathways in chondrocytic cells. Articular chondrocytes, isolated from porcine knee cartilage, and the SW1353 cell line were cultured on either type II collagen-coated or -uncoated plates in the presence or absence of TGF-beta1. Expression of pSMAD 2, pSMAD 3, pFAK(Y397) and pFAK(Y925) in articular chondrocytes and the SW1353 cell line was analyzed by immunoblotting. Cell proliferation rates and glycosaminoglycan (GAG) content was determined after treatment with type II collagen or/and TGF-beta1. For inhibition study, human FAK-specific RNA small interference (siFAK) in SW1353 cell line was performed. In this study, expression of pSMAD 2, pSMAD 3, pFAK(Y397) and pFAK(Y925) were synergistically increased by co-treatment with type II collagen and TGF-beta1 in articular chondrocytes. The proliferation of porcine articular chondrocytes and GAG secretion in SW1353 cells were synergistically increased by co-stimulation with type II collagen and TGF-beta1. Synergistically increased expression and nuclear translocation of pSMAD 2 and pSMAD 3 and GAG secretion of SW1353 cells were significantly inhibited by siFAK transfection. Therefore, we suggest that FAK-SMAD 2/3 mediates signal crosstalk between type II collagen and TGF-beta1 and regulates GAG secretion in chondrocytic cells.

The prevalence of liver and intestinal helminth infections, including Opisthorchis, Haplorchis, Phaneropsolus, hookworms, Enterobius, and Taenia, was surveyed in Khammouane province, Lao PDR. Fecal specimens were collected from 1,242 people (590 men and 652 women) in 3 Mekong riverside villages and were examined by the Kato-Katz thick smear technique. The overall helminth egg positive rate was 81.1%. The positive rate for small trematode eggs, including Opisthorchis viverrini, heterophyids, and lecithodendriids, was 81.1% and the positive rate for hookworms was 6.7%. To obtain adult worms, 35 people who were positive for small trematode eggs were treated with 20-30 mg/kg praziquantel and 10-15 mg/kg pyrantel pamoate, and then purged. Diarrheic stools were collected from 33 of these people and searched for helminth parasites using a stereomicroscope. Mixed infections with various helminths (Haplorchis taichui, Haplorchis yokogawai, Prosthodendrium molenkampi, Phaneropsolus bonnei, echinostomes, hookworms, Trichostrongylus spp., Trichuris trichiura, Enterobius vermicularis, and/or Taenia saginata) were found. The total number of helminth specimens collected was 20,907 (approximately 634 per person). The most common species was H. taichui, followed by P. molenkampi, O. viverrini, P. bonnei, E. vermicularis, hookworms, and Trichostrongylus spp. These results show that diverse species of intestinal nematodes, trematodes, and cestodes are infecting humans in Khammouane province, Lao PDR.

Open reading frame 11 (ORF11) of Kaposi's sarcoma-associated herpesvirus belongs to a herpesviral homologous protein family that is conserved among members of the gamma-herpesvirus subfamily. Little is known about the function of ORF11 and how this viral gene is regulated in KSHV life cycle. In this study, we have characterized the major transcript of the ORF11 gene, which is located adjacent and in the opposite orientation to a well-characterized viral IL-6 gene. Northern blot analysis revealed that the ORF11 gene is lytic viral gene with delayed-early expression kinetics. We have determined the 5′ and 3′ untranslated region of the unspliced ORF11 transcript and identified both the transcription start site and the transcription termination site. Core promoter region, representing ORF11 promoter activity, was mapped to a 160 nt fragment 5′ most proximal to the transcription start site. A functional TATA box was identified in the core promoter region. We also found that the characterized ORF11 gene promoter region is not responsive to Rta, the KSHV lytic switch protein. Our data help to elucidate transcription regulation of the KSHV ORF11 gene and to understand the biology of ORF11 in KSHV life cycle.

