Hs in a villus-trough unit compared with the intestine of purchase T0901317 theTamaoki et al. Cell Biosci (2016) 6:Page 8 offabp1 H3K36me1 15 10 5 0 15 10 5 0 15 10 5 0 10 5a afabp2acdx2afxrabAbBb aCb aDab a20 10 PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/27488460 0aEarpla a5 05 05 0bFb abGb abHRelative input ( ) H3K36me2 H3K36meIJb a ab5aabaa5 0Kab a10 5 0Lab a10Mb a0abNb a aOb ab a5 010 0PQb a a0 10 5Ra ab bSb a aTa a aRNAPIIS2 Pabab55fed fasted refedfed fasted refedfed fasted refed0 fed fasted refed fed fasted refedFig. 5 Epigenetic modifications on fabp1, fabp2, cdx2 and fxr genes in the intestines of fed, fasted and refed X. laevis. Chromatin samples were prepared from the intestines from the frogs that were fed for 22 days (fed), fasted for 22 days (fasted), or fasted for 21 days and then refed for 1 day (refed). Signals of ChIP on fabp1 (A, F, K and P ), fabp2 (B, G, L and Q), cdx2 (C, H, M and P), fxr (D, I, N and S) and rpl8 (E, J, O and T) genes were detected by qPCR following immunoprecipitation with antibodies against H3K36me1 (A ), H3K36me2 (F ), H3K36me3 (K ), and RNAPIIS2P (P ). Primers used in qPCR are shown in Additional file 6: Table S3. Each value is the mean ?SEM (n = 8). Distinct letters denote significantly different means (p < 0.05). These experiments were repeated at least two times, with similar resultsfasted frogs, suggesting the appearance of the epithelial cells that begin to proliferate and differentiated at various positions in a villus-trough unit to generate new troughs within 1 day after refeeding. Future studies need to address what characteristics these cells have and how these cells appeared in response to refeeding.Intestinal functions and gene expressionFasting down-regulated and refeeding recovered not only the functions of the intestinal epithelial cells but also the motility of the digestive tract. The activities of intestinal alkaline phosphatase, aminopeptidase, glucoamylase and maltase declined during fasting and quickly recovered to the feeding levels within 1 day after refeeding. Although the level of transcripts of these enzymes declined during fasting, the transcript levels of these enzymes hardly increased with refeeding, suggesting the presence of post-transcriptional regulation. Alternatively, as intestinal alkaline phosphatase, aminopeptidase and maltaseglucoamylase are membrane proteins that are subject toglycosylation, sulfation or phosphorylation at specific residues, regulation of such post-translational modifications may be another possibility to explain the discordance between the enzyme activity and transcription level. The expression of the fabp1, fabp2, fabp6 and rbp2 genes, which are involved in the cellular transport of fatty acids and retinol, was also quickly recovered or up-regulated from a down-regulated state of fasting within 1 day after refeeding. In contrast, mammalian intestines need at least 2? days after refeeding to recover the protein or transcript amounts of the alkaline phosphatase and glucose transporter, GLUT2 [22, 23]. The X. laevis intestine may have some mechanisms by which the enzyme activities and the transcription of the genes for fatty acid and retinol transport proteins respond quickly and selectively to refeeding. The presence of digesta in the rectum of the frogs that fasted for 5 months suggests an increase in digesta retention times during fasting. A similar observation has also been reported in the green striped burrowing frog, Cyclorana alboguttata during 3 months ofTamaoki et al. Cell Biosci (2016) 6:Page 9 of.