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Human Amniotic Fluid Mesenchymal Stem Cells from Second- and Third-Trimester Amniocentesis: Differentiation Potential, Molecular Signature, and Proteome Analysis.

Savickiene J, Treigyte G, Baronaite S, Valiuliene G, Kaupinis A, Valius M, Arlauskiene A, Navakauskiene R - Stem Cells Int (2015)

Bottom Line: Using two-stage protocol, MSCs were successfully cultured and exhibited typical stem cell morphological, specific cell surface, and pluripotency markers characteristics.The analysis of the functional and expression patterns of 250 high abundance proteins selected from more than 1400 demonstrated the similar proteome of cultured and differentiated AF-MSCs but the unique changes in their expression profile during cell differentiation that may help the identification of key markers in differentiated cells.Our results provide evidence that human amniotic fluid of second- and third-trimester contains stem cells with multilineage potential and may be attractive source for clinical applications.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology, Vilnius University, LT-08662 Vilnius, Lithuania.

ABSTRACT
Human amniotic fluid stem cells have become an attractive stem cell source for potential applications in regenerative medicine and tissue engineering. The aim of this study was to characterize amniotic fluid-derived mesenchymal stem cells (AF-MSCs) from second- and third-trimester of gestation. Using two-stage protocol, MSCs were successfully cultured and exhibited typical stem cell morphological, specific cell surface, and pluripotency markers characteristics. AF-MSCs differentiated into adipocytes, osteocytes, chondrocytes, myocytes, and neuronal cells, as determined by morphological changes, cell staining, and RT-qPCR showing the tissue-specific gene presence for differentiated cell lineages. Using SYNAPT G2 High Definition Mass Spectrometry technique approach, we performed for the first time the comparative proteomic analysis between undifferentiated AF-MSCs from late trimester of gestation and differentiated into myogenic, adipogenic, osteogenic, and neurogenic lineages. The analysis of the functional and expression patterns of 250 high abundance proteins selected from more than 1400 demonstrated the similar proteome of cultured and differentiated AF-MSCs but the unique changes in their expression profile during cell differentiation that may help the identification of key markers in differentiated cells. Our results provide evidence that human amniotic fluid of second- and third-trimester contains stem cells with multilineage potential and may be attractive source for clinical applications.

No MeSH data available.


Highly up- and downregulated proteins in second-trimester AF-MSCs undergoing (a) myogenic, (b) adipogenic, (c) osteogenic, and (d) neurogenic differentiation. Each bar corresponds to the expression fold differences higher than six or lower than eight in each differentiating population as compared with undifferentiated AF-MSCs.
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fig5: Highly up- and downregulated proteins in second-trimester AF-MSCs undergoing (a) myogenic, (b) adipogenic, (c) osteogenic, and (d) neurogenic differentiation. Each bar corresponds to the expression fold differences higher than six or lower than eight in each differentiating population as compared with undifferentiated AF-MSCs.

Mentions: For a comparison of the proteome profile, we presented a sample of normal cultured AF-MSCs from second-trimester (16 weeks, passage 5), cultured under appropriate conditions to induce myogenic, adipogenic, osteogenic, and neurogenic differentiation. The SYNAPT G2 High Definition Mass Spectrometry based analysis demonstrated 1423 proteins expressed in AF-MSCs. The relative protein expression ratios were calculated for each differentiation, and proteins with the expression ratio above 1.5 compared with undifferentiated control were selected and presented in Supplementary Table 1 (see Supplementary Table 1 in Supplementary Material available online at http://dx.doi.org/10.1155/2015/319238). The abbreviated and full names of the proteins, the accession numbers, the theoretical molecular weight and pI values, Max score and the number of reported peptides, and the percentage coverage of the identified proteins are listed in the table. Figure 4(a) represents the functional classification of 250 selected proteins from AF-MSCs differentiated toward myogenic, adipogenic, osteogenic, and neurogenic lineages with the expression ratio above 1.5 versus control. The groups of selected proteins were related to cell growth and differentiation (12%), regulation (13%), cell signalling/communication (9%), and transcription/translation (8%). Other groups represent metabolic (22%), transport (18%), structural (6%), and immune response (5%) proteins, including 7% of unknown proteins. Among 250 of high abundance proteins selected (Figure 4(b)), 91, 89, 96, and 87 were upregulated and 89, 105, 106, and 81 downregulated in AF-MSCs undergoing myogenic, adipogenic, osteogenic, and neurogenic differentiation, respectively. A number of proteins were absent in undifferentiated AF-MSCs, including latexin (LXN, involved in negative regulation of peptidase activity), growth differentiation factor 6 (GDF6, involved in multicellular organism development), Ig mu heavy chain disease protein (MUCB, an extracellular vesicular exosome), alpha crystallin B chain (CRYAB, involved in metabolic processes), glycogen synthase kinase 3 beta (GSK3β, involved in glycogen metabolism and signalling), and ATP dependent RNA helicase DDX 19A (DD19A, involved in many biological, metabolic, and transport processes) (Supplementary Table 1). Next we identified significant changes in the expression levels of up- and downregulated proteins in differentiated cells compared with undifferentiated control (Figure 5).


