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DNA transfection of mononuclear cells in muscle tissue
Author(s) -
Grønevik Eirik,
Tollefsen Stig,
Sikkeland Liv Ingunn Bjoner,
Haug Terje,
Tjelle Torunn Elisabeth,
Mathiesen Iacob
Publication year - 2003
Publication title -
the journal of gene medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.689
H-Index - 91
eISSN - 1521-2254
pISSN - 1099-498X
DOI - 10.1002/jgm.416
Subject(s) - transfection , electroporation , microbiology and biotechnology , spleen , dna vaccination , biology , lymph node , green fluorescent protein , peripheral blood mononuclear cell , immune system , immunology , cell culture , in vitro , gene , biochemistry , genetics , immunization
Background Genes encoding non‐self proteins may be injected into skeletal muscles in vivo to obtain induction of cellular and humoral immune responses against the encoded antigens (DNA vaccination). Bone marrow derived professional antigen‐presenting cells (APCs) play a key role in the induction of immunity by DNA vaccination. In the present work we have investigated whether the APCs are transfected by DNA injection into muscle. Methods DNA encoding green fluorescent protein (GFP) was injected into rat and mouse limb muscle and followed by electroporation. Whole mount muscle tissue with GFP‐positive mononuclear cells (MNCs) were treated with immunocytochemical markers specific for leukocytes, and studied with fluorescent microscopy. To detect transfected cells migrating to peripheral lymphoid tissue RT‐PCR was applied on RNA isolated from the draining popliteal lymph node and spleen. Lymphoid tissue was also analyzed with real‐time PCR for distribution of the injected plasmid. Results MNCs were transfected after intramuscular DNA injection, and, following DNA injection with electroporation, the number of GFP‐positive MNCs increased 6‐fold in rats and 14‐fold in mice. None of the GFP‐positive MNCs were stained with leukocyte‐specific antibodies. Even though GFP encoding DNA was detected in the popliteal lymph node, no RNA encoding GFP was found in the lymph node or spleen. However, MHC II‐positive cells in the muscle tissue appeared preferentially around the transfected MNCs. Conclusions Many MNCs in the muscle are transfected after intramuscular DNA injection. Electroporation significantly increases the number of transfected MNCs. None of the observed transfected MNCs however were leukocytes. MHC II‐positive cells accumulated around transfected MNCs; this suggests that transfer of antigen from transfected MNCs to APCs may contribute to the immune response. Copyright © 2003 John Wiley & Sons, Ltd.