Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-30a regulates chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells through targeting Sox9

doi: 10.3892/etm.2019.8148

Figure Lengend Snippet: Characterization of hBMSCs. (A) MSCs from the human bone marrow were isolated and cultured. Alizarin red staining, Oil red O staining and toluidine blue staining were performed to assess the osteogenic, adipogenic and chondrogenic differentiation potential, respectively. (B) The cell membrane markers of MSCs were determined by flow cytometry. Surface markers CD45, CD14, CD44, CD105, CD29 and CD34 were detected and analyzed (n=3). MSCs, mesenchymal stem cells; hBMSCs, bone marrow-derived mesenchymal stem cells; CD, cluster of differentiation.

Article Snippet: Briefly, hBMSCs were incubated with Human Mesenchymal Stem Cell Growth Medium (HUXMX-90011; Cyagen Biosciences, Inc., kit components included human mesenchymal stem cell basal medium, 10% human mesenchymal stem cell-qualified fetal bovine serum, 1% penicillin-streptomycin and 2 mmol/l glutamine), after isolation from bone marrow at a 37°C and 5% CO 2 incubator.

Techniques: Isolation, Cell Culture, Staining, Membrane, Flow Cytometry, Derivative Assay

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-30a regulates chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells through targeting Sox9

doi: 10.3892/etm.2019.8148

Figure Lengend Snippet: Differential expression of Sox9 and SOX9-associated miRNAs during chondrogenic differentiation of MSCs. (A) Western blotting and (B) quantification of protein expression. β-actin was used as an internal reference. One-way ANOVA, *P<0.05 between two indicated groups; # P<0.05 vs. day 0 group. (C-F) After induction of chondrogenic differentiation (7–21 days), the levels (C) miR-30a, (D) miR-195, (E) miR-216b and (F) miR-15a were determined by reverse transcription-quantitative PCR. Data are presented as mean ± SD of three independent experiments. One-way ANOVA, *P<0.05 vs. day 0. SOX9, transcription factor SOX-9; MSCs, mesenchymal stem cells; miR or miRNA, microRNA.

Article Snippet: Briefly, hBMSCs were incubated with Human Mesenchymal Stem Cell Growth Medium (HUXMX-90011; Cyagen Biosciences, Inc., kit components included human mesenchymal stem cell basal medium, 10% human mesenchymal stem cell-qualified fetal bovine serum, 1% penicillin-streptomycin and 2 mmol/l glutamine), after isolation from bone marrow at a 37°C and 5% CO 2 incubator.

Techniques: Quantitative Proteomics, Western Blot, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: MicroRNA-30a regulates chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells through targeting Sox9

doi: 10.3892/etm.2019.8148

Figure Lengend Snippet: Effect of miR-30a on SOX9 expression. (A) Alignment of sequences of the wild-type SOX9, miR-30a and mutant SOX9 3′UTR. The lines between the sequences shows how complementary the sequences are. The mutant SOX9 3′UTR contains 5 nucleotide mutations, which are indicated in green. (B) Relative luciferase activity was assayed after the 293 cells were co-transfected with a random DNA sequence or the miR-30a mimic or the negative control mimic and the reporter plasmid with the wild-type or mutant SOX9 3′UTR. Data are presented as mean ± SD of three independent experiments. One-way ANOVA, *P<0.05 versus control. (C) MSCs were transfected with either miR-30a mimic or transfection reagents, the levels of miR-30a were determined by reverse transcription-quantitative PCR. (D) Western blotting and (E) quantification of protein expression. β-actin was used as an internal reference. Data are presented as mean ± SD of three independent experiments. One-way ANOVA, *P<0.05 vs. control. SOX9, transcription factor SOX-9; MSCs, mesenchymal stem cells; miR or miRNA, microRNA; NC, negative control; UTR, untranslated region.

Article Snippet: Briefly, hBMSCs were incubated with Human Mesenchymal Stem Cell Growth Medium (HUXMX-90011; Cyagen Biosciences, Inc., kit components included human mesenchymal stem cell basal medium, 10% human mesenchymal stem cell-qualified fetal bovine serum, 1% penicillin-streptomycin and 2 mmol/l glutamine), after isolation from bone marrow at a 37°C and 5% CO 2 incubator.

Techniques: Expressing, Mutagenesis, Luciferase, Activity Assay, Transfection, Sequencing, Negative Control, Plasmid Preparation, Control, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot