Efficient culture of CD8(+) T cells from the islets of NOD mice and their use for the study of autoreactive specificities.
During progression to type 1 diabetes (T1D), the pancreatic islets of humans and of the widely studied mouse model of T1D, the nonobese diabetic (NOD) mouse, are infiltrated by cells of the immune system. Here we report that infiltrated mouse islets ("translucent islets") can be identified visually. We demonstrate the use of an efficient method for the isolation and culture of the islet-infiltrating cells of NOD mice, which results in a high percentage of CD8(+) T cells after seven days, with minimal manipulation. We show that islet-infiltrating cells exit the islets soon after they are placed in culture and can be used in flow cytometric experiments several hours later. Importantly, we demonstrate that the cultured cells are antigen-responsive and that specificities present at the beginning of the culture are generally still present on day seven. In addition, some reactivities are undetected without culture, supporting the validity of the seven-day expansion period. This technique greatly facilitates investigations of the CD8(+) T cell reactivities that play a pivotal role in the demise of pancreatic beta cells leading to the development of T1D.
Jarchum
Irene
I
Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Takaki
Toshiyuki
T
DiLorenzo
Teresa P
TP
D000818:Animals; D001324:Autoantigens / Q000276:immunology; D015551:Autoimmunity; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000473:pathology*; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental / Q000276:immunology / Q000473:pathology*; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology / Q000473:pathology*; D006801:Humans; D050417:Insulin-Secreting Cells / Q000276:immunology / Q000473:pathology*; D051379:Mice; D016688:Mice, Inbred NOD
D001324:Autoantigens
Diabetes. 1994 Mar;43(3):505-9
8314025
J Clin Invest. 1993 Nov;92(5):2313-22
8227346
Diabetologia. 1989 May;32(5):282-9
2666213
Diabetologia. 2003 Mar;46(3):305-21
12687328
Adv Immunol. 1992;51:285-322
1323922
Transplantation. 1985 Oct;40(4):437-8
2996187
J Immunol. 2003 Jul 1;171(1):185-95
12816997
Eur J Immunol. 1996 Aug;26(8):1762-9
8765018
Diabetes. 2007 Oct;56(10):2551-60
17620420
J Immunol. 2006 Mar 1;176(5):3257-65
16493087
J Immunol. 2006 Feb 1;176(3):1637-44
16424193
J Immunol. 1999 Nov 15;163(10):5708-14
10553102
Diabetes. 1994 Apr;43(4):599-606
8024653
Curr Protoc Immunol. 2001 May;Chapter 15:15.9.1-15.9.23
18432739
Int Immunol. 1995 Jan;7(1):139-46
7718510
J Clin Invest. 2005 Jul;115(7):1879-87
15937548
Proc Natl Acad Sci U S A. 2008 Apr 29;105(17):6374-9
18430797
Clin Immunol. 2008 Jun;127(3):359-65
18358785
J Immunol. 2008 Apr 15;180(8):5430-8
18390725
J Exp Med. 2004 Jun 7;199(11):1467-77
15184500
J Immunol. 1994 Aug 1;153(3):1360-77
7913115
Diabetes. 2007 Mar;56(3):613-21
17327428
J Immunol. 2007 Jun 1;178(11):7458-66
17513797
J Immunol Methods. 1995 Nov 16;187(1):163-9
7490452
Diabetes. 2008 May;57(5):1312-20
18305140
J Immunol. 2004 Dec 1;173(11):6727-34
15557165
Nat Med. 2005 Jun;11(6):645-52
15908957
J Clin Invest. 1992 Nov;90(5):1901-10
1385478