Journal: Frontiers in Molecular Neuroscience

Article Title: Ptbp2 re-expression rescues axon growth defects in Smn-deficient motoneurons

doi: 10.3389/fnmol.2024.1393779

Figure Lengend Snippet: Ptbp2 interacts with Smn in motoneurons. (A) Representative images of Smn-Ptbp2 PLA signal in cultured motoneurons at DIV 6 using anti-Smn and anti-Ptbp2 antibodies. Motoneuron morphology was visualized with anti-Tubb3 antibody. Scale bars, 10 and 5 μm (magnified areas). (B) Co-immunoprecipitation of Smn by anti-Ptbp2 from motoneuron lysate pre-treated with RNase A as indicated. (C) Representative images showing Smn immunofluorescence and Hnrnpr FISH in cultured motoneurons at DIV 6. Arrowheads indicate colocalization of Smn and Hnrnpr in granules. Scale bars, 10 and 2 μm (magnified areas). (D) Fluorescence intensity profiles of Smn and Hnrnpr at the location indicated by arrow 4 in (C) . (E) Representative images showing Ptbp2 and Smn immunofluorescence and Hnrnpr FISH in cultured motoneurons at DIV 6. Scale bars, 10 and 2 μm (magnified areas). (F) Fluorescence intensity profiles of Ptbp2, Smn and Hnrnpr at the location indicated by a line in (E) .

Article Snippet: Cells were fixed again for 10 min at room temperature in PLP, washed with DPBS, and stained with FITC-conjugated anti-Tubb3 antibody (130-131-158, Miltenyi Biotec).

Techniques: Cell Culture, Immunoprecipitation, Immunofluorescence, Fluorescence

Journal: Scientific Reports

Article Title: The Role of Transthyretin in Oligodendrocyte Development

doi: 10.1038/s41598-020-60699-8

Figure Lengend Snippet: TTR influences OPC differentiation. Representative images of neurospheres in 3D culture, isolated from SVZ derived NSCs of P21 ( A ) wild type and ( B ) TTR null mice. Scale bar 5 mm. ( C ) Quantitation of the potency of the NSC colonies isolated from the SVZ of P21 mice by neural colony-forming cell assays. There was a significant increase in the number of colonies with an average diameter ≥ 2 mm that were isolated from the SVZ of P21 TTR null mice when compared with wild type at the same age. Increases corresponded with a significant decrease in the number of colonies with an average diameter < 2 mm that were isolated from TTR null mouse SVZ compared to wild type. The colonies ≥ 2 mm in diameter are “NSC derived” and have self-renewal and multi-potential capabilities. Colonies < 2 mm diameter are “progenitor derived”. All data were expressed as the mean ± SEM. Statistical comparisons were performed using a one-way ANOVA with post hoc analysis using Sidak’s test as appropriate (data derived from n = 4 independent experiments per genotype and P < 0.05 was considered statistically significant). ( D ) Characterization of NSCs isolated from the SVZ of P21 TTR null mice and differentiated into the three neural lineages: oligodendrocytes, astrocytes and neurons. Nuclei were stained with DAPI; oligodendrocytes were stained with anti-Olig2; neurons were stained with anti-β-III-tubulin; astrocytes were stained with anti-GFAP. Lack of TTR promotes NSCs to differentiate into glial precursor cells. Differentiation assay for NSCs isolated from the SVZ of P21 TTR null mice showed a greater proportion of cells differentiating into a glial lineage, whereas the equivalent cells from wild type mice had a greater proportion differentiating into a neuronal lineage. All data were expressed as the mean ± SEM. Statistical comparisons were performed using a one-way ANOVA with post hoc analysis using Sidak’s test as appropriate (n = 3 independent experiments and P < 0.05 was considered statistically significant). (Modified from ).

Article Snippet: Cell pellets were collected and incubated with primary antibodies diluted in blocking solution: mouse anti-Olig2 (1:200; Merck Millipore, MAPN50), mouse anti-beta III tubulin (1:150; Merck Millipore, MAB1637), mouse anti-GFAP (1:200; Merck Millipore, MAB360), rabbit anti-TTR (1:300;; ABBIOTC, 250892) and incubated overnight at 4 °C.

