Mitochondrial Apoptotic Pathways: A Mechanism for Low Androgen-induced Vascular Endothelial Injury in Male Rats

Abstract

The aim of the study was to investigate the role of mitochondrial apoptotic pathways in vascular endothelial injury in male rats with low androgen. 8 week-old adult male Sprague-Dawley (SD) rats were randomly divided into 3 groups (n=6/each group): control group, castrated group (low androgen), and replacement group (given androgen after castration). After 10 weeks, endothelial structure was observed by general light microscope and transmission electron microscope (TEM) respectively. Isolated mitochondria and mitochondrial membrane potential (MMP) were detected by fluorescence to access mitochondrial function. Chromatin degradation was measured by terminal deoxynucleotidyl transferase-mediated deoxyuridine-biotin nick end labeling (TUNEL) staining method. The mRNA and protein of bcl-2, cytochrome C (Cyt C), caspase-9, and caspase-3 were analyzed for apoptosis. Cell shrinkage and condensed chromatin, less mitochondria and a fall in MMP levels were observed in the castrated group, along with more apoptotic endothelial cells. Bcl-2 level was reduced and the expression of caspase-9, caspase-3 and Cyt C were elevated in the castrated group (p<0.05). But there was no significant difference between the replacement group and the control group (p>0.05). It was concluded that low androgen caused vascular endothelial damage. It may be, at least in part, related with the activating mitochondrial apoptotic pathways.

Key words

low androgen - endothelial cell - mitochondria - apoptosis

References

• 1 Harman SM. Testosterone in older men after the Institute of Medicine Report: where do we go from here?.  Climacteric. 2005;  8 124-135
• 2 Wu FC, Tajar A, Pye SR, Silman AJ, Finn JD, O’Neill TW, Bartfai G, Casanueva F, Forti G, Giwercman A, Huhtaniemi IT, Kula K, Punab M, Boonen S, Vanderschueren D. Hypothalamic-pituitary-testicular axis disruptions in older men are differentially linked to age and modifiable risk factors: the European Male Aging study.  J Clin Endocrinol Metab. 2008;  93 2737-2745
• 3 Raynor MC, Carson CC, Pearson MD, Nix JW. Androgen deficiency in the aging male: a guide to diagnosis and testosterone replacement therapy.  Can J Urol. 2007;  14 63-68
• 4 Yeap BB, Hyde Z, Almeida OP, Norman PE, Chubb SAP, Jamrozik K, Flicker L, Hankey GJ. Lower testosterone levels predict incident stroke and transient ischemic attack in older men.  J Clin Endocrinol Metab. 2009;  94 2353-2359
• 5 Akishita M, Hashimoto M, Ohike Y, Ogawa S, Iijima K, Eto M, Ouchi Y. Low testosterone level is an independent determinant of endothelial dysfunction in men.  Hypertens Res. 2007;  30 1029-1034
• 6 Gimbrone Jr MA. Vascular endothelium: An integrator of pathophysiologic stimuli in atherosclerosis.  Am J Cardiol. 1995;  75 67B-70B
• 7 Schächinger V, Britten MB, Zeiher AM. Prognostic impact of coronary vasodilator dysfunction on adverse long-term outcome of coronary heart disease.  Circulation. 2000;  101 1899-1906
• 8 Hill JM, Zalos G, Halcox JPJ, Schenke WH, Waclawiw MA, Quyyumi AA, Finkel T. Circulating endothelial progenitor cells, vascular function and cardiovascular risk.  N. Engl J Med. 2003;  348 593-600
• 9 Lu YL, Kuang L, Zhu H, Wu H, Wang X-F, Pang Y-P, Wang N-J, Yu D-L. Changes in aortic endothelium ultrastructure in male rats following castration, replacement with testosterone and administration of 5alpha-reductase inhibitor.  Asian J Androl. 2007;  9 843-847
• 10 Desagher S, Martinou JC. Mitochondria as the central control point of apoptosis.  Cell Biology. 2000;  10 369-377
• 11 Bialik S, Cryns VL, Drincic A, Miyata S, Wollowick AL, Srinivasan A, Kitsis RN. The mitochondrial apoptotic pathway is activated by serum and glucose deprivation in cardiac myocytes.  Circ Res. 1999;  85 403-414
• 12 Oshitari T, Yamamoto S, Hata N, Roy S. Mitochondria- and caspase-dependent cell death pathway involved in neuronal degeneration in diabetic retinopathy.  Br J Ophthalmol. 2008;  92 552-556
• 13 Reers M, Smith TW, Chen LB. J-aggregate formation of a carbocyanine as a quantitative fluorescent indicator of membrane potential.  Biochemistry.. 1991;  30 4480-4486
• 14 Scragg JL, Dallas ML, Peers C. Molecular requirements for L-type Ca2+ channel blockade by testosterone.  Cell Calcium. 2007;  42 11-15
• 15 Seyrek M, Yildiz O, Ulusoy HB, Yildirim V. Testosterone relaxes isolated human radial artery by potassium channel opening action.  J Pharmacol Sci. 2007;  103 309-316
• 16 Kimura N, Mizokami A, Oonuma T, Sasano H, Nagura H. Immunocytochemical localization of androgen receptor with polyclonal antibody in paraffin-embedded human tissues.  J Histochem Cytochem. 1993;  41 671-618
• 17 Yu J, Akishita M, Eto M, Ogawa S, Son B-K, Kato S, Ouchi Y, Okabe T. Androgen Receptor-Dependent Activation of Endothelial Nitric Oxide Synthase in Vascular Endothelial Cells: Role of Phosphatidylinositol 3-Kinase/Akt Pathway.  Endocrinology. 2010;  151 1822-1828
• 18 Karsan A, Harlan JM. Modulation of endothelial cell apoptosis: mechanisms and pathophysiological roles.  J Atheroscler Thromb. 1996;  3 75-80
• 19 Mohamad N, Gutiérrez A, Núñez M, Cocca C, Martín G, Cricco G, Medina V, Rivera E, Bergoc R. Mitochondrial apoptotic pathways.  Biocell. 2005;  29 149-161
• 20 Mignotte B, Vayssiere J-L. Mitochondria and apoptosis.  Eur J Biochem. 1998;  252 1-15
• 21 Wong WW, Puthalakath H. Bcl-2 family proteins: the sentinels of the mitochondrial apoptosis pathway.  IUBMB Life. 2008;  60 390-397
• 22 Zhou H, Ma Y, Zhou Y, Liu Z, Wang K, Chen G. Effects of magnesium sulfate on neuron apoptosis and expression of caspase, bax and bcl-2 after cerebral ischemia-reperfusion injury.  Chin Med J. 2003;  116 1532-1534
• 23 Krajewski S, Krajewska M, Ellerby LM, Welsh K, Xie Z, Deveraux QL, Salvesen GS, Bredesen DE, Rosenthal RE, Fiskum G, Reed JC. Release of caspase-9 from mitochondria during neuronal apoptosis and cerebral ischemia.  Proc Natl Acad Sci USA. 1999;  96 5752-5757
• 24 Hou J, Chong ZZ, Shang YC, Maiese K. FOXO3a governs early and late apoptotic endothelial programs during elevated glucose through mitochondrial and caspase signaling.  Mol Cell. 2010;  321 194-206

Correspondence