Human Amniotic Fluid Mesenchymal Stem Cells from Second- and Third-Trimester Amniocentesis: Differentiation Potential, Molecular Signature, and Proteome Analysis.

Savickiene J, Treigyte G, Baronaite S, Valiuliene G, Kaupinis A, Valius M, Arlauskiene A, Navakauskiene R - Stem Cells Int (2015)

Highly up- and downregulated proteins in second-trimester AF-MSCs undergoing (a) myogenic, (b) adipogenic, (c) osteogenic, and (d) neurogenic differentiation. Each bar corresponds to the expression fold differences higher than six or lower than eight in each differentiating population as compared with undifferentiated AF-MSCs.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4553339&req=5

fig5: Highly up- and downregulated proteins in second-trimester AF-MSCs undergoing (a) myogenic, (b) adipogenic, (c) osteogenic, and (d) neurogenic differentiation. Each bar corresponds to the expression fold differences higher than six or lower than eight in each differentiating population as compared with undifferentiated AF-MSCs.
Mentions: For a comparison of the proteome profile, we presented a sample of normal cultured AF-MSCs from second-trimester (16 weeks, passage 5), cultured under appropriate conditions to induce myogenic, adipogenic, osteogenic, and neurogenic differentiation. The SYNAPT G2 High Definition Mass Spectrometry based analysis demonstrated 1423 proteins expressed in AF-MSCs. The relative protein expression ratios were calculated for each differentiation, and proteins with the expression ratio above 1.5 compared with undifferentiated control were selected and presented in Supplementary Table 1 (see Supplementary Table 1 in Supplementary Material available online at http://dx.doi.org/10.1155/2015/319238). The abbreviated and full names of the proteins, the accession numbers, the theoretical molecular weight and pI values, Max score and the number of reported peptides, and the percentage coverage of the identified proteins are listed in the table. Figure 4(a) represents the functional classification of 250 selected proteins from AF-MSCs differentiated toward myogenic, adipogenic, osteogenic, and neurogenic lineages with the expression ratio above 1.5 versus control. The groups of selected proteins were related to cell growth and differentiation (12%), regulation (13%), cell signalling/communication (9%), and transcription/translation (8%). Other groups represent metabolic (22%), transport (18%), structural (6%), and immune response (5%) proteins, including 7% of unknown proteins. Among 250 of high abundance proteins selected (Figure 4(b)), 91, 89, 96, and 87 were upregulated and 89, 105, 106, and 81 downregulated in AF-MSCs undergoing myogenic, adipogenic, osteogenic, and neurogenic differentiation, respectively. A number of proteins were absent in undifferentiated AF-MSCs, including latexin (LXN, involved in negative regulation of peptidase activity), growth differentiation factor 6 (GDF6, involved in multicellular organism development), Ig mu heavy chain disease protein (MUCB, an extracellular vesicular exosome), alpha crystallin B chain (CRYAB, involved in metabolic processes), glycogen synthase kinase 3 beta (GSK3β, involved in glycogen metabolism and signalling), and ATP dependent RNA helicase DDX 19A (DD19A, involved in many biological, metabolic, and transport processes) (Supplementary Table 1). Next we identified significant changes in the expression levels of up- and downregulated proteins in differentiated cells compared with undifferentiated control (Figure 5).

Bottom Line: Using two-stage protocol, MSCs were successfully cultured and exhibited typical stem cell morphological, specific cell surface, and pluripotency markers characteristics.The analysis of the functional and expression patterns of 250 high abundance proteins selected from more than 1400 demonstrated the similar proteome of cultured and differentiated AF-MSCs but the unique changes in their expression profile during cell differentiation that may help the identification of key markers in differentiated cells.Our results provide evidence that human amniotic fluid of second- and third-trimester contains stem cells with multilineage potential and may be attractive source for clinical applications.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology, Vilnius University, LT-08662 Vilnius, Lithuania.

ABSTRACT
Human amniotic fluid stem cells have become an attractive stem cell source for potential applications in regenerative medicine and tissue engineering. The aim of this study was to characterize amniotic fluid-derived mesenchymal stem cells (AF-MSCs) from second- and third-trimester of gestation. Using two-stage protocol, MSCs were successfully cultured and exhibited typical stem cell morphological, specific cell surface, and pluripotency markers characteristics. AF-MSCs differentiated into adipocytes, osteocytes, chondrocytes, myocytes, and neuronal cells, as determined by morphological changes, cell staining, and RT-qPCR showing the tissue-specific gene presence for differentiated cell lineages. Using SYNAPT G2 High Definition Mass Spectrometry technique approach, we performed for the first time the comparative proteomic analysis between undifferentiated AF-MSCs from late trimester of gestation and differentiated into myogenic, adipogenic, osteogenic, and neurogenic lineages. The analysis of the functional and expression patterns of 250 high abundance proteins selected from more than 1400 demonstrated the similar proteome of cultured and differentiated AF-MSCs but the unique changes in their expression profile during cell differentiation that may help the identification of key markers in differentiated cells. Our results provide evidence that human amniotic fluid of second- and third-trimester contains stem cells with multilineage potential and may be attractive source for clinical applications.

No MeSH data available.