Protection Against Autoimmune Myocarditis by Gene Transfer of Interleukin-10 by Electroporation

Abstract

Background-Although immunosuppressive therapy for myocarditis has attracted a great deal of attention, its effectiveness is controversial. Interleukin (IL)-10 has a variety of immunomodulatory properties. Among the nonviral techniques for gene transfer in vivo, the direct injection of plasmid DNA into muscle is simple, inexpensive, and safe. Methods and Results-We examined the applicability of murine IL-10 (mIL-10) gene transfer to the treatment of rats with experimental autoimmune myocarditis. Nine-week-old Lewis rats were inoculated with pig myosin (day 0). A plasmid vector expressing mIL-10 cDNA (800 g per rat) was transferred into the tibialis anterior muscles by electroporation 3 times (5 days before immunization and at days 4 and 13); control rats received empty plasmid. Electroporation increased the serum mIL-10 levels to Ͼ250 pg/mL. The 21-day survival rate in rats treated with mIL-10 cDNA was higher (15 of 15; 100%) than that of the control group (9 of 15; 60% In a rat model in which myocarditis was induced by purified cardiac myosin, T cells were reported to play an important role in inducing myocarditis. [3][4][5] Immune dysfunction associated with autoimmune disease may be related to an imbalance between T helper type 1 and 2 cells. 6 The T helper type 2-associated cytokine interleukin (IL)-10 has a variety of immunomodulatory properties, including the inhibition of T helper type 1 cells and the production of proinflammatory cytokines. 7,8 Recent reports have suggested that the immunosuppressive effects associated with IL-10 are effective in suppressing the rejection of transplanted organs and immune complex diseases, and clinical trials of IL-10 have been performed in patients with these disorders. 9,10 Electroporation has been widely used to introduce DNA into various types of cells in vitro. Gene transfer by electroporation in vivo has been effective for introducing DNA into mouse skin, chick embryos, rat liver, and murine melanoma and muscle. [11][12][13] We previously showed that gene transfer into muscles by electroporation in vivo can be used to deliver cytokines systemically. 13,14 In the present study, we applied this method for the delivery of IL-10 in a rat model of autoimmune myocarditis. Methods AnimalsNine-week-old male Lewis rats were injected with the antigenadjuvant emulsion in their foot pads according to the procedure described previously. [3][4][5] The morbidity of experimental autoimmune myocarditis was 100% in rats immunized by this method. [3][4][5] Throughout the studies, all animals were treated in accordance with the guidelines for animal experiments of our institute. 5 Construction of Mouse IL-10 Expression VectorMouse IL-10 (mIL-10) cDNA cloned by polymerase chain reaction was inserted into the unique Xho I site between the cytomegalovirus immediate early enhancer-chicken ␤-actin hybrid promoter and rabbit ␤-globin poly A site of the pCAGGS expression plasmid. 13,14 The resulting plasmid, pCAGGS-IL-10, was grown in Escherichia coli DH 5␣ and prepared using plasmid purifica...

Inhibition of mast cells by interleukin‐10 gene transfer contributes to protection against acute myocarditis in rats

“…Likewise, gene transfer of IL-10 in our protocol may also have provided long-term exposure. As reported earlier, in EAM rats, mouse IL-10 levels are over 80 pg/ml until day 10 after a single administration of mouse IL-10 cDNA, which is greater than the peak value of serum rat IL-10 during the natural course of progression of acute myocarditis in this animal model (42AE4 pg/ml) [36]. Also in the present study, administration of mouse IL-10 cDNA on days 7, 12 and 17 resulted in elevation of the serum IL-10 level on day 21 compared to that in the group V animals (which received only empty plasmid on those days), which is again much greater than the peak value of serum rat IL-10 during the natural course of progression of acute myocarditis.…”

Central Neurotranspeptide, Alpha-Melanocyte-Stimulating Hormone (.ALPHA.-MSH) is Upregulated in Patients with Congestive Heart Failure

“…Inflammation is reduced by α-MSH via direct and indirect mechanisms (25) (27,28 (30,31), α-MSH may be one of the therapeutic targets of CHF in the future (32,33). IL-10 is also considered to be one of the therapeutic targets for acute myocarditis and heart failure (12,32,33).…”

“…IL-10 is also considered to be one of the therapeutic targets for acute myocarditis and heart failure (12,32,33). Interestingly, the induction of IL-10 was reported as a major effect of anti-inflammatory effects by α-MSH in monocytes (18) …”

Non-viral gene delivery in skeletal muscle: a protein factory

“…Applications of this technique have been extended from treating muscular dystrophy to using muscle for systemic delivery of therapeutic proteins. [97][98][99][100][101] The mechanisms by which this method enhances transgene expression appear to be relatively simple. Electroporation is effective only when plasmid DNA is injected into the muscle prior to, not after, electroporation.…”