RESUMEN
In this study, we investigated whether multipotent (human-bone-marrow-derived mesenchymal stem cells [hBM-MSCs]) and pluripotent stem cells (murine-induced pluripotent stem cells [iPSCs] and murine embryonic stem cells [ESCs]) respond to nanocomposite fibrous mats of poly(L-lactic acid) (PLLA) loaded with 1 or 8 wt % of calcium-deficient nanohydroxyapatite (d-HAp). Remarkably, the dispersion of different amounts of d-HAp to PLLA produced a set of materials (PLLA/d-HAp) with similar architectures and tunable mechanical properties. After 3 weeks of culture in the absence of soluble osteogenic factors, we observed the expression of osteogenic markers, including the deposition of bone matrix proteins, in multi/pluripotent cells only grown on PLLA/d-HAp nanocomposites, whereas the osteogenic differentiation was absent on stem-cell-neat PLLA cultures. Interestingly, this phenomenon was confined only in hBM-MSCs, murine iPSCs, and ESCs grown on direct contact with the PLLA/d-HAp mats. Altogether, these results indicate that the osteogenic differentiation effect of these electrospun PLLA/d-HAp nanocomposites was independent of the stem cell type and highlight the direct interaction of stem cell-polymeric nanocomposite and the mechanical properties acquired by the PLLA/d-HAp nanocomposites as key steps for the differentiation process.
Asunto(s)
Calcio/química , Células Madre Embrionarias/química , Ácido Láctico/química , Células Madre Mesenquimatosas/química , Nanocompuestos/química , Células Madre Pluripotentes/química , Polímeros/química , Animales , Supervivencia Celular , Durapatita/química , Electroquímica , Células Madre Embrionarias/citología , Humanos , Células Madre Mesenquimatosas/citología , Ratones , Tamaño de la Partícula , Células Madre Pluripotentes/citología , PoliésteresRESUMEN
OBJECTIVE: Nitric oxide (NO) production and Ca(2+) homeostasis are key determinants for the control of many cell functions. NO is known to be a mediator of Ca(2+) homeostasis in a highly complex and cell-specific manner and although Ca(2+) homeostasis has been explored in human oral cancer cells, the exact mechanisms are not completely understood. In this study we investigated the impact of exogenous NO on [Ca(2+)]c homeostasis in PE/CA-PJ15 cells. DESIGN: Cells were treated with S-nitrosocysteine as NO-donor and the determinations of cytosolic Ca(2+) concentrations were performed using FURA-2 AM. Carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP) and oligomycin were used to challenge mitochondrial functionality, whereas thapsigargin (TG) and La(3+) were employed to perturb intracellular calcium levels. RESULTS: NO derived from S-nitrosocysteine (CySNO) induced a dose-dependent reduction of cytosolic calcium [Ca(2+)]c whereas oxy-haemoglobin (oxyHb) completely counteracted this effect. Subsequently, we assessed possible relationships between NO and cellular structures responsible for Ca(2+) homeostasis. We found that uncoupling of mitochondrial respiration with carbonyl-cyanide-4-(trifluoromethoxy)-phenylhydrazone (FCCP) and oligomycin strongly reduced the effect of NO on [Ca(2+)]c. Moreover, we found that during this mitochondrial energetic deficit, the effect of NO on [Ca(2+)]c was also reduced in the presence of La(3+) or thapsigargin. CONCLUSIONS: NO induces a concentration-dependent [Ca(2+)]c reduction in PE/CA-PJ15 human oral cancer cells and potentiates mitochondrial Ca(2+) buffering in the presence of TG or La(3+). Further, we show that exogenous NO deregulates Ca(2+) homeostasis in PE/CA-PJ15 cells with fully energized mitochondria.
Asunto(s)
Señalización del Calcio/efectos de los fármacos , Óxido Nítrico/farmacología , Carbonil Cianuro p-Trifluorometoxifenil Hidrazona/farmacología , Supervivencia Celular/efectos de los fármacos , Cisteína/análogos & derivados , Cisteína/farmacología , Citosol/química , Relación Dosis-Respuesta a Droga , Fura-2/farmacología , Homeostasis/efectos de los fármacos , Humanos , Mitocondrias , Oligomicinas/farmacología , Oxihemoglobinas/farmacología , S-Nitrosotioles/farmacología , Tapsigargina/farmacología , Células Tumorales CultivadasRESUMEN
The interaction between stem cells and biomaterials with nanoscale topography represents a main route in the roadmap for tissue engineering-based strategies. In this study, we explored the interface between human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and hydrogenated amorphous carbon (a-C:H) film designed with uniform, groove, or grid nanopatterns. In either case, hBM-MSCs preserved growth rate and multi-differentiation properties, suggesting that the films were biocompatible and suitable for stem cell culture. hBM-MSCs responded to different nanopattern designs with specific changes of microtubule organization. In particular, the grid pattern induced a square-localized distribution of alpha-tubulin/actin fibers, whereas the groove pattern exerted a more dynamic effect, associated with microtubule alignment and elongation.