Journal: Frontiers in cell and developmental biology

Article Title: miR-129-5p Inhibits Bone Formation Through TCF4.

doi: 10.3389/fcell.2020.600641

Figure Lengend Snippet: FIGURE 6 | Rescue effect of miR-129-5p inhibitor on postmenopausal osteoporosis. (A) Representative images showing calvarial mineral apposition rate of C57BL/6 mice after OVX and inhibitor-129-5p treatment. Scale bar: 10 µm. BL (baseline): sacrifice before RNA treatment. Sham: sham OVX operation group. OVX: OVX group. Mock: transfection reagent control group. inhibit-NC: inhibitor-NC–treated group. inhibit-129: inhibitor-129-5p–treated group. (B) Calvarial mineral apposition rates of C57BL/6 mice after OVX and inhibitor-129-5p treatment (mean ± SD, ∗∗∗P < 0.001). (C) Expression of OCN in calvarial tissues of C57BL/6 mice after OVX and inhibitor-129-5p treatment, as detected by immunohistochemical staining. Scale bar: 50 µm. (D) Quantification of relative integrated optical density (IOD) values of OCN immunostaining using Image-Pro Plus 6.0 software (mean ± SD, ∗∗∗P < 0.001). (E) Expression of OXTERIX in calvarial tissues of C57BL/6 mice after OVX and inhibitor-129-5p treatment, as detected by immunohistochemical staining. Scale bar: 50 µm. (F) Quantification of relative IOD values of OXTERIX immunostaining using Image-Pro Plus 6.0 software (mean ± SD, ∗∗∗P < 0.001). (G) Expression of RUNX2 in calvarial tissues of C57BL/6 mice after OVX and inhibitor-129-5p treatment, as detected by immunohistochemical staining. Scale bar: 50 µm. (H) Quantification of relative IOD values of RUNX2 immunostaining using Image-Pro Plus 6.0 software (mean ± SD, ∗∗∗P < 0.001).

Article Snippet: Sections (5 μm in thickness) were dewaxed, immersed in the distilled water, blocked in 5% goat serum (CWBIO, CW0130) in PBS, and then incubated overnight at 4◦C with primary antibodies against OCN (Rabbit pAb, 1:200, Santa Cruz Biotech, sc365797, Dallas, TX, United States), OSTERIX (Rabbit pAb, 1:50; Proteintech, 12593-1-AP), and TCF4 (Rabbit pAb, 1:100, Proteintech, 22337-1-AP), respectively.

Techniques: Transfection, Control, Expressing, Immunohistochemical staining, Staining, Immunostaining, Software

Journal: Frontiers in cell and developmental biology

Article Title: miR-129-5p Inhibits Bone Formation Through TCF4.

doi: 10.3389/fcell.2020.600641

Figure Lengend Snippet: FIGURE 7 | miR-129-5p and osteogenic transcript factor expression levels of OVX mice after inhibitor-129-5p treatment. (A) miR-129-5p level in calvarial tissue of C57BL/6 mice after OVX and inhibitor-129-5p treatment, as detected by reverse transcriptase-polymerase chain reaction (RT-PCR; mean ± SD, ∗∗P < 0.01, ∗∗∗P < 0.001). BL (baseline): sacrifice before RNA treatment. Sham: sham OVX operation group. OVX: OVX group. Mock: transfection reagent control group. inhibit-NC: inhibitor-NC–treated group. inhibit-129: inhibitor-129-5p–treated group. (B–F) Ocn, Osterix, Tcf4, Tcf7, and Lef1 expression levels in calvarial tissue of C57BL/6 mice after OVX and inhibitor-129-5p treatment, as detected by RT-PCR (mean ± SD, ∗∗P < 0.01, ∗∗∗P < 0.001).

Article Snippet: Sections (5 μm in thickness) were dewaxed, immersed in the distilled water, blocked in 5% goat serum (CWBIO, CW0130) in PBS, and then incubated overnight at 4◦C with primary antibodies against OCN (Rabbit pAb, 1:200, Santa Cruz Biotech, sc365797, Dallas, TX, United States), OSTERIX (Rabbit pAb, 1:50; Proteintech, 12593-1-AP), and TCF4 (Rabbit pAb, 1:100, Proteintech, 22337-1-AP), respectively.

Techniques: Expressing, Reverse Transcription, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Transfection, Control

Journal: Stem Cells International

Article Title: The Histone Demethylase KDM3B Promotes Osteo-/Odontogenic Differentiation, Cell Proliferation, and Migration Potential of Stem Cells from the Apical Papilla

doi: 10.1155/2020/8881021

Figure Lengend Snippet: KDM3B enhanced the osteo-/odontogenic differentiation potential of SCAPs. (a) The knockdown efficiency of KDM3B in SCAPs was tested by western blot. (b) KDM3B knockdown significantly depressed the ALP activity in SCAPs. (c) The Alizarin red staining and (d) the quantitative calcium analysis showed that KDM3B knockdown reduced the mineralization capacity of SCAPs compared with the control group. (e, f) Real-time RT-PCR analysis confirmed that KDM3B knockdown reduced the expression of (e) RUNX2 and (f) OSX in SCAPs. (g) Western blot analysis showed the expression of RUNX2 and OSX in the KDM3B knockdown group and the control group. Histone H3 served as an internal control. (h) Western blot analysis revealed that the expression of DSPP and OCN was decreased after KDM3B was knocked down. Histone H3 served as an internal control. (i) The KDM3B overexpression was tested by western blot. (j) KDM3B overexpression significantly enhanced the ALP activity in SCAPs. (k, l) The results of (k) the Alizarin red staining and (l) the quantitative calcium analysis revealed that KDM3B overexpression enhanced the mineralization capacity of SCAPs compared with the control group. (m, n) Real-time RT-PCR analysis revealed that KDM3B overexpression increased the expression of (m) RUNX2 and (n) OSX in SCAPs. (o) Western blot analysis showed the expression of RUNX2 and OSX in the KDM3B overexpression group and the control group. Histone H3 served as an internal control. (p) Western blot analysis revealed that the expression of OCN and DSPP was enhanced after KDM3B was overexpressed. Histone H3 served as an internal control. Statistical significance was determined using Student's t -test. All error bars represent SD ( n = 3). P < 0.05. ∗∗ P ≤ 0.01.

Article Snippet: The primary antibodies used in the study are as follows: KDM3B antibody (cat no. 19915-1-AP, Rabbit Polyclonal, USA), RUNX2 antibody (cat no. ab76956, Abcam), OSX antibody (cat no. ab209484, Abcam), OCN antibody (cat no. bs-4917R, Bioss, China), DSPP antibody (cat no. bs10316R, Bioss, China), and histone H3 antibody (cat no. SC-56616, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Western Blot, Activity Assay, Staining, Quantitative RT-PCR, Expressing, Over Expression

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Bioinspired Protein/Peptide Loaded 3D Printed PLGA Scaffold Promotes Bone Regeneration

doi: 10.3389/fbioe.2022.832727

Figure Lengend Snippet: The immumohistochemical staining for detecting the expression of Runx2, OCN and COL-1 at 4 (A) and 12 (B) weeks (400 ×).

Article Snippet: Additionally, the expression of osteogenic factors including runt-related transcription factor 2 (Runx2) (Ab-AF5186,Affinity), osteocalcin (OCN) (OM266707, OmnimAbs)and Collagen I antibody (bs-10423R,Bioss) were analyzed through immunohistochemistry.

Techniques: Staining, Expressing