ABSTRACT
The purpose of the study was to evaluate the radioprotective effects of Gongronema latifolio (GL) leaf extract on a whole-body irradiated wistar albino rats. A prospective experimental and cross-sectional design was adopted for this study and it included a control group and experimental group. Part of the control group (normal control NC) was not irradiated neither was it administered with GL extract but the other part (experimental control EC) was only exposed to graded radiation doses (GRDs). In the experimental group, the pre-treatment group (PRT) received GL extract orally before being exposed to GRDs while post-treatment group were exposed to GRDs before receiving GL extract orally.
Phytochemical analysis of GL extract was done to re-determine the bioactive constituents of the extract. Physical changes were observed and recorded in all the groups using weight loss as an index. The blood samples of the animal groups were collected before and after irradiation (IR) for following analysis namely liver function test (LFT) {which includes-Alkaline phosphase (ALP), Alanine amino-transferase (ALT), Aspatate amino-transferase (AST)}, and antioxidant enzymes tests like Malondialdehyde (MDA), Glutathione (GSH), Catalase (CAT) and Superoxide dismutase (SOD)}.
The result of the phytochemical analysis revealed the presence of the following bioactive agents- alkaloids (3.11mg/g), tannins (2.43mg/g), flavonoids (1.31mg/g), phenols (1.10mg/g) and saponin (0.8mg/g). Body weight of the rats exposed to 6Gy in EC (51g) significantly (p<0.05) decreased when compared to NC (115g) and PRT (70g) but not significantly (p>0.05) different from PST (60g) group. ALP mean levels recorded in rats exposed to 4Gy increased (p<0.05) significantly in EC (74iU/L) when compared to PRT (37iU/L), PST (43iU/L) and NC (39iU/L) group on day 8 after IR. ALT mean level for rats exposed to 4Gy elevated (p<0.05) significantly in EC (50iU/L) relatively to PRT (31.67iU/L), PST (38.67iU/L) and NC (37iU/L) on day 8 after IR. MDA activity levels for rats exposed to 6Gy significantly (p<0.05) increased in EC (70%) relatively to PRT (35%), PST (59%) and NC (36%) on day 8 after IR. For rats exposed to 2Gy, GSH % activities decreased (p<0.05) significantly in EC (26%) when compared to PRT (59%) and NC (69%) on day 8 after IR. For rats exposed to 4Gy, CAT % activities significantly (p<0.05) decreased in EC (31%), PRT (49%) and PST (44%) relatively to NC (79%) on day 8 after IR. For rats exposed to 2Gy, SOD % activities decreased significantly in EC (29.33%), PRT (50.67%) and PST (40.67) when compared to NC (75%) on day 8 after IR.
Consequently, the result obtained suggested that GL extract emeroliates oxidative stress induced by ionizing radiation, thus affirming its radioprotective potentials. The result also demonstrated that the extract was more effective in PRT group relatively to PST group
CHAPTER ONE
INTRODUCTION
1.1 Background of the study
With the discovery of x-rays in 1895 and radioactivity in 1896, the biologic effects were also observed shortly after. Within the first six months of its use in treating patients, several cases of erythema, dermatitis and alopecia were already reported among x-ray operators and their patients. The first report of a skin cancer ascribed to x-rays was reported in 1902, followed eight years later by experimental confirmation, Bushberget al., (2002).
Radiation medicine is one of the major sources of ionizing radiation due to its numerous applications in the hospital. Other sources of radiation exposure include radon in houses, contamination from weaponstesting sites, nuclear accidents and cosmic rays.Today, ionizing radiation is not only employed in treatment of diseases and industry but also in developing new varieties of high-yielding crops and enhancing storage period of food materials. Radiotherapy is one of the common sources of ionizing radiation and more so one of the most common modality used for treating human cancer. About 80% of cancer patients need radiotherapy at some time or the other either for curative or palliative purpose, Cherupally et al., (2001). It is essentially used in the treatment of a number of malignancies, but frequently its use is limited due to its adverse effects on normal tissue.
The effects of radiation on human cells/tissue can be divided into somatic and genetic effects. Somatic effects are harms exposed individual suffer during their life time such as radiation induced cancers, opacification of the eye etc, while genetic effects are radiation induced mutation to an individual genes and DNA that can contribute to the birth defective descendants Podgorsak, (2005). Somatic effects of radiation exposure can be classified as either stochastic or non-stochastic. A stochastic effect is the effect in which the probability of the effect, rather than its severity, increases with dose. Radiation-induced cancer and genetic effects are stochastic in nature. Stochastic effect is believed not to have a dose threshold. In non-stochastic effect, there is a threshold dose below which the effect is not seen. Cataract, erythema, fibrosis and hematopoietic damage are some of the non-stochastic effects that can result from large radiation exposure.
Radiation interactions that produce biologic changes are classified as either direct or indirect action. The change takes place by direct action if biologic macromolecules such as deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or proteins become ionized or excited by an ionizing particle or photon passing through them or near them.The DNA damages caused per Gray are about 1000 single strand breaks (SSB), 40 double strand breaks and 950 base depurination. Roughly 4.4 x 107 single strand breaks, 1.4 x 107 double strand breaks and 1.1 x 107base lesion per year occur per mammalian cell, Fleck, et al,.(1999). Indirect effects are the result of radiation interactions within the medium (e.g. cytoplasm or water) which creates highly reactive free radicals chemical that in turn interact with the target molecule, Bushberget al., 2002). Because 70% to 85% of the mass of living system is composed of water, the vast majority of radiation-induced damage from medical irradiation is mediated through indirect action on water molecules. Exposure of biological tissues to ionizing radiation immediately leads to ionization and excitation of their constituent atoms. The molecules where the atoms reside then dissociate, resulting in so called free radicals, Mayles et al., (2007). This free radicals are reactive oxygen species such as hyoxyl radical (OH), superoxide radicals ( ), singlet oxygen and peroxyl radicals (ROO) in irradiated tissue that incite several pathophysiological changes in the body, Maurya, et al., (2011).
