Journal: Frontiers in Immunology

Article Title: In silico design of novel precision vaccine targeting sclerostin epitopes for osteoporosis prevention and treatment

doi: 10.3389/fimmu.2025.1644437

Figure Lengend Snippet: Screening and analysis of high-affinity epitopes on SOST. (A) ELISA experiments were conducted to identify SOST fragments with strong binding affinity for ROMO, revealing that SOST 114–143 and SOST 143–173 exhibit significantly higher affinity ( P <0.01). (B) A schematic diagram delineating the binding functional regions associated with the high-affinity fragments of SOST. (C) ELISA results indicate that SOST 131–163 displays the highest affinity for ROMO ( P <0.01), thereby identifying it as a potent functional epitope of SOST. (D-a) SOST 131–163 fragment (highlighted in yellow) is located within the loop3 domain of SOST protein. (D-b) Docking studies indicate that SOST 131–163 fragment interacts with ROMO light chain, yielding a binding free energy of -25.8 kcal/mol and an interface area of 712.9 Ų. (D-c) Additionally, SOST 131–163 fragment can bind to the ROMO heavy chain, resulting in a binding free energy of -33.19 kcal/mol and an interface area of 451.6 Ų. (E) CTL epitopes within SOST 131–163 sequence include two strong binder epitopes and four weak binder epitopes. (F) HTL epitopes in SOST 131–163 sequence comprise one strong binder epitope and four weak binder epitopes. Predictions of B cell epitopes for SOST 131–163 sequence are illustrated, including predicted linear B cell epitopes (G) and predicted discontinuous B cell epitopes (H) .

Article Snippet: In brief, 1 μg/mL of human SOST protein (MedChemExpress Inc.) was coated onto the wells of MaxiSorp microtiter plates (Thermo Fisher Scientific Inc.) and incubated overnight at 4°C.

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, Functional Assay, Sequencing

Journal: Cells

Article Title: Sclerostin Alters Tumor Cell Characteristics of Oral Squamous Cell Carcinoma and May Be a Key Player in Local Bone Invasion

doi: 10.3390/cells13020137

Figure Lengend Snippet: Immunohistochemical staining protocol.

Article Snippet: Sclerostin , Mouse, monoclonal, clone AbD09097_h/mIgG 2a, 1:1200 , HIER (pH 9) , Dako EnVision FLEX , BioRad, Hercules, CA, USA (HCA230Z).

Techniques: Immunohistochemical staining, Staining

Journal: Cell reports

Article Title: Osteocyte intrinsic TGFβ signaling regulates bone quality through perilacunar/canalicular remodeling

doi: 10.1016/j.celrep.2017.10.115

Figure Lengend Snippet: (A–D) qPCR analysis of PLR genes Mmp13, Mmp14 and Ctsk and Serpine1 upon TGFβ (5ng/mL) treatment in MLO-Y4 (A, B) and OCY454 (C, D) cells. (n=3 replicates/group). (E, F) Intracellular pH (pHi) of MLO-Y4 cells after 3 days of TGFβ (5ng/ml), TβRI inhibitor SB-431542 (10 μM), or recombinant sclerostin (rhSCL, 10 ng/ml). The representative image (E) shows the shift in the emission peak from 580 nm to 640 nm after TGFβ treatment of MLO-Y4 cells. Scale bar, 100 μm). TGFβ-induced acidification is blocked by SB-431542 (F) (n=4 replicates/group). Error bars indicate mean ± SD of 3 independent experiments, *p<0.05 different from control mRNA, a-p<0.05 different from control pHi, b-p<0.05 different from TGFβ pHi, and c-p<0.05 different from rhSCL pHi. Statistics calculated from Student’s t test.

Article Snippet: For treatment, cells were cultured in α-MEM containing 0.5–1% fetal bovine serum, supplemented with 5 ng/ml TGFβ1 (Humanzyme, HZ-1011), 10 μM SB431542 (Sigma, S4317) or 10 ng/ml recombinant human sclerostin (rhSCL, R&D Systems) for the indicated times.

Techniques: Recombinant, Control

Journal: Cell reports

Article Title: Osteocyte intrinsic TGFβ signaling regulates bone quality through perilacunar/canalicular remodeling

doi: 10.1016/j.celrep.2017.10.115

Figure Lengend Snippet: (A, B) TβRII-stained osteocytes (A) (arrow, scale bar, 50 μm) in the femoral cortical bone from WT and TβRIIocy−/− mice (8-week old males) were quantified as percentage of positively stained osteocytes normalized to total bone area (B) (n=5 mice/group) (C) qPCR analysis of TβRII and Serpine1 in WT and TβRIIocy−/− femoral bones. (n=8–10 mice/group). (D, E) Silver nitrate stained images of WT and TβRIIocy−/− femoral cortical bone shows the osteocyte lacuno-canalicular network (D) and canalicular length (E) (scale bar, 20 μm, n=5 mice/group). (F, G) qPCR analysis of PLR genes, Mmp2, Mmp13, Mmp14, Ctsk, and Acp5 (F) and OCY-specific genes, Sost, Dmp1 and Phex (G) in the WT and TβRIIocy−/− bones (n=8–10 mice/group) (H, I) IHC of MMP13, MMP14, CTSK and H&E staining of WT and TβRIIocy−/− femoral cortical bone. Arrows in the image indicate positively stained osteocytes (H) that were quantified and normalized to total bone area (I), (n=4 mice/group).(J–M) SRμT shows volume (J), degree of anisotropy (K), orientation (L) and mineralization (N) of osteocyte lacunae of WT and TβRIIocy−/− bone (n=3–4 mice/group). Error bars indicate mean ± SEM with *p<0.05 compared to WT from Student’s t test.

