AMELIORATIVE EFFECT OF BLUE BERRY AGAINST MULTIPLE ORGAN DAMAGE BY γ-IRRADIATION IN ADULT MALE ALBINO RATS

AMELIORATIVE EFFECT OF BLUE BERRY AGAINST MULTIPLE ORGAN DAMAGE BY

γ-IRRADIATION IN ADULT MALE ALBINO RATS

By

Aziza K. Omer, Gehan A. Youssef, AzzaH. Abd El-Wahab,

Asmaa F. Yousf and * Samy M. Elsayed Aly

Physiology Department, Faculty of Medicine (Girls) Al-Azhar University

and *Egyptian Atomic Energy Authority

ABSTRACT

Background: Blue berry fruit are rich in bioactive constituents, including flavonoids, anthocyanins, phenolic acids, stilbenes, and tannins, as well as nutritive compounds such as sugars, essential oils, carotenoids, vitamins and minerals. Numerous scientific studies provide ample evidence that bioactive compounds have the potential to decrease the risk of cardiovascular and degenerative diseases. Whole body exposure to ionizing radiation induces oxidative stress through generation of free radicals which often trigger chain reactions mediated tissue damage.

Objective: This work was done to study the potential protective effect of blue berry against injury of  heart, liver and kidney caused by exposure to gamma irradiation .

Material and Methods: Forty adult male albino rats were used and divided into four equal groups: Group 1 served as control group, group 2 received blue berry juice (BBJ) 1ml/100g body wt. by oral intubation gavage for 14 consecutive days , group 3 was exposed to single dose of 7Gy whole body gamma-irradiation, and group 4 received BBJ (1ml/100g body wt.) by oral gavage for 14 consecutive days before irradiation.

All groups of rats were anesthetized and sacrificed on the last day after treatment and/or irradiation. Blood samples were taken from all groups to estimate lactate dehydrogenase (LDH), creatine phosphokinase (CPK), interleukin-6 (IL-6), total cholesterol (TC), triacylglycerol (TAG), high density lipoprotein cholesterol (HDL-C), malondialdehyde (MDA), alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), urea, creatinine, cystatin and lipocalin.

Results: BBJ administration before exposure to g-irradiation significantly reduced serum LDH, CPK, IL-6, TC, TAG, and MDA versus a significant increase of serum HDL-C. Moreover, amelioration of liver and kidney function test was observed by significant decrease in ALT, AST, ALP, serum urea, creatinine, cyctatin-C and lipocalin-2 in comparison with irradiated group.

Conclusion: The study pointed out to the promising positive role of blue berry supplementation as a natural product to reduce oxidative stress and protect body organs from gamma irradiation. 

Key words: Oxidative stress, blue berry, γ- radiation, lipid profile, liver enzymes.

INTRODUCTION

      Radiation has been used increasingly in medicine and industry to help with diagnosis, treatment and technology. Radiations have tremendous therapeutic benefits for human. However, it is also associated with the risk of serious adverse effects (Saada et al., 2009).

     The absorption of ionizing radiation by living cells can directly disrupt atomic structures, producing chemical and biological changes. It can also act indirectly through radiolysis of water, thereby generating reactive chemical species known as free radicals. Together, the direct and indirect effects of radiation initiate a series of biochemical and molecular signaling events that may repair the damage or culminate in permanent physiological changes or cell death due to oxidative stress (Azzam et al., 2012).

     Oxidative stress is caused by an imbalance between the production of reactive oxygen species (ROS) and theability of a biological system to eliminate ROS or repair the resulting damage(Abd el-Aal, 2012). Once this imbalance takes place, cellular molecules such as nucleic acids, proteins, structural carbohydrates, and lipids may be damaged by oxidative modifications. Also, ROS play a causative role in numerous disease pathologies such as cancer, ischemia, and degenerative diseases such as aging, atherosclerosis, arthritis and neurodege-neration (Farag and Darwish , 2016):

      Evidence of oxidative injury is proved from measurements of biochemical markers of lipid peroxidation and protein oxidation. Lipid peroxidation is believed to be an important cause of destruction and damage to cell membranes and has been shown to be a contributing factor to the development of oxygen radicals-mediated tissue damage. However, cells are equipped with several natural enzymatic and non-enzymatic antioxidant defenses (Azab et al., 2011). The exposure of the human body to ionizing radiation leads to depletion of these endogenous antioxidants and ultimately to the development of systemic diseases (El-Missiry et al., 2007).

     Antioxidants can prevent or inhibit the oxidation of a target biomolecules. Dieticians actually prescribe diets containing antioxidants for conditions resistant to conventional pharmacological treatments (John et al., 2011).

     Keeping a close watch on the amount of antioxidants consumed and their action against free radicals is ,therefore, of the essence in maintaining a condition of health. Numerous epidemiological studies have revealed the existence of a positive correlation between eating fruit and the reduction of heart disorders, cancer and other degenerative pathologies (Basu et al., 2010-b). The various kinds of fruit investigated and described in the literature notably include blue berries, which have a strong antioxidant capacity (El-Nekeety et al., 2007).

     The berry fruits, including blueberry, cranberry, raspberry and strawberry, are known to be good sources of a number of bioactive compounds such as vitamins and phenolics having an antioxidant activity (Paredes-Lopez et al., 2010)

     The blue berry (Wild blueberry, Vaccinium angustifolium) contain higher level of polyphenols such as anthocyanins, flavonols, tannins and phenolic acids which show the potentiality to decrease the risk of cardiovascular and degenera-tive diseases, and to prevent cancer through their biological activities (Yi et al., 2005). However, there is a limited number of studies on the effect of blueberry in irradiation induced multiple organ damage.

