Preconditioning of mesenchymal stromal cells with low-intensity ultrasound: influence on chondrogenesis and directed SOX9 signaling pathways

Background: Continuous low-intensity ultrasound (cLIUS) facilitates the chondrogenic differentiation of human mesenchymal stromal cells (MSCs) in the absence of exogenously added transforming growth factor-beta (TGFβ) by upregulating the expression of transcription factor SOX9, a master regulator of chondrogenesis. The present study evaluated the molecular events associated with the signaling pathways impacting SOX9 gene and protein expression under cLIUS. Methods: Human bone marrow-derived MSCs were exposed to cLIUS stimulation at 14 kPa (5 MHz, 2.5 Vpp) for 5 min. The gene and protein expression of SOX9 was evaluated. The specificity of SOX9 upregulation under cLIUS was determined by treating the MSCs with small molecule inhibitors of select signaling molecules, followed by cLIUS treatment. Signaling events regulating SOX9 expression under cLIUS were analyzed by gene expression, immunofluorescence staining, and western blotting. Results: cLIUS upregulated the gene expression of SOX9 and enhanced the nuclear localization of SOX9 protein when compared to non-cLIUS-stimulated control. cLIUS was noted to enhance the phosphorylation of the signaling molecule ERK1/2. Inhibition of MEK/ERK1/2 by PD98059 resulted in the effective abrogation of cLIUS-induced SOX9 expression, indicating that cLIUS-induced SOX9 upregulation was dependent on the phosphorylation of ERK1/2. Inhibition of integrin and TRPV4, the upstream cell-surface effectors of ERK1/2, did not inhibit the phosphorylation of ERK1/2 and therefore did not abrogate cLIUS-induced SOX9 expression, thereby suggesting the involvement of other mechanoreceptors. Consequently, the effect of cLIUS on the actin cytoskeleton, a mechanosensitive receptor regulating SOX9, was evaluated. Diffused and disrupted actin fibers observed in MSCs under cLIUS closely resembled actin disruption by treatment with cytoskeletal drug Y27632, which is known to increase the gene expression of SOX9. The upregulation of SOX9 under cLIUS was, therefore, related to cLIUS-induced actin reorganization. SOX9 upregulation induced by actin reorganization was also found to be dependent on the phosphorylation of ERK1/2. Conclusions: Collectively, preconditioning of MSCs by cLIUS resulted in the nuclear localization of SOX9, phosphorylation of ERK1/2 and disruption of actin filaments, and the expression of SOX9 was dependent on the phosphorylation of ERK1/2 under cLIUS

Comparison of the Commercial Color LCD and the Medical Monochrome LCD Using Randomized Object Test Patterns

Workstations and electronic display devices in a picture archiving and communication system (PACS) provide a convenient and efficient platform for medical diagnosis. The performance of display devices has to be verified to ensure that image quality is not degraded. In this study, we designed a set of randomized object test patterns (ROTPs) consisting of randomly located spheres with various image characteristics to evaluate the performance of a 2.5 mega-pixel (MP) commercial color LCD and a 3 MP diagnostic monochrome LCD in several aspects, including the contrast, resolution, point spread effect, and noise. The ROTPs were then merged into 120 abdominal CT images. Five radiologists were invited to review the CT images, and receiver operating characteristic (ROC) analysis was carried out using a five-point rating scale. In the high background patterns of ROTPs, the sensitivity performance was comparable between both monitors in terms of contrast and resolution, whereas, in the low background patterns, the performance of the commercial color LCD was significantly poorer than that of the diagnostic monochrome LCD in all aspects. The average area under the ROC curve (AUC) for reviewing abdominal CT images was 0.717±0.0200 and 0.740±0.0195 for the color monitor and the diagnostic monitor, respectively. The observation time (OT) was 145±27.6 min and 127±19.3 min, respectively. No significant differences appeared in AUC (p = 0.265) and OT (p = 0.07). The overall results indicate that ROTPs can be implemented as a quality control tool to evaluate the intrinsic characteristics of display devices. Although there is still a gap in technology between different types of LCDs, commercial color LCDs could replace diagnostic monochrome LCDs as a platform for reviewing abdominal CT images after monitor calibration

In situ study of fiber structure development of poly(butylene terephthalate) in a continuous laser-heated drawing process

