Journal: bioRxiv

Article Title: Senescence-induced reparative fibroblasts enable scarless wound healing in aged murine skin

doi: 10.1101/2025.04.17.648896

Figure Lengend Snippet: (A) Schematic diagram of study design, multi-omics analysis, and validation cohort. (B) Hematoxylin-eosin (H&E) and Masson staining for scar tissues, displaying morphological differences between the young and aged groups. Scale bar, 200 um. (C) Immunofluorescence staining for Sox9 (marker of the bulge) and Ki67 (marker of de novo HF) in the scar tissues of the aged group. Scale bar, 50um. (D) Quantification analysis for numbers of de novo HFs (n=5 per group). Significances were analyzed using bidirectional t-tests. (E-F) Diagram of ECM ultrastructure analysis(E). TSNE plots of unsupervised cluster analysis of ECM characteristics in each group with representative Sirius red staining images. Scale bar, 50um. (F-H) Bulk RNA sequencing for scar tissues at 28 dpw(n=3 per group). Gene ontology enrichment analysis showing the top biological process terms based on differentially expressed genes (p<0.05, |log2FC|>1)(G). Gene set enrichment analysis (GSEA) showing regeneration-associated terms (left) and heatmap of corresponding genes (right)(H). Abbreviation: dpw, day post wound. HF, hair follicle. Epi, epidermis. ECM, extracellular matrix. TSNE, T-Distributed Stochastic Neighbor Embedding. EGFR, epidermal growth factor receptor. Statistical thresholds followed p = 0.05 convention (*p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001), with nonsignificant (ns) determinations requiring p > 0.05.

Article Snippet: Ki-67( ) and SOX9( ) were utilized for observation of hair follicle, while CRABP1(12588-1-AP, proteintech), a-SMA(67735-1-Ig, proteintech), PRSS35(GTX123037, GeneTex), PTN(HA722055, HUABIO), and EREG(ER65835, HUABIO) were employed to assess the fibroblast subcluster.

Techniques: Biomarker Discovery, Staining, Immunofluorescence, Marker, RNA Sequencing

Journal: bioRxiv

Article Title: Senescence-induced reparative fibroblasts enable scarless wound healing in aged murine skin

doi: 10.1101/2025.04.17.648896

Figure Lengend Snippet: (A) Schematic diagram showing treatment design in vivo and vitro assays. (B)Hematoxylin-eosin (H&E) and Masson staining for scar tissues of control, PTN, and EREG treatment samples. Scale bar, 200 um. (C-D) Immunofluorescence staining for Sox9 (yellow) and Ki67 (purple) in the scar tissues of the EREG treatment group. Dotted lines signed morphology of de novo HF. Scale bar, 50um (C). Quantification analysis for numbers of de novo HFs (n=5 per group). Significances were utilized using bidirectional t-tests (D). (E) TSNE plots of unsupervised cluster analysis of ECM characteristics in each group with representative Sirius red staining images. Scale bar, 100um. (F) Immunofluorescence staining for CRABP1 (green, marker of papilla fibroblast), and PRSS35 (red, marker of reparative fibroblast) in the scar tissues. Scale bar, 50um(left). Quantification analysis for the ratio of marker protein area/DAPI area (n=5 per group). Significances were utilized using bidirectional t-tests(middle). Colocation plot showing colocalization of CRABP1/PRSS35 assessed by Pearson correlation coefficient(right). (G) Immunofluorescence staining for F-actin (red, cytoskeleton) and a-SMA (green, marker of fibroblast activation) in murine fibroblast lineage (L929 cells). Scale bar as shown in images. The difference in mean fluorescence intensity was utilized by bidirectional t-tests (n=5). (H-I) TSNE plots of unsupervised cluster analysis of ECM characteristics in the EREG group with representative Sirius red staining images. Scale bar, 100um(H). Quantification analysis for the ratio of collagen deposition area in scar tissues assessed by bidirectional t-tests (n=5). Abbreviation: HF, hair follicle. Epi, epidermis. EREG_R, EREG_regeneration area. EREG_S, EREG_scar area.a-SMA, a-smooth muscle actin. P value (*p < 0.05; **p < 0.01; ***p < 0.001), with nonsignificant (ns) determinations requiring p > 0.05.

Article Snippet: Ki-67( ) and SOX9( ) were utilized for observation of hair follicle, while CRABP1(12588-1-AP, proteintech), a-SMA(67735-1-Ig, proteintech), PRSS35(GTX123037, GeneTex), PTN(HA722055, HUABIO), and EREG(ER65835, HUABIO) were employed to assess the fibroblast subcluster.

Techniques: In Vivo, Staining, Control, Immunofluorescence, Marker, Activation Assay, Fluorescence

Journal: Frontiers in oncology

Article Title: DNA Polymerase Iota Promotes Esophageal Squamous Cell Carcinoma Proliferation Through Erk-OGT-Induced G6PD Overactivation.

doi: 10.3389/fonc.2021.706337

Figure Lengend Snippet: FIGURE 3 | Pol i activates G6PD through OGT-promoted O-GlcNAcylation. (A) The relative mRNA level of POLI and OGT in Pol i differentially expressed TE-1 and KYSE-150 cells. (B) the protein level of Pol i, OGT and G6PD. (C) O-GlcNAcylation of G6PD was detected after G6PD immunoprecipitation in ESCC cells. (D) the promoter of OGT (-2000 to +500 bp) was cloned into pGL-4 vector. The pGL4-OGT and internal control reporter vector pRL-TK were co-transfected into TE-1 and KYSE-150 cells. The relative OGT promoter activity was detected by dual-luciferase reporter assay. (E) Immunohistochemical staining of Pol i, OGT and protein O- GlcNAcylation in paraffin-embedded ESCC tissues. Scale bar = 100 mm. (F) level of OGT and protein O-GlcNAcylation in ESCC and adjacent tissues based on IHC score. (G) The correlation between Pol i and OGT, and the correlation between OGT and protein O-GlcNAcylation were evaluated based on IHC score in 114 tumor tissue samples. (H) Survival analysis based on the IHC score of protein O-GlcNAcylation in 114 ESCC samples. Kaplan–Meier survival analysis was applied. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: After blocking with 5% non-fat milk, the membranes were incubated with primary antibodies against bactin (Beyotime Biotechnology), Pol i (Proteintech, Rosemont, IL, USA), G6PD (Abcam, Cambridge, MA, USA), OGT (Proteintech), O-GlcNAc (Invitrogen Life Technologies, Carlsbad, CA, USA), Erk and p-Erk (Cell Signaling Technology, Danvers, Massachusetts, USA) at 4°C overnight.

Techniques: Immunoprecipitation, Clone Assay, Plasmid Preparation, Control, Transfection, Activity Assay, Luciferase, Reporter Assay, Immunohistochemical staining, Staining

Journal: Frontiers in oncology

Article Title: DNA Polymerase Iota Promotes Esophageal Squamous Cell Carcinoma Proliferation Through Erk-OGT-Induced G6PD Overactivation.

doi: 10.3389/fonc.2021.706337

Figure Lengend Snippet: FIGURE 6 | Pol i promotes ESCC cell proliferation through G6PD activation in vivo. (A) the tumor volume of xenograft nude mice with different Pol i expression. Mice were divided into three groups when the average tumor size grew up to 300 mm3. 5 mg/kg polydatin was intraperitoneally injected every other day. Same volume of normal saline was used as control. (B) relative G6PD activity in tumor tissue. (C) hematoxylin and eosin (HE) staining and immunohistochemical staining of Pol i, OGT and protein O-GlcNAcylation in tumors. **P < 0.01, ***P < 0.001. Scale bar = 100 mm.

Article Snippet: After blocking with 5% non-fat milk, the membranes were incubated with primary antibodies against bactin (Beyotime Biotechnology), Pol i (Proteintech, Rosemont, IL, USA), G6PD (Abcam, Cambridge, MA, USA), OGT (Proteintech), O-GlcNAc (Invitrogen Life Technologies, Carlsbad, CA, USA), Erk and p-Erk (Cell Signaling Technology, Danvers, Massachusetts, USA) at 4°C overnight.

Techniques: Activation Assay, In Vivo, Expressing, Injection, Saline, Control, Activity Assay, Staining, Immunohistochemical staining

Journal: Cancer biology & therapy

Article Title: circ_C20orf11 enhances DDP resistance by inhibiting miR-527/YWHAZ through the promotion of extracellular vesicle-mediated macrophage M2 polarization in ovarian cancer.

doi: 10.1080/15384047.2021.1959792

Figure Lengend Snippet: Figure 5. EVs mediate M2-type macrophage polarization in ovarian cancer. (a) EVs were isolated from the supernatant of SKOV3/DDP cells and characterized via electron microscopy and (b) nanoparticle tracking. (C) EV-specific markers (CD9, TSG101 and CD63) were measured via western blotting. (d) EVs were labeled with PKH67 dye and cultured with THP-1 cells. Representative fluorescence images are shown. (e) After SKOV3/DDP and A2780/DDP cells were transfected with adenoviral circ_C20orf11 (si- SKOV3/DDP-Exo+si-C20orf11) or its vector control (si-SKOV3/DDP-Exo+si-NC), the cells were cultured with EVs from SKOV3/DDP cells. IL-10 expression was detected using an ELISA. Untreated THP-1 cells served as the control. (f) An ELISA was performed to detect IL-6 expression. (g) Relative mRNA expression of TNF-α, IL-6 and iNOS. (H) mRNA expression of CD206, IL-10 and Arg-1 in relation to GAPDH and U6 expression. n = 3. *P < .05, ** P < .01, *** P < .001.

