Evaluating the Nephro-protective and Hepatoprotective Effects of Hypoestes rosea in Acetaminophen - Induced Toxicity in Albino Rats by Ogregade in Journal of Clinical Case Reports Medical Images and Health Sciences

Evaluating the Nephro-protective and Hepatoprotective Effects of Hypoestes rosea in Acetaminophen Induced Toxicity in Albino Rats by Ogregade in Journal of Clinical Case Reports Medical Images and Health Sciencence 

Abstract

Hypoestes rosea leaves are in use as traditional medicine in most of the South – South states in  Nigeria and Western part of Cameroun for the treatment of various diseased conditions such as fever in children, anaemia, malaria in children etc. Despite its uses, there are scanty studies evaluating its organ protective effects. Therefore, this study evaluates the nephro-protective and hepatoprotective effects of Hypoestes rosea in acetaminophen-induced toxicity in Albino rats. The study objectives are to evaluate the protective effect of Hypoestes rosea on the kidney and the liver of albino rats. Acetaminophen being a commonly and popularly used analgesic and antipyretic drug, at high doses may be injurious and deleterious to vital organs of the body affecting the liver and kidney and the effects of aqueous extract of Hypoestes rosea (AEHr) on liver function parameters and kidney function parameters of acetaminophen induced-toxicity in albino rats were evaluated using acute (15 days) and sub-chronic (30 days) duration of study and study group comprising of prophylactic (pre-treatment) and therapeutic (post- treatment) phases with six experimental groups in each phase. A total of 112 adult apparently healthy Albino rats weighing (180-220g) were used for this study,  divided into six experimental groups of extract control (EC), negative control (NC), positive control (PC), AEHr100mg/kg b w., AEHr 200mg/kg b w., and AEHr 300mg/kg b w. groups each of six rats. At the end of the study period, chloroform anaesthesia in a desiccator was done after which blood samples were collected through jugular puncture for liver and kidney function parameters. Results showed that acetaminophen induced toxicity in albino rats caused nephrotoxicity and hepatotoxicity as evidenced by the elevated potassium,, urea & creatinine and reduced bicarbonate from the renal function parameters and also as evidenced by significant elevation of total and conjugated bilirubin and liver enzymes with a significantly reduced total protein and albumin levels from the liver function parameters when compared with other experimental groups .  However, various concentrations of aqueous extract of Hypoestes rosea in a dose dependent pattern at the different treatment phases at acute and sub-chronic period was able to restore the injury and nephrotoxicity and hepatotoxicity caused by acetaminophen induction to normal. Therefore, the results of this study suggest that Hypoestes rosea have active ingredients responsible for its nephro - protective and hepatoprotective properties in rats and should be subjected for further studies using higher mammals.

Keywords: Hypoestes rosea, acetaminophen, experimental groups, aqueous extract of Hypoestes rosea (AEHr), nephro-toxicity, hepatotoxicity, hepato-protective, nephro-protective, liver function parameters, kidney function parameters.

Introduction

The kidneys are vital organs that functions in maintaining homeostasis which is made possible with management of fluid levels, electrolyte balance, waste excretion, reabsorption of nutrients, maintaining pH, osmolality, regulation of blood pressure and secretion of active compounds. it is prone to stimuli or drugs causing nephrotoxicity, [1].

The liver is also a vital organ essential for maintaining homeostasis and metabolic integrity in the body, harboring important functions associated with regulation of carbohydrate, lipid, amino acid, and hormone metabolism, synthesis and degradation of plasma proteins, glycogen synthesis, storage of vitamins and metals, secretion of bile, xenobiotic metabolism and playing major role in the metabolism and removal of drugs among others. [2].

Several studies have demonstrated the induction of hepatocellular and or renal damage by acetaminophen overdose in experimental animals and humans. [3]. Drug-induced nephrotoxicity is increasingly recognized as a significant contributor to kidney disease including acute kidney injury (AKI) and chronic kidney disease (CKD). Nephrotoxicity has a wide spectrum, reflecting damage to different nephron segments based upon individual drug mechanisms. [4]. Both glomerular and tubular injuries are known targets for drug toxicity and may result in acute or chronic functional changes [5].Acetaminophen is generally safe at recommended doses but because the drug is available without prescription, it is potentially more dangerous than other similar drugs when used in excess or overdose [6]. Metabolically, acetaminophen is detoxified in the liver by oxidation through a minor cytochrome p450EI-mediatedpathway to produce a highly reactive cytotoxic metabolite, N-acetyl benzoquinone mine (NAPQI), which liver reduced glutathione (GSH) converts to a water soluble harmless product, mercapturic acid, [7]. The liver defense system succumbs to acetaminophen drug burden following the depletion of glutathione to pave the way for NAPQI accumulation, and oxidative stress ensues [8]

Natural plants products and their derivatives or herbal drugs have gained importance and popularity in recent years because it is considered safe, efficacious and cheap, [9]. Therefore, interest in the utilization of alternative medicines for the treatment of renal and hepatic diseases has been increased [10], since renal and liver diseases are important problem all over the world and it is increasing day after day. Plants have been used as a folkloric source of medicinal agents since the beginning of mankind. Hypoestes rosea is one of such plants with acclaimed folk medicinal usage and reported to possess anti- inflammatory, anti- cancer, anti- malarial and antioxidant properties [6,9,11,12,13]. The leaves are therefore medicinal plant products since it contains active organic ingredients employed in the treatment of diseases.