The structural development of poly(butylene terephthalate) (PBT) fibers was analyzed using in situ wide angle X-ray diffraction and fiber temperature measurements during CO2 laser-heated drawing, in which the necking position on the running fiber could be fixed by CO2 laser irradiation. The measured parameters were determined as functions of the elapsed time after necking with a time resolution of 0.3 ms. The as-spun PBT fibers, which exhibited a low-oriented alpha-crystalline structure, were drawn to a draw ratio of 5 using laser heating. The (001') reflection, which indicates a quasi-smectic fibrillar structure, was not observed before crystallization in contrast to measurements of poly(ethylene terephthalate) (PET) and poly(ethylene 2,6-naphthalene dicarboxylate) (PEN). The alpha-crystal was transformed into an oriented beta-form crystal at the necking position, and the developed beta-crystallites exhibited increased size and altered orientation <2 ms after necking. The fiber temperature increased rapidly at around T-g, and the rearrangement of the beta-crystal primarily occurred as the fiber's temperature rose from 100 to 160 degrees C. The oriented beta-crystal of the drawn fiber transformed into the oriented beta-crystal when the drawing tension was released. Polymer Journal (2012) 44, 1030-1035; doi: 10.1038/pj.2012.65; published online 18 April 2012ArticlePOLYMER JOURNAL. 44(10):1030-1035 (2012)journal articl

Output from VIP cells of the mammalian central clock regulates daily physiological rhythms

The suprachiasmatic nucleus (SCN) circadian clock is critical for optimising daily cycles in mammalian physiology and behaviour. The roles of the various SCN cell types in communicating timing information to downstream physiological systems remain incompletely understood, however. In particular, while vasoactive intestinal polypeptide (VIP) signalling is essential for SCN function and whole animal circadian rhythmicity, the specific contributions of VIP cell output to physiological control remains uncertain. Here we reveal a key role for SCN VIP cells in central clock output. Using multielectrode recording and optogenetic manipulations, we show that VIP neurons provide coordinated daily waves of GABAergic input to target cells across the paraventricular hypothalamus and ventral thalamus, supressing their activity during the mid to late day. Using chemogenetic manipulation, we further demonstrate specific roles for this circuitry in the daily control of heart rate and corticosterone secretion, collectively establishing SCN VIP cells as influential regulators of physiological timing

Duplication of a well-conserved homeodomain-leucine zipper transcription factor gene in barley generates a copy with more specific functions

Three spikelets are formed at each rachis node of the cultivated barley (Hordeum vulgare ssp. vulgare) spike. In two-rowed barley, the central one is fertile and the two lateral ones are sterile, whereas in the six-rowed type, all three are fertile. This characteristic is determined by the allelic constitution at the six-rowed spike 1 (vrs1) locus on the long arm of chromosome 2H, with the recessive allele (vrs1) being responsible for the six-rowed phenotype. The Vrs1 (HvHox1) gene encodes a homeodomain-leucine zipper (HD-Zip) transcription factor. Here, we show that the Vrs1 gene evolved in the Poaceae via a duplication, with a second copy of the gene, HvHox2, present on the short arm of chromosome 2H. Micro-collinearity and polypeptide sequences were both well conserved between HvHox2 and its Poaceae orthologs, but Vrs1 is unique to the barley tribe. The Vrs1 gene product lacks a motif which is conserved among the HvHox2 orthologs. A phylogenetic analysis demonstrated that Vrs1 and HvHox2 must have diverged after the separation of Brachypodium distachyon from the Pooideae and suggests that Vrs1 arose following the duplication of HvHox2, and acquired its new function during the evolution of the barley tribe. HvHox2 was expressed in all organs examined but Vrs1 was predominantly expressed in immature inflorescence

Relationships between Levels of Serum IgE, Cell-Bound IgE, and IgE-Receptors on Peripheral Blood Cells in a Pediatric Population

