Noggin maintains pluripotency of human embryonic stem cells grown on Matrigel.
OBJECTIVE
Spontaneous differentiation of human embryonic stem cell (hESC) cultures is a major concern in stem cell research. Physical removal of differentiated areas in a stem cell colony is the current approach used to keep the cultures in a pluripotent state for a prolonged period of time. All hESCs available for research require unidentified soluble factors secreted from feeder layers to maintain the undifferentiated state and pluripotency. Under experimental conditions, stem cells are grown on various matrices, the most commonly used being Matrigel.


METHODS
We propose an alternative method to prevent spontaneous differentiation of hESCs grown on Matrigel that uses low amounts of recombinant noggin. We make use of the porosity of Matrigel to serve as a matrix that traps noggin and gradually releases it into the culture to antagonize bone morphogenetic proteins (BMP). BMPs are known to initiate differentiation of hESCs and are either present in the conditioned medium or are secreted by hESCs themselves.


RESULTS
hESCs grown on Matrigel supplemented with noggin in conditioned medium from feeder layers (irradiated mouse embryonic fibroblasts) retained both normal karyotype and markers of hESC pluripotency for 14 days. In addition, these cultures were found to have increased cell proliferation of stem cells as compared to hESCs grown on Matrigel alone.


CONCLUSIONS
Noggin can be utilized for short term prevention of spontaneous differentiation of stem cells grown on Matrigel.
Chaturvedi
G
G
Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA.
Simone
P D
PD
Ain
R
R
Soares
M J
MJ
Wolfe
M W
MW
D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors; D002352:Carrier Proteins / Q000008:administration & dosage / Q000235:genetics / Q000494:pharmacology*; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D018997:Chorionic Gonadotropin, beta Subunit, Human / Q000378:metabolism; D003094:Collagen / Q000494:pharmacology*; D017077:Culture Media, Conditioned / Q000494:pharmacology; D003692:Delayed-Action Preparations / Q000494:pharmacology; D004338:Drug Combinations; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D007797:Laminin / Q000494:pharmacology*; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D011509:Proteoglycans / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000494:pharmacology; D013997:Time Factors; D000099176:Noggin Protein
D019485:Bone Morphogenetic Proteins; D002352:Carrier Proteins; D018997:Chorionic Gonadotropin, beta Subunit, Human; D003094:Collagen; D017077:Culture Media, Conditioned; D003692:Delayed-Action Preparations; D004338:Drug Combinations; D007797:Laminin; D050814:Octamer Transcription Factor-3; D011509:Proteoglycans; D011994:Recombinant Proteins; D000099176:Noggin Protein; C494173:POU5F1 protein, human; C059485:matrigel