Differential regulation of morphology and stemness of mouse embryonic stem cells by substrate stiffness and topography

The maintenance of stem cell pluripotency or sternness is crucial to embryonic development and differentiation. The mechanical or physical microenvironment of stem cells, which includes extracellular matrix stiffness and topography, regulates cell morphology and stemness. Although a growing body of evidence has shown the importance of these factors in stem cell differentiation, the impact of these biophysical or biomechanical regulators remains insufficiently characterized. In the present study, we applied a micro-fabricated polyacrylamide hydrogel substrate with two elasticities and three topographies to systematically test the morphology, proliferation, and sternness of mESCs. The independent or combined impact of the two factors on specific cell functions was analyzed. Cells are able to grow effectively on both polystyrene and polyacrylamide substrates in the absence of feeder cells. Substrate stiffness is predominant in preserving stemness by enhancing Oct-4 and Nanog expression on a soft polyacrylamide substrate. Topography is also a critical factor for manipulating sternness via the formation of a relatively flattened colony on a groove or pillar substrate and a spheroid colony on a hexagonal substrate. Although topography is less effective on soft substrates, it plays a role in retaining cell sternness on stiff, hexagonal or pillar-shaped substrates. mESCs also form, in a timely manner, a 3D structure on groove or hexagonal substrates. These results further the understanding of stem cell morphology and stemness in a microenvironment that mimics physiological conditions. (C) 2014 Elsevier Ltd. All rights reserved

Planck Scale Boundary Conditions and the Higgs Mass

If the LHC does only find a Higgs boson in the low mass region and no other new physics, then one should reconsider scenarios where the Standard Model with three right-handed neutrinos is valid up to Planck scale. We assume in this spirit that the Standard Model couplings are remnants of quantum gravity which implies certain generic boundary conditions for the Higgs quartic coupling at Planck scale. This leads to Higgs mass predictions at the electroweak scale via renormalization group equations. We find that several physically well motivated conditions yield a range of Higgs masses from 127-142 GeV. We also argue that a random quartic Higgs coupling at the Planck scale favors M_H > 150 GeV, which is clearly excluded. We discuss also the prospects for differentiating different boundary conditions imposed for \lambda(M_{pl}) at the LHC. A striking example is M_H = 127\pm 5 GeV corresponding to \lambda(M_{pl})=0, which would imply that the quartic Higgs coupling at the electroweak scale is entirely radiatively generated.Comment: 12 pages, 5 figures; references added and other minor improvements, matches version published in JHE

Maximal Spontaneous Photon Emission and Energy Loss from Free Electrons

Free electron radiation such as Cerenkov, Smith--Purcell, and transition radiation can be greatly affected by structured optical environments, as has been demonstrated in a variety of polaritonic, photonic-crystal, and metamaterial systems. However, the amount of radiation that can ultimately be extracted from free electrons near an arbitrary material structure has remained elusive. Here we derive a fundamental upper limit to the spontaneous photon emission and energy loss of free electrons, regardless of geometry, which illuminates the effects of material properties and electron velocities. We obtain experimental evidence for our theory with quantitative measurements of Smith--Purcell radiation. Our framework allows us to make two predictions. One is a new regime of radiation operation---at subwavelength separations, slower (nonrelativistic) electrons can achieve stronger radiation than fast (relativistic) electrons. The second is a divergence of the emission probability in the limit of lossless materials. We further reveal that such divergences can be approached by coupling free electrons to photonic bound states in the continuum (BICs). Our findings suggest that compact and efficient free-electron radiation sources from microwaves to the soft X-ray regime may be achievable without requiring ultrahigh accelerating voltages.Comment: 7 pages, 4 figure

Gastric Lavage in Acute Organophosphorus Pesticide poisoning (GLAOP) – a randomised controlled trial of multiple vs. single gastric lavage in unselected acute organophosphorus pesticide poisoning

BACKGROUND: Organophosphorus (OP) pesticide poisoning is the most common form of pesticide poisoning in many Asian countries. Guidelines in western countries for management of poisoning indicate that gastric lavage should be performed only if two criteria are met: within one hour of poison ingestion and substantial ingested amount. But the evidence on which these guidelines are based is from medicine overdoses in developed countries and may be irrelevant to OP poisoning in Asia. Chinese clinical experience suggests that OP remains in the stomach for several hours or even days after ingestion. Thus, there may be reasons for doing single or multiple gastric lavages for OP poisoning. There have been no randomised controlled trials (RCTs) to assess this practice of multiple lavages. Since it is currently standard therapy in China, we cannot perform a RCT of no lavage vs. a single lavage vs. multiple lavages. We will compare a single gastric lavage with three gastric lavages as the first stage to assess the role of gastric lavage in OP poisoning. METHODS/DESIGN: We have designed an RCT assessing the effectiveness of multiple gastric lavages in adult OP self-poisoning patients admitted to three Chinese hospitals within 12 hrs of ingestion. Patients will be randomised to standard treatment plus either a single gastric lavage on admission or three gastric lavages at four hour intervals. The primary outcome is in-hospital mortality. Analysis will be on an intention-to-treat basis. On the basis of the historical incidence of OP at the study sites, we expect to enroll 908 patients over three years. This projected sample size provides sufficient power to evaluate the death rate; and a variety of other exposure and outcome variables, including particular OPs and ingestion time. Changes of OP level will be analyzed in order to provide some toxic kinetic data. DISCUSSION: the GLAOP study is a novel, prospective cohort study that will explore to the toxic kinetics of OP and effects of gastric lavage on it. Given the poor information about the impact of gastric lavage on clinical outcomes for OP patients, this study can provide important information to inform clinical practice

