Superparamagnetic Iron Oxide Particles (VSOPs) Show Genotoxic Effects but No Functional Impact on Human Adipose Tissue-Derived Stromal Cells (ASCs).

Adipose tissue-derived stromal cells (ASCs) represent a capable source for cell-based therapeutic approaches. For monitoring a cell-based application in vivo, magnetic resonance imaging (MRI) of cells labeled with iron oxide particles is a common method. It is the aim of the present study to analyze potential DNA damage, cytotoxicity and impairment of functional properties of human (h)ASCs after labeling with citrate-coated very small superparamagnetic iron oxide particles (VSOPs). Cytotoxic as well as genotoxic effects of the labeling procedure were measured in labeled and unlabeled hASCs using the MTT assay, comet assay and chromosomal aberration test. Trilineage differentiation was performed to evaluate an impairment of the differentiation potential due to the particles. Proliferation as well as migration capability were analyzed after the labeling procedure. Furthermore, the labeling of the hASCs was confirmed by Prussian blue staining, transmission electron microscopy (TEM) and high-resolution MRI. Below the concentration of 0.6 mM, which was used for the procedure, no evidence of genotoxic effects was found. At 0.6 mM, 1 mM as well as 1.5 mM, an increase in the number of chromosomal aberrations was determined. Cytotoxic effects were not observed at any concentration. Proliferation, migration capability and differentiation potential were also not affected by the procedure. Labeling with VSOPs is a useful labeling method for hASCs that does not affect their proliferation, migration and differentiation potential. Despite the absence of cytotoxicity, however, indications of genotoxic effects have been demonstrated.

Toxicity and Functional Impairment in Human Adipose Tissue-Derived Stromal Cells (hASCs) Following Long-Term Exposure to Very Small Iron Oxide Particles (VSOPs).

Magnetic nanoparticles (NPs), such as very small iron oxide NPs (VSOPs) can be used for targeted drug delivery, cancer treatment or tissue engineering. Another important field of application is the labelling of mesenchymal stem cells to allow in vivo tracking and visualization of transplanted cells using magnetic resonance imaging (MRI). For these NPs, however, various toxic effects, as well as functional impairment of the exposed cells, are described. The present study evaluates the influence of VSOPs on the multilineage differentiation ability and cytokine secretion of human adipose tissue derived stromal cells (hASCs) after long-term exposure. Human ASCs were labelled with VSOPs, and the efficacy of the labelling was documented over 4 weeks in vitro cultivation of the labelled cells. Unlabelled hASCs served as negative controls. Four weeks after labelling, adipogenic and osteogenic differentiation was histologically evaluated and quantified by polymerase chain reaction (PCR). Changes in gene expression of IL-6, IL-8, VEGF and caspase 3 were determined over 4 weeks. Four weeks after the labelling procedure, labelled and unlabelled hASCs did not differ in the gene expression of IL-6, IL-8, VEGF and caspase 3. Furthermore, the labelling procedure had no influence on the multidifferentiation ability of hASC. The percentage of labelled cells decreased during in vitro expansion over 4 weeks. Labelling with VSOPs and long-term intracellular disposition probably have no influence on the physiological functions of hASCs. This could be important for the future in vivo use of iron oxide NPs.

Isolation of a novel compound (MIMO2) from the methanolic extract of Moringa oleifera leaves: protective effects against vanadium-induced cytotoxity.

Moringa oleifera is reported to be a miracle plant, with positive effects on practically every system in the animal body. The methanolic extract of Moringa oleifera leaves was fractionated using liquid-liquid fractionation, column chromatography and preparative high-performance liquid chromatography (HPLC). Bioassay guided fractionation using Ferric Reducing Antioxidant Power (FRAP) was used to determine the fraction with the highest antioxidative power. Chemical structure was elucidated with nuclear magnetic resonance (NMR) spectroscopy. FRAP showed that the pure compound, butyl p-hydroxyphenyl-acetate (MIMO2) exhibited an antioxidant activity higher than TEMPOL (positive control). Vanadium is a metal, which as a salt has been shown to be a neurotoxicant; and was therefore used to assess the efficacy of MIMO2 in this experiment. HT22 (immortalized mouse hippocampal) cells were used for cell culture. The Comet assay showed a statistically significant reduction (p < .05) in DNA damage when 0.25 and 0.5 µM MIMO2 as well as 0.1 and 0.2 mg of the methanolic extract of Moringa oleifera leaves (MO) were used in combination with 200 µM vanadium (sodium metavanadate). Analogously, a reduced formation of superoxide was observed using dihydroethidium (2,7-Diamino-10-ethyl-9-phenyl-9,10-dihydrophenanthridine-DHE) stain after 0.5 µM MIMO2 and 0.063 mg MO were used in combination with vanadium 100 µM. MIMO2 and MO gave a statistically significant (p < .05) protective effect against vanadium toxicity on neuronal cells. Further assays may need to be performed to assess the extent of protection that MIMO2 may offer, and also to better understand its mechanisms of action.

Chromosomal aberrations and deoxyribonucleic acid single-strand breaks in adipose-derived stem cells during long-term expansion in vitro.

BACKGROUND AIMS: Adipose-derived stem cells (ASCs) are a promising mesenchymal cell source for tissue engineering approaches. To obtain an adequate cell amount, in vitro expansion of the cells may be required in some cases. To monitor potential contraindications for therapeutic applications in humans, DNA strand breaks and chromosomal aberrations in ASCs during in vitro expansion were examined. METHODS: After isolation of ASC from human lipoaspirates of seven patients, in vitro expansion over 10 passages was performed. Cells from passages 1, 2, 3, 5 and 10 were used for the alkaline single-cell microgel electrophoresis (comet) assay to detect DNA single-strand breaks and alkali labile as well as incomplete excision repair sites. Chromosomal changes were examined by means of the chromosomal aberration test. RESULTS: During in vitro expansion, ASC showed no DNA single-strand breaks in the comet assay. With the chromosomal aberration test, however, a significant increase in chromosomal aberrations were detected. CONCLUSIONS: The study showed that although no DNA fragmentation could be determined, the safety of ASC cannot be ensured with respect to chromosome stability during in vitro expansion. Thus, reliable analyses for detecting ASC populations, which accumulate chromosomal aberrations or even undergo malignant transformation during extensive in vitro expansion, must be implemented as part of the safety evaluation of these cells for stem cell-based therapy.