International Scholarly Research Notices

International Scholarly Research Notices / 2012 / Article

Research Article | Open Access

Volume 2012 |Article ID 309084 | 6 pages | https://doi.org/10.5402/2012/309084

Examination of Haematotoxicity of Fixed-Dose Highly Active Antiretroviral Drug in Albino Wistar Rats

Academic Editor: G. Biala
Received16 Sep 2012
Accepted18 Oct 2012
Published30 Dec 2012

Abstract

Highly active antiretroviral therapy (HAART) is considered toxic and has other life-threatening side effects. Our aim was to evaluate the haematotoxic effects of lamivudine, zidovudine, and nevirapine fixed-dose combinations in Albino Wistar rats. Fifty (50) three (3) months old male Albino Wistar rats weighing between 200 and 250 g were randomly assigned to five (5) groups (A, B, C, D, and E). Group A served as control. Two (2 mLs) of venous blood was aseptically collected on Days 5, 10, 15, 20, and 25 of treatment. Red blood cell (RBC) mean value recorded statistically significant increase ( ) in groups B and C when compared with the control group on Day 5. However, there was a statistically significant decrease ( ) in RBC, haemoglobin concentration (Hb), packed cell volume (PCV), and some red cell indices on Day 10. In addition there was no statistically significant difference ( ) in all the parameters evaluated when the test group was compared with the control on Day 25. Furthermore, there was a time-related statistically significant increase ( ) in the two major blood cells—RBC and platelet counts. From the result of this present study, it can be concluded that HAART when administered in fixed-dose combinations have no subacute haematotoxic effects.

1. Introduction

Antiretroviral drugs are medication for treatment of infection by retroviruses, primarily human immunodeficiency virus (HIV). When several such drugs, typically three or four, are taken in combination, the approach is known as highly active antiretroviral therapy, or HAART. The American National Institute of Health and other organizations recommend offering antiretroviral treatment to all patients with AIDS. Because of the complexity of selecting and following a regimen, the severity of the side effects and the importance of compliance to prevent viral resistance, such organizations emphasize the importance of involving patients in therapy choices and recommend analyzing the risks and the potential benefits to patients with low viral loads [1].

There is no doubt that HAART has been the most important progress in the therapy of HIV-infected patients in the last decade. A growing number of observations suggest that the beneficial effects of HAART also include improvement of HIV-related haematological complications [2]. Current guidelines for treatment of human immune-deficiency virus (HIV) infection recommend the combination of three antiretroviral agents, two reverse transcriptase inhibitors (RTIs) plus one protease inhibitor, or the association of three RTIs [1, 3]. These regimens of HAART have dramatically reduced the morbidity and mortality of HIV infection [2].

The values of haematologic parameters are affected by a number of factors even in apparently healthy populations. These factors include age, sex, ethnic background, body build, and social, nutritional, and environmental factors, especially altitude [46]. It has also been shown in several studies that some of the heamatological parameters exhibit considerable variations at different periods of life and disease conditions [79]. Although HAART has dramatically improved survival in HIV-infected patients, precautions still need to be taken to prevent HAART-related haematotoxicity and other life-threatening side-effects. Due to the fact that there are no available preclinical safety data on the treatment with the combinations of lamivudine, zidovudine, and nevirapine in animals, the aim of the present study was to examine the potential haematotoxicity of combinations of lamivudine, zidovudine, and nevirapine in Albino Wistar rats.

2. Materials and Methods

2.1. Drugs and Sources

The three (3) fixed-dose antiretroviral combination drugs used for the study were obtained from one of the accredited United States Presidential Emergency Plan for AIDS Relief (PEPFAR) centers in Enugu, Enugu State, Nigeria. The HAART considered in the study was a fixed-dose combinations (FDCs) of lamividine (150 mg), zidovudine (300 mg), and nevarapine (200 mg) per tablet. The drug was manufactured by Aurobindo Pharma Limited, Unit III, survey No. 313, Bachupally village, Quthubullapur Mandal, Ranga Reddy District (A.P) India.

2.2. Animal Care and Handling

This was according to the guidelines issued by World Health Organization (WHO), Geneva, Switzerland and the Indian National Science Academy (INSA), New Delhi, India on the care and use of laboratory animals. The Albino Wistar rats which were three (3) months old and weighing 200–250 g were selected from the colony maintained under the controlled conditions of temperature ( °C), humidity, and light (12 hours of light and dark) in the Animal House of University of Nigeria, Enugu Campus (UNEC), Enugu, Enugu State, Nigeria. The animals had free access to food (Standard pellet diet, starter feed) and clean tap water. The animals were housed in standard environmental conditions in wire-mesh-bottomed stainless steel cages.

3. Experimental Design

Fifty (50) three (3) months old male Albino Wistar rats weighing between 200–250 g were randomly assigned to five (5) groups (A, B, C, D, and E) according to similar body weight with ten (10) animals per group. Groups B, C, D, and E received graded doses (3, 6, 12, and 24 mg/Kg body weight, resp.) once daily for twenty five (25) days. Group A served as control. All administrations were by oral intubation.

3.1. Blood Sample Collection and Analysis

Two (2 mLs) of venous blood was aseptically collected on Days 5, 10, 15, 20, and 25 of treatment after overnight fasting from each animal into tripotassium ethylene diamine tetra acetic acid (K3EDTA) anticoagulant bottle. This was immediately mixed by gentle inversion and used in the determination of the haematological profile using a haematology auto analyser (Sysmex KX-2IN) following the manufacturer’s guidelines. The samples were analysed within two hours of collection. Two (2) animals were bled painlessly under chloroform anesthesia by ocular puncture through the retro-bulber plexus of the medial canthus.

3.2. Data Analysis

Data were analysed using Students’ “t”-test and one-way analysis of Variance (ANOVA) to determine significant differences between means. was regarded as significant.

4. Results

Red blood cells (RBCs) count mean value recorded statistically significant increase ( ) in groups B (3 mg/kg body weight) and group C (6 mg/kg body weight) when compared with the control group. However, total white blood cell (WBC) count showed statistically significant decrease ( ) in group B when compared with the control group. RBC and mean cell haemoglobin (MCH) mean values revealed dose dependent statistically significant decrease ( and ), respectively, in the one-way analysis of variance (ANOVA) when groups B, C, D, and E were compared on Day 5 (Table 1).


ParameterGroup A
0 mg/Kg
Group B
3 mg/Kg
Group C
6 mg/Kg
Group D
12 mg/Kg
Group E
24 mg/Kg

RBC (×1012/L) β ββ
HB (g/dL)
PCV (L/L)
MCHC (g/dL)
MCV (fL)
MCH (pg)
RETICS (%)
PLAT (×109/L)
WBC (×109/L)
NEUT (%)
LYMPH (%)
EOS (%)
MONO (%)
BAS (%)

RBC: red blood cell, HB: haemoglobin concentration, PCV: packed cell volume, MCHC: mean cell haemoglobin concentration, MCV: mean cell volume, MCH: mean cell haemoglobin, RETICS: reticulocytes, PLAT: platelets, WBC: white blood cell, NEUT: neutrophils, LYMPH: lymphocytes, EOS: eosinophils, BAS: basophils.
Compared with the control group , .
One way ANOVA for groups B, C, D, and E, .

However, RBC, haemoglobin (Hb), and packed cell volume (PCV) recorded statistically significant decrease ( ) in group E (24 mg/kg body weight) when compared with the control group. Also, mean cell volume (MCV) and MCHC revealed statistically significant decrease ( and ) when compared with the control group. Neutrophil count showed a statistically significant increase ( ) in group B when compared with the control group, while lymphocyte count recorded statistically significant decrease ( ) when compared with the control group. MCV revealed a dose-dependent statistically significant increase ( ) in the one way ANOVA when groups B, C, D, and E were compared on Day 10 (Table 2).


ParameterGroup A
0 mg/Kg
Group B
3 mg/Kg
Group C
6 mg/Kg
Group D
12 mg/Kg
Group E
24 mg/Kg

RBC (×1012/L)
HB (g/dL)
PCV (L/L)
MCHC (g/dL)
MCV (fL) β
MCH (pg)
RETICS (%)
PLAT (×109/L)
WBC (×109/L)
NEUT (%)
LYMPH (%)
EOS (%)
MONO (%)
BAS (%)

RBC: red blood cell, HB: haemoglobin concentration, PCV: packed cell volume, MCHC: mean cell haemoglobin concentration, MCV: mean cell volume, MCH: mean cell haemoglobin, RETICS: reticulocytes, PLAT: platelets, WBC: white blood cell, NEUT: neutrophils, LYMPH: lymphocytes, EOS: eosinophils, BAS: basophils.
*Compared with the control group * , ** .
βOne way ANOVA for groups B, C, D, and E β .

Only platelet count mean value recorded a dose-dependent statistically significant increase ( ) when all the test groups were compared on Day 15 (Table 3).


ParameterGroup A
0 mg/Kg
Group B
3 mg/Kg
Group C
6 mg/Kg
Group D
12 mg/Kg
Group E
24 mg/Kg

RBC (×1012/L)
HB (g/dL)
PCV (L/L)
MCHC (g/dL)
MCV (fL)
MCH (pg)
RETICS (%)
PLAT (×109/L)
WBC (×109/L)
NEUT (%)
LYMPH (%)
EOS (%)
MONO (%)
BAS (%)

RBC: red blood cell, HB: haemoglobin concentration, PCV: packed cell volume, MCHC: mean cell haemoglobin concentration, MCV: mean cell volume, MCH: mean cell haemoglobin, RETICS: reticulocytes, PLAT: platelets, WBC: white blood cell, NEUT: neutrophils, LYMPH: lymphocytes, EOS: eosinophils, BAS: basophils.
βOne way ANOVA for groups B, C, D, and E β .

In addition, there was no statistically significant difference ( ) in all the parameters evaluated when the test groups was compared with the control group on Day 20 (Table 4).


ParameterGroup A
0 mg/Kg
Group B
3 mg/Kg
Group C
6 mg/Kg
Group D
12 mg/Kg
Group E
24 mg/Kg

RBC (×1012/L)
HB (g/dL)
PCV (L/L)
MCHC (g/dL)
MCV (fL)
MCH (pg)
RETICS (%)
PLAT (×109/L)
WBC (×109/L)
NEUT (%)
LYMPH (%)
EOS (%)