Journal: Journal of Dental Sciences

Article Title: Laser-modified titanium surfaces induce sex-dimorphic secretion of angiogenic factors by gingiva-derived mesenchymal stromal cells

doi: 10.1016/j.jds.2025.09.013

Figure Lengend Snippet: Laser-modified titanium upregulates angiogenesis-related genes in human gingiva-derived mesenchymal stromal cells (GMSCs). Assay for transposase-accessible chromatin sequencing (ATAC-seq) and RNA-sequencing (RNA-seq) were performed on GMSCs cultured on machined, lasered, and SLA (sand-blasted, large-grit, acid-etched) titanium discs for 72 h (n = 2 per group; age-matched 50-year-old female and male). (A) Venn diagrams show differentially upregulated (UP) and downregulated (DOWN) genes in the lasered group and SLA group compared to the machined group, respectively. (B) Volcano plot highlights differentially expressed angiogenesis-related genes. (C) Gene Ontology (GO) enrichment analysis of genes upregulated in the lasered group versus the machined group. (D) Thirty-three genes were identified at the intersection of GO:0016477 (cell migration) and genes upregulated in both female and male GMSCs on laser-modified versus machined surfaces. The heatmap shows shared migration-related genes upregulated in both sexes. (E) Chromatin accessibility and RNA expression levels of CCN1 and EDIL3. F, female. M, male. CCN1, cellular communication network factor 1. EDIL3, EGF like repeats and discoidin domains 3.

Article Snippet: Subsequently, the medium was replaced with fresh exosome-free α-MEM and cells were cultured for an additional 48 h. Conditioned medium was collected for the tube formation assay, and GMSCs were harvested for RNA extraction. mRNA expression was validated using TaqMan Fast Advanced Master Mix (4444556, Invitrogen) with TaqMan Gene Expression Assays (Invitrogen) for CYR61 (Hs00155479_m1), EDIL3 (Hs00964112_m1), and GAPDH (Hs02786624_g1), according to the manufacturer's protocol.

Techniques: Modification, Derivative Assay, Sequencing, RNA Sequencing, Cell Culture, Migration, RNA Expression

Journal: Journal of Dental Sciences

Article Title: Laser-modified titanium surfaces induce sex-dimorphic secretion of angiogenic factors by gingiva-derived mesenchymal stromal cells

doi: 10.1016/j.jds.2025.09.013

Figure Lengend Snippet: Laser-modified titanium surfaces upregulate CCN1 expression in human gingiva-derived mesenchymal stromal cells (GMSCs). Relative gene expression levels of ADM2, CCN1, EDIL3, and β-actin in GMSCs cultured on three different titanium disc surfaces: machined, lasered, and SLA (sand-blasted, large-grit, acid-etched). (A) CCN1 mRNA levels were significantly upregulated in the laser-treated and SLA groups compared to the machined group. (B) and (C) No significant differences in ADM2 and EDIL3 expression were observed among the three groups. (D) β-actin expression was significantly increased in the laser-treated and SLA groups compared to the machined group. Data are means ± standard deviation (n = 8 per group, 4 females and 4 males). Paired t-test. ∗ P < 0.05, ∗∗∗∗ P < 0.0001. mRNA, messenger ribonucleic acid. CCN1, cellular communication network factor 1. ADM2, adrenomedullin 2. EDIL3, EGF like repeats and discoidin domains 3. RPS18, ribosomal protein S18.

Article Snippet: Subsequently, the medium was replaced with fresh exosome-free α-MEM and cells were cultured for an additional 48 h. Conditioned medium was collected for the tube formation assay, and GMSCs were harvested for RNA extraction. mRNA expression was validated using TaqMan Fast Advanced Master Mix (4444556, Invitrogen) with TaqMan Gene Expression Assays (Invitrogen) for CYR61 (Hs00155479_m1), EDIL3 (Hs00964112_m1), and GAPDH (Hs02786624_g1), according to the manufacturer's protocol.

Techniques: Modification, Expressing, Derivative Assay, Gene Expression, Cell Culture, Standard Deviation

Journal: Journal of Dental Sciences

Article Title: Laser-modified titanium surfaces induce sex-dimorphic secretion of angiogenic factors by gingiva-derived mesenchymal stromal cells

doi: 10.1016/j.jds.2025.09.013

Figure Lengend Snippet: Sex-dimorphic secretion of EDIL3 and CCN1 by human gingiva-derived mesenchymal stromal cells (GMSCs). ELISA assays were performed to quantify CCN1 and EDIL3 levels in conditioned medium collected from GMSCs cultured on machined, lasered, and SLA (sand-blasted, large-grit, acid-etched) titanium discs for 72 h. (A) ELISA results from conditioned medium without Triton X treatment. Data are mean ± standard deviation (n = 8; four females and four males). Dotted line: paired t-test. Solid line: unpaired t-test. ∗ P < 0.05. (B) ELISA results from conditioned medium treated with Triton X. Data are mean ± standard deviation (n = 8; four females and four males). Dotted line: paired t-test. Solid line: unpaired t-test. ∗ P < 0.05, ∗∗ P < 0.01. F, female. M, male. CCN1, cellular communication network factor 1. EDIL3, EGF like repeats and discoidin domains 3.

