Transplantation of bone marrow genetically engineered to express proinsulin II protects against autoimmune insulitis in NOD mice.
BACKGROUND
Type 1 diabetes (T1D) is a T-cell-dependent autoimmune disease resulting from destructive inflammation (insulitis) of the insulin-producing pancreatic beta-cells. Transgenic expression of proinsulin II by a MHC class II promoter or transfer of bone marrow from these transgenic mice protects NOD mice from insulitis and diabetes. We assessed the feasibility of gene therapy in the NOD mouse as an approach to treat T1D by ex vivo genetic manipulation of normal hematopoietic stem cells (HSCs) with proinsulin II followed by transfer to recipient mice.


METHODS
HSCs were isolated from 6-8-week-old NOD female mice and transduced in vitro with retrovirus encoding enhanced green fluorescent protein (EGFP) and either proinsulin II or control autoantigen. Additional control groups included mice transferred with non-manipulated bone marrow and mice which did not receive bone marrow transfer. EGFP-sorted or non-sorted HSCs were transferred into pre-conditioned 3-4-week-old female NOD mice and insulitis was assessed 8 weeks post-transfer.


RESULTS
Chimerism was established in all major lymphoid tissues, ranging from 5-15% in non-sorted bone marrow transplants to 20-45% in EGFP-sorted bone marrow transplants. The incidence and degree of insulitis was significantly reduced in mice receiving proinsulin II bone marrow compared to controls. However, the incidence of sialitis in mice receiving proinsulin II bone marrow and control mice was not altered, indicating protection from insulitis was antigen specific.


CONCLUSIONS
We show for the first time that ex vivo genetic manipulation of HSCs to express proinsulin II followed by transplantation to NOD mice can establish molecular chimerism and protect from destructive insulitis in an antigen-specific manner.
Chan
James
J
Department of Immunology, Monash University, Commercial Road, Prahran, Victoria 3181, Australia.
Clements
Warren
W
Field
Judith
J
Nasa
Zeyad
Z
Lock
Peter
P
Yap
Felicia
F
Toh
Ban-Hock
BH
Alderuccio
Frank
F
D000818:Animals; D001483:Base Sequence; D016026:Bone Marrow Transplantation; D017931:DNA Primers / Q000235:genetics; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics / Q000473:pathology / Q000517:prevention & control*; D005260:Female; D015870:Gene Expression; D005818:Genetic Engineering; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D018380:Hematopoietic Stem Cell Transplantation; D007515:Islets of Langerhans / Q000473:pathology; D051379:Mice; D016688:Mice, Inbred NOD; D011384:Proinsulin / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic; D018183:Transplantation Chimera
D017931:DNA Primers; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D011384:Proinsulin