Journal: Experimental cell research

Article Title: Tumor stroma interaction is mediated by monocarboxylate metabolism

doi: 10.1016/j.yexcr.2017.01.013

Figure Lengend Snippet: Glycolytic flux and lactate metabolism. (A) The extracellular acidification rate (ECAR; mean±SD) of MDA-MB-231 cells and CAFs, measured by Seahorse analyzer, reveals that MDA-MB-231 cells are more glycolytic than CAFs. The extracellular acidification rate ECAR measures proton excretion (representing cellular glycolysis) over time in units mpH/min where 1 mpH = 4.3 pmole excreted H+. (B) Extracellular glucose consumption and lactate production of MDA-MB-231 cells and stromal cells confirm the higher glycolytic activity of MDA-MB-231 cells compared to CAFs observed by Seahorse Analyzer analysis. Data are displayed as mean±SD (n = 3). (C) The glucose uptake is significantly higher in MDA-MB-231 cells (MDAs) than in MSCs or CAFs, in good agreement with the higher aerobic glycolysis observed in the cancer cells. Data are displayed as mean±SD (n = 4). (D) CAFs take up and metabolize lactate as well as secrete lactate oxidation metabolites, as shown by 13C MR spectroscopy on cell extracts and CCM. Signal assignments are: α-KG – α-ketoglutarate, Glu – glutamate; Ala – alanine; Lac – lactate; Pyr – pyruvate; α-Glc & β-Glc – α-glucose and β-glucose; “-C” followed by number – position of 13C labeling as a result of metabolic conversion of exogenous 10 mM 3-13C-L-lactate. ** p<0.005, *** p<0.0005 by two-tailed, unpaired, unequal variance Student’s T-test; Abbreviations: MDAs – MDA-MB-231 cells; MSCs: human mesenchymal stem cells; CAFs: cancer-associated fibroblasts; CCM – CAF-conditioned medium;

Article Snippet: Bone marrow-derived mesenchymal stem cells (MSCs) were purchased from Lonza Walkersville, Inc. (Walkersville, MD; Lonza Group Ltd.).

Techniques: Activity Assay, Spectroscopy, Labeling, Two Tailed Test

Journal: Disease Models & Mechanisms

Article Title: Intramyocardial angiogenetic stem cells and epicardial erythropoietin save the acute ischemic heart

doi: 10.1242/dmm.033282

Figure Lengend Snippet: EPO induced synergistic angiogenesis with rat cardiac CD45 − CD44 + DDR2 + MSCs and facilitated cardiomyogenic differentiation. (A,B) Representative immunostaining images (A) from co-cultures of Qtracker ® -labeled (red) cardiac CD45 − CD44 + DDR2 + MSCs and cardiomyocytes (CM). MSCs (arrows) show partially positive expression for cardiomyogenic transcription factors GATA4 (yellow) and Nkx2.5 (green). Quantified GATA4 and Nkx2.5 signal intensities (B) illustrated promotion of cardiomyogenic differentiation in co-cultured MSCs compared with mono-cultured MSCs and strongest Nkx2.5 signal expression in co-cultured MSCs by continuous EPO stimulation. The average signal intensities from MSCs in mono-culture were arbitrarily given a value of 1 (2°). * P <0.05 versus cardiac CD45 − CD44 + DDR2 + MSC mono-culture. # P <0.05. Scale bars: 10 µm. Blue, DAPI in nuclei. (C-E) Representative phase-contrast microscopy images (C) from HUVECs in untreated mono-culture (C1, HUVEC control), in co-culture with rat cardiac CD45 − CD44 + DDR2 + MSCs (C2, HUVECs+MSCs), in MSC-conditioned medium (C3, Cond. DMEM-10), in EPO-supplemented medium (C4, DMEM-10+EPO) and in EPO-supplemented MSC-conditioned medium (C5, Cond. DMEM-10+EPO) with clear tubuli and network formations. Quantified HUVEC branching (D) and junctional network formation (E) illustrated synergistic angiogenetic potential of EPO and paracrine factors from cardiac CD45 − CD44 + DDR2 + MSCs. * P <0.05. Scale bars: 100 µm. Mean±s.e.m.

Article Snippet: Relative fluorescence signal intensities of intranuclear Nkx2.5 and GATA4 in Qtracker ® -labeled cardiac MSCs were quantified under standardized confocal laser scanning microscope settings (ELYRA PS.1 LSM 780, Carl Zeiss) and application of ZEN 2011 software (blue edition, Carl Zeiss).

Techniques: Immunostaining, Labeling, Expressing, Cell Culture, Microscopy, Co-Culture Assay