Evaluation of the Effect of Nigella Sativa L. Seeds Melanin on Acetaminophen-Induced Hepatotoxicity and Nephrotoxicity in Rats

Abstract

Backgrounds: Recently, biopolymer melanin had been discovered and isolated from the outer black coat of the Nigella sativa L. (NSM) seeds. Objective: This study was designed to evaluate the hepatoprotective and/or the nephroprotective effects of the NSM against Acetaminophen (APAP)-induced liver and renal injuries in albino Wistar rats compared with N-Acetylcysteine (ACC), which pharmaceutically well-known as an antidote against APAP toxicities, and to investigate the safety of NSM on in vitro cell lines. As biological, the increases in demands to maintain a state of organs and/or all tissue balances within a specific range of the surrounding life; the zone of the melanin biomedical uses. However, data on NSM due to its recently discovered and using for research purposes appears to be lacked. In fact, there is no study that we are aware of, reporting any information about the NSM for any kind of liver/kidney protector in experimental rats’ species. Notably, using of Melanin might open a new avenue into how hepatocytes and/or kidney cells with NSM intraperitoneal (I.P.) injection rats may communicate. Method: four experiments were done; the first experiment was conducted to evaluate the effect of NSM on APAP hepatotoxicity in rats compared with ACC; the antidote drug against APAP toxicities in markets by using 30 albino rats for this purpose which were randomly arranged into 6 groups, each of 5 rats, group 1 served as untreated control; group 2 acted as acetaminophen control, while group 3 were given APAP antidote doses at 140 mg/kg/rats. Groups 4, 5 and 6 were treated with single daily doses of NSM at 20, 50 and 100 mg/kg/rats, respectively, one hour later all groups (2, 3, 4, 5 and 6) except group 1, received single daily oral doses of acetaminophen at 500 mg/kg/rats, all doses continued for14 days. In the second experiment, the evaluations of NSM against acetaminophen-induced nephrotoxicity in rats were done. Thirty albino rats were used and evenly divided into 6 groups each of 5 rats. Group 1 served as control group, group 2 served as APAP control group, and group 3 given ACC as an antidote for APAP toxicity (140 mg/kg/rats). Groups 4, 5 and 6 were received NSM doses at 20, 50, and 100 mg/kg/rats, respectively, and continued for 14 days. After 1 hour all groups except group 1 received oral doses of APAP at 500mg/Kg/rats, for 14 days. The third experiment was designed to study the toxicity of NSM on in vitro tests using both HEp-2 and Vero cells by allotted to two groups; group 1 HEp-2 cells and Vero cells served as control cell lines the other cell lines were treated with different NSM extracts concentrations in daily doses ranging from (8،16،32،64،125،250،500،1000،2500μg/mL). Morphological signs were observed according to changes in incubated cell lines for three days. Then after incubation, results were collected to find out the cytotoxicity avoidance following cells treated with NSM and viability% of live in vitro cells measure by using MTT assay. Last experiment which was designed to study the toxicity of the NSM by using twenty albino rats after allotted to three groups; group 1 normal control received distilled water, groups 2 and 3 were given NSM (i.p.) in daily doses (50 and 100 mg/kg) continued for 10 days. Clinical sign was observed. Rats were sacrificed and blood sample was collected for blood work study and serum analyze, there were no changes in almost all parameters in all groups treated with NSM compared to control. Tissue specimens of liver, kidney, intestine and testes were taken for histopathology to support the previous results. Results: Clinical signs were regularly observed and rats were sacrificed after 14 days and the blood samples were collected for biochemical analysis for liver enzymes of serum markers such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphates (ALP) and thus bilirubin were significantly increased in APAP-treated rats compared to the control, and in groups treated with NSM extracts and ACC were significantly decreased compared to the control APAP rats. The biochemical observations were correlated with histopathological examination of rat liver sections. Serum concentrations of urea, creatinine, cholesterol and total lipids were significantly increased, while total albumin and protein were significantly decreased in APAP-treated rats when compared to control group. In groups treated with NSM, all this concentration of sera were significantly altered and became closer to the concentrations of the rats in control groups. Histological examinations confirmed these results when kidney of APAP groups showed severe hemorrhage, congestion, shrinking of the glomeruli with necrosis of renal tubules, while in groups 4, 5 and 6 which were received ACC and the NSM kidney sections showed mild congestion and lymphocytic infiltration particularly with NSM at 50 mg/kg. Conclusion: In fact, current experiment substantiates with the constellation of several biochemicals, hematological, histological and morphological measurements. NSM able to possess hepatorenal protection against APAP-induced liver and kidney damages and even in much better than ACC. NSM when injected (I.P.) on in vivo and in vitro up to high concentration.

Keywords: Melanin; Nigella sativa seeds; Acetaminophen; N-Acetylcysteine; Ahepatotoxicity; nephrotoxicity.