Journal: Science Advances

Article Title: GSK3β and Plk1 sequentially phosphorylate ATP–citrate lyase to promote homologous recombination

doi: 10.1126/sciadv.aeg1097

Figure Lengend Snippet: ( A to E ) HeLa cells were transfected with the indicated siRNAs. Forty-eight hours posttransfection, cells were irradiated (2 Gy) and processed for immunoblotting (A) or immunofluorescence staining 4 hours later. Representative images are shown (B). Quantification of the percentage of γH2AX-positive cells with >5 BRCA1 foci (C), BRCA1 foci intensity in γH2AX-positive cells (D), and nuclear γH2AX intensity (E) is shown. Data are presented as means ± SD from three independent experiments [ n > 300 cells per condition for (C) and (E); n > 150 cells per condition for (D)]. ( F to I ) HeLa cells were transfected with the indicated siRNAs. Forty-eight hours posttransfection, cells were irradiated (2 Gy) and processed for immunoblotting (F) or immunofluorescence staining 4 hours later. Representative images are shown (G). Quantification of the percentage of cyclin A2–positive cells with >5 γH2AX foci (H) and nuclear γH2AX intensity in cyclin A2–positive cells (I) is shown. Data are presented as means ± SD from three independent experiments [ n > 300 cells per condition for (H) and (I)]. ( J to L ) HeLa cells were transfected and irradiated as in (F) and processed for immunofluorescence staining 4 hours later. Representative images are shown (J). Quantification of the percentage of cyclin A2–positive cells with >5 BRCA1 foci (K) and BRCA1 foci intensity in cyclin A2–positive cells (L) is shown. Data are presented as means ± SD from three independent experiments [ n > 300 cells per condition for (K); n > 150 cells per condition for (L)]. Scale bars, 10 μm (B, G, and J). A.U., arbitrary units (D, E, I, and L). N.S., not significant. See also fig. S4.

Article Snippet: Primary antibodies used were as follows: ACLY (Abcam, ab40793), Flag tag (GenScript, A01868; for immunoblotting), GST (Sigma-Aldrich, G7781), HA tag (BioLegend, 16B12), Myc-tag (05-724, Millipore), MBP (New England BioLabs, E8032), pan-pSer/Thr (ABclonal, AP0893), pan-Ser-pThr-Pro (Cell Signaling Technology, D73F6; 5243), GSK3α (ABclonal, A19060), GSK3β (ABclonal, A6164), ACLY-pS455 (Cell Signaling Technology, 4331), BRCA1 (Santa Cruz Biotechnology, sc-6954), γH2AX (Abcam, ab81299), cyclin A2 (ABclonal, A19036), cyclin A2 (Proteintech, 66391-1-Ig), 53BP1 (made in-house), pan-AKT (ABclonal, A18120), RAD51 (made in-house), histone H4 (Sigma-Aldrich, 07-108), H4K16ac (Abcam, ab109463), α-tubulin (ABclonal, AC012), GFP (Abmart, M20004), Plk1 (Abcam, ab17057), and Plk1 phospho-T210 (BD Transduction, 558400).

Techniques: Transfection, Irradiation, Western Blot, Immunofluorescence, Staining

Journal: Science Advances

Article Title: GSK3β and Plk1 sequentially phosphorylate ATP–citrate lyase to promote homologous recombination

doi: 10.1126/sciadv.aeg1097

Figure Lengend Snippet: ( A to D ) Parental HeLa cells and HeLa cells stably expressing ACLY-GFP (WT or S442A) were transfected with control or ACLY siRNA. Forty-eight hours posttransfection, cells were irradiated (2 Gy) and processed for immunoblotting (A) or immunofluorescence staining 4 hours later. Representative images are shown (B). Quantification of the percentage of γH2AX-positive cells with >5 BRCA1 foci (C) and BRCA1 foci intensity in γH2AX-positive cells (D) is shown. Data are presented as means ± SD from three independent experiments [ n > 300 cells per condition for (C); n > 150 cells per condition for (D)]. ( E to H ) Parental HeLa cells and HeLa cells stably expressing ACLY-GFP (WT or S442A) were transfected with control or ACLY siRNA. Forty-eight hours posttransfection, cells were irradiated (2 Gy) and processed for immunoblotting (E) or immunofluorescence staining 4 hours later. Representative images are shown (F). Quantification of the percentage of cyclin A2–positive cells with >5 BRCA1 foci (G) and BRCA1 foci intensity in cyclin A2–positive cells (H) is shown. Data are presented as means ± SD from three independent experiments [ n > 300 cells per condition for (G); n > 150 cells per condition for (H)]. Scale bars, 10 μm (B and F). See also figs. S7 and S8.

