EF1α and RPL13a represent normalization genes suitable for RT-qPCR analysis of bone marrow derived mesenchymal stem cells

<p>Abstract</p> <p>Background</p> <p>RT-qPCR analysis is a widely used method for the analysis of mRNA expression throughout the field of mesenchymal stromal cell (MSC) research. Comparison between MSC studies, both <it>in vitro </it>and <it>in vivo</it>, are challenging due to the varied methods of RT-qPCR data normalization and analysis. Therefore, this study focuses on putative housekeeping genes for the normalization of RT-qPCR data between heterogeneous commercially available human MSC, compared with more homogeneous populations of MSC such as MIAMI and RS-1 cells.</p> <p>Results</p> <p>Eight genes including; <it>ACTB, B2M, EF1α, GAPDH, RPL13a, YWHAZ, UBC </it>and <it>HPRT1 </it>were tested as possible housekeeping genes based on their expression level and variability. <it>EF1α </it>and <it>RPL13a </it>were validated for RT-qPCR analysis of MIAMI cells during expansion in varied oxygen tensions, endothelial differentiation, neural precursor enrichment, and during the comparison with RS-1 cells and commercially available MSC. <it>RPL13a </it>and <it>YWHAZ </it>were validated as normalization genes for the cross-species analysis of MIAMI cells in an animal model of focal ischemia. <it>GAPDH</it>, which is one of the most common housekeeping genes used for the normalization of RT-qPCR data in the field of MSC research, was found to have the highest variability and deemed not suitable for normalization of RT-qPCR data.</p> <p>Conclusions</p> <p>In order to make comparisons between heterogeneous MSC populations, as well as adult stem cell like MSC which are used in different laboratories throughout the world, it is important to have a standardized, reproducible set of housekeeping genes for RT-qPCR analysis. In this study we demonstrate that <it>EF1α</it>, <it>RPL13a </it>and <it>YWHAZ </it>are suitable genes for the RT-qPCR analysis and comparison of several sources of human MSC during <it>in vitro </it>characterization and differentiation as well as in an <it>ex vivo</it> animal model of global cerebral ischemia. This will allow for the comparative RT-qPCR analysis of multiple MSC populations with the goal of future use in animal models of disease as well as tissue repair.</p

Time-Expanded F-OTDR based on binary sequences

In this paper, the capabilities of time-expanded phase-sensitive optical time-domain reflectometry (TE F-OTDR) using binary sequences are demonstrated. We present a highly flexible and integrable TE F-OTDR approach that allows a customized distributed optical fiber sensor (range, spatial resolution, and acoustic sampling) by simply changing the length of the binary sequence and the reference clock frequencies of the binary sequence generators. The here presented architecture eliminates the need for the cumbersome arbitrary signal generators used to date to create the dual-comb spectra for interrogating the fiber. In this approach, the use of large binary sequences allows us to obtain dual combs in a simple and cost-effective way. Spatial resolution of ~1 cm is achieved, attaining ~15, 000 independent measurements points along the interrogated fiber, with a capability of sensing ~30, 000 measurements points

"I won't be staying here for long": a qualitative study on the retention of migrant nurses in Ireland

<p>Abstract</p> <p>Background</p> <p>Although international nurse recruitment campaigns have succeeded in attracting large numbers of migrant nurses to countries such as Ireland, where domestic supply has not kept pace with demand, the long-term success of such initiatives from a workforce planning perspective will depend on the extent to which these nurses can be retained in destination countries.</p> <p>Methods</p> <p>This paper draws on qualitative, in-depth interviews undertaken with 21 migrant nurses in Ireland, focusing specifically on their future migration intentions.</p> <p>Results</p> <p>Our findings indicate that more than half of the respondents are considering migration onwards, for the most part because the destination country has failed to provide them with sufficient stability, particularly in terms of citizenship and family reunification. In considering onward migration, factors outside the health system were of most concern to those interviewed.</p> <p>Conclusion</p> <p>This demonstrates the need for destination countries to take a broader and more long-term approach to international nurse recruitment, rather than regarding it as an inexpensive way to fill gaps within the health care system.</p

