Protective Effects of Garlic and Cinnamon Oils on Hepatocellular Carcinoma in Albino Rats

Abstract

Natural oils are traditional medicinal herbs, which have attracted interests for its potential anti-inflammatory and anticancer activities. The present work is aimed at evaluating the protective effect of garlic oil and cinnamon oil on diethylnitrosamine- (DENA-) and 2-acetylaminofluorene- (2-AAF-) induced p53 gene mutation and hepatocarcinogenesis in rats. Forty male albino rats were divided into 4 equal groups: control, hepatocellular carcinoma (HCC), garlic oil-HCC, and cinnamon oil-HCC. The HCC-induced group showed a significant decrease in the body mass and a significant elevation in the liver weight, alpha-fetoprotein (AFP), liver enzymes, hepatic malondialdehyde (MDA), and p53 protein expression levels as well as genetic mutations in intron 5 of p53 gene in the form of Single-Nucleotide Polymorphisms (SNPs) and insertions. In addition, the glutathione (GSH) level and superoxide dismutase (SOD) activities were increased. While HCC rats pretreated with garlic oil or cinnamon oil were significantly reversed, these destructive actions increased GSH and SOD levels. The HCC-induced group showed histopathological features of liver cancer including hypercellularity, nuclear hyperchromasia, mitotic figures, and preneoplastic foci. On the other hand, HCC rats pretreated with garlic oil or cinnamon oil revealed partial reversal of normal liver architecture. The present findings proposed that these natural oils have the ability to improve liver function, significantly reduced the liver toxicity and HCC development. However, further sophisticated studies are recommended before their use as conventional therapeutics for HCC treatment.

Figure 1

Body weight of rats in different experimental groups throughout the weeks of experiment.

Figure 2

Tumor marker alpha fetoprotein (AFP) levels in serum of experimented rats.

Figure 3

A gross liver of experimental groups, control rats showing normal liver, HCC-induced rats showing that grayish-white pin point foci with variable sizes gave the external surface of the liver a rough, nodular, and irregular appearance, garlic oil-HCC rats showing nearly normal liver size and morphology, and cinnamon oil-HCC rats showing nearly normal liver size and morphology.

Figure 4

Liver tissue in each group of rats (H&E staining, 200), control rats showing a normal hexagonal lobular pattern of hepatic tissue, HCC-induced rats showing that delicate fibrous connective tissue bands cause distortion of the normal architectural pattern of the hepatic parenchyma, garlic oil-HCC rats showing nearly preserved architecture, and cinnamon oil-HCC rats showing partial reversal of normal liver architecture.

Figure 5

p53 expression levels (IHC) of different studied experimental groups at 3 and 5 months of the experiment.

Figure 6

PCR amplification results of p53 gene exons 5, 6, and 7 in the control group. Lane 1: 100 bp DNA ladder. Lane 2: 265 bp PCR amplicon representing a part of intron 4, exon 5, and part of intron 5 of rat p53 gene. Lane 3: 273 bp PCR amplicon representing a part of intron 5, exon 6, and part of intron 6 of rat p53 gene. Lane 4: 188 bp PCR amplicon representing a part of intron 6, exon 7, and part of intron 7 of rat p53 gene.

Figure 7

PCR amplification results of p53 gene exons 5, 6, and 7 in the HCC-induced group. Lane 1: 100 bp DNA ladder. Lane 2: 265 bp PCR amplicon representing a part of intron 4, exon 5, and part of intron 5 of rat p53 gene. Lane 3: 275 bp PCR amplicon representing a part of intron 5, exon 6, and part of intron 6 of rat p53 gene. Lane 4: 188 bp PCR amplicon representing a part of intron 6, exon 7, and part of intron 7 of rat p53 gene.

Figure 8

Nucleotide sequence of the amplified PCR product (265 bp) representing exon 5 of rat p53 gene in the control group.

Figure 9

Nucleotide sequence of the amplified PCR product (265 bp) representing exon 5 of rat p53 gene in the HCC-induced group.

Figure 10

Nucleotide sequence of the amplified PCR product (273 bp) representing exon 6 of rat p53 gene in the control group.

Figure 11

Nucleotide sequence of the amplified PCR product (275 bp) representing exon 6 of rat p53 gene in the HCC-induced group.

Figure 12

Nucleotide sequence of the amplified PCR product (188 bp) representing exon 7 of rat p53 gene in the control group.

Figure 13

Nucleotide sequence of the amplified PCR product (188 bp) representing exon 7 of rat p53 gene in the HCC-induced group.

Figure 14

Direct sequencing of the amplified PCR product (265 bp) representing exon 5 of rat p53 gene in both control (a) and HCC-induced (b) groups showing an A to C transversion.

Figure 15

Nucleotide sequence alignment of the amplified PCR product (265 bp) representing exon 5 of rat p53 gene in both control and HCC-induced groups showing an A to C transversion at position 1289 of intron 5.

Figure 16

Amino acid sequence alignment of rat p53 gene, exon 5 in both control and HCC-induced groups showing complete homogeneity.

Figure 17

Direct sequencing of the amplified PCR product representing exon 6 of rat p53 gene in both control (a) and HCC-induced (b) groups showing two insertion mutations of the nucleotides T and C, three T to C transitions, and one C to T transition.

Figure 18

Nucleotide sequence alignment of the amplified PCR product representing exon 6 of rat p53 gene in both control and cancer-induced groups showing two insertion mutations of the nucleotides T and C at position numbers 1331 and 1338 of intron 5, respectively, three T to C transitions at positions 1332 (T 1332 C), 1336 (T 1336 C), and 1342 (T 1342 C), respectively, of intron 5, and one C to T transition at position 1341 (C 1341 T) of intron 5.

Figure 19

Amino acid sequence alignment of rat p53 gene, exon 6 in both control and cancer-induced groups showing complete homogeneity.

Figure 20

Nucleotide sequence alignment of the amplified PCR product (188 bp) representing exon 7 of rat p53 gene in both control and cancer-induced groups showing complete homogeneity.

Figure 21

Amino acid sequence alignment of rat p53 gene, exon 7 in both control and cancer-induced groups showing complete homogeneity.