Human dental pulp derived from exfoliated deciduous teeth has been described as a promising alternative source of multipotent stem cells. While these cells share certain similarities with mesenchymal stem-like cells (MSC) isolated from other tissues, basically they are still poorly characterized. In this study, for the first time, a proteomic map of abundantly expressed proteins in stromal cells derived from the dental pulp of human exfoliated deciduous teeth (SHED) was established. We also analyzed proteomic signatures of 2 clonal strains derived from SHEDs by single-cell cloning. The SHEDs were established from enzyme-disaggregated deciduous dental pulp from 6-year-old children. They had typical fibroblastoid morphology and high colony-forming efficiency index (16.4%). Cloning was performed at the second passage using limiting dilution in a 96-well plate (0.3 cell/well). Differentiation assessment revealed strong osteogenic but no adipogenic potential of the SHEDs in either clonal strain. The cells expressed characteristic antigens of MSC-like cells, including CD73, CD90, CD105, CD146, and did not express hematopoietic markers CD14, CD34, and CD45, as assessed with FACS analysis. For proteomic studies, cytosolic and nuclear proteins were analyzed with 2-dimensional gel electrophoresis (2-DE) and identified using matrix-assisted laser desorption/ionization (MALDI)-time of fl ight (TOF)-mass spectrometry (MS). All proteins were identified with high level of confidence (the lowest sequence coverage was 27%). Identification of highly expressed proteins in SHEDs revealed proteomic profiles very similar to that of MSC-like cells derived from other tissues. We also found a high degree of similarity between proteomic signatures of primary SHEDs and clonal cell strains. Thus, our data confirm a close resemblance between SHEDs and MSC-like cells from other tissues and may serve as starting point for creating-comprehensive proteomic maps.

A b s t r a c t In the study, honey from oilseed rape Brassica napus L., and both hand-collected (winter rape Visby and Cult) and bee-collected pollen of oilseed rape were analyzed for their proteome content, in order to see if any plant proteins were present to allow the proteo-typing of the oilseed rape honey. Proteins were fractionated by two-dimensional gel electrophoresis (2DE), stained by Coomassie blue and then analyzed by mass spectrometry. All identified proteins were divided into few groups due to their biological function. In 2DE gels with separated proteins from blossom honey, only bee (Apis mellifera) main proteins (Major royal jelly protein 1-5 and Glucosidase) were found. So we analyzed all proteins using gel-free based analysis with the SYNAPT G2 high definition mass spectrometry. We identified proteins that were present in both oilseed rape pollen and honey (Bna, Polygalacturonase, Non-specific lipid-transfer protein, GAPDH and others). We believe that these proteins are important for the nutritional value of plant pollen-enriched honey and further research is required on honey and honeybee pollen protein.Keywords: Brasica napus L., honey, honeybee pollen, proteomics, rape In recent years, there has been an increased demand for honeybee pollen as a healthy food supplement. The content of protein, fat, phosphorus, iron and vitamins (E, D, B 12 ) makes pollen's nutritive value comparable to dried beans or beef (Erlund, 2004). The unique properties of pollen were one of the reasons for using it as a pharmaceutical preparation. It was also important whether pollen was gathered by hand directly from the flower or by bees which is particularly rich in vitamins with a far higher concentration of pantothenic acid. The German Federal Board of Health has officially recognized the use of pollen as medicine; pollen has been helpful when administered in cases of chronic prostatitis (Cai et al., 2014; Wagenlehner et al., 2009). Some chronic diseases such as cancer, coronary, and neurological degeneration have been reported to be a consequence of oxidative damage (Babizhayev, Vishnyakova, & Yegorov, 2014; Jiang, Sun, & Chen, 2016). The therapeutic potential of honey is almost always associated with the antioxidant capacity against reactive oxygen species (Ferreira et al., 2009 Al-Meeri, & Al-Habori, 2002; Tonks et al., 2003;Brudzynski & Kim, 2011; Nasir et al., 2010; Erejuwa, Sulaiman, & Wahab, 2012). Oilseed rape (Brassica napus) is one of the most cultivated crop plants around the world. It is an important source of oil and medicinal components. The cultivation rate of oilseed rape around the globe has been increasing over the last ten years with about 31.5 Mha of lands used to cultivate in 2010 (Gulden, Warwick,& Thomas, 2008; Islam et al., 2013). Rape seed (Brassica napus L.) crops and honey are very good and commonly used bee food. Its pollen contains such amino acids as Threonine, Valine and Methionine and 23% to 24% crude protein, but the exact content is not exactly known. Honey differs in its com...

Background and Objectives Human amniotic fluid-derived mesenchymal stem cells (AF-MSCs) may be a valuable source for cardiovascular tissue engineering and cell therapy. The aim of this study is to verify angiotensin II and transforming growth factor-beta 1 (TGF- β 1) as potential cardiomyogenic differentiation inducers of AF-MSCs. Methods and Results AF-MSCs were obtained from amniocentesis samples from second-trimester pregnant women, isolated and characterized by the expression of cell surface markers (CD44, CD90, CD105 positive; CD34 negative) and pluripotency genes ( OCT4 , SOX2 , NANOG , REX1 ). Cardiomyogenic differentiation was induced using different concentrations of angiotensin II and TGF- β 1. Successful initiation of differentiation was confirmed by alterations in cell morphology, upregulation of cardiac genes-markers NKX2-5 , TBX5 , GATA4 , MYH6 , TNNT2 , DES and main cardiac ion channels genes (sodium, calcium, potassium) as determined by RT-qPCR. Western blot and immunofluorescence analysis revealed the increased expression of Connexin43, the main component of gap junctions, and Nkx2.5, the early cardiac transcription factor. Induced AF-MSCs switched their phenotype towards more energetic and started utilizing oxidative phosphorylation more than glycolysis for energy production as assessed using Agilent Seahorse XF analyzer. The immune analysis of chromatin-modifying enzymes DNMT1, HDAC1/2 and Polycomb repressive complex 1 and 2 (PRC1/2) proteins BMI1, EZH2 and SUZ12 as well as of modified histones H3 and H4 indicated global chromatin remodeling during the induced differentiation. Conclusions Angiotensin II and TGF- β 1 are efficient cardiomyogenic inducers of human AF-MSCs; they initiate alterations at the gene and protein expression, metabolic and epigenetic levels in stem cells leading towards cardiomyocyte-like phenotype formation.

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