Supplemental Information. Materials and methods.

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1 Supplemental Information Materials and methods. Cell culture. hmscs were isolated from bone marrow of 3 male donors, undergoing orthopedic surgery (mean age 69.7). Cells were cultured in high glucose DMEM medium (Gibco BRL, Life Technologies, Inc., Gaithersburg, MD), supplemented with 12.5% FCS (Gibco), 50 µg/ml gentamycin and 50 µg/ml of ascorbic acid. The ability to differentiate into 3 different mesenchymal lineages was proved for each donor. For all experiments a concentration of 5000 cell/cm² was used. Differentiation experiments. The labeling concentrations of 0, 2.5 and 5 nm were used of each QD type. Adipogenic differentiation was induced in monolayer in DMEM supplemented with 0.5 mm 1-methyl-3-isobuthylxanthine, 100 nm dexamethason, 10 µg/ml insulin, 100µM indomethacin and 15% FCS. Differentiation was detected by accumulation of visible lipid droplets and confirmed by oil red O staining. Osteogenesis was induced in monolayer in DMEM containing 100 nm dexamethasone, 0.05 mm ascorbate-2-phosphate, 10 mm β-glycerophosphat and 15% FCS. After three weeks Van Kossa staining showed mineralization. Long-term cocultures. Human dermal fibroblasts were isolated from the human foreskin. The cells were obtained from de-epidermized dermis using Dispase II 2.5 mg/ml (Boehringer Mannheim, Mannheim, Germany) for 24 h at 4 C. Small pieces were tripsinized for 10 min in 0.08% tripsin, after washing with 10% FCS in DMEM and filtration, the cells were taken for cultivation. Chondrocytes were obtained from the cartilage and synovial cells from synovial tissue, both from bovine knee joint. Tissues were mechanically minced and finally enzymatic digested upon 24 h incubation in an orbital mixer incubator (180 rpm) at 37 C in high glucose Dulbecco s modified Eagle s medium (DMEM, Gibco, Karlsruhe, Germany) containing 10% FCS, 100 U/ml penicillin and 100 µg/ml streptomycin (both Biochrom AG, Berlin, Germany) and 2 mg/ml collagenase A (Roche, Mannheim, Germany). After incubation the cell suspension was passed through a 70 µm nylon mesh filter (Becton Dickinson, Heidelberg, Germany). The filtrate was centrifuged at 800 g for 8 min, the supernatant removed, and the cell

2 pellet was resuspended in DMEM. Cells were counted and their viability was determined in a haemocytometer via the trypan-blue (Invitrogen, Karlsruhe, Germany) exclusion test. Each cell type was mixed with hmsc immediately before plating 1:1 and cultured as described above. Cytotoxicity, metabolic and cell proliferation assays. Cytotoxic/membrane integrity was analyzed using LDH cytotoxic assay kit (Roche, Basel, Switzerland) at timepoints 24, 48 and 72 h after labeling. BrdU incorporation (colorimetric) was done with BrdU proliferative assay kit (Roche, Basel, Switzerland). An alamar blue test was made as described. 17 MTT assay was done as described elsewhere. All measures were made at time-points till 3 week in continuous culture and analyzed with ELISA reader Safire (Tecan Trading AG, Zürich, Switzerland). Transmission electron microscopy was done using Zeiss EM 10 microscope as described. 18 LC3 immunocytology. The cells, labeled with QD, were seeded in 24-well plates. After 24 and 72 h postlabeling cells were fixed with 4% formaldehyde in PBS for 15 min. After permeabilization unspecific binding sites were saturated with 1.5% normal goat serum (DakoCytomation, Glostrup, Denmark) for 30 min, then cells were incubated with the rabbit polyclonal anti-lc3 antibody (Santa Cruz Biotech., Santa Cruz, US) diluted 1:2000 in PBS containing 0.25% BSA (Sigma, Deisenhoven, Germany) overnight at 4 C. The binding site in QD525-labeled cells was detected with a CY3-conjugated second antibody (Dianova, Hambourg, Germany), in QD605- labeled cells with a CY2-conjugated, both diluted 1:100 in PBS and incubated for 30 min. Additionally, the cells nuclei were stained with 4'-6-Diamidino-2-phenylindole (DAPI; Serva, Heidelberg, Germany). Negative controls were incubated with rabbit IgG 1 and the second antibody, respectively. Photodocumentation. Microscopic observations were performed using Carl Zeiss Axiovert 200 microscope equipped with camera and using Axiovision software.

3 Statistic. Experiments were mean of 3 6 independent measurements. The data are shown as mean ± SEM. Statistical analysis was performed by using the Student s t test (twotailed). Differences were taken as statistically significant at p 0.05.

4 Supplemental Figures Fig.S1. LDH cytotoxic/membrane stability assay. 24 h after labeling procedure. No significant differences between LDH activity of QD labeled cells and unlabeled control were found. * p 0.05 Fig.S2. Alamar blue metabolic assay. Day 3 after labeling procedure. The differences between labeled and unlabeled

5 Fig.S3. Proliferative assay via BrdU incorporation. Day 3 after labeling procedure. The differences between labeled and unlabeled Fig.S4. MTT metabolic/proliferative assay. Day 7 after labeling procedure. The differences between labeled and unlabeled