Journal: The Journal of Cell Biology

Article Title: FGF-23–Klotho signaling stimulates proliferation and prevents vitamin D–induced apoptosis

doi: 10.1083/jcb.200803024

Figure Lengend Snippet: Signaling events induced by FGF-23–Klotho. Immunoblotting showing that FGF-23 or Klotho alone have no effect on kinase activity in PTEC or FHs74Int cells. Combined effects of FGF-23 and Klotho show increased phosphorylation of ERK1/2, p38, JNK, AKT, IκB, and GSK-3β. α-Tubulin was used as a loading control.

Article Snippet: Human renal PTEC were obtained from Clontech Laboratories, Inc. FHs74Int human small intestine epithelial cells were acquired from the American Type Culture Collection.

Techniques: Western Blot, Activity Assay, Phospho-proteomics, Control

Journal: The Journal of Cell Biology

Article Title: FGF-23–Klotho signaling stimulates proliferation and prevents vitamin D–induced apoptosis

doi: 10.1083/jcb.200803024

Figure Lengend Snippet: FGF-23–Klotho prevents vitamin D–induced apoptosis. (A) ELISA analysis of 1α-hydroxylase expression showing no significant changes in PTEC cells exposed to FGF-23 or Klotho alone but greatly decreased levels when exposed to both FGF-23 and Klotho. Small molecule inhibitors against Ras and PI3K were sufficient to provide marginal rescue of this decrease in expression. No significant changes were found for treatment of FHs74Int cells. Graphs represent mean ± SD ( n = 3). *, P < 0.05. (B) Flow cytometry analysis for TUNEL staining of cells exposed to exogenous vitamin D showing that it caused extremely high levels of apoptosis. Addition of FGF-23 and Klotho was sufficient to rescue most of the vitamin D–induced apoptosis, whereas FGF-23 or Klotho alone did not. PI3K inhibitor prevented this rescue, whereas Ras inhibitor had no effect. (C) ELISA for active caspase-3 levels, showing the same patterns as observed with the TUNEL analysis. Graphs represent mean ± SD ( n = 3). *, P < 0.001.

Article Snippet: Human renal PTEC were obtained from Clontech Laboratories, Inc. FHs74Int human small intestine epithelial cells were acquired from the American Type Culture Collection.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Flow Cytometry, TUNEL Assay, Staining

Journal: Pediatric Research

Article Title: Hydrocortisone and Indomethacin Negatively Modulate EGF-R Signaling in Human Fetal Intestine

doi: 10.1203/pdr.0b013e318155ac3b

Figure Lengend Snippet: Figure 1. EGF-induced activation of EGF-R signaling pathways in FHs74Int cells. (A) Stimulation with EGF resulted in the appearance of phosphorylated EGF-R in the cell membrane (arrows). (B) FHs74Int cells were treated as indicated and effects of EGF were controlled in the presence of ZD1839. UV-treatment was used as a positive control for Jnk and p38 activation.

Article Snippet: Human small intestine epithelial FHs74Int cells (ATCC, Manassas, VA; CCL-241) were cultured in a humidified 5% CO2 atmosphere at 37°C in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FCS, 2 mM glutamine, 100 unit/mL penicillin, 100 g/mL streptomycin, nonessential amino acids, sodium pyruvate (0.5 mM), oxaloacetic acid (1 mM), and insulin (0.2 units/mL).

Techniques: Activation Assay, Protein-Protein interactions, Membrane, Positive Control

Journal: Pediatric Research

Article Title: Hydrocortisone and Indomethacin Negatively Modulate EGF-R Signaling in Human Fetal Intestine

doi: 10.1203/pdr.0b013e318155ac3b

Figure Lengend Snippet: Figure 2. HC potentiated EGF-induced proliferation of FHs74Int cells. Fhs74Int cells were cultured and cell numbers were counted on d 3 (white bars), 6 (shaded bars) and 9 (black bars). EGF alone and in combination with HC synergistically increased cell proliferation, whereas ZD1839 inhibited these effects. PD and LY inhibited EGF-induced cell proliferation. †Untreated vs EGF-treated; ‡EGF vs EGFHC, EGFHCInd, EGFHCIndZD, EGFPD, or EGFLY; *p 0.05.

