Journal: Molecular Oncology

Article Title: STIM2 is involved in the regulation of apoptosis and the cell cycle in normal and malignant monocytic cells

doi: 10.1002/1878-0261.13584

Figure Lengend Snippet: STIM2 knockdown alters store‐operated calcium entry (SOCE) and decreases cell proliferation in THP1 and OCI‐AML3. All data were obtained (western blot, calcium imaging, isoform, RT‐qPCR, and cell count) at 48 h postinfection for siRNA and 72 h for shRNA. (A) In THP1 protein expression of STIM2 cells (si#1: 0.407 ± 0.219; si#2: 0.410 ± 0.123) as determined by western blot relative to GAPDH ( n = 3). (B, C) The STIM2.2/STIM2.1 ratio was performed by quantitative PCR in THP1 (si#1: 4.54 ± 3.259; si#2: 6.120 ± 0.9966) (B) ( n = 4) and in OCI‐AML3 (sh#1: 6.98 ± 3.84; sh#2: 5.60 ± 2.55) (C) ( n = 5). (D) Representative trace of SOCE measured with the ratio F340/F380 in THP1 cells after STIM2 silencing ( n = 4) (siCONTROL n = 57; si#1 n = 72; si#2 n = 90). Cells were exposed to 1 μ m Thapsigargin in the absence of Ca 2+ which depletes the intracellular calcium (Ca 2+ ). Extracellular Ca 2+ concentration was then brought to 2 m m to induce SOCE. (E–G) Quantification of SOCE (siCONTROL: 53.63 ± 49.61; si#1: 93.33 ± 51.78; si#2: 134.2 ± 53.02) ( n = 6) (siCONTROL n = 87; si#1 n = 175; si#2 n = 112) (E), Thapsgiargin (TG)‐response (siCONTROL: 744 ± 142; si#1: 816 ± 229; si#2: 740 ± 257) ( n = 4) (F) and basal calcium (siCONTROL: 0.808 ± 0.108; si#1: 0.890 ± 0.169; si#2: 0.811 ± 0.123) ( n = 4) (G). (H) STIM2 protein expression in THP‐1 cells (sh#1: 0.08 ± 0.02; sh#2: 0.03 ± 0.01) was determined by western blot relative to GAPDH ( n = 3). (I) STIM2 protein expression in OCI‐AML3 cells (sh#1: 0.220 ± 0.0693; sh#2: 0.130 ± 0.03) was determined by western blot relative to GAPDH ( n = 3). (J, K) In THP1 cells, proliferation was assessed by trypan blue cell count and then reported to the numbers (Nb) of GFP cells at each cytometry point (Days 1 – 3 – 5 (D1 – 3 – 5)) ( n = 3) (J) and in OCI‐AML3 ( n = 3) (K). All numeric values are presented as the mean values ± standard error of the mean. P ‐values are calculated using one‐way ANOVA, *** P < 0.001; ** P < 0.01; * P < 0.05; NS, not significant.

Article Snippet: For differentiation into monocyte‐like cells, THP1 and OCI‐AML3 cells were treated with 1 μ m 1,25(OH) 2 D 3 (Calbiochem, Millipore, Burlington, MA, USA) for 72 h. To suppress the function of p53 protein, α‐pifithrin (PFT‐α) (TargetMol, Wellesley Hills, MA, USA) was used at 10 μ m in the medium.

Techniques: Knockdown, Western Blot, Imaging, Quantitative RT-PCR, Cell Characterization, shRNA, Expressing, Real-time Polymerase Chain Reaction, Concentration Assay, Cytometry

