Molecular Medicine

, Volume 19, Issue 1, pp 65–71 | Cite as

Effect of Recombinant α1-Antitrypsin Fc-Fused (AAT-Fc) Protein on the Inhibition of Inflammatory Cytokine Production and Streptozotocin-Induced Diabetes

  • Siyoung Lee
  • Youngmin Lee
  • Kwangwon Hong
  • Jaewoo Hong
  • Suyoung Bae
  • Jida Choi
  • Hyunjhung Jhun
  • Areum Kwak
  • Eunsom Kim
  • Seunghyun Jo
  • Charles A. Dinarello
  • Soohyun Kim
Research Article


α1-Antitrypsin (AAT) is a member of the serine proteinase inhibitor family that impedes the enzymatic activity of serine proteinases, including human neutrophil elastase, cathepsin G and neutrophil proteinase 3. Here, we expressed recombinant AAT by fusing the intact AAT gene to the constant region of IgG1 to generate soluble recombinant AAT-Fc protein. The recombinant AAT-Fc protein was produced in Chinese hamster ovary (CHO) cells and purified using mini-protein A affinity chromatography. Recombinant AAT-Fc protein was tested for antiinflammatory function and AAT-Fc sufficiently suppressed tumor necrosis factor (TNF)-α-induced interleukin (IL)-6 in human peripheral blood mononuclear cells (PBMCs) and inhibited cytokine-induced TNFα by different cytokines in mouse macrophage Raw 264.7 cells. However, AAT-Fc failed to suppress lipopolysaccharide-induced cytokine production in both PBMCs and macrophages. In addition, our data showed that AAT-Fc blocks the development of hyperglycemia in a streptozotocin-induced mouse model of diabetes. Interestingly, we also found that plasma-derived AAT specifically inhibited the enzymatic activity of elastase but that AAT-Fc had no inhibitory effect on elastase activity.



This work was supported by National Research Foundation of Korea grant funded by the Korea government (MEST: 2012R1A2A1A01001791 and WCU: R33-2008-000-10022-0). S Kim received support from the Konkuk University research support program.


