Journal: Frontiers in Immunology

Article Title: Hyaluronic acid−CD44 signaling from decidual stromal cells orchestrates dNK1 differentiation and immune tolerance in early pregnancy

doi: 10.3389/fimmu.2026.1777567

Figure Lengend Snippet: HA-induced activation of the canonical Wnt pathway upregulates FOSL2 to expand the dNK1-like subpopulation. (A) Analysis of transcriptional activity and expression levels of transcription factors in three dNK subsets using scRNA-seq data from normal decidual tissue ( n = 11). (B) Transcriptional activity and expression levels of FOSL2 in decidual tissues from normal pregnancy ( n = 5) and RSA groups ( n = 3). (C) Changes in NK92MI cells of the five transcription factors ( STAT3, MAFB, HES1, FOSL2, ETV5 ) characterized by high transcriptional activity and expression in the dNK1 subset following HMW-HA treatment, as determined by RT−qPCR ( n = 6 per group). (D) FOSL2 expression in NK92MI cells following HMW-HA treatment, assessed by WB ( n = 3 per group). (E) Wnt1 and β-catenin protein expression in NK92MI cells treated with or without HMW-HA in the presence or absence of CD44 blockade ( n = 3 per group). (F) FOSL2 expression in HA−treated NK92MI cells after addition of CD44−blocking antibody or the Wnt pathway inhibitor IWP−2 to the culture system ( n = 3 per group). Data are expressed as mean ± SD; * P < 0.05; ** P < 0.01; ns, not significant. EP, early pregnancy; SA, spontaneous abortion; RSA, recurrent spontaneous abortion.

Article Snippet: To determine the signaling pathways involved, we used an anti-CD44 blocking antibody (30 μg/mL; BioxCell, BE0262) to block CD44, and IWP-2 (50 μM; MedChemExpress, HY-13912) to inhibit the canonical Wnt pathway.

Techniques: Activation Assay, Activity Assay, Expressing, Quantitative RT-PCR, Blocking Assay

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) Schematic representation of the R26-WntVis reporter. H2B–EGFP expression is driven by seven tandem TCF/LEF binding elements upstream of a minimal thymidine kinase promoter, enabling visualization of canonical Wnt/β-catenin signaling activity. (B) Sagittal frozen sections from R26-WntVis mice at postnatal day 14 (P14) were analyzed by fluorescence microscopy. H2B–EGFP–positive nuclei are enriched in the superficial (sz; open arrowheads) and fibrocartilage (fc; closed arrowheads) zones and largely absent from the deeper chondrocartilage (cc) zone, indicating spatial restriction of canonical Wnt activity to the upper compartments. Dashed boxes indicate regions shown at higher magnification. Scale bar, 100 µm. Data are representative of three biologically independent mice. (C) Single-cell RNA sequencing was performed on enzymatically dissociated mandibular condylar cartilage from P16 R26-WntVis mice. Uniform Manifold Approximation and Projection (UMAP) visualization identified nine transcriptionally distinct clusters following quality control filtering and shared nearest neighbor clustering. (D) Feature plots show expression of representative marker genes used to annotate fibrocartilage ( Col1a1 ) and chondrocartilage ( Col2a1 ) populations, enabling classification of major cartilage compartments. (E) Feature plot and dot plot analyses were used to examine H2B–EGFP reporter expression across clusters. Cluster 5 shows marked enrichment of H2B–EGFP–positive cells and was designated as the MC-progenitor cluster, indicating a Wnt-responsive progenitor-like population. (F) KEGG pathway enrichment analysis was performed on differentially expressed genes in the MC-progenitor cluster relative to differentiated cartilage clusters. Cell-cycle–associated pathways are significantly enriched, consistent with a proliferative transcriptional state. (G) Trajectory inference was performed using Monocle3 to assess lineage relationships among clusters. The MC-progenitor cluster (cluster 5) is positioned at the root of a bifurcating trajectory leading toward fibrocartilage and chondrocartilage lineages, supporting its role as an upstream progenitor population.

Article Snippet: Pharmacological treatments were performed using the Foxm1 inhibitor RCM-1, the β-catenin inhibitor XAV-939, or a canonical Wnt pathway activator (Wnt agonist 1) (all from Selleck Chemicals, Japan).

Techniques: Expressing, Binding Assay, Activity Assay, Fluorescence, Microscopy, Single Cell, RNA Sequencing, Control, Marker

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: Isolated Wnt-responsive cells were cultured in vitro and treated with the Wnt pathway inhibitor XAV-939 (1 μM or 5 μM), the Foxm1 inhibitor RCM-1 (4 μM or 20 μM), or the Wnt agonist Wnt agonist 1 (0.2 nM or 1 nM). Cell proliferation was quantified by measuring total cell number after treatment. Pharmacological inhibition of canonical Wnt signaling using XAV-939 significantly reduced cell expansion compared with control cultures. Similarly, inhibition of Foxm1 using RCM-1 markedly suppressed proliferation. In contrast, activation of canonical Wnt signaling by Wnt agonist 1 increased cell numbers relative to controls. These findings support a cooperative role for canonical Wnt signaling and Foxm1 in promoting proliferative capacity of Wnt-responsive cells in vitro . Bars represent mean ± s.d. Each dot represents one biologically independent sample. Statistical significance was determined by one-way ANOVA followed by multiple comparisons testing. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.

Article Snippet: Pharmacological treatments were performed using the Foxm1 inhibitor RCM-1, the β-catenin inhibitor XAV-939, or a canonical Wnt pathway activator (Wnt agonist 1) (all from Selleck Chemicals, Japan).

Techniques: Isolation, Cell Culture, In Vitro, Inhibition, Control, Activation Assay