LEAD ACETATE: A DANGEROUS ENVIRONMENTAL TOXICANT THAT IS AMELIORATED BY THE AQUEOUS EXTRACT OF FICUS

To investigate the effects of lead toxicity on the histoarchiteture of the vagina and cervix of adult female Wistar rats and the currative role of FicusVogelii ( Fv)in lead toxicity. The experiment lasted for a period of 21 days involving 20 adult female albino Wistar rats with an average weight of 145g. The rats were divided into five (5) groups A, B, C, D and E. A is negative control group that received normal saline(why?), B is positive group that received lead acetate solution, C received Aqueous extract of Fv (100g/kg) and lead acetate solution,D receivedaqueous extract of Fv (300g/kg) and lead acetate solutiongroup while E received aqueous extract of Fv only. Examination of the vagina and cervixhistologically revealed various damages such as, necrosis, oedema and reduction is vaginal gland size suspected to be as a result of lead toxicity. These changes were ameliorated by the administration of Fv extract. Exposure to lead also produced significant reduction in blood supply to the vagina and cervix which caused the mucosa to necrotize. These structural changes correlated with the level of exposure in the vagina. They were mainly oedema, necrosis and denudation of the vaginal walls. These alterations can make the vagina and cervix acidic which have been implicated as one of the cause of infertility in females. The results of this study suggested that treatment with F. Vogelii has an ameliorative effect.


INTRODUCTION
Toxicity is very deadly to various part of human body which adversely affects the reproductive system leading to impaired reproduction 22 . It has been reported that the female reproductive system functions in vitro!!! can be modified by exposure to a very low level of lead 1,6 . Longer and more variable menstrual cycles have been found in lead treated female Rhesus monkeys 12 . The vagina is an important organ of reproduction in the female. It is also referred to as birth canal where the foetus passes during birth. Its importance in reproduction cannot be over emphasized especially to women. The Semen is deposited in the vaginal vault during intercourse and thespermatozoa make their way into the external os of the cervical canal, pass through the cervical canal into the uterine cavity, and then continue through the uterine cavity into the uterine tubes where fertilization occurs in the ampulla 3,20 . This led us to examine the effect of lead acetate on the vaginal wall female Wistar rats. The extent of natural fertility that takes place is invariably connected to the motility of the spermatozoa and one of the things that affect sperm motility is vaginal toxicity (acidity)????. The toxicity of the vagina which can affect its histoarchiteture also causes the death of the spermatozoa and thereby impedes the fertility.
The cervix is a narrow and cylindrical part of the uterus which communicates with the uterine body via the internal os and opens into the vagina at the external os 3,21 . During the movement of the spermatozoa, it moves from the vagina into the cervix and so anything that affects the histology of the cervix likely to alter the structure of the cervical mucus which protects the sperm from naturally acidic environment of the vagina and get it safely to the ovum. It has been noted that animals have protective mechanism in the form of antioxidant nutrients 25 , vitamins and several enzymes. Antioxidants may play an important role in abating some hazardous effects of lead 6 . This study seeks to assess the lead acetate induced histological damages on vagina and cervix; and to ascertain the protective effects of aqueous extract ofF. Vogelii on female Wistar rats as an experimental model.

Collection and Authentication of Plant materials
The leaves of F. vogeliiwas collectedfrom Enyibichiri Ndufu-Alike Ikwo (wrong name) a town where Federal University Ndufu-Alike Ikwo (FUNAI) is located in Ikwo Local Government Area of Ebonyi State and authenticatedin Botany Department of the University of Nigeria Nsukka (UNN).

Preparation of the extracts
Those leaves were washed and dried in ventilated room and thereafter, were crushed into powder using pestle and mortar, and passed through mesh sieve to get the fine powders. The powdered form was soaked in water for 48 hours and filtered using a white muslin cloth to remove debrisand then re-filtered with filter paper to obtain a homogenous clear filtrate which was concentrated in vacuo, using a rotary evaporator at <40 o C to yield a sticky paste 16 . This was stored under refrigeration until it was required. All preparations were performed at the Department of Anatomy, Faculty of Basic Medical Sciences, Federal University Ndufu-Alike, Ikwo (FUNAI), Ebonyi State, Nigeria.

Animal Procurement and Housing
Twenty (20) adult female Wistar rats with average weight of 145g were procured from the animal house of the Department of Pharmacology, University of Nigeria Enugu Campus (UNEC) and maintained in the Animal House of Anatomy Department of Faculty of Basic Medical Sciences of the same University. The animals were housed in netted cages, fed with grower's mash and allowed water ad libitum with acclimatization period of one week (7 days).