High-throughput genotyping of single nucleotide polymorphisms with rolling circle amplification

BACKGROUND: Single nucleotide polymorphisms (SNPs) are the foundation of powerful complex trait and pharmacogenomic analyses. The availability of large SNP databases, however, has emphasized a need for inexpensive SNP genotyping methods of commensurate simplicity, robustness, and scalability. We describe a solution-based, microtiter plate method for SNP genotyping of human genomic DNA. The method is based upon allele discrimination by ligation of open circle probes followed by rolling circle amplification of the signal using fluorescent primers. Only the probe with a 3' base complementary to the SNP is circularized by ligation. RESULTS: SNP scoring by ligation was optimized to a 100,000 fold discrimination against probe mismatched to the SNP. The assay was used to genotype 10 SNPs from a set of 192 genomic DNA samples in a high-throughput format. Assay directly from genomic DNA eliminates the need to preamplify the target as done for many other genotyping methods. The sensitivity of the assay was demonstrated by genotyping from 1 ng of genomic DNA. We demonstrate that the assay can detect a single molecule of the circularized probe. CONCLUSIONS: Compatibility with homogeneous formats and the ability to assay small amounts of genomic DNA meets the exacting requirements of automated, high-throughput SNP scoring

Selectivity of biopolymer membranes using HepG2 cells

Bioartificial liver (BAL) system has emerged as an alternative treatment to bridge acute liver failure to either liver transplantation or liver regeneration. One of the main reasons that the efficacy of the current BAL systems was not convincing in clinical trials is attributed to the lack of friendly interface between the membrane and the hepatocytes in liver bioreactor, the core unit of BAL system. Here, we systematically compared the biological responses of hepatosarcoma HepG2 cells seeded on eight, commercially available biocompatible membranes made of acetyl cellulose&ndash;nitrocellulose mixed cellulose (CA&ndash;NC), acetyl cellulose (CA), nylon (JN), polypropylene (PP), nitrocellulose (NC), polyvinylidene fluoride (PVDF), polycarbonate (PC) and polytetrafluoroethylene (PTFE). Physicochemical analysis and mechanical tests indicated that CA, JN and PP membranes yield high adhesivity and reasonable compressive and/or tensile features with friendly surface topography for cell seeding. Cells prefer to adhere on CA, JN, PP or PTFE membranes with high proliferation rate in spheriod-like shape. Actin, albumin and cytokeratin 18 expressions are favorable for cells on CA or PP membrane, whereas protein filtration is consistent among all the eight membranes. These results further the understandings of cell growth, morphology and spreading, as well as protein filtration on distinct membranes in designing a liver bioreactor.</p

A 115-bp MethyLight assay for detection of p16 (CDKN2A) methylation as a diagnostic biomarker in human tissues

<p>Abstract</p> <p>Background</p> <p><it>p16 </it>Methylation is a potential biomarker for prediction of malignant transformation of epithelial dysplasia. A probe-based, quantitative, methylation-specific PCR (MSP) called MethyLight may become an eligible method for detecting this marker clinically. We studied oral mucosa biopsies with epithelial dysplasia from 78 patients enrolled in a published 4-years' followup cohort, in which cancer risk for patients with <it>p16 </it>methylation-positive dysplasia was significantly higher than those without <it>p16 </it>methylation (by 150-bp MSP and bisulfite sequencing; +133 ~ +283, transcription starting site, +1). The <it>p16 </it>methylation status in samples (<it>N </it>= 102) containing sufficient DNA was analyzed by the 70-bp classic (+238 ~ +307) and 115-bp novel (+157 ~ +272) MethyLight assays, respectively.</p> <p>Results</p> <p><it>p16 </it>Methylation was detectable in 75 samples using the classic MethyLight assay. The methylated-<it>p16 </it>positive rate and proportion of methylated-<it>p16 </it>by the MethyLight in MSP-positive samples were higher than those in MSP-negative samples (positive rate: 37/44 vs. 38/58, <it>P</it>=0.035, two-sided; proportion [median]: 0.78 vs. 0.02, <it>P <</it>0.007). Using the published results of MSP as a golden standard, we found sensitivity, specificity, and accuracy for this MethyLight assay to be 70.5%, 84.5%, and 55.0%, respectively. Because amplicon of the classic MethyLight procedure only partially overlapped with the MSP amplicon, we further designed a 115-bp novel MethyLight assay in which the amplicon on the sense-strand fully overlapped with the MSP amplicon on the antisense-strand. Using the 115-bp MethyLight assay, we observed methylated-<it>p16 </it>in 26 of 44 MSP-positive samples and 2 of 58 MSP-negative ones (<it>P </it>= 0.000). These results were confirmed with clone sequencing. Sensitivity, specificity, and accuracy using the 115-bp MethyLight assay were 59.1%, 98.3%, and 57.4%, respectively. Significant differences in the oral cancer rate were observed during the followup between patients (≥60 years) with and without methylated-<it>p16 </it>as detected by the 115-bp MethyLight assay (6/8 vs. 6/22, P = 0.034, two-sided).</p> <p>Conclusions</p> <p>The 115-bp MethyLight assay is a useful and practical assay with very high specificity for the detection of <it>p16 </it>methylation clinically.</p

