Journal: Biology of reproduction

Article Title: Differential endometrial gene expression in pregnant and nonpregnant sows.

doi: 10.1095/biolreprod.109.082321

Figure Lengend Snippet: FIG. 3. Immunohistochemical localization of IL6R (a and b), LIFR (d and e), IL11RA (g and h), MUC4 (j and k), ERBB3 (m and n), FGF9 (p and q), FGFR3 (s and t), in endometrial tissue from nonpregnant (a, d, g, j, m, p, and s) and pregnant (b, e, h, k, n, q, and t) sows. Corresponding negative controls: IL6R preadsorption control (c), LIFR preadsorption control (f), IL11RA preadsorption control (i), murine IgG2a Isotype control (l), ERBB3 preadsorption control (o), murine IgG3 Isotype control (r), and murine IgG2a Isotype control (u). DG, deep glands; LE, luminal epithelium; S, stroma; SG, superficial glands. Bar ¼ 200 lm (a–f and j–u) and 500 lm (g–i).

Article Snippet: Primary antibody Manufacturer Product code Source Clonality Isotype Concentration (lg/ml) HIERa Primary antibody incubationb Anti-human LIFR R&D Systems (Minneapolis, MN) AF-249-NA Goat Polyclonal IgG 0.50 Ph 9 2 h RT Anti-human IL6R Santa Cruz (Santa Cruz, CA) sc-661 Rabbit Polyclonal IgG 0.10 Ph 6 2 h RT Anti-human IL11RA Santa Cruz sc-993 Rabbit Polyclonal IgG 8.00 Ph 9 2 h RT Anti-human FGF9 Santa Cruz sc-8413 Mouse Monoclonal IgG3 20.00 Ph 9 2 h RT Anti-human FGFR3 Santa Cruz sc-13121 Mouse Monoclonal IgG2a 8.00 Ph 9 2 h RT Anti-human MUC4 Abnova (Taipei City, Taiwan) H00004585-M07 Mouse Monoclonal IgG2a 0.20 Ph 6 2 h RT Anti-human ERBB3 Santa Cruz sc-285-G Goat Polyclonal IgG 0.67 Ph 9 o/n 48C a HIER, heat induced epitope retrieval. b RT, room temperature; o/n, over night.

Techniques: Immunohistochemical staining, Control

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Downregulation of miRNA-214 in cancer-associated fibroblasts contributes to migration and invasion of gastric cancer cells through targeting FGF9 and inducing EMT

doi: 10.1186/s13046-018-0995-9

Figure Lengend Snippet: MiR-214 inhibited the tumor-promoting ability of CAFs by directly targeting FGF9. a Ten predicted genes as potential targets of miR-214 were analyzed by qRT-PCR. Five of them were overexpressed in CAFs compared to NFs, in which FGF9 represented the most prominent one. b The protein expression levels of FGF9 in NFs and CAFs. c and d The migration and invasion abilities of cultured GC cells were suppressed after adding FGF9 neutralizing antibody into CAF-CM. e Among five upregulated genes as mentioned above, only FGF9 mRNA expression was reduced in CAF miR-214 compared to CAF NC . f FGF9 protein expression in CAFs was also suppressed in CAF miR-214 compared to CAF NC . g Two predicted binding sites between miR-214 and FGF9 3’UTR. h The relative luciferase activity was significantly suppressed in cells co-transfected with wild-type binding site vectors of FGF9 3’UTR in the presence of pre-miR-214 (* P < 0.05, ** P < 0.01)

Article Snippet: After incubation with bovine serum albumin for 20 min at room temperature, the slides were incubated with primary antibody FGF9 (1:150, R&D systems) overnight at 4 °C and then with HRP-conjugated secondary antibody at room temperature for 30 min. DAB was used for staining.

Techniques: Quantitative RT-PCR, Expressing, Migration, Cell Culture, Binding Assay, Luciferase, Activity Assay, Transfection

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Downregulation of miRNA-214 in cancer-associated fibroblasts contributes to migration and invasion of gastric cancer cells through targeting FGF9 and inducing EMT

doi: 10.1186/s13046-018-0995-9

Figure Lengend Snippet: Correlation between FGF9 expression and clinicopathological parameters of gastric adenocarcinomas

Article Snippet: After incubation with bovine serum albumin for 20 min at room temperature, the slides were incubated with primary antibody FGF9 (1:150, R&D systems) overnight at 4 °C and then with HRP-conjugated secondary antibody at room temperature for 30 min. DAB was used for staining.