Article Snippet: For treatment, cells were cultured in α-MEM containing 0.5–1% fetal bovine serum, supplemented with 5 ng/ml TGFβ1 (Humanzyme, HZ-1011), 10 μM SB431542 (Sigma, S4317) or 10 ng/ml recombinant human sclerostin (rhSCL, R&D Systems) for the indicated times.

Techniques: Staining

Journal: Clinical Journal of the American Society of Nephrology

Article Title: Sclerostin and Dickkopf-1 in Renal Osteodystrophy

doi: 10.2215/cjn.06550810

Figure Lengend Snippet: Figure 1. | (A and B) Scatter plot of serum concentrations of sclerostin (pg/ml) at different levels of Ac.f. (A) and different levels of BFR/BS (B).

Article Snippet: Fifty microliters of serum was loaded per well, incubated overnight at 4°C, washed, and incubated for 1 hour at 37°C, followed by incubation at 4°C for 1 hour with a biotinylated polyclonal anti-sclerostin antibody (BAF 1406; R&D Systems) diluted to a concentration of 0.5 g/ml in dilution buffer.

Techniques:

Journal: International Journal of Molecular Medicine

Article Title: Icaritin promotes the osteogenesis of bone marrow mesenchymal stem cells via the regulation of sclerostin expression

doi: 10.3892/ijmm.2020.4470

Figure Lengend Snippet: Icaritin (1 µ M) increases mRNA levels of osteogenic marker genes. (A) SOST , (B) OCN , (C) Runx2 , (D) Alp and (E) β-catenin at the different time points of osteogenesis of human bone marrow-derived mesenchymal stem cells. DMSO was used as the control group. Data are presented as the mean ± standard deviation (n=3). * P<0.05 and ** P<0.01 vs. control group at the same stage. SOST, sclerostin; OCN, osteocalcin; Runx2, RUNX family transcription factor 2; Alp, alkaline phosphatase.

Article Snippet: The membrane was incubated with antibodies targeting β-actin (1:1,000; cat. no. 4970S; Cell Signaling Technology, Inc.), osteocalcin (OCN; 1:1,000; cat. no. MAB1419; R&D Systems, Inc.), RUNX family transcription factor 2 (Runx2; 1:1,000; cat. no. 12556; Cell Signaling Technology, Inc.), ALP (1:1,000; cat. no. AF2910; R&D Systems, Inc.), β-catenin (1:1,000; cat. no. 8480S; Cell Signaling Technology, Inc.) and SOST (1:1,000; cat. no. MAB1406; R&D Systems, Inc.) overnight at 4°C.

Techniques: Marker, Derivative Assay, Control, Standard Deviation

Journal: International Journal of Molecular Medicine

Article Title: Icaritin promotes the osteogenesis of bone marrow mesenchymal stem cells via the regulation of sclerostin expression

doi: 10.3892/ijmm.2020.4470

Figure Lengend Snippet: SOST overexpression reverses icaritin-induced osteogenesis of hBMSCs. (A) Western blot analysis for the protein level of SOST. (B) The mRNA expression of SOST in BMSCs, BMSCs-vector, and BMSCs-SOST. (C) Mineralization in cultured hBMSCs in BMSCs-vector and BMSCs-SOST groups with or without icaritin were detected at day 14. Magnification, ×10. (D) The mRNA levels of OCN, Runx2, Alp and β-actin were determined by reverse transcription quantitative polymerase chain reaction. Data are presented as mean ± standard deviation (n=3). ** P<0.01 vs. BMSCs group; # P<0.05 and ## P<0.01 vs. BMSCs-vector group. SOST, sclerostin; BMSCs, bone marrow-derived mesenchymal stem cells; OCN, osteocalcin; Runx2, RUNX family transcription factor 2; Alp, alkaline phosphatase.

Article Snippet: The membrane was incubated with antibodies targeting β-actin (1:1,000; cat. no. 4970S; Cell Signaling Technology, Inc.), osteocalcin (OCN; 1:1,000; cat. no. MAB1419; R&D Systems, Inc.), RUNX family transcription factor 2 (Runx2; 1:1,000; cat. no. 12556; Cell Signaling Technology, Inc.), ALP (1:1,000; cat. no. AF2910; R&D Systems, Inc.), β-catenin (1:1,000; cat. no. 8480S; Cell Signaling Technology, Inc.) and SOST (1:1,000; cat. no. MAB1406; R&D Systems, Inc.) overnight at 4°C.

Techniques: Over Expression, Western Blot, Expressing, Plasmid Preparation, Cell Culture, Reverse Transcription, Real-time Polymerase Chain Reaction, Standard Deviation, Derivative Assay