Journal: Frontiers in Immunology

Article Title: Bridging aging, immunity, and atherosclerosis: novel insights into senescence-related genes

doi: 10.3389/fimmu.2025.1557266

Figure Lengend Snippet: Construction of diagnostic nomogram and evaluation of diagnostic performance. (A) A diagnostic nomogram based on three characteristic genes was constructed. Each gene corresponded to a score and the total score of the three genes was used to predict the risk of AS ( n =32 biologically replicated experiments.). (B) The calibration curves were constructed to evaluate the accuracy of the nomogram ( n =32 biologically replicated experiments.). (C) DCA was performed to assess the net benefit of AS diagnostic decisions predicted by the nomograms ( n =32 biologically replicated experiments.). (D, E) The expression comparisons of the three hub genes (PDLIM1, PARP14 and SEL1L3) in the GSE43292 ( n =32 biologically replicated experiments.) and GSE100927 (AS groups, n =29 biologically replicated experiments; control groups, n =12 biologically replicated experiments.) training datasets. (G, F) ROC curves showing the performance of the three genes in the GSE43292 ( n =32 biologically replicated experiments.) and GSE100927 (AS groups, n =29 biologically replicated experiments; control groups, n =12 biologically replicated experiments.) datasets. (H-J) The expression comparisons of the three hub genes (PDLIM1, PARP14 and SEL1L3) in the GSE28829 (advanced groups, n =16 biologically replicated experiments; early groups, n =13 biologically replicated experiments.), GSE120521 ( n =4 biologically replicated experiments.) and GSE41571 (unstable groups, n =5 biologically replicated experiments; stable groups, n =6 biologically replicated experiments.) validation datasets. (K-M) . ROC curves showing the performance of the three genes in the GSE28829 (advanced groups, n =16 biologically replicated experiments; early groups, n =13 biologically replicated experiments.), GSE120521 ( n =4 biologically replicated experiments.) and GSE41571 (unstable groups, n =5 biologically replicated experiments; stable groups, n =6 biologically replicated experiments.) validation datasets. Statistics performed by Independent sample T-test. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: For immunofluorescence staining, the samples were incubated overnight at 4°C with the following antibodies: PDLIM1 polyclonal antibody (1:1000; Cat# 11674-1-AP; Proteintech), PARP14 polyclonal antibody (1:7000; Cat# bs-19886R; Bioss), SEL1L3 polyclonal antibody (1:5000; Cat# bs-19626R; Bioss), F4/80 FITC-labeled polyclonal secondary antibody (1:2000; Cat# GB113373 ; Servicebio), CD80 polyclonal antibody (1:1000; Cat# GB114055 ; Servicebio), CD163 monoclonal antibody (1:3000; Cat# GB15340; Servicebio), CD36 polyclonal antibody (1:5000; Cat# GB112562 ; Servicebio).

Techniques: Diagnostic Assay, Construct, Expressing, Control, Biomarker Discovery

Journal: Frontiers in Immunology

Article Title: Bridging aging, immunity, and atherosclerosis: novel insights into senescence-related genes

doi: 10.3389/fimmu.2025.1557266

Figure Lengend Snippet: Characterized genes of single gene GSEA. (A, D, E) KEGG analysis of genes in the PDLIM1 low-expression group, PARP14 and SEL1L3 high-expression group using GSEA (top 5). (B, C, F) KEGG analysis of genes in the PDLIM1 high-expression group, PARP14 and SEL1L3 low-expression group using GSEA (top 5).

Article Snippet: For immunofluorescence staining, the samples were incubated overnight at 4°C with the following antibodies: PDLIM1 polyclonal antibody (1:1000; Cat# 11674-1-AP; Proteintech), PARP14 polyclonal antibody (1:7000; Cat# bs-19886R; Bioss), SEL1L3 polyclonal antibody (1:5000; Cat# bs-19626R; Bioss), F4/80 FITC-labeled polyclonal secondary antibody (1:2000; Cat# GB113373 ; Servicebio), CD80 polyclonal antibody (1:1000; Cat# GB114055 ; Servicebio), CD163 monoclonal antibody (1:3000; Cat# GB15340; Servicebio), CD36 polyclonal antibody (1:5000; Cat# GB112562 ; Servicebio).