Background: Elevated serum immunoglobulin (Ig) E is a diagnostic marker of immediate-type allergic reactions. We hypothesize that serum IgE does not necessarily reflect total body IgE because in vivo IgE can be bound to cell surface receptors such as FcεRI and FcεRII (CD23). The aim of this study was to analyze the relationships between levels of serum IgE, cell-bound IgE, and IgE-receptors on peripheral blood cells in a pediatric population. Methodology: Whole blood samples from 48 children (26 boys, 22 girls, mean age 10,3±5,4 years) were analyzed by flow cytometry for FcεRI, CD23, and cell-bound IgE on dendritic cells (CD11c+MHC class II+), monocytes (CD14+), basophils (CD123+MHC class II-) and neutrophils (myeloperoxidase+). Total serum IgE was measured by ELISA and converted into z-units to account for age-dependent normal ranges. Correlations were calculated using Spearman rank correlation test. Principal Findings: Dendritic cells, monocytes, basophils, and neutrophils expressed the high affinity IgE-receptor FcεRI. Dendritic cells and monocytes also expressed the low affinity receptor CD23. The majority of IgE-receptor positive cells carried IgE on their surface. Expression of both IgE receptors was tightly correlated with cell-bound IgE. In general, cell-bound IgE on FcεRI+ cells correlated well with serum IgE. However, some patients carried high amounts of cell-bound IgE despite low total serum IgE levels. Conclusion/Significance: In pediatric patients, levels of age-adjusted serum IgE, cell-bound IgE, and FcεRI correlate. Even in the absence of elevated levels of serum IgE, cell-bound IgE can be detected on peripheral blood cells in a subgroup of patients

Association of CCR2-CCR5 Haplotypes and CCL3L1 Copy Number with Kawasaki Disease, Coronary Artery Lesions, and IVIG Responses in Japanese Children

BACKGROUND: The etiology of Kawasaki Disease (KD) is enigmatic, although an infectious cause is suspected. Polymorphisms in CC chemokine receptor 5 (CCR5) and/or its potent ligand CCL3L1 influence KD susceptibility in US, European and Korean populations. However, the influence of these variations on KD susceptibility, coronary artery lesions (CAL) and response to intravenous immunoglobulin (IVIG) in Japanese children, who have the highest incidence of KD, is unknown. METHODOLOGY/PRINCIPAL FINDINGS: We used unconditional logistic regression analyses to determine the associations of the copy number of the CCL3L1 gene-containing duplication and CCR2-CCR5 haplotypes in 133 Japanese KD cases [33 with CAL and 25 with resistance to IVIG] and 312 Japanese controls without a history of KD. We observed that the deviation from the population average of four CCL3L1 copies (i.e., <or>four copies) was associated with an increased risk of KD and IVIG resistance (adjusted odds ratio (OR)=2.25, p=0.004 and OR=6.26, p=0.089, respectively). Heterozygosity for the CCR5 HHF*2 haplotype was associated with a reduced risk of both IVIG resistance (OR=0.21, p=0.026) and CAL development (OR=0.44, p=0.071). CONCLUSIONS/SIGNIFICANCE: The CCL3L1-CCR5 axis may play an important role in KD pathogenesis. In addition to clinical and laboratory parameters, genetic markers may also predict risk of CAL and resistance to IVIG

Circulating CD133+CD34+ progenitor cells inversely correlate with soluble ICAM-1 in early ischemic stroke patients

<p>Abstract</p> <p>Background and Purpose</p> <p>Both endothelial progenitor cells (EPC) and markers of neuroinflammation are candidate biomarkers for stroke severity and outcome prediction. A relationship between EPC and neuroinflammatory markers in early stroke is not fully elucidated. The objectives were to investigate correlations between EPC and neuroinflammation markers (adhesion molecules ICAM-1, VCAM-1, E-selectin, tumor necrosis factor (TNF)-α, interleukin (IL)-6, endothelin (ET)-1, markers of tissue injury (matrix metalloproteinases (MMP)-9 and tissue inhibitor of matrix metalloproteinases (TIMP)-1) in early stroke patients.</p> <p>Methods</p> <p>We prospectively recruited symptomatic patients with ischemic cerebrovascular disease. We assessed stroke severity by using of acute (diffusion-weighted imaging (DWI) and final lesion volumes (fluid attenuated inversion recovery (FLAIR). We measured serum soluble ICAM-1, VCAM-1, E-selectin, MMP-9, TIMP-1 and plasma TNF-α, IL-6, ET-1 by ELISA, and quantified EPC in mononuclear fraction of peripheral blood on days 1 and 3 in 17 patients (mean(SD) age 62(14), with admission National Institutes of Health Stroke Scale (NIHSS) 10(8)) selected from 175 patients with imaging confirmed ischemic stroke. Non-parametric statistics, univariate and multivariate analysis were used.</p> <p>Results</p> <p>Only ICAM-1 inversely correlated with EPC subset CD133+CD34+ on day 1 (Spearman r = -0.6, p < 0.01) and on day 3 (r = -0.967, p < 0.001). This correlation remained significant after adjustment for age and NIHSS (beta -0.992, p < 0.004), for glucose and systolic blood pressure (beta -0.86, p < 0.005), and for white blood cells and hematocrit (beta -1.057, p < 0.0001) on day 3. MMP-9 (r = 0.509, p < 0.04) and MMP-9/TIMP-1 (r = 0.59, p < 0.013) on day 1 correlated with acute lesion volume. Both IL-6 (r = 0.624, p < 0.01) and MMP-9/TIMP-1 (r = 0.56, p < 0.02) correlated with admission NIHSS.</p> <p>Conclusion</p> <p>Our study showed that high ICAM-1 is associated with low CD133+CD34+subset of EPC. Biomarkers of neuroinflammation may predict tissue injury and stroke severity in early ischemia.</p

