Effect of Artemisinin-Based Combination Therapy on some Selected Liver Function Indices of Pregnant Wistar Albino Rats

Ugian E.A; Dasofunjo K; Nwangwa J.N; Asuk A.A; Akam M.S; Ajing E.N; Ugoanyanwu F.U. Department of Medical Biochemistry, Cross River University of Technology, Okuku, Cross River State, Nigeria. Department of Physiology, University of Calabar, Calabar, Cross River State, Nigeria Department of Biochemistry, University of Nigeria, Nsukka, Enugu State, Nigeria Department of Biochemistry, University of Calabar, Calabar, Cross River State, Nigeria.


INTRODUCTION
Malaria is a vector-borne infectious disease caused by different strains of the protozoan parasites of the genius plasmodium.Malaria still remains one of the most deadly infections in the tropical and subtropical regions of the world despite various control programmes (Okafor et al., 2013).Each year, there are about 216 million episode of acute malaria illness, 655,000 mortality and 91% occurred in the African region (CDC, 2012).The increasing resistance to the available pharmacological agents with antimalarial potentials call for more findings on malarial control in endemic countries (Ridley, 2002).One of the chemotherapeutic drugs recommended by the World Health Organization (WHO) in the management of malaria are artemisinin-based combination therapy which prevents the development of gametophyte in Plasmodium vivax, .Plasmodium ovale, Plasmodium malarae and the early stage of Plasmodium falciparium.
Therefore, since relevant information about the effect of artemisinin-based combination therapy on the hepatobilliary system of pregnant women have not been fully elucidated, hence, this present research is focussed on the effect of artemisinin-based therapy on some selected liver function indices of pregnant wistar albino rats.

SAMPLES
The drug sample (Coartem) was obtained from Rabana pharmacy, Calabar while other samples and reagents used were of analytical grade.

Animals and treatment protocols:
Fifteen (15) pregnant wistar albino rats weighing 200-220g were used for this work.The animals were obtained from the animal holding unit of the Department of Biochemistry, University of Calabar, Calabar, Cross River State.The animals were housed in plastic cages and were allowed acclimatization period of Seven ( 7) days in a well-ventilated room with a temperature and pressure of 29±2 O C and70% respectively.The pregnant wistar albino rats were maintained with rat chow (vital feeds Ltd) and water ad libitum.
The animals were exposed to 12 hours light-dark cycle and handled according to standard protocols.At the end of acclimatization period, they were divided into three groups A, B and C of five (5) pregnant rats each.Group A served as control while B and C were the test groups.The control group was treated with 0.5mls of normal saline while B and C were treated with 0.5mls corresponding to 3 and 6 mg/kg body weight of the drug respectively and were administered intra-peritoneally for seven (7) days.
The pregnant wistar albino rats were sacrificed 24 hours after the last administration in accordance with the guide lines of the European Convention for the protection of vertebrate animals and other scientific purposes-ETS-123 (European Treaty Series, 2005).

Preparation of Serum
The animals were anaesthetized in a jar containing cotton wool soaked in ether.When the animals became unconscious, they were brought out quickly of the jar, the abdominal region was opened along the linear Alba and diaphragm cut with scalpel blade to expose the organs and blood was collected into a sterile sample container by cardiac puncture.Blood was collected into a clean, dry centrifuge tube and allowed to clot for 30 minutes before centrifuging at 300rpm x 10min using Uniscope Laboratory Centrifuge.
The serum was thereafter aspirated into clean, dry, sample bottles using Pasteur pipette and was kept or stored in sample bottles and used within 12hours of preparation as described by (Malomo, 2000).Later it was transferred into specimen bottles before being used for biochemical analysis.

Statistical Analysis
Statistically analysed data used was presented as mean ± SD of five (5) determinations.Statement analysis was carved out using one way analysis of variance (ANOVA).Differences were statistically significant at P<0.05 (Mahajan, 1997).

RESULTS
The results below depict the effect of administration of artemsinin-based combination therapy on some selected liver function indices of pregnant wistar albino rats.Table 1 showed a significant increase (P<0.05) in serum ALT, AST and ALP.The ratio of AST: ALT and AST: ALP produced a significant (P<0.05)increase following the administration of the drug to the pregnant wistar albino rats (Table2).More so, artemisinin-based combination therapy produces a significant (P<0.05)elevation on both total and conjugated serum bilirubin level at both 3mg and 6mg per kg body weight when compared with control (Table 3).

DISCUSSION
The hepatocyte membrane distortion is associated with membrane leakage of the hepatocyte cytosolic contents which is manifested by significant elevation of serum/plasma enzymes of acute hepatocellular damage namely ALT, AST and ALP as a marker of hepatocellular damage (Bhaltacharyya et al., 2003).However of this marker enzymes, ALT is the most reliable.AST is known to be abundant in the cardiac muscles, skeletal muscles, kidneys and testes.Thus, any disease state affecting any of these extra hepatic tissues, significantly elevates the serum level of enzymes (Olayinka and Ore, 2013).Therefore, the observed significant increase in serum ALT, AST and ALP when compared with the control at both dosages of the treated animals suggest that the drug might induce hepatic damage or hepatotoxicity in the pregnant albino rats.These findings are similar to the findings of other researchers (Farombi et al., 2000;Obi et al., 2004;Pari and Anuli, 2005).Researchers have shown that other antimalarial drugs such as chloroquine, amodiaquine, quinine and halofrantrine were also reported to elevate serum ALT and ALP and may induce hepatic damage (Nwanjo and Oze, 2007;Adaramoye et al., 2008;Udobre et al., 2009).The observed elevation AST: ALT and AST: ALP ratios, are indications of hepatic injury (Nyblom et al., 2004;Nyblom et al., 2006) .
Likewise, the significant increase in the serum level of both total bilirubin and conjugated bilirubin is an indication that the drug (ACTs) might induce injury to the hepatic tissue or caused conjugated hepatobilliary injury on the pregnant wistar albino rats which might also signal an inimical effect on the foetus.Though pregnancy itself induces some abnormalities on the liver, including cholestasis (resulting in raised conjugated bilirubin and ALP) and in cases of pre-eclampia, the HELLP Syndrome (Haemolysis elevated liver enzymes and low platelet) Jamjute et al., 2009).It might thus become necessary to exercise special caution in the usage of artemether-lumefantrine in the management of malaria in pregnancy.Artesunate -sulfadoxine/pyrimethamine might thus continue to remain the drug of choice in the intermittent prevention of malaria in pregnancy (Schantz et al., 2009).Thus, it is logical to conclude that various alterations in the selected biochemical parameters studied above showed that artemisinin-based combination therapy might induce hepatotoxicity or hepatobilliary injury in pregnancy.

Table . 1
: Effect of artemisinin-based combination therapy on serum enzymes.