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Bridging aging, immunity, and atherosclerosis: novel insights into senescence-related genes

doi: 10.3389/fimmu.2025.1557266

Figure Lengend Snippet: Single-cell sequencing reveals complex cellular features of AS lesions. (A) UMAP plot showing cell distribution from carotid and AS plaque samples. (B) Umap plot showing cellular composition in the AS microenvironment, colored according to cell types. (C) Proportion of major cell types between Control and AS groups. (D) Violin plot showing typical genealogical marker genes used to identify different cell types. (E-G) Uamp plot (left) and violin plot (right) showing the distribution and expression of PDLIM1, PARP14 and SEL1L3 in different cell clusters of AS.

Article Snippet: For immunofluorescence staining, the samples were incubated overnight at 4°C with the following antibodies: PDLIM1 polyclonal antibody (1:1000; Cat# 11674-1-AP; Proteintech), PARP14 polyclonal antibody (1:7000; Cat# bs-19886R; Bioss), SEL1L3 polyclonal antibody (1:5000; Cat# bs-19626R; Bioss), F4/80 FITC-labeled polyclonal secondary antibody (1:2000; Cat# GB113373 ; Servicebio), CD80 polyclonal antibody (1:1000; Cat# GB114055 ; Servicebio), CD163 monoclonal antibody (1:3000; Cat# GB15340; Servicebio), CD36 polyclonal antibody (1:5000; Cat# GB112562 ; Servicebio).

Techniques: Sequencing, Control, Marker, Expressing

Journal: Frontiers in Immunology

Article Title: Bridging aging, immunity, and atherosclerosis: novel insights into senescence-related genes

doi: 10.3389/fimmu.2025.1557266

Figure Lengend Snippet: Expression of biomarkers in mice AS tissues. (A) Representative images of aortic root sections stained with HE and quantitative analysis of plaque area. Representative images and protein levels of immunofluorescence staining for (B, C) PARP14, (D, E) PDLIM1, (F, G) SEL1L3, (B, D, F, H) F4/80, (I, J) CD80, (I, K) CD163, (I, L) CD36 in mice aortic root, respectively. Scale bar, 50 μm. Statistics performed by independent sample T-test. * P < 0.05, ** P < 0.01, *** P < 0.001, n =4 biologically replicated experiments, n =3 technically repeated experiments.

Article Snippet: For immunofluorescence staining, the samples were incubated overnight at 4°C with the following antibodies: PDLIM1 polyclonal antibody (1:1000; Cat# 11674-1-AP; Proteintech), PARP14 polyclonal antibody (1:7000; Cat# bs-19886R; Bioss), SEL1L3 polyclonal antibody (1:5000; Cat# bs-19626R; Bioss), F4/80 FITC-labeled polyclonal secondary antibody (1:2000; Cat# GB113373 ; Servicebio), CD80 polyclonal antibody (1:1000; Cat# GB114055 ; Servicebio), CD163 monoclonal antibody (1:3000; Cat# GB15340; Servicebio), CD36 polyclonal antibody (1:5000; Cat# GB112562 ; Servicebio).

Techniques: Expressing, Staining, Immunofluorescence

Journal: Cell Death and Differentiation

Article Title: MicroRNA-17-92 regulates myoblast proliferation and differentiation by targeting the ENH1/Id1 signaling axis

