Journal: Aging (Albany NY)

Article Title: The long non-coding RNA FLJ46906 binds to the transcription factors NF-κB and AP-1 and regulates expression of aging-associated genes

doi: 10.18632/aging.101528

Figure Lengend Snippet: Expression of the lncRNA FLJ46906 increases with aging in fibroblasts. ( A ) The expression of FLJ46906 in fibroblasts from older donors (63, 68, and 70 years old, aged in-vivo ), as measured by qPCR, is 2.8 fold higher than in fibroblasts from young donors (18, 23, and 27 years old; n = 3 (triplicate samples from each donor), mean ± SD, *p < 0.05). ( B ) Neonatal fibroblasts aged in-vitro by longterm culture show increasing expression of FLJ46906 with increasing population doublings (PD), as measured by qPCR (n = 3 (cells from triplicate tissue culture dishes), mean ± SD, *p < 0.05, **p < 0.01).

Article Snippet: Adult primary human dermal fibroblasts established from abdominal skin of donors aged 18, 23, 27, 63, 68, and 70 years were purchased from Kurabo (Japan).

Techniques: Expressing, In Vivo, In Vitro

Journal: Aging (Albany NY)

Article Title: The long non-coding RNA FLJ46906 binds to the transcription factors NF-κB and AP-1 and regulates expression of aging-associated genes

doi: 10.18632/aging.101528

Figure Lengend Snippet: The lncRNA FLJ46906 does not regulate the expression of neighboring genes. ( A ) Genome map around FLJ46906 gene locus on chromosome 6. ( B ) The expression of two of the FLJ46906’s neighboring genes is not affected by knockdown of FLJ46906, as determined by qPCR; two other neighboring genes are not expressed in neonatal fibroblasts (n = 3, mean ± SD).

Article Snippet: Adult primary human dermal fibroblasts established from abdominal skin of donors aged 18, 23, 27, 63, 68, and 70 years were purchased from Kurabo (Japan).

Techniques: Expressing, Knockdown

Journal: Mechanisms of ageing and development

Article Title: A new model to investigate UVB-induced cellular senescence and pigmentation in melanocytes

doi: 10.1016/j.mad.2020.111322

Figure Lengend Snippet: UVB induces senescence in human epidermal melanocytes. (A) Representative pictures of control and UVB-irradiated mela-nocytes stained for SA-β-Gal on D9 of the experiment. Scale bar 50 μm. (B) Percentage of SA-β-Gal positive melanocytes. For each group at least 300 cells were analyzed. (C) p21, pp53 (serin 15), p53 and Lamin B1 protein expression of UVB-irradiated and the respective control cells were analyzed by western blot on days 4 and 9 of the experiment. Representative pictures are shown. Lysates from HDFs passage 35 and HDFs treated with 33 μM Cisplatin were used as positive controls. GAPDH was used as a protein loading control. (D-E) – Analysis of relative intensity of the Western blot on days 4 (D) and 9 (E) was performed in ImageJ software. Results are presented as mean values ± SD of at least 3 independent experiments. * < 0.05; ** < 0.01; *** < 0.001 and n.s. no statistical significance.

Article Snippet: Human dermal fibroblasts (HDFs) derived from neonatal foreskin (ATCC, Manassas, USA #SCRC-1042), HEK293-T cells and U2OS cells were cultivated with normal DMEM supplemented with 10 % FCS, 2 % glutamine and 1 % penicillin/streptomycin.

Techniques: Control, Irradiation, Staining, Expressing, Western Blot, Software

Journal: Mechanisms of ageing and development

Article Title: A new model to investigate UVB-induced cellular senescence and pigmentation in melanocytes

doi: 10.1016/j.mad.2020.111322

Figure Lengend Snippet: Presence of UVB-irradiated HDFs induces multinucleation and modulate melanogenesis of melanocytes in a co-culture system. (A) Schematic representation of the experimental setup for co-culture. (B) Analysis of cell surface area was performed in ImageJ. At least 30 cells from three independent experiments were analyzed. (C) Percentage of multinucleated cells was calculated for melanocytes cultivated in the presence of UVB-irradiated or control HDFs on D3. (D) Intracellular melanin content measurement was performed on D3 of co-culture. Bars represent mean values of three independent experiments ± SD. (E) Representative pictures of melanocytes stained by FM. Scale bar 50 μm. Insets display detail of cytoplasmic regions containing melanosomes (F) The FM stained melanocytes were separated into groups according to the number of melanosomes per cell. Results are presented as mean values ± SD of three independent replicates. * < 0.05; ** < 0.01; *** < 0.001 and n.s. no statistical significance.

Article Snippet: Human dermal fibroblasts (HDFs) derived from neonatal foreskin (ATCC, Manassas, USA #SCRC-1042), HEK293-T cells and U2OS cells were cultivated with normal DMEM supplemented with 10 % FCS, 2 % glutamine and 1 % penicillin/streptomycin.

Techniques: Irradiation, Co-Culture Assay, Control, Staining

Journal: Biomedicines

Article Title: A Platform for Testing the Biocompatibility of Implants: Silicone Induces a Proinflammatory Response in a 3D Skin Equivalent

doi: 10.3390/biomedicines12010224

Figure Lengend Snippet: Schematic overview of experimental methods for construction of the 3D skin equivalent and 3D skin equivalent with an implant using a three-dimensional collagen gel culture. A single cell suspension of human fibroblasts (HDFp) obtained from confluent monolayers was passaged and cultivated in a type I collagen gel matrix (HDFp-collagen gel) ( A ). Silicone samples were cut out using a biopsy punch ( B ) and implanted in HDFp–collagen gel constructs ( C ). After solidification of HDFp–collagen gel constructs, keratinocytes were placed on the gel matrix to build up a three-dimensional skin-like structure ( D ). The skin constructs were cultured for another 3 days for keratinocytes to expand ( E ). The following air-lift culturing technique ( F ) allowed for cell differentiation. Eventually, the 3D equivalent had a skin-specific stratification and morphology ( G , H ).

