[Expression changes of protein kinase Calpha during differentiation of mouse embryonic stem cells into neuron-like cells].
OBJECTIVE
To study the expression of protein kinase Calpha (PKCalpha) during differentiation of mouse embryonic stem cells (ESC)into neuron-like cells in an attempt to elucidate their role in signaling.


METHODS
ES-D3 cells were subjected to an 18-day induction procedure which consisted of 4 days of culture as embryoid bodies (EBs) without all-trans retinoic acid (RA) followed by 14 days of culture in the presence of RA, then the EBs were plated separately onto gelatin-coated 6-well culture dishes coated glass coverslip for immunohistochemistry, onto 100-mm-diameter bacteriological (nontissue culture) dish for Western blot assay and RT-PCR, and were cultivated and collected cells for 1, 3, 5, 7 and 14 days in the presence of RA.


RESULTS
Staining of PKCalpha was strong and distributed throughout the cells, especially in the cytoplasm and membrane. Western blot analysis exhibited a band of approximately 84 kD with the antibody specific to PKCalpha. After induction of RA, PKCalpha was present in significantly lower levers in induced ESC as compared with that in ES-D3 cells at first, then PKCalpha isoform was found in slightly higher amounts and resumed on 14th day.


CONCLUSIONS
This study was characterized the PKCalpha isoenzyme profile in RA-induced differentiation of ESC into neuron-like cells and suggests that PKCalpha may play a very important role in differentiation of mouse ESC along the neuronal pathway.
Gao
Qianying
Q
Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, China.
Ge
Jian
J
Wang
Zhichong
Z
Chen
Huiyi
H
Huang
Danping
D
Yuan
Zhaohui
Z
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology; D004627:Embryonic Induction; D051379:Mice; D009474:Neurons / Q000166:cytology*; D011493:Protein Kinase C / Q000096:biosynthesis* / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000201:enzymology; D014212:Tretinoin / Q000494:pharmacology
D014212:Tretinoin; D011493:Protein Kinase C