Journal: Development (Cambridge, England)

Article Title: The evolution of basal progenitors in the developing non-mammalian brain

doi: 10.1242/dev.127100

Figure Lengend Snippet: Distinct characteristics of basal mitotic cells and Tbr2 + cells in the developing chicken pallium. (A,B) Distribution of phosphorylated histone H3 (PH3) + cells in the developing mouse (A, E15.5) and chick (B, E8) telencephalon. Arrows indicate mitotic cells on the basal side of the pallia. (C,D) The proportion of the PH3 + cells that were also Pax6 + , Sox2 + or EdU + on the basal side of the chicken pallium. Arrows, indicate PH3 + /Pax6 + cells (C, magnified by insets) and Sox2 + /EdU + cells (D). Mean±s.d. (E,F) Distribution of Tbr2 + cells in the developing mouse (E, E12.5) and chick (F, E8) telencephalon. (G-J) Labeling of Tbr2 + cells by electroporation (EP) with a GFP expression vector. Electroporation was performed at E4 and the chick embryos were collected at E7. Arrowheads (I,J), GFP + /Tbr2 + cells. (K-P) Characterization of Tbr2 + cells in the developing chick pallium. The sections were stained with anti-Pax6 (K), anti-BrdU (L,M), anti-PH3 (N) or anti-NeuN (O,P) antibodies and an anti-Tbr2 antibody. Arrowhead (N), a PH3 + cell that does not overlap with Tbr2. DP, dorsal pallium; DVR, dorsal ventricular ridge; MP, medial pallium; NCx, neocortex; GE, ganglionic eminence; SVZ, subventricular zone; VZ, ventricular zone. Scale bars: 200 µm in B (for A,B), E (for E,F), G; 50 µm in C (for C,D), K (for K-P); 10 µm in I (for I,J).

Article Snippet: Manipulations of gene function were performed with expression vectors including pTK38_mCherry-LGN-C (Addgene, ID46346), pCMS-EGFP-Cdk4, pCMS-EGFP, pDSV-RFPnls-CyclinD, pDSV-RFPnls, pCAG-Eomesodermin-MycDDK (Tbr2; Origene ORF clone MR210179 was subcloned into the Hin dIII/ Bgl I sites of pCAGRB), pCAG-Cre.

Techniques: Labeling, Electroporation, Expressing, Plasmid Preparation, Staining

Journal: Development (Cambridge, England)

Article Title: The evolution of basal progenitors in the developing non-mammalian brain

doi: 10.1242/dev.127100

Figure Lengend Snippet: Experimental amplification of basal mitotic cells and Tbr2 + cells in the developing chick pallium. (A) Diagram of the LGN and LGN-C proteins. C-terminal TPR repeats were replaced with mCherry in LGN-C. (B,C) LGN-C increased the number of PH3 + cells on the basal side of the chicken pallium at the expense of APs (arrowheads in B). (D-G) Overexpression of Cdk4 and cyclin D1 (collectively 4D) in the developing chick pallium. (D) The structures of the 4D vectors. Co-expression of GFP and RFP in the same cells was induced by electroporation of 4D vectors. (E-G) Overexpression of 4D vectors increased the number of Tbr2 + cells. (H-J) Overexpression of Tbr2 in the developing chicken pallium. (H) Structure of the expression vectors containing GFP and Tbr2. GFP and Tbr2 were co-expressed in the same cells upon electroporation. (I,J) Overexpression of Tbr2 suppressed Sox2 expression in APs but did not change the number of APs and BPs. All bar charts show mean±s.d. ** P <0.01 (Student's t -test); n.s., not significant. Scale bars: 50 µm in B,E (for E,F); 25 µm in D; 15 µm in H; 10 µm in I.

Article Snippet: Manipulations of gene function were performed with expression vectors including pTK38_mCherry-LGN-C (Addgene, ID46346), pCMS-EGFP-Cdk4, pCMS-EGFP, pDSV-RFPnls-CyclinD, pDSV-RFPnls, pCAG-Eomesodermin-MycDDK (Tbr2; Origene ORF clone MR210179 was subcloned into the Hin dIII/ Bgl I sites of pCAGRB), pCAG-Cre.

Techniques: Amplification, Over Expression, Expressing, Electroporation

Journal: Development (Cambridge, England)

Article Title: The evolution of basal progenitors in the developing non-mammalian brain

doi: 10.1242/dev.127100

Figure Lengend Snippet: A possible scenario of changes in pallial progenitor subtypes during amniote evolution. In mammals, apical progenitors (APs) proliferate and generate basal progenitors (BPs), which include Tbr2 + intermediate progenitors (IPs) and other types of BPs [such as basal radial glial cells (bRGs)]. In birds, both BPs and Tbr2 + cells exist, but these cells are distinct populations. We propose that a few subapical mitotic cells that have already evolved in ancestral amniotes are at the origin of BPs, although de novo BPs appeared in several taxa independently.