Techniques: Isolation, Derivative Assay, Quantitation Assay, Staining, Differentiation Assay, Modification

Journal: Communications Biology

Article Title: Precise CAG repeat contraction in a Huntington’s Disease mouse model is enabled by gene editing with SpCas9-NG

doi: 10.1038/s42003-021-02304-w

Figure Lengend Snippet: a The procedure of neuronal differentiation of ES cells. For in vitro differentiation assay, we used ES clones transfected with prototype SpCas9-NG (not published), not ES clones described in Fig. . b Immunostaining of differentiated embryoid bodies. White arrows in the high magnified image of R6/2 clone #2 indicate HTT aggregates. All scale bars indicate 100 µm: the EB in genome-edited #2 (top) was almost twice larger than the others. c The differentiation scores based on a percentage of the β III tubulin staining in the circumference of embryoid bodies; 1: 0–25%; 2: 25–50%; 3: 50–75%; 4: 75–100%. Statistical analysis was performed using a two-tailed Mann–Whitney U-test ( p -value = 0.012).

Article Snippet: After three washes with PBS, the cells were incubated with Alexa Fluor 555 conjugated goat anti-mouse IgG (A21422, Thermo Fisher Scientific; 1:4000) at room temperature for 2 h. The cells were then incubated with a mouse anti-β III tubulin mouse antibody (T8660, Merck; 1:1000) at 4 °C overnight.

Techniques: In Vitro, Differentiation Assay, Clone Assay, Transfection, Immunostaining, Staining, Two Tailed Test, MANN-WHITNEY

Journal: Anatomical record (Hoboken, N.J. : 2007)

Article Title: Comparative morphology and histology of the nasal fossa in four mammals: gray squirrel, bobcat, coyote and white-tailed deer

doi: 10.1002/ar.23352

Figure Lengend Snippet: Epithelial lining of the bobcat nasal fossa. Sagittal view of segmented MRI scans of the bobcat nasal airway showing five coronal sections (b–f) selected for morphological and histological representation (A). The five coronal sections along the rostrocaudal axis illustrate the maxilloturbinal (B–C), nasomaxillary (D) and ethmoturbinal (E–F) regions of the bobcat nasal fossa. Magnified views show the nasoturbinal (G), maxilloturbinal (H, J) and ethmoturbinal (I) covered with nonsensory epithelium and packed with Alcian blue labeled goblet cells at rostral regions of the nasal fossa. Olfactory epithelium covered the nasoturbinal (K), septum (L), and ethmoturbinal (L) in more caudal regions and had the characteristic epithelial thickness and Bowman’s glands in the lamina propria (M) and labeled with β-tubulin III antibody (N–O). nt = nasoturbinal; mt = maxilloturbinal; s = septum; et = ethmoturbinals; ob = olfactory bulb; oe = olfactory epithelium; bg = Bowman’s gland; ne = nonsensory epithelium. Scale bar: B–F = 2 mm; G–L, N = 100 µm; and M, O = 50 µm.

Article Snippet: To further visualize the OE, we conducted immunohistochemistry with a mouse monoclonal anti-neuronal class III β-tubulin antibody (TMJ1, Babco) as has been used to label olfactory neurons in rodents ( Roskams et al., 1998 ), cats (Lishcka et al., 2008) and humans ( Ronnett et al., 2003 ), suggesting reactivity of this antibody in diverse mammalian samples.

Techniques: Labeling

Journal: Anatomical record (Hoboken, N.J. : 2007)

Article Title: Comparative morphology and histology of the nasal fossa in four mammals: gray squirrel, bobcat, coyote and white-tailed deer

doi: 10.1002/ar.23352

Figure Lengend Snippet: Epithelial lining of the coyote nasal fossa. Sagittal view of segmented MRI scans of the coyote nasal airway showing five coronal sections (b–f) selected for morphological and histological representation (A). The five coronal sections along the rostrocaudal axis illustrate the maxilloturbinal (B–C), nasomaxillary (D) and ethmoturbinals (E–F) regions of the coyote nasal fossa. Nonsensory epithelium covered the nasoturbinal (G), maxilloturbinal (H–K), and ethmoturbinal (L) at the rostral regions. Thick olfactory epithelium (L), clearly labeled with β-tubulin antibody (N–O), covered the septum, nasoturbinal, and ethmoturbinal at the caudal regions of the nasal fossa. nt = nasoturbinal; mt = maxilloturbinal; s = septum; et = ethmoturbinal; fs = frontal sinus; ob = olfactory bulb; oe = olfactory epithelium; ne = nonsensory epithelium. Scale bar: B–F = 2 mm; G–L, N = 100 µm; and M, O = 50 µm.

Article Snippet: To further visualize the OE, we conducted immunohistochemistry with a mouse monoclonal anti-neuronal class III β-tubulin antibody (TMJ1, Babco) as has been used to label olfactory neurons in rodents ( Roskams et al., 1998 ), cats (Lishcka et al., 2008) and humans ( Ronnett et al., 2003 ), suggesting reactivity of this antibody in diverse mammalian samples.

Techniques: Labeling