Techniques: Expressing

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Downregulation of miRNA-214 in cancer-associated fibroblasts contributes to migration and invasion of gastric cancer cells through targeting FGF9 and inducing EMT

doi: 10.1186/s13046-018-0995-9

Figure Lengend Snippet: FGF9 expression in primary tumor ( a – d ) and lymph node metastatic sites ( e – h ) of GC. a The high expression of FGF9 in CAFs, but low expression in tumor cells. b CAFs and tumor cells were both positive for FGF9. c The high expression of FGF9 in tumor cells, but low staining in CAFs. d CAFs and tumor cells were both negative for FGF9. e and f FGF9 staining was high in CAFs but low in tumor cells of both intestinal type ( e ) and diffuse type ( f ). g and h CAFs and tumor cells were both positive for FGF9 ( g , intestinal type and h , diffuse type). i and j The proportion of high expressed FGF9 in CAFs and tumor cells of primary tumor ( i ) and lymph node metastatic sites ( j ). The percentage of high expressed FGF9 in LNCAFs ( k ) and LNT ( l ) of intestinal-type and diffused and mixed-type

Article Snippet: After incubation with bovine serum albumin for 20 min at room temperature, the slides were incubated with primary antibody FGF9 (1:150, R&D systems) overnight at 4 °C and then with HRP-conjugated secondary antibody at room temperature for 30 min. DAB was used for staining.

Techniques: Expressing, Staining

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Downregulation of miRNA-214 in cancer-associated fibroblasts contributes to migration and invasion of gastric cancer cells through targeting FGF9 and inducing EMT

doi: 10.1186/s13046-018-0995-9

Figure Lengend Snippet: High FGF9 expression in lymph node metastatic sites CAFs was associated with poor prognosis in patients with GC. a – c The Kaplan–Meier survival analysis revealed that the FGF9 level in primary tumor CAFs, primary tumor cells, and lymph node metastatic sites tumor cells was not associated with prognosis. d High FGF9 expression in lymph node metastatic site CAFs was significantly associated with poor prognosis in patients with GC

Article Snippet: After incubation with bovine serum albumin for 20 min at room temperature, the slides were incubated with primary antibody FGF9 (1:150, R&D systems) overnight at 4 °C and then with HRP-conjugated secondary antibody at room temperature for 30 min. DAB was used for staining.

Techniques: Expressing

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Downregulation of miRNA-214 in cancer-associated fibroblasts contributes to migration and invasion of gastric cancer cells through targeting FGF9 and inducing EMT

doi: 10.1186/s13046-018-0995-9

Figure Lengend Snippet: High FGF9 level in lymph node metastatic site CAFs and tumor cells were associated with poor prognosis in diffuse and mixed–type GC. a and b High FGF9 level in lymph node metastatic site CAFs or tumor cells were not associated with prognosis in intestinal-type GC. c and d High FGF9 level in lymph node metastatic CAFs or tumor cells was associated with poor prognosis in diffuse and mixed–type GC

Article Snippet: After incubation with bovine serum albumin for 20 min at room temperature, the slides were incubated with primary antibody FGF9 (1:150, R&D systems) overnight at 4 °C and then with HRP-conjugated secondary antibody at room temperature for 30 min. DAB was used for staining.

Techniques:

Journal: eLife

Article Title: Hair follicle dermal condensation forms via Fgf20 primed cell cycle exit, cell motility, and aggregation

