Journal: Pediatric research

Article Title: Lyophilization of human amniotic fluid is feasible without affecting biological activity.

doi: 10.1038/s41390-019-0632-0

Figure Lengend Snippet: Fig. 2 Cellular proliferation of FHs74, small intestinal epithelial cell line, as measured by absorbance at 490 nm of growth media, lipopolysaccharide, and three preparations of human amniotic fluid (hAF): control (C), filtered (F), and lyophilized (L). (*p < 0.01 represents significant difference in IEC proliferation in C vs F vs L; **p < 0.001 represents significant difference in IEC proliferation in L vs F). The data represent mean of quadruplicate measurements.

Article Snippet: Pediatric Research _#####################_ Suppression of inflammatory response with hAF IL-8 was measured selectively because of its reported role in modulating LPS-mediated inflammatory processes and the fact that this chemokines is significantly induced by LPS in NEC models.11 Human small intestine epithelial cells FHs74 (ATCC, Manassas, VA) were cultured to 80% confluence in 6-well plates.

Techniques: Control

Journal: The FASEB Journal

Article Title: CHRFAM7A: a human-specific α 7-nicotinic acetylcholine receptor gene shows differential responsiveness of human intestinal epithelial cells to LPS

doi: 10.1096/fj.14-268037

Figure Lengend Snippet: Identification of CHRFAM7A. A) RT-PCR of CHRFAM7A, CHRNA7, and GAPDH of mRNA isolated from human epithelial cell lines reveals the presence of transcripts in HEK (HEK293l and HEK293W), liver cancer (SKHep), ovarian cancer [OvCar8 (2) and OvCar8(6)], pancreatic (PANC1 and DU145), colon cancer (HCT116), prostate cancer (PC3), and lung cancer (H1299) epithelial cells. B) Overlapping the 3′ and 5′sequences obtained with the primer (bold, underlined) from CaCo2 cells revealed the nucleotide sequence of human epidermal CHRFAM7A that has the exons A (blue) and B (red) of FAM7 and the exons 5–10 of CHRNA7 (black), which when translated, reveals the unique 27 amino acid sequence of CHRFAM7A (C) that distinguishes the human-specific gene from the amino termini found in CHRNA7 variants. D) The CHRFAM7A PCR primers shown in the text were selected to detect the nucleotide sequences that encode the unique CHRFAM7A peptide (C), whereas the CHRNA7A primers enable detection of transcript variants 1 and 2 mRNAs that encode α7nAChRs that differ in their amino terminus by the underlined peptide sequence in (D). The common 386 amino acid sequence that is shared by CHRFAM7A and both CHRNA7 variants is shown in (E).

Article Snippet: Normal human small intestine epithelial cells (FHs-Int-74) were also obtained from the ATCC (CCL-241).

Techniques: Reverse Transcription Polymerase Chain Reaction, Isolation, Sequencing

Journal: The FASEB Journal

Article Title: CHRFAM7A: a human-specific α 7-nicotinic acetylcholine receptor gene shows differential responsiveness of human intestinal epithelial cells to LPS

doi: 10.1096/fj.14-268037

Figure Lengend Snippet: CHRFAM7A is expressed in epithelial cells. PC3 cells were transfected with plasmid encoding a CHRFAM7A-DDK fusion protein and the following day lysed and immunoblotted with antibodies to the DDK tag (A). Untransfected cells were used as control, and molecular weights (kiloDaltons) were determined with molecular weight standards. In (B), RT-PCR was used to detect CHRNA7 and CHRFAM7A expression in gut 1) CaCo2, 2) KM12, 3) HT29, 4) KM20, 5) LS174, 6) HCT116, 7) SW24, and 8) Colo205 epithelial cells. In 3 instances (CaCoT, HCT116T, and KM20L), the same cell line from 2 alternative sources was analyzed. As shown in (C) and (D), both CHRNA7 and CHRFAM7A were measured by qRT-PCR and the relative expression levels compared to that measured in CaCo2 cells.

Article Snippet: Normal human small intestine epithelial cells (FHs-Int-74) were also obtained from the ATCC (CCL-241).

Techniques: Transfection, Plasmid Preparation, Control, Molecular Weight, Reverse Transcription Polymerase Chain Reaction, Expressing, Quantitative RT-PCR

Journal: The FASEB Journal

Article Title: CHRFAM7A: a human-specific α 7-nicotinic acetylcholine receptor gene shows differential responsiveness of human intestinal epithelial cells to LPS

doi: 10.1096/fj.14-268037

Figure Lengend Snippet: Differential regulation of CHRNA7 and CHRFAM7A by LPS treatment of gut epithelial cells. Triplicate cultures of 1) CaCo2, 2) KM12, 3) HT29, 4) KM20, 5) LS174, 6) HCT116, 7) SW24, 8) Colo205, and 9) FHs or T84 epithelial cells were treated for 3 hours with LPS as described in the text. A) The effects of LPS on CHRNA7 gene expression measured by qPCR, normalized to GAPDH, and changes from controls assessed using the ∆∆Ct method. B) The cDNA prepared from the same cell lysates was assessed for CHRFAM7A gene expression. In (C) and (D), the ratios of CHRFAM7A expression to CHRNA7 in control and LPS-stimulated cells were compared.

Article Snippet: Normal human small intestine epithelial cells (FHs-Int-74) were also obtained from the ATCC (CCL-241).

Techniques: Gene Expression, Expressing, Control

Journal: Clinical immunology (Orlando, Fla.)

Article Title: CVID Enteropathy is Characterized by Exceeding Low Mucosal IgA Levels and Interferon-Driven Inflammation Possibly Related to the Presence of a Pathobiont

doi: 10.1016/j.clim.2018.09.008

Figure Lengend Snippet: Interferons Drive Inhibition of Lipid Metabolism in Epithelial Cells. (a) Expression of IFNB1 and IFN type I dependent genes (RSAD2, OAS2, MX1) in duodenal biopsies from of CVID patients with (red), without (blue) enteropathy and control subjects (green), all genes passed false discovery rate <0.03 comparing E-CVID to other two groups. (b) expression of IFNG and IFN type II dependent genes (GBP1, IDO, TAP1); (c) Gene expression ratios between E-CVID and control individuals (y axis) and between interferon treated versus untreated human epithelial cells (x axis; n=3 per group), left panel both types of interferons; middle panel interferon type I; right panel interferon type II. (d) Expression of genes defined as lipid metabolism by Gene Ontology and downregulated in E-CVID, representative genes are indicated; left panel: median values in E-CVID and noE-CVID tissues; right panel: individual values in cells treated and untreated with both interferons, red is high and blue is low expression; (e) intracellular cholesterol levels in epithelial cells treated or untreated with both types of interferons data presented as mean±s.e.m.; each column number of biological replicates equals 14 (**p<0.0001 by one-tailed Wilcoxon test); (f) model of CVID enteropathy.

Article Snippet: Human epithelial cell line (FHs 74 Int) was acquired from ATCC and maintained according to ATCC protocol using Hybri-Care Medium (cat# ATCC® 46-XTM) with addition of 10% FBS (ATCC, cat# 30–2020, lot: 62144240), 5% Penicillin Streptomycin (Corning, REF 30–002-CI) and 30 ng/ml epidermal growth factor (EGF; Tonbo Biosciences, cat# 21–8356-M001).

Techniques: Inhibition, Expressing, Control, Gene Expression, One-tailed Test