     The aim of the present study was to investigate the radio-modifying effect of a blueberry juice against possible hazards of irradiation on heart, liver and kidney in adult  male albino rats.

MATERIAL AND METHODS

(I) Experimental animal and design: The present study was carried out on forty adult male albino rats weighing 130–160 g . Rats were acclimatized for 14 days in the animal house of Egyptian Atomic Energy Authority, under normal conditions, temperature range of 25 ± 5°C, and regular light/dark cycle in cages (150x60x50 cm- 5rats/cage). Animals were allowed free access of water and fed on a diet of normal rats chow.

Animals were divided randomly into fourequal  groups:

Group 1: Untreated control group.

Group 2: Rats received blue berry juice (BBJ) 1ml/kg body wt. by oral intubation gavage for 14 consecutive days (Zhong et al., 2015).

Group 3: Rats were exposed to whole body gamma irradiation at 7 Gy applied as single sublethal dose (Abd El-Azime and Ossman, 2012).

Group 4: Rats received BBJ 1ml/100g body wt. by oral gavage for 14 consecutive days before irradiation. This group was exposed to whole body γ-irradiation in the 14^th day of experiment as a single dose of 7Gy.

(II) Preparation of blue berry juice (BBJ): Blue berry fruit was purchased from local marked and prepared according to the method described by Wang et al. (2013). Fresh BBJ was prepared by homogenization of the fresh fruit right before the start of experiments (1 ml of BBJ contained about 2 g of dried blueberry).

(III) Irradiation Source and Technique: Irradiation was performed using a Canadian gamma cell-40 (¹³⁷Cs), at the Egyptian Atomic Energy Authority, Nasr City, Cairo, Egypt. Rats were placed individually into cages, each cage is 40×60×40 cm in size. Animals were whole body exposed to 7 Gy delivered as a single sublethal dose at dose rate of 0.45 Gy/minute (Abd El-Azime and Ossman, 2012).

(IV) Blood Sampling: All groups of rats were anesthetized on the next day after treatment and/or irradiation. Blood samples were obtained from the retro-orbital plexus veins from the individual rat (Simmons and Brick, 1970) by fine capillary heparinized tubes. Blood was collected into a plain centrifuge tube for serum preparation to assay the biochemi-cal parameters of blood including some cardiac enzymes, liver function tests, kidney function tests and the rest of biochemical parameters of inflammation and oxidative stress. Serum was separated in individual ependorf tubes and stored at -40^°C until biochemical measurements.

(V) Serum Assay: The separated sera were analyzed for estimation of:

Lipid profile and lipid peroxide byproduct:

- Serum total cholesterol (TC) and triacyl-glycerol (TAG) were estimated as described in the method of Sharma et al. (1987).

- Serum HDL was measured according to the method described by Grillo et al. (1981).

- Serum MDA was measured according to the method described by Liu et al. (2000).

   Biomarkers of cardiac injury and inflammation:

- Serum creatine phosphokinase kinase (CPK) was estimated according to Tietz (1995)

- Serum LDH was determined according to Guder et al. (2009).

- Serum IL-6 was determined by solid phase sandwich Enzyme-Linked Immuno-sorbent Assay (ELISA) kit (supplied by Gamma trade) using an ELISA reader (Triturus, Grifols Diagno-tistic, Barcelona, Spain) according to the manufacturer's instructions (Kaminska et al., 2000).

Biomarkers of liver function: Serum Liver transaminases (AST, ALT) and alkaline phosphatase (ALP) were determined as described by Ochei and Kolhackar (2008).

Biomarkers of kidney function:

- Serum urea and creatinine were determined according to the methods described by Henry et al. (1974) and Tabacco et al. (1979) respectively.

- Serum cystatin-C was determined by Enzo Cystatin-C ELISA kits (Haves-Zburof, 2011).

- Serum lipocalin-2 was determined by ELISA kit which allows for the in vitro quantitative determination of lipocalin-2 concentrations in serum from Sigma (Bolignano et al., 2008).

Statistical Analysis: was done using statistic package for social science version 16 (SPSS, 16) for windows. A One-way analysis of variance (ANOVA) for a completely randomized design and Duncan’s multiple range tests were used to analyze experimental data. Results were expressed as Mean ± Standard error of mean (S.E.M) and statistically analyzed using Tukey multiple comparison tests. Results were considered significant at P < 0.05.

RESULTS

    Whole body γ-irradiation of rats at a single dose of 7Gy induced significant increase in the serum levels of lipid profile; serum total cholesterol (TC) and triacylglycerol (TAG) and lipid peroxide byproduct malondialdehyde (MDA) versus a significant decrease in serum high density lipoprotein cholesterol (HDL-C) in comparison to control group. On the other hand, blue berry supplemented rats showed insignificant change in all the tested parameters, if compared to control group.  Blue berry administration before irradiation of rats resulted in improvement of lipid profile state as there was a significant decrease in serum TC, TAG and MDA while HDL-C was increased significantly in comparison with γ-irradiated rats. Moreover, blue berry administration before irradiation could return serum TAG to normal levels as there was insignificant difference (p˃0.05) in TAG when compared to blue berry only administered group. But, serum TC and MDA still showing a significant increase and serum HDL-C still showing significant decrease relative to the corresponding control group (Table 1).

