[page 12] [Geriatric Care 2016; 2:5722] The effect of erythropoietin on calcium levels during hypoxia reoxygenation injury in rats Constantinos Tsompos, 1 Constantinos Panoulis, 2 Konstantinos Toutouzas, 3 Aggeliki Triantafyllou, 4 George Zografos, 3 Apostolos Papalois 5 1 Department of Obstetrics & Gynecology, Mesologi County Hospital, Etoloakarnania; 2 Department of Obstetrics & Gynecology, Aretaieion Hospital, Athens University, Attiki; 3 Department of Surgery, Ippokrateion General Hospital, Athens University, Attiki; 4 Department of Biologic Chemistry, Athens University, Attiki; 5 Experimental Research Centre ELPEN Pharmaceuticals, S.A. Inc., Co. Attiki, Greece Abstract This experimental study examined the effect of erythropoietin (Epo) on rat model and partic- ularly in a hypoxia-reoxygenation protocol. The effect of that molecule was studied biochemical- ly using blood mean calcium levels (Ca ++ ). Forty rats of mean weight 247.7 g were used in the study. Ca ++ levels were measured at 60 min (groups A and C) and at 120 min (groups B and D) of reoxygenation. Erythropoietin was admin- istered only in groups C and D. Epo administra- tion non-significantly decreased the Ca ++ levels by 0.56%±1.13% (P=0.5761). Reoxygenation time non-significantly increased the Ca ++ levels by 0.65%±1.12% (P=0.5281). However, Epo administration and reoxygenation time togeth- er non-significantly decreased the Ca ++ levels by 0.34%±0.68% (P=0.6095). Epo administra- tion whether it interacted or not with reoxy- genation time had non-significant decreasing short-term effects on calcium levels. Perhaps, a longer study time than 2 h or a higher Epo dose may reveal more significant effects. Introduction Permanent or transient damage with seri- ous implications on adjacent organs and cer- tainly on patients’ health may be due to tissue hypoxia and reoxygenation (HR). Although important progress has been made regarding the usage of erythropoietin (Epo) in managing this kind of damages, satisfactory answers have not been given yet to fundamental ques- tions, as, by what velocity this factor acts, when should it be administered and at what dosage. The particularly satisfactory action of Epo in stem blood cells recovery has been noted in several performed experiments. However, just few relative reports were found concerning Epo trial in ischemia reperfusion (IR) experiments, not covering completely this particular matter. A meta-analysis of 23 pub- lished seric variables, 1-4 coming from the same experimental setting, tried to provide a numer- ic evaluation of the Epo efficacy at the same endpoints (Table 1). Furthermore, several pub- lications addressed trials of other similar mol- ecules of growth factors, which the studied molecule also belongs to. The aim of this experimental study was to examine the effect of Epo on rat model and particularly in an HR protocol. The effect of that molecule was studied by measuring the blood mean calcium (Ca ++ ) levels. Hypocalcemia although seriously underdiag- nosed 5 in geriatric units may be deteriorated by Epo administration. Materials and Methods Animal preparation This experimental study was licensed by Veterinary Address of East Attiki Prefecture under 3693/12-11-2010 and 14/10-1-2012 deci- sions. All consumables, equipment and sub- stances, were a courtesy of Experimental Research Centre of ELPEN Pharmaceuticals Co. Inc. S.A. at Pikermi, Attiki. Accepted stan- dards of humane animal care were adopted for Albino female Wistar rats. Pre-experimental normal housing in laboratory for 7 days includ- ed ad libitum diet. Post-experimental awaken- ing and preservation of the rodents was not permitted, even if euthanasia was needed. They were randomly delivered to four experi- mental groups by 10 animals in each one. Hypoxia for 45 min followed by reoxygenation for 60 min (group A). Hypoxia for 45 min fol- lowed by reoxygenation for 120 min (group B). Hypoxia for 45 min followed by immediate Epo intravenous (IV) administration and reoxy- genation for 60 min (group C). Hypoxia for 45 min followed by immediate Epo IV administra- tion and reoxygenation for 120 min (group D). The molecule Epo dosage was 10 mg/kg body weight of animals. Prenarcosis preceded of continuous intra- experimental general anesthesia, oxygen supply, electrocardiogram and acidometry of animals. 1-4 The protocol of HR was followed. Hypoxia was caused by laparotomic forceps clamping inferior aorta over renal arteries for 45 min. Reoxygenation was induced by removing the clamp and reestablishment of inferior aorta patency. The molecules were administered at the time of reoxygenation, through catheter- ized inferior vena cava. The Ca ++ levels meas- urements were performed at 60 min of reoxy- genation (for groups A and C) and at 120 min of reoxygenation (for groups B and D). The mean weight of the forty (40) female Wistar albino rats used was 247.7 g [standard devia- tion (Std. Dev): 34.99172 g], with min weight ≥165 g and max weight ≤320 g. Rats’ weight could be potentially a confusing factor, e.g., the more obese rats to have higher Ca ++ levels. This assumption was investigated. Model of hypoxia-reoxygenation injury Control groups Twenty control rats [mean mass 252.5 g (Std. Dev: 39.31988 g)] experienced hypoxia for 45 min followed by reoxygenation. Group A Reoxygenation lasted for 60 min (n=10 con- trols rats) mean mass 243 g (Std. Dev: 45.77724 g), mean Ca ++ levels 10.53 mg/dL (Std. Dev: 0.3465705 mg/dL) (Table 2). Group B Reoxygenation lasted for 120 min (n=10 controls rats) mean mass 262 g (Std. Dev: Geriatric Care 2016; volume 2:5722 Correspondence: Tsompos Constantinos, Department of Obstetrics & Gynecology, Mesologi County Hospital, Nafpaktou street, Mesologi 30200, Etoloakarnania, Greece. Tel.: +302631360237 / +306946674264 - Fax: +302106811215. E-mail: tsomposconstantinos@gmail.com Key words: Hypoxia; erythropoietin; calcium; reoxygenation; hypocalcemia; dyscalcemia; hypercalcemia. Acknowledgments: this study was funded by Scholarship by the Experimental Research Center ELPEN Pharmaceuticals (E.R.C.E), Athens, Greece. The research facilities for this project were provided by the aforementioned institution. Received for publication: 31 December 2015. Revision received: 17 February 2016. Accepted for publication: 24 March 2016. This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License (CC BY-NC 4.0). ©Copyright C. Tsompos et al., 2016 Licensee PAGEPress, Italy Geriatric Care 2016; 2:5722 doi:10.4081/gc.2016.5722 Non-commercial use only