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 Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 30  |  Issue : 3  |  Page : 141-145

The effect of the antioxidant drug "U-74389G" on amylase during ischemia reperfusion injury in rats


1 Department of Obstetrics and Gynecology, Mesologi County Hospital, Etoloakarnania, Greece
2 Department of Obstetrics and Gynecology, Aretaeio Hospital, Athens University, Attiki, Greece
3 Department of Surgery, Ippokrateio General Hospital, Athens University, Attiki, Greece
4 Department of Biologic Chemistry, Athens University, Medical School, Attiki, Greece
5 Exprerimental Research Center, ELPEN Pharmaceuticals, Co. Inc S.A., Pikermi, Attiki, Greece

Date of Web Publication28-Sep-2016

Correspondence Address:
Tsompos Constantinos
Department of Obstetrics and Gynecology, Mesologi County Hospital, Nafpaktou Street, Mesologi 30200, Etoloakarnania
Greece
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0972-4958.191177

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  Abstract 

Background: This experimental study examined the effect of the antioxidant drug "U-74389G" on a rat model and particularly in an ischemia-reperfusion protocol. Settings and Design: The beneficial effect or noneffectiveness of that molecule was studied biochemically using blood mean amylase (A) levels. Materials and Methods: Forty rats of mean weight 231.875 g were used in the study. The levels were measured at 60 min of reperfusion (Groups A and C) and at 120 min of reperfusion (Groups B and D) with the administration of the drug U-74389G in Groups C and D. Results: U-74389G administration significantly decreased the predicted A levels by 8.40 ± 2.02% (P = 0.0001). Reperfusion time kept nonsignificantly increased the predicted A levels by 1.68 ± 2.44% (P = 0.4103). However, U-74389G administration and reperfusion time together produced a significant combined effect in keeping decreased the predicted A levels by 4.67 ± 1.26% (P = 0.0005). Conclusions: U-74389G administration whether it interacted or not with reperfusion time has a significant decreasing beneficial restoring short-term effect on amylase levels.

Keywords: Amylase, ischemia, reperfusion, U-74389G


How to cite this article:
Constantinos T, Constantinos P, Konstantinos T, Aggeliki T, George Z, Apostolos P. The effect of the antioxidant drug "U-74389G" on amylase during ischemia reperfusion injury in rats. J Med Soc 2016;30:141-5

How to cite this URL:
Constantinos T, Constantinos P, Konstantinos T, Aggeliki T, George Z, Apostolos P. The effect of the antioxidant drug "U-74389G" on amylase during ischemia reperfusion injury in rats. J Med Soc [serial online] 2016 [cited 2020 Oct 29];30:141-5. Available from: https://www.jmedsoc.org/text.asp?2016/30/3/141/191177


  Introduction Top


Permanent or transient damage with serious implications on adjacent organs and systems may be due to tissue ischemia-reperfusion (IR). The use of U-74389G in IR has been a challenge for many years. However, although the progress was significant, several practical questions have not clarified. They include: (a) How potent U-74389G should be, (b) when should it be administered, and (c) at what optimal dose U-74389G should be administered. The promising effect of U-74389G in tissue protection has been noted in several IR studies. U-74389G or also known as 21-[4-(2,6-di-1-pyrrolidinyl-4-pyrimidinyl)-1-piperazinyl]-pregna-1, 4, 9 (11)-triene-3,20-dione maleate salt is an antioxidant which prevents both arachidonic acid-induced and iron-dependent lipid peroxidation. [1] It protects against IR injury in animal heart, liver, and kidney models. These membrane-associating antioxidants are particularly effective in preventing permeability changes in brain microvascular endothelial cells monolayers. [2] A meta-analysis of 15 published seric variables coming from the same experimental setting tried to provide a numeric evaluation of the U-74389G efficacy at the same endpoints [Table 1]. Several publications addressed the trials of other similar antioxidant molecules to which the studied molecule U-74389G belongs to.
Table 1: The U-74389G influence (±standard deviation) on the levels of some seric variables concerning reperfusion time

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The aim of this experimental study was to examine the effect of U-74389G on the rat model and particularly in a pancreas IR protocol. The beneficial effect or noneffectiveness of that molecule was studied by measuring the blood mean amylase (A) levels, fully complied with SMART criteria. The time variable of SMART criteria concerned the mentioned hours since every pre-/post-treatment and a measurement was performed on the same day.