Article Snippet: After centrifugation (1,000 rpm for 8 min at 4°C), the isolated cells were stained using a CD206 antibody (cat. no. 18704-1-AP, Proteintech) according to the manufacturer’s instructions.

Techniques: Isolation, Electron Microscopy, Western Blot, Labeling, Cell Culture, Fluorescence, Transfection, Plasmid Preparation, Control, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Cancer biology & therapy

Article Title: circ_C20orf11 enhances DDP resistance by inhibiting miR-527/YWHAZ through the promotion of extracellular vesicle-mediated macrophage M2 polarization in ovarian cancer.

doi: 10.1080/15384047.2021.1959792

Figure Lengend Snippet: Figure 6. Silencing of circ_C20orf11 enhances sensitivity to DDP in vivo. Xenotransplantation studies were performed with SKOV3 cells. (a) Tumor volume was measured every 5 days for 30 days. (b) Representative images of tumor formation. (c) Tumor weight was measured. (d) qPCR analysis of C20orf11, miR-527 and YWHAZ expression. (e) qPCR results showing CD206, IL-10 and Arg-1 expression. (f) Western blotting detection of YWHAZ PD-L1 is presented. n = 3. *P < .05, ** P < .01, *** P < .001.

Article Snippet: After centrifugation (1,000 rpm for 8 min at 4°C), the isolated cells were stained using a CD206 antibody (cat. no. 18704-1-AP, Proteintech) according to the manufacturer’s instructions.

Techniques: In Vivo, Expressing, Western Blot

Journal: Cancer biology & therapy

Article Title: circ_C20orf11 enhances DDP resistance by inhibiting miR-527/YWHAZ through the promotion of extracellular vesicle-mediated macrophage M2 polarization in ovarian cancer.

doi: 10.1080/15384047.2021.1959792

Figure Lengend Snippet: Figure 7. Serum EV-circ_C20orf11 levels are upregulated in ovarian patients. Patients were considered DDP resistant if they showed no significant clinical effect or had progressive disease after receiving one cycle of DDP treatment. The remaining patients were considered DDP sensitive. Real-time qPCR analysis of (a) C20orf11 and (b) miR-527 expression in relation to GAPDH and U6 expression in DDP-sensitive ovarian cancer tissue (S), DDP-resistant ovarian cancer tissue (R), and healthy ovarian tissue (Normal). (c) Flow cytometry detection and quantification of CD206-positive cells in DDP-sensitive ovarian cancer tissue (S), DDP-resistant ovarian cancer tissue (R), and healthy ovarian tissue (Normal). (d) Real-time qPCR analysis of CD206 expression in relation to GAPDH and U6 expression in DDP-sensitive ovarian cancer tissue (S), DDP- resistant ovarian cancer tissue (R), and healthy ovarian tissue (Normal). (e) The expression level of C20orf11 in ovarian cancer tissue was negatively correlated with that of miR-527. (f) The expression level of miR-527 in ovarian cancer tissue was negatively correlated with that of CD206. (g) Representative image of serum EVs detected using an electron microscope. (h) EVs were measured using nanoparticle tracking. (i) EV markers were analyzed via western blotting. (j) The abundance of C20orf11 in serum EVs was assessed using qPCR. (k) Kaplan-Meier survival curves of patients with ovarian cancer with high and low C20orf11 expression. n = 3. *P < .05, ** P < .01.

Article Snippet: After centrifugation (1,000 rpm for 8 min at 4°C), the isolated cells were stained using a CD206 antibody (cat. no. 18704-1-AP, Proteintech) according to the manufacturer’s instructions.

