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Beneficial Effects of Black Seed Oil on Blood, Hormone Profile, Sperm Parameters and Histotexture of Testis of Male Mice Exposed to Bisphenol-a

Khaled Mahmud Sujan
Department of Physiology, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh

Mohammad Alam Miah
Department of Physiology, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh

Afrina Mustari
Department of Physiology, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh

Md. Kamrul Islam
Department of Physiology, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh

Abstract/ Executive Summary:

Bisphenol-A (BPA) is an endocrine disruptor that is widely used in the manufacture of polycarbonate plastics, epoxy resins and dental sealants. It has adverse effects on reproduction. Black seed oil is frequently used as folk medicine for the promotion of good health and treatment of many ailments. The study was carried out to investigate the effect of Bisphenol-A (BPA) and black seed oil (BSO) on hematological parameters, hormone, sperm parameters and patho-physiological changes of testis in mice. A total of 15 male Swiss Albino mice (Mus musculus), of 25-28 days were used and randomly divided into 3 groups consisting 5 mice in each group. Group A served as vehicle control whereas group B and group C were as BPA and BPA plus BSO group respectively. Both BPA @ 50 mg/kg/day and BSO @ 1ml/kg/day were administered orally. Results revealed that BPA caused reduced hematological parameters (Hb, TEC, and PCV) in mice. Inclusion of BSO in BPA treated mice prevented the alterations of those values. Serum T4 and testosterone concentration were significantly (p<0.01) decreased in the BPA-treated mice whereas BSO dramatically improved the levels of these two hormones. There was reduction in the sperm count and sperm motility along with increased head and tail abnormalities in BPA treated mice. BSO reduced the abnormal alterations of sperm parameters. Histopathological analysis of testis demonstrated that there were degenerative and necrotic changes in the seminiferous tubules in mice upon treatment with BPA. It can be concluded from this study that BPA has harmful effects on hematology, hormonal assay, sperm parameters and testis. These harmful effects could be alleviated by ingestion of black seed oil.

Keywords: Bisphenol-A, Black Seed Oil, Mice, Hematology, T4, Testosterone, Sperm and Testis

Location: The Department of Physiology, Bangladesh Agricultural University, Mymensingh, Bangladesh

Start Date: 01-02-2018

End Date: 25-04-2018

Program Area: Pest Management

Commodity: Mice

Objectives:

To investigate the counteraction effects of BSO on physiological alteration in BPA treated male mice.

Methodology:

Experimental design: The experiment was conducted in the Department of Physiology, Bangladesh Agricultural University, Mymensingh, from 1 February to 25 April 2018. In this study, total 15 male Swiss Albino mice (Mus musculus), aged 25-28 days with an average body weight of 27.4 g were used. The mice used for this study were purchased from ICDDR’B, Dhaka. At first, mice were randomly divided into 3 groups viz., A, B and C consisting 5 mice in each group. Group A served as vehicle control (received only basal mouse pellet feed mixed with sunflower oil) whereas group B was administered BPA @ 50 mg/kg body weight daily, formulated in sunflower oil (as vehicle for BPA), while group C received BPA and BSO @ 50 mg/kg bw/day and 1ml/kg bw/day, respectively. The experiment was carried out for a period of 12 weeks. They were reared in a compartmentalized square wooden cages wrapped with wire mesh under controlled conditions of temperature (26-30) °C and relative humidity of 70-80% with natural day light. Bisphenol-A (BPA) was purchased from Sigma-Aldrich Company, USA. The pure oil extract of Nigella sativa L. seeds was bought from local market in Mymensingh. Ethical approval The present study and all experimental procedures were approved and performed according to the guidelines for the care and use of animals as established by Animal Welfare and Experimentation Ethics Committee, Bangladesh Agricultural University, Mymensingh [AWEEC/BAU/2019(36)]. Blood collection and serum preparation Blood samples were collected by cardiac puncture at 12th week by sacrificing the mice. About 1 to 1.5 ml blood was collected and transferred half of blood into anticoagulant containing eppendrop tube and the remaining half of blood was transferred to another tube without anticoagulant for serum preparation. The serum samples were separated by centrifugation and collected by using 200 µl pipettes. Serum samples were stored in capped tube at -20ºC for hormonal analysis. Hematological parameters Hemoglobin (Hb), Packed Cell Volume (PCV), Total Erythrocyte Count (TEC) and Erythrocyte indices: mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC) were performed as per methods indicated by Ghai (2012). Hormonal assay The hormonal parameters: Serum Testosterone and Serum Thyroxine (T4) were determined by using Testosterone Radioimmunoassay Kit and T4 Radioimmunoassay Kit respectively at the Institute of Nuclear Medicine & Allied Sciences (INMAS), Mymensingh Medical College. Sperm physiological parameters At the end of the experiment, all the mice were sacrificed by using anesthesia and testicles were removed to collect epididymis for evaluation of sperm. Epididymal sperm motility was made by transferring a small amount of the diluted suspension (PBS) on a pre-warmed slide and then applying a cover slip and observed at low power of the microscope. At least ten widely-spaced fields were examined to provide an estimate of the percentage of motile sperms. Sperm motility was recorded in percentage (%) (Saalu et al., 2010). Histopathology The testis (3) from each group of mice were collected after completely removal of blood by perfusion with phosphate buffered saline and kept in 10% neutral buffered formalin for 15 days. The well-fixed tissues were processed, sectioned and stained with Hematoxylin and Eosin (H & E) for histo-pathological as per standard procedure Banchroft et al. (1996) in collaboration with the Department of Pathology, Mymensingh Medical College, Mymensingh. The stained slides were observed under Optika Vision Lite 21 and photographs of these slides were taken at the Department of Physiology, Bangladesh Agricultural University. Statistical analysis The data was placed and stored in Microsoft Excel- 2010 and imported to the software IBM SPSS Statistics 20 for analysis. One-way analysis of variance (ANOVA) was used to determine the effect of different parameters. Descriptive statistics analysis was done to measure the mean, standard deviation and standard error and p value of different parameters. Because of using multiple comparisons, the corrected p value was calculated adjusted at 0.01 and 0.05 considered for level of significance.

Source of Information: J Bangladesh Agril Univ 19(2): 000–000, 2021; ISSN 1810-3030 (Print) 2408-8684 (Online)

Article Link (Online Journal): https://doi.org/10.5455/JBAU.68360

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The research findings suggest that BPA caused hematological alterations and suppressed the sexual behavior of male mice, which might be attributed to a decrease in testosterone and T₄ levels. It affects the germ cells, leading to spermatogenesis and reduction in sperm count, sperm motility percentage, and increased head and tail abnormality percentage. BSO might play a vital role in preventing BSA-induced infertility and reproductive health hazards by regulating the hypothalamo-pituitary-gonadal hormone axis.

Knowledge Management: Journal