Journal: Bioactive Materials

Article Title: Sulfated polysaccharide prevents senescent adipocyte-driven osteonecrosis by stem cell fate reprogramming

doi: 10.1016/j.bioactmat.2025.11.039

Figure Lengend Snippet: Schematic illustration of the senescence-regulatory mechanisms of the sulfated polysaccharide in the glucocorticoid-induced bone marrow microenvironment. Bone marrow senescence plays a critical role in the pathogenesis of osteonecrosis. Glucocorticoids act on bone marrow target cells—adipocytes—to initiate primary bone marrow senescence via triggering a positive feedback loop through the prostaglandin/PPARγ/INK signaling axis. Subsequently, these senescent adipocytes propagate SASP factors to adjacent healthy cells through paracrine signaling or direct cell–cell contact, leading to secondary senescence. Sulfated chitosan (SCS) reprograms the lineage commitment bias of LepR + MSCs by activating the IGF-1/PI3K/Akt/mTOR signaling cascade, suppressing adipogenic differentiation and lipid biosynthesis pathways. SCS attenuates the spread of primary adipocyte senescence into secondary senescence, limiting the progressive amplification of the senescence cascade. Ultimately, this strategy halts the onset of senescence-driven osteonecrosis at an early stage and preserves the functional stability of the bone marrow microenvironment.

Article Snippet: Furthermore, to explore the molecular mechanisms by which SCS regulates MSCs lineage bias, bone marrow supernatant was collected on day 7 following co-treatment with SCS and MPS, and ELISA assays for IGF-1 (R&D Systems, MG100) and BMP-2 (R&D Systems, DBP200) were performed as described above.

Techniques: Amplification, Functional Assay

Journal: Bioactive Materials

Article Title: Sulfated polysaccharide prevents senescent adipocyte-driven osteonecrosis by stem cell fate reprogramming

doi: 10.1016/j.bioactmat.2025.11.039

Figure Lengend Snippet: SCS modulates mesenchymal stem cell lineage bias via activation of the IGF-1/PI3K/Akt/mTOR signaling pathway. ( A ) Quantitative analysis of osteocyte morphology in the trabecular bone matrix of the bone marrow at week 6 after MPS treatment with or without SCS, in the presence of various neutralizing antibodies (NAbs) and antagonistic proteins. ( B ) ELISA analysis of IGF-1 and BMP-2 levels in the femoral bone marrow and peripheral serum at day 7 following SCS treatment under MPS conditions. ( C and D ) Western blot analysis of phospho-PI3K, phospho-Akt, and phospho-mTOR (C), as well as phospho-Smad1/5/8, phospho-ERK, and phospho-p38 (D), in CD45 − Ter119 − CD31 − LepR + MSCs after 15-min stimulation with conditioned medium (CM) derived from bone marrow fluid at day 7 following SCS treatment. ( E – G ) Representative flow cytometry plots (E, F) and quantitative analysis (G) of CD45 − CD31 − Sca-1 + CD24 − adipocyte progenitor cells (APCs), CD45 − CD31 − Sca-1 + CD24 + MSCs (E), and CD45 − CD31 − Sca-1 − PDGFRα + (Pα + ) osteoprogenitor cells (OPCs) (F) from femoral bone marrow at day 14 post-MPS induction with or without combined treatment using SCS and IGF-1 NAb or Noggin. ( H and I ) Representative SA-β-Gal staining images (green) of the femur (H), and corresponding quantification (I), at week 4 following MPS treatment with SCS in combination with IGF-1 NAb or DMH1. Insets show magnified views of bone marrow (BM) and trabecular bone matrix (TBM) regions. (Scale bars, 100 μm and 25 μm) ( J ) qPCR analysis of 12 senescence-associated markers in ex vivo femoral bone tissues at week 4 following MPS treatment with SCS in combination with IGF-1 NAb or DMH1. ( K ) Representative Oil Red O staining images of CD45 − Ter119 − CD31 − LepR + MSCs sorted from femurs at day 7 following MPS treatment with SCS in combination with LY294002 or LDN-193189, after in vitro adipogenic induction. (Scale bars, 50 μm and 25 μm) ( L and M ) γ-H2A.X and telomere-associated DNA damage foci (TAFs) co-localization analysis (L), and corresponding quantification (M), in CD45 − Ter119 − CD31 + arteriolar ECs sorted from femurs at day 28 following MPS treatment with SCS in combination with rapamycin or LDN-193189, using immuno-FISH staining. (Scale bars, 7 μm and 1 μm) ( N and O ) Sequential fluorescent labeling using calcein (N) and quantification of mineral apposition rate (O) in femurs treated with SCS and MPS for 4 weeks, with or without LY294002 and/or GW9662. (Scale bars, 50 μm) ( P ) ELISA analysis of five senescence-associated cytokines in femoral bone marrow at day 28 following MPS treatment with SCS in combination with rapamycin and/or T0070907. ( Q and R ) Representative t-distributed stochastic neighbor embedding (t-SNE) plots (Q) from flow cytometric analysis of CD45 − CD31 − Sca-1 + CD24 − APCs, CD45 − CD31 − Sca-1 + CD24 + MSCs, CD45 − CD31 − Sca-1 − Pα + OPCs, CD45 − Ter119 − CD31 + arteriolar ECs, and CD45 − Ter119 − Emcn + sinusoidal ECs at day 14 following MPS treatment with SCS in combination with IGF-1 and/or rosiglitazone, and quantitative analysis of APCs (R) ( S ) Heatmap showing the fluorescent intensity distribution of Lamin-B1 expression across five cellular subpopulations as identified in the t-SNE clustering plot. ∗ P < 0.05 vs. IgG (empty lacunae); # P < 0.05 vs. IgG (filled lacunae). ∗ P < 0.05 vs. SCS; # P < 0.05 vs. SCS + IGF-1 NAb. Data are presented as mean ± SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001; ns, not significant. Statistical significance was determined using an unpaired two-tailed Student's t -test ( B ), or one-way ANOVA with Tukey's post hoc test ( A, G, I, J, O, P and R ).