Journal: Molecular Oncology

Article Title: STIM2 is involved in the regulation of apoptosis and the cell cycle in normal and malignant monocytic cells

doi: 10.1002/1878-0261.13584

Figure Lengend Snippet: STIM2 knockdown induced apoptosis in THP1 and OCI‐AML3 cells. (A, B) Cell viability study was assessed in flow cytometry by annexin V/DAPI labeling and compared with shSCRAMBLE cells (Day 1—shSCRAMBLE: 4.14 ± 0.420; sh#1: 10.9 ± 10.5; sh#2: 13.3 ± 2.16) (Day 3—shSCRAMBLE: 5.43 ± 0.48; sh#1: 46.1 ± 18.6; sh#2: 25.5 ± 18.5) (Day 5—shSCRAMBLE: 9.68 ± 0.365; sh#1: 65.4 ± 16.1; sh#2: 67.6 ± 10.4) in THP1 ( n = 3) (A) and in OCI‐AML3 (Day 1—shSCRAMBLE: 5.09 ± 0.175; sh#1: 4.49 ± 0.924; sh#2: 2.72 ± 0.655) (Day 3—shSCRAMBLE: 3.27 ± 0.667; sh#1: 26.7 ± 7.47; sh#2: 2.3 ± 0.524) (Day 5—shSCRAMBLE: 5.13 ± 0.242; sh#1: 87.2 ± 1.15; sh#2: 22.3 ± 2.52) ( n = 3) (B). (C) In THP1 cell line, quantification of antiapoptotic proteins was performed by western blot (Bcl‐2—sh#1: 0.553 ± 0.0208; sh#2: 0.330 ± 0.0361) (Bcl‐XL—sh#1: 0.497 ± 0.0971; sh#2: 0.403 ± 0.150) (MCL‐1—sh#1: 0.642 ± 0.0351; sh#2: 0.590 ± 0.123) relative to GAPDH and compared with shSCRAMBLE ( n = 3). (D) Quantification of proapoptotic proteins was performed by western blot (BAX—sh#1: 2.43 ± 0.252; sh#2: 2.6 ± 0.5) (BAD—sh#1: 2.63 ± 1.01; sh#2: 3.03 ± 0.718) relative to GAPDH and compared with shSCRAMBLE ( n = 3). (E) Membrane and mitochondrial proteins were quantified by western blot (caspase‐8—sh#1: 1.42 ± 0.393; sh#2: 1.52 ± 0.407) (caspase‐9—sh#1: 0.103 ± 0.0351; sh#2: 0.0633 ± 0.0208) (cleaved (Cl) caspase‐9—sh#1: 2.11 ± 0.431; sh#2: 2.07 ± 0.347) relative to GAPDH and compared with shSCRAMBLE ( n = 3). (F) Apoptosis effector proteins were quantified by western blot (caspase‐3—sh#1: 0.553 ± 0.185; sh#2: 0.457 ± 0.0874) (cleaved caspase‐3—sh#1: 2.71 ± 0.499; sh#2: 1.57 ± 0.248) (PARP—sh#1: 0.717 ± 0.136; sh#2: 0.593 ± 0.204) (cleaved PARP—sh#1: 2.16 ± 0.320; sh#2: 1.84 ± 0.140) relative to GAPDH and compared with shSCRAMBLE ( n = 3). All numeric values are presented as the mean values ± standard error of the mean. P ‐values are calculated using one‐way ANOVA, ***P < 0.001; ** P < 0.01; * P < 0.05.

Article Snippet: For differentiation into monocyte‐like cells, THP1 and OCI‐AML3 cells were treated with 1 μ m 1,25(OH) 2 D 3 (Calbiochem, Millipore, Burlington, MA, USA) for 72 h. To suppress the function of p53 protein, α‐pifithrin (PFT‐α) (TargetMol, Wellesley Hills, MA, USA) was used at 10 μ m in the medium.

Techniques: Knockdown, Flow Cytometry, Labeling, Western Blot, Membrane

Journal: Molecular Oncology

Article Title: STIM2 is involved in the regulation of apoptosis and the cell cycle in normal and malignant monocytic cells

