Chemopreventive efficacy of geraniol against 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

Figures & data

Figure 1. Chemical structure of geraniol.

Figure 2. (A) Gross appearance of oral squamous cell carcinoma (indicated by arrow) in DMBA-alone-painted hamsters. (B–F) Histopathological changes in the buccal mucosa of control and experimental hamsters in each group (hematoxylin and eosin, ×400). (B) Well-differentiated squamous cell carcinoma exhibiting epithelial and keratin pearls in DMBA-alone-treated group 1 hamsters. (C) Buccal pouch epithelium from group 2 hamsters (DMBA + geraniol) exhibiting moderate keratosis and hyperplasia. (D–F) Buccal pouch epithelium from group 3 (geraniol alone 250 mg/kg b.w.), group 4 (liquid paraffin + corn oil), and group 5 (untreated control) hamsters showed normal cellular architecture with no signs of cell proliferation.

Table 1. Tumor incidence, tumor number, tumor volume, and tumor burden in control and experimental hamsters

Groups	Treatment	Tumor incidence (oral squamous cell carcinoma)	Total number of tumors/hamsters	Tumor volume (mm^3)	Tumor burden (mm^3)
1.	DMBA	100%	28/(10)	364.50 ± 42.40	1020.60 ± 98.50
2.	DMBA + geraniol (125 mg/kg b.w.)	50%	12/(5)	74.15 ± 5.60	177.96 ± 13.60
3.	DMBA + geraniol (250 mg/kg b.w.)	0%	0	0	0
4.	DMBA + geraniol (500 mg/kg b.w.)	0%	0	0	0

Values are expressed as mean ± SD for 10 hamsters in each group. Tumor volume was measured using the formula (where D₁, D₂, and D₃ are the three diameters (mm³) of the tumor). Tumor burden was calculated by multiplying tumor volume and the number of tumors/animal.

Table 2. Histopathological changes in the buccal mucosa of control and experimental hamsters (n = 10/group)

Group	Treatment	Keratosis	Hyperplasia	Dysplasia	Squamous cell carcinoma
1.	DMBA	++ +	++ +	++ +	++ +
2.	DMBA + geraniol (250 mg/kg b.w.)	++	++	+	−
3.	Geraniol alone (250 mg/kg b.w.)	−	−	−	−
4.	Corn oil + liquid paraffin	−	−	−	−
5.	Control	−	−	−	−

−, no change; +, mild; ++, moderate; ++ +, severe.

Table 3. The levels of LPO by-products (TBARS, LOOH, and CD) in the plasma and buccal mucosa of control and experimental hamsters

		Plasma			Buccal mucosa
Groups	Treatment/parameters	TBARS (nmol/ml)	LOOH (mM/ml)	CD (μM/ml)	TBARS (nmol/100 mg protein)	LOOH (mmol/mg protein)	CD (mg/mg protein)
1.	DMBA	4.96 ± 0.49^a	0.92 ± 0.06^a	2.02 ± 0.16^a	49.72 ± 4.93^a	24.02 ± 2.12^a	21.62 ± 2.02^a
2.	DMBA + geraniol (250 mg/kg b.w.)	3.07 ± 0.28^b	0.52 ± 0.06^b	1.12 ± 0.18^b	70.68 ± 5.26^b	29.02 ± 2.88^b	26.18 ± 2.20^b
3.	Geraniol alone (250 mg/kg b.w.)	2.68 ± 0.20^c	0.34 ± 0.03^c	0.80 ± 0.07^c	79.86 ± 4.37^c	32.24 ± 3.02^c	29.02 ± 1.38^c
4.	Corn oil + liquid paraffin	2.64 ± 0.21^c	0.36 ± 0.03^c	0.82 ± 0.08^c	79.75 ± 6.66^c	31.42 ± 2.98^c	28.76 ± 1.46^c
5.	Control	2.74 ± 0.23^c	0.34 ± 0.02^c	0.78 ± 0.08^c	78.38 ± 5.69^c	32.12 ± 2.92^c	30.22 ± 1.56^c

Values are expressed as mean ± SD for 10 hamsters in each group.

^a–cValues not sharing a common superscript letter (a, b, and c) differ significantly at P < 0.05 (DMRT).

