Journal: Genes & development

Article Title: ATR inhibition induces synthetic lethality in mismatch repair-deficient cells and augments immunotherapy.

doi: 10.1101/gad.351084.123

Figure Lengend Snippet: Figure 2. ATRi preferentially induces DNA damage in MMR-d cells. (A,B) U2OS cells were transfected with control, MHL1, or MSH2 siRNA for 2 d; pulse-labeled with EdU for 20 min; and then treated with 2 µM ATRi (AZD6738) for 4 h. γ-H2AX and biotinylated EdU were analyzed by immunofluorescence. Representative images are shown in A. Frac- tions of cells positive for γ-H2AX and EdU were quantified as shown in B. Scale bar, 20 µm. (C) U2OS cells transfected with control, MLH1, or MSH2 siRNA were treated with 10 μM CDK1i (RO-3360) for 20 h followed by 2 μM ATRi (AZD6738) for 4 h. Fractions of cells positive for γ-H2AX and EdU were quantified. (D) U2OS cells were transfected with control, MLH1, MSH1, and MUS81 siRNAs as indicat- ed and then treated with 2 μM ATRi (AZD6738) for 4 h. Fractions of cells positive for γ-H2AX and EdU were quantified. Scale bar, 20 µm. (E) U2OS cells were transfected with control or MLH1 siRNA and treated with 2 μM ATRi (AZD6738) for 4 h. Cells were analyzed by PLA using γ-H2AX antibody, MSH2 antibody, or both. The numbers of PLA foci in individual cells were quantified. (Red bars) Mean PLA foci per nucleus in cell popula- tions. (F) U2OS cells were treated as in E and analyzed by PLA using γ-H2AX antibody, PCNA antibody, or both. The numbers of PLAs in individual cell foci were quantified as in E.

Article Snippet: For the γ-H2AX-PCNA PLA, U2OS cells transfected with control or MLH1 siRNA were incubated with ATRi AZD6738 for 4 h. Subsequently, cells were fixed in ice-cold methanol and permeabilized in 0.5% Triton X, and PLAwas performed using the Duolink PLA kit (Sigma) according to themanufacturer’s instructions using the γ-H2AX (Cell Signaling Technologies) and PCNA (Santa Cruz Biotechnologies) antibodies.

Techniques: Transfection, Control, Labeling, Immunofluorescence

Journal: Oncogenesis

Article Title: PRAS40 promotes colorectal cancer stemness by enhancing glycolysis through triggering PGK1 acetylation

doi: 10.1038/s41389-025-00594-x

Figure Lengend Snippet: A The expression of AKT1S1 mRNA in COAD and READ samples from TCGA. B The expression of AKT1S1 mRNA in CRC samples from GSE89076 . C – F IHC analyses with anti-PRAS40 antibody. Representative images C , H-scores D , E , and overall survival rate of CRC patients F . G–N colon cancer formation in Akt1s1 +/+ and Akt1s1 -/- mice. Genotyping results G , western blotting analyses in colon tissues H , body weights I , representative images J , quantitation of tumors K , L , HE staining M and IHC staining with anti-PCNA antibody N . Data represent the mean ± SD. ** P < 0.01; *** P < 0.001.

Article Snippet: Antibodies were purchased for detection of PRAS40 (2691S), PCNA (13110), PCAF (3378 T) (Cell Signaling); PGK1 (17811-1-AP), CD133 (66666-1-Ig), α-Tubulin (66031-1-Ig), β-Actin (66009-1-Ig), CD44 (15675-1-AP) (Proteintech); SIRT7 (ab259968), anti-phosphotryosine (ab179530) (Abcam); and pan-Acetyl (sc-8649)(Santa Crus Biotechnology).

