U937 cells were treated with 200nM of PMA . After 3 days, the PMA-containing media was removed and replaced with fresh RPMI 1640 medium for a further 5 days. The final yield was very less.
If I understand your question correctly, you would like to increase the number of differentiated U937 cells in your culture. If this is the case, one can try increasing the concentration of PMA or increase the incubation time. The extent of differentiation depends upon additional factors such as cell density and serum growth factors.
Hi Keshavardini,
Replying to your request on how to increase the amount of differentiated U937 cells, I’ve recalled an old paper from Sloan-Kettering Division, Cornell University group of Nynke Y. Rotset al. entitled “Induced Differentiation of U937 Cells by 1,25-dihydroxyvitamin D3 Involves Cell Cycle Arrest in G1 That Is Preceded by a Transient Proliferative Burst and an Increase in Cyclin Expression”, published in Blood 93(8), 1999. In a nutshell, the authors observed almost 2-fold increased amount of differentiated U937 cells. These human myelomonocytic cells (clone 4) were routinely maintained in RPMI-1640 medium supplemented with 10% fetal bovine serum and 5 mmol/L L-glutamine. In the study, they presented evidence demonstrating that cell cycle arrest accompanies differentiation in U937 cells treated with 1,25(OH)2D3. This effect was facilitated in part by elevations in the levels of CDK inhibitors p21 and p27, which in the case of p21. Dr. Rotset and his colleagues demonstrated a direct transcriptionally regulated target of 1,25(OH)2D3 through its cognate nuclear receptor VDR. Paradoxically, growth arrest and differentiation of this cell line was found to be preceded by two transient proliferative bursts that were accompanied by increases in the levels cyclin A, D1, and E, as well as p21-associated kinase activity, shortly after the addition 1,25(OH)2D3.
Best wishes,
Ilya
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