Table (1): Effect of Blue berry administration on serum lipid profile [total cholesterol (mg/dl), triacylglycerol (mg/dl) and high density lipoprotein (mg/dl)] and malondialdehyde (nmol/ml) of irradiated rats.

a: significant values compared to control.                                 b: significant values compared to irradiated group.

c: significant values compared to blue berry group.

     Gamma-irradiated rats exhibited a significant elevation in serum biomarkers of cardiac injury and inflammation [creatine phosphokinase (CPK), lactate dehydrogenase (LDH) and interleukin- 6 (IL-6)] compared to control group. On the other hand, blue berry supplemented rats showed insignificant change in all the tested parameters, if compared to control group, blue berry treated rats before being γ-irradiated demonstrated an obvious decrease in serum levels of CPK, LDH and IL-6 in comparison to untreated γ-irradiated group. Moreover, Blue berry pretreated γ-irradiated rats could return all the measured cardiac parameters to the normal levels as there were insignificant differences, if compared to their respec-tive control group (Table 2).

Table (2): Effect of Blue berry administration on serum biomarkers of cardiac injury and inflammation and malondialdehyde of  irradiated rats.

a: significant values compared to control.                                 b: significant values compared to irradiated group.

c: significant values compared to Blue berry group.

     Gamma-irradiated rats exhibited a significant increase in serum biomarkers of liver function tests (ALT, AST and ALP) in comparison with control group. On the other hand, Blue berry supplemented rats showed insignificant change in all the tested parameters, if compared to control group. Meanwhile, blue berry pretreated γ-irradiated rats could improve liver functions as there was a significant decrease in serum levels of ALT, AST and ALP in comparison with untreated γ-irradiated animal group. However, their levels still higher than normal as they were significantly higher than their corresponding control group (Table 3).

Table (3): Effect of Blue berry administration on serum biomarkers of liver function [alanine transaminase (U/L), aspartate transaminase (U/L) and alkaline phosphatase (U/L)] of  irradiated rats..

a: significant values compared to control.                              b: significant values compared to irradiated group.

c: significant values compared to Blue berry group.

     Gamma-irradiation of rats produced a significant increase in serum biomarkers of kidney function tests (urea, creatinine, cystatin-C and lipocalin-2) relative to control group. On contrary, blue berry supplemented rats showed insignificant change in all the tested parameters, if compared to control group. Blue berry pretreated γ-irradiated rats exhibited a significant decrease in serum levels of urea, creatinine, cystatin-C and lipocalin-2 compared with γ-irradiated rats. Moreover, it could return all of them, except cystatin-C, to normal levels as there were insignificant change as regard serum levels of urea, creatinine and lipocalin-2 but serum level of cystatin-C still significantly higher as compared to their respective control group (Table 4).

Table (4): Effect of Blue berry administration on serum biomarkers of kidney function [urea (mg/dl), creatinine (mg/dl), cystatin-C (ng/ml) and lipocalin-2 (pg/dl)] of irradiated rats.

a: significant values compared to control.                                 b: significant values compared to irradiated group.

c: significant values compared to Blue berry group.

DISCUSSION

      In the present work, the level of TC, TAG and MDA in serum was significantly higher in irradiated group than the control group. Remarkable decrease was observed in the concentration of HDL-C in the serum of γ-irradiated rats. These findings were in line with Farag and Darwish (2016) who attributed these findings to the generation of free radicals resulting to the peroxidation of membrane lipids which justifies a state of liver dysfunction. Abou-Safi et al. (2005) and Adaramoye (2010) observed that the elevation in serum lipid fractions might result from ionizing radiation ability to accelerate other pathways of cholesterol formation like increasing its rate of biosynthesis in the liver and other tissues, or destruction of cell membrane by radiation and also to disturbance of LDL cholesterol receptors, leading to hypercholesterolemia. The recorded elevated level of triglycerides correlated with the previous findings of Makhlouf and Makhlouf (2012) who observed that, after irradiation, insulin level increases and synthesis of triglycerides increases in both adipose tissues and liver which are accompanied by an acceleration of fatty acids mobilization from fat depots to blood. Also, they suggested that oxidative stress might be an important determinant of altered lipid metabolism due to radiation exposure. Other possible mechanism of increased TAG levels might be related to the decrease in lipoprotein lipase activity in adipose tissue, leading to a reduction in the uptake of triglycerides (Jedidi et al., 2003).

     The increase in MDA level might be attributed to the high level of oxidative stress and the overproduction of ROS associated with irradiation which interact with the polyunsaturated fatty acids in the lipid portion of biological membranes, initiating the lipid peroxidation and finally damaging the cell membranes (Mansour and Tawfik, 2012). Yilmaz and Yilmaz (2006) reported that MDA is a major biomarker of oxidative damage to living cells is increased lipid peroxidation and commonly measured parameter of lipid damage after ionizing radiation exposure. Lipid peroxdation is believed to be an important cause of destruction and damage to cell membranes and has been suggested to be a contributing factor to the development of oxygen radicals-mediated tissue damage.

     The present study showed that blueberry administration before irradiation of rats resulted in a significant decrease in serum level of TC, TAG and MDA with concomitant significant increment in the serum HDL-C compared to irradiated rats. The recorded data correlated with previous findings of Jensen et al. (2008) and Baus et al. (2010-a) who detected that  berry anthocyanins may exert cardiopro-tective effects by reducing oxidative stress and inflammation through inhibitory effects of the anthocyanin fraction of berry extract on NO biosynthesis and inducible iNOS protein expression, thereby decreasing the inflammatory response in macrophages and inhibiting the formation of foam cells.