doi: 10.1038/cdd.2016.56

Figure Lengend Snippet: miR-17, -20a and -92a downregulate ENH1 protein expression by directly targeting its 3′UTR and ENH1 promotes myogenic differentiation but inhibits cell proliferation. ( a ) ENH1 protein expression during C2C12 myoblast myogenesis. C2C12 mouse myoblasts were cultured in growth medium (GM) and then switched to differentiation medium (DM) for 1–7 days. Beta-actin served as the loading control. ( b ) ENH1 protein expression in skeletal muscle during regeneration after CTX injury. Beta-tubulin served as the loading control. M1 and M2, mouse 1 and mouse 2. ( c ) Schematic illustration of the predicted binding sites for miR-17 and -20a (2234-2257) and for miR-92a (2791-2811) in the 3′UTR of ENH1. The wild -type miRNA-binding sites (orange) were modified to complementary sequences (green) to construct the mutated ENH1 3′UTR. ( d ) ENH1 3′UTR or 3′UTR Mut constructs were co-transfected with the miR-17, -20a or -92a mimics into 293 A cells. The data were normalized to the Renilla luciferase activity. The luciferase activity in cells transfected with miR-NC was set to 1.0. The error bars depict the means±S.D. of three measurements. *** P <0.001. ( e ) Western blot analysis of ENH1 protein in the C2C12 myoblasts transfected with miR-17, -20a, -92a or NC mimics and ( f ) with miR-17, -20a, -92a or NC inhibitors. Beta-actin served as the loading control. The data are presented as the means of the samples from three different cell samples. ( g ) Western blot confirms the efficiency of si-ENH1 on ENH1 protein expression in myoblasts. The rest is as in e . ( h and i ) si-ENH1 blocks C2C12 myoblast differentiation. C2C12 myoblasts were transfected with si-ENH1 or si-NC and stimulated to differentiate. Immunostaining of MHC ( h ) and western blot ( i ) of myogenin showing that si-ENH1 resulted in smaller myotubes and a decrease of myogenin protein at DM4. Scale bar, 200 μ m. The rest is as in e . ( j and k ) ENH1 modulates cell proliferation. C2C12 myoblasts were lentivirally transfected with si-ENH1 or infected with vectors expressing ENH1 and maintained in GM. The EdU incorporation assay was performed for the analysis of cell proliferation. The silencing of ENH1 ( j ) enhanced EdU incorporation compared with the control (si-NC), whereas the overexpression of ENH1 ( k ) inhibited cell proliferation. The error bars depict the means±S.D. of three measurements. *** P <0.001

Article Snippet: The following primary antibodies were used: β -actin (1 : 10 000; Proteintech, Wuhan, Hubei, China), β -tubulin (1 : 5,000; Proteintech), E2F1 (1 : 600; Santa Cruz Biotechnology, Santa Cruz, CA, USA), ENH1 (1 : 300; Proteintech), Id1 (1 : 100; CalBioreagents, San Mateo, CA, USA), myogenin (1 : 5,000; Abcam, Cambridge, MA, USA) and PCNA (1 : 2,000; Proteintech).

Techniques: Expressing, Cell Culture, Control, Binding Assay, Modification, Construct, Transfection, Luciferase, Activity Assay, Western Blot, Immunostaining, Infection, Over Expression

Journal: Cell Death and Differentiation

Article Title: MicroRNA-17-92 regulates myoblast proliferation and differentiation by targeting the ENH1/Id1 signaling axis

doi: 10.1038/cdd.2016.56

Figure Lengend Snippet: miR-17-92 and ENH1 modulate Id1 subcellular distribution. ( a ) Western blot analysis shows that the Id1 protein is downregulated during C2C12 myoblast myogenesis. ( b ) Nuclear and cytoplasmic Id1 proteins are downregulated in C2C12 cells at DM4. Alpha-tubulin and TATA-binding protein (TBP) served as the loading controls. The data are presented as the means of the samples from three different cell samples. ( c ) Localization of exogenous Flag-Id1 in C2C12 myoblasts (cultured in GM) and in differentiating C2C12 cells at DM4, as detected by Flag-Id1 immunostaining (red). Nuclei were stained blue with DAPI. Scale bar, 50 μ m. ( d ) si-ENH1 increases the nuclear Flag-Id1 in C2C12 myoblasts. C2C12 myoblasts were co-transfected with Flag-Id1 and si-ENH1 and transferred to DM for 4 days. The rest is as in c . ( e ) Western blot analysis of the whole-cell, nuclear and cytoplasmic Id1 proteins in C2C12 myoblasts transfected with si-ENH1 or NC. The rest is as in b . ( f ) Overexpression of miR-17, -20a or -92a induces the accumulation of Flag-Id1 in the nucleus. C2C12 myoblasts were co-transfected with Flag-Id1 and miRNA mimics and transferred to DM for 4 days. The rest is as in c . ( g ) Overexpression of miR-17, -20a or -92a regulates the protein levels of ENH1 and Id1 in C2C12 cells that were transfected with miRNA mimics or NC at GM and at DM4. Beta-actin served as the loading control. The rest is as in b . ( h ) Western blot analysis of nuclear and cytoplasmic Id1 in the C2C12 myoblasts transfected with a mixture of miR-17, -20a and -92a mimics or NC. The rest is as in b . ( i ) Id1 mRNA is upregulated in C2C12 myoblasts stably expressed miRNA as indicated. The data are presented as the means±S.D. of the samples from three different cell samples. * P <0.05 and ** P <0.01