Animal Grouping and Experimental Design
The rats used in this experiment were randomly assigned into five groupsas follows: A, B, C, D and E containing four (4) animals per group. The rats in this group received lead acetate (1.5mg/kg) for 7 7 days and 24 hours later exposed to aqueous extract of F. vogelii (100g/kg) till the end of experiment. The dosage was calculated based on the rats' weight. Group D: Lead acetate solution & aqueous F. vogelii extract (higher dose):The rats in this group received lead acetate (1.5mg/kg) for 7 7 days and 24 hours later exposed to aqueous extract of F. vogelii (300g/kg) till the end of experiment. The dosage was calculated based on the rats' weight. Group E: Aqueous extract of F.vogelii only: The rats in this group received 300mg/kg of aqueous extract of F.vogeliionly daily for 14days. NOTE: The extracts and lead acetate solution were administered by oral intubation using oral gavage (cannula). The experiment lasted for a period of 21 days.

Histological Study
At the end of the experiment, the rats were starved overnight and anaesthetized with chloroform and then decapitated 17 . The animals were dissected, and the vagina and cervixwere quicklyharvested and fixed in bouin's fluid for routine histological procedures. The tissue was processed and embedded in paraffin wax. Thin sections (5-6μm) were obtained and stained using haematoxylin and eosin (H&E) and were examined under light microscope to determine the histological changesthat occurred.

RESULTS
The results of the histological studies carried out in adult female Wistar rats' vagina and cervix at the end of 21 days are presented in the figures below as histological slides.

The vagina
When the vagina was microscopically examined, the negative control and extract group revealed the presence of a normal structure of vaginal mucosa and glands with no alterations (Figures 1A and 1E). Following exposure to 1.5mg/kg lead acetate (positive control), some areas of the vaginapresented optically empty spacesin the tissue, as well as diffuse oedemas and vaginal wall denudation ( Figure 1B), necrotic zones, diffused oedemas andvaginal glands necrosiswere also visibly present. After the animals were treated with the extract of Fv, there were signs of recovery revealed by the vagina. This is presented in Figures 1C and 1D below.

The cervix
The microscopic examination of the negative control and extract group revealed the presence of a normal structure of uterine cervix mucosa and glands with no alterations (Figures 2A and 2E). as the animals were exposed to 1.5mg/kg lead acetate (positive control), some areas of the cervixpresented optically empty spaces, as well as diffuse oedemas and cervical wall denudation ( Figure 2B), necrotic zones, andcervical glands necrosis were also visibly. Following the administration of the aqueous extract of F. vogelii, the cervix showed signs of recovery as presented in Figures 2C and 2D below.

Discussion
Most researches on female specimens' exposure to lead has focused on clinically visible effects such as miscarriage, premature delivery, and infant mortality in humans and animals 24 and effects of lead on the fallopian tubes, ovaries and uterus 4,5 . F. vogelii is a vegetable that is locally used to treat some illnesses and have also been implicated by so many researchers as an herbal remedy for some human diseases 7,9,10 . Works on vaginal and cervical toxicity are sparse, yet many authors have identified the toxic effects of lead on the uterus, fallopian tube and ovaries and went ahead to provethat it can hamper reproduction 14,19,24 .
In this study, we noticed that all the histology of the organs and tissues under investigation were seriously altered. This caused changes in the reproductive system integrity leading to their deformations. These changes in the vagina make it toxic and increase its acidity making the environment dangerous for the survival of sperm. The major structural changes as found in the vagina were diffuse edema, necrosis in the vagina and cervix, optical empty spaces, denudation of the vaginal and cervical mucosa, and vaginal and cervical gland necrosis as shown in the figures above. The administration of the extractappeared to neutralize the effects of lead on the organs. The fatty change that appeared in the vagina with the oedema disappeared following the administration of the extractas seen in figures 1C and 1D. The main changes in the cervix are diffuse oedema, cervical gland necrosis, optical empty spaces, denudation of the cervical mucosa, and cervical gland necrosis. There was restoration of the muscle and mucosa integrity of the cervix following the administration of the extract of F.vogelii. It can be noted that the effects of lead on reproductive systems are complex and sex-specific, and they seem to involve multiple locations on the hypothalamic-pituitary-gonadal axis, confirming our findings on female rats. Women's occupational exposure to lead is undoubtedly related to reproductive impairments 23 .Mostimpairments in the nervoussystem, the kidneys, the reproductive system and every other parts of the body as reported in other literatures show the importance and versatility oflead effects in mammals 2,8,11,13,14,15,18 .
The structural alterationsas noticed in the present study in thevagina and cervixadult rats have been able to demonstrate the poisonous effects of lead on the vagina and cervix. With the alterations of the cervical walls, the secretion of cervical mucus that helps the sperm's move towards the egg will be hampered seriously this in turn leads to infertility in female.With the results ofthis research, we wish to recommend the use of this leave Ficus vogelii as vegetable for families and its inclusion as an herbal medicine for the treatment of reproductive toxicity especially that of lead toxicity in the female laboratory animals.