Techniques: Expressing, Flow Cytometry, Microscopy, Western Blot

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Involvement of kisspeptin in androgen-induced hypothalamic endoplasmic reticulum stress and its rescuing effect in PCOS rats.

doi: 10.1016/j.bbadis.2021.166242

Figure Lengend Snippet: Fig. 4. Kisspeptin rescues DHT-induced ER stress in GT1–7 cells. (a-b) Western blots for ATF6, IRE1α, Perk, BIP p-eIF2α and eIF2α (a) and qPCR analysis of GADD34 and Xbp1s/Xbp1 (b) in GT1–7 cells 30 min after treatment with DHT (10 nM), Kp10 (a kiss1 peptide, 10 nM) or DMSO as control (ctrl). n = 3. ** P < 0.01, ***P < 0.001, t-test. (c-d) Western blots (c) or qPCR analysis (d) for ATF4, CHOP, PDI and XBP1 in GT1–7 cells 24 h after treatment with DHT (10 nM), Kp10 (10 nM), or DMSO as Ctrl. n = 3. ** P < 0.01, *** P < 0.001, t-test. (e) qPCR analysis of ATF4, BIP, PDI and CHOP in GT1–7 cells after treatment with thapsigargin (TG, an ER stress inducer, 20 nM), Kp10 (10 nM) or vehicle control (Ctrl). n = 3. *** P < 0.001, t-test. GAPDH was used as a loading control for all western blots. Error bars are smaller than symbol size except where shown.

Article Snippet: Antibodies used are: IRE1a (1:500, catalog 3294, Cell Signaling Technology [CST], USA), Perk (1:500, catalog 5683, CST), BIP (1:500, catalog 3177, CST), PDI (1:500, catalog 3501,CST), Phospho-eIF2α (1:500, catalog 9721, CST), eIF2α (1:500, catalog 9722, CST), Bax (1:500, catalog 2772, CST), ATF6 (1:500, catalog 15,794–1-AP, ProteinTech, USA), ATF4 (1:500, catalog 10,835–1-AP, ProteinTech), KISS1 (1:500, catalog H-048-46PA5-106920, Thermo Fisher, USA), β-actin (1:5000, catalog A8481, Sigma-Aldrich), Xbp1 (1:500, catalog ab37152, Abcam, USA), Phospho-IRS (1:500, catalog 3203, CST), PI3K (1:500 catalog 4249, CST), Phospho-GSK-3β (1:500,catalog 5558,CST)GSK-3β (1:500, catalog 9315, CST) and GAPDH (1:5000, catalog sc-47,724, Santa Cruz, USA).

Techniques: Western Blot, Control

Journal: Biochimica et biophysica acta. Molecular basis of disease

Article Title: Involvement of kisspeptin in androgen-induced hypothalamic endoplasmic reticulum stress and its rescuing effect in PCOS rats.

doi: 10.1016/j.bbadis.2021.166242

Figure Lengend Snippet: Fig. 5. Effect of kisspeptin on UPR in the hypo thalamus of DHT-induced PCOS rats. (a-b) qPCR analysis of PDI, ATF6, Traf2, Xbp1s/Xbp1 (a) and immunofluorescence labeling for Xbp1 (b) in the hypothalamus of DHT-induced PCOS (PCOS_DHT) rats 24 h after kp10 (100 nM,10 μl per rat) was intracerebroventricularly (i.c.v) injected at day 21 post DHT treatment. n = 3. * P < 0.05, ** P < 0.01, t-test. 3v, third ventricle. Scale bar, 20 μm. (c) qPCR analysis of BIP, PDI, Xbp1s/Xbp1 in the hypothalamus of PCOS_DHT rats 24 h after salubrinal (100 nM, 10 μl per rat) was injected (i.c.v) at day 21 post DHT treat ment. n = 4–9. * P < 0.05, t-test.

Article Snippet: Antibodies used are: IRE1a (1:500, catalog 3294, Cell Signaling Technology [CST], USA), Perk (1:500, catalog 5683, CST), BIP (1:500, catalog 3177, CST), PDI (1:500, catalog 3501,CST), Phospho-eIF2α (1:500, catalog 9721, CST), eIF2α (1:500, catalog 9722, CST), Bax (1:500, catalog 2772, CST), ATF6 (1:500, catalog 15,794–1-AP, ProteinTech, USA), ATF4 (1:500, catalog 10,835–1-AP, ProteinTech), KISS1 (1:500, catalog H-048-46PA5-106920, Thermo Fisher, USA), β-actin (1:5000, catalog A8481, Sigma-Aldrich), Xbp1 (1:500, catalog ab37152, Abcam, USA), Phospho-IRS (1:500, catalog 3203, CST), PI3K (1:500 catalog 4249, CST), Phospho-GSK-3β (1:500,catalog 5558,CST)GSK-3β (1:500, catalog 9315, CST) and GAPDH (1:5000, catalog sc-47,724, Santa Cruz, USA).

Techniques: Immunofluorescence, Labeling, Injection