Techniques: Expressing, Western Blot, Quantitation Assay, Staining, Immunohistochemistry

Journal: Scientific Reports

Article Title: Establishment and Characterization of Human Germline Stem Cell Line with Unlimited Proliferation Potentials and no Tumor Formation

doi: 10.1038/srep16922

Figure Lengend Snippet: ( A ) RT-PCR showed the expression of UCHL1, RET, GPR125, GFRA1, PLZF, MAGEA4, THY1, PCNA and SV40 in the immortalized human male germline stem cells. ACTB was used as a loading control of total RNA, whereas RNA without RT (RT-) but with PCR served as a negative control. ( B,C ) Western blots revealed the expression of SV40, PCNA, UCHL1, and RET in the human SSC line ( B ) at different passages and primary human SSCs ( C ). ACTB was utilized as a loading control of loading proteins, while replacement of primary antibodies with PBS served as negative controls (NC). Notes: passage 2 (P2); passage 5 (P5); passage 8 (P8); S1, S2, and S3 indicated 3 independent experiments of primary human SSCs.

Article Snippet: After permeabilization with 0.4% Triton X-100 and blocking with 5% donkey serum (Maibio), the sections were incubated with primary antibodies, including PCNA (Santa Cruz), UCHL1 (AbD Serotec), GPR125 (Abcam), and HumNuc (Abcam) at a 1:100 dilution overnight at 4 °C.

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Control, Negative Control, Western Blot

Journal: Scientific Reports

Article Title: Establishment and Characterization of Human Germline Stem Cell Line with Unlimited Proliferation Potentials and no Tumor Formation

doi: 10.1038/srep16922

Figure Lengend Snippet: ( A,B ) The expression of eGFP in the control seminiferous tubules of recipient mice without cell transplantation ( A ) and with transplantation of the immortalized human male germline stem cells ( B ). Scale bars in ( A,B ) = 200 μm. ( C–H ) Immunohistochemisty illustrated the expression of PCNA ( C ), UCHL1 ( D ), GPR125 ( E ), MAGEA4 ( F ), and HumNuc ( G ), as well as co-expression of UCHL1 and HumNuc ( H ) in the seminiferous tubules of recipient mice with transplantation of the immortalized human male germline stem cells. Scale bars in ( C–E ) = 20 μm; scale bar in ( F,G ) = 30 μm; scale bar in ( H ) = 10 μm.

Article Snippet: After permeabilization with 0.4% Triton X-100 and blocking with 5% donkey serum (Maibio), the sections were incubated with primary antibodies, including PCNA (Santa Cruz), UCHL1 (AbD Serotec), GPR125 (Abcam), and HumNuc (Abcam) at a 1:100 dilution overnight at 4 °C.

Techniques: Expressing, Control, Transplantation Assay

Journal: Scientific Reports

Article Title: Establishment and Characterization of Human Germline Stem Cell Line with Unlimited Proliferation Potentials and no Tumor Formation

doi: 10.1038/srep16922

Figure Lengend Snippet: The primer sequences of genes used for RT-PCR.

Article Snippet: After permeabilization with 0.4% Triton X-100 and blocking with 5% donkey serum (Maibio), the sections were incubated with primary antibodies, including PCNA (Santa Cruz), UCHL1 (AbD Serotec), GPR125 (Abcam), and HumNuc (Abcam) at a 1:100 dilution overnight at 4 °C.

Techniques: Sequencing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Super enhancer-driven LINC01013 mediates hypoxia-induced mitochondrial dysfunction by HSPA9 to determine pulmonary arterial smooth muscle cell fate

doi: 10.1007/s00018-025-06071-3

Figure Lengend Snippet: Silencing of LINC01013 inhibited hypoxia-induced hPASMC proliferation and inflammation. A , B hPASMC proliferation was determined by CCK8 and EdU incorporation assays ( n = 6). EdU (red), DAPI (blue), scale bar, 50 μm. C Western blotting analysis of PCNA, cyclin A and cyclin D in hPASMCs ( n = 6). D RT‒qPCR analysis showed the mRNA levels of TNF-α and IL-6 after LINC01013 knockdown, β-actin was used as a internal reference gene ( n = 6). E Western blotting analysis of TNF-α and IL-6 in hPASMCs. ( n = 6). All values are presented as the mean ± SEM. Statistical analysis was performed with one-way ANOVA. * p < 0.05, ** p < 0.01, *** p < 0.001. Nor, normoxia; Hyp, hypoxia; NC, negative control; si, siRNA