Figure 3. The status of enzymatic antioxidants (GPx, SOD, and CAT) and non-enzymatic antioxidants (vitamin E and GSH) in the plasma of control and experimental hamsters. (1) DMBA, (2) DMBA + geraniol (250 mg/kg b.w.), (3) Geraniol alone (250 mg/kg b.w.), (4) Corn oil + liquid paraffin, (5) Control. Bars are mean ± SD for 10 hamsters in each group. ^a–cValues that do not share a common superscript letter (a, b, and c) in the same column differ significantly at P < 0.05 (DMRT). U^A – micromoles of glutathione utilized/minute; U^B – the amount of enzymes required to inhibit 50% nitroblue-tetrazolium (NBT) reduction; U^C – micromoles of H₂O₂ utilized/second.

Figure 4. The status of enzymatic antioxidants (GPx, SOD, and CAT) and non-enzymatic antioxidants (vitamin E and GSH) in the buccal mucosa of control and experimental hamsters. (1) DMBA, (2) DMBA + geraniol (250 mg/kg b.w.), (3) Geraniol alone (250 mg/kg b.w.), (4) Corn oil + liquid paraffin, (5) Control. Bars are mean ± SD for 10 hamsters in each group. ^a–cValues that do not share a common superscript letter (a, b, and c) in the same column differ significantly at P < 0.05 (DMRT). U^A – micromoles of glutathione utilized/minute; U^B – the amount of enzymes required to inhibit 50% nitroblue-tetrazolium (NBT) reduction; U^C – micromoles of H₂O₂ utilized/second.

Table 4. Activities of phase I detoxification enzymes in the liver of control and experimental hamsters

		Liver		Buccal mucosa
Groups	Treatment/parameters	Cytochrome P450 (U^X /mg protein)	Cytochrome b5 (U^Y/mg protein)	Cytochrome P450 (U^X /mg protein)	Cytochrome b5 (U^Y/mg protein)
1.	DMBA	1.70 ± 0.12^a	1.98 ± 0.18^a	3.50 ± 0.28^a	0.76 ± 0.06^a
2.	DMBA + geraniol (250 mg/kg b.w.)	0.76 ± 0.06^b	1.08 ± 0.16^b	2.00 ± 0.20^b	0.42 ± 0.04^b
3.	Geraniol alone (250 mg/kg b.w.)	0.69 ± 0.04^c	0.98 ± 0.08^c	1.84 ± 0.16^c	0.40 ± 0.04^c
4.	Corn oil + liquid paraffin control	0.66 ± 0.04^c	0.92 ± 0.06^c	1.80 ± 0.14^c	0.38 ± 0.03^c
5.	Control	0.68 ± 0.04^c	0.94 ± 0.08^c	1.78 ± 0.14^c	0.34 ± 0.02^c

Values are expressed as mean ± SD for 10 hamsters in each group.

^a–cValues not sharing a common superscript letter (a, b, and c) differ significantly at P < 0.05 (DMRT).

X – micromoles of cytochrome P450.

Y – micromoles of cytochrome b5.

Figure 5. The activities of phase II detoxification enzymes (GST, GR, and DTD) and GSH in the liver of control and experimental hamsters. (1) DMBA, (2) DMBA + geraniol (250 mg/kg b.w.), (3) Geraniol alone (250 mg/kg b.w.), (4) Corn oil + liquid paraffin, (5) Control. Bars are mean ± SD for 10 hamsters in each group. ^a–cValues not sharing a common superscript letter (a, b, and c) in the same column differ significantly at P < 0.05 (DMRT). U^A – micromoles of 1-chloro-2,4-dinitrobenzene-reduced glutathione conjugate formed/minute. U^B – micromoles of NADPH oxidized/hour. U^C – micromoles of 2,6-dichlorolindophenol reduced/minute.

Figure 6. The status of phase II detoxification enzymes (GST and GR) and glutathione content (GSH, GSSG, and GSH/GSSG ratio) in the buccal mucosa of control and experimental hamsters. (1) DMBA, (2) DMBA + geraniol (250 mg/kg b.w.), (3) Geraniol alone (250 mg/kg b.w.), (4) Corn oil + liquid paraffin, (5) Control. Bars are mean ± SD for 10 hamsters in each group. ^a–cValues not sharing a common superscript letter (a, b, and c) in the same column differ significantly at P < 0.05 (DMRT). U^A – micromoles of 1-chloro-2,4-dinitrobenzene-reduced glutathione conjugate formed/minute. U^B – micromoles of NADPH oxidized/hour.