Fig. 9. The Kazal1(–) cellular phenotype is enhanced after amputation. (A,B) Confocal images of cells isolated from (a-j) head-regenerating tips (HR) and (k-o) foot-regenerating tips (FR) of H. vulgaris, 1 hour after bisection following 3 exposures to (a-e) empty vector or (f-o) Kazal1 dsRNAs. Cells were stained with Hoechst 33342 dye (blue) and immunodetected with the anti-hyCREB (red), anti-RSK (green) antibodies. Bars, 16 µm. (B) Staging of the Kazal1(–) endodermal phenotype. Stage (st.) 0, normal, elongated shape, basal nucleus, homogenous RSK-staining; st.1, perinuclear vacuolization; st. 2, multiple non-confluent cytoplasmic vacuoles; st. 3, confluent vacuoles occupying most of the cytoplasm. Small digestive vacuoles that provide false red staining (star) were occasionally noted. (C) Frequencies of the respective phenotypes in control and Kazal1(–) hydra during head- and foot-regeneration. For each condition at least 300 endodermal epithelial cells were counted.