Additional file 2: Table S1. of High-throughput immunophenotypic characterization of bone marrow- and cord blood-derived mesenchymal stromal cells reveals common and differentially expressed markers: identification of angiotensin-converting enzyme (CD143) as a marker differentially expressed between adult and perinatal tissue sources

Mean and CV values of log2 FI for each of 246 investigated markers. (DOCX 22 kb

Selected genes for testing DNA microarray outcomes in quantitative real-time PCR.

<p>Selected genes for testing DNA microarray outcomes in quantitative real-time PCR.</p

MSC morphology and flow cytometry analysis after QMR stimulation.

<p>A) The images were obtained after 10 minutes of QMR stimulation at Day 5 (first cycle of treatment) and at B) Day 12 (second cycle of treatment). Scale bar = 100 μm. Total magnification = 100X. One representative experiment was shown. C) Five colour combination of monoclonal antibodies was used to verify MSC identity according to the above listed surface markers of a representative sample. Grey line = unstained control (CTL-). Blue line = sham-exposed control (CTL). Green line = QMR setting 80. Red line = QMR setting 40.</p

Cellular viability and proliferation after QMR treatment.

<p>A) Histograms represent the % of cellular viability after two cycles of QMR treatment at the different settings compared to the sham-exposed controls determined by flow cytometry. Data were shown as mean ± SD of three independent experiments; B) Percentages of cellular proliferation on the controls were obtained by WST-1 assay after 72 hours. Data were represented as mean ± SD of n = 6 independent experiments. No statistical differences were found between conditions.</p

Best enrichment gene lists.

<p>Analysis of functional gene enrichment using ToppFun tool (application of ToppGene Suite) for A) up-regulated and B) down-regulated DEG between 40 QMR setting and control with significant enrichment (dotted line) for FDR B&H q-value <0.01); C) Comparative analysis of up-regulated (green bar) and down-regulated (red bar) functional gene enrichments using IPA software with significant enrichment (dotted line) for -log2 (B-H p-value) >2.</p

<i>In-vitro</i> analysis of Quantum Molecular Resonance effects on human mesenchymal stromal cells

<div><p>Electromagnetic fields play an essential role in cellular functions interfering with cellular pathways and tissue physiology. In this context, Quantum Molecular Resonance (QMR) produces waves with a specific form at high-frequencies (4–64 MHz) and low intensity through electric fields. We evaluated the effects of QMR stimulation on bone marrow derived mesenchymal stromal cells (MSC). MSC were treated with QMR for 10 minutes for 4 consecutive days for 2 weeks at different nominal powers. Cell morphology, phenotype, multilineage differentiation, viability and proliferation were investigated. QMR effects were further investigated by cDNA microarray validated by real-time PCR. After 1 and 2 weeks of QMR treatment morphology, phenotype and multilineage differentiation were maintained and no alteration of cellular viability and proliferation were observed between treated MSC samples and controls. cDNA microarray analysis evidenced more transcriptional changes on cells treated at 40 nominal power than 80 ones. The main enrichment lists belonged to development processes, regulation of phosphorylation, regulation of cellular pathways including metabolism, kinase activity and cellular organization. Real-time PCR confirmed significant increased expression of MMP1, PLAT and ARHGAP22 genes while A2M gene showed decreased expression in treated cells compared to controls. Interestingly, differentially regulated MMP1, PLAT and A2M genes are involved in the extracellular matrix (ECM) remodelling through the fibrinolytic system that is also implicated in embryogenesis, wound healing and angiogenesis. In our model QMR-treated MSC maintained unaltered cell phenotype, viability, proliferation and the ability to differentiate into bone, cartilage and adipose tissue. Microarray analysis may suggest an involvement of QMR treatment in angiogenesis and in tissue regeneration probably through ECM remodelling.</p></div

Relative gene expressions using quantitative real-time PCR.

<p>Expression of 8 genes selected by cDNA microarray was illustrated after n = 6 independent experiments using TBP as representative reference gene; mean ± SD; * p<0.05; ** p<0.01.</p

Adipogenic, osteogenic and chondrogenic differentiation after QMR cycles of stimulation.

<p>Panels display one representative experiment showing the final outcome in MSC multilineage differentiation after 21 days of induction. QMR-treated (at 40 and 80 nominal powers) and untreated samples (CTL+) were induced to differentiation. Osteogenic differentiation after one cycle (A) and two cycles (B) of QMR stimulation was assessed using Alizarin Red. Adipogenic (C, D) and chondrogenic (E, F) differentiation were detected using Oil Red O and Alcian Blue stainings, respectively. Scale bar = 100 μm. Total magnification = 100x.</p

Additional file 5: Table S2. of Generation of mesenchymal stromal cells from cord blood: evaluation of in vitro quality parameters prior to clinical use

List of antibodies used for immunophenotypic analysis. (DOCX 14 kb

Additional file 10: Figure S7. of Generation of mesenchymal stromal cells from cord blood: evaluation of in vitro quality parameters prior to clinical use

SL-CBMSC immunophenotypic analysis. Characterization of SL-MSC (n = 6) by flow cytometry using a panel of 14 cell surface markers. Boxes extend from 25th percentile to the 75th percentile, the middle line represents median value and the whiskers extend from minimum to maximum values. Data are displayed as rMFI on the unstained control. (DOCX 104 kb