Pluripotent stem cell manufacturing system and method for producing induced pluripotent stem cells

Abstract

The present invention provides a stem cell manufacturing system comprising: a pre-transfer cell solution sending channel 20 through which a solution containing cells flows; an inducer solution sending mechanism 21 which sends a pluripotency inducer into the pre-transfer cell solution sending channel 20; an inducer transfer apparatus 30 which is connected to the pre-transfer cell solution sending channel 20 and transfers the pluripotency inducer into the cells to produce cells harboring the inducer; a cell cluster production apparatus 40 which cultures the cells harboring the inducer to produce a plurality of cell clusters consisting of stem cells; and a packaging apparatus 100 which sequentially packages the plurality of cell clusters.

1 . A method for producing induced pluripotent stem cells, comprising:
 preparing somatic cells; and   transferring reprogramming factor RNAs into the somatic cells by a lipofection method.   
     
     
         2 . The method for producing induced pluripotent stem cells according to  claim 1 , wherein the somatic cells are blood cells. 
     
     
         3 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are monocytes. 
     
     
         4 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are hematopoietic stem/progenitor cells. 
     
     
         5 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are CD34-positive. 
     
     
         6 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are blood cells separated on condition that the cells are CD34-positive. 
     
     
         7 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are CD3-positive. 
     
     
         8 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the blood cells are blood cells separated on condition that the cells are CD3-positive. 
     
     
         9 . The method for producing induced pluripotent stem cells according to  claim 1 , wherein the reprogramming factor RNAs comprise Oct3/4mRNA, Sox2 mRNA, Klf4 mRNA, and c-Myc mRNA. 
     
     
         10 . The method for producing induced pluripotent stem cells according to  claim 9 , wherein the reprogramming factor RNAs further comprise at least one member selected from the group consisting of GLIS1 mRNA, FOXH1 mRNA, L-MYC mRNA, and p53-dn mRNA. 
     
     
         11 . The method for producing induced pluripotent stem cells according to  claim 9 , wherein the reprogramming factor RNAs further comprise LIN28A mRNA or LIN28B mRNA. 
     
     
         12 . The method for producing induced pluripotent stem cells according to  claim 1 , wherein an siRNA lipofection reagent or an mRNA lipofection reagent is used in the lipofection with the reprogramming factor RNAs. 
     
     
         13 . The method for producing induced pluripotent stem cells according to  claim 1 , wherein at least one member selected from Lipofectamine® RNAiMAX transfection reagent, Lipofectamine® MessengerMAX transfection reagent, Stemfect® RNA transfection reagent, and ReproRNA® transfection reagent is used in the lipofection with the reprogramming factor RNAs. 
     
     
         14 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the number of the blood cells for the lipofection with the reprogramming factor RNAs is 1 to 1·10 8 . 
     
     
         15 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the amounts of the reprogramming factor RNAs for the lipofection with the reprogramming factor RNAs are 5 ng to 50 μg per run. 
     
     
         16 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the amount of the lipofection reagent for the lipofection with the reprogramming factor RNAs is 0.1 μL to 500 μL per run. 
     
     
         17 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the lipofection with the reprogramming factor RNAs is performed for 0.1 hours or longer and 24 hours or shorter per run. 
     
     
         18 . The method for producing induced pluripotent stem cells according to  claim 2 , wherein the lipofection with the reprogramming factor RNAs is performed a plurality of times. 
     
     
         19 . The method for producing induced pluripotent stem cells according to  claim 1 , wherein the medium used for the lipofection with the reprogramming factor RNAs is Opti-MEM®. 
     
     
         20 . The method according to  claim 3 , further comprising separating the monocytes from blood using a filter.