Vernonia Amygdalina Del (Bitter Leaf) extract ameliorates isoniazid (INH) induced liver injury in Swiss Albino Mice

Liver plays a central role in the metabolism of drugs. Drug clearance and transformation exposes liver to toxic injury. Antitubercular drugs have been found to be hepatotoxic and potentially lead to drug-induced liver injury. Isoniazid is one of the most hepatotoxic first line antitubercular drugs. Conventional drugs used in the treatment of liver disease are often inadequate and a search for supplementation or alternative drugs for the treatment of hepatic damage is indispensible. Therefore our study aims to investigate the hepatoprotective potential of Vernonia Amygdalina Del (bitter leaf) extract against Isoniazid-induced liver injury in Swiss Albino Mice. Treatment of Mice orally with Vernonia Amygdalina Del extract at dose of 250mg/kg and 375 mg/kg significantly lowered (P<0.05) the serum level of liver enzymes in Isoniazid pretreated mice. The hepatoptotective activity of the extract found to be comparable with the standard drug, Silymarin (100 mg/kg, P.o.). Moreover, treatment with the extract significantly alleviated Isoniazid induced hepatic injury as supported by the photomicrographs of liver section of mice. The data shows aqueous Vernonia Amygdalina Del extract has a very promising hepatoprotective potential against isoniazid-induced liver injury.


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Liver is the largest organ of the body weighing approximately 1500g, located in the 46 upper right quadrant of the abdomen, anterior to the right kidney and inferior to the diaphragm [1]. The organ is responsible for over 500 metabolic functions and maintenance of Isoniazid has been widely used for treatment of tuberculosis. However, if the drug is 68 consumed in overdose or for a long period of time susceptible individuals might suffer sever 69 hepatotoxicity. Its effect may range from mild increase in serum liver enzyme activity to 70 sever forms of hepatocellular necrosis or intrahepatic cholestasis [8]. Mild and transient 71 serum enzyme increases occur in 10-20% of the patients taking the drug and severe 72 hepatotoxicity occurs in about 1-3% of patients [9]. Isoniazid is the most hepatotoxic 73 antitubercular drug [7,8]. The drug received a black box warning from the US Food and 74 Drug Administration (FDA) due to its high incidence of adverse drug reactions in resulting 75 hepatocyte injury [10]. Various metabolites of isoniazid have been suggested as being 76 hepatotoxic, including hydrazine, monoacetyl hydrazine, acetylisoniazid and isonicotinic acid 77 [11]. Administration of acetylhydrazine or acetylisoniazid in rats leads to the production of 78 reactive alkylating species that covalently bind to liver proteins, causing hepatocyte injury 79 [12]. In another study, Isoniazid induced toxicity resulted a significant elevation in the level 80 of Liver enzymes, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and 81 alkaline phosphatase (ALP) [13]. Therefore in this study we used Isoniazid to induce 82 hepatotoxicity in Swiss Albino Mice and the doses of Isoniazid used to induce hepatotoxicity 83 was adopted from Chen et al., 2011 [14]. 84 Although Isoniazid (INH) is known to be potentially hepatotoxic, because of its 85 efficacy the drug remains a mainstay for the treatment of tuberculosis [15]. Conventional 86 drugs used in the treatment of liver disease are often inadequate. It is therefore search for 87 supplementation/ alternative drugs for the treatment of hepatic damage caused by anti-88 tubercular drugs.

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A total of 200g shade dried leaves were powdered in an electrical grinder and soaked 91 in 1.8 litres of distilled water. The filtrate was frozen and lyophilized to dryness. After 92 drying, a dark brown extract of Vernonia Amygdalina was stored at 4 0 C until tested in the 93 animals. The animals were assigned to one of six groups, each group consisting of six mice.

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The control mice were orally injected with sterile saline (1 mL/kg

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The reagent cuvette was incubated at a temperature of (37ºC ± 0.5 ºC) for the duration 181 of the test, then 50 µl of serum sample was mixed with 500µl of the working reagent and The reagent cuvette was incubated at (37 o C ± 0.5 o C) for the duration of the test.

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Then 20 µl of serum sample was mixed with 500 µl of the working reagent and incubated for 224 1min at 37 o C. The absorbance was read at 405 nm exactly after 1, 2, and 3 minutes and from 225 the reading the mean absorbance change per minute was calculated. The rate of the reaction is 226 directly proportional to the alkaline phosphatase activity. Finally the activity of alkaline 227 phosphatase in the sample was calculated from the mean absorbance change per minute.

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The level of ALT, AST, ALP and total protein were analyzed using Auto lab 229 18 Analyzer (fully automated chemistry analyzer, Italy).      Table 3 indicated that Vernonia Amygdalinadel treatment resulted in a significant decrease 340 (p<0.01) in liver/body weight % in isoniazid-pretreated group compared to only Isoniazid 341 treated group. This is probably due to the healing of liver to perform all its physiological 342 functions including burn the fat it had accumulated inside its cells.  properties, which are mediated by antioxidant activity in mice in vivo.

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There was a problem of developing an animal model of isoniazid-induced 430 hepatotoxicity. Because of its short half-life, it was found that smaller, more frequent doses 431 of isoniazid lead to greater hepatotoxicity than one large dose. But we managed this 432 challenge by providing the mice with the minimum toxic dose of the drug. The other 433 challenge was it is recommended to give the drug in food because isoniazid has a short half-434 life, and this provided a more consistent blood level than once-a-day oral gavages. This 435 method of drug administration produced blood levels in mice that were comparable to the 436 isoniazid in humans. However, the mice were not voluntary to eat the pellet mixed with the 437 drug. So we managed the problem by giving the drug in a solution via oral gavage.