doi: 10.1002/1878-0261.13584

Figure Lengend Snippet: Blockage of cell cycle induced by STIM2 knockdown in THP1 and OCI‐AML3 cells. In THP1, the cell cycle was studied by flow cytometry (FCM) and western blot. (A, B) The FCM revealed an increase in the number of cells blocked in G2/M phase (shSCRAMBLE—19.77 ± 4.398; sh#1: 35.33 ± 6.047; sh#2: 32.80 ± 5.274) ( n = 3). (C, D) In THP1, quantification of the key proteins of G2/M transition were performed by western blot (CDK1—sh#1: 0.507 ± 0.0231; sh#2: 0.617 ± 0.0493) (cyclin B1—sh#1: 0.467 ± 0.0611; sh#2: 0.460 ± 0.0721) relative to GAPDH ( n = 3) (C) and in OCI‐AML3 (CDK1—sh#1: 0.527 ± 0.0451; sh#2: 0.393 ± 0.0643) (cyclin B1—sh#1: 0.370 ± 0.145; sh#2: 0.400 ± 0.0265) ( n = 3) (D). (E, F) CDC25c was quantified by western blot in THP1 (CDC25c—sh#1: 0.350 ± 0.201; sh#2: 0.353 ± 0.216) ( n = 3) (E) and in OCI‐AML3 (CDC25c—sh#1: 0.400 ± 0.0854; sh#2: 0.450 ± 0.0800) relative to GAPDH ( n = 3) (F). All numeric values are presented as the mean values ± standard error of the mean. P ‐values are calculated using one‐way ANOVA, *** P < 0.001; ** P < 0.01; * P < 0.05.

Article Snippet: For differentiation into monocyte‐like cells, THP1 and OCI‐AML3 cells were treated with 1 μ m 1,25(OH) 2 D 3 (Calbiochem, Millipore, Burlington, MA, USA) for 72 h. To suppress the function of p53 protein, α‐pifithrin (PFT‐α) (TargetMol, Wellesley Hills, MA, USA) was used at 10 μ m in the medium.

Techniques: Knockdown, Flow Cytometry, Western Blot

Journal: Molecular Oncology

Article Title: STIM2 is involved in the regulation of apoptosis and the cell cycle in normal and malignant monocytic cells

doi: 10.1002/1878-0261.13584

Figure Lengend Snippet: Apoptosis and cell blockage induced by STIM2 KD are related to DNA double‐strand breaks and p53 induction. (A, B) Expression of p53 and p21 in THP1 leukemic line with STIM2 KD was performed by western blot (p53—sh#1: 2.35 ± 0.560; sh#2: 2.75 ± 0.542) (p21—sh#1: 1.64 ± 0.214; sh#2: 2.12 ± 0.639) ( n = 3) (A) and in OCIA‐AML3 (p53—sh#1: 2.69 ± 0.745; sh#2: 2.79 ± 0.735) (p21—sh#1: 3.52 ± 0.671; sh#2: 3.27 ± 0.439) relative to GAPDH ( n = 3) (B), P ‐values are calculated using one‐way ANOVA. (C) Cell viability was performed by Annexin/DAPI (Ann/DAPI) labeling in THP1 cells after STIM2 knockdown (KD), treated or not with pifithrin alpha (PFT‐α) (shSCRAMBLE −PFT‐α: 91.43 ± 1.582 – shSCRAMBLE +PFT‐α: 89.8 ± 1.9; sh#1 −PFT‐α: 47.27 ± 8.223 – sh#1 +PFT‐α: 65.67 ± 4.839; sh#2 −PFT‐α: 42.17 ± 6.772 – sh#2 +PFT‐α: 67.57 ± 9.097) ( n = 3), P ‐values are calculated using two‐way ANOVA. (D) Cell cycle analysis in THP1 cells after STIM2 KD, treated or not with PFT‐α was performed by flow cytometry (FCM) and compared with cells transduced with shSCRAMBLE (shSCRAMBLE −PFT‐α: 16.8 ± 3.64 – shSCRAMBLE +PFT‐α: 19.8 ± 1.69; sh#1 −PFT‐α: 47.2 ± 5.15 – sh#1 +PFT‐α: 34.7 ± 2.89; sh#2 −PFT‐α: 47.1 ± 2.95 – sh#2 +PFT‐α: 35.6 ± 3.69) ( n = 5); P ‐values are calculated using two‐way ANOVA. (E) Quantification of p53 and CDC25c at protein level was performed by western blot in THP1 cells after STIM2 KD treated or not with PFT‐α (p53—sh#1 −PFT‐α: 2.84 ± 0.9958 – sh#1 +PFT‐α: 1.237 ± 0.05508; sh#2 −PFT‐α: 2.737 ± 1.051 – sh#2 +PFT‐α: 1.297 ± 0.2196) (CDC25c—sh#1 −PFT‐α: 0.5033 ± 0.08083 – sh#1 +PFT‐α: 1.013 ± 0.1692; sh#2 ‐PFT‐α: 0.4833 ± 0.1850 – sh#2 +PFT‐α: 0.9533 ± 0.1069) ( n = 3), P ‐values are calculated using two‐way ANOVA. (F, G) p‐H2AXγ quantification was performed in THP1 by western blot (sh#1: 5.63 ± 1.38; sh#2: 3.45 ± 0.546) ( n = 3) (F) and in OCI‐AML3 (sh#1: 2.8 ± 0.725; sh#2: 3.08 ± 1.04) relative to GAPDH ( n = 3) (G), P ‐values are calculated using one‐way ANOVA. (H) In THP‐1 cell line treated or not with PFT‐α, p‐H2AXγ quantification was performed by western blot (sh#1 −PFT‐α: 4.13 ± 0.176 – sh#1 +PFT‐α: 3.8 ± 1.55; sh#2 −PFT‐α: 4.14 ± 1.14 – sh#2 +PFT‐α: 3.6 ± 1.31) relative to GAPDH ( n = 3), P ‐values are calculated using two‐way ANOVA. All numeric values are presented as the mean values ± standard error of the mean. *** P < 0.001; ** P < 0.01; * P < 0.05.

