We also analyzed the content of

We also analyzed the content of HGF and angiopoietin-1 in these secretase signaling and found that intracellular content of HGF was 2.6 times lower in MSCs transfected with pre-miR-92a comparing to scramble transfected cells; however, angiopoietin content within MSCs did not change significantly (see Fig. 2). We collected conditioned medium of MSCs transfected with pre-miR-92a, anti-miR-92a or scramble oilgos, applied it to HUVEC and analyzed their viability. Conditioned medium of transfected MSCs did not affect the viability of HUVEC (see Fig. 3), which was about 90%. Addition of recombinant HGF but not angiopoietin-1 to the conditioned medium of MSCs transfected with pre-miR-92a restored its ability to stimulate the tube formation by HUVEC (see Fig. 4). We also examined if the suppressive effect of conditioned medium of MSCs, which overexpress miR-92a, could be mediated by a direct transfer of this microRNA to endothelial cells by extracellular vesicles. We removed these vesicles from conditioned medium by ultracentrifugation and analyzed the effect of cleared medium on tube formation. Removal of extracellular vesicles completely abrogated the ability of conditioned medium to induce tube formation (see Fig. 5). Data interpretation and discussion can be found in [1].
2 Experimental design, materials and methods
Acknowledgments This study was supported by subsidy #14.607.21.0045 from the Russian Ministry of Education and Science (GrantID RFMEFI60714×0004).
Data HeLa cells were transfected with pCIC-RPSAwt or pCIC-RPSAmut and incubated in control, oxidizing (with H2O2) or reducing (with NAC) conditions. RPSA expression was then analyzed by immunofluorescence. The expression was cytoplasmic and nuclear (Fig. 1). Overlap of RPSA and the 40S ribosomal subunit RPS6 was assessed by co-immunofluorescence in HeLa cells overexpressing RPSAwt or RPSAmut (Fig. 2). Western blot analysis of endogenous RPSA expression in ribosomal extracts was performed in HeLa cells in control or oxidizing (with H2O2) conditions (Fig. 3). H2O2 treatment decreased RPSA ribosomal expression.
Experimental design, materials and methods
Conflict of interest
Acknowledgments We thank Telmo Nunes for assistance with confocal imaging and Margarida Gama Carvalho for the RPS6 antibody. This work was supported by Fundação para a Ciência e a Tecnologia (FCT), Grants PTDC/BIA-PRO/101624/2008, PEst-OE/QUI/UI0612/2013 and UID/Multi/00612/2013. Filipe Vilas-Boas was supported by FCT, individual fellowship SFRH/BPD/74715/2010. Carla Real acknowledges Ciência 2008 Program (FCT). The funder had no role in study design; data collection, analysis, and interpretation; decision to publish, or preparation of the manuscript.
Specifications table
Value of the data
Data In order to investigate the effects of several adjustments to the preanalytical phase of quantitative cfDNA measurements, the growth medium of cultured cancer cells was used as a source of cfDNA. The data in this report was obtained by amplifying cfDNA with real-time PCR, after it had been extracted under different preanalytical conditions. The data is presented in a supplementary file as a single table, which includes several quantitative measurements of cfDNA following modifications to the standard protocol followed. These changes are described in Table 1.
Experimental design, materials and methods
Acknowledgments AB and JA were supported by post-graduate scholarships from the National Research Foundation (NRF), South Africa. The financial assistance of the NRF is hereby acknowledged. Opinions expressed and conclusions arrived at, are those of the authors and are not to be attributed to the NRF.
Specifications Table
Value of the data
Data
Acknowledgments We would like to thank the NIH AIDS Reagents Program for providing TZM-bl and U38 cells. This work was partly supported by Grants from the National Institutes of Health to G.D.K (MH081780 and MH094160).