Journal: Non-coding RNA Research

Article Title: CircSMAD4 shapes matrix-remodeling TAMs in lung adenocarcinoma

doi: 10.1016/j.ncrna.2026.03.003

Figure Lengend Snippet: circSMAD4 drives tumor-educated M2-like polarization of macrophages and promotes tumor-cell aggressiveness. (A) Workflow for generating TC-hMDMs and TC-BMDMs, circSMAD4 knockdown, and downstream functional assays. (B) RT–qPCR analysis of M1-associated markers (MHC-II [HLA-DRA in TC-hMDMs; H2-Ab1 in TC-BMDMs], NOS2, and CD86) and M2-associated markers (CD163, CD206, and ARG1) in TC-hMDMs and TC-BMDMs. (C) Representative flow-cytometry histograms for HLA-DR, iNOS, CD86, CD163, CD206, and ARG1 in TC-hMDMs. Gating strategy and marker thresholds were defined based on FMO controls (see ). (D) Flow-cytometry quantification of marker-positive cells in TC-hMDMs and TC-BMDMs. (E) ELISA of IL-10, TGF-β, and iNOS in culture supernatants. (F) CCK-8 assays of A549 and LLC cells. (G) Colony-formation assays of A549 and LLC cells with quantification. (H) Bioluminescence-based growth readouts of patient-derived LUAD organoids (PDO #1 and PDO #2) after co-culture with TC-hMDMs. (I) Immunoblot analysis of EMT-related proteins (E-cadherin, N-cadherin, Vimentin) in A549 and LLC cells. (J) Transwell migration and invasion assays of A549 and LLC cells with quantification. Scale bar, 50 μm. ∗P < 0.05; ∗∗P < 0.01; ∗∗∗P < 0.001; ∗∗∗∗P < 0.0001; ns, not significant.

Article Snippet: Sections were incubated with primary antibodies against Ki-67 (Servicebio, Cat# GB111499 ), E-cadherin (Proteintech, Cat# 20874-1-AP), and Vimentin (Proteintech, Cat# 10366-1-AP).

Techniques: Knockdown, Functional Assay, Quantitative RT-PCR, Flow Cytometry, Marker, Enzyme-linked Immunosorbent Assay, CCK-8 Assay, Derivative Assay, Co-Culture Assay, Western Blot, Migration

Journal: Non-coding RNA Research

Article Title: CircSMAD4 shapes matrix-remodeling TAMs in lung adenocarcinoma

doi: 10.1016/j.ncrna.2026.03.003

Figure Lengend Snippet: circSMAD4 depletion in macrophages restrains LUAD growth and metastasis in vivo. (A) Schematic of orthotopic lung implantation and experimental metastasis models using LLC cells mixed with BMDMs expressing shNC or sh-circSMAD4. (B) Representative images of orthotopic lung tumors. (C) Tumor weight of orthotopic implants. (D) Overall survival of mice bearing orthotopic tumors. (E) Immunofluorescence showing F4/80 and circSMAD4 signals in tumor tissues. Scale bar, 50 μm. (F, G) Representative Ki-67 IHC staining and quantification in orthotopic tumors. Scale bar, 50 μm. (H) Representative bioluminescence images of lung tumor burden in the metastasis model. (I) Tumor weight in the metastasis model. (J) Overall survival of mice in the metastasis model. (K–M) Representative IHC staining and quantification of E-cadherin and vimentin in tumors. Scale bar, 50 μm. ∗P < 0.05; ∗∗P < 0.01; ∗∗∗P < 0.001; ∗∗∗∗P < 0.0001; ns, not significant.

Article Snippet: Sections were incubated with primary antibodies against Ki-67 (Servicebio, Cat# GB111499 ), E-cadherin (Proteintech, Cat# 20874-1-AP), and Vimentin (Proteintech, Cat# 10366-1-AP).

Techniques: In Vivo, Expressing, Immunofluorescence, Immunohistochemistry

Journal: Genes & Diseases

Article Title: Identification of PLXNC1 as a novel biomarker for consensus molecular subtype 4 in colorectal cancer

doi: 10.1016/j.gendis.2025.101974

Figure Lengend Snippet: PLXNC1 knockdown inhibits gene signatures characteristic of CMS4 colorectal cancer. (A) qRT-PCR analysis was applied to examine the relative mRNA expression of genes related to epithelial–mesenchymal transition (EMT), complement, angiogenesis, and immunosuppression. GAPDH was a normalization control. (B) Protein expression and quantification results of SNAIL, Vimentin, and E-cadherin in colorectal cancer cells were determined by western blotting. (C) Immunofluorescence analysis of the levels of E-cadherin (the epithelial marker, red) and Vimentin (the mesenchymal marker, green) proteins in the control and PLXNC1-silenced LoVo cells. The nuclei were stained with Hoechst (blue). Scale bars = 75 μm ∗∗∗ p < 0.001, ∗∗ p < 0.01, and ∗ p < 0.05.