Article Snippet: For differentiation into monocyte‐like cells, THP1 and OCI‐AML3 cells were treated with 1 μ m 1,25(OH) 2 D 3 (Calbiochem, Millipore, Burlington, MA, USA) for 72 h. To suppress the function of p53 protein, α‐pifithrin (PFT‐α) (TargetMol, Wellesley Hills, MA, USA) was used at 10 μ m in the medium.

Techniques: Expressing, Western Blot, Labeling, Knockdown, Cell Cycle Assay, Flow Cytometry, Transduction

Journal: Advances in Ophthalmology Practice and Research

Article Title: RUNX1 promotes pathological retinal angiogenesis through von Willebrand factor

doi: 10.1016/j.aopr.2025.11.006

Figure Lengend Snippet: RUNX1 inhibition decreases pathological angiogenesis in OIR mouse retinas. (A) Timeline of the OIR model with or without Ro5-3335 intravitreal injection. (B) Representative images of the retinal flat-mounts stained with IB4 from OIR + Vehicle group, OIR + Ro5-3335, and OIR + Ranibizumab group after OIR induction at P17. Neovascular and avascular areas are indicated by the white dots and white area, respectively. Scale bar = 200 μm. (C) Neovascular area and avascular area were quantified. ∗ P < 0.05; ∗∗ P < 0.01. n.s., no significant, n = 4.

Article Snippet: The membrane was blocked in Tris-buffered saline containing 5% skim milk and 0.1% Tween-20 and probed overnight at 4 °C with primary antibodies against RUNX1 and GAPDH antibody (Cat#19555-1-AP, Cat#60004-1-Ig, 1:500, Proteintech, Wuhan, China).

Techniques: Inhibition, Injection, Staining

Journal: Advances in Ophthalmology Practice and Research

Article Title: RUNX1 promotes pathological retinal angiogenesis through von Willebrand factor

doi: 10.1016/j.aopr.2025.11.006

Figure Lengend Snippet: Functional analysis of the RUNX1 inhibition in the mouse model of OIR. (A) Venn diagram identifying the numbers of differentially expressed proteins among four comparisons: (OIR-PBS v.s. RA Up = 295; OIR-PBS v.s. RA Down = 170; OIR-Ro5-3335 v.s. OIR-PBS Up = 117; OIR-Ro5-3335 v.s. OIR-PBS Down = 115). (B) Clustered heatmap displaying differential protein expression profiles across RA, OIR + Vehicle, and OIR + Ro5-3335 group. (C) In the top 10 of enrichment analysis of the molecular function. (D) In the top 10 of enrichment analysis of the biological processes. (E) In the top 10 of enrichment analysis of the cellular components. (F) In the top 10 pathways of the KEGG enrichment analysis. (G) GSEA enrichment plot of the ECM-receptor interaction pathway. (H) Relative abundance of Vwf, Itga1 and Lama2 in RA, OIR + Vehicle, and OIR + Ro5-3335 group. (I) Vwf protein levels in RA, OIR + Vehicle, and OIR + Ro5-3335 group. ∗ P < 0.05; ∗∗ P < 0.01; ∗∗∗ P < 0.001. n = 3 in each group.

Article Snippet: The membrane was blocked in Tris-buffered saline containing 5% skim milk and 0.1% Tween-20 and probed overnight at 4 °C with primary antibodies against RUNX1 and GAPDH antibody (Cat#19555-1-AP, Cat#60004-1-Ig, 1:500, Proteintech, Wuhan, China).

Techniques: Functional Assay, Inhibition, Expressing

Journal: Advances in Ophthalmology Practice and Research

Article Title: RUNX1 promotes pathological retinal angiogenesis through von Willebrand factor

doi: 10.1016/j.aopr.2025.11.006

Figure Lengend Snippet: Impact of vWF knockdown on the cell motility effects in RUNX1-OE HUVECs. (A) A schematic presentation of human RUNX1 vector. The protein levels of RUNX1 in HUVECs after RUNX1-OE transfection. (B, C) Wound-healing assays were performed to assess HUVECs migration. Scale bar = 500 μm. (D, E) Tube formation assays were performed to assess HUVEC differentiation into tube-like structures. Scale bar = 200 μm ∗ P < 0.05; ∗∗ P < 0.01. n = 3 in each group.

Article Snippet: The membrane was blocked in Tris-buffered saline containing 5% skim milk and 0.1% Tween-20 and probed overnight at 4 °C with primary antibodies against RUNX1 and GAPDH antibody (Cat#19555-1-AP, Cat#60004-1-Ig, 1:500, Proteintech, Wuhan, China).

Techniques: Knockdown, Plasmid Preparation, Transfection, Migration

Journal: International Journal of Clinical and Experimental Pathology

Article Title: Knockdown of eukaryotic translation initiation factor 3 subunit B inhibits cell proliferation and migration and promotes apoptosis by downregulating WNT signaling pathway in acute myeloid leukemia

doi:

Figure Lengend Snippet: EIF3B relative expression in human AML cell lines. EIF3B mRNA relative expression (A) and protein expression (B) were increased in AML-193, OCI-AML2, and KG-1 cell lines compared to human primary BMMC. The comparison of EIF3B mRNA relative expression between human AML cell lines and human primary BMMC was conducted using one-way ANOVA followed by Dunnett’s multiple comparisons test. P < 0.05 was considered significant. NS, non-significant; *P < 0.05, **P < 0.01, ***P < 0.001. EIF3B, eukaryotic translation initiation factor 3 subunit B; AML, acute myeloid leukemia; BMMC, bone marrow mononuclear cells.

Article Snippet: Cell culture Human AML cell lines AML-193, HL-60, OCI-AML2 and KG-1 were purchased from Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany), AML-193 cells were cultured in 90% Iscove’s Modified Dulbecco’s Medium (Gibco, USA) and 10% fetal bovine serum (FBS) (Gibco, USA), and HL-60, OCI-AML2 and KG-1 cells were cultured in 90% Roswell Park Memorial Institute 1640 Medium (Gibco, USA) and 10% FBS (Gibco, USA) under 95% air and 5% CO 2 at 37°C.

Techniques: Expressing, Comparison

Journal: Development (Cambridge, England)

Article Title: Termination of cell-type specification gene programs by the miR-183 cluster determines the population sizes of low-threshold mechanosensitive neurons.

doi: 10.1242/dev.165613

Figure Lengend Snippet: Fig. 1. The population sizes of TRKB+ and TRKC+ neurons are altered in the P0 miRCKO mice. (A-C) The percentage of TRKB+ neurons was increased in the miRCKO mice. Immunostaining for TRKB on DRG sections of control Wnt1-Cre mice (A) and the miRCKO mice (B) as well as quantification of the percentage of TRKB+ neurons in L3-L5 ganglions (C, n=3, 4). (D) The number of total neurons (ISL1+) in L5 DRG was not altered in the miRCKO mice compared with controls (n=3, 4; not significant). (E-G) The percent of total TRKC neurons were decreased in the miRCKO mice. Double immunostaining for TRKC and RUNX3 on DRG sections of Wnt1-Cre mice (E) and miR CKO mice (F), arrow heads in E and F point to NF3 (TRKC+RUNX3-) neurons, inset are higher magnifications of areas outlined in E and F. (G) Quantification of the percentage of total TRKC, TRKC+RUNX3+ (not significant) and TRKC+RUNX3- neurons in L3-L5 ganglions (n=4, 4). Data shown in mean±s.e.m. and analyzed by t-test and P<0.05 is statistically significant. *P<0.05, **P<0.01. Scale bar =µ.