Article Snippet: The following primary antibodies were used: β -actin (1 : 10 000; Proteintech, Wuhan, Hubei, China), β -tubulin (1 : 5,000; Proteintech), E2F1 (1 : 600; Santa Cruz Biotechnology, Santa Cruz, CA, USA), ENH1 (1 : 300; Proteintech), Id1 (1 : 100; CalBioreagents, San Mateo, CA, USA), myogenin (1 : 5,000; Abcam, Cambridge, MA, USA) and PCNA (1 : 2,000; Proteintech).

Techniques: Western Blot, Binding Assay, Cell Culture, Immunostaining, Staining, Transfection, Over Expression, Control, Stable Transfection

Journal: Cell Death and Differentiation

Article Title: MicroRNA-17-92 regulates myoblast proliferation and differentiation by targeting the ENH1/Id1 signaling axis

doi: 10.1038/cdd.2016.56

Figure Lengend Snippet: Ad-miR-20a delays skeletal muscle regeneration after the CTX injury. C57BL/6 mice were injected in their tibialis anterior muscles with CTX. Adenoviruses expressing miR-20a (Ad-miR-20a) or control (Ad-miR-NC) were injected into the tibialis anterior muscles 10 days prior to the CTX injection. The tibialis anterior muscles were collected on 0, 1, 3, 5 and 10 days post-CTX injury. ( a ) Quantitative PCR shows that the injection of Ad-miR-20a increases miR-20a in the mouse tibialis anterior muscles 10 days after the adenoviruses injection. qRT-PCR of miR-20a normalized to SNORNA234 is shown relative to control adenovirus-injected samples. The error bars depict the means±S.D. of the samples from six mice. * P <0.05. ( b , left) Injection of Ad-miR-20a in the tibialis anterior muscles downregulates the ENH1 protein level as detected by western blot on post-CTX injury day 0 and 5 (top panel). M1, mouse 1; M2, mouse 2; M3, mouse 3. Beta-tubulin served as the loading control. (Right) The ENH1 proteins were quantified, and the data were expressed relative to the control adenovirus-injected (Ad-miR-NC) samples. The data are presented as the means±S.D. of the samples from five mice. ** P <0.01 and *** P <0.001. ( c ) H&E images of the tibialis anterior muscle cross-sections of mice injected with Ad-miR-NC or Ad-miR-20a on 5 and 10 days post-CTX injury. Arrows indicate the regenerating myofibers. Scale bar, 200 μ m. ( d , left) Immunostaining of Ki67 in the tibialis anterior muscle on day 3 after the CTX injury. Scale bar, 200 μ m. (Right) Quantification of Ki67-positive cells from 10 random fields per sample. The error bars depict the means±S.D. of the samples from six mice. * P <0.05. ( e , left) Western blot for PCNA in the tibialis anterior muscles on day 1 after the CTX injury. (Right) Quantification of PCNA protein levels. Ad-miR-20a increased the protein levels of PCNA in the tibialis anterior muscle. The results were normalized to beta-tubulin and expressed relative to the control. The error bars depict the means±S.D. of samples from five mice. * P <0.05. ( f ) Ad-miR-20a downregulated the protein expression of myogenin in the tibialis anterior muscles on day 3 post-CTX injury. Western blot (left) and the quantification (right) of myogenin in the tibialis anterior muscles. The rest is as in e . ( g ) Immunostaining of MHC in regenerating fibers on day 5 post-CTX injury. Scale bar, 200 μ m. ( h ) Analysis of the cross-sectional areas of regenerating fibers on day 5 post injury using the ImageJ software. Only myofibers containing centralized nuclei were measured. The error bars depict the means±S.D. of samples from five mice. ( i ) Average area of the cross-sections of regenerating fibers on day 5 post-CTX injury. Approximately eight random fields were captured, and >1500 fibers were measured per sample. The error bars depict the means±S.D. of samples from five mice. ** P <0.01