  Materials and methods Top


Animal preparation

This basic experimental research was licensed by Veterinary Address of East Attiki Prefecture under 3693/12-11-2010 and 14/10-1-2012 decisions. Every consumable, equipment and substance used, was a grant of Experimental Research Centre of ELPEN Pharmaceuticals Co., Inc., S.A. at Pikermi, Attiki. Accepted standards of humane animal care were adopted for Albino female Wistar rats. Normal housing included ad libitum diet in laboratory 7 days before the experiment. Postexperimental awakening and preservation of animals were not permitted. Rats were randomly delivered to four experimental groups by ten animals in each one using following protocols of IR: Ischemia for 45 min followed by reperfusion for 60 min (Group A), ischemia for 45 min followed by reperfusion for 120 min (Group B), ischemia for 45 min followed by immediate U-74389G intravenous (IV) administration and reperfusion for 60 min (Group C), and ischemia for 45 min followed by immediate U-74389G IV administration and reperfusion for 120 min (Group D). The dose height of 10 mg/kg was determined by related experiments. Flessas et al. improved the histological score in intestinal tissue after U-74389G administration in pigs. [3] Bimpis et al. neuroprotected the brain itself from strokes by U-74389G administration in a porcine model. [4] Tsaroucha et al. attenuated the liver damage after U-74389G administration in a swine model. [5] Andreadou et al. protected the rat small intestine from oxidative damage after the administration of U-74389G. [6]

The detailed preceded prenarcotic and general anesthesiologic techniques are described in related references. [7],[8] Oxygen supply, electrocardiogram, and acidimetry were continuously provided during whole experiment performance. The protocol of IR was followed. Ischemia was caused by laparotomic clamping inferior aorta over renal arteries with forceps for 45 min. Reperfusion was induced by removing the clamp and reestablishment of inferior aorta patency. After exclusion of a blood flow, the protocol of IR was applied as described above for each of the experimental group. U-74389G was administered at the time of reperfusion through catheterized inferior vena cava. The A levels were determined at 60 th min of reperfusion (for A and C groups) and at 120 th min of reperfusion (for B and D groups). Forty female Wistar albino rats were used (mean weight 231.875 g (standard deviation [SD]: 36.59703 g], with minimum weight 165 g and maximum weight 320 g. Rats' weight could be potentially a confusing factor, for example, more obese rats to have higher A levels. This suspicion was also investigated.

Model of ischemia-reperfusion injury

Control groups


Twenty control rats (mean mass 252.5 g [SD: 39.31988 g]) suffered by ischemia for 45 min followed by reperfusion.

Group A: Reperfusion lasted for 60 min (n = 10 controls rats) mean mass 243 g (SD: 45.77724 g), mean A levels 1674 IU/L (SD: 282.0162 IU/L) [Table 2].
Table 2: Weight and amylase mean levels and standard deviation of groups

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Group B: Reperfusion lasted for 120 min (n = 10 controls rats) mean mass 262 g (SD: 31.10913 g), mean A levels 1877 IU/L (SD: 354.5749 IU/L) [Table 2].

Lazaroid (L) group

Twenty L rats (mean mass 211.25 g [SD: 17.53755 g]) suffered by ischemia for 45 min followed by reperfusion in the beginning of which 10 mg U-74389G/kg body weight were IV administered.

Group C: Reperfusion lasted for 60 min (n = 10 L rats) mean mass 212.5 g (SD: 17.83411 g), mean A levels 1514.1 IU/L (SD: 172.8303 IU/L) [Table 2].

Group D: Reperfusion lasted for 120 min (n = 10 L rats) mean mass 210 g (SD: 18.10463 g), mean A levels 1444.7 IU/L (SD: 102.79 IU/L) [Table 2].