Article Snippet: Primary antibodies used were as follows: ACLY (Abcam, ab40793), Flag tag (GenScript, A01868; for immunoblotting), GST (Sigma-Aldrich, G7781), HA tag (BioLegend, 16B12), Myc-tag (05-724, Millipore), MBP (New England BioLabs, E8032), pan-pSer/Thr (ABclonal, AP0893), pan-Ser-pThr-Pro (Cell Signaling Technology, D73F6; 5243), GSK3α (ABclonal, A19060), GSK3β (ABclonal, A6164), ACLY-pS455 (Cell Signaling Technology, 4331), BRCA1 (Santa Cruz Biotechnology, sc-6954), γH2AX (Abcam, ab81299), cyclin A2 (ABclonal, A19036), cyclin A2 (Proteintech, 66391-1-Ig), 53BP1 (made in-house), pan-AKT (ABclonal, A18120), RAD51 (made in-house), histone H4 (Sigma-Aldrich, 07-108), H4K16ac (Abcam, ab109463), α-tubulin (ABclonal, AC012), GFP (Abmart, M20004), Plk1 (Abcam, ab17057), and Plk1 phospho-T210 (BD Transduction, 558400).

Techniques: Stable Transfection, Expressing, Transfection, Control, Irradiation, Western Blot, Immunofluorescence, Staining

Journal: iScience

Article Title: Endogenous phosphatidylcholine 16:0/20:4 increases to promote liver regeneration in hepatectomy

doi: 10.1016/j.isci.2026.116962

Figure Lengend Snippet: Regeneration was advanced in the CDAHFD-treated PHx mouse model (A) Body weight changes in the CD subgroups. (B–C) Liver weight and liver index changes in the CD subgroups. (D–G) Relative mRNA levels of Cyclin D1 , Cyclin B1 , Cyclin E1 , and Ki 67 in the CD subgroups. (H) Hepatic protein levels of pathways involved in regeneration. (I–K) Gray value levels of hepatic proliferative proteins. Data were expressed as mean ± SD. One-way ANOVA between multiple groups ( n = 5, ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001, compared with CD-D0 subgroup).

Article Snippet: Cyclin D1 Rabbit mAb , ABclonal , Cat# A19038, RRID: AB_2862530.

Techniques:

Journal: iScience

Article Title: Endogenous phosphatidylcholine 16:0/20:4 increases to promote liver regeneration in hepatectomy

doi: 10.1016/j.isci.2026.116962

Figure Lengend Snippet: PC metabolism correlated well with the regeneration peak in the CDAHFD-treated PHx mouse model (A–B) The mRNA levels of Cyclin D1 and Cyclin E1 . (C–F) The mRNA levels of Pcyt1a , Pemt , Chka , and Cept1 . (G) Spearman’s correlation analysis between proliferation phenotype and PC synthesis in the CDAHFD-treated PHx mice. Data were expressed as mean ± SD. One-way ANOVA between multiple groups. n = 5, ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001, compared with CDAHFD-D0 subgroup.

Article Snippet: Cyclin D1 Rabbit mAb , ABclonal , Cat# A19038, RRID: AB_2862530.

Techniques:

Journal: iScience

Article Title: Endogenous phosphatidylcholine 16:0/20:4 increases to promote liver regeneration in hepatectomy

doi: 10.1016/j.isci.2026.116962

Figure Lengend Snippet: PC (16:0/20:4) facilitated the proliferation of Hepa 1-6 and the mouse model (A–B) CCK8 determination of Hepa 1-6 cells after 24 and 48 h of PC (16:0/20:4) challenge. (C–D) Number of Hepa 1-6 cells after 24 and 48 h of PC (16:0/20:4) challenge. (E) H&E staining of Hepa 1-6 cells after 24 and 48 h of PC (16:0/20:4) challenge. Scale bars, 50 μm. (F–I) The mRNA levels of Cyclin D1 , Ki 67 , Cyclin B1 , and Cyclin E1 . (J–L) The mRNA levels of Cyclin D1 , E1 , and Pcna determined using qPCR in the mouse liver challenged with PC (16:0/20:4). (M) Hepatic protein levels of CYCLIN D1 and PCNA in the mouse liver challenged with PC (16:0/20:4). Data were expressed as mean ± SD. One-way ANOVA between multiple groups. n = 5, ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001, compared with 0 μM group.