Hypoestes rosea commonly called ‘polka dot plant’, ‘freckle face’ and ‘morning glory lobelia’ is a broad - leafed flowering evergreen plant that belong to the kingdom; Plantae, Phylum; Tracheophyta, class; Magnoliopsida, order; Lamiales, family; Acanthaceaa, sub - family; Acanthoideae, Tribe; Ruellieae, sub tribe; Justiciinae and genus Hypoestes. Hypoestes phyllostachya ‘rosea’ is a tropical sub- shrub, a native to Madagascar, but found in most parts of the world especially West Africa. It has scientifically been proven to contain phytochemicals such as flavonoids, deterpenes and sterols, balsam, carbohydrates, monosaccharides reducing sugars, tannins and saponins [14].

However, there has not being adequate scientific data to support the nephro-protective and hepatoprotective potentials of Hypoestes rosea and provide information on its mechanism of action. This study therefore provides information on the ability of aqueous extract of Hypoestes rosea leaves to protect the kidney and the liver against acetaminophen-induced hepatocellular damage in albino rats.

MATERIALS AND METHODS

Plant Collection, Identification and Authentication

Fresh Hypoestes rosea leaves were collected from Ulakwo -1 in Etche LGA (4°59’ 27.00’’ N, 7°03 16 00’’E) Rivers state in Nigeria. It was identified by Dr. Osiyemi Seun 22/04/2019 with FHI no.: 112295 at the Taxonomy section of the Forest herbarium unit in the Forestry Research Institute of Nigeria, Ibadan.

Method of Extraction and Preparation of AEHr

The leaves of Hypoestes rosea were removed from the stem, washed and air dried under shade at room temperature for fourteen days (2 weeks) and then milled into powder. 450g of Hypoestes rosea powder were macerated in 1000 ml of water to dissolve for 48hr in a flask, the extract was decanted and then filtered through Whatman No. 1 filter paper to obtain a clear extract. The aqueous extract was further concentrated at 60°C using a rotary evaporator and dried using a freezer drier. The resulting crude extract which weighed 214 g was stored in a refrigerator maintained at 4-18°C until the analysis was over. The extracts were later weighed and reconstituted in distilled water to give the required doses of 100, 200 and 300 mg/kg body weight that were used in the study.

Collection of Experimental animals and Acclimatization

Albino rats were considered the animals of choice for this study because of its availability, cost, genetic make - up, its handling technique and the nature of the study. Adult apparently healthy albino rats weighing (180 – 220grams) were used. The rats were purchased from the Experimental Animal Unit of the Department of Human Physiology, University of Port- Harcourt. The rats were contained in conservative wire mesh cages under standard laboratory conditions. After the collection of the animals, they were weighed, identified and kept in wire gauge cages under favourable condition for two weeks. The animals were receiving food and water libitum and handled regularly so as to acclimatize with the environment. One hundred and fifty-six (112) albino rats 12 weeks’ old rats were used in this study. All animals handling protocols were in accordance with institutional guidelines for laboratory animals. (Ethic Reference Number PM/27/08/2011/MAA (R) and OECD guidelines.

Reagent’s Requisition and Preparation

Acetaminophen was purchased from Sigma Aldrich. They were prepared following standard procedures.

Experimental Design

Animal grouping

A total of one hundred and twelve (112) adult albino rats were assigned by weight into eighteen (18) groups and allowed to acclimatize for (fourteen) 14 days (2 weeks).The duration of the study was fifteen (15) days acute and thirty (30) days sub-chronic study. Eight (8) albino rats each were assigned for the two (2) positive control groups and six (6) albino rats each were assigned to the other groups.

Experimental Grouping and treatment regimen

The study groups comprised of two treatment phases each, prophylactic (Pre-treatment) and therapeutic (Post-treatment) phases, duration of treatment (Acute and Sub-chronic) with six experimental groups in each of the phases. In the prophylactic (pre-treatment) phases, the Albino rats were administered with AEHr before acetaminophen induction while in the therapeutic (post-treatment) phases, the albino rats were treated with AEHr after acetaminophen induction. The groups are as follows:

Group 1. Negative control (NC): Apparently healthy rats receiving de-ionized water and normal feed only.

Group 2.  Positive control (PC): 500mg/kg b w. acetaminophen induced rats at 14th day in acute and 29th day in Sub-chronic study.

Group 3. Extract Control (EC): Apparently healthy rats that received AHEr 100mg/kg b w. orally daily for fifteen (15) days and thirty (30) days.

Group 4. Acetaminophen induced treatment group aqueous extract of Hypoestes rosea of 100 mg/kg b w.

Group 5. Acetaminophen induced treatment group aqueous extract of Hypoestes rosea of 200 mg/kg b w.

Group 6. Acetaminophen induced treatment group aqueous extract of Hypoestes rosea of 300 mg/kg b w.

 Sample collection

Rats were anaesthetized using chloroform and were sacrificed on the 15th and the 30th days after an overnight fast. Blood samples were collected by puncture of the jugular vein and put into lithium heparin bottles for analyses of  liver and renal function parameters.

Laboratory analysis

The laboratory analysis was done using Mindray Biochemical analyzer (Model BS 120) using timed end point at the Research Laboratory of the departments of Biochemistry and Physiology, University of Port-Harcourt, Port -Harcourt.

Quality Control

Quality was adhered following standard operating procedures and good laboratory and best practices.

Ethical consideration

This study was carried out in accordance with the Guidelines of the Organization for Economic Cooperation and Development (OECD) 2001 and ethical approval was obtained from the University and Departmental Committee for Research and Ethics, University of Port Harcourt.

Data Analysis

Data were analyzed using SPSS version 23, they were presented as Mean ±SEM. Variations between were determined using Analysis of variance (ANOVA) and Tukey Test of Multiple Comparison used to differentiate variations in means between groups. P-values less than 0.05 (P<0.05) were considered statistically significant.

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