IL-2-independent generation of FOXP3(+)CD4(+)CD8(+)CD25(+) cytotoxic regulatory T cell lines from human umbilical cord blood.
OBJECTIVE
Since the existence of mouse naturally occurring CD4(+)CD25(+) T regulatory (Treg) cells was demonstrated, a variety of human Treg subsets have been identified as distinct T cell populations. Here we show the establishment of novel Treg cell lines possessing unique characteristics.


METHODS
Novel Treg cell lines, designated HOZOT, were generated by coculturing human umbilical cord blood cells with mouse stromal cell lines in the absence of exogenous IL-2 or other cytokines. HOZOT were characterized and compared with CD4(+)CD25(+) Treg cells in terms of the CD phenotype, FOXP3 expression, suppressor activity against allogeneic MLR, anergy property, and IL-10 production.


RESULTS
HOZOT were generated and expanded as normal lymphoblastoid cells with cytotoxic activity against the cocultured stromal cells. HOZOT consisted of three subpopulations as defined by phenotype: CD4(+)CD8(+), CD4(+)CD8(dim), and CD4(-)CD8(+). All three subpopulations showed both suppressor and cytotoxic activities. While HOZOT's expression of FOXP3, CD25, GITR, and cytoplasmic CTLA-4 implied a similarity to naturally occurring CD4(+)CD25(+) Treg cells, these two Treg cells differed in IL-2 responsiveness and IL-10 production.


CONCLUSIONS
Our studies introduce a new method of generating Treg cells in an IL-2-independent manner and highlight a unique Treg cell type with cytotoxic activity and a phenotype of FOXP3(+)CD4(+)CD8(+)CD25(+).
Nakamura
Shuji
S
Cell Biology Institute, Research Center, Hayashibara Biochemical Laboratories, Inc., Okayama, Okayama, Japan. shnakamu@hayashibara.co.jp
Suzuki
Motoyuki
M
Sugimoto
Akira
A
Tsuji-Takayama
Kazue
K
Yamamoto
Mayuko
M
Otani
Takeshi
T
Inoue
Toshiya
T
Harashima
Akira
A
Okochi
Ayumi
A
Motoda
Ryuichi
R
Yamasaki
Fumiyuki
F
Orita
Kunzo
K
Kibata
Masayoshi
M
D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis*; D000943:Antigens, Differentiation / Q000096:biosynthesis; D060908:CTLA-4 Antigen; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D018920:Coculture Techniques; D003601:Cytotoxicity Tests, Immunologic; D005260:Female; D005312:Fetal Blood / Q000166:cytology; D051858:Forkhead Transcription Factors / Q000096:biosynthesis*; D053450:Glucocorticoid-Induced TNFR-Related Protein; D006801:Humans; D016130:Immunophenotyping; D016753:Interleukin-10 / Q000096:biosynthesis / Q000378:metabolism; D007376:Interleukin-2 / Q000494:pharmacology*; D051379:Mice; D017475:Receptors, Nerve Growth Factor / Q000096:biosynthesis; D018124:Receptors, Tumor Necrosis Factor / Q000096:biosynthesis; D050378:T-Lymphocytes, Regulatory / Q000166:cytology* / Q000276:immunology* / Q000378:metabolism
D015703:Antigens, CD; D000943:Antigens, Differentiation; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; C556707:Ctla4 protein, mouse; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D053450:Glucocorticoid-Induced TNFR-Related Protein; C508609:IL10 protein, human; D007376:Interleukin-2; D017475:Receptors, Nerve Growth Factor; D018124:Receptors, Tumor Necrosis Factor; C491405:TNFRSF18 protein, human; D016753:Interleukin-10