Loss of p21Waf1/Cip1/Sdi1 enhances intestinal stem cell survival following radiation injury.
The microcolony assay following gamma irradiation (IR) is a functional assay of intestinal stem cell fate. The cyclin-dependent kinase (CDK) inhibitor p21(Waf1/Cip1/Sdi1) (p21) regulates cell cycle arrest following DNA damage. To explore the role of p21 on stem cell fate, we examined the effects of p21 deletion on intestinal crypt survival following IR and expression of the stem/progenitor cell marker Musashi-1 (Msi-1) and the antiapoptotic gene survivin. Intestinal stem cell survival in adult wild-type (WT) and p21(-/-) mice was measured using the microcolony assay. Msi-1, p21, and survivin mRNA were measured using real-time PCR and immunohistochemical analysis. Laser capture microdissection (LCM) was used to isolate mRNA from the crypt stem cell zone. No differences in radiation-induced apoptosis were observed between WT and p21(-/-) mice. However, increased crypt survival (3.0-fold) was observed in p21(-/-) compared with WT mice 3.5 days after 13 Gy. Msi-1 and survivin mRNA were elevated 12- and 7.5-fold, respectively, in LCM-dissected crypts of p21(-/-) compared with WT mice. In conclusion, deletion of p21 results in protection of crypt stem/progenitor cells from IR-induced cell death. Furthermore, the increase in crypt survival is associated with increased numbers of Msi-1- and survivin-expressing cells in regenerative crypts.
George
Robert J
RJ
Department of Internal Medicine, Division of Gastroenterology, Washington University School of Medicine, St. Louis, Missouri, USA.
Sturmoski
Mark A
MA
May
Randal
R
Sureban
Sripathi M
SM
Dieckgraefe
Brian K
BK
Anant
Shrikant
S
Houchen
Courtney W
CW
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects; D002470:Cell Survival / Q000528:radiation effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D005260:Female; D051034:Inhibitor of Apoptosis Proteins; D007421:Intestine, Small / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008938:Mitosis / Q000528:radiation effects; D009419:Nerve Tissue Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D012097:Repressor Proteins; D013234:Stem Cells / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects*; D000077022:Survivin; D013997:Time Factors; D014916:Whole-Body Irradiation
C117749:Birc5 protein, mouse; C493957:Cdkn1a protein, mouse; D050759:Cyclin-Dependent Kinase Inhibitor p21; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; C104786:Msi1h protein, mouse; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; D000077022:Survivin