Journal: Genes & Diseases

Article Title: Targeting MDK alleviates bone loss via dual regulation of osteogenic differentiation and inflammatory cytokine expression

doi: 10.1016/j.gendis.2025.101931

Figure Lengend Snippet: Recombinant MDK protein inhibits osteogenic differentiation in vitro in a dose-dependent manner. (A) Cell viability after treating MC3T3-E1 cells with recombinant MDK protein after 48 h, assessed using the CCK-8 assay. Inter-group comparisons were analyzed by one-way ANOVA. (B) Western blotting analysis of ALP, RUNX2, OSX, and OCN expression levels following MDK treatment (7 days). (C–F) Reverse transcription PCR analysis of mRNA expression levels of Alpl , Runx2 , Sp7 , and Bglap in MC3T3-E1 cells following MDK treatment (7 days). β-actin served as the internal control. Inter-group comparisons were analyzed by one-way ANOVA. (G, H) ALP staining and activity assays were performed after inducing MC3T3-E1 cells with recombinant MDK protein (0–600 ng/mL) for 14 days. Inter-group comparisons were analyzed by one-way ANOVA. (I, J) ARS staining and quantitative analysis were conducted after inducing MC3T3-E1 cells with recombinant MDK protein (0–600 ng/mL) for 21 days. Inter-group comparisons were analyzed by one-way ANOVA. Scale bar, 100 μm ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001; “ns” indicates non-significant differences.

Article Snippet: Procell Life Science & Technology Co., Ltd. (Wuhan, China) supplied the pre-osteoblastic cell line MC3T3-E1 Subclone 14.

Techniques: Recombinant, In Vitro, CCK-8 Assay, Western Blot, Expressing, Reverse Transcription, Control, Staining, Activity Assay

Journal: Genes & Diseases

Article Title: Targeting MDK alleviates bone loss via dual regulation of osteogenic differentiation and inflammatory cytokine expression

doi: 10.1016/j.gendis.2025.101931

Figure Lengend Snippet: MDK suppresses osteoblast differentiation via the PI3K/AKT signaling pathway. (A, B) Western blot detection of the effect of recombinant MDK protein on the protein expression of molecules in the PI3K/AKT signaling pathway during the differentiation of MC3T3-E1 to osteoblasts (7 days). Inter-group comparisons were analyzed by a two-tailed unpaired Student's t -test (for normally distributed data with equal variance). (C, D) ALP, RUNX2, and OSX expression levels were detected by Western blotting. MC3T3-E1 cells were pretreated with 30 μM LY294002. Osteogenic differentiation was induced for 7 days. Inter-group comparisons were analyzed by one-way ANOVA. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001; “ns” indicates non-significant differences.

Article Snippet: Procell Life Science & Technology Co., Ltd. (Wuhan, China) supplied the pre-osteoblastic cell line MC3T3-E1 Subclone 14.

Techniques: Western Blot, Recombinant, Expressing, Two Tailed Test

Journal: Genes & Diseases

Article Title: Targeting MDK alleviates bone loss via dual regulation of osteogenic differentiation and inflammatory cytokine expression

doi: 10.1016/j.gendis.2025.101931

Figure Lengend Snippet: Recombinant MDK protein triggers the activation of inflammatory cytokines through the NF-κB signaling pathway. (A, B) IL-6, TNFα, and IL-1β expression levels were detected using Western blotting. MC3T3-E1 cells were treated with recombinant MDK protein (600 ng/mL). Osteogenic differentiation was induced for 7 days. Inter-group comparisons were analyzed by a two-tailed unpaired Student's t -test (for normally distributed data with equal variance). (C, D) Western blotting analysis of NF-κB signaling pathway molecules in MC3T3-E1 cells treated with recombinant MDK protein for 7 days during osteoblastic differentiation. Inter-group comparisons were analyzed by two-tailed unpaired Student's t -test (for normally distributed data with equal variance). (E, F) IL-6 and IL-1β expression levels were detected using Western blotting. MC3T3-E1 cells were pretreated with 10 μM BAY 11–7082. Osteogenic differentiation was induced for 7 days. Inter-group comparisons were analyzed by one-way ANOVA. ∗ p < 0.05 and ∗∗ p < 0.01.

Article Snippet: Procell Life Science & Technology Co., Ltd. (Wuhan, China) supplied the pre-osteoblastic cell line MC3T3-E1 Subclone 14.

Techniques: Recombinant, Activation Assay, Expressing, Western Blot, Two Tailed Test

Journal: Calcified Tissue International

Article Title: Dual Osteoprotective Actions of the Kefir Peptide KFP-1: Enhancement of Bone Formation and Suppression of Bone Resorption in Cells and Murine Models

doi: 10.1007/s00223-026-01487-w

Figure Lengend Snippet: KFP-1 promotes osteoblastic differentiation and mineralization in BMMSCs and MC3T3-E1 cells. BMMSCs and MC3T3-E1 cells were induced to differentiate into osteoblasts with varying concentrations of KFP-1. A Representative ALP staining images of BMMSCs on Day 14 (scale bar: 1 mm). B Quantitative analysis of ALP staining by DMSO solubilization and measuring absorbance at 562 nm. C ALP activity in BMMSCs measured on Days 7 and 14 with 20 μg/mL KFP-1. D ALP activity in MC3T3-E1 cells on Day 14 with different KFP-1 concentrations. E Representative alizarin red staining images of BMMSCs on Day 21 (scale bar: 0.5 mm). F Quantification of mineralization by solubilizing bound alizarin red with 10% cetylpyridinium chloride and measuring absorbance at 405 nm. Data are mean ± SD (n ≥ 3). * p < 0.05, *** p < 0.001 vs. control (without KFP-1)

Article Snippet: The MC3T3-E1 cell line (EP-CL-0378; Elabscience Biotechnology Inc., Houston, TX, USA) was cultured in complete α-MEM growth medium.

Techniques: Staining, Activity Assay, Control