human trophoblast cell line jeg 3  (ATCC)

Journal: bioRxiv

Article Title: Trophoblast ferroptosis restricts SARS-CoV-2 spread in the placenta

doi: 10.64898/2026.01.26.701742

Figure Lengend Snippet: (A) Viral titers of SARS-CoV-2 Delta variant in supernatants from JEG-3 cells infected over 48 hours with an MOI of 1. Titers were determined via plaque assay. (B-E) Protein expression of iron transport and storage proteins in SARS-CoV-2-infected JEG-3 cells at 6, 24, and 48 hpi, as determined by western blotting. (F) Schematic of regulation of iron transport and storage expression by iron regulatory proteins (IRPs). (G,H) Protein expression of IRPs in infected JEG-3 cells, as determined by western blotting. (I) Impact of excess iron supplementation (FAC) on viral titers in JEG-3 cells, as determined via plaque assay. (J-N) Protein expression of ferroptosis markers and inhibitors in SARS-CoV-2-infected JEG-3 cells over course of infection. Expression was measured by western blotting. (O) Cytotoxicity of SARS-CoV-2 Delta variant in JEG-3 cells as measured by LDH release over course of infection. (P) Cytotoxicity of JEG-3 cells treated with increasing concentrations of the FSP1 inhibitor, iFSP1, with or without infection. Cytotoxicity was determined via LDH assay. (Q) Cytotoxicity of small molecules iFSP1 and RSL3 in JEG-3 cells with or without SARS-CoV-2 infection, as measured by LDH release. (R) Viral titers in supernatants from JEG-3 cells infected with SARS-CoV-2 for 48 hours, following pre-treatment and post-inoculation exposure to iFPS1 and RSL3. Viral titers were determined via plaque assay. Data represents average of n=3 independent replicates per group ± SEM. Significance determined by Student’s unpaired t-test except for P and Q which were determined by two-way ANOVA and R which was determined by one-way ANOVA. For all tests, *p<0.05, **p<0.01, ***p<0.001, ****p<<0.001.

Article Snippet: The human trophoblast cell line JEG-3 (ATCC HTB-36) was cultured in DMEM/F-12 (Gibco, 11330032) supplemented with 10% Fetal Bovine Serum (FBS) (Gibco, 16140071).

Techniques: Variant Assay, Infection, Plaque Assay, Expressing, Western Blot, Lactate Dehydrogenase Assay

Journal: Placenta

Article Title: Reduced SMEK1 regulates trophoblast migration and invasion in fetal growth restriction.

doi: 10.1016/j.placenta.2025.02.005

Figure Lengend Snippet: Fig. 1. SMEK1 was downregulated in FGR placenta Western Blots (A) and corresponding quantification (B) of SMEK1 in control and FGR placentas. (C) Immunofluorescent staining of SMEK1(red), CK7(green) in the indicated placentas. Scale bar, 100 μm. (D) Data of the red fluorescence intensity of SMEK1. (E) Immunofluorescent staining of SMEK1(red), CK7(green) in first- trimester decidual. Scale bar, 100 μm. Western Blots and corresponding quantification for SMEK1 in JEG3 cells following transfection with plasmid vector (Vec- tor) SMEK1-overexpressing construct (OE-SMEK1) (F, G), SMEK1 siRNA(si-SMEK1) or scrambled siRNA (si-NC) (G, H). Data are presented as mean ± SD based on three independently repeated experiments, and comparison between groups is carried out with Student’s t-test. *p < 0.05, **, p < 0.01, ***, p < 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The human trophoblast cell line JEG3 obtained from ATCC (American Type Culture Collection), was incubated in a constant temperature chamber (at 37 ◦C with 5 % CO2) with Dulbecco’s Modified Eagle Medium supplemented with 10 % fetal bovine serum and 1 % penicillinstreptomycin.

Techniques: Western Blot, Control, Staining, Fluorescence, Transfection, Plasmid Preparation, Construct, Comparison

Journal: Placenta

Article Title: Reduced SMEK1 regulates trophoblast migration and invasion in fetal growth restriction.

doi: 10.1016/j.placenta.2025.02.005

Figure Lengend Snippet: Fig. 2. The proliferation and apoptosis of JEG3 cells. (A–D) EDU assay and corresponding quantification of EdU-positive cells in JEG3 cells after 48h of transfected of plasmid vector (Vector) or SMEK1-overexpressing construct (OE-SMEK1), SMEK1 siRNA(si-SMEK1), scrambled siRNA (si-NC). Scale bar, 100 μm. (E–H). Flow cytometry and analysis revealed the rates of cell apoptosis. Western Blots and corresponding quantification for BCL2, BAX and Cleaved-Caspase3 in JEG3 cells following transfection with plasmid vector (Vector) SMEK1-overexpressing construct (OE-SMEK1) (I, J), SMEK1 siRNA(si-SMEK1) or scrambled siRNA (si-NC) (K, L).Data are presented as mean ± SD based on three independently repeated experiments, and comparison between groups is carried out with Student’s t-test. *p < 0.05, **, p < 0.01, ***, p < 0.001.

