Journal: Cell reports

Article Title: SILAC Analysis Reveals Increased Secretion of Hemostasis-Related Factors by Senescent Cells

doi: 10.1016/j.celrep.2019.08.049

Figure Lengend Snippet: (A) A schematic outlining the strategy for SILAC-based identification of SASP proteins. See for time lines of how cells were made quiescent or senescent and how quiescent and senescent conditioned media (CM) were produced. (B) Reactome pathway analysis of proteins secreted at >2-fold levels by senescent compared with quiescent HCA2 cells. Pathways related to hemostasis are identified by boxes. (C) Heatmap showing individual levels of proteins, identified by SILACs, that participate in hemostasis and are secreted by quiescent (QUI) and senescent (SEN [IR]) HCA2 normal human fibroblasts. (D) CM were collected from quiescent (QUI) and senescent (SEN [IR]) cells, concentrated, and analyzed using western blotting for the indicated SASP factors. Cells remaining on the plate at the time of CM collection were counted for normalization. Mindin protein levels and Ponceau S staining were used as loading controls. Fold changes are mean ± SEM. *p < 0.05 and **p < 0.01. See also and .

Article Snippet: WI-38 and IMR-90 are normal primary human embryonic lung fibroblasts (obtained from the American Type Culture Collection).

Techniques: Multiplex sample analysis, Produced, Western Blot, Staining

Journal: Cell reports

Article Title: SILAC Analysis Reveals Increased Secretion of Hemostasis-Related Factors by Senescent Cells

doi: 10.1016/j.celrep.2019.08.049

Figure Lengend Snippet: (A) mRNA was extracted from HCA2 cells made quiescent (QUI) or induced to senesce by doxorubicin (SEN [DOXO]), and analyzed using qPCR for the indicated genes, using actin mRNA for normalization. The data are shown in a.u., and the bars show mean ± SEM. *p < 0.05 and **p < 0.01, two-tailed, unpaired t test. (B) CM were collected from cells treated as in (A), concentrated and analyzed using western blotting for the indicated SASP factors. Cells were counted for normalization. Mindin protein levels and Ponceau S staining were used as loading controls. (C) IMR-90 fibroblasts were depleted of mtDNA by culture in ethidium bromide until senescent (21 days) (labeled as MiDAS). RNA was extracted and analyzed using qPCR for the indicated factors. Heatmap represents individual values for three experiments. *p < 0.05, one-way ANOVA. See also .

Article Snippet: WI-38 and IMR-90 are normal primary human embryonic lung fibroblasts (obtained from the American Type Culture Collection).

Techniques: Two Tailed Test, Western Blot, Staining, Labeling

Journal: Cell reports

Article Title: SILAC Analysis Reveals Increased Secretion of Hemostasis-Related Factors by Senescent Cells

doi: 10.1016/j.celrep.2019.08.049

Figure Lengend Snippet:

Article Snippet: WI-38 and IMR-90 are normal primary human embryonic lung fibroblasts (obtained from the American Type Culture Collection).

Techniques: Recombinant, Modification, Sequencing, Multiplex sample analysis, Labeling, Enzyme-linked Immunosorbent Assay, Sandwich ELISA, Reverse Transcription, Software

Journal: Biotechnology and bioengineering

Article Title: Engineering dense tumor constructs via cellular contraction of extracellular matrix hydrogels.

doi: 10.1002/bit.28561

Figure Lengend Snippet: FIGURE 1 Harnessing fibroblast contractility to engineer dense carcinoma constructs. (a) Schematic illustration of fibroblast‐driven contraction of collagen fibers around cancer spheroids. Blue shapes represent fibroblasts, black lines represent collagen fibers, and green clusters represent spheroids. (b) Side by side comparison of H&E‐stained sections of dense breast carcinoma constructs (left) and biopsied mammary ductal carcinoma tissue (right). Scale bars = 80 μm. (c) Time‐course imaging of gross contraction and compaction of dense carcinoma constructs. Scale bars = 200 μm. (d) Box plot of the relative area change over time for gels seeded at 2.5 × 105 HLF/mL (n = 6) and 5 × 105/mL (n = 7). **p < 0.01. ***p < 0.001. See Section 2 for details of statistical analyses. (e) Box plot of the corresponding increase in estimated collagen density over time for gels seeded for acellular gels and 5 × 105 HLF/mL (n = 5 per group).

Article Snippet: Normal human lung fibroblasts (HLF) (American Type Culture Collection [ATCC] and Lonza) were cultured in fibroblast basal medium with low serum kit (ATCC) and 2% fetal bovine serum (FBS, ATCC).

Techniques: Construct, Comparison, Staining, Imaging

Journal: Biotechnology and bioengineering

Article Title: Engineering dense tumor constructs via cellular contraction of extracellular matrix hydrogels.

doi: 10.1002/bit.28561

Figure Lengend Snippet: FIGURE 2 Effect of fibroblast cell type and seeding density on tissue contraction. (a–c) Time‐course imaging of gross contraction and compaction of dense carcinoma constructs formed using several types of fibroblasts. Constructs were seeded at 5 × 105/mL (n = 3 per group). Scale bars = 3 mm. Similar kinetics and magnitude of contraction using human dermal fibroblasts (HDF, a), human lung cancer associated fibroblasts (CAF, b), and human lung fibroblasts (HLF, c). (d and e) Time‐course imaging of gross contraction and compaction for up to 3 weeks in culture. Scale bars = 3 mm. Constructs were seeded at 0.5 × 105 HLF/mL (n = 6, panel d) and 5 × 105/mL (n = 7, e). (f) Relative area change over time as a measure of contraction for each type of fibroblast during 7 days of culture as shown in (a–c). No significant differences were measured. HDF constructs were slightly less contracted at 7 days, matching the qualitatively larger size in time course images. (g) Relative area change over time as a measure of contraction. Similar final volumes were reached with each seeding density. Contraction was more gradual with a seeding density of 0.5 × 105 HLF/mL.