Article Snippet: Subsequently, the medium was replaced with fresh exosome-free α-MEM and cells were cultured for an additional 48 h. Conditioned medium was collected for the tube formation assay, and GMSCs were harvested for RNA extraction. mRNA expression was validated using TaqMan Fast Advanced Master Mix (4444556, Invitrogen) with TaqMan Gene Expression Assays (Invitrogen) for CYR61 (Hs00155479_m1), EDIL3 (Hs00964112_m1), and GAPDH (Hs02786624_g1), according to the manufacturer's protocol.

Techniques: Derivative Assay, Enzyme-linked Immunosorbent Assay, Cell Culture, Standard Deviation

Journal: Journal of Dental Sciences

Article Title: Laser-modified titanium surfaces induce sex-dimorphic secretion of angiogenic factors by gingiva-derived mesenchymal stromal cells

doi: 10.1016/j.jds.2025.09.013

Figure Lengend Snippet: Laser-modified titanium promotes the secretion of extracellular vesicles enriched with CCN1 and EDIL3 from human gingiva-derived mesenchymal stromal cells (GMSCs). (A) Primary GMSCs from one female and one male donor were cultured on machined, lasered, and SLA discs for 72 h, followed by ATAC-seq and RNA-seq analysis. Heatmaps show exosome-related genes. M, machined. L, lasered. S, SLA (sand-blasted, large-grit, acid-etched). (B) Primary GMSCs were cultured on machined, laser-treated, and SLA (sand-blasted, large-grit, acid-etched) titanium discs for 72 h. The conditioned medium was collected and centrifuged at 300× g for 10 min. The supernatant was then filtered through a 0.22 μm membrane, and particle size was analyzed using nanoparticle tracking analysis (NTA). Data are presented as mean values (n = 8 per group; 4 females and 4 males). (C) Western blot analysis was performed to evaluate CCN1, EDIL3 and ADM2 protein levels in cells, extracellular vehicles (EVs), and non-extracellular vesicle (non-EV) fractions derived from GMSCs (51-year-old male, 51M, and 63-year-old female, 63F) were cultured on machined (M), lasered (L), and SLA (sand-blasted, large-grit, acid-etched, S) titanium discs for 72 h. Laser-modified surfaces increased CCN1 in male GMSC EVs and EDIL3 in female EVs compared to machined surfaces. Lower panels show Ponceau S staining of the corresponding PVDF membranes. Results shown are representative of at least three independent experiments. CCN1, cellular communication network factor 1. ADM2, adrenomedullin 2. EDIL3, EGF like repeats and discoidin domains 3. TSG101, tumor susceptibility gene 101.

Article Snippet: Subsequently, the medium was replaced with fresh exosome-free α-MEM and cells were cultured for an additional 48 h. Conditioned medium was collected for the tube formation assay, and GMSCs were harvested for RNA extraction. mRNA expression was validated using TaqMan Fast Advanced Master Mix (4444556, Invitrogen) with TaqMan Gene Expression Assays (Invitrogen) for CYR61 (Hs00155479_m1), EDIL3 (Hs00964112_m1), and GAPDH (Hs02786624_g1), according to the manufacturer's protocol.

Techniques: Modification, Derivative Assay, Cell Culture, RNA Sequencing, Membrane, Western Blot, Staining

Journal: Journal of Dental Sciences

Article Title: Laser-modified titanium surfaces induce sex-dimorphic secretion of angiogenic factors by gingiva-derived mesenchymal stromal cells

doi: 10.1016/j.jds.2025.09.013

Figure Lengend Snippet: Silencing CCN1 or EDIL3 in gingiva-derived mesenchymal stromal cells (GMSCs) impairs HUVEC angiogenesis. GMSCs were seeded on machined, lasered, or SLA (sand-blasted, large-grit, acid-etched) titanium discs and transfected with siRNAs targeting CCN1 or EDIL3 for 24 h. (A) Relative mRNA expression of CCN1 and EDIL3 after siRNA transfection. NC, negative control siRNA. (B) Representative images of HUVEC tube formation in response to conditioned medium (CM) collected from female (F) and male (M) GMSCs. Images were captured using the ImageXpress Pico system. Scale bars = 500 μm. (C) Quantification of tube formation (branch points, total length, segments, and nodes) using ImageJ. Data are mean ± standard deviation (n = 2, one female and one male). One-way ANOVA analysis with Tukey's multiple comparison test. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001, ∗∗∗∗P < 0.0001. ns, not significant.

Article Snippet: Subsequently, the medium was replaced with fresh exosome-free α-MEM and cells were cultured for an additional 48 h. Conditioned medium was collected for the tube formation assay, and GMSCs were harvested for RNA extraction. mRNA expression was validated using TaqMan Fast Advanced Master Mix (4444556, Invitrogen) with TaqMan Gene Expression Assays (Invitrogen) for CYR61 (Hs00155479_m1), EDIL3 (Hs00964112_m1), and GAPDH (Hs02786624_g1), according to the manufacturer's protocol.