Article Snippet: The antibody against CEBPB (PB9171, BA0670, 1:500, Boster, Wuhan, China), H3K27ac (A7253, 1:500, ABclonal, Wuhan, China), H3K4me1 (A2355, 1:500, Wuhan, China), PCNA (A00125, 1:500, Boster, Wuhan, China), Cyclin A (PB0515, 1:500, Boster, Wuhan, China), Cyclin D (BM4272, 1:500, Boster, Wuhan, China), IL-6 (AF7236, 1:500, Beyotime, Shanghai, China), TNF-α (AF8208, 1:500, Beyotime, Shanghai, China), PKM2 (4053, 1:1000, Cell Signaling, MA, US), HK II (66974-1-Ig, 1:1000, Proteintech, IL, USA), PDH (2784, 1:1000, Cell Signaling, MA, US), HSPA9 (14887-1-AP, 1:5000, Proteintech, IL, USA), VDAC1 (10866-1-AP, 1:5000, Proteintech, IL, USA), and β-actin (TA-09, 1:1000, ZSGB‐BIO, Beijing, China) was incubated at 4 °C overnight, followed by incubation with appropriate horseradish peroxidase-conjugated secondary antibodies at room temperature for 1 h, and proteins were visualized with enhanced chemiluminescence reagents.

Techniques: Western Blot, Knockdown, Negative Control

Journal: ACS Omega

Article Title: Pterostilbene-Isothiocyanate Inhibits Proliferation of Human MG-63 Osteosarcoma Cells via Abrogating β-Catenin/TCF-4 Interaction—A Mechanistic Insight

doi: 10.1021/acsomega.3c02732

Figure Lengend Snippet: Effect of PTER-ITC on transcription and translation of different factors. (A) Effect of PTER-ITC on transcription level of proliferation ( PCNA ), migration ( MMP-2/9 ), metastatic ( E/N-cadherin ), and apoptotic ( Caspase-3 and PARP-1 ) markers. (B, C) Representative immunoblot images showing the effect of PTER-ITC and ZA treatments on PCNA, MMP-2/9, E/N-cadherin, active Caspase-3, and cleaved PARP-1 markers (B) along with their corresponding quantitative densitometric analysis (C). Data represented as mean ± SE of three independent experiments. Statistical significance was determined through two-way ANOVA and represented as *, **, and *** for p < 0.05, 0.01, and 0.001, respectively, when compared to respective control.

Article Snippet: Antibodies against proliferative cell nuclear antigen (PCNA) (E-AB-10010), cleaved poly [ADP-ribose] polymerase-1 (PARP-1) (E-AB-30059), matrix metalloproteinase-2 (MMP-2) (E-AB-32054), matrix metalloproteinase-9 (MMP-9) (E-AB-70247), active Caspase-3 (E-AB-22115), TCF-4 (E-AB-60206), and β-actin (E-AB-20058) were purchased from Elabscience (Texas, USA).

Techniques: Migration, Western Blot, Control

Journal: ACS Omega

Article Title: Pterostilbene-Isothiocyanate Inhibits Proliferation of Human MG-63 Osteosarcoma Cells via Abrogating β-Catenin/TCF-4 Interaction—A Mechanistic Insight

doi: 10.1021/acsomega.3c02732

Figure Lengend Snippet: List of Primers Used in This Study

Article Snippet: Antibodies against proliferative cell nuclear antigen (PCNA) (E-AB-10010), cleaved poly [ADP-ribose] polymerase-1 (PARP-1) (E-AB-30059), matrix metalloproteinase-2 (MMP-2) (E-AB-32054), matrix metalloproteinase-9 (MMP-9) (E-AB-70247), active Caspase-3 (E-AB-22115), TCF-4 (E-AB-60206), and β-actin (E-AB-20058) were purchased from Elabscience (Texas, USA).

Techniques: Sequencing