Disparities and risks of sexually transmissible infections among men who have sex with men in China: a meta-analysis and data synthesis.

BACKGROUND: Sexually transmitted infections (STIs), including Hepatitis B and C virus, are emerging public health risks in China, especially among men who have sex with men (MSM). This study aims to assess the magnitude and risks of STIs among Chinese MSM. METHODS: Chinese and English peer-reviewed articles were searched in five electronic databases from January 2000 to February 2013. Pooled prevalence estimates for each STI infection were calculated using meta-analysis. Infection risks of STIs in MSM, HIV-positive MSM and male sex workers (MSW) were obtained. This review followed the PRISMA guidelines and was registered in PROSPERO. RESULTS: Eighty-eight articles (11 in English and 77 in Chinese) investigating 35,203 MSM in 28 provinces were included in this review. The prevalence levels of STIs among MSM were 6.3% (95% CI: 3.5-11.0%) for chlamydia, 1.5% (0.7-2.9%) for genital wart, 1.9% (1.3-2.7%) for gonorrhoea, 8.9% (7.8-10.2%) for hepatitis B (HBV), 1.2% (1.0-1.6%) for hepatitis C (HCV), 66.3% (57.4-74.1%) for human papillomavirus (HPV), 10.6% (6.2-17.6%) for herpes simplex virus (HSV-2) and 4.3% (3.2-5.8%) for Ureaplasma urealyticum. HIV-positive MSM have consistently higher odds of all these infections than the broader MSM population. As a subgroup of MSM, MSW were 2.5 (1.4-4.7), 5.7 (2.7-12.3), and 2.2 (1.4-3.7) times more likely to be infected with chlamydia, gonorrhoea and HCV than the broader MSM population, respectively. CONCLUSION: Prevalence levels of STIs among MSW were significantly higher than the broader MSM population. Co-infection of HIV and STIs were prevalent among Chinese MSM. Integration of HIV and STIs healthcare and surveillance systems is essential in providing effective HIV/STIs preventive measures and treatments. TRIAL REGISTRATION: PROSPERO NO: CRD42013003721

Transplacentally Acquired Maternal Antibody against Hepatitis B Surface Antigen in Infants and its Influence on the Response to Hepatitis B Vaccine

BACKGROUND: Passively acquired maternal antibodies in infants may inhibit active immune responses to vaccines. Whether maternal antibody against hepatitis B surface antigen (anti-HBs) in infants may influence the long-term immunogenicity of hepatitis B vaccine remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: Totally 338 pairs of mothers and children were enrolled. All infants were routinely vaccinated against hepatitis B based on 0-, 1- and 6-month schedule. We characterized the transplacental transfer of maternal anti-HBs, and compared anti-HBs response in children of mothers with or without anti-HBs. In a prospective observation, all 63 anti-HBs positive mothers transferred anti-HBs to their infants; 84.1% of the infants had higher anti-HBs concentrations than their mothers. One and half years after vaccination with three doses of hepatitis B vaccine, the positive rate and geometric mean concentration (GMC) of anti-HBs in 32 infants with maternal anti-HBs were comparable with those in 32 infants without maternal antibody (90.6% vs 87.5%, P = 0.688, and 74.5 vs 73.5 mIU/ml, P = 0.742, respectively). In a retrospective analysis, five and half years after vaccination with three doses vaccine, the positive rates of anti-HBs in 88 children of mothers with anti-HBs ≥1000 mIU/ml, 94 children of mothers with anti-HBs 10-999 mIU/ml, and 61 children of mothers with anti-HBs <10 mIU/ml were 72.7%, 69.2%, and 63.9% (P = 0.521), respectively; anti-HBs GMC in these three groups were 38.9, 43.9, and 31.7 mIU/ml (P = 0.726), respectively. CONCLUSIONS/SIGNIFICANCE: The data demonstrate that maternal anti-HBs in infants, even at high concentrations, does not inhibit the long-term immunogenicity of hepatitis B vaccine. Thus, current hepatitis B vaccination schedule for infants will be still effective in the future when most infants are positive for maternal anti-HBs due to the massive vaccination against hepatitis B