Article Snippet: Fixed cells were incubated with primary antibody against E-cadherin (1:50; sc-8426; Santa Cruz Biotechnology, California, USA) or vimentin (1:100; BM0135; Boster, Wuhan, China) at 4 °C overnight, followed by incubation with secondary antibodies (1:200; Boster, Wuhan, China) at room temperature for 1 h. The nuclei of cells were stained with Hoechst for 10 min.

Techniques: Knockdown, Quantitative RT-PCR, Expressing, Control, Western Blot, Immunofluorescence, Marker, Staining

Journal: Genes & Diseases

Article Title: Identification of PLXNC1 as a novel biomarker for consensus molecular subtype 4 in colorectal cancer

doi: 10.1016/j.gendis.2025.101974

Figure Lengend Snippet: PLXNC1 promotes tumor growth and metastasis in vivo . (A) Magnetic resonance imaging (MRI) assessment of wild-type (WT) mice carrying subcutaneous tumors. (B) Tumor volumes calculated by MRI. The tumor volume = area of tumor in each slice × slice thickness. (C) Representative images of tumors from WT mice after subcutaneous inoculation of SL4 cells infected with the control vector or sh-PLXNC1 vector. (D) Mice were killed 14 days following subcutaneous injection, and tumors were dissected and weighed. (E) Bioluminescence images showing tumor metastasis by tracking luciferase-expressing SL4 cells in sh-NC and sh-PLXNC1 groups. (F) Histogram showing the bioluminescent signal intensity analyzed by the IVIS System. (G) MRI inspection of hepatic tumor metastasis of colon cancer after intrasplenic injection of SL4 cells in sh-NC and sh-PLXNC1 groups. (H) Gross examination of hepatic tumor metastasis of colon cancer after intrasplenic injection of SL4 cells in sh-NC and sh-PLXNC1 groups. (I) Mice were sacrificed on day 10 after intrasplenic injection, and livers were excised and weighed. (J) Immunohistochemical staining was used to detect the expression levels of PCNA, CD31, TGF-β, vimentin, β-catenin, and E-Cadherin in tumor tissues from sh-NC and sh-PLXNC1 groups. Scale bars = 50 μm. (K) Quantification of immunohistochemical staining in subcutaneous tumor tissues from sh-NC and sh-PLXNC1 groups. (L) Immunohistochemical staining showing expression of epithelial–mesenchymal transition (EMT) markers in liver metastasis tumor tissues from sh-NC and sh-PLXNC1 groups. (M) Quantification of immunohistochemical staining in liver metastasis tumor tissues from sh-NC and sh-PLXNC1 groups. ∗∗∗ p < 0.001, ∗∗ p < 0.01, and ∗ p < 0.05; ns, not significant.

Article Snippet: Fixed cells were incubated with primary antibody against E-cadherin (1:50; sc-8426; Santa Cruz Biotechnology, California, USA) or vimentin (1:100; BM0135; Boster, Wuhan, China) at 4 °C overnight, followed by incubation with secondary antibodies (1:200; Boster, Wuhan, China) at room temperature for 1 h. The nuclei of cells were stained with Hoechst for 10 min.

Techniques: In Vivo, Magnetic Resonance Imaging, Infection, Control, Plasmid Preparation, Injection, Luciferase, Expressing, Immunohistochemical staining, Staining

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Positive and negative control cells for both marker panels. ( A ) HeLa (white arrows): in the first panel positive for cytokeratin (CK), Vimentin (Vim), VEGF and in the second panel positive for p16INK4A, negative for PD-L1 and CD45. ( B ) T98G (red arrows): positive for Vim. ( C ) CaSki (yellow arrows): positive for CK, Vim, VEGF, PD-L1 and p16INK4A. ( D ) MCF-7 (green arrows): positive for CK. All cell lines were negative for CD45. Only hematopoietic cells showed CD45 positive staining. Scale bar = 10 μm. Images were processed using Zeiss ZEN 3.7 software and the figure was assembled using Microsoft PowerPoint.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Negative Control, Marker, Staining, Software

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Representative images of patient-derived DTCs with three different profiles. ( A ) DTC positive for Vim, VEGF and p16, negative for CK, PD-L1 and CD45. ( B ) CK and Vim positive while negative for VEGF, PD-L1, p16 and CD45. ( C ) DTC positive for Vim and PD-L1, negative for all other markers. Scale bar = 10 μm. Images were processed using Zeiss ZEN 3.7 software and figure was assembled using Microsoft PowerPoint.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Derivative Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Proportion of DTC Phenotypes. The majority of detected DTCs (70%, n = 174) was positive for Vimentin without CK expression, hence revealing a mesenchymal phenotype. An epithelial phenotype was found in 13% ( n = 31) of the DTCs and 15% ( n = 37) presented epithelial–mesenchymal transition (EMT)-like properties. Chart generated using Microsoft Excel.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Expressing, Generated

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Sequential multi-parameter immunofluorescent staining. After applying the first antibody panel (CK, Vim and VEGF), the slides were scanned and DTC profiles were assessed. Subsequently, releasable fluorochrome-conjugates were digested and the slides were subjected to the second antibody panel (PD-L1, p16, CD45) followed by scanning and DTC detection. Created with BioRender.com.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Staining