Article Snippet: The following primary antibodies were used: β -actin (1 : 10 000; Proteintech, Wuhan, Hubei, China), β -tubulin (1 : 5,000; Proteintech), E2F1 (1 : 600; Santa Cruz Biotechnology, Santa Cruz, CA, USA), ENH1 (1 : 300; Proteintech), Id1 (1 : 100; CalBioreagents, San Mateo, CA, USA), myogenin (1 : 5,000; Abcam, Cambridge, MA, USA) and PCNA (1 : 2,000; Proteintech).

Techniques: Injection, Muscles, Expressing, Control, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Immunostaining, Software

Journal: Cell Death and Differentiation

Article Title: MicroRNA-17-92 regulates myoblast proliferation and differentiation by targeting the ENH1/Id1 signaling axis

doi: 10.1038/cdd.2016.56

Figure Lengend Snippet: A model of the E2F1/miR-17-92/ENH1/Id1 regulatory axis in myogenesis. In the proliferating myoblasts, E2F1 upregulates the transcription of miR-17-92. miR-17, -20a and -92a, sequester Id1 in the nucleus by targeting its cytoplasmic sequestration factor, ENH1. Id1 binds to myogenic regulatory factor (MRF) and E protein, which prevents the heterodimerization of MRF and E protein and subsequently inhibits myogenic differentiation. Upon the stimulation of differentiation, the low expression of miR-17-92 results in a high level of ENH1, which reduces the abundance of Id1 protein in the nucleus, possibly by sequestrating the Id1 into the cytoplasm. The complex of MRF and E protein binds to the promoters of target muscle genes, thereby driving their expressions and inducing myogenic differentiation

Article Snippet: The following primary antibodies were used: β -actin (1 : 10 000; Proteintech, Wuhan, Hubei, China), β -tubulin (1 : 5,000; Proteintech), E2F1 (1 : 600; Santa Cruz Biotechnology, Santa Cruz, CA, USA), ENH1 (1 : 300; Proteintech), Id1 (1 : 100; CalBioreagents, San Mateo, CA, USA), myogenin (1 : 5,000; Abcam, Cambridge, MA, USA) and PCNA (1 : 2,000; Proteintech).

Techniques: Expressing

Journal: Cancer biology & medicine

Article Title: MDM1 overexpression promotes p53 expression and cell apoptosis to enhance therapeutic sensitivity to chemoradiotherapy in patients with colorectal cancer.

doi: 10.20892/j.issn.2095-3941.2024.0540

Figure Lengend Snippet: Figure 7 Working model: MDM1 regulates therapeutic sensitivity to chemoradiotherapy in colorectal cancer. The sensitivity of CRC cells to chemoradiotherapy increased after MDM1-OE and decreased after MDM1-KO. MDM1-OE eliminate binding of YBX1 and TP53 promoter, upregulate TP53 transcription, and make CRC cells more sensitive to chemoradiation. MDM1-KO downregulate TP53 expression and cell apoptosis, leading to treatment resistance. Combination of inhibitors inducing apoptosis and chemoradiotherapy restored the sensitivity of MDM1-KO cells to cancer therapy because CRC cells have low MDM1 expression.

Article Snippet: Antibodies for co-immunoprecipitation of rabbit immunoglobulin G (30000- 0-AP), MDM1 (17575-1-AP), and YBX1 (20039-1-AP) were purchased from Proteintech.

Techniques: Binding Assay, Expressing