Article Snippet: Normal human lung fibroblasts (HLF) (American Type Culture Collection [ATCC] and Lonza) were cultured in fibroblast basal medium with low serum kit (ATCC) and 2% fetal bovine serum (FBS, ATCC).

Techniques: Imaging, Construct

Journal: Biotechnology and bioengineering

Article Title: Engineering dense tumor constructs via cellular contraction of extracellular matrix hydrogels.

doi: 10.1002/bit.28561

Figure Lengend Snippet: FIGURE 5 Local anisotropy is similar in constructs formed with various types of contractile cells. (a) H&E‐staining of sectioned dense carcinoma constructs formed with cancer associated fibroblasts (CAF), human dermal fibroblasts (HDF), and human lung fibroblasts (HLF). Seven days, scale bars = 30 μm. (b) Collagen type I staining of sectioned dense carcinoma constructs formed with CAF, HDF, and HLF. Seven days, scale bars = 20 μm. (c) Box plot of the relative fluorescent intensity in binned regions of interest at the specified distances from spheroid for dense carcinoma constructs formed with CAF (white), HDF (light gray), and HLF (dark gray) (n = 9 per group). *p < 0.05, **p < 0.01. One way ANOVA. (d) Box plot of fiber orientation relative to spheroid tangent line and distance from spheroid for dense carcinoma constructs. Same shading as (d) to indicate fibroblast type; CAF (white), HDF (light gray), and HLF (dark gray) (n = 9 per group). One way ANOVA. ns, not significant.

Article Snippet: Normal human lung fibroblasts (HLF) (American Type Culture Collection [ATCC] and Lonza) were cultured in fibroblast basal medium with low serum kit (ATCC) and 2% fetal bovine serum (FBS, ATCC).

Techniques: Construct, Staining

Journal: Nanoscale Advances

Article Title: Drug delivery with a pH-sensitive star-like dextran-graft polyacrylamide copolymer

doi: 10.1039/d2na00353h

Figure Lengend Snippet: Viability of the normal human lung fibroblast HEL299 cells, treated for 48 h with: (a) – free Dox or D- g -PAAan-Dox in Dox-equivalent concentrations, (b) – free Cis or D- g -PAAan-Cis in Cis-equivalent concentrations; * p < 0.05 compared to free drugs.

Article Snippet: Lung carcinoma human A549 and normal human lung fibroblast HEL299 cell lines were purchased from AddexBio Technologies (San Diego, CA, USA).

Techniques:

Journal: Nanoscale Advances

Article Title: Drug delivery with a pH-sensitive star-like dextran-graft polyacrylamide copolymer

doi: 10.1039/d2na00353h

Figure Lengend Snippet: IC 50 of free drugs and drug-loaded D- g -PAAan nanoparticles on HEL299 cell viability

Article Snippet: Lung carcinoma human A549 and normal human lung fibroblast HEL299 cell lines were purchased from AddexBio Technologies (San Diego, CA, USA).

Techniques:

Journal: Inflammation

Article Title: The Priming Potential of Interferon Lambda-1 for Antiviral Defense in the Oral Mucosa

doi: 10.1007/s10753-022-01624-1

Figure Lengend Snippet: Type III IFN receptor expression in human gingival keratinocytes. A : Detection of IL10R2 , IFNLR1 , and GAPDH mRNA in various keratinocytes and fibroblasts with (+) or without (-) cDNA by RT-PCR. M, marker. B : Quantitative analysis of IFNLR1 mRNA expression by real-time PCR in various keratinocytes and fibroblasts. Data represent the mean ± standard deviation (SD) of triplicate assays. C : Detection of cell surface IFN-λR1 by flow cytometry. The black line represents IFN-λR1, and the gray area shows the isotype control. A representative histogram from three independent experiments is shown. Values are the mean fluorescence intensity of IFN-λR1 expression and shown as a fold increase in IFN-λR1 expression from that with the control. D : HGK were treated with the indicated concentrations of IFN-λ1 for 30 min. Whole-cell lysates prepared from these cells were immunoblotted with anti-phospho-STAT1 (pSTAT1), anti-STAT1, or anti-GAPDH antibodies. A representative blot is shown. The bar graph shows the integrated signal intensities of the pSTAT1/STAT1 ratio. Data represent the mean ± SD of triplicate assays. * P < 0.05 and ** P < 0.01 versus untreated cells (0) (Dunnett’s multiple comparison test). E : Human gingival sections were stained with an isotype and anti-IFN-λR1 antibody. Sections were counterstained with hematoxylin. Representative images of 3 samples each are shown. The right image is a high-magnification image of the square area. A-D : Data show a representative of at least three independent experiments. OBA-9; an immortalized human gingival keratinocyte cell line. HGK; human primary gingival keratinocytes. HaCaT; an immortalized human skin keratinocyte cell line. HEK; human primary epidermal keratinocytes. HGF; human primary gingival fibroblasts. HFL-III; normal human lung fibroblasts.

Article Snippet: Normal human lung fibroblasts were provided by the Japanese Collection of Research Bioresources Cell Bank.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Marker, Real-time Polymerase Chain Reaction, Standard Deviation, Flow Cytometry, Control, Fluorescence, Comparison, Staining