Article Snippet: It was based on fibroblasts (HDFp, primary human dermal fibroblasts, CELLnTEC Advanced Cell Systems AG, Bern, Switzerland), supplemented by human epidermal keratinocyte cells (HDEK, human epidermal keratinocytes, CELLnTEC).

Techniques: Suspension, Construct, Cell Culture, Cell Differentiation

Journal: Biomedicines

Article Title: A Platform for Testing the Biocompatibility of Implants: Silicone Induces a Proinflammatory Response in a 3D Skin Equivalent

doi: 10.3390/biomedicines12010224

Figure Lengend Snippet: Expression of tight junctions in unaffected human skin ( a , b ) and the 3D skin equivalent with ( e , f ) and without implant ( c , d ). Immunofluorescence of ZO-1 ( a,c,e ) and E-Cadherin ( b,d,f ). The staining of ZO-1 (red) and E-cadherin (green) clearly showed the cell membranes of keratinocytes in the epidermis. The cell nuclei were counterstained with 4,6-diamidino-2-phenylindole (DAPI) (blue). Human fibroblasts showed no expression of ZO-1 or E-cadherin at cell membranes. The red box marks the area shown in detail to its right. Arrows signify TJ. Controls in which the primary antibody was omitted showed no specific staining. Scale bar = 50 μm.

Article Snippet: It was based on fibroblasts (HDFp, primary human dermal fibroblasts, CELLnTEC Advanced Cell Systems AG, Bern, Switzerland), supplemented by human epidermal keratinocyte cells (HDEK, human epidermal keratinocytes, CELLnTEC).

Techniques: Expressing, Immunofluorescence, Staining

Journal: Stem Cell Research & Therapy

Article Title: Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

doi: 10.1186/s13287-015-0149-3

Figure Lengend Snippet: Skin fibroblast donor information

Article Snippet: Primary dermal fibroblast cultures from PBD-ZSD patients and controls were obtained from the Kennedy Krieger Institute and Coriell Institute Cell Repositories (CIRC), respectively.

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

doi: 10.1186/s13287-015-0149-3

Figure Lengend Snippet: Gene expression and epigenetic profiles of fibroblasts and iPSCs. a Dendrogram depicting unsupervised hierarchical clustering analysis of gene expression data from PBD-ZSD patient and healthy control skin fibroblasts and iPSCs. Analysis was based on log2-transformed gene expression scores from 575 probe sets with coefficient of variation (CV) greater than 0.25 and conducted using average linkage and Euclidean distance. Color bar represents log-2 transformed gene expression values. b Dendrogram depicting results of unsupervised hierarchical clustering analysis of DNA methylation data from PBD-ZSD patient and healthy control skin fibroblasts and iPSCs. Analysis was based on 4073 DNA methylation assays interrogating autosomal CpG loci with CV greater than 0.75 and the 10th largest and smallest β-value being greater than 0.6 and less than 0.4, respectively, in order to represent the most variable loci. Clustering was performed using average linkage and Pearson dissimilarity distance. Color bar represents β-values

Article Snippet: Primary dermal fibroblast cultures from PBD-ZSD patients and controls were obtained from the Kennedy Krieger Institute and Coriell Institute Cell Repositories (CIRC), respectively.

Techniques: Gene Expression, Control, Transformation Assay, DNA Methylation Assay

Journal: Stem Cell Research & Therapy

Article Title: Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

doi: 10.1186/s13287-015-0149-3

Figure Lengend Snippet: Estimated mtDNA genome copy number in PBD-ZSD patient and healthy control-derived cells. The numbers of mtDNA genomes per cell were estimated by quantitative PCR in triplicate. Total DNA isolated from human 143B cells and human 143B rho cells (devoid of mtDNA) served as positive (+) and negative (−) controls. Fibroblast and iPSC DNA samples were analyzed on different 96-well plates with embedded positive controls ( grey ) yielding similar results (330 and 267 mtDNA genomes per cell, respectively). All data from the fibroblast ( light blue : controls; dark blue : patients) and iPSC ( light red : controls; dark red : patients) plates were normalized so that internal positive control provided an estimate of 297 mtDNA genomes per cell (average of mtDNA estimates above). Error bars represent the high and low estimates of mtDNA genomes per cell in a given sample. Fibroblast control4 is derived from fibroblast culture AG04446 obtained from a 48-year-old healthy male donor (Coriell Cell Repositories). iPSC induced pluripotent stem cell

Article Snippet: Primary dermal fibroblast cultures from PBD-ZSD patients and controls were obtained from the Kennedy Krieger Institute and Coriell Institute Cell Repositories (CIRC), respectively.

Techniques: Control, Derivative Assay, Real-time Polymerase Chain Reaction, Isolation, Positive Control

Journal: Stem Cell Research & Therapy

Article Title: Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

doi: 10.1186/s13287-015-0149-3

Figure Lengend Snippet: Relative saturated very long chain fatty acid levels in cultured patient- and control-derived cells

Article Snippet: Primary dermal fibroblast cultures from PBD-ZSD patients and controls were obtained from the Kennedy Krieger Institute and Coriell Institute Cell Repositories (CIRC), respectively.

Techniques: Cell Culture, Control