Article Snippet: The following primary antibodies were used: mouse antibodies against ISL1 (DSHB, USA, 1:100), SHOX2 (Santa Cruz, USA, 1:400), NFH (neurofilament, heavy polypeptide) (CloneN52; Sigma-aldrich, St. Louis, MO, USA, 1:500); rabbit antibodies against RUNX3 (gifts from Thomas Jessell, Columbia University Medical Center, 1:300), ISL1 (gifts from Thomas Jessell, Columbia University Medical Center, 1:250), NFH (Millipore, USA, 1:200), TRKA (Millipore, USA, 1:500), TRKC (Cell signaling, USA, 1:500), NECAB2 (Proteintech Europe, 1:1000), CALB1 (Millipore, USA, 1:500) and chicken-TRKB (kind gift from Louis F. Reichardt, 1:2000); goat antibodies against TRKA (R&D systems, Minneapolis, MN, USA, 1:500), TRKB (R&D systems, Minneapolis, MN, USA, 1:500), TRKC (R&D, 1:500), Ret (R&D, 1:100) and GFP (Abcam, USA, 1:500); Guinea pig antibodies against TLX3 (kind gift from Carmen Birchmeier).

Techniques: Immunostaining, Control, Double Immunostaining

Journal: International journal of molecular medicine

Article Title: Brain death induces the alteration of liver protein expression profiles in rabbits.

doi: 10.3892/ijmm.2014.1806

Figure Lengend Snippet: Figure 5. Re‑identification of altenation of runt‑related transcription factor 1 (RUNX1) proteins by immunohistochemistry and western blot analysis. (A) Representative images (x100) of immunohistochemical analysis of (a‑d) control samples and (e‑h) paired brain death groups. The protein expression of RUNX1 stained in the nucleus in each brain death group is clearly decreased as compared to the control group at 2, 4, 6 and 8 h after brain death. (B) Representative results of western blot analysis of paired brain death groups and control samples with β‑actin as an internal control. A significant decrease in a time‑dependent manner and compared with the control groups was observed for the brain death groups. *P<0.05 indicates statistical significance compared with the previous groups. △P<0.05 indicates statistical significance compared with the control groups.

Article Snippet: The membrane was then blocked with 5% non-fat dry milk overnight and probed with primary rabbit polyclonal antibody against RUNX1 (1:500, Boster Biological Engineering, Co., Ltd.).

Techniques: Immunohistochemistry, Western Blot, Immunohistochemical staining, Control, Expressing, Staining

Journal: International journal of molecular medicine

Article Title: Brain death induces the alteration of liver protein expression profiles in rabbits.

doi: 10.3892/ijmm.2014.1806

Figure Lengend Snippet: Figure 4. Identification of alteration of runt‑related transcription factor 1 (RUNX1) protein expression by MALDI‑TOF/TOF tandem mass spectrometry. Acquired spectra were searched against a Mascot search engine based on the Swiss‑Prot protein database. MS/MS spectrum of the precursor ion with m/z 2425.2 for peptide 335ILPPCTNASTGAALLNPSLPSQSDVVETEGSHSNSPTNMPPARLEEAVWRPY386 with corresponding peak values was identi fied as RUNX1. Inset is the Swiss‑Prot protein score from the Swiss‑Prot database.

Article Snippet: The membrane was then blocked with 5% non-fat dry milk overnight and probed with primary rabbit polyclonal antibody against RUNX1 (1:500, Boster Biological Engineering, Co., Ltd.).

Techniques: Expressing, Mass Spectrometry, Tandem Mass Spectroscopy