Project description:The transcription factor Zinc finger protein 148 (Zfp148) interacts physically with the tumor suppressor p53, but the siginficance of this interaction is not known. We recently showed that knockout of Zfp148 in mice leads to ectopic activation of p53 in tissues and cultured fibroblasts, suggesting that Zfp148 represses p53 activity. Here we hypothesized that targeting Zfp148 would unleash p53 activity and protect against cancer development, and test this idea in the APCMin/+ mouse model of intestinal adenomas. Crypt-enriched tissues were isolated by laser microdissection (PALM) from the small intestines (proximal) of Zfp148gt/+APCMin/+ and Zfp148+/+APCMin/+ mice for RNA extraction and hybridization to Affymetrix microarrays.
Project description:Genome wide expression profiling to determine the overlap of Affymetrix-signals with SOLID sequencing RNA was extracted using the Qiagen RNeasy kit following the manufacturers guidelines, arrays were prepared and hybridized following the Affymetrix protocol. Mus musculus samples from small intestine and colon, to be compared to transcript data aquired with other techniques
Project description:We collected whole genome testis expression data from hybrid zone mice. We integrated GWAS mapping of testis expression traits and low testis weight to gain insight into the genetic basis of hybrid male sterility.
Project description:The transcription factor Zinc finger protein 148 (Zfp148) interacts physically with the tumor suppressor p53, but the siginficance of this interaction is not known. We recently showed that knockout of Zfp148 in mice leads to ectopic activation of p53 in tissues and cultured fibroblasts, suggesting that Zfp148 represses p53 activity. Here we hypothesized that targeting Zfp148 would unleash p53 activity and protect against cancer development, and test this idea in the APCMin/+ mouse model of intestinal adenomas.
Project description:Background & Aims: We have recently established long-term culture conditions under which single crypts or stem cells derived from murine small intestine expand over long periods of time. Growing crypts undergo multiple crypt fission events, whilst simultaneously generating villus-like epithelial domains in which all differentiated cell types are present. We have now adapted the culture conditions to grow similar epithelial organoids from mouse colon and human small intestine and colon. Methods: Based on the murine small intestinal culture system, we optimized the murine and human colon culture system. Results: Addition of Wnt3A to the growth factor cocktail allowed mouse colon crypts to expand indefinitely. Further addition of nicotinamide, a small molecule Alk inhibitor and a p38 inhibitor was essential for long-term human small intestine and colon culture. The culture system also allowed growth of murine Apcmin adenomas, human colorectal cancer and human esophageal metaplastic Barrett’s epithelium. Conclusion: The culture technology should be widely applicable as a research tool for infectious, inflammatory and neoplastic pathologies of the human gastrointestinal tract. Moreover, regenerative applications may become feasible with ex vivo expanded intestinal epithelia. Human organoids were grown embedded in Matrigel in HISC (Human intestinal stem cell culture) medium. Additionally, human small intestinal crypts and villi were isolated independently from a freshly operated sample. RNA was isolated using the RNeasy Micro kit (Qiagen). Samples were labled according to Agilent guidelines with Cy3, whereas human reference RNA (Stratagene) was labeled in Cy5. Feature Extraction Software was used to extract and normalize data.
Project description:Background & Aims: We have recently established long-term culture conditions under which single crypts or stem cells derived from murine small intestine expand over long periods of time. Growing crypts undergo multiple crypt fission events, whilst simultaneously generating villus-like epithelial domains in which all differentiated cell types are present. We have now adapted the culture conditions to grow similar epithelial organoids from mouse colon and human small intestine and colon. Methods: Based on the murine small intestinal culture system, we optimized the murine and human colon culture system. Results: Addition of Wnt3A to the growth factor cocktail allowed mouse colon crypts to expand indefinitely. Further addition of nicotinamide, a small molecule Alk inhibitor and a p38 inhibitor was essential for long-term human small intestine and colon culture. The culture system also allowed growth of murine Apcmin adenomas, human colorectal cancer and human esophageal metaplastic Barrett’s epithelium. Conclusion: The culture technology should be widely applicable as a research tool for infectious, inflammatory and neoplastic pathologies of the human gastrointestinal tract. Moreover, regenerative applications may become feasible with ex vivo expanded intestinal epithelia.