Article Snippet: Furthermore, to explore the molecular mechanisms by which SCS regulates MSCs lineage bias, bone marrow supernatant was collected on day 7 following co-treatment with SCS and MPS, and ELISA assays for IGF-1 (R&D Systems, MG100) and BMP-2 (R&D Systems, DBP200) were performed as described above.

Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay, Western Blot, Derivative Assay, Flow Cytometry, Staining, Ex Vivo, In Vitro, Labeling, Expressing, Two Tailed Test

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Representative images of IHC staining. IHC staining of TNBC specimens was performed as described in the Methods section with antibodies to IR, IGF-1R, pErk1/2, and FOXO3a. The images are shown in 40× magnification. Abbreviations: IGF-1R, IGR-1 receptor; IHC, immunohistochemistry; IR, insulin receptor; pErk1/2, phosphorylated Erk1/2; TNBC, triple negative breast cancer.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Immunohistochemistry

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Representative images of protein localization for IR, IGF1R, FOXO3a, and pErk1/2. IHC staining of TNBC specimens was performed and evaluated for localization of proteins. The images are shown in 40× magnification. Abbreviations: IGF-1R, IGR-1 receptor; IHC, immunohistochemistry; IR, insulin receptor; pErk1/2, phosphorylated Erk1/2; TNBC, triple negative breast cancer.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Immunohistochemistry

Journal: Journal of the Endocrine Society

Article Title: Insulin receptor expression and its association with hyperinsulinemia in triple negative breast cancer

doi: 10.1210/jendso/bvag041

Figure Lengend Snippet: Overall survival for women with breast cancer based on IR and IGF-1R protein expression in KMPlot.com . KMPlot.com was used to generate overall survival Kaplan-Meier plots. The dataset used was not from our cohort but from Tang and colleagues ; n = 65 patients had IR (A) and IGF1R (B) protein expression available. The patients were split using the “auto select best cutoff” percentile option in KMPlot. No other restrictions were added to the analysis. The plots show hazard ratios with 95% confidence intervals in parentheses . Abbreviations: IGF-1R, IGR-1 receptor; IR, insulin receptor.

Article Snippet: Primary antibodies were anti-IR Ab (ab137747, Abcam, Cambridge, MA, RRID:AB_3717506) 1:300 dilution, anti-IGF-1R Ab (#3027, Cell Signaling, Danvers, CA, RRID:AB_2122378) 1:200 dilution, anti-FOXO3a Ab (#12829, Cell Signaling, RRID:AB_2636990) 1:3200 dilution, and anti-pErk1/2 (Thr202/Tyr204) Ab (#4370, Cell Signaling, RRID:AB_2315112) 1:400 dilution.

Techniques: Expressing