BackgroundHuman dental mesenchymal stem cells (MSCs) are considered as highly accessible and attractive MSCs for use in regenerative medicine, yet some of their features are not as well characterized as other MSCs. Hypoxia-preconditioning and hypoxia-inducible factor 1 (HIF-1) alpha overexpression significantly improves MSC therapeutics, but the mechanisms involved are not fully understood. In the present study, we characterize immunomodulatory properties of dental MSCs and determine changes in their ability to modulate adaptive and innate immune populations after HIF-1 alpha overexpression.MethodsHuman dental MSCs were stably transduced with green fluorescent protein (GFP-MSCs) or GFP-HIF-1 alpha lentivirus vectors (HIF-MSCs). A hypoxic-like metabolic profile was confirmed by mitochondrial and glycolysis stress test. Capacity of HIF-MSCs to modulate T-cell activation, dendritic cell differentiation, monocyte migration, and polarizations towards macrophages and natural killer (NK) cell lytic activity was assessed by a number of functional assays in co-cultures. The expression of relevant factors were determined by polymerase chain reaction (PCR) analysis and enzyme-linked immunosorbent assay (ELISA).ResultsWhile HIF-1 alpha overexpression did not modify the inhibition of T-cell activation by MSCs, HIF-MSCs impaired dendritic cell differentiation more efficiently. In addition, HIF-MSCs showed a tendency to induce higher attraction of monocytes, which differentiate into suppressor macrophages, and exhibited enhanced resistance to NK cell-mediated lysis, which supports the improved therapeutic capacity of HIF-MSCs. HIF-MSCs also displayed a pro-angiogenic profile characterized by increased expression of CXCL12/SDF1 and CCL5/RANTES and complete loss of CXCL10/IP10 transcription.ConclusionsImmunomodulation and expression of trophic factors by dental MSCs make them perfect candidates for cell therapy. Overexpression of HIF-1 alpha enhances these features and increases their resistance to allogenic NK cell lysis and, hence, their potential in vivo lifespan. Our results further support the use of HIF-1 alpha-expressing dental MSCs for cell therapy in tissue injury and immune disorders.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-017-0659-2) contains supplementary material, which is available to authorized users.
Human induced pluripotent stem cell-derived cardiomyocytes hold great promise for regenerative medicine and in vitro screening. Despite displaying key cardiomyocyte phenotypic characteristics, they more closely resemble foetal/neonatal cardiomyocytes and further characterisation is necessary. Combining the use of tandem mass tags to label cell lysates, followed by multiplexing, we have determined the effects of short term (30 day) in vitro culture on human induced pluripotent stem cell derived cardiomyocyte protein expression. We found that human induced pluripotent stem cell derived cardiomyocytes exhibit temporal changes in global protein expression; alterations in protein expression were pronounced during the first 2 weeks following thaw and dominated by reductions in proteins associated with protein synthesis and ubiquitination. Between 2 and 4 weeks proceeding thaw alterations in protein expression were dominated by metabolic pathways, indicating a potential temporal metabolic shift from glycolysis towards oxidative phosphorylation. Timedependent changes in proteins associated with cardiomyocyte contraction, excitationcontraction coupling and metabolism were detected. While some were associated with expected functional outcomes in terms of morphology or electrophysiology, others such as metabolism did not produce the anticipated maturation of human induced pluripotent stem cell derived cardiomyocytes. In several cases, a predicted outcome was not clear because of the concerted changes in both stimulatory and inhibitory pathways. Nevertheless, clear development of human induced pluripotent stem cell derived cardiomyocytes over this time period was evident.
The spontaneously hypertensive rat (SHR) is a well-characterised model for studies of hypertension and atrial arrhythmias but little is known about the electrophysiological properties of the left ventricle (LV) and their relation with ventricular arrhythmias in this model. To investigate the mechanisms behind electrophysiological abnormalities in the LV and their links to the morphological substrate we used myocardial slices, a multicellular preparation which allows the investigation of functional and structural properties in the same tissue location. Vibratome-cut myocardial slices (300mm thick) were prepared from 20 month-old SHR and aged-matched control LVs, tangentially to the epicardial surface. Slices were point-stimulated and analysed using a multi-electrode array system; conduction velocity (CV) and field potential duration (FPD), an index of action potential duration, were measured. Longitudinal CV was lower in the SHR compared with controls (SHR: 2954 cm/s, n=12 slices; C: 4556 cm/s, n=22 slices; p<0.05), but transverse CV was unchanged. FPD was not statistically different between the two groups and showed a similar restitution curve. A high number of fractionated field potentials were observed at similar levels in both groups (SHR: 107520 ms, n=6 slices; C: 7452 ms, n=6 slices; p>0.05). In support of the lower CV, Western blotting analysis revealed a reduction in the gap junctional protein connexin43 expression in the SHR (p<0.01). A high amount of interstitial fibrosis, quantified by Sirius red/fast green collagen quantitative staining kit was found in both groups at similar levels (P>0.05) and could be ascribed to aging as this was not detected in slices from 2 month old rats (p <0.05). These results suggest that down regulation of connexin 43 and not myocardial fibrosis is associated with electrophysiological LV abnormalities in a rat model of chronic hypertension.
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