Article Snippet: Human small intestine epithelial FHs74Int cells (ATCC, Manassas, VA; CCL-241) were cultured in a humidified 5% CO2 atmosphere at 37°C in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FCS, 2 mM glutamine, 100 unit/mL penicillin, 100 g/mL streptomycin, nonessential amino acids, sodium pyruvate (0.5 mM), oxaloacetic acid (1 mM), and insulin (0.2 units/mL).

Techniques: Cell Culture

Journal: Pediatric Research

Article Title: Hydrocortisone and Indomethacin Negatively Modulate EGF-R Signaling in Human Fetal Intestine

doi: 10.1203/pdr.0b013e318155ac3b

Figure Lengend Snippet: Figure 3. Activation of signal transduction pathways in FHs74Int cells by EGF and HC. (A) HC had no effect on EGF-R activation or EGF-induced Erk- and Akt-activation. (B) Expression of p21 and cyclinD1 was up-regulated by EGF, while HC had no effect on them.

Article Snippet: Human small intestine epithelial FHs74Int cells (ATCC, Manassas, VA; CCL-241) were cultured in a humidified 5% CO2 atmosphere at 37°C in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FCS, 2 mM glutamine, 100 unit/mL penicillin, 100 g/mL streptomycin, nonessential amino acids, sodium pyruvate (0.5 mM), oxaloacetic acid (1 mM), and insulin (0.2 units/mL).

Techniques: Activation Assay, Transduction, Expressing

Journal: Pediatric Research

Article Title: Hydrocortisone and Indomethacin Negatively Modulate EGF-R Signaling in Human Fetal Intestine

doi: 10.1203/pdr.0b013e318155ac3b

Figure Lengend Snippet: Figure 6. EGF up-regulated COX-2 mRNA and PGE2 protein levels. FHs74Int cells were grown as indicated without (A) or with (B) pretreatment with ZD, PD, and LY. (C) PGE2 leads to activation of EGF-R, Erk, and Akt pathways. †Untreated vs EGF-treated; ‡EGF vs EGFHC; *p 0.05.

Article Snippet: Human small intestine epithelial FHs74Int cells (ATCC, Manassas, VA; CCL-241) were cultured in a humidified 5% CO2 atmosphere at 37°C in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FCS, 2 mM glutamine, 100 unit/mL penicillin, 100 g/mL streptomycin, nonessential amino acids, sodium pyruvate (0.5 mM), oxaloacetic acid (1 mM), and insulin (0.2 units/mL).

Techniques: Activation Assay

Journal: Pediatric Research

Article Title: Hydrocortisone and Indomethacin Negatively Modulate EGF-R Signaling in Human Fetal Intestine

doi: 10.1203/pdr.0b013e318155ac3b

Figure Lengend Snippet: Figure 7. EGF and HC increased migration of FHs74Int cells. (A) FHs74Int cells were grown in Transwell chambers as indicated. Relative migration rates compared with untreated cells (C, control) are shown. †Untreated vs EGF-, HC-, and PGE2-treated; ‡EGF vs EGFHC and EGFHCInd; *p 0.05. (B) Morphology of the FHs74Int cells changed toward fibroblastoid during treatment with EGF and HC. Without any treatment, FHsInt74 cells were cuboidal and expressed cortical actin (white solid arrows). Upon EGF- and HC-treatment, they become elongated and actin was reorganized into stress- fibers (open white arrowheads), which was inhibited by the addition of Ind.

Article Snippet: Human small intestine epithelial FHs74Int cells (ATCC, Manassas, VA; CCL-241) were cultured in a humidified 5% CO2 atmosphere at 37°C in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FCS, 2 mM glutamine, 100 unit/mL penicillin, 100 g/mL streptomycin, nonessential amino acids, sodium pyruvate (0.5 mM), oxaloacetic acid (1 mM), and insulin (0.2 units/mL).

Techniques: Migration, Control