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ABSTRACT: Previous studies have demonstrated that mesenchymal stromal cells (MSCs) enhance cell survival through upregulation and secretion of stanniocalcin-1 (STC1). This study shows that MSC-derived STC1 promotes survival of lung cancer cells by uncoupling oxidative phosphorylation, reducing intracellular reactive oxygen species (ROS), and shifting metabolism towards a more glycolytic metabolic profile. MSC-derived STC1 upregulated uncoupling protein 2 (UCP2) in injured A549 cells in an STC1-dependent manner. Knockdown of UCP2 reduced the ability of MSCs and recombinant STC1 (rSTC1) to reduce cell death in the A549 population. rSTC1-treated A549 cells displayed decreased levels of ROS, mitochondrial membrane potential (MMP), and increased lactate production, all of which were dependent on the upregulation of UCP2. Our data suggest that MSCs can promote cell survival by regulating mitochondrial respiration via STC1.
Molecular Therapy 12/2011; 20(2):417-23. · 6.87 Impact Factor
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ABSTRACT: Promyelocytic leukemia protein nuclear bodies (PML NBs) have been implicated in host immune response to viral infection. PML NBs are targeted for degradation during reactivation of herpes viruses, suggesting that disruption of PML NB function supports this aspect of the viral life cycle. The Epstein-Barr virus (EBV) Latent Membrane Protein 1 (LMP1) has been shown to suppress EBV reactivation. Our finding that LMP1 induces PML NB immunofluorescence intensity led to the hypothesis that LMP1 may modulate PML NBs as a means of maintaining EBV latency. Increased PML protein and morphometric changes in PML NBs were observed in EBV infected alveolar epithelial cells and nasopharyngeal carcinoma cells. Treatment with low dose arsenic trioxide disrupted PML NBs, induced expression of EBV lytic proteins, and conferred ganciclovir susceptibility. This study introduces an effective modality to induce susceptibility to ganciclovir in epithelial cells with implications for the treatment of EBV associated pathologies.
Virology 07/2011; 416(1-2):86-97. · 3.35 Impact Factor
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ABSTRACT: The Epstein-Barr virus (EBV) encoded Latent Membrane Protein 1 (LMP1) has been shown to increase the expression of promyelocytic leukemia protein (PML) and the immunofluorescent intensity of promyelocytic leukemia nuclear bodies (PML NBs). PML NBs have been implicated in the modulation of transcription and the association of reporter plasmids with PML NBs has been implicated in repression of reporter activity. Additionally, repression of various reporters in the presence of LMP1 has been noted. This study demonstrates that LMP1 suppresses expression of reporter activity in a dose responsive manner and corresponds with the LMP1 induced increase in PML NB intensity. Disruption of PML NBs with arsenic trioxide or a PML siRNA restores reporter activity. These data offer an explanation for previously conflicting data on LMP1 signaling and calls attention to the possibility of false-positives and false-negatives when using reporter assays as a research tool in cells expressing LMP1.
Virology Journal 01/2011; 8:461. · 2.34 Impact Factor
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ABSTRACT: The epithelial cell response to stress involves the transmission of signals between contiguous cells that can be visualized as a calcium wave. In some cell types, this wave is dependent on the release of extracellular trinucleotides from injured cells. In particular, extracellular ATP has been reported to be critical for the epithelial cell response to stress and has recently been shown to be upregulated in tumors in vivo.
Here, we identify stanniocalcin-1 (STC1), a secreted pleiotrophic protein, as a critical mediator of calcium wave propagation in monolayers of pulmonary (A549) and prostate (PC3) epithelial cells. Addition of STC1 enhanced and blocking STC1 decreased the distance traveled by an extracellular ATP-dependent calcium wave. The same effects were observed when calcium was stimulated by the addition of exogenous ATP. We uncover a positive feedback loop in which STC1 promotes the release of ATP from cells in vitro and in vivo.
The results indicated that STC1 plays an important role in the early response to mechanical injury by epithelial cells by modulating signaling of extracellular ATP. This is the first report to describe STC1 as a modulator or purinergic receptor signaling.
PLoS ONE 01/2010; 5(4):e10237. · 4.09 Impact Factor
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ABSTRACT: Multipotent stromal cells (MSCs) have been shown to reduce apoptosis in injured cells by secretion of paracrine factors, but these factors were not fully defined. We observed that coculture of MSCs with previously UV-irradiated fibroblasts reduced apoptosis of the irradiated cells, but fresh MSC conditioned medium was unable reproduce the effect. Comparative microarray analysis of MSCs grown in the presence or absence of UV-irradiated fibroblasts demonstrated that the MSCs were activated by the apoptotic cells to increase synthesis and secretion of stanniocalcin-1 (STC-1), a peptide hormone that modulates mineral metabolism and has pleiotrophic effects that have not been fully characterized. We showed that STC-1 was required but not sufficient for reduction of apoptosis of UV-irradiated fibroblasts. In contrast, we demonstrated that MSC-derived STC-1 was both required and sufficient for reduction of apoptosis of lung cancer epithelial cells made apoptotic by incubation at low pH in hypoxia. Our data demonstrate that STC-1 mediates the antiapoptotic effects of MSCs in two distinct models of apoptosis in vitro.
