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Efficient retina formation requires suppression of both Activin and BMP signaling pathways in pluripotent cells.

Wong KA, Trembley M, Abd Wahab S, Viczian AS - Biol Open (2015)

Bottom Line: We determined the effect of these treatments on retina formation using the Animal Cap Transplant (ACT) assay; in which treated pluripotent cells were transplanted into the eye field of host embryos.We found that inhibition of Activin signaling, in the presence of BMP signaling inhibition, promotes efficient retinal specification in Xenopus tissue, mimicking the affect of adding Noggin alone.In whole embryos, we found that the eye field marker, rax, expanded when adding both dominant-negative Smad1 and Smad2, as did treating the cells with both dorsomorphin and SB431542.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Physiology, SUNY Upstate Medical University, Syracuse, NY 13210, USA The Center for Vision Research, SUNY Eye Institute, Upstate Medical University, Syracuse, NY 13210, USA.

No MeSH data available.


Related in: MedlinePlus

Follistatin is less efficient than Cerberus or Noggin at specifying retina.(A–C) YFP+ donor cells expressing Noggin, Cerberus, and Follistatin all contribute to the retina. However, Noggin- and Cerberus-treated cells formed retina in all animals while cells expressing Follistatin had a significantly lower retinal integration efficiency (D). On retinal sections, dashed white lines lie on outer and inner plexiform layers. Green, YFP donor cells; red, rod photoreceptor marker XAP2; blue, DAPI staining. YFP, 500 pg, n = 29; Nog, 20 pg, n = 65; Cerberus, 1.6 ng, n = 77; Follistatin, 1200 pg, n = 53; N = 3. Scale bars, 50 µm. Error bars  =  ±s.e.m.; ***p<0.001. Dashed red line on the graph marks Noggin/Cerberus treatment, highlighting the difference from Follistatin treatment.
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f07: Follistatin is less efficient than Cerberus or Noggin at specifying retina.(A–C) YFP+ donor cells expressing Noggin, Cerberus, and Follistatin all contribute to the retina. However, Noggin- and Cerberus-treated cells formed retina in all animals while cells expressing Follistatin had a significantly lower retinal integration efficiency (D). On retinal sections, dashed white lines lie on outer and inner plexiform layers. Green, YFP donor cells; red, rod photoreceptor marker XAP2; blue, DAPI staining. YFP, 500 pg, n = 29; Nog, 20 pg, n = 65; Cerberus, 1.6 ng, n = 77; Follistatin, 1200 pg, n = 53; N = 3. Scale bars, 50 µm. Error bars  =  ±s.e.m.; ***p<0.001. Dashed red line on the graph marks Noggin/Cerberus treatment, highlighting the difference from Follistatin treatment.

Mentions: If blocking both the BMP and Activin pathways are necessary to transform pluripotent tissue into retina, then adding a known antagonist of both pathways, like Cerberus (Piccolo et al., 1999), should generate retinal tissue as efficiently as Noggin. Using the ACT assay, we observed Cerberus-expressing pluripotent cells formed retina with a similar frequency as Noggin (Cerberus, 98±2%, n = 77; Noggin, 100%, n = 65, Fig. 7A,B). Conversely, if we used an antagonist that favored blocking the Activin pathway over the BMP pathway, like Follistatin (Thompson et al., 2005), we hypothesized that less of these treated animal caps to become eye tissue. Animal caps were isolated from embryos injected with an increasing concentration of Follistatin RNA (400 pg, 800 pg, 1200 pg, and 1600 pg) and the retinal integration efficiency was tested by ACT assay (supplementary material Fig. S3). Animal caps injected with 1600 pg did not survive, but those injected with lower concentrations formed retina in 21±10%, 41±11%, and 50±3% of animals, respectively. To be sure that the exogenous Follistatin can still function as a neural inducer, we scored the same sections for brain formation based on morphology and cellular location. The percentage of animals that contained transplanted cells found in the brain were comparable between Noggin- (80±6%; n = 80) and Follistatin-treated (74±10%; n = 60) cells. Thus, factors that repressed both BMP and Activin signals, such as Noggin and Cerberus, were more efficient retinal-inducers than Follistatin, an Activin-biased antagonist.