Article Snippet: The human trophoblast cell line JEG3 obtained from ATCC (American Type Culture Collection), was incubated in a constant temperature chamber (at 37 ◦C with 5 % CO2) with Dulbecco’s Modified Eagle Medium supplemented with 10 % fetal bovine serum and 1 % penicillinstreptomycin.

Techniques: EdU Assay, Transfection, Plasmid Preparation, Construct, Flow Cytometry, Western Blot, Comparison

Journal: Placenta

Article Title: Reduced SMEK1 regulates trophoblast migration and invasion in fetal growth restriction.

doi: 10.1016/j.placenta.2025.02.005

Figure Lengend Snippet: Fig. 4. PI3K/AKT/mTOR pathway was suppressed by knockdown of SMEK1 in JEG3 cells. (A) Bulb map of KEGG analysis for the DEGs of JEG3 cells transfected of SMEK1 siRNA compared to scrambled siRNA. Western Blots (B) and corresponding quantification (C) of p-AKT, AKT, and mTOR in JEG3 cells transfected of scrambled siRNA (si-NC) and SMEK1 siRNA(si-SMEK1). Western Blots (D) and corresponding quantification (E) of E-cadherin, N-cadherin, and Vimentin in JEG3 cells transfected of scrambled siRNA (si-NC) and SMEK1 siRNA(si-SMEK1). (F) FITC-phalloidin staining of actin filament in the JEG3 cells after 48h of transfected of scrambled siRNA (si-NC) and SMEK1 siRNA(si-SMEK1). Scale bar, 10 μm. (G) Data of the FITC fluorescence intensity of F-actin. Data are presented as mean ± SD based on three independently repeated experiments, and comparison between groups is carried out with Student’s t-test. *p < 0.05, **, p < 0.01, ***, p < 0.001.

Article Snippet: The human trophoblast cell line JEG3 obtained from ATCC (American Type Culture Collection), was incubated in a constant temperature chamber (at 37 ◦C with 5 % CO2) with Dulbecco’s Modified Eagle Medium supplemented with 10 % fetal bovine serum and 1 % penicillinstreptomycin.

Techniques: Knockdown, Transfection, Western Blot, Staining, Fluorescence, Comparison

Journal: Placenta

Article Title: Reduced SMEK1 regulates trophoblast migration and invasion in fetal growth restriction.

doi: 10.1016/j.placenta.2025.02.005

Figure Lengend Snippet: Fig. 5. SMEK1 regulated cell migration and invasion through PI3K/AKT/mTOR pathway in JEG3 cells. Western Blots (A) and corresponding quantification (B) of E- cadherin, N-cadherin in JEG3 cells transfected of scrambled siRNA (si-NC), SMEK1 siRNA(si-SMEK1) or treated with Recilisib (50 μM, 24h). Wound healing migration assay (C) and corresponding quantification (D) of JEG3 cells transfected of scrambled siRNA (si-NC), SMEK1 siRNA(si-SMEK1) or treated with Recilisib. Scale bar, 200 μm. Transwell insert assay (E) and corresponding quantification (F) of JEG3 cells transfected of scrambled siRNA (si-NC), SMEK1 siRNA(si-SMEK1) or treated with Recilisib. Scale bar, 50 μm. In each group, 5 views were randomly selected and the invaded cells were quantified. Data are presented as mean ± SD based on three independently repeated experiments, and comparison between groups is carried out with Student’s t-test. *p < 0.05, **, p < 0.01, ***, p < 0.001.

Article Snippet: The human trophoblast cell line JEG3 obtained from ATCC (American Type Culture Collection), was incubated in a constant temperature chamber (at 37 ◦C with 5 % CO2) with Dulbecco’s Modified Eagle Medium supplemented with 10 % fetal bovine serum and 1 % penicillinstreptomycin.

Techniques: Migration, Western Blot, Transfection, Comparison