Platelet-Rich Plasma Promotes the Proliferation of Human Muscle Derived Progenitor Cells and Maintains Their Stemness

Human muscle-derived progenitor cells (hMDPCs) offer great promise for muscle cell-based regenerative medicine; however, prolonged ex-vivo expansion using animal sera is necessary to acquire sufficient cells for transplantation. Due to the risks associated with the use of animal sera, the development of a strategy for the ex vivo expansion of hMDPCs is required. The purpose of this study was to investigate the efficacy of using platelet-rich plasma (PRP) for the ex-vivo expansion of hMDPCs. Pre-plated MDPCs, myoendothelial cells, and pericytes are three populations of hMDPCs that we isolated by the modified pre-plate technique and Fluorescence Activated Cell Sorting (FACS), respectively. Pooled allogeneic human PRP was obtained from a local blood bank, and the effect that thrombin-activated PRP-releasate supplemented media had on the ex-vivo expansion of the hMDPCs was tested against FBS supplemented media, both in vitro and in vivo. PRP significantly enhanced short and long-term cell proliferation, with or without FBS supplementation. Antibody-neutralization of PDGF significantly blocked the mitogenic/proliferative effects that PRP had on the hMDPCs. A more stable and sustained expression of markers associated with stemness, and a decreased expression of lineage specific markers was observed in the PRP-expanded cells when compared with the FBS-expanded cells. The in vitro osteogenic, chondrogenic, and myogenic differentiation capacities of the hMDPCs were not altered when expanded in media supplemented with PRP. All populations of hMDPCs that were expanded in PRP supplemented media retained their ability to regenerate myofibers in vivo. Our data demonstrated that PRP promoted the proliferation and maintained the multi-differentiation capacities of the hMDPCs during ex-vivo expansion by maintaining the cells in an undifferentiated state. Moreover, PDGF appears to be a key contributing factor to the beneficial effect that PRP has on the proliferation of hMDPCs. © 2013 Li et al

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Light regulation of metabolic pathways in fungi

Light represents a major carrier of information in nature. The molecular machineries translating its electromagnetic energy (photons) into the chemical language of cells transmit vital signals for adjustment of virtually every living organism to its habitat. Fungi react to illumination in various ways, and we found that they initiate considerable adaptations in their metabolic pathways upon growth in light or after perception of a light pulse. Alterations in response to light have predominantly been observed in carotenoid metabolism, polysaccharide and carbohydrate metabolism, fatty acid metabolism, nucleotide and nucleoside metabolism, and in regulation of production of secondary metabolites. Transcription of genes is initiated within minutes, abundance and activity of metabolic enzymes are adjusted, and subsequently, levels of metabolites are altered to cope with the harmful effects of light or to prepare for reproduction, which is dependent on light in many cases. This review aims to give an overview on metabolic pathways impacted by light and to illustrate the physiological significance of light for fungi. We provide a basis for assessment whether a given metabolic pathway might be subject to regulation by light and how these properties can be exploited for improvement of biotechnological processes

Internationally recruited nurses from India and the Philippines in the United Kingdom: the decision to emigrate.

