Abstract
In the present study we aimed to establish an animal model of dexamethasone (DEX)-induced apoptosis in the thymus of rats. The degree of apoptosis was determined in the same animals at 6 and 11 h after a single administration of DEX (5 mg/kg, ip) by (a) in vivo biodistribution of the uptake of [123I]Annexin V, a biomarker of the early stages of apoptosis; (b) in vitro evaluation of the apoptotic index (percentage of number of apoptotic cells versus total number of cells) in the form of DNA fragmentation, on tissue sections using in situ oligo ligation (ISOL). ISOL demonstrated a 62- and 90-fold increase of apoptotic index at 6 and 11 h after DEX administration respectively, in the outer part of the thymic lobule (cortex) and a 25- and 54-fold increases in the inner part of the thymic lobule (medulla) in the corresponding treatment groups. In the biodistribution study, [123I]Annexin V uptake was significantly increased in the thymus of rats 11 h after DEX administration (by 1.3- to 1.4-fold) and significantly decreased at the 6-h time point. We conclude that the specificity of the apoptotic signal provided by isotopic methods in vivo would always require confirmation by complementary in vitro techniques that verify the assessment of ongoing apoptosis accurately.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ahmed SA, Sriranganathan N (1994) Differential effects of dexamethasone on the thymus and spleen: alterations in programmed cell death, lymphocyte subsets and activation of T cells. Immunopharmacology 28:55–66
Allen RT, Hunter WJ, Agrawal DK (1997) Morphological and biochemical characterization and analysis of apoptosis. J Pharmacol Toxicol Methods 37:215–228
Belkebir-Mesbah D, Bonnefont-Rousselot D, Frey-Fressart V, Moinard C, Delattre J, Vasson M-P (1999) Consequences of treatment with dexamethasone in rats on the susceptibility of total plasma and isolated lipoprotein fractions to copper oxidation. Endocrine 10:233–242
Blankenberg FG, Strauss HW (2001) Will imaging of apoptosis play a role in clinical care? A tale of mice and men. Apoptosis 6:117–123
Blankenberg FG, Katsikis PD, Tait JF, Davis E, Naumovski L, Ohtsuki K, Kopiwoda S, Abrams MJ, Darkes M, Robbins RC, Maecker HT, Strauss HW (1998) In vivo detection and imaging of phosphatidylserine expression during programmed cell death. Proc Natl Acad Sci USA 95:6349–6354
Claman HN (1972) Corticosteroids and lymphoid cells. N Engl J Med 287:388–397
Compton MM, Cidlowski JA (1986) Rapid in vivo effects of glucocorticoids in the integrity of rat lymphocyte genomic deoxyribonucleic acid. Endocrinology 118:38–45
Didenko VV, Hornsby PJ (1999) Presence of double-strand breaks with single-base 3′ overhangs in cells undergoing apoptosis but not necrosis. J Cell Biol 26:1251–1258
Gavrieli Y, Sherman Y, Ben-Sasson SA (1992) Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol 119:493–501
Green AM, Steinmetz ND (2002) Monitoring apoptosis in real time. Cancer J 8:82–92
Ichiyoshi H, Kiyozuka Y, Kishimoto Y, Fukuhara S, Tsubura A (2003) Massive telomere loss and telomerase RNA expression in dexamethasone-induced apoptosis in mouse thymocytes. Exp Mol Pathol 75:178–186
Keen HG, Dekker BA, Disley L, Hastings D, Lyons S, Reader AJ, Ottewell P, Watson A, Zweit J (2005) Imaging apoptosis in vivo using 124-annexin V and PET. Nucl Med Biol 32:395–402
Kerr JFR, Wyllie AH, Currie AR (1972) Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer 26:239–257
Lahorte C, Slegers G, Philippe J, van de Wiele C, Dierckx RA (2001) Synthesis and in vitro evaluation of 123I-labelled human recombinant annexin V. Biomol Eng 17:51–53
Lahorte CMM, Vanderheyden J-L, Steinmetz N, van de Wiele C, Dierckx RA, Slegers G (2004) Apoptosis-detecting radioligands: current state of the art and future perspectives. Eur J Nucl Med Mol Imaging 31:887–919
Mann CL, Cidlowski JA (2001) Glucocorticoids regulate plasma membrane potential during rat thymocyte apoptosis in vivo and in vitro. Endocrinology 142:421–429
Martin SJ, Reutelinsperger CPM, McGahon AJ, Rader JA, Van Schie RC, Laface DM, Green DR (1995) Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl-2 and Abl. J Exp Methods 182:1545–1556
Ohtsuki K, Akashi K, Aoka Y, Blankenberg FG, Kopiwoda S, Tait JF, Strauss HW (1999) Technetium-99m HYNIC-annexin V: a potential radiopharmaceutical for the in vivo detection of apoptosis. Eur J Nucl Med 26:1251–1258
Proskuryakov SY, Konoplyannikov AG, Gabai VL (2003) Necrosis: a specific form of programmed cell death? Exp Cell Res 283:1–16
Saraste A, Pulkki K (2000) Morphologic and biochemical hallmarks of apoptosis. Cardiovasc Res 45:528–537
Schimmer AD, Welsh K, Pinilla C, Wang Z, Krajewska M, Bonneau MJ, Pedersen IM, Kitada S, Scott FL, Bailly-Maitre B, Glinsky G, Scudiero D, Sausville E, Salvesen G, Nefzi A, Ostresh JM, Houghten RA, Reed JC (2004) Small-molecule antagonists of apoptosis suppressor XIAP exhibit broad antitumor activity. Cancer Cell 5:25–35
Sun X-M, Dinsdale D, Snowden RT, Cohen GM, Skilleter DN (1992) Characterisation of apoptosis in thymocytes isolated from dexamethasone-treated rats. Biochem Pharmacol 44:2131–2137
Surh CD, Sprent J (1994) T-cell apoptosis detected in situ during positive and negative selection in the thymus. Nature 372:100–103
van den Eijnde SM, Boshart L, Reutelingsperger CPM, De Zeeuw CI, Vermeij-Keers C (1997) Phosphatidylserine plasma membrane asymmetry in vivo: a pancellular phenomenon which alters during apoptosis. Cell Death Differ 4:311–316
Acknowledgements
The authors wish to thank Filomena Mattner, Paula Berghofer and Emma Millard, for assistance on the day of the biodistribution experiment.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Zavitsanou, K., Nguyen, V., Greguric, I. et al. Detection of apoptotic cell death in the thymus of dexamethasone treated rats using [123I]Annexin V and in situ oligonucleotide ligation. J Mol Hist 38, 313–319 (2007). https://doi.org/10.1007/s10735-007-9104-7
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10735-007-9104-7