Statistical analysis

Rats of each group were compared by weight and A level with each other by statistical paired t-tests [Table 3]. Any significant difference among A levels was investigated whether owed in potent significant weight correlations. The application of generalized linear models (glms) with dependent variable the A levels and independent variables the U-74389G or no drug, the reperfusion time, and both variables in combination was followed. Inserting the rats weight also as an independent variable at glms analysis, a very significant relation results in (P = 0.0074), so as to further investigation was needed. The predicted A values adjusted for rats weight were calculated and are depicted at [Table 4]. The differences between predicted mean A values as calculated by paired t-tests are depicted at [Table 5]. A second application of glms with dependent variable the predicted A levels and independent variables the U-74389G administration or no, the reperfusion time and their interaction was followed.
Table 3: Statistical significance of mean values difference for groups after statistical paired t-test application

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Table 4: Predicted mean amylase levels and standard deviation of groups

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Table 5: Statistical significance of predicted mean amylase values difference for groups after statistical paired t-test application

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  Results Top


The first glm resulted in U-74389G administration significantly decreased the A levels by 296.1 IU/l (−458.3629-−133.8371 IU/L] (P = 0.0007). This finding was in accordance with the results of paired t-test (P = 0.0011). Reperfusion time kept nonsignificantly increased the A levels by 66.8 IU/L (−121.0922-254.6922 IU/L] (P = 0.4761) in accordance also with paired t-test (P = 0.4302). However, U-74389G administration and reperfusion time together produced a significant combined effect in keeping decreased the A levels by 174.1273 IU/L [−272.8323-−75.42225 IU/L] (P = 0.0010). Reviewing the above and [Table 3], [Table 6] sums up concerning the decreasing influence of U-74389G along with reperfusion time. The second glm resulted in U-74389G administration significantly decreased the predicted A levels by 137.2091 IU/L [−202.0351-−72.38315 IU/L] (P = 0.0001). This finding was in accordance with the results of paired t-test (P = 0.0002). Reperfusion time kept nonsignificantly increased the predicted A levels by 27.44174 IU/L (−50.9901-105.8736 IU/L] (P = 0.4831) in accordance also with paired t-test (P = 0.3375). However, U-74389G administration and reperfusion time together produced a significant combined effect in keeping decreased the predicted A levels by 76.35332 IU/L (−116.8222-−35.8844 IU/L) (P = 0.0005). Reviewing the above and [Table 5], [Table 7] and [Table 8] sum up concerning the decreasing influence of U-74389G along with reperfusion time.
Table 6: The decreasing influence of U-74389G in connection with reperfusion time

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Table 7: The decreasing influence of U-74389G in connection with reperfusion time

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Table 8: The percentage decreasing influence of U-74389G in connection with reperfusion time

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  Discussion Top


Examples are described herein concerning whether pancreatic ischemia can influence the amylase levels. Fisher et al. predicted [9] decreased alteration of salivary alpha-amylase (SAA) levels by higher baseline ones in induced-stressed worries than healthy controls. Schneeberger et al. found no difference [10] of serum amylase values in 6 months observation period pancreas transplant graft function between mean cold ischemia time of 10 h histidine-tryptophan-ketoglutarate and University of Wisconsin solutions. Woessner et al. noted [11] a well-known increase in SAA levels through the infusion of 6% hydroxyethyl starch 130/0.4 than a crystalloid solution as a continuous infusion over 4 days in acute ischemic stroke patients. Viola et al. demonstrated [12] pancreatic IR damage by increased SAA levels than sham-operated rats. Freiburghaus et al. observed [13] no difference for SAA levels between ischemic pancreatic branches of the splenic artery than sham-operated ones with saline until 9 weeks in rats. Chronic pancreatitis follows acute necrotizing pancreatitis (ANP). Fiolet et al. measured [14] the extracellular volume decreased by about 20% by the noncardiac enzyme alpha-amylase in suspensions of isolated rat ventricular myocytes after 30 min total IR. Kφltringer et al. found [15] significantly increased the fasting levels of SAA in patients with II b stage peripheral arterial occlusive disease after 500 ml Elohaest administration during the first 4 treatment days. Spormann et al. analyzed [16] alpha-amylase in an acute pancreatitis of ischemic rats until 24 h postoperatively. Letko et al. found [17] SAA significantly increased 24 h after acute ischemic pancreatitis than the control cases. Sokolowski et al. found [18] the alpha-amylase release dependent on the duration of ischemia within 24 h postoperatively in acute pancreatitis rats. Sokolowski et al. caused [19] a drastic 2.5-fold increase in initial phase alpha-amylase levels, remaining 24 h postoperatively at a distinct pathological level in ischemic acute pancreatitis rats. Fleischer et al.[20] did not alter alpha-amylase levels after pancreatic 20 min intervals IR in mongrel dogs serum. Scheele et al. remarked an extreme initial rise in the levels of alpha-amylase gradually normalized in postoperative course of a [21] complete transected hepatoduodenal ligament occurred in a 59-year-old female patient. The majority of the above clinical situations simply confirm the reverse alteration of salivary and serum alpha-amylase levels upon pancreatic IR establishment.

