Journal: bioRxiv

Article Title: A Wnt-responsive fibrocartilage progenitor system coordinates postnatal mandibular condylar cartilage growth

doi: 10.64898/2026.03.25.714159

Figure Lengend Snippet: (A) Differential gene expression analysis was performed comparing H2B–EGFP –positive and H2B–EGFP –negative cells within the MC-progenitor cluster identified by single-cell RNA sequencing. Genes are ranked by statistical significance. Foxm1 is significantly enriched in the H2B–EGFP –positive population, whereas Tgfb1 is enriched in the H2B–EGFP –negative population, indicating divergent transcriptional programs associated with Wnt activity. (B) Feature plots were generated to visualize expression of representative genes across mandibular condylar cartilage populations. Wnt-responsive cells show enriched expression of Foxm1 and IGF signaling–related genes ( Igf1 , Igf1r , Igfbp4 , Igfbp7 ), whereas Wnt-low populations express Tgfb1 and related factors ( Igf2r , Igfbp5 , Igfbp6 ), supporting distinct signaling states. (C) Western blot analysis was performed in isolated Wnt-responsive cells transfected with control vector or constitutively active β-catenin (S33Y). Cells were stimulated with recombinant IGF1 for the indicated time points. β-catenin activation enhances Foxm1 expression and downstream mitogenic signaling, including ERK and IGF1R phosphorylation, indicating that β-catenin promotes proliferative signaling responses. (D) Co-immunoprecipitation was performed to assess interaction between β-catenin and Foxm1. Cell lysates immunoprecipitated with anti–β-catenin antibody show enrichment of Foxm1 compared with control IgG, indicating a physical association between β-catenin and Foxm1. (E,F) Histological and immunofluorescence analyses were performed on mandibular condyles from control and Axin2 CreERT2 ;Ctnnb1 fl/+ ;Foxm1 fl/+ compound heterozygous mice at P42. H&E staining reveals reduced fibrocartilage thickness, and Ki67 staining shows decreased proliferative activity, indicating cooperative effects of β-catenin and Foxm1 in maintaining fibrocartilage proliferation. Scale bar, 100 μm. (G) Quantification of fibrocartilage thickness and Ki67-positive cells was performed. Compound heterozygous mice show reduced fibrocartilage thickness and decreased proliferation compared with controls. Data are presented as mean ± s.d. Each dot represents one biologically independent animal. (H,I) Histological and immunofluorescence analyses were performed on mandibular condyles from control and Axin2 CreERT2 ;Foxm1 fl/fl mice at P42. Foxm1 deletion results in marked condylar hypoplasia and reduced proliferative activity, indicating a critical role for Foxm1 in fibrocartilage growth. Scale bar, 100 μm. (J) Quantification of cartilage thickness and proliferative indices was performed in Foxm1 conditional knockout mice. Foxm1 deficiency significantly reduces cartilage growth and proliferation. Data are presented as mean ± s.d. Each dot represents one biologically independent animal. Statistical significance was assessed using two-tailed Student’s t-test. n.s., not significant; **P < 0.01; ****P < 0.0001. Abbreviations: sz, superficial zone; fc, fibrocartilage zone; cc, chondrocartilage zone.

Article Snippet: Cells were stimulated with recombinant mouse IGF1 (Cat No. 791-MG-050, R&D systems) for 0, 30, 60, or 180 min.

Techniques: Gene Expression, Single Cell, RNA Sequencing, Activity Assay, Generated, Expressing, Western Blot, Isolation, Transfection, Control, Plasmid Preparation, Recombinant, Activation Assay, Phospho-proteomics, Immunoprecipitation, Immunofluorescence, Staining, Knock-Out, Two Tailed Test

Journal: bioRxiv

Article Title: A Wnt-responsive fibrocartilage progenitor system coordinates postnatal mandibular condylar cartilage growth

doi: 10.64898/2026.03.25.714159

Figure Lengend Snippet: (A) RNAscope in situ hybridization showing Foxm1 transcript localization within the fibrocartilage compartment of the mandibular condyle. (B) RNAscope detection of Igf1 transcripts enriched in the superficial region of the fibrocartilage layer. (C) Violin plot comparing Foxm1 expression between H2B-EGFP –positive and H2B-EGFP –negative cells within the MC-progenitor cluster. Scale bars: 100 μm.

Article Snippet: Cells were stimulated with recombinant mouse IGF1 (Cat No. 791-MG-050, R&D systems) for 0, 30, 60, or 180 min.

Techniques: RNAscope, In Situ Hybridization, Expressing

Journal: Frontiers in Aging Neuroscience

Article Title: Western diet-induced visceral adipose tissue inflammation promotes Alzheimer’s disease pathology via microglial activation in a mouse model

doi: 10.3389/fnagi.2025.1644988

Figure Lengend Snippet: Identification of epididymal white adipose tissue (eWAT)-associated pro-inflammatory cytokines using AB array and QuantSeq 3′-mRNA sequencing. (A) Heatmap of cytokine expression in eWAT using the AB array. A total of 115 cytokines were significantly upregulated in the WD group compared with those in the control group, as indicated by the color scale representing relative expression levels (blue: downregulation, red: upregulation). (B) Heatmap of cytokine expression in eWAT determined using QuantSeq 3′-mRNA sequencing. In this analysis, 132 cytokines showed significant upregulation in the WD group compared with those in the control group, with similar color coding for relative expression changes. (C) A subset of five cytokines, including CCL8, CCL9, CXCL13, and IL-18, were consistently increased across both datasets (AB array for eWAT and QuantSeq for plasma) and were identified as eWAT-associated pro-inflammatory cytokines contributing to systemic and neuroinflammation in the WD group. (D) Protein–protein interaction (PPI) network analysis of differentially expressed genes (DEGs) was constructed using known interaction data. The network highlights significant interactions among key pro-inflammatory cytokines, illustrating the interconnected roles of these factors in inflammatory signaling. (E) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the DEGs, with significant pathways related to inflammation, such as cytokine–cytokine receptor interaction and the chemokine signaling pathway. The x -axis denotes the number of genes associated with each pathway, while the size and color of the dots represent gene count and statistical significance, respectively.

