Journal: Advanced healthcare materials

Article Title: Designing Inherently Photodegradable Cell-Adhesive Hydrogels for 3D Cell Culture.

doi: 10.1002/adhm.202100632

Figure Lengend Snippet: Figure 2. A) Live/dead staining of normal human dermal fibroblasts (NHDF) with calcein-AM (green, live cells) and propidium iodide (red, dead cells) embedded in GelMA-free PEGMA/PEGDMA (Gel-N0) and GelMA-containing (Gel-N10) hydrogels on day 1, 7, 14 postencapsulation, followed by imaging with confocal microscopy (Leica TCS SPE, scale bar: 200 µm). B-i) NHDF embedded in Gel-N10 hydrogels on day 1 postencapsulation. Comparison of HepG2 cells encapsulated in (ii) Gel-N10 and (iii) Gel-N0 hydrogels (scale bar: 100 µm). C) Proliferation behavior of encapsulated HepG2 cells in Gel-N10 and Gel-N0 hydrogels over a period of 7 d assessed by PrestoBlue assay (n = 3). D) Toxicity evaluation of Gel-N10 photodegradation products by MTT proliferation assay with HeLa cells after 72 h exposure (n = 3). The concentration of nondiluted degradation products was 53 mg mL−1. Data are presented as mean ± SD and statistically evaluated by (C) Student’s t-test and (D) one-way ANOVA. *, **, *** represent p < 0.05, 0.01, and 0.001, respectively.

Article Snippet: Normal human dermal fibroblasts (NHDF), human liver cancer cells (HepG2), and HeLa cells were purchased from PromoCell GmbH (Heidelberg, Germany).

Techniques: Staining, Imaging, Confocal Microscopy, Comparison, Prestoblue Assay, Proliferation Assay, Concentration Assay

Journal: Genes to Cells

Article Title: Derivation of human differential photoreceptor cells from adult human dermal fibroblasts by defined combinations of CRX, RAX , OTX2 and NEUROD

doi: 10.1111/gtc.12127

Figure Lengend Snippet: Induction of retina-specific genes in human dermal fibroblasts by the retroviral infection of genes for defined transcription factors. (A) RT-PCR analysis for photoreceptor-specific genes in cultured human dermal fibroblasts (NHDF) obtained from Lonza after gene transfer of several kinds of transcription factors. Recoverin, blue opsin and PDE6c genes were up-regulated by CRN transduction. ‘Negative control’: amplified water as a negative control. ‘GFP’: cultured fibroblasts after retroviral gene transfer of the GFP gene as another negative control. ‘w/o’: cultured fibroblasts without gene transfer as the other negative control. ‘SPPO’: SOX2, POU1F1, PAX6 and OTX2 . ‘SPO’: SOX2, POU1F1 and OTX2 . ‘CRN’: CRX, RAX and NEUROD . ‘Human retina’: human retinal tissue as a positive control. The amount of cDNA as a template was a half in the positive control. (B) RT-PCR analysis for photoreceptor-specific genes in cultured human dermal fibroblasts (NHDF) obtained from Promo Cell after gene transfer of several transcription factors. Recoverin, blue opsin and PDE6c genes were up-regulated by CRN or CRNO transduction. ‘w/o’: cultured fibroblasts without gene transfer as a negative control. ‘GFP’: cultured fibroblasts after retroviral gene transfer of the GFP gene as another negative control. ‘CRNO’: CRX, RAX, NEUROD and OTX2 . (C) RT-PCR analysis for photoreceptor-specific genes in cultured human dermal fibroblasts (HDF-a) obtained from ScienCell after gene transfer of several transcription factors. Recoverin, blue opsin and PDE6C genes were up-regulated by CRN or CRNO transduction. Expression levels of blue opsin were increased by additional OTX2 gene transduction. ‘w/o’: cultured fibroblasts without gene transfer as a negative control. ‘GFP’: cultured fibroblasts after retroviral gene transfer of the GFP gene as another negative control. ‘CRNO’: CRX , RAX , NEUROD and OTX2 . ‘1’, ‘2’ and ‘3’ mean independently cultured, transfected and harvested cells by the same combination of CRN genes. (D) Immunocytochemistry using antibodies to rhodopsin and blue opsin (green). Nuclei were stained with DAPI (blue). Experiments were carried out at 2 weeks after infection. The cells in the left panel and the right panel are CRN-infected Fib#2 and Fib#1, respectively. Scale bars represent 10 μm.

Article Snippet: Three strains of cultured human dermal fibroblasts were used: one was obtained from Lonza (NHDF), another was from Promo Cell (NHDF) and the other was from ScienCell (HDF-a).

Techniques: Infection, Reverse Transcription Polymerase Chain Reaction, Cell Culture, Transduction, Negative Control, Amplification, Positive Control, Expressing, Transfection, Immunocytochemistry, Staining

Journal: Oncotarget

Article Title: Evaluation of an Actinomycin D/VX-680 aurora kinase inhibitor combination in p53-based cyclotherapy

doi:

Figure Lengend Snippet: (A) Pretreatment with low dose ActD decreases the proportion of 8N cells caused by VX-680 in NHDF cells. ActD was added 24 hours post-seeding. VX-680 was added 48 hours post-seeding. Cells were harvested 96 hours after seeding. (B) NHDFs were treated with 4 nM ActD for 72 hours, fixed and stained with Giemsa (top panel). In the bottom panel, ActD was removed from the medium and cells were allowed to proliferate for 6 days before Giemsa staining.

Article Snippet: NHDF cells were bought from Promocell.

Techniques: Staining

Journal: Oncotarget

Article Title: Evaluation of an Actinomycin D/VX-680 aurora kinase inhibitor combination in p53-based cyclotherapy

doi:

Figure Lengend Snippet: (A and B) NHDF cells were treated for 48 hours with VX-680 alone (labelled as no preinc.), or treated for 48 hours with VX-680 after preincubating with the indicated amounts of ActD for 24 hours. Thereafter cells were fixed, stained Giemsa (A) and counted (B). Arrows in (A) indicate cells with abnormal nuclei. (C and D) NHDF cells treated as in A and B were allowed to recover for 6 days in drug-free medium. The growth of the cultures in drug-free medium is quantified in panel (C) by counting the total number of cells per field before and after the recovery period. The average number of cells per field with aberrant nuclei before and after the recovery period is shown in panel (D). Error bars correspond to standard deviations.

Article Snippet: NHDF cells were bought from Promocell.

Techniques: Staining

Journal: Oncotarget

Article Title: Evaluation of an Actinomycin D/VX-680 aurora kinase inhibitor combination in p53-based cyclotherapy

doi:

Figure Lengend Snippet: (A) NHDF cells were treated with the indicated amounts of ActD for 4 or 12 hours. p53 and p21 were analysed by Western blotting. α-tubulin was detected as a loading control. (B) Example of nuclear abnormalities in NHDF cells expressing a dominant negative form of p53. The percentage of cells with this sort of aberrant nuclei was 2.5 fold higher than in the corresponding cells with intact p53. Severely damaged cells, such as the one in the center of this picture, only appeared in the fibroblasts with dominant negative p53.

Article Snippet: NHDF cells were bought from Promocell.

Techniques: Western Blot, Expressing, Dominant Negative Mutation