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Fig. 2 TSA impairs growth and disrupts morphogenesis of exocrine pancreas in zebrafish larvae with hyperacetylation of nucleosomal histones.
WT zebrafish larvae were incubated in the absence or presence of 165 nM TSA (added at 48h.p.f.) for 24hours and then analyzed. (A) Exocrine pancreas (arrows) was analyzed by in situ hybridization using trypsin anti-sense riboprobes. Pancreatic acinar morphology by immunohistochemistry using anti-cadherin (Cad) antibodies, followed by transverse histological sectioning. e.p. exocrine pancreas; i, intestine. (B) The larvae were pulse-labeled with BrdU and analyzed by immunohistochemistry using anti-BrdU antibodies, followed by transverse histological sectioning. The number (#) of DAPI+ nuclei, the number (#) of BrdU+ nuclei, and the proportion of cells in S phase (% BrdU+ nuclei) were determined. Result is presented as the mean + s.d., and * indicates statistical significance, # trend of statistical significance. (C) Total protein was extracted and analyzed for acetylated and total histones H3 and H4 by immunoblotting. Anti-total histones H3 and H4 antibodies and anti-actin antibodies were used as internal controls.