The effects of 5% A. indica aqueous extract (AI), or more commonly as Neem, on
hepatocarcinogenesis induced Spraque-Dawley male rats were investigated.
Hepatocarcinogenesis was induced in rats by employing a two carcinogen
system: an intraperitoneal injection of 200 mg/kg diethyl nitrosamine (DEN) as
initiator; followed by 0.02% of 2-acetylaminofluorene (AAF) in rat chow for two
weeks to promote carcinogenesis. The rats were then left for two weeks to allow
hepatic preneoplastic lesions to occur. The plant extract was prepared in 5% w/v
in distilled water. Fresh leaves were collected, blended and mixed with distilled
water. Twenty male rats Spraque-Dawley weighmg 150-250g were acclimatized
for 1 week before use. The rats were divided into four groups of five rats each.
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The groups were: DENIAAF-iduced rats (C), DENIAAF-iduced rats treated with
5% A. indica (CAI), normal control rats (N) and normal rats treated with 5% A.
indica extract group (NAI). The rats in group N and NAI were not induced with
cancer however were injected once intraperitoneally with corn oil and act as
normal control. The plant extract was fed to CAI and NAI groups to study its
effects on both cancer and normal groups, respectively.
In this study several parameters were evaluated as means of determining the
effects of A1 on DENIAAF-induced hepatocarcinogenesis in rats. Body and liver
weight profiles, foremost, hepatic lesions were scored in rats induced with
DENIAAF cacinogens especially in the portal and lobular regions of the liver
sections examined for histology. Loss of normal cell organization was also
observed once the hepatocarcinogenesis was induced. In addition to histological
observations, the TUNEL Assay, liver antioxidant enzyme Glutathione S
transferase (GS-T), Glutathione Peroxidase (GPx) in the serum and liver, tumor
marker alpha-fetoprotein (AFP) in serum and molecular detection of AFP and
albumin genes expressions were conducted. The observation of the lesion
scoring have shown significant difference (p<0.05) between DENIAAF and
normal control groups (N, NAI). Histologically there were sigruficant changes in
the lesion scoring of the liver in portal and lobular region in DENIAAF induced
group (C) compared to the DENJAAF treated with A.indica (CAI). TUNEL assay
supported that there was more numbers of apoptotic cells in the liver of (CAI)
group compared to(C) group. The liver enzymes (GST & GPx) activity was
measured and the result for both glutathione Stransferase and glutathione
peroxidase were sigruficantly (p<0.05) higher in the (C) compared to the other
groups (CAI, N, NAI). This result revealed that A. indica extract could reduce the
activity of liver and serum GPx and GST enzymes of rats during
hepatocarcinogenesis. However, the results of body and liver weight profiles
showed that the CAI group was not significantly different (pN.05) from N, C
and NAI groups.
Alpha fetoprotein (AFP), a notable liver tumor marker, level was measured. The
DENIAAF induced group (C) showed the highest increase in AFP levels while
in CAI group illustrated significant (pc0.05) decrease in AFP level. There was no
significant (pM.05) difference between N, NAI and CAI group.
Mokdar detection of gene expression was done by RT-PCR for a-fetoprotein
and albumin specific genes. However, the expression of the AFP gene was
observed only in DEN/AAF induced group (C). Albumin gene expression was
observed in all the study groups C, N, NAI and CAI proving the hepatic nature
of the studied tissue and used as a housekeeping control gene in the RT-PCR
experiments.
As a conclusion, A. indica (Neem) has revealed a chemopreventive capability by
regressing the hepatacarcinogenesis induced by DENJAAF carcinogens. This
capability can be seen from the modulating effects of the plant in the biological
indicators used in this study which can encourage the researchers to consider
the A. indica (Neem) for further on mechanism and toxicology study