Article Snippet: Manipulations of gene function were performed with expression vectors including pTK38_mCherry-LGN-C (Addgene, ID46346), pCMS-EGFP-Cdk4, pCMS-EGFP, pDSV-RFPnls-CyclinD, pDSV-RFPnls, pCAG-Eomesodermin-MycDDK (Tbr2; Origene ORF clone MR210179 was subcloned into the Hin dIII/ Bgl I sites of pCAGRB), pCAG-Cre.

Techniques:

Journal: Cancer Science

Article Title: NKG2D defines tumor‐reacting effector CD8 + T cells within tumor microenvironment

doi: 10.1111/cas.15050

Figure Lengend Snippet: Expression of transcription factors in tumor antigen‐specific CD8 + T cells in the immune control effector phase. B16OVA‐Luc2 cells (10 5 cells/mouse) were used to sc inoculate vaccinated C57BL/6 mice. B16OVA‐Luc2 cells (10 5 cells/mouse) were used to sc inoculate vaccinated C57BL/6 mice. At the indicated time‐points, TIL were collected and subjected to flow cytometry (Day 0 = tumor inoculation). Representative histograms (left) and summaries (right) of Eomes (A) and T‐bet (B) expression in OVATet + T cells are shown. C, Representative contour plots (left) and summaries (middle) of TOX expression (TOX hi ) on OVATet + T cells in TIL are shown. The percentage of TOX hi OVATet + T cells isolated from the indicated sources is shown (right). Data are presented as the mean ± SEM

Article Snippet: For intracellular staining, cells were fixed and permeabilized in accordance with the manufacturer's instructions (FoxP3/Transcription factor staining buffer set, eBioscence) before being stained with fluorophore‐labeled antibodies: anti‐mouse eomesodermin (Eomes, clone REA116, MACS Miltenyi Biotec), T‐bet (4B10, BioLegend), and TOX (REA473, Miltenyi Biotec).

Techniques: Expressing, Control, Flow Cytometry, Isolation

Journal: Pediatric Diabetes

Article Title: Integrating Physical Activity Strategies to Lower Hyperglycaemia in Structured Education Programmes for Children and Young People with Type 1 Diabetes Improves Glycaemic Control without Augmenting the Risk of Hypoglycaemia

doi: 10.1155/2023/2519368

Figure Lengend Snippet: Outcome measures by activity groups.

Article Snippet: Data over a 3 month period (starting 3 months after education) on TIR, TAR, TAR2 (>13.9 mmol/L), time below range (TBR), and TBR2 (<3.0 mmol/L) were collected from Dexcom Clarity and Libre View (downloaded at 6-month clinical review).

Techniques: Activity Assay

Journal: Pediatric Diabetes

Article Title: Integrating Physical Activity Strategies to Lower Hyperglycaemia in Structured Education Programmes for Children and Young People with Type 1 Diabetes Improves Glycaemic Control without Augmenting the Risk of Hypoglycaemia

doi: 10.1155/2023/2519368

Figure Lengend Snippet: Pairwise comparisons of activity groups (low, mild, and moderate). All significant differences between groups are indicated with p values: (a) HbA1c (mmol/mol), (b) time in range (TIR, 3.9–10.0 mmol/L), (c) time above range (TAR, >10.0 mmol/L), (d) time above range 2 (TAR2, >13.9 mmol/L), (e) time below range (TBR2, <3.9 mmol/L), and (f) time below range 2 (TBR2, <3.0 mmol/L).

Article Snippet: Data over a 3 month period (starting 3 months after education) on TIR, TAR, TAR2 (>13.9 mmol/L), time below range (TBR), and TBR2 (<3.0 mmol/L) were collected from Dexcom Clarity and Libre View (downloaded at 6-month clinical review).

