StemRegenin 1 increases the number of CD34+ cells after 5 to 7 days with an EC50 of 120 nM. Culture of mPB CD34+ cells with cytokines plus SR1 (1 μM) for 7 days increases the number of CD34+, CD133+, and CD90+ hematopoietic stem and progenitor cell populations 2.6-, 2.3-, and 10-fold, respectively compared to control cells. Continued culture with SR1 (1 μM) for 3 weeks leads to an 11-fold increase in total nucleated cells (TNC), a 73-fold increase in CD34+ cells as compared to control cultures, and a 1118-fold increase in CD34+ cells relative to input cells. Culture of 1×103 cord blood CD34+ cells for 5 weeks with SR1 (1 μM) results in the production of 1.69×106 colony forming cells. SR1-induces CD34+ cell expansion acts by binding and antagonizing AhR as evident by decreased CYP1B1 and AHRR mRNA levels. SR1 (1 μM) treatment accelerates the proliferation of CD34+ cells and decreased the expression levels of VentX in human CD34+ cells. Ectopic expression of VentX prevents SR1-induced expansion of CD34+ cells. Sequential coculture with bone morphogenetic protein 4 (20 ng/mL), PGE2 (2 μM), and SR1 (0.75 μM) lead to robust Macaca nemestrina iPSC hematopoietic progenitor cell formation. CD34(+)CD38(-)Thy1(+)CD45RA(-)CD49f(+) cells isolated on the basis of CD34 expression and cultured in SR1 (0.75 μM) expands 3-fold and maintained this long-term repopulating HSC phenotype.
StemRegenin-1(SR1)常見(jiàn)問(wèn)題:
1. 如何使用SR1,使用濃度多少?
大多數(shù)培養(yǎng)條件下,稀釋SR1儲(chǔ)存液(如10 mM DMSO) 1:10,000,加入感興趣的造血干細(xì)胞培養(yǎng)基內(nèi)獲得工作液濃度為1 μM。微量濃度的DMSO(0.01%)對(duì)細(xì)胞培養(yǎng)物*。但對(duì)于非傳統(tǒng)的培養(yǎng)基,如含高濃度人血清的培養(yǎng)基,建議進(jìn)行一組不同濃度SR1預(yù)實(shí)驗(yàn),因?yàn)镾R1的蛋白結(jié)合特性仍未知。