Abstract
5-Bromo-2ʹ-deoxyuridine (BrdU) is a marker that is widely used to label S-phase cells in neurobiological research in most common doses 50 or 100 mg/kg per single intraperitoneal (i.p.) injection. However, the important data regarding its pharmacokinetics in rodents are still missing. The aim of our study was to investigate the BrdU level in serum after a single i.p. injection to adult rats (doses: 50 or 100 mg/kg) and adult mice (50 mg/kg). The animals were killed at selected time-points after the BrdU injection, and proliferating tumour cells (cell lines HCT-116 and HL-60) were co-cultivated with isolated blood sera. BrdU incorporated in the DNA of the S-phase tumour cells was stained with an anti-BrdU antibody and analysed using flow cytometry. In rats, the efficacies of BrdU labelling of S-phase cells in both in vitro and in vivo conditions were compared in the 50 and 100 mg/kg groups. According to our results, BrdU was in saturated concentration to label almost all S-phase cells for 60 min in both doses and was detectable in blood serum until 120 min after the single i.p. injection. However, the 100 mg/kg dose of BrdU did not provide a prolonged staining period to offset the potentially higher toxicity in comparison with the 50 mg/kg dose. In mice, due to their faster metabolism, the concentration of BrdU in blood serum was sufficient to label the whole population of S-phase cells for only 15 min after the i.p. injection, then dropped rapidly.
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Barker JM, Charlier TD, Ball GF, Balthazart J (2013) A new method for in vitro detection of bromodeoxyuridine in serum: a proof of concept in a songbird species, the canary. PLoS One 8. doi:10.1371/journal.pone.0063692
Cameron HA, McKay RDG (2001) Adult neurogenesis produces a large pool of new granule cells in the dentate gyrus. J Comp Neurol 435:406–417
Cooper-Kuhn CM, Kuhn HG (2002) Is it all DNA repair? methodological considerations for detecting neurogenesis in the adult brain. Dev Brain Res 134:13–21
Duque A, Rakic P (2011) Different effects of bromodeoxyuridine and [3h]thymidine incorporation into DNA on cell proliferation, position, and fate. J Neurosci 31:15205–15217. doi:10.1523/jneurosci.3092-11.2011
Eadie BD, Redila VA, Christie BR (2005) Voluntary exercise alters the cytoarchitecture of the adult dentate gyrus by increasing cellular proliferation, dendritic complexity, and spine density. J Comp Neurol 486:39–47
Hayes NL, Nowakowski RS (2000) Exploiting the dynamics of S-phase tracers in developing brain: interkinetic nuclear migration for cells entering versus leaving the S-phase. Dev Neurosci 22:44–55
Horner PJ et al (2000) Proliferation and differentiation of progenitor cells throughout the intact adult rat spinal cord. J Neurosci 20:2218–2228
Kolb B, Pedersen B, Ballermann M, Gibb R, Whishaw IQ (1999) Embryonic and postnatal injections of bromodeoxyuridine produce age-dependent morphological and behavioral abnormalities. J Neurosci 19:2337–2346
Kriss JP, Revesz L (1962) The distribution and fate of bromodeoxyuridine and bromodeoxycytidine in the mouse and rat. Cancer Res 22:254–265
Kriss JP, Maruyama Y, Tung LA, Bond SB, Revesz L (1963) The fate of 5-bromodeoxyuridine, 5-bromodeoxycytidine, and 5-iododeoxycytidine in man. Cancer Res 23:260–268
Kuan HY, Smith DE, Ensminger WD, Knol JA, DeRemer SJ, Yang Z, Stetson PL (1996) Regional pharmacokinetics of 5-bromo-2′-deoxyuridine and 5-fluorouracil in dogs: hepatic arterial versus portal venous infusions. Cancer Res 56:4724–4727
Kuhn HG, Cooper-Kuhn CM (2007) Bromodeoxyuridine and the detection of neurogenesis. Curr Pharm Biotechnol 8:127–131. doi:10.2174/138920107780906531
Mandyam CD, Harburg GC, Eisch AJ (2007) Determination of key aspects of precursor cell proliferation, cell cycle length and kinetics in the adult mouse subgranular zone. Neuroscience 146:108–122. doi:10.1016/j.neuroscience.2006.12.064
Mikeš J, Ševc J, Košuth J, Matiašová A, Daxnerová Z, Fedoročko P (2014) Flow cytometric method for estimation of 5-bromo-2′-deoxyuridine content in rat serum. Phys Res (in press)
Nowakowski RS, Rakic P (1974) Clearance rate of exogenous 3H thymidine from the plasma of pregnant rhesus monkeys. Cell Tissue Kinet 7:189–194
Nowakowski RS, Lewin SB, Miller MW (1989) Bromodeoxyuridine immunohistochemical determination of the lengths of the cell cycle and the DNA-synthetic phase for an anatomically defined population. J Neurocytol 18:311–318
Packard DS Jr, Menzies RA, Skalko RG (1973) Incorporation of thymidine and its analogue, bromodeoxyuridine, into embryos and maternal tissues of the mouse. Differentiation 1:397–404
Phuphanich S, Levin VA (1985) Bioavailability of bromodeoxyuridine in dogs and toxicity in rats. Cancer Res 45:2387–2389
Rubini JR, Cronkite EP, Bond VP, Fliedner TM (1960) The metabolism and fate of tritiated thymidine in man. J Clin Invest 39:909–918
Russo A et al (1984) Pharmacological evaluation of intravenous delivery of 5-bromodeoxyuridine to patients with brain tumors. Cancer Res 44:1702–1705
Sekerková G, Ilijic E, Mugnaini E (2004) Bromodeoxyuridine administered during neurogenesis of the projection neurons causes cerebellar defects in rat. J Comp Neurol 470:221–239. doi:10.1002/cne.11016
Ševc J, Daxnerová Z, Haňová V, Koval J (2011) Novel observations on the origin of ependymal cells in the ventricular zone of the rat spinal cord. Acta Histochem 113:156–162. doi:10.1016/j.acthis.2009.09.007
Staroscik RN, Jenkins WH, Mendelsohn ML (1964) Availability of tritiated thymidine after intravenous administration. Nature 202:456–458
Taupin P (2007) BrdU immunohistochemistry for studying adult neurogenesis: paradigms, pitfalls, limitations, and validation. Brain Res Rev 53:198–214. doi:10.1016/j.brainresrev.2006.08.002
Acknowledgments
This work was supported by the Scientific Grant Agency of the Ministry of Education of the Slovak Republic No. VEGA 1/0322/11 and 1/0967/12. The authors are grateful to Eva Pástorová and Viera Balážová for assistance with technical procedures.
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All the authors indicate no actual or potential conflict of interest including any financial, personal or other relationships with other people or organisations.
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Matiašová, A., Ševc, J., Mikeš, J. et al. Flow cytometric determination of 5-bromo-2ʹ-deoxyuridine pharmacokinetics in blood serum after intraperitoneal administration to rats and mice. Histochem Cell Biol 142, 703–712 (2014). https://doi.org/10.1007/s00418-014-1253-7
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DOI: https://doi.org/10.1007/s00418-014-1253-7