Techniques: Activity Assay

Journal: bioRxiv

Article Title: Lmx1a is a master regulator of the cortical hem

doi: 10.1101/2022.10.25.513532

Figure Lengend Snippet: (A) Normalized read counts for Tbr2 from the RNAseq experiment . (B-D) Arrowheads indicate Tbr2+ cells in the CH (CR cells, ) (B), which exhibited lower intensity of Tbr2 immunofluorescence in e13 Lmx1a-/- embryos compared to controls (B, C). In contrast, Tbr2+ cells in the hippocampal primordium (hippocampal intermediate progenitors, ) (arrow in B) had similar Tbr2 immunofluorescence intensity in control and Lmx1a-/- embryos (B, D), suggesting that Lmx1a loss reduces Tbr2 expression specifically in cells arising in the CH. (Dashed lines demarcate the CH boundaries, identified by Lmx1a immunostaining of adjacent sections, as described in the Materials and Methods). (E-F) Reduced HF length (dashed line) and aberrant accumulation of CR cells at the FDJ surface (arrowheads) in Lmx1a/Tbr2 double-gene heterozygotes ( Lmx1a+/-;Lmx1a-Cre;Tbr2 + F mice – Lmx1a+/- mice in which one copy of Tbr2 was deleted specifically in the CH), but not singlegene heterozygotes, compared to wild type controls at P3. ***p<0.001, n=3 mice per genotype. (G, H) Normal cell cycle exit of progenitors in the CH of Lmx1a/Tbr2 double heterozygous embryos. 24 h after BrdU injection, progenitors that exited the cell cycle were BrdU+/Ki67-(green, arrows); progenitors that re-entered the cell cycle were BrdU+/Ki67+ (yellow, arrowheads). n=3-4 mice per genotype. Scale bars: 50 μm (B, G); 200 μm (E).

Article Snippet: Mice used in this study include Lmx1a null ( Lmx1a drJ , Jackson Laboratory strain #000636) ( ; ; ), Wnt3a null (Jackson Laboratory strain #004581) , Tbr2 floxed (Jackson Laboratory strain #017293) , and Lmx1a-Cre (BAC transgenic mice that express a GFP-tagged Cre protein under the control of the Lmx1a regulatory elements, referred to as Lmx1a-Cre throughout the paper) ( ).

Techniques: Immunofluorescence, Control, Expressing, Immunostaining, Injection

Journal: bioRxiv

Article Title: Lmx1a is a master regulator of the cortical hem

doi: 10.1101/2022.10.25.513532

Figure Lengend Snippet: Lmx1a promotes expression (solid arrow) of secreted Wnt3a, which non-autonomously (dashed arrow) regulates the development of the hippocampal DG (proliferation in the DNe, the transhilar scaffold, and the number and input resistance of DG neurons). Lmx1a represses the expression of the cortical selector gene Lhx2 (solid bar), segregating the CH from the adjacent hippocampal field. Lmx1a positively regulates the expression of a wide range of CH markers. It also promotes the exit of CH progenitors from the cell cycle and their differentiation into CR cells by activating the expression of Cdkn1a , and promotes migration of CR cells via Tbr2 , which is necessary for the HF and transhilar scaffold formation.

Article Snippet: Mice used in this study include Lmx1a null ( Lmx1a drJ , Jackson Laboratory strain #000636) ( ; ; ), Wnt3a null (Jackson Laboratory strain #004581) , Tbr2 floxed (Jackson Laboratory strain #017293) , and Lmx1a-Cre (BAC transgenic mice that express a GFP-tagged Cre protein under the control of the Lmx1a regulatory elements, referred to as Lmx1a-Cre throughout the paper) ( ).

Techniques: Expressing, Migration

Journal: bioRxiv

Article Title: Assembly of neuron- and radial glial cell-derived extracellular matrix molecules promotes radial migration of developing cortical neurons

doi: 10.1101/2022.09.03.506497

Figure Lengend Snippet: Histochemical analysis of radial glial cells, intermediate progenitor cells, and the morphology of radial fibers. ( A ) Immunostaining of Pax6-positive radial glial cells and Tbr2-positive intermediate progenitor cells in the VZ of WT and DKO cerebral cortices at E16. Bar graphs show the numbers of Pax6- and Tbr2-positive cells per 150 × 150 μm 2 . N = 5 mice per group. Mean ± SD; ns > 0.05; the Student’s t -test. ( B ) Immunostaining of nestin-positive radial fibers in WT and DKO cerebral cortices at E16. ( C ) High magnification images of mCherry-positive bipolar neurons and radial fibers in the WT and DKO at E16, 2 days after in utero labeling. Bipolar neurons in the SP/IZ-attached radial fiber, irrespective of genotypes. Scale bars represent 20 μm (A), 50 μm (B), and 5 μm (C).

Article Snippet: Sections were incubated at 4°C overnight with biotinylated HA-binding protein (b-HABP) (1:400, Hokudo), anti-NCAN (sheep, 1:400, R&D), anti-TNC (rat, 1:400, R&D), anti-Pax6 (rabbit, 1:400, Invitrogen), anti-Tbr2 (mouse, 1:400, Wako), anti-Nestin (mouse, 1:400, Millipore), and Alexa Fluor 488-conjugated anti GFP (rabbit, 1:1000, Invitrogen).

Techniques: Immunostaining, In Utero, Labeling