     Another possible mechanism of the beneficial effect of BBJ pretreatment may be due to its high content of phytochemi-cals, mainly flavonols, phenolic acids and anthocyanins (Brito et al., 2014). Also, Qin et al. (2009) proposed that the elevated HDL-C may be partially mediated via the inhibition of  cholesteryl ester transfer protein (CETP) by anthocyanins. CETP is a plasma protein that mediates the removal of cholesteryl esters from HDL in exchange for a triacylglycerol molecule derived primarily from either LDL, VLDL, or chylomicrons. Moreover, Ikeda et al. (2005) have indicated that polyphenols could exert their lipid lowering properties through various mechanisms, namely by slowing down triacylglycerol absorption through inhibition of pancreatic lipase, increasing cholesterol excretion in feces, attenuating hepatic lipid accumulation through activation of adenosine monophosphate (AMP)-activated protein kinase suppres-sing hepatic secretion of apo-lipoprotein B100 and increasing expression of LDL receptors in the liver.

     Effects of whole body γ-irradiation of rats were accompanied by  alteration of traditional serum biomarkers of cardiac injury, elevated  activity  of creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) with concomitant increase in pro-inflammatory cytokine interleukein-6 (IL-6) compared with control group, indicating occurrence of cardiac injury. These results and conclusion were in accordance with Guida et al. (2016) who indicated that IR induces pro-inflammatory processes in which tumor necrosis factor (TNF-α), interferon (IFN-γ) and interleukin levels are altered, eventually leading to inflammatory disorders. Also, the current findings were consistent with several previous studies proved by Mansour & Tawfik ( 2012), Freitas et al. (2013), and Elkady & Ibrahim (2014). 

    g-irradiation induced-cardiotoxicity might be produced by stress from acute radiation response. The cardiomyocyte plasma membrane loses its fluidity, causing extravasations of cytosolic enzymes (e.g. CPK and LDH) to circulating blood, indicating a pathological condition. In addition, the heart is in spite of being a vital organ, it generates intense oxidative imbalances because of its intense activity. Moreover, the heart presents a less potent antioxidant system when compared to other body tissues (Elkady and Ibrahim, 2014).

     Moreover, radiation-induces cardiovas-cular disruption might be due to endothelial damage via production of reactive oxygen species (Hatoum et al., 2006). In addition, the inflammatory vascular damage due to radiation is also via oxidative stress and activation of nuclear factor-kappa B (Weintraub et al., 2010). Hayashi et al. (2005) proposed the indirect association of inflammation with radiation-induced vascular damage comes from studies showing elevated levels of the proinflammatory cytokines IL-6, CRP, TNF-α, and INF-γ and also increased the levels of the anti-inflammatory cytokine IL-10, in the Japanese atomic bomb survivors. Moreover, data regarding the main networks activated after IR exposure, cytokines exhibit pivotal roles in invasiveness and fibrosis radiation-related and radio-immune combined therapies (Maggio et al., 2015).

      The current results showed a radiopro-tective potential of blueberry supplemen-tation before γ-irradiation in the tested parameters (LDH, CPK and IL-6) relative to the control γ-irradiated group. These results were in agreement with Blumeberg et al. (2015) who reported that consumption of a purified mixture of anthocyanins (i.e. bioactive polyphenol of edible berries) led to significant decreases in C-reactive protein (CRP), soluble vascular cell adhesion molecule1 (sVCAM-1), and plasma IL-1β in patients with hypercholesterolemia. Ashour et al. (2011) detected that bilberry significantly inhibits doxorubicin-induced elevations in serum activity of LDH, CPK and CK-MB, as well as troponin I levels.

     The putative mechanism of radiomodi-fying action of berries resides in its bioactive contents especially anthocya-nins.  Anthocyanins act as novel cardio-protectants by maintaining vascular permeability, reducing inflamatory responses and platelet aggregation, and offer superior vascular protection compared to other cardioprotective drugs (Duthie et al., 2006 and Neto, 2007). Also, Freitas et al. (2013) suggested that phenolic and flavonoids found in  fruits as black grape juice are great antioxidants because their molecular structures contain hydroxyl groups which provide electrons to stabilize free radicals and ROS, restoring the redox state of cardiomyocytes.

     The results of the current study revealed marked elevation in liver transaminases (ALT and AST) and alkaline phosphatase (ALP) in γ-irradiated animals compared to control group, which could be attributed to hepatic damage induced by gamma rays. The liver, which was used for bio-transformation and detoxification of materials is particularly susceptible to oxidative stress. These findings stand in well agreement with several previous studies of El-Missiry et al. (2007),  Eydan et al. (2011) and Farag & Darwish (2016) who suggested to be due to the damage of structural integrity of the liver  resulted in the leakage of these enzymes from the cytosol into the blood stream.

    The mechanism of g-irradiation induced liver injury clarified by Kalpana et al. (2011) who suggested that levels of the lipid peroxidation indicator MDA have been shown to increase in a variety of biological systems including the liver after gamma radiation exposure. Also, Choi et al. (2007) reported that lipid peroxidation reactions can occur at  both of cell membrane and mitochondria membranes, and either can subsequently trigger cell death through apoptosis (caspase-mediated apoptotic cell death) and/or autophagy (LC-3-mediated autophagic cell death). Marnett (1991) demonstrated that such free radical-mediated lipid peroxidation is harmful not only because damaged lipids disrupt membrane structure and function, but also because the process produces potentially mutagenic and carcinogenic by products.

     In the present work, BBJ consumption by rats before irradiation decreased the serum liver enzymes (ALT, AST and ALP) when compared to irradiated rats, indicating hepato-protective properties of the BBJ. The observed decrease in the activity of these hepatic enzymes of BBJ supplemented group before irradiation could be attributed to the high content of naturally occurring polyphenols with antioxidant and free radical scavenging activity. The present data agreed with Wang et al. (2013) who demonstrated that BBJ consumption by rats attenuates CCl4-induced hepatotoxicity evaluated by increased expression of metallothionein, increased SOD activity, reduced oxidative stress, and decreased levels of α-smooth muscle actin and collagen III in the liver.  These suggest that attenuation of hepatotoxicity by BBJ might be through its enhancement of the antioxidative capability of the liver. Ozcelik et al. (2014) also reported that consumption of blue berry significantly decreases liver arginase activity and ornithine levels in addition to serum transaminases with marked increase in nitric oxide and glutathione levels during the acetamino-phen-induced liver injury. Therefore, hepatoprotective effect efforded by blue berry can be attributed to its antioxidant and anti-inflammatory activities.

     In the present study, elevation of serum urea and creatinine was observed in γ-irradiated rats when compared to control group. These results came similar to previous investigations by Barakat et al. (2011) and Abd El Kader et al. (2015). These effects caused by the interaction of cellular membranes with gamma rays or action of free radicals may be related to extensive breakdown of renal architecture (Hussein, 2008).

     The observed increase in serum urea could be due to increase in glutamate dehydrogenase enzyme as a result of irradiation, and this may increase carbamayl phosphate synthetase activity leading to increase in urea concentration (Ramadan et al., 2001). Robbins et al. (2001) reported the impaired detoxifica-tion function of the liver by irradiation could also contribute in the increase of urea in the blood. Ferguson and Waikar (2012) reported that creatinine produces through metabolic processes in the muscles and is freely filtered by the glomerulus without renal tubular reabsorption, and increased levels in serum is an indicator of reduced GFR and or increased muscle protein catabolism. Abdelhalim and Moussa (2013) reported elevation of serum creatinine level of the irradiated rats group might indicate nephritis and renal dysfunction due to impairment of glomerular selective properties, and  proposed that oxidative stress is linked to the organ damage following exposure to ionizing radiation. El-Kashef and Saada (1988) attributed the increase in serum creatinine to the interaction of ionizing radiation with the sites of biosynthesis.

      The current results indicated a signifi-cant increase in serum cystatin C and lipocalin-2 levels in γ-irradiated rats compared to control group, indicating kidney injury. The present data were in accordance with previous study of Vijayasimha et al. (2013). Parikh and Devarajan (2008) reported that cystatin-C is assigned as a more reliable marker of GFR than creatinine, and can detect mild to moderate kidney affection. Cystatin-C is one of the major protectors of the intact glomerular basement membrane from damage by cysteine proteinases.  Also, lipocalin-2 was identified as one of the earliest and most robustly induced proteins in the kidney after toxic kidney injury (Devarajan, 2006).

     BBJ supplemention of rats prior to whole body γ-irradiation had a marked decrease in kidney function biomarkers (serum urea, creatinine, cystatin-C and lipocalin-2) when compared to γ-irradia-ted control group, indicating nephropro-tective potential of BBJ. The present findings were consistent with the study of Nair et al. (2014) who indicated that blueberry could protect against renal structural injury and dysfunction in metabolic syndrome animal model by inhibiting Toll-like receptors (TLR4) signaling-driven inflammation. This is, at least in part, a possible cause for the progression of glomerular and tubular injury, thereby contributing to renal dysfunction. Also, Elks et al. (2011) reported feeding of BB-enriched diet attenuate the development of hyper-tension-induced renal injury in rat model, proved by lowered blood pressure, preserved renal hemodynamics, and improved redox status in kidneys.

CONCLUSION

     BBJ may serve as radioprotector against γ-irradiation induced cardio-hepato-nephrotoxicity possibly by amelio-rating dyslipidemia, inhibiting lipid peroxidation, oxidative stress and inflam-mation and enhancing the antioxidant activity.

REFERENCES

1. Abd El-Aal, H.A  (2012): Lipid peroxidation end-products as a key of oxidative stress: effect of antioxidant on their production and transfer of free radicals, lipid peroxidation. Zoology Department, Faculty of Science, Mansoura University, Egypt  INTCHE. Sci. Technol. Med., 3: 63-88.

2. Abd El-azime, A.S. and Ossman, N.N. (2012): Protective effects of Hawthorn (Cartagus Linn) against radiation induced damage in rats. Radiation biology department, food irradiation departement, national center for radiation research and technology, authority, Cairo, Egypt. Arab Journal of Nuclear Science and Applications,  46(1): (304-312).

3. Abdelhalim, M.A. and Moussa, S.A. (2013): The biochemical changes in rats' blood serum levels exposed to different gamma radiation doses. Afr. J. Pharm. Pharmacol.,7(15):785-792.

4. Abd El-Kader, M.A.; El-kafrawy, M.H.; Tolba, A.M.A.; Ali, M.M. and Mohamed, A.S. (2015): Evaluation of Taurine Role on Some Biochemical and Histological Alterations in γ-Irradiated Rats. Int. J. Pharm. Sci. Rev. Res., 47: 263-271.

5. Abou-Safi, H.M.; Ashry, O.M. and Kafafy, Y.A. (2005): Effect of N-acetyl-L- cysteine and α-tocopheroldministration on endogenous antioxidant protection of liver DNA and RNA and plasma lipid profile in γ-irradiated rats.Egypt. J. Rad. Sci. Applic., 18(1): 81-96.

6. Adaramoye, O.A. (2010): Protective Effect of Kolaviron, a Biflavonoid from Garcinia kola Seeds, in Brain of Wistar Albino Rats Exposed to Gamma-Radiation. Biol. Pharm. Bull., 33: 260-266.

7. Ashour, O.M.; Elberry, A.A.; Alahdal,A.M.;  Al Mohamadi, A.M.; Nagy, A.A.; Abdel-Naim, A.B.; Abdel-Sattar, E.A. and Mohamadin, A.M. (2011): Protective effect of bilberry (Vaccinium myrtillus) against doxorubicin-induced oxidative cardiotoxicity in rats. Med. Sci. Monit., 17(4): BR110–BR115.

8. Azab, K.S; Bashandy, M.; Salem, M.; Ahmed, O.; Tawfik, Z.  and  Helal, H. (2011): Royal jelly modulates oxidative stress and tissue injury in gamma irradiated male Wister Albino rats. N. Am. J. Med. Sci., 3(6): 268–276.

9. Azzam, E.I.; Gerin, J.P. and Pain, D. (2012): Ionizing radiation-induced metabolic oxidative stress and prolonged cell injury. Cancer Lett., 327(0): 48-60.

10. Barakat, I.A.H.; Abbas, O.A.; Ayad, S. and Hassan, A.M. (2011): Evaluation of radioprotective effects of wheat germ oil in male rats, J. Am. Sci., 7(2): 664-673.

11. Basu, A.; Du, M.; Leyva, M.J.; Sanchez, K.;  Betts; N.M.; Wu, M.; Aston, C.E. and Lyons, T.J. (2010-a): Blueberries decrease cardiovas-cular risk factors in obese men and women with metabolic syndrome. J Nutr., 140(9):1582-1587.

12. Basu, A.; Michael, R. and Timothy, J.L. (2010-b): Berries: emerging impact on cardiovascular health. Nutr Rev., 68(3): 168–177.

13. Blumberg, J.B.; Camesano, T.A.; Cassidy, A.; Kris-Etherton,  P.; Howell, A.; Manach, C.; Ostertag, L.M.; Sies, H.; Skulas-Ray, A. and Vita, J.A. (2015): A polyphenol-rich cranberry extract protects from diet-induced obesity, insulin resistance and intestinal inflammation in association with increased Akkermansia. J Food Composition and Analysis, 42:56-62.

14. Bolignano, D.; Lacquaniti, A.; Coppolino, G.; Campo, S.; Arena, A. and Buemi, M. (2008): Neutrophil gelatinase-associated lipocalin reflects the severity of renal impairment in subjects affected by chronic kidney disease. Kidney Blood Press. Res., 31:255-258.

15. Brito, A.; Areche, C.; Sepulveda, B.; Kennelly, E.J. and Simirgiotis, M.J. (2014): Anthocyanin Characterization, Total Phenolic Quantification and Antioxidant Features of Some Chilean Edible Berry Extracts. Molecules, 19: 10936-10955.

16. Choi, A.O.; Cho, S.J.; Desbarats, J. and Lovric, J. (2007): Maysinger D. Quantum dot-induced cell death  involves Fasupregulation and lipid peroxidation in human neuroblastoma cells. J. Nanobiotechnology,12; 5:1-13.

17. Devarajan, P. (2006): Update on mechanisms of ischemic acute kidney injury. J. Am. Soc. Nephrol., 17: 1503-1520.

18. Duthie, S.J.; Jenkinson, A.M.; Crozier, A.; Mullen, W.; Pirie, L. and Kyle, J. (2006): The effects of cranberry juice consumption on antioxidant status and biomarkers relating to heart disease and cancer in healthy human volunteers. Eur. J. Nutr., 45:113–122.

19. Elkady,  A.A. and Ibrahim, R.M. ( 2014 ): Royal jelly attenuates ionizing radiation induced cardiac toxicity in male albino rats. Med. J. Cairo Univ., 82 (1): 13-18.

20. El-Kashef, H. and Saada, H. (1988): Changes in the level of urea, creatine and creatinine in the liver and serum of irradiated rats. Isotope and Rad. Res., 20: 43-60.

21. Elks, C.M.; Reed, S.D.; Mariappan, N.; Hale, B.S.; Joseph, J.A.; Ingram, D.K. and  Francis, J. (2011): A Blueberry-enriched diet attenuates nephropathy in a rat model of hypertension via reduction in oxidative stress. PLoS ONE, 6 (9): e24028 (1-10).

22.  El-Missiry, M.A.;  Fayed, T.A.;  El-Sawy, M.R. and El-Sayed A.A. (2007): Ameliorative effect of melatonin against gamma-irradiation-induced oxidative stress and tissue injury. Ecotoxicol Environ Saf., 66(2): 278 - 286.

23. El-Nekeety, A.A.; El-Kholy, W.; Abbas, N.F.; Ebaid, A.; Amra, H.A. and Abdel-Wahhab, M.A. (2007): Efficacy of royal jelly against the oxidative stress of fumonisin in rats. Toxicon., 50: 256–269.

24. Eydan, D.M.; Ursel, B.G.; Ilgici, B.B.; An, B.C. and Zbek, N.O. (2011): Protective effect of lycopene against radiation-induced hepatic toxicity in rats. J Int. Med. Res., 39: 1239 – 1252.

25. Farag, M.F. and Darwish, M.M. (2016):  Possible impact of antioxidant properties of cocoa (Theobroma Cacao L.) against irradiation-induced some biochemical disorders in rats. Arab J. Nuclear Sci. Applications, 94 (1): 151-158.

26. Ferguson, M.A. and Waikar, S.S. (2012): Established and emerging markers of kidney function. Clinical chemistry, 58: 680-689.

27. Freitas, R.B.; Boligon,A.A.; Rovani, B.T.; Piana, M.; Brum, T.F.; Jesus, R.S.; Rother, F.C.; Alves, N.M.; Rocha, J.B.; Athayde, M.L.; Barrio, J.P.; Andrade, E.R. and Bauerman, L.F. (2013): Effect of Black Grape Juice against Heart Damage from Acute Gamma TBI in Rats. Molecules, 18: 12154-12167.

28. Grillo, F.; Izzo, C.; Mazzotti, G. and  Murador, E. (1981): Improved method for determination of high-density-lipoprotein cholesterol II. Enzymic determination of cholesterol in high-density lipoprotein fractions with a sensitive reagent. Clin. Chem., 27 (3): 375-9.

29. Guder, W.G.; Narayanan, S.; Wisser, H. and Zawta B. (2009): Samples: From the patient to the laboratory, 4^th ed., Pbl. Weinheim: Wiley-Blackwell.

30. Guida, M.S.; El-Aal, A.A.; Kafafy, Y.; Salama, S.F.; Badr, B.M. and Badr, G. (2016): Thymoquinone rescues t lymphocytes from gamma irradiation-induced apoptosis and exhaustion by modulating pro-inflammatory cytokine levels and PD-1, Bax, and Bcl-2 Signaling. Cell Physiol. Biochem., 38(2): 786-800.

31. Hatoum, O.A.; Otterson, M.F. and Kopelman, D. (2006): Radiation induces endothelial dysfunction inmurine intestinal arterioles via enhanced production of reactive oxygen species. Arterioscler. Thromb. Vasc. Biol., 26 (2): 287–294.

32. Haves-Zburof, D. (2011):Cathepsins and their endogenous inhibitors cystatins: expression and modulation in multiple sclerosis. J. Cell. Mol. Med., 15: 2421.

33. Hayashi, T.; Morishita, Y. and Kubo, Y. (2005): Long-term effects of radiation dose on inflammatory markers in atomic bomb survivors. Am. J.  Medicine, 118 (1): 83–86.

34. Henry, R.J.; Cannon, D.C. and Winkelman, J.W. (1974): Clinical Chemistry:  principles  and  techniques  2nd  ed.,  New  York,  Harper  and  Row, 422-424.

35. Hussein, E.M. (2008): Modulatory role of arabic gum in gamma rays-induced damages in rats. J. Rad. Res. Appl. Sci.,1(2): 427-441.

36. Ikeda, I.;, Tsuda, K.; Suzuki, Y.; Kobayashi, M.; Unno, T.; Tomoyori, H.; Goto, H.; Kawata, Y.; Imaizumi, K.; Nozawa, A. and Kakuda, T. (2005): Tea catechins with a galloyl moiety suppress postprandial hyper-triacylglycerolemia by delaying lymphatic transport of dietary fat in rats. J Nutr.,135(2): 155-9.

37. Jedidi, I.; Therond, P.; Zarev, S.; Cosson, C.; Couturier, M. and Massot, C. (2003): Paradoxical protective effect of aminoguanidine toward low-density lipoprotein oxidation: inhibition of apolipoprotein B fragmentation without preventing its carbonylation. Mecha-nism of action of aminoguanidine. Biochem., 42: 11356–11365.

38. Jensen, G.S.; Wu, X. and Patterson, K.M. (2008): In vitro and in vivo antioxidant and anti-inflammatory capacities of an antioxidant-rich fruit and berry juice blend. Results of a pilot and randomized, double-blinded, placebo-controlled, crossover study. J Agric Food Chem., 56:8326–8333.

39. John, E.B.; John, E.M. and John, W.H. (2011): Radiation as a risk factor for cardiovas-cular disease. Antioxid. Redox Signal., 15(7): 1945–1956.

40. Kalpana, K.B.; Devipriya, N.; Srinivasan, M.; Vishwanathan, P.; Thayalan, K. and Menon, V.P. (2011): Evaluating the radioprotective effect of hesperidin in the liver of Swiss albino mice. Eur. J. Pharmacol., 658:206-12.

41. Kaminska, J., Kowalska, M., Nowacki, M., Chwalinski, M., Rysinska, A. and Fuksiewicz, M. (2000): CRP, TNF-α, IL-1ra, IL-6, IL-8 and IL-10 in blood serum of colorectal cancer patients.Pathology Oncology Research, 6: 38-41.

42. Liu, J.; Yeo, H.C.; Overvik-Douki, E.; Hagen, T.; Doniger, S.J.; Chu, D.W.; Brooks, G.A. and Ames, B.N. (2000):Chronically and acutely exercised rats: biomarkers of oxidative stress and endogenous antioxidants. J. Appl. Physiol., 89: 21-28.

43. Maggio, F.M.; Minafra, L.; Forte, G.I.; Cammarata, F.P.; Lio, D.; Messa, C.; Gilardi, M.C. and Bravata, V. (2015): Portrait of inflammatory response to ionizing radiation treatment. J. Inflamm., 12(14):1-11.

44. Makhlouf, R. and Makhlouf, I. (2012): Evaluation of the effect of spirulina against gamma irradiation induced oxidative stress and tissue injury in rats. Int. J. Appli. Sci. Engin. Res., 1(2): 152-164.

45. Mansour, H.H. and Tawfik, S.S. (2012): Early treatment of radiation-induced heart damage in rats by caffeic acid phenethyl ester. Eur. J. Pharmacol., 692 (1-3): 46-51.

46. Marnett, L.J. (1999): Lipid peroxidation-DNA damage by malondialdehyde. Mutat. Res., 424:83-95.

47. Nair, A.R.; Elks, C.M.; Vila, J.; Piero, F.D.; Paulsen, D.B. and Francis, J. (2014): A blueberry-enriched diet improves renal function and reduces oxidative stress in metabolic syndrome animals: potential mechanism of TLR4-MAPK signaling pathway. PLoSONE, 9(11): e111976 (1-12).

48. Neto, C.C. (2007): Cranberry and blueberry: evidence for protective effects against cancer and vascular diseases. Mol. Nutr. Food Res., 51: 652–64.

49. Ochei, J. and Kolhatkar, A. (2008): Medical laboratory science: theory and practice. Pbl., Tata McGraw–Hill Publishing Company Limited, New York: pp. 637-745.

50. Ozcelik, E.; Uslu, S.; Burukoglu, D. and Musmul, A. (2014): Chitosan and blueberry treatment induces arginase activity and inhibits nitric oxide production during acetaminophen-induced hepatotoxicity. Pharmacogn. Mag., 10 (Suppl 2): S217–S224.

51. Paredes-Lopez, O.; Cervantes-Ceja, M.K.; Vigna-Perez, M.; Hernandez-Perez, T. (2010): Berries improving human health and healthy aging and promoting quality life – a review. Plant Foods Hum. Nutr., 65: 299-308.

52. Parikh, C.R. and Devarajan, P. (2008): New biomarkers of acute kidney injury. Crit. Care Med., 36 (4supp1): S159-165.

53. Qin, Y.; Xia, M.; Ma, J.; Hao, Y.T.; Liu, J.; Mou, H.Y.; Cao, L. and Ling, W. (2009): Anthocyanin supplementation improves serum LDL- and HDL-cholesterol concentrations associated with the inhibition of cholesteryl ester transfer protein in dyslipidemic subjects. Am. J. Clin. Nutr., 90:485–92.

54. Ramadan, L.A.; Shouman, S.A.; Sayed-Ahmed, M.M. and El-Habit, O.H. (2001): Modulation of radiation-induced organs toxicity by cremophorel in experimental animals. Pharmacol. Res., 43:185-191.

55. Robbins, S.; Mally, Y.; Davis, C. and Bonsib, S. (2001): The rate of the tubule interstitum in irradiation induced renal fills . Rad. Res., 155: 481.

56. Saada, H.N.; Said, U.Z.; Meky, N.H. and Abd El Azime, A.S. (2009): Grape seed extract Vitisvinifera protects against radiation-induced oxidative damage and metabolic disorders in rats. PhytotherRes. ,23(3):434-8.

57. Sharma, A.; Artiss, J.D. and, Zak B. (1987): A method for the sequential colorimetric determination of serum triglycerides and cholesterol. Clin. Biochem., 20 (3): 167-72.

58. Simmons, M.L. and Brick, J.O. (1970): The laboratory mouse. selection and management. Pbl., Prentice-Hall, Englewood Cliffs, New Jersey, pp. 153-156.

59. Tabacco,  A.;  Meiattini,  F.;  Moda,  E. and  Tarli, P. (1979): Simplified enzymatic colorimetric  serum  urea  nitrogen  determina-tion.  Clin.  Chem.,  25:  336-337.

60. Tietz, N.W. (1995): Clinical guide to laboratory tests. 3^rded. PbL. Philadelphia, PA: WB Saunders, pp. 806-809.

61. Vijayasimha, M.; Padma1, V.V.; Mujumdar, S.K. and Satyanarayana, P.V. (2013): Evaluation of the diagnostic perfor-mance of new markers for acute kidney injury associated with contrast administration. Journal of Mahatma Gandhi Institute of Medical Sciences, 18(2):116-121.

62. Wang, Y.; Cheng, M.; Zhang, B.; Nie, F. and Jiang, H. (2013): Dietary supplementation of blueberry juice enhances hepatic expression of metallothionein and attenuates liver fibrosis in rats. PLoS ONE, 8(3): e58659.

63. Weintraub, N.L.; Jones, W.K. and Manka, D. (2010): Understanding radiation-induced vascular disease. J. Am. College of Cardiol., 55(12): 1237–1239.

64. Yi, W.; Fischer, J.; Krewer, G. and Akoh, C.C. (2005): Phenolic compounds from blueberries can inhibit colon cancer cell proliferation and induce apoptosis. J. Agric. Food Chem., 53, 7320–7329.

65. Yilmaz, S. and Yilmaz, E. (2006): Review: Effects of melatonin and vitamin E on oxidative-antioxdative status in rats exposed to irradiation. Toxicol., 222 (1-2): 1- 7.

66. Zhong,Y.; Wang,Y.;  Guo, J.; Chu, H.; Gao, Y. and Pang, L. (2015): Blueberry improves the therapeutic effect of etanercept on patients with juvenile idiopathic arthritis: phase III study. Tohoku J. Exp. Med., 237, 183-191.