Techniques: Derivative Assay, Transfection, Expressing, Negative Control, Standard Deviation, Comparison

Journal: Cell reports

Article Title: A retinoic acid:YAP1 signaling axis controls atrial lineage commitment

doi: 10.1016/j.celrep.2025.115687

Figure Lengend Snippet: (A) Simplified scheme of an E7.75 embryo. E7.75 Yap1 cKO (Sox2cre: Yap1 flox/flox) and heterozygous control (Sox2cre: Yap1 flox/+) embryos were processed for scRNA-seq analysis. (B) Unbiased clustering identified 19 clusters in these embryos, annotated as indicated in (E) using known markers (see also ). (C) UMAP plot shows Yap1 expression levels in indicated samples. Red arrowhead marks the extraembryonic (ExE) (Sox2 − Rhox5 + ; it does not express the cre). (D) UMAP plot of normalized number of cells in control and Yap1 cKO embryos. (E) Dot plot graph shows the number of DEGs in Yap1 cKO embryos compared with the control embryos in each cluster. Note that the bigger the dot, the more DEGs in the given population (abs(Log2FC) > 0.25, adj p < 0.05). (F) Heatmap shows DEGs in the SHF of Yap1 cKO embryos versus control embryos. Genes relevant for heart development are highlighted. Genes in red are involved in SHF patterning and RA activity. UMAP plots show the expression levels of indicated genes. (G) Violin plots show expression levels of indicated genes in each cluster, in controls and Yap1 cKO embryos. Dotted box highlights the SHF cluster. Adjusted p value, based on Bonferroni correction is below 0.05. * p < 0.05, ** p < 0.001, *** p < 0.0001. (H) Western blot (WB) analysis of E7.75 control and Yap1 cKO embryos. Analyzed proteins are indicated. GAPDH was used as loading control. Number of embryos loaded per lane is shown on top. (I) Graphs show RT-qPCR analysis of relevant DEGs identified in the scRNA-seq analysis. The analyzed genes are indicated. ( n = 3) Data are represented as mean ± SEM. Statistical analysis: Student’s t test. * p < 0.05, ** p < 0.01, *** p < 0.001. (J) WB of E8.25 control and Yap1 cKO embryos blotted against the indicated proteins. GAPDH was used as loading control. The number of embryos loaded per lane is indicated.

Article Snippet: The Yap1 fl/fl ( Yap1 tm1.1Dupa /J) and the Sox2cre mice (B6.Cg-Edil3/J) were purchased from Jackson laboratories.

Techniques: Control, Expressing, Activity Assay, Western Blot, Quantitative RT-PCR

Journal: Cell reports

Article Title: A retinoic acid:YAP1 signaling axis controls atrial lineage commitment

doi: 10.1016/j.celrep.2025.115687

Figure Lengend Snippet: (A) Simplified scheme of an E7.75 embryo. E7.75 Yap1 cKO (Sox2cre: Yap1 flox/flox) and heterozygous control (Sox2cre: Yap1 flox/+) embryos were processed for scRNA-seq analysis. (B) Unbiased clustering identified 19 clusters in these embryos, annotated as indicated in (E) using known markers (see also ). (C) UMAP plot shows Yap1 expression levels in indicated samples. Red arrowhead marks the extraembryonic (ExE) (Sox2 − Rhox5 + ; it does not express the cre). (D) UMAP plot of normalized number of cells in control and Yap1 cKO embryos. (E) Dot plot graph shows the number of DEGs in Yap1 cKO embryos compared with the control embryos in each cluster. Note that the bigger the dot, the more DEGs in the given population (abs(Log2FC) > 0.25, adj p < 0.05). (F) Heatmap shows DEGs in the SHF of Yap1 cKO embryos versus control embryos. Genes relevant for heart development are highlighted. Genes in red are involved in SHF patterning and RA activity. UMAP plots show the expression levels of indicated genes. (G) Violin plots show expression levels of indicated genes in each cluster, in controls and Yap1 cKO embryos. Dotted box highlights the SHF cluster. Adjusted p value, based on Bonferroni correction is below 0.05. * p < 0.05, ** p < 0.001, *** p < 0.0001. (H) Western blot (WB) analysis of E7.75 control and Yap1 cKO embryos. Analyzed proteins are indicated. GAPDH was used as loading control. Number of embryos loaded per lane is shown on top. (I) Graphs show RT-qPCR analysis of relevant DEGs identified in the scRNA-seq analysis. The analyzed genes are indicated. ( n = 3) Data are represented as mean ± SEM. Statistical analysis: Student’s t test. * p < 0.05, ** p < 0.01, *** p < 0.001. (J) WB of E8.25 control and Yap1 cKO embryos blotted against the indicated proteins. GAPDH was used as loading control. The number of embryos loaded per lane is indicated.

Article Snippet: Sox2 cre/+ B6.Cg-Edil3/J , Jackson Research Laboratory , RRID:IMSR_JAX:00845.

Techniques: Control, Expressing, Activity Assay, Western Blot, Quantitative RT-PCR