Journal: Laboratory Investigation; a Journal of Technical Methods and Pathology

Article Title: A murine mesenchymal stem cell model for initiating events in osteosarcomagenesis points to CDK4/CDK6 inhibition as a therapeutic target

doi: 10.1038/s41374-021-00709-z

Figure Lengend Snippet: Murine MSCs ( A ) that have spontaneously transformed after long-term culture (B6_4, wildtype (WT) Ink4ab; B6_7, deletion (DEL) of Ink4ab; B6_10, wildtype (WT) Ink4ab) or B MSCs from Ink4ab −/− mice after transformation were treated with palbociclib for 72 h after which relative cell viability and IC 50 values were determined. Wildtype MSCs were derived from FVB mice. Data points represent the mean of three experiments performed ± standard deviation.

Article Snippet: For 2D cultures, human osteosarcoma cell lines or murine MSCs were seeded (between 3000 and 6000 cells per well) into 96-well plates and after 24 h treated with PBS or Palbociclib (dissolved in PBS, PD-0332991, Selleckchemicals, Houston, TX, USA) in concentrations ranging from 0.01 μM to 100 μM.

Techniques: Transformation Assay, Derivative Assay, Standard Deviation

Journal: Laboratory Investigation; a Journal of Technical Methods and Pathology

Article Title: A murine mesenchymal stem cell model for initiating events in osteosarcomagenesis points to CDK4/CDK6 inhibition as a therapeutic target

doi: 10.1038/s41374-021-00709-z

Figure Lengend Snippet: A Osteosarcoma cell lines were treated with palbociclib for 72 h after which relative cell viability and IC 50 values were determined. Cell lines MG63 and 143BHOS (green) have loss of p16, whereas other OS cell lines have intact p16 (blue). Data points represent the mean of three experiments performed ± standard deviation. B Western blot showing expression levels of Rb in osteosarcoma cell lines. GAPDH was used as a loading control. C p16 protein expression status of each osteosarcoma cell line as published was correlated with IC 50 values from the current study. n.s. not statistically significant. D CDK4 and CDK6 RNA expression levels as published elsewhere of each cell line did not correlate with IC 50 values from the current study. Each dot represents one cell line.

Article Snippet: For 2D cultures, human osteosarcoma cell lines or murine MSCs were seeded (between 3000 and 6000 cells per well) into 96-well plates and after 24 h treated with PBS or Palbociclib (dissolved in PBS, PD-0332991, Selleckchemicals, Houston, TX, USA) in concentrations ranging from 0.01 μM to 100 μM.

Techniques: Standard Deviation, Western Blot, Expressing, Control, RNA Expression

Journal: Laboratory Investigation; a Journal of Technical Methods and Pathology

Article Title: A murine mesenchymal stem cell model for initiating events in osteosarcomagenesis points to CDK4/CDK6 inhibition as a therapeutic target

doi: 10.1038/s41374-021-00709-z

Figure Lengend Snippet: A Hematoxylin and eosin staining of osteosarcoma MCTS treated with palbociclib for 72 h. Scalebar represents 20 µm. Inset shows SATB2 staining. B Relative cell viability and IC 50 values were determined after treatment with palbociclib. C Cleaved caspase 3 and Ki67 staining and quantification of 3D cultured MCTS of OS cell lines treated with 10 µM palbociclib (Palb) or PBS. Scale bar represents 20 µm. n.s. not statistically significant, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: For 2D cultures, human osteosarcoma cell lines or murine MSCs were seeded (between 3000 and 6000 cells per well) into 96-well plates and after 24 h treated with PBS or Palbociclib (dissolved in PBS, PD-0332991, Selleckchemicals, Houston, TX, USA) in concentrations ranging from 0.01 μM to 100 μM.

Techniques: Staining, Cell Culture

Journal: The Journal of Neuroscience

Article Title: Memory-Related Synaptic Plasticity Is Sexually Dimorphic in Rodent Hippocampus

doi: 10.1523/JNEUROSCI.0801-18.2018

Figure Lengend Snippet: ERα is present at higher concentrations in female than in male CA1 synapses. A, Top left, Deconvolved two-photon microscopic images of immunofluorescent labeling were used to construct a 3D montage of the CA1 sample field (shown); one can see that a subpopulation of PSD-95-IR contacts (green) also contains ERα immunoreactivity (double labeling appears yellow). Scale bar, 2 μm. Top right, Image of a single double-labeled PSD shows the spatial relationship of areas occupied by PSD-95 (green) and ERα (red) immunoreactivities and the extent of overlap (merge, yellow). Scale bar, 0.1 μm. Bottom, Image shows the montage from the same z-stack illustrated in the top left but with the top of that panel rotated away from the viewer to show double labeling of the same puncta (arrows) from a different 3D viewpoint. B, The density frequency distribution for ERα-IR (colocalized with PSD-95) shows a greater rightward skew in females relative to males (p < 0.0001, F(19,646) = 17.28; males, n = 12; female, n = 24). C, Bar graph shows the percentage of double-labeled synapses with high concentrations of ERα immunolabeling (density units of ≥90) normalized to the mean male value shows that there were far more dense ERα-IR synapses in females than in males (***p = 0.0001, t(34) = 4.40). D, E, Density frequency distributions for all synapse-sized clusters of ERβ (D) and GPER1 (E) immunoreactivities colocalized with PSD-95 in the CA1 SR sample field (n = 12/group, females in diestrus). For ERβ-IR (D), there was a significant interaction between sex and immunolabeling density (p = 0.0005, F(21,462) = 2.4) because of a slightly greater proportion of synapses with low-density ERβ-IR (density units 83–88) in males than in females. There was no effect of sex on the numbers of densely ERβ-IR synapses (density units, ≥90; p = 0.12; t(22) = 1.6). For GPER1-IR, there were no group differences (GPER1: p = 0.89; F(20,440) = 0.33). F, Deconvolved epifluorescence images shows that ERβ and GPER1 (red) are both localized to synapse-sized puncta in CA1 SR and that some of those are colocalized with PSD-95 (green; doubles appear yellow). Scale bar, 2 μm.

Article Snippet: Stacks were deconvolved using AutoQuant version 2.2.1 (Media Cybernetics) and constructed into a 3D montage using Imaris (Bitplane; Lauterborn et al., 2016 ).

Techniques: Labeling, Construct, Immunolabeling