Stem Cells 04/2009; 27(3):670-81. · 7.78 Impact Factor
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ABSTRACT: Multipotent stromal cells (MSCs) have been shown to reduce apoptosis in injured cells by secretion of paracrine factors, but these factors were not fully defined. We observed that coculture of MSCs with previously UV-irradiated fibroblasts reduced apoptosis of the irradiated cells, but fresh MSC conditioned medium was unable reproduce the effect. Comparative microarray analysis of MSCs grown in the presence or absence of UV-irradiated fibroblasts demonstrated that the MSCs were activated by the apoptotic cells to increase synthesis and secretion of stanniocalcin-1 (STC-1), a peptide hormone that modulates mineral metabolism and has pleiotrophic effects that have not been fully characterized. We showed that STC-1 was required but not sufficient for reduction of apoptosis of UV-irradiated fibroblasts. In contrast, we demonstrated that MSC-derived STC-1 was both required and sufficient for reduction of apoptosis of lung cancer epithelial cells made apoptotic by incubation at low pH in hypoxia. Our data demonstrate that STC-1 mediates the antiapoptotic effects of MSCs in two distinct models of apoptosis in vitro. STEM CELLS2009;27:670–681
Stem Cells 03/2009; 27(3):670 - 681. · 7.78 Impact Factor
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ABSTRACT: Multipotent stromal cells (MSCs) have been shown to reduce apoptosis in injured cells by secretion of paracrine factors, but these factors were not fully defined. We observed that co-culture of MSCs with previously UV irradiated fibroblasts reduced apoptosis of the irradiated cells, but fresh MSC conditioned media was unable reproduce the effect. Comparative Microarray analysis of MSCs grown in the presence or absence of UV irradiated fibroblasts demonstrated that the MSCs were activated by the apoptotic cells to increase synthesis and secretion of stanniocalcin-1 (STC-1), a peptide hormone that modulates mineral metabolism and has pleiotrophic effects that have not been fully characterized. We showed that STC-1 was required but not sufficient for reduction of apoptosis of UV-irradiated fibroblasts. In contrast, we demonstrated that MSC-derived STC-1 was both required and sufficient for reduction of apoptosis of lung cancer epithelial cells made apoptotic by incubation at low pH in hypoxia. Our data demonstrate that STC-1 mediates the anti-apoptotic effects of MSCs in two distinct models of apoptosis in vitro. ______________________________________________________________________________ Author contributions: G.B.: Conception and design, collection and/or assembly of data, analysis and interpretation, manuscript writing and revisions; S.O.: Conception and design, collection and/or assembly of data, analysis and interpretation, manuscript writing; F.F.: Collection of data; J.F.: Collection of data; C.G.: Conception and design; R.P.: Conception and design; G.D.: Conception and design, provision of study material or patients; D.E.S.: Conception and design, provision of study material or patients; D.P.: Conception and design, analysis and interpretation, manuscript writing, Final approval of manuscript. Gregory Block and Shinya Ohkouchi contributed equally to this work.
Stem Cells 01/2009; · 7.78 Impact Factor
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ABSTRACT: Whereas the PML protein has been reported to have both transcriptional coactivator and corepressor potential, the contribution of the PML nuclear body (PML NB) itself to transcriptional regulation is not well understood. Here we demonstrate that plasmid DNA artificially tethered to PML or the PML NB-targeting domain of Sp100 is preferentially localized to PML NBs. Using the tethering technique, we targeted a simian virus 40 promoter-driven luciferase reporter plasmid to PML NBs, resulting in the repression of the transgene transcriptional activity. Conversely, the tethering of a cytomegalovirus promoter-containing reporter plasmid resulted in activation. Targeting a minimal eukaryotic promoter did not affect its activity. The expression of targeted promoters could be modulated by altering the cellular concentration of PML NB components, including Sp100 and isoforms of the PML protein. Finally, we demonstrate that ICP0, the promiscuous herpes simplex virus transactivator, increases the level of transcriptional activation of plasmid DNA tethered to the PML NB. We conclude that when PML NB components are artificially tethered to reporter plasmids, the PML NB contributes to the regulation of the tethered DNA in a promoter-dependent manner. Our findings demonstrate that transient transcription assays are sensitive to the subnuclear localization of the transgene plasmid.
Molecular and Cellular Biology 01/2007; 26(23):8814-25. · 5.53 Impact Factor
