GSK-3β inhibition promotes engraftment of ex vivo-expanded hematopoietic stem cells and modulates gene expression.
Glycogen synthase kinase-3β (GSK-3β) has been identified as an important regulator of stem cell function acting through activation of the wingless (Wnt) pathway. Here, we report that treatment with an inhibitor of GSK-3β, 6-bromoindirubin 3'-oxime (BIO) delayed cell cycle progression by increasing cell cycle time. BIO treatment resulted in the accumulation of late dividing cells enriched with primitive progenitor cells retaining the ability for sustained proliferation. In vivo analysis using a Non-obese diabetic/severe combined immunodeficient (NOD/SCID) transplantation model has demonstrated that pretreatment with BIO promotes engraftment of ex vivo-expanded hematopoietic stem cells. BIO enhanced the engraftment of myeloid, lymphoid and primitive stem cell compartments. Limiting dilution analysis of SCID repopulating cells (SRC) revealed that BIO treatment increased human chimerism without increasing SRC frequency. Clonogenic analysis of human cells derived from the bone marrow of transplant recipient mice demonstrated that a higher level of human chimerism and cellularity was related to increased regeneration per SRC unit. Gene expression analysis showed that treatment with BIO did not modulate the expression of canonical Wnt target genes upregulated during cytokine-induced cell proliferation. BIO increased the expression of several genes regulating Notch and Tie2 signaling downregulated during ex vivo expansion, suggesting a role in improving stem cell engraftment. In addition, treatment with BIO upregulated CDK inhibitor p57 and downregulated cyclin D1, providing a possible mechanism for the delay seen in cell cycle progression. We conclude that transient, pharmacologic inhibition of GSK-3β provides a novel approach to improve engraftment of expanded HSC after stem cell transplantation.
Sydney Cord and Marrow Transplant Facility, Centre for Children's Cancer and Blood Disorders, Sydney Children's Hospital, Randwick, Sydney, Australia.
Kap-Hyoun
KH
Ko
Tiffany
T
Holmes
Patricia
P
Palladinetti
Emma
E
Song
Robert
R
Nordon
Tracey A
TA
O'Brien
Alla
A
Dolnikov
D000818:Animals; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D046528:Chimerism / Q000187:drug effects; D019938:Cyclin D1 / Q000378:metabolism; D050761:Cyclin-Dependent Kinase Inhibitor p57 / Q000378:metabolism; D005312:Fetal Blood / Q000166:cytology; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors*; D000071679:Glycogen Synthase Kinase 3 beta; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods*; D006412:Hematopoietic Stem Cells / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D010091:Oximes / Q000494:pharmacology*; D015398:Signal Transduction / Q000235:genetics; D051153:Wnt Proteins / Q000378:metabolism*
C483321:6-bromoindirubin-3'-oxime; D050761:Cyclin-Dependent Kinase Inhibitor p57; D007211:Indoles; D010091:Oximes; D051153:Wnt Proteins; D019938:Cyclin D1; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3