Alhan et al. induced [22] ANP, which resulted in significant pancreatic necrosis, serum A levels increase, and serum PO 2 decreases in rats. The usage of U-74389G inhibited the serum PO 2 decreases and indicated a limited effect on the course of ANP. Therefore, it may be used in the treatment of lung injury during acute pancreatitis.


  Conclusions Top


U-74389G administration whether it interacted or not with reperfusion time has a significant decreasing beneficial restoring short-term effect on amylase levels.

Financial support and sponsorship

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.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

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Flessas I, Bramis I, Menenakos E, Toutouzas K, Agrogiannis G, Patsouris E, et al. Effects of lazaroid U-74389G on intestinal ischemia and reperfusion injury in porcine experimental model. Int J Surg 2015;13:42-8.  Back to cited text no. 3
    
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Bimpis A, Papalois A, Tsakiris S, Zarros A, Kalafatakis K, Botis J, et al. Activation of acetylcholinesterase after U-74389G administration in a porcine model of intracerebral hemorrhage. Metab Brain Dis 2012;27:221-5.  Back to cited text no. 4
    
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Tsaroucha AK, Papalois A, Vernadakis S, Adamopoulos S, Papadopoulos K, Lambropoulou M, et al. The effect of U-74389G on liver recovery after acute liver ischemia-reperfusion injury in a swine model. J Surg Res 2009;151:10-4.  Back to cited text no. 5
    
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Andreadou I, Poussios D, Papalois A, Gavalakis N, Aroni K, Gazouli M, et al. Effect of U-74389G (21-lazaroid) on intestinal recovery after acute mesenteric ischemia and reperfusion in rats. In Vivo 2003;17:463-8.  Back to cited text no. 6
    
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Tsompos C, Panoulis C, Toutouzas K, Zografos G, Papalois A. The acute effect of the antioxidant drug "U-74389g" on platelet distribution width during hypoxia reoxygenation injury in rats. J Neurol Stroke 2015;3:111.  Back to cited text no. 7
    
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Tsompos C, Panoulis C, Toutouzas K, Zografos G, Papalois A. Antioxidant 21-aminosteroid "U-74389G" ameliorates the short-time effect of hypoxia-reoxygenation on the platelet count in rats. Folia Med Cracov 2015;55:25-34.  Back to cited text no. 8
    
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Woessner R, Grauer MT, Dieterich HJ, Bepperling F, Baus D, Kahles T, et al. Influence of a long-term, high-dose volume therapy with 6% hydroxyethyl starch 130/0.4 or crystalloid solution on hemodynamics, rheology and hemostasis in patients with acute ischemic stroke. Results of a randomized, placebo-controlled, double-blind study. Pathophysiol Haemost Thromb 2003;33:121-6.  Back to cited text no. 11
    
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Költringer P, Langsteger W, Lind P, Klima G, Pierer G, Reisecker F, et al. Kinetic effects of 6% hydroxyethyl starch 200.000/0.6-0.66 in hypervolemic hemodilution. Acta Med Austriaca 1991;18 Suppl 1:20-2.  Back to cited text no. 15
    
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Sokolowski A, Spormann H, Urbahn H, Letko G. Contribution of pancreatic edema and short-term ischemia to experimental acute pancreatitis in the rat. II. Behaviour of serum parameters. Z Exp Chir Transplant Kunstliche Organe 1986;19:331-9.  Back to cited text no. 19
    
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22.
Alhan E, Türkyilmaz S, Erçin C, Kural BV. Effects of lazaroid U-74389G on acute necrotizing pancreatitis in rats. Eur Surg Res 2006;38:70-5.  Back to cited text no. 22
    



 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6], [Table 7], [Table 8]



 

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