Article Snippet: Recombinant mouse cytokines CCL8 (PMC1104) and CXCL13 (PMC1614) were purchased from Peprotech (Rocky Hill, NJ, USA), and CCL9 (463-MG) and IL-18 (9139) were purchased from R&D Systems (Minneapolis, MN, USA).

Techniques: Ab Array, Sequencing, Expressing, Control, Clinical Proteomics, Construct

Journal: Frontiers in Aging Neuroscience

Article Title: Western diet-induced visceral adipose tissue inflammation promotes Alzheimer’s disease pathology via microglial activation in a mouse model

doi: 10.3389/fnagi.2025.1644988

Figure Lengend Snippet: Validation of epididymal white adipose tissue (eWAT)-associated pro-inflammatory cytokines. A Western diet (WD) significantly increased the levels of eWAT-associated pro-inflammatory cytokines in both eWAT and plasma. (A–E) Western blot analysis validated a marked increase in the expression of eWAT-associated pro-inflammatory cytokines in the WD group compared with that in the controls. Specifically, the levels of CCL8, CCL9, CXCL13, and IL-18 were significantly upregulated in the eWAT of WD group, indicating heightened inflammatory activity in this tissue. (F–I) Corresponding ELISA analysis of plasma samples showed increased concentrations of these cytokines in the WD group, with significant increases observed for CCL8, CCL9, CXCL13, and IL-18. Importantly, in the eWAT-removal group, plasma levels of these cytokines were reduced to levels comparable to the sham, confirming that eWAT is a primary source of the increased circulating cytokines observed in the WD group. Both the control (normal diet-fed) and the WD group underwent the same surgical procedure without eWAT removal. The results are presented as mean ± SD with a sample size of n = 9 for western blot quantification and n = 5 for plasma ELISA assay. Asterisks denote statistically significant differences compared with the control group, with * p < 0.05 and *** p < 0.001, ns = no significant difference, One-way analysis of variance followed by Dunnett’s post hoc test.

Article Snippet: Recombinant mouse cytokines CCL8 (PMC1104) and CXCL13 (PMC1614) were purchased from Peprotech (Rocky Hill, NJ, USA), and CCL9 (463-MG) and IL-18 (9139) were purchased from R&D Systems (Minneapolis, MN, USA).

Techniques: Biomarker Discovery, Western Blot, Clinical Proteomics, Expressing, Activity Assay, Enzyme-linked Immunosorbent Assay, Control

Journal: Frontiers in Aging Neuroscience

Article Title: Western diet-induced visceral adipose tissue inflammation promotes Alzheimer’s disease pathology via microglial activation in a mouse model

doi: 10.3389/fnagi.2025.1644988

Figure Lengend Snippet: Epididymal white adipose tissue (eWAT)-associated pro-inflammatory cytokines induce significant neurotoxicity in HT22 cells. (A) The viability of HT22 hippocampal neuronal cells was not affected after 24 h of direct culture with eWAT-associated cytokines (CCL8, CCL9, CXCL13, and IL-18) at varying concentrations (25, 50, and 100 ng/mL). The results indicate no direct cytotoxic effects of these cytokines on hippocampal neurons under these conditions. (B) To evaluate the cytotoxicity of eWAT-associated cytokine in BV2 microglia, each cytokine was treated at a concentration of 50, 100, and 200 ng/ml. eWAT-associated cytokines did not induce cytotoxicity in BV2 microglial cells after 24 h of exposure, but they significantly increased microglial proliferation. (C) Treatment with each cytokine at a concentration of 100 ng/mL and analysis by ELISA showed that eWAT-associated pro-inflammatory cytokines significantly increased IL-6 secretion in BV2 microglial cells compared to those of untreated controls. (D) Intracellular reactive oxygen species (ROS) levels were measured in BV2 cells using a DCFDA fluorescence assay. Treatment with eWAT-associated cytokines led to a significant increase in ROS production, indicating oxidative stress–mediated microglial activation. (E) Treatment of HT22 hippocampal neurons with conditioned medium (CM) containing 100 ng/mL of each eWAT-associated cytokine to BV2 microglia significantly decreased cell viability, demonstrating the neurotoxic effect of cytokine-activated microglia on neurons. The data are presented as mean ± SD, with sample sizes ranging from n = 3 to n = 6 per group. Statistical significance is indicated by asterisks (*, compared with the control at 24 h): * p < 0.05, ** p < 0.01, and **** p < 0.0001, ns = no significant difference, One-way analysis of variance followed by Dunnett’s post hoc test.

Article Snippet: Recombinant mouse cytokines CCL8 (PMC1104) and CXCL13 (PMC1614) were purchased from Peprotech (Rocky Hill, NJ, USA), and CCL9 (463-MG) and IL-18 (9139) were purchased from R&D Systems (Minneapolis, MN, USA).

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Fluorescence, Activation Assay, Control