Journal: Development (Cambridge, England)

Article Title: Integrins are required for tissue organization and restriction of neurogenesis in regenerating planarians

doi: 10.1242/dev.139774

Figure Lengend Snippet: Impaired cell migration after β1-int RNAi. (A) Sublethal γ-irradiation assay with 12.5 Gy. See text for details. (B,C) Double FISH against smedwi-1 (red) and NB.32.1g (green) reveals a significant net shift of neoblasts and NB.32.1g + progenitor cells (green arrows) towards the posterior in β1-int RNAi fragments 10 dpa. Fragments were segmented into four zones based on their distance from the anterior epidermis: Zone 1, up to 50 μm; Zone 2, 50-100 μm; Zone 3, 100-150 μm; Zone 4, >150 μm. The average percentage (+s.d.) smedwi-1 + and NB.32.1g + cells out of the total cell number, of eight animals from each condition in every zone are plotted in C. (D) Cell transplantation assay. See text for details. (E,F) FISH against smedwi-1 (green) reveals a significant decrease in neoblast dispersion for all β1-int RNAi combinations. Average distances between anterior- and posterior-most neoblast of seven animals are plotted in F in boxplots. The dark horizontal lines within the boxes represent the median for each condition, with the box representing the 25th and 75th percentiles and the whiskers indicating minimum and maximum values. The gray area marks the average distance (permanent line: ∼193.2 µm; n =7 animals) of transplanted ctrl neoblasts in uncut ctrl animals with s.d. (dashed lines: ∼±81.8 µm; n =7 animals). The composite image in E was generated using the customized tile scan function of Zeiss AxioVision software. Statistical significance (Student's t -test) is indicated (* P ≤0.05; ** P ≤0.01; *** P ≤0.001). DNA is blue (Hoechst). dpinj, days post injection; dpirr, days post irradiation; Gy, gray. Scale bars: 100 µm (B); 250 µm (E).

Article Snippet: Composite images in D were generated using the customized tile scan function of Zeiss AxioVision software.

Techniques: Migration, Irradiation, Transplantation Assay, Generated, Software, Injection

Journal: Development (Cambridge, England)

Article Title: Integrins are required for tissue organization and restriction of neurogenesis in regenerating planarians

doi: 10.1242/dev.139774

Figure Lengend Snippet: Impaired muscle and gut regeneration in β1-int RNAi planarians. (A) Anti-MYHC immunostaining (green) of sagittal sections of transversally amputated ctrl and β1-int RNAi trunk fragments at 20 dpa. Images show anterior and posterior regeneration sites. White arrows point to poorly structured muscle fibers; asterisks indicate gut lumen; white dashed line indicates approximate line of amputation. DS, digestive system; CNS, central nervous system; ant, anterior; post, posterior. (B-C″) Whole-mount immunostaining with an anti-PHOSPHOTYROSINE (P-TYR) antibody (green) indicate impaired branching of the gut (white arrows) in anterior and posterior blastemas of β1 - int RNAi trunk fragments at 20 dpa compared with ctrl RNAi fragments. Composite images were generated using the customized tile scan function of Zeiss AxioVision software. High magnification images of the anterior and posterior regeneration blastema of β1-int or ctrl RNAi fragments (B′,B″,C′,C″) are taken from the boxed areas as indicated. (D) Scheme for evaluation of gut branches. abl, anterior branch length; al, anterior length; pbl, posterior branch length; pl, posterior length. (E) Average number of secondary (2ary), tertiary (3ary) and quaternary (4ary) per anterior or posterior primary branch length of eight fragments are plotted. (F) Length of primary (1ary) gut branches in relation to anterior or posterior animal length respectively is plotted. Error bars represent s.d. of eight fragments for each RNAi condition. Statistical significance (Student's t -test) is indicated (* P ≤0.05; ** P ≤0.01; *** P ≤0.001; n.s. not significant). DNA is blue (Hoechst). Scale bars: 100 µm (A); 250 µm (B,C); 50 µm (B′,B″,C′,C″).

Article Snippet: Composite images in D were generated using the customized tile scan function of Zeiss AxioVision software.

Techniques: Immunostaining, Generated, Software

Journal: Development (Cambridge, England)

Article Title: Integrins are required for tissue organization and restriction of neurogenesis in regenerating planarians

doi: 10.1242/dev.139774

Figure Lengend Snippet: Ectosphere formation in anterior blastemas of β1-int RNAi planarians. (A,A′) FISH against pc2 (green) on ctrl and β1-int RNAi trunk fragments at 10 dpa. White box indicates magnified area shown in A′, from another z -position within the ectosphere. The composite image in A′ was generated using the customized tile scan function of Zeiss AxioVision software. (B,B′) Anti - SYNAPSIN immunostaining (green) on β1-int RNAi trunk fragment at 10 dpa. Orthogonal view shows three ectospheres in the anterior blastema at different positions (yellow arrowhead in xy view, white arrowhead in yz view, red arrowhead in xz view). White box indicates magnified area shown in B′. DNA (Hoechst) is blue. Scale bars: 150 µm (A); 100 µm (B).

Article Snippet: Composite images in D were generated using the customized tile scan function of Zeiss AxioVision software.

Techniques: Generated, Software, Immunostaining

Journal: Development (Cambridge, England)

Article Title: Integrins are required for tissue organization and restriction of neurogenesis in regenerating planarians

doi: 10.1242/dev.139774

Figure Lengend Snippet: Ectosphere formation depends on neoblasts and anterior cues. (A) Ectospheres visualized by Hoechst staining (gray) in untreated or irradiated (3 dpirr) β1-int RNAi planarians. Average areas with s.d. of at least seven animals per time point are displayed. (B) EdU-pulse chase (green) of sphere-forming cells 7 days post injection (dpinj) and 14 dpa. The red arrowhead points to an anti-H3P + mitotic cell. (C) FISH against smedwi-1 (green) in combination with immunostaining against H3P (red arrowhead) reveals neoblasts in the periphery of ectospheres (white arrowheads) in anterior regenerating β1-int RNAi fragments 10 dpa. (D) Ectospheres, visualized by FISH against pc2 (green) in regenerating trunk fragments of double RNAi planarians (ctrl;ctrl, β1-int;ctrl , β1-int;β-cat1 , β1-int;APC , β1-int;ndk ) at 15 or 20 dpa. White boxes highlight ectospheres in Hoechst (gray) channel. White arrowheads point to ectopic neural protrusions, described as brain primordium for APC RNAi animals . Red boxes in schemes illustrate areas of the images in the regeneration blastema of A-C. DNA (Hoechst) is blue. Composite images in D were generated using the customized tile scan function of Zeiss AxioVision software. Scale bars: 100 µm (A-C); 250 µm (D).

Article Snippet: Composite images in D were generated using the customized tile scan function of Zeiss AxioVision software.

Techniques: Staining, Irradiation, Pulse Chase, Injection, Immunostaining, Generated, Software