doi: 10.7554/eLife.36468

Figure Lengend Snippet: ( A ) Quantification of scratch wound closure of E13.5 growth-arrested primary dermal fibroblasts treated with FGF9 (200 ng/ml) or FGF9 and SU5402 (20 µM). At 24 hr, FGF9 treatment resulted in greater wound-closure at 8 hr (36.17 ± 5.04%, p=0.0490) and at 24 hr (97.11 ± 3.09%, p=0.0133) compared to DMSO control (all treatments n = 5 experiments, each performed with freshly extracted primary dermal fibroblast cell population). Wound closure was not altered when cells were treated with both FGF9 and SU5402 inhibitor (p=0.2563). ( B ) Quantification of transwell migration assay of E13.5 primary dermal fibroblasts. Migration was significantly increased when FGF9 (200 ng/ml) was added to lower or both upper and lower chambers (p=0.0003 and p=0.0010, respectively; for both n = 6 experiments, each performed with freshly extracted primary dermal fibroblast cell population). No statistical difference was observed between FGF9 treatments (p=0.4033). ( C ) Quantification of nuclei density in E13.5 dermis explants treated for 3 hr with beads loaded with FGF9 (100 µg/ml) or 0.1% BSA vehicle control. Density measured from a single optical slice at mid-bead. FGF9 bead induces an increase in density within 15 µm radius from the bead relative to BSA control (p=0.033, n = 7 explants), but not between 15 and 30 µm radius (p=0.236, n = 7 explants). ( D ) Whole-mount RNA in situ hybridization of dermal samples treated with FGF9 beads for 3, 8, and 16 hr. Induction of Spry4 and Dusp6 , but not Sox2 expression (purple) was observed around the bead at all time points ( n indicates induction/total samples, induction was tested in two independent experiments with skin samples derived from at least two different litters.). ( E ) 2 hr EdU incorporation into dermis organ cultures after overnight incubation with BSA (left), FGF20 (center), FGF9 (right) loaded beads. Note the increased number of proliferating cells around the FGF9 bead ( n = 5 explants). Error bars represent SD. *, p≤0.05; **, p≤0.01; ***, p≤0.001. Scale bar = 30 µm. See also and . 10.7554/eLife.36468.027 Figure 6—figure supplement 1—source data 1. Values used to quantify FGF9-induced cellular changes. Values used to quantify fibroblast wound closure in the presence of DMSO, SU5402, FGF9 +DMSO, or FGF9 +SU5402 . Values used to quantify E13.5 primary fibroblast transwell migration in control, FGF9 in lower chamber, and FGF9 in seeding and lower chambers . Values used to quantify fibroblast density in response to BSA or FGF9-loaded beads at 0–15 µm and 15–30 µm distance from the bead .

Article Snippet: Peptide, recombinant protein , FGF9 human recombinant protein , R and D Systems , 273-F9 , .

Techniques: Control, Transwell Migration Assay, Migration, RNA In Situ Hybridization, Expressing, Derivative Assay, Incubation

Journal: eLife

Article Title: Hair follicle dermal condensation forms via Fgf20 primed cell cycle exit, cell motility, and aggregation

doi: 10.7554/eLife.36468

Figure Lengend Snippet: 3 hr incubation of BSA-, FGF20- or FGF9-loaded beads with E13.5 wildtype ( A, B, C ) or Fucci mKO ( D ) dermises. Cdkn1a expression was assayed in these samples using ( A ) whole-mount RNA in situ hybridization of dermal samples. Cdkn1a expression (purple) was not observed around the bead (0/20 each condition, in four independent experiments with skin samples derived from four different litters.) ( B ) section radioactive in situ hybridization (0/5 each condition in two independent experiments with skin samples from two different litters), and ( C ) section in situ hybridization (0/5 each condition in two independent experiments with skin samples from two different litters). ( D ) Cell cycle exit was assessed using Fucci mKO reporter allele (representing G 0 /G 1 cell cycle phase) in the 30 µm surrounding the center of the bead (0/10 each condition in two independent experiments with skin samples from two different litters). ( E ) Quantification of percent total cells surrounding the bead positive for Fucci-mKO . No significant difference was observed between any of the groups. Error bars represent SD. Scale bar = 50 µm. See also and . 10.7554/eLife.36468.029 Figure 6—figure supplement 2—source data 1. Values used to quantify FGF20 or FGF9 induced Fucci-mKO expression. Values used to quantify the percent of total cells expressing Fucci-mKO 30 µm surrounding the center of the bead .

Article Snippet: Peptide, recombinant protein , FGF9 human recombinant protein , R and D Systems , 273-F9 , .

Techniques: Incubation, Expressing, RNA In Situ Hybridization, Derivative Assay, In Situ Hybridization