Efficient retina formation requires suppression of both Activin and BMP signaling pathways in pluripotent cells.

Wong KA, Trembley M, Abd Wahab S, Viczian AS - Biol Open (2015)

Follistatin is less efficient than Cerberus or Noggin at specifying retina.(A–C) YFP+ donor cells expressing Noggin, Cerberus, and Follistatin all contribute to the retina. However, Noggin- and Cerberus-treated cells formed retina in all animals while cells expressing Follistatin had a significantly lower retinal integration efficiency (D). On retinal sections, dashed white lines lie on outer and inner plexiform layers. Green, YFP donor cells; red, rod photoreceptor marker XAP2; blue, DAPI staining. YFP, 500 pg, n = 29; Nog, 20 pg, n = 65; Cerberus, 1.6 ng, n = 77; Follistatin, 1200 pg, n = 53; N = 3. Scale bars, 50 µm. Error bars  =  ±s.e.m.; ***p<0.001. Dashed red line on the graph marks Noggin/Cerberus treatment, highlighting the difference from Follistatin treatment.
© Copyright Policy - open-access
Related In: Results  -  Collection

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f07: Follistatin is less efficient than Cerberus or Noggin at specifying retina.(A–C) YFP+ donor cells expressing Noggin, Cerberus, and Follistatin all contribute to the retina. However, Noggin- and Cerberus-treated cells formed retina in all animals while cells expressing Follistatin had a significantly lower retinal integration efficiency (D). On retinal sections, dashed white lines lie on outer and inner plexiform layers. Green, YFP donor cells; red, rod photoreceptor marker XAP2; blue, DAPI staining. YFP, 500 pg, n = 29; Nog, 20 pg, n = 65; Cerberus, 1.6 ng, n = 77; Follistatin, 1200 pg, n = 53; N = 3. Scale bars, 50 µm. Error bars  =  ±s.e.m.; ***p<0.001. Dashed red line on the graph marks Noggin/Cerberus treatment, highlighting the difference from Follistatin treatment.
Mentions: If blocking both the BMP and Activin pathways are necessary to transform pluripotent tissue into retina, then adding a known antagonist of both pathways, like Cerberus (Piccolo et al., 1999), should generate retinal tissue as efficiently as Noggin. Using the ACT assay, we observed Cerberus-expressing pluripotent cells formed retina with a similar frequency as Noggin (Cerberus, 98±2%, n = 77; Noggin, 100%, n = 65, Fig. 7A,B). Conversely, if we used an antagonist that favored blocking the Activin pathway over the BMP pathway, like Follistatin (Thompson et al., 2005), we hypothesized that less of these treated animal caps to become eye tissue. Animal caps were isolated from embryos injected with an increasing concentration of Follistatin RNA (400 pg, 800 pg, 1200 pg, and 1600 pg) and the retinal integration efficiency was tested by ACT assay (supplementary material Fig. S3). Animal caps injected with 1600 pg did not survive, but those injected with lower concentrations formed retina in 21±10%, 41±11%, and 50±3% of animals, respectively. To be sure that the exogenous Follistatin can still function as a neural inducer, we scored the same sections for brain formation based on morphology and cellular location. The percentage of animals that contained transplanted cells found in the brain were comparable between Noggin- (80±6%; n = 80) and Follistatin-treated (74±10%; n = 60) cells. Thus, factors that repressed both BMP and Activin signals, such as Noggin and Cerberus, were more efficient retinal-inducers than Follistatin, an Activin-biased antagonist.

Bottom Line: We determined the effect of these treatments on retina formation using the Animal Cap Transplant (ACT) assay; in which treated pluripotent cells were transplanted into the eye field of host embryos.We found that inhibition of Activin signaling, in the presence of BMP signaling inhibition, promotes efficient retinal specification in Xenopus tissue, mimicking the affect of adding Noggin alone.In whole embryos, we found that the eye field marker, rax, expanded when adding both dominant-negative Smad1 and Smad2, as did treating the cells with both dorsomorphin and SB431542.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Physiology, SUNY Upstate Medical University, Syracuse, NY 13210, USA The Center for Vision Research, SUNY Eye Institute, Upstate Medical University, Syracuse, NY 13210, USA.

No MeSH data available.


Related in: MedlinePlus