Generating Double Knockout Mice to Model Genetic Intervention for Diabetic Cardiomyopathy in Humans
Diabetes is a rapidly increasing disease that enhances the chances of heart failure twofold to fourfold (as compared to age and sex matched nondiabetics) and becomes a leading cause of morbidity and mortality. There are two broad classifications of diabetes: type1 diabetes (T1D) and type2 diabetes (T2D). Several mice models mimic both T1D and T2D in humans. However, the genetic intervention to ameliorate diabetic cardiomyopathy in these mice often requires creating double knockout (DKO). In order to assess the therapeutic potential of a gene, that specific gene is either overexpressed (transgenic expression) or abrogated (knockout) in the diabetic mice. If the genetic mice model for diabetes is used, it is necessary to create DKO with transgenic/knockout of the target gene to investigate the specific role of that gene in pathological cardiac remodeling in diabetics. One of the important genes involved in extracellular matrix (ECM) remodeling in diabetes is matrix metalloproteinase-9 (Mmp9). Mmp9 is a collagenase that remains latent in healthy hearts but induced in diabetic hearts. Activated Mmp9 degrades extracellular matrix (ECM) and increases matrix turnover causing cardiac fibrosis that leads to heart failure. Insulin2 mutant (Ins2+/−) Akita is a genetic model for T1D that becomes diabetic spontaneously at the age of 3–4 weeks and show robust hyperglycemia at the age of 10–12 weeks. It is a chronic model of T1D. In Ins2+/− Akita, Mmp9 is induced. To investigate the specific role of Mmp9 in diabetic hearts, it is necessary to create diabetic mice where Mmp9 gene is deleted. Here, we describe the method to generate Ins2+/−/Mmp9−/− (DKO) mice to determine whether the abrogation of Mmp9 ameliorates diabetic cardiomyopathy.
Key wordsIns2+/− Akita Mmp9 Diabetes Heart failure Cardiomyopathy
This work is supported by National Institute of Health grants HL-113281 and HL116205 to Paras Kumar Mishra.
- 3.Mathew V, Gersh BJ, Williams BA, Laskey WK, Willerson JT, Tilbury RT, Davis BR, Holmes DR Jr (2004) Outcomes in patients with diabetes mellitus undergoing percutaneous coronary intervention in the current era: a report from the Prevention of REStenosis with Tranilast and its Outcomes (PRESTO) trial. Circulation 109:476–480PubMedCrossRefGoogle Scholar
- 4.Pignone M, Alberts MJ, Colwell JA, Cushman M, Inzucchi SE, Mukherjee D, Rosenson RS, Williams CD, Wilson PW, Kirkman MS (2010) Aspirin for primary prevention of cardiovascular events in people with diabetes: a position statement of the American Diabetes Association, a scientific statement of the American Heart Association, and an expert consensus document of the American College of Cardiology Foundation. Diabetes Care 33:1395–1402PubMedCentralPubMedCrossRefGoogle Scholar
- 5.Rota M, LeCapitaine N, Hosoda T, Boni A, De AA, Padin-Iruegas ME, Esposito G, Vitale S, Urbanek K, Casarsa C, Giorgio M, Luscher TF, Pelicci PG, Anversa P, Leri A, Kajstura J (2006) Diabetes promotes cardiac stem cell aging and heart failure, which are prevented by deletion of the p66shc gene. Circ Res 99:42–52PubMedCrossRefGoogle Scholar
- 6.Sarwar N, Gao P, Seshasai SR, Gobin R, Kaptoge S, Di AE, Ingelsson E, Lawlor DA, Selvin E, Stampfer M, Stehouwer CD, Lewington S, Pennells L, Thompson A, Sattar N, White IR, Ray KK, Danesh J (2010) Diabetes mellitus, fasting blood glucose concentration, and risk of vascular disease: a collaborative meta-analysis of 102 prospective studies. Lancet 375:2215–2222PubMedCrossRefGoogle Scholar
- 7.Schramm TK, Gislason GH, Kober L, Rasmussen S, Rasmussen JN, Abildstrom SZ, Hansen ML, Folke F, Buch P, Madsen M, Vaag A, Torp-Pedersen C (2008) Diabetes patients requiring glucose-lowering therapy and nondiabetics with a prior myocardial infarction carry the same cardiovascular risk: a population study of 3.3 million people. Circulation 117:1945–1954PubMedCrossRefGoogle Scholar
- 11.Stoy J, Edghill EL, Flanagan SE, Ye H, Paz VP, Pluzhnikov A, Below JE, Hayes MG, Cox NJ, Lipkind GM, Lipton RB, Greeley SA, Patch AM, Ellard S, Steiner DF, Hattersley AT, Philipson LH, Bell GI (2007) Insulin gene mutations as a cause of permanent neonatal diabetes. Proc Natl Acad Sci U S A 104:15040–15044PubMedCentralPubMedCrossRefGoogle Scholar
- 13.Simsek DG, Aycan Z, Ozen S, Cetinkaya S, Kara C, Abali S, Demir K, Tunc O, Ucakturk A, Asar G, Bas F, Cetinkaya E, Aydin M, Karaguzel G, Orbak Z, Siklar Z, Altincik A, Okten A, Ozkan B, Ocal G, Semiz S, Arslanoglu I, Evliyaoglu O, Bundak R, Darcan S (2013) Diabetes care, glycemic control, complications, and concomitant autoimmune diseases in children with type 1 diabetes in Turkey: a multicenter study. J Clin Res Pediatr Endocrinol 5:20–26PubMedCentralPubMedCrossRefGoogle Scholar
- 19.Barengo NC, Trejo R, Sposetti G (2013) Prevalence of type 2 diabetes in Argentina 1979-2012. Diabetes Metab Res Rev, Epub. July, 16Google Scholar
- 27.Buralli S, Dini FL, Ballo P, Conti U, Fontanive P, Duranti E, Metelli MR, Marzilli M, Taddei S (2010) Circulating matrix metalloproteinase-3 and metalloproteinase-9 and tissue Doppler measures of diastolic dysfunction to risk stratify patients with systolic heart failure. Am J Cardiol 105:853–856PubMedCrossRefGoogle Scholar
- 29.Ducharme A, Frantz S, Aikawa M, Rabkin E, Lindsey M, Rohde LE, Schoen FJ, Kelly RA, Werb Z, Libby P, Lee RT (2000) Targeted deletion of matrix metalloproteinase-9 attenuates left ventricular enlargement and collagen accumulation after experimental myocardial infarction. J Clin Invest 106:55–62PubMedCentralPubMedCrossRefGoogle Scholar
- 30.Lindsey ML, Escobar GP, Dobrucki LW, Goshorn DK, Bouges S, Mingoia JT, McClister DM Jr, Su H, Gannon J, MacGillivray C, Lee RT, Sinusas AJ, Spinale FG (2006) Matrix metalloproteinase-9 gene deletion facilitates angiogenesis after myocardial infarction. Am J Physiol Heart Circ Physiol 290:H232–H239PubMedCrossRefGoogle Scholar