Article Snippet: Cyclin D1 Rabbit mAb , ABclonal , Cat# A19038, RRID: AB_2862530.

Techniques: Staining

Journal: iScience

Article Title: Endogenous phosphatidylcholine 16:0/20:4 increases to promote liver regeneration in hepatectomy

doi: 10.1016/j.isci.2026.116962

Figure Lengend Snippet: PC (16:0/20:4) facilitated the proliferation of Huh-7 cells (A–B) CCK8 determination of Huh-7 cells after 24 and 48 h of PC (16:0/20:4) challenge. (C–D) The number of Huh-7 cells after 24 and 48 h of PC (16:0/20:4) challenge. (E) H&E staining of Huh-7 cells after 24 and 48 h of PC (16:0/20:4) challenge. Scale bars, 50 μm. (F–I) The relative mRNA levels of Cyclin D1 , Ki 67 , Cyclin B1 , and Cyclin E1 . Data were expressed as mean ± SD. One-way ANOVA between multiple groups. n = 5, ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001, compared with 0 μM group.

Article Snippet: Cyclin D1 Rabbit mAb , ABclonal , Cat# A19038, RRID: AB_2862530.

Techniques: Staining

Journal: iScience

Article Title: Endogenous phosphatidylcholine 16:0/20:4 increases to promote liver regeneration in hepatectomy

doi: 10.1016/j.isci.2026.116962

Figure Lengend Snippet: PC (16:0/20:4) facilitated the proliferation of human organoids and the clinical evidence (A) Representative images of growing human liver organoids on day 1 and day 4 after PC (16:0/20:4) challenge. Scale bars, 250 μm. (B) Proliferation of growing human liver organoids determined by CellTiter-Glo3D ( n = 3/group). (C–E) The mRNA levels of Cyclin D1 , Ki 67 , and Cyclin B1 after 3 days of PC (16:0/20:4) challenge ( n = 3/group). (F) Concentration of the serum PC (16:0/20:4) among patients with MASLD in comparison with that in healthy volunteers ( n = 30–32/group). (G) Concentration of the serum PC (16:0/20:4) increased in PHx patients whose samples on day 1 were not missing. (H) Concentration of the serum PC (16:0/20:4) increased in PHx patients whose samples on day 1 were missing. Data were expressed as mean ± SD. One-way ANOVA between multiple groups. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001, compared with 0 μM group.

Article Snippet: Cyclin D1 Rabbit mAb , ABclonal , Cat# A19038, RRID: AB_2862530.

Techniques: Concentration Assay, Comparison

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Hsa_circ_0044097 Serves as a Promising Biomarker of Atherosclerosis and Its Effects on Vascular Smooth Cell Proliferation and Migration

doi: 10.1002/jbt.71031

Figure Lengend Snippet: Hsa_circ_0044097 affects the proliferation and migration of HASMCs. (A) RT‐qPCR was used to verify the transfection effect of pcDNA3.1‐hsa_circ_0044097. (B) Overexpression of hsa_circ_0044097 inhibits the levels of IL‐6 and TNF‐α in HASMCs induced by ox‐LDL. (C) Overexpression of hsa_circ_0044097 inhibited the cell proliferation of HASMCs induced by ox‐LDL. (D) Overexpression of hsa_circ_0044097 inhibited PCNA and Cyclin D1 expression in HASMCs cells induced by ox‐LDL. (E) Overexpression of hsa_circ_0044097 inhibited the cell migration of HASMCs induced by ox‐LDL. (F) Overexpression of hsa_circ_0044097 inhibited MMP‐9 and OPN expression in HASMCs cells induced by ox‐LDL. ** p < 0.01, *** p < 0.001.

Article Snippet: After blocking with non‐fat milk, the membranes were incubated overnight at 4°C with primary antibodies against PCNA (1:1000, HY‐ P80268 , MCE, Shanghai, China), Cyclin D1 (1:500, HY‐ P80633 , MCE, Shanghai, China), MMP9 (1:500, HY‐ P80756 , MCE, Shanghai, China), osteopontin (OPN) (1:500, HY‐ P86670 , MCE, Shanghai, China) and GAPDH (1:10000, HY‐P80137, MCE, Shanghai, China).