BACKGROUND: The United Kingdom has recruited nurses from countries with a reported surplus in their nursing workforce, such as India and the Philippines. However, little is known about the decision to emigrate made by nurses from these countries. One theory suggests that individuals weigh the benefits and costs of migration: the push and pull factors. This paper challenges the restricted economic focus of this predominant theory and compares the diverse motivations of nurses from different countries as well as those of nurses with previous migratory experience and first-time migrants. METHODS: This research was undertaken in a National Health Service acute trust in London by means of a qualitative interpretative approach. Data were collected through face-to-face longitudinal and cross-sectional interviews with internationally recruited nurses from India (n=6) and the Philippines (n=15); and analysis of their narratives was used to generate data about their expectations and experiences. Data were analysed by means of a framework approach that allowed for intra-case and cross-case analysis. RESULTS: From an individual perspective, nurses in this study reported economic reasons as the main trigger for migration in the first instance. Yet this doesn't entirely explain the decision to move from previous migratory destinations (e.g. Saudi Arabia) where economic needs are already fulfilled. In these cases migration is influenced by professional and social aspirations that highlight the influence of the cultural environment--specifically some religious and gender-related issues. Family support and support from migratory networks in the country of origin and destination were also important elements conducive to and supportive of migration. Nurses from India report coming to the United Kingdom to stay, while Filipina nurses come as temporary migrants sending remittances to support their families in the Philippines. CONCLUSION: This study shows the diverse motivations of nurses from different countries and with different migratory backgrounds and provides evidence that factors other than economic factors influence nurses' decision to emigrate. This information can help developing countries increase retention of this essential and often scarce resource and can also help the United Kingdom's National Health Service to improve the experience of internationally recruited nurses and therefore increase their retention in the United Kingdom

Early pregnancy diagnosis in goats by determination of pregnancy-associated glycoprotein concentrations in plasma samples.

peer reviewedDifferent RIA systems available for measuring the concentrations of pregnancy-associated glycoproteins (PAGs) in dairy goats were compared in order to evaluate their accuracy in early pregnancy diagnosis. Plasma concentrations of PAGs were determined by 3 heterologous RIA systems with a bovine PAG standard and tracer in combination with antisera anti-ovine PAG (RIA 1), anti-caprine PAG55 + 62 (RIA 2), anti-caprine PAG55 + 59 (RIA 3), and by 2 homologous RIA systems that employed caprine PAG55 + 62 and caprine PAG55 + 59 and their specific antisera (RIAs 4 and 5, respectively). In all of the RIAs, the mean concentrations of PAGs were significantly higher (P 99% for RIAs 2, 3, 4 and 5. The RIAs for PAGs depend on proteins from the placenta being present in maternal plasma and require only a single sample of blood, to distinguish pregnant goats from those that fail to return to estrus for other reasons. The homologous and semi-heterologous assays are highly accurate as early as Day 21 of pregnancy

Milk production correlates negatively with plasma levels of pregnancy-associated glycoprotein (PAG) during the early fetal period in high producing dairy cows with live fetuses

This study was designed to establish possible factors affecting plasma pregnancy-associated glycoprotein (PAG) concentrations during early pregnancy in high producing dairy cows with live fetuses. Blood samples were obtained on days 35, 42, 49, 56 and 63 of gestation from 80 lactating cows in two herds carrying live fetuses. Radioimmunoassay systems were used to determine PAG (RIA-497 and RIA-706) and progesterone concentrations. We evaluated the effects on PAG concentrations of herd, lactation number, sire of fetus, day of gestation, fetus number, plasma progesterone and milk production at each time point established, along with possible paired interactions. Mean milk production per cow approached 41 kg during the study period. PAG concentrations were not affected by herd, lactation number or plasma progesterone concentration. Significant positive effects on PAG concentrations were shown by the gestation day, and the interaction between day of gestation and twin pregnancy. Significant differences between bulls and a significant negative correlation between milk production and PAG values on day 63 of pregnancy were also detected. Proportions of blood samples showing undetectable PAG levels and false negative diagnoses throughout the study period were significantly higher (P < 0.001) using the RIA-497 system (2.5% and 5.3%, respectively) compared to RIA-706 (0% and 0.8%, respectively). Our findings suggest that PAG concentrations during the early fetal period are related to the day of gestation, milk production, number of fetuses and sire of fetus in high producing dairy cows. Under our working conditions, the RIA-706 method was better at detecting plasma PAG molecules than the RIA-497 system. (c) 2005 Elsevier Inc. All rights reserved