  1. 1.
    Lewis EC, Shapiro L, Bowers OJ, Dinarello CA. (2005) Alpha1-antitrypsin monotherapy prolongs islet allograft survival in mice. Proc. Natl. Acad. Sci. U. S. A. 102:12153–8.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Song S, et al. (2004) Recombinant adeno-associated virus-mediated alpha-1 antitrypsin gene therapy prevents type I diabetes in NOD mice. Gene Ther. 11:181–6.CrossRefPubMedGoogle Scholar
  3. 3.
    Lu Y, et al. (2006) Alpha1-antitrypsin gene therapy modulates cellular immunity and efficiently prevents type 1 diabetes in nonobese diabetic mice. Hum. Gene Ther. 17:625–34.CrossRefPubMedGoogle Scholar
  4. 4.
    Shahaf G, et al. (2011) Alpha-1-antitrypsin gene delivery reduces inflammation, increases T-regulatory cell population size and prevents islet allograft rejection. Mol. Med. 17:1000–11.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Zhang B, et al. (2007) Alpha1-antitrypsin protects beta-cells from apoptosis. Diabetes. 56:1316–23.CrossRefPubMedGoogle Scholar
  6. 6.
    Lewis EC, et al. (2008) Alpha1-antitrypsin monotherapy induces immune tolerance during islet allograft transplantation in mice. Proc. Natl. Acad. Sci. U. S. A. 105:16236–41.CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Koulmanda M, et al. (2008) Curative and beta cell regenerative effects of alpha1-antitrypsin treatment in autoimmune diabetic NOD mice. Proc. Natl. Acad. Sci. U. S. A. 105:16242–7.CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Pileggi A, et al. (2008) Alpha-1 antitrypsin treatment of spontaneously diabetic nonobese diabetic mice receiving islet allografts. Transplant. Proc. 40:457–8.CrossRefPubMedGoogle Scholar
  9. 9.
    Lisowska-Myjak B, Pachecka J, Antoniewicz B, Krawczyk A, Jozwik A. (1995) Alpha-1-antitrypsin, albumin and whole protein in meconium and stools during the first days of life in the neonate [in Polish]. Pediatr. Pol. 70:819–26.PubMedGoogle Scholar
  10. 10.
    Sharma K, et al. (2005) Two-dimensional fluorescence difference gel electrophoresis analysis of the urine proteome in human diabetic nephropathy. Proteomics. 5:2648–55.CrossRefPubMedGoogle Scholar
  11. 11.
    Lisowska-Myjak B, Pachecka J. (2003) Antigenic and functional levels of alpha-1-antitrypsin in serum during normal and diabetic pregnancy. Eur. J. Obstet. Gynecol. Reprod. Biol. 106:31–5.CrossRefPubMedGoogle Scholar
  12. 12.
    Lisowska-Myjak B, Pachecka J, Kaczynska B, Miszkurka G, Kadziela K. (2006) Serum protease inhibitor concentrations and total antitrypsin activity in diabetic and non-diabetic children during adolescence. Acta Diabetol. 43:88–92.CrossRefPubMedGoogle Scholar
  13. 13.
    Hashemi M, Naderi M, Rashidi H, Ghavami S. (2007) Impaired activity of serum alpha-1-antitrypsin in diabetes mellitus. Diabetes Res. Clin. Pract. 75:246–8.CrossRefPubMedGoogle Scholar
  14. 14.
    Yaghmaei M, et al. (2009) Serum trypsin inhibitory capacity in normal pregnancy and gestational diabetes mellitus. Diabetes Res. Clin. Pract. 84:201–4.CrossRefPubMedGoogle Scholar
  15. 15.
    Kueppers F, Briscoe WA, Bearn AG. (1964) Hereditary deficiency of serum alpha-L-antitrypsin. Science. 146:1678–9.CrossRefPubMedGoogle Scholar
  16. 16.
    Eriksson S. (1964) Pulmonary emphysema and alpha1-antitrypsin deficiency. Acta. Med. Scand. 175:197–205.CrossRefPubMedGoogle Scholar
  17. 17.
    Blanco I, Lara B, de Serres F. (2011) Efficacy of alpha1-antitrypsin augmentation therapy in conditions other than pulmonary emphysema. Orphanet. J. Rare Dis. 6:14.CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Cox DW, Woo SL, Mansfield T. (1985) DNA restriction fragments associated with alpha 1-antitrypsin indicate a single origin for deficiency allele PI Z. Nature. 316:79–81.CrossRefPubMedGoogle Scholar
  19. 19.
    Carroll TP, et al. (2010) Evidence for unfolded protein response activation in monocytes from individuals with alpha-1 antitrypsin deficiency. J. Immunol. 184:4538–46.CrossRefPubMedGoogle Scholar
  20. 20.
    Nita I, Hollander C, Westin U, Janciauskiene SM. (2005) Prolastin, a pharmaceutical preparation of purified human alpha1-antitrypsin, blocks endotoxin-mediated cytokine release. Respir. Res. 6:12.CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Al-Omari M, et al. (2011) Acute-phase protein alpha1-antitrypsin inhibits neutrophil calpain I and induces random migration. Mol. Med. 17:865–74.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Subramaniyam D, et al. (2008) TNF-alpha-induced self expression in human lung endothelial cells is inhibited by native and oxidized alpha1-antitrypsin. Int. J. Biochem. Cell Biol. 40:258–71.CrossRefPubMedGoogle Scholar
