Low-level tsg1 knockdown causes a ventral tail vein edema and cell death in anterior structures, but does not alter patterning during early somitogenesis. Uninjected embryo (A); a wild-type embryo injected with 8 ng MO1 (B) displays a slightly reduced head and a large edema in, or dilation of, the ventral tail vein (arrowhead), in which the blood accumulates and circulation slows. (C) The vein edema can be rescued by injection of tsg1 mRNA (10% with edema phenotype, n=133, not shown) or tsg2 mRNA (8% with edema phenotype, n=106), whereas the reduction in head size is unaltered. Scale bars in A-C show the diameter of the eye as a measure of the extent of anterior tissues. TUNEL analysis as a measure of cell death in 24 hours post fertilization (hpf) uninjected wild-type (D) and 8 ng MO1-injected sibling (E) embryos. Cell death is not rescued by co-injection of tsg mRNA (F). (D-F) Anterior towards the left, dorsal view posterior to the MHB boundary. (G-N) Whole-mount in situ hybridization of uninjected embryos (G-J) or embryos injected with 8 ng MO1 (K-N). (G,K) Expression of pax2.1 (arrowhead) and krox20 (bracket) in the mid/hindbrain (MHB) boundary and rhombomeres 3 and 5, respectively. (H,L) More posterior views of the embryos in G and K. pax2.1 expression in the pronephric system (arrowheads) and myoD in anterior somitic mesoderm is unchanged. (I,M) Lateral views of untreated or injected embryos showing no alterations in anterior expression of pax2.1 in the eye field (asterisk). (J,N) Expression of bmp4 anteriorly (asterisk) and in the tail bud (arrowhead) at the three-somite stage is normal in injected embryos. Insets show tail bud views. (G-I,K-M) Eight somites. (J,N) Three somites. (G,H,K,L) Dorsal views with anterior upwards. (I,J,M,N) Lateral views, anterior is upwards.
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