Expansion of CD4(+)CD25 (+) regulatory T cells from cord blood CD4(+) cells using the common γ-chain cytokines (IL-2 and IL-15) and rapamycin.
Rapamycin has important roles in the modulation of regulatory T cells. We tried to expand CD4(+)CD25(+) regulatory T cells (Treg cells) from umbilical cord blood (CB) CD4-positive cells using interleukin (IL)-15 or IL-2 with transforming growth factor (TGF)-β and rapamycin. We were able to obtain more than 500-fold expansion of CD4(+)CD25(+) cells from CB CD4(+) cells using IL-15 and TGF-β with rapamycin. These expanded CD4(+)CD25(+) cells expressed forkhead box P3 (FoxP3) mRNA at a level about 100-fold higher and could suppress allogeneic mixed lymphocyte culture (MLC) by more than 50%. Early after rapamycin stimulation, CB CD4(+) cells showed increased expression of FoxP3 and a serine/threonine kinase Pim2 and sustained expression of negative phosphoinositide 3-kinase regulator phosphatase and tensin homolog deleted on chromosome 10 (PTEN). On the other hand, CD4(+)CD25(+) cells expanded with rapamycin for 8 days showed much higher levels of FoxP3 mRNA expression and decreased expression of PTEN. A comparison of IL-15 stimulation and IL-2 stimulation showed slightly higher efficiency of IL-15 for expansion of CD4(+)CD25(+) cells, and for FoxP3 expression, IL-15 also showed significantly higher efficacy for inhibition of MLC. The combination of the common γ-chain cytokine IL-15, TGF-β, and rapamycin may be a useful means for expanding Treg cells. Pim2 expression early after stimulation with rapamycin may be important for conferring rapamycin resistance for growth of Treg cells. IL-15 is not less useful than IL-2 for expansion of Treg cells.
Department of Hematology and Oncology, Hokkaido University Graduate School of Medicine, N15 W7, Kita-Ku, Sapporo, 060-8638, Japan.
Shinsuke
S
Asanuma
Junji
J
Tanaka
Junichi
J
Sugita
Mizuha
M
Kosugi
Souichi
S
Shiratori
Kentarou
K
Wakasa
Yusuke
Y
Shono
Akio
A
Shigematsu
Takeshi
T
Kondo
Takahiko
T
Kobayashi
Masahiro
M
Asaka
Masahiro
M
Imamura
D015704:CD4 Antigens / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D004338:Drug Combinations; D005312:Fetal Blood / Q000166:cytology* / Q000378:metabolism; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D053631:Interleukin Receptor Common gamma Subunit / Q000502:physiology; D019409:Interleukin-15 / Q000494:pharmacology*; D007376:Interleukin-2 / Q000494:pharmacology*; D053645:Interleukin-2 Receptor alpha Subunit / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D020123:Sirolimus / Q000494:pharmacology*; D050378:T-Lymphocytes, Regulatory / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
D015704:CD4 Antigens; D004338:Drug Combinations; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; C508017:IL2RA protein, human; D053631:Interleukin Receptor Common gamma Subunit; D019409:Interleukin-15; D007376:Interleukin-2; D053645:Interleukin-2 Receptor alpha Subunit; C490587:PIM2 protein, human; D011518:Proto-Oncogene Proteins; D017346:Protein Serine-Threonine Kinases; D020123:Sirolimus