  23. 23.
    Janciauskiene S, et al. (2008) Alpha1-antitrypsin inhibits the activity of the matriptase catalytic domain in vitro. Am. J. Respir. Cell Mol. Biol. 39:631–7.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Ryu CJ, Padlan EA, Jin BR, Yoo OJ, Hong HJ. (1996) A humanized antibody with specificity for hepatitis B surface antigen. Hum. Antibodies Hybridomas. 7:113–22.CrossRefPubMedGoogle Scholar
  25. 25.
    Eden E, et al. (1997) Atopy, asthma, and emphysema in patients with severe alpha-1-antitrypysin deficiency. Am. J. Respir. Crit. Care Med. 156:68–74.CrossRefPubMedGoogle Scholar
  26. 26.
    Griffith ME, Lovegrove JU, Gaskin G, Whitehouse DB, Pusey CD. (1996) C-antineutrophil cytoplasmic antibody positivity in vasculitis patients is associated with the Z allele of alpha-1-antitrypsin, and P-antineutrophil cytoplasmic antibody positivity with the S allele. Nephrol. Dial. Transplant. 11:438–43.CrossRefPubMedGoogle Scholar
  27. 27.
    King MA, et al. (1996) Alpha 1-antitrypsin deficiency: evaluation of bronchiectasis with CT. Radiology. 199:137–41.CrossRefPubMedGoogle Scholar
  28. 28.
    Ma H, et al. (2010) Intradermal alpha1-antitrypsin therapy avoids fatal anaphylaxis, prevents type 1 diabetes and reverses hyperglycaemia in the NOD mouse model of the disease. Diabetologia. 53:2198–204.CrossRefPubMedGoogle Scholar
  29. 29.
    Toldo S, et al. (2011) Alpha-1 antitrypsin inhibits caspase-1 and protects from acute myocardial ischemia-reperfusion injury. J. Mol. Cell. Cardiol. 51:244–51.CrossRefPubMedGoogle Scholar
  30. 30.
    Marcondes AM, et al. (2011) Inhibition of IL-32 activation by alpha-1 antitrypsin suppresses alloreactivity and increases survival in an allogeneic murine marrow transplantation model. Blood. 118:5031–9.CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Tawara I, et al. (2012) Alpha-1-antitrypsin monotherapy reduces graft-versus-host disease after experimental allogeneic bone marrow transplantation. Proc. Natl. Acad. Sci. U. S. A. 109:564–9.CrossRefPubMedGoogle Scholar
  32. 32.
    Kalis M, Kumar R, Janciauskiene S, Salehi A, Cilio CM. (2010) Alpha 1-antitrypsin enhances insulin secretion and prevents cytokine-mediated apoptosis in pancreatic beta-cells. Islets. 2:185–9.CrossRefPubMedGoogle Scholar
  33. 33.
    Kuttler B, et al. (2007) Ex vivo gene transfer of viral interleukin-10 to BB rat islets: no protection after transplantation to diabetic BB rats. J. Cell Mol. Med. 11:868–80.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Heywood DM, Mansfield MW, Grant PJ. (1996) Levels of von Willebrand factor, insulin resistance syndrome, and a common vWF gene polymorphism in non-insulin-dependent (type 2) diabetes mellitus. Diabet. Med. 13:720–5.CrossRefPubMedGoogle Scholar
  35. 35.
    Laakso M, Malkki M, Kekalainen P, Kuusisto J, Deeb SS. (1995) Polymorphisms of the human hexokinase II gene: lack of association with NIDDM and insulin resistance. Diabetologia. 38:617–22.CrossRefPubMedGoogle Scholar
  36. 36.
    Lisowska-Myjak B, Sygitowicz G, Wolf B, Pachecka J. (2001) Serum alpha-1-antitrypsin concentration during normal and diabetic pregnancy. Eur. J. Obstet. Gynecol. Reprod. Biol. 99:53–6.CrossRefPubMedGoogle Scholar
  37. 37.
    Sandstrom CS, et al. (2008) An association between type 2 diabetes and alpha-antitrypsin deficiency. Diabet. Med. 25:1370–3.PubMedGoogle Scholar
  38. 38.
    Talmud PJ, et al. (2003) Progression of atherosclerosis is associated with variation in the alpha1-antitrypsin gene. Arterioscler. Thromb. Vasc. Biol. 23:644–9.CrossRefPubMedGoogle Scholar

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Authors and Affiliations

  • Siyoung Lee
    • 1
  • Youngmin Lee
    • 2
  • Kwangwon Hong
    • 1
  • Jaewoo Hong
    • 1
  • Suyoung Bae
    • 1
  • Jida Choi
    • 1
  • Hyunjhung Jhun
    • 1
  • Areum Kwak
    • 1
  • Eunsom Kim
    • 1
  • Seunghyun Jo
    • 1
  • Charles A. Dinarello
    • 3
  • Soohyun Kim
    • 1
  1. 1.Laboratory of Cytokine Immunology, Department of Biomedical Sciences and TechnologyKonkuk UniversitySeoulKorea
  2. 2.Department of Medicine, Pusan Paik Hospital, College of MedicineInje UniversityBusanKorea
  3. 3.Department of MedicineUniversity of Colorado Denver, AuroraDenverUSA

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