Techniques: Migration, Quantitative RT-PCR, Transfection, Over Expression, Expressing

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Hsa_circ_0044097 Serves as a Promising Biomarker of Atherosclerosis and Its Effects on Vascular Smooth Cell Proliferation and Migration

doi: 10.1002/jbt.71031

Figure Lengend Snippet: Hsa_circ_0044097 affects the proliferation and migration of HASMCs by miR‐3918. (A) MiR‐3918 expression was upregulated in AS patients. (B) The effect of transfection with miR‐3918 mimics/inhibitors on the luciferase activity of circ‐WT and circ‐MUT. (C) The RIP experiment verified the interaction between hsa_circ_0044097 and miR‐3918 in cells. (D) The Spearman correlation analysis for hsa_circ_0044097 and miR‐3918. (E) The transfection effects of pcDNA3.1‐hsa_circ_0044097 and miR‐3918 mimics were verified by RT‐qPCR. (F) The miR‐3918 mimic reversed the inhibitory effect of overexpressed hsa_circ_0044097 on the inflammatory factors (IL‐6 and TNF‐α) induced by ox‐LDL in HASMCs. (G) The miR‐3918 mimic reversed the inhibitory effect of overexpressed hsa_circ_0044097 on the proliferation induced by ox‐LDL in HASMCs. (H) The miR‐3918 mimic reversed the inhibitory effect of overexpressed hsa_circ_0044097 on the PCNA and Cyclin D1 expression induced by ox‐LDL in HASMCs. (I) The miR‐3918 mimic reversed the inhibitory effect of overexpressed hsa_circ_0044097 on the migration induced by ox‐LDL in HASMCs. (J) The miR‐3918 mimic reversed the inhibitory effect of overexpressed hsa_circ_0044097 on the MMP‐9 and OPN expression induced by ox‐LDL in HASMCs. *** p < 0.001.

Article Snippet: After blocking with non‐fat milk, the membranes were incubated overnight at 4°C with primary antibodies against PCNA (1:1000, HY‐ P80268 , MCE, Shanghai, China), Cyclin D1 (1:500, HY‐ P80633 , MCE, Shanghai, China), MMP9 (1:500, HY‐ P80756 , MCE, Shanghai, China), osteopontin (OPN) (1:500, HY‐ P86670 , MCE, Shanghai, China) and GAPDH (1:10000, HY‐P80137, MCE, Shanghai, China).

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

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Hsa_circ_0044097 Serves as a Promising Biomarker of Atherosclerosis and Its Effects on Vascular Smooth Cell Proliferation and Migration

doi: 10.1002/jbt.71031

Figure Lengend Snippet: Hsa_circ_0044097 affects the proliferation and migration of HASMCs through the miR‐3918/CBS axis. (A) Venn diagram of the downstream target genes of miR‐3918. (B) CBS expression was downregulated in AS patients. (C) The effect of transfection with miR‐3918 mimics/inhibitors on the luciferase activity of CBS‐WT and CBS‐MUT. (D) The RIP experiment verified the interaction between CBS and miR‐3918 in cells. (E) The Spearman correlation analysis for CBS and miR‐3918. (F) The transfection effects of pcDNA3.1‐hsa_circ_0044097, miR‐3918 mimics, and pcDNA3.1‐CBS were verified by RT‐qPCR. (G) The ELISA kit measured the levels of inflammatory factors (IL‐6 and TNF‐α). (H) The CCK‐8 method evaluated the proliferation ability of cells. (I) Western blot was used to detect the protein expression levels of PCNA and Cyclin D1. (J) The Transwell method evaluated the migration ability of cells. (K) Western blot was used to detect the protein expression levels of MMP‐9 and OPN. *** p < 0.001.

Article Snippet: After blocking with non‐fat milk, the membranes were incubated overnight at 4°C with primary antibodies against PCNA (1:1000, HY‐ P80268 , MCE, Shanghai, China), Cyclin D1 (1:500, HY‐ P80633 , MCE, Shanghai, China), MMP9 (1:500, HY‐ P80756 , MCE, Shanghai, China), osteopontin (OPN) (1:500, HY‐ P86670 , MCE, Shanghai, China) and GAPDH (1:10000, HY‐P80137, MCE, Shanghai, China).

Techniques: Migration, Expressing, Transfection, Luciferase, Activity Assay, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Western Blot