A Small Peptide Modeled after the NRAGE Repeat Domain Inhibits XIAP-TAB1-TAK1 Signaling for NF-κB Activation and Apoptosis in P19 Cells

In normal growth and development, apoptosis is necessary to shape the central nervous system and to eliminate excess neurons which are not required for innervation. In some diseases, however, apoptosis can be either overactive as in some neurodegenerative disorders or severely attenuated as in the spread of certain cancers. Bone morphogenetic proteins (BMPs) transmit signals for regulating cell growth, differentiation, and apoptosis. Responding to BMP receptors stimulated from BMP ligands, neurotrophin receptor-mediated MAGE homolog (NRAGE) binds and functions with the XIAP-TAK1-TAB1 complex to activate p38MAPK and induces apoptosis in cortical neural progenitors. NRAGE contains a unique repeat domain that is only found in human, mouse, and rat homologs that we theorize is pivotal in its BMP MAPK role. Previously, we showed that deletion of the repeat domain inhibits apoptosis, p38MAPK phosphorylation, and caspase-3 cleavage in P19 neural progenitor cells. We also showed that the XIAP-TAB1-TAK1 complex is dependent on NRAGE for IKK-α/β phosphorylation and NF-κB activation. XIAP is a major inhibitor of caspases, the main executioners of apoptosis. Although it has been shown previously that NRAGE binds to the RING domain of XIAP, it has not been determined which NRAGE domain binds to XIAP. Here, we used fluorescence resonance energy transfer (FRET) to determine that there is a strong likelihood of a direct interaction between NRAGE and XIAP occurring at NRAGE's unique repeat domain which we also attribute to be the domain responsible for downstream signaling of NF-κB and activating IKK subunits. From these results, we designed a small peptide modeled after the NRAGE repeat domain which we have determined inhibits NF-κB activation and apoptosis in P19 cells. These intriguing results illustrate that the paradigm of the NRAGE repeat domain may hold promising therapeutic strategies in developing pharmaceutical solutions for combating harmful diseases involving excessive downstream BMP signaling, including apoptosis

Municipal health expectancy in Japan: decreased healthy longevity of older people in socioeconomically disadvantaged areas

BACKGROUND: Little is known about small-area variation in healthy longevity of older people and its socioeconomic correlates. This study aimed to estimate health expectancy at 65 years (HE65) at the municipal level in Japan, and to examine its relation to area socio-demographic conditions. METHODS: HE65 of municipalities (N = 3361) across Japan was estimated by a linear regression formula with life expectancy at 65 years and the prevalence of those certificated as needing nursing care. The relation between HE65 and area socio-demographic indicators was examined using correlation coefficients. RESULTS: The estimated HE65 (years) ranged from 13.13 to 17.39 for men and from 14.84 to 20.53 for women. HE65 was significantly positively correlated with the proportion of elderly and per capita income, and negatively correlated with the percentage of households of a single elderly person, divorce rate, and unemployment rate. These relations were stronger in large municipalities (with a population of more than 100,000) than in small and medium-size municipalities. CONCLUSION: A decrease in healthy longevity of older people was associated with a higher percentage of households of a single elderly person and divorce rate, and lower socioeconomic conditions. This study suggests that older people in urban areas are susceptible to socio-demographic factors, and a social support network for older people living in socioeconomically disadvantaged conditions should be encouraged