מנהל המערך הקרדיולוגי- פרופ' גד קרן,M.D.
מנהלת המעבדה- מיכל אנטין, PhD / Cardiology division
Head of Cardiology division– Gad Keren, M.D.
Lab manager– Michal Entin-Meer, Ph.D.
1. Myocardial infarction (MI) induction in Rats:
performed by permanent ligation of the left anterior descending coronary artery (LAD). Brief protocol: Animals are anesthetized with Ketamine and Xylazine, orally intubated and artificially ventilated with a respirator. A small oblique thoracotomy is performed lateral to the midsternal line in the third costal space to expose the heart. After the pericardium is opened, the proximal left anterior descending artery (LAD) branch of the left coronary artery is legated. Echocardiography is test using a pediatric probe (15MHz) [1].
2. Cardiorenal syndrome:
The model is based on the experience gathered in Prof Gad Keren laboratory for induction of myocardial infarction, and in the laboratory of Prof. Doron Schwartz from the Nephrology department at the Sourasky Medical center for induction of renal insufficiency [2].
Briefly, male Wister rats undergo 5/6 kidney removal followed by LAD ligation 3 week later. This model may be successfully applied for assessing safety and therapeutic potential of novel drugs for patients suffering from combined cardio-renal illnesses. Alternatively, the model can be used for assessment of gene therapy, such as microRNA administration.
3. Experimental autoimmune myocarditis (EAM):
induced by injection of 1 mg of porcine cardiac myosin dissolved in an equal volume of Complete Freud’s adjuvant containing 10 mg/ml heat-killed mycobacterium tuberculosis to their foot pads [3-6]. EAM , if left untreated, progresses toward dilated cardiomyopathy.
4. In vivo matrigel assays for studying angiogenesis [7-8]:
Briefly, cells (2 X 106) in 50 ml of PBS are mixed with 400 ml of liquid Matrigel and are subcutaneously injected into mice (n = 5). After 8 days, mice are being sacrificed, and 5-mm-thick frozen sections are being prepared from the excised plugs. Immunohistochemical assessment of vascular density us then performed by CD31 and hematoxylin co-staining. Quantitative analysis is performed under a light microscope (X20) in high-power fields (n = 10), employing Image J software (identifiable vessel number ! mean vessel area). Additional sections are stained with DAPI, and optical fields (n = 5) are analyzed by fluorescence microscopy (X20 and X40).
5. Various ex-vivo and in vitro techniques:
Organ isolation, advance use of flow cytometry, tissue culture including: viral infection, and various assays such as: adhesion, migration, and apoptosis [7-11]; Immuno-assays including identification of regulatory T cells [12-13]; access to clinical samples of cardiac patients.
Refrenses:
1. George, J., Tyrphostin AG-556 reduces myocardial infarct size and improves cardiac performance in the rat. Exp Mol Pathol, 2003. 74(3): p. 314-8.
2. Schwartz, D., Glomerular arginine transport is attenuated in chronic renal failure in rats. J Med, 2003. 34(1-6): p. 113-20.
3. George, J., Suppression of myosin-induced and adoptively transferred myocarditis by prior treatment with complete Freund's adjuvant. Cardiovasc Pathol, 2004. 13(4): p. 221-4.
4. George, J., The effect of early and late treatment with the tyrphostin AG-556 on the progression of experimental autoimmune myocarditis. Exp Mol Pathol, 2004. 76(3): p. 234-41.
5. George, J., The effect of intravenous immunoglobulins on the progression of experimental autoimmune myocarditis in the rat. Exp Mol Pathol, 2001. 71(1): p. 55-62.
6. Pando, R., The Ras antagonist farnesylthiosalicylic acid ameliorates experimental myocarditis in the rat. Cardiovasc Pathol, 2009.
7. Barzelay, A., A potential role for islet-1 in post-natal angiogenesis and vasculogenesis. Thromb Haemost, 2010. 103(1): p. 188-97.
8. Ben-Shoshan, J., Hypoxia-inducible factor-1alpha and -2alpha additively promote endothelial vasculogenic properties. J Vasc Res, 2009. 46(4): p. 299-310.
9. Ben-Shoshan, J., Endothelial progenitor cells as therapeutic vectors in cardiovascular disorders: from experimental models to human trials. Pharmacol Ther, 2007. 115(1): p. 25-36.
10. Ben-Shoshan, J., Hypoxia controls CD4+CD25+ regulatory T-cell homeostasis via hypoxia-inducible factor-1alpha. Eur J Immunol, 2008. 38(9): p. 2412-8.
11. Ben-Shoshan, J., Constitutive expression of HIF-1alpha and HIF-2alpha in bone marrow stromal cells differentially promotes their proangiogenic properties. Stem Cells, 2008. 26(10): p. 2634-43.
12. Mor, A., Altered status of CD4(+)CD25(+) regulatory T cells in patients with acute coronary syndromes. Eur Heart J, 2006. 27(21): p. 2530-7.
13. Mor, A., Role of naturally occurring CD4+ CD25+ regulatory T cells in experimental atherosclerosis. Arterioscler Thromb Vasc Biol, 2007. 27(4): p. 893-900.
For further information:
Gad Keren, M.D..
Cardiology division
Head of Cardiology division
6 Weizmann St, 64239, ISRAEL
Tel: 972-3-6974762
email:
Michal Entin-Meer, Ph.D .
Cardiology division
Lab manager
6 Weizmann St, 64239, ISRAEL
Tel: 972-3-6974762
email: