@2024 Afarand., IRAN

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ISSN: 2252-0805    The Horizon of Medical Sciences 2017;23(4):293-299

Background
Silver nanoparticles are small particles of silver in the range of 1 to 100 nanometers and today they have found extensive applications in various sciences, especially biology and medicine [1]. Due to the widespread use of silver nanoparticles, concerns about their biological effects on the environment and human health have been generated [2].
Previous Studies
… [3]. It has been shown that silver nanoparticles accumulate in various tissues after entering the bloodstream, causing damage to the liver, kidneys, and damage of testicular tissue [4]. Research has shown that silver nanoparticles have toxic effect on ovarian tissue and by activating the caspase 3 through increasing free radicals from silver nanoparticles, it activates internal pathway of apoptosis. Therefore, the activation of the pathway of apoptosis and the formation of stress-oxidative conditions affect ovulation [5]. … [6-17]. Crocus sativus L is a perennial herb and from the iridas family. Crocin glycosides consists of carotenoid called crocin and sugar which creates the color of saffron. Crocin, Crostin, and Safranal are known as active biological agent of saffron [18] which has antioxidant properties and are capable of eliminating free radicals and can lead to significant reductions in oxidative damage in ischemic tissues [19]. … [20-23].
Aim(s)
Considering the growing trend of the use of medicinal plants and the antioxidant properties of crocin, the present study was conducted to determine the effect of crocin on Bax/Bcl-2 ratio, lipid peroxidation and antioxidant enzymes activity in chicken embryo liver tissue treated with silver nanoparticles.
Research type
This study is experimental.
Research society, place and time
This study was performed on Ross 308 chick embryo with an average weight of 84.15±4.22 gram.
Sampling method and number
45 Ross 308 eggs with embryo were randomly divided into five groups of nine included one control and four experimental groups.
Used devices&materials
Control samples were treated with 10ml injection of saline solution on day 10 of incubation. All experimental groups 1, 2, 3, and 4 were treated in the first step on day 10 of incubation with 0.5 ml of silver nanoparticle (Sigma Aldrich, Germany) 200 ppm and the amount of 60 nm in a single injection. Then, experimental groups 2, 3, and 4 were treated with concentrations of 100, 200, and 300 micrograms per milliliter on day 12 of incubation with 0.5 ml injection of crocin (Sigma-Aldrich, France) respectively. It should be noted that the concentration of 900 microgram per milliliter of crocin was considered as LD 50 (fatal dose). Also, the ED50 (effective dose) of crocin was in the range of 50 to 400 μg / ml. Therefore the concentrations of 100, 200, and 300 μg / ml were chosen for treatment. Eggs were placed in an incubation machine (M-32) at a temperature of 38 ° C and a relative humidity of 65% for a period of 20 days [24]. On the 10th and 12th days of incubation, at first, the place of embryo amniotic bag was determined using a neurobiological method. All single injections were performed by a syringe with needle number 25 to amniotic fluid. At first, the injection site was disinfected with alcohol and then injection was performed. In the next step, to prevent the entering of microbes, the injection site was blocked by paraffin [25]. On the day 20th of incubation, the chick embryos were removed from the eggs and then liver tissue was removed from the body of the embryos. The liver tissue was then washed with saline solution and then homogenized with Tris buffer (Sigma-Aldrich, Germany) for 5 minutes with a T25 digital ULTRA-TURRAX Model homogenizer (IKA; Germany). The homogenized solution was centrifuged by a refrigerated centrifuge model Z366 (Hermie, Germany). To prevent degradation of enzymes and proteins, all steps were carried out at a temperature of 4 ° C (refrigerated centrifuges) and 0.5 mM phenylmethylsulfonyl fluoride (Sigma-Alderich, Germany) was used as a protease inhibitor [26]. After centrifugation, a clear overlay solution was separated from the underside of the sediment and was used to measure the parameters Bax, Bcl-2, superoxide dismutase, glutathione peroxidase, catalase, and malondialdehyde. Bcl-2 cytoplasmic surface with a sensitivity of 9.375 pg/ml and range of 15.625-1000 pg/ml, Bax with a sensitivity of 188 ng/ml and a range of 0.312-20 ng/ml, SOD with sensitivity of 9.375 pgr/ml and range of 15.6-1000 pgr/ml, GPX with sensitivity of 18.75 pgr/ml and range of 31.25-2000 pgr/ml, CAT with sensitivity of 18.75 international milliunit per milliliter, and MDA with sensitivity of 18.75 ngr/ml and range of 31.25-2000 ngr/ml in the liver tissue with ELISA using ELISA reader model 2100 (Stat Fax, USA) and special kits (Finetest, China) were measured. It should be noted that the measurements were made according to the method in the manual of the manufacture kit. Data were analyzed by SPSS 20 software. Given that the results were small, the assumption of the natural distribution of data was investigated by Kolmogorov Smirnov test. One-way analysis of variance was used to compare the means between the studied groups and Tukey's post hoc test was used to compare the couples' group data. Also, data was reported as mean (mean ± standard deviation).
Findings by Text
In the group treated with 200 ppm silver nanoparticles compared with the control group, Bcl-2 levels and superoxide dismutase, glutathione peroxidase and catalase levels of the liver tissue significantly decreased, and the Bax and malondialdehyde levels were significantly increased (p<0.05). In treated groups with silver nanoparticles of 200 ppm with concentrations of 100, 200, and 300 μg / ml of crocin compared to the groups treated with silver nanoparticles of 200 ppm, Bcl-2 and superoxide dismutase, glutathione peroxidase and catalase levels of liver tissue were increased as dependent on the injectable dose, and the Bax and malondialdehyde levels decreased, depending on injectable dose (p<0.05; Table 1).
Main comparison to the similar studies
… [27-38].In a study that investigated the protective effect of crocin on renal function of diabetic rats, it was found that crocin-dependent administration could reduce serum levels of malondialdehyde in diabetic rats. Crocin was also reported to be effective in reducing stress-induced oxidative stress and reducing lipid peroxidation in reducing renal damage caused by diabetic nephropathy [39]. A study was conducted to determine the protective effect of crocin on sperm DNA damage in rats treated with cyclophosphamide. The results showed that crocin was able to reduce the amount of malondialdehyde and to reduce the oxidative damage of DNA. Crocin has also been reported to reduce free radicals induced by cyclophosphamide, protecting unsaturated fatty acids contained in sperm membrane and preventing the onset of a series of chemical reactions called lipid peroxidation [40]. … [41-45].
Suggestions
Additional studies are recommended for accurately understanding the molecular pathway of crocin in stress-oxidative reduction and inhibition of apoptosis induced by silver nanoparticles in the liver tissue.
Limitations
From the limitations of this study, the lack of necessary means to accurately recognize the mechanism of the crocin effect in reducing apoptosis, lipid peroxidation, and enhancement of antioxidant activity in chick embryo liver tissue treated with silver nanoparticles can be mentioned.
Conclusions
Crocin-induced injection leads to reduction of oxidative stress, decrease of lipid peroxidation and reduction of apoptosis in the chick embryo liver tissue through decreasing of toxicity of silver nanoparticles.
Acknowledgments
The author of this article thank The Young and Elite Scholars Club of The Islamic Azad University, Mashhad Branch.
Conflict of interest
Non-declared
Ethical Permissions
All stages of this study were designed and implemented in accordance with guidelines and international rules of working with laboratory animals under the supervision of the Young and Elite Research Club of Islamic Azad University, Mashhad Branch in 2016.
Funding Sources
This research was sponsored by the Young and Elite Research Club of Islamic Azad University.

TABLES and CHARTS

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