Compressive mechanical stimuli are crucial in regenerating cartilage with tissue engineering, which creates a need for scaffolds that can maintain their mechanical integrity while delivering mechanical signals to adherent cells during strain applications. With these goals in mind, the aim of this study was to develop a mechano-active scaffold that facilitated effective cartilaginous tissue formation under dynamic physiological environments. Using a gel-pressing method, we fabricated a biodegradable and highly-elastic scaffold from poly(L-lactide-co-epsilon-caprolactone) (PLCL; 5:5), with 85% porosity and a 300-500-microm pore size, and we compared it to control scaffolds made of rigid polylactide (PLA) or poly(lactide-co-glycolide) (PLGA). After tensile mechanical tests and recovery tests confirmed the elasticity of the PLCL scaffolds, we seeded them with rabbit chondrocytes, cultured them in vitro, and subcutaneously implanted them into nude mice for up to eight weeks. The PLCL scaffolds possessed a completely rubber-like elasticity, were easily twisted and bent, and exhibited an almost complete (over 97%) recovery from applied strain (up to 500%); the control PLA scaffolds showed little recovery. In vitro and in vivo accumulations of extracellular matrix on the cell-PLCL constructs demonstrated that they could not only sustain but also significantly enhance chondrogenic differentiation. Moreover, the mechanical stimulation of the dynamic in vivo environment promoted deposition of the chondral extracellular matrix onto the PLCL. In contrast, on the PLA scaffolds, most of the chondrocytes had de-differentiated and formed fibrous tissues. In a rabbit defect model, the groups treated with PLCL scaffolds exhibited significantly enhanced cartilage regeneration compared to groups harboring an empty control or PLGA scaffolds. These results indicated that the mechano-active PLCL scaffolds effectively delivered mechanical signals associated with biological environments to adherent chondrocytes, suggesting that these elastic PLCL scaffolds could successfully be used for cartilage regeneration.

Background MCAO has been widely used to produce ischemic brain lesions. The lesions induced by MCAO tend to be variable in size because of the variance in the collateral blood supply found in the mouse brain. Methods We modified the rat photothrombosis model for use in mice. Male C57BL/6 mice were subjected to focal cerebral ischemia by photothrombosis of cortical microvessels. Cerebral infarction was produced by intraperitoneal injection of rose bengal, a photosensitive dye, and by focal illumination through the skull. Motor impairment was assessed by the accelerating rotarod and staircase tests. The brain was perfusion fixed for histologic determination of infarct volume 4 weeks after stroke. Results The lesion was located in the frontal and parietal cortex and the underlying white matter was partly affected. A relatively constant infarct volume was achieved 1 month after photothrombosis. The presence of the photothrombotic lesion significantly impaired the motor performance as measured by the rotarod and staircase tests. Our findings show that photothrombotic infarction in mice is highly reproducible in size and location. Conclusion This procedure can provide a simple model of cerebral infarction for a unilateral motor cortex lesion. In addition, it can provide a suitable model for the study of potential neuroprotective and therapeutic agents in human stroke.

DNA double-strand breaks can be introduced by exogenous agents or during normal cellular processes. Genes belonging to the RAD52 epistasis group are known to repair these breaks in budding yeast. Among these genes, RAD52 plays a central role in homologous recombination and DNA double-strand break repair. Despite its importance, its mechanism of action is not yet clear. It is known, however, that the human homologue of Rad52 is capable of binding to DNA ends in vitro. Herein, we show that Rad22 protein, a Rad52 homologue in the fission yeast Schizosaccharomyces pombe, can similarly bind to DNA ends at double-strand breaks. This end-binding ability was demonstrated in vitro by electron microscopy and by protection from exonuclease attack. We also showed that Rad22 specifically binds near double-strand break associated with mating type switching in vivo by chromatin immunoprecipitation analysis. This is the first evidence that a recombinational protein directly binds to DNA double-strand breaks in vivo.