Free radicals can diffuse in the cell, producing damage at locations remote from their origin. They may inactive cellular mechanisms directly or via damage to genetic material (DNA and RNA), and they are believed to be the primary cause of biologic damage from low linear energy transfer (LET) radiation, Bushberg et al., (2002). It is estimated that two-thirds of DNA damage is caused indirectly by scavengeable radicals (Root and Okada, 1972), as reported in lobachevsky, et al., (n.d).Generally ionizing radiation causes either excitation or ionization or both to atoms and molecules which lead to the following conditions.
- Generation of free radicals as mentioned earlier.
- Breaking of chemical bonds.
- Formation of new chemical bonds and cross-linkage between macromolecules.
- Damage to biomolecules (e.g. DNA, RNA, Lipids, Proteins) which controls or regulates vital cell processes.
The detrimental consequences of irradiation (IR) of cells and tissues can be encountered in cancer radiation therapy. Apart from normal tissue damage, another issue associated with cancer radiotherapy is the potential for emergency of secondary radiation-induced cancers, affecting more than 1% of patients (Hall, 2006).Severally protective mechanisms have been adopted in radiotherapy to reduce oxidative stress in patients and it includes;
- Physical protection (E.g. Conformal radiotherapy, intensity modulated radiotherapy IMRT etc).
- Biological protection (E.g. hyperfractionation and Ultrafractionation).
Attempts have also been made to protect personnel working inradiation medicine departments, radiopharmaceutical centers, nuclear power operations, aviations, uranium miners and other sources of ionizing radiation through the provision of the following; personal dosimeter, shielding devices, radiation detection equipment and other safety procedures, policies etc so as to ensure safety of patient, occupational staff and the general public. But the truth is thationizing radiation and radioactive substances are natural and permanent features of theenvironment,and thus the risks associatedwith radiation exposure can only be restricted and cannot be eliminated entirely.
Consequently, attempt to mitigate radiation toxicity in normal cells/tissues and in a whole organism are of significant clinical importance and an area of active research, considering the fact that ionizing radiation is on increase in numerous aspect of human life. There isexigency to develop and improve on another protective mechanism aside from the ones mention earlier that can mitigatenormal tissues from toxic effects of radiation.It has also been considered realizable that radiation therapy for cancer patients could be enhanced by the use of radioprotectors to protect normal tissues from unwanted radiation exposure.
Radioprotectors are compounds that are designed either to mitigate or prevent the damage caused by radiation in normal tissue. These compounds are often antioxidants and must be present before or at the time of radiation for effectiveness. It has also been found in the studies that chemical agents given after radiation exposure may assist in DNA repair activities, reduce inflammation and persistent radiation-induced oxidative stress and facilitate death pathways (apoptosis) of damaged cells, Kumud et al.,(2014). Other agents, termed mitigators, may be used to minimize toxicity even after radiation has been delivered, Deborah et al., (2010).
A number of compounds have been evaluated under the anti-irradiation drug development program, in 1948 for the first time, Patt et al., reported that cysteine is an effective radioprotector and showed that it can protect mice from harmful effects of total body x-ray irradiation when administered before radiation exposure. Badr et al., (1999) in their study suggested that melatonin administration confers protection against damage inflicted by radiation when given prior to exposure to irradiation and not after, and supports the contention that melatonin radioprotection is achieved by its ability as a scavenger for free radicals generated by ionizing radiation. The radioprotective effect of abana, following a total body irradiation was studied by Baliga et al., (2004). Their result indicates that the radioprotective activity of abana may be due to free radical scavenging and increase GSH levelin the irradiated mice.
Several chemical compounds have been synthesized and tested for their radioprotective ability (Sweeney, 1979). The major disadvantage of some these compounds has been their high toxicity at the optimum protective dose (Sweeney,1979), which forestall their effective use in man.
Gongronema latifolium (GL) is an edible plant, less toxic, relatively cheap and available, thus, it is considered a possible radioprotective material. This study therefore aims at providing information on the radioprotective effects of GL on wistar albino rats whose whole-bodies were exposed to different doses of radiation.
- Objectives of the study
- This research investigates the possible radioprotective effect of Gongronema latifolio(GL) extract on a whole-body irradiated wistar rats through the following specific objectives.
- Tore-determine the phytochemical constituent of GL extract, so as to find out the bioactive constituents of the leaves.
- To observe any physical changes following graded doses of radiation to wistar albino rats.
- To determine any radioprotective effects of GL by measuring changes in liver enzymes following exposure to graded radiation doses (GRDs).
- To determine lipid oxidative degradation using malondialdehyde (MDA) as an index for radiation damage in un-irradiated and radiated animal groups.
- To determine the scavenging of free electron activity in all the animal groups following exposure to graded radiation doses by measuring the antioxidant enzymes.
- To compare the radioprotective effects of GL extract in both pre-treated animals and post-treated animals exposed to radiation.
1.3 Justification of the study
So far it is only few compounds that are radioprotectors registered forhuman use that has shown good radioprotective effects.However, they have significant shortcomings including relatively high toxicity and unfavorable routes of administration, which negatively affect their application and efficacy,(Lirenet al., (2010).
- This very study will be a contribution in the search for new cost effective and relatively less toxic radioprotectors.
- This study will provide information on whether latifolio can serve as prophylactic agent, mitigator or therapeutic agents, in whole-body irradiated rats.
- Results obtained in this study will also contribute significantly to the growing search for radioprotectors.