Genetic variants of ALDH2-rs671 and CYP2E1-rs2031920 contributed to risk of hepatocellular carcinoma susceptibility in a Chinese population

Figures & data

Table 1 Baseline characteristics of the HCC and control groups

Factors	HCC (%) (n=300)	Control (%) (n=292)	OR (95% CI)	P-value
Sex			1.11 (0.67–1.86)	0.685
Male	265 (88.3)	261 (87.0)
Female	35 (11.7)	31 (27.0)
Age (years)			1.62 (0.93–2.84)	0.091
≤60	265 (88.3)	270 (92.5)
>60	35 (11.7)	22 (7.5)
Race			0.44 (0.30–0.64)	<0.001
Han	245 (81.7)	193 (64.3)
Minority	55 (18.3)	99 (35.7)
Drinking status			1.03 (0.72–1.47)	0.875
Yes	214 (71.3)	210 (71.9)
No	86 (28.7)	82 (28.1)
Smoking status			0.92 (0.67–1.27)	0.623
Yes	154 (51.3)	144 (49.3)
No	146 (48.7)	148 (50.7)
Tumor family history			0.60 (0.34–1.04)	0.070
Yes	36 (11.6)	22 (7.5)
No	264 (88.4)	270 (92.5)
HBsAg status			0.02 (0.01–0.03)	<0.001
+	255 (85.0)	30 (10.3)
−	45 (15.0)	262 (89.7)

Abbreviations: HCC, hepatocellular carcinoma; OR, odds ratio.

Table 2 Joint-effects analysis between ALDH2 or CYP2E1 and drinking status on HCC susceptibility

Groups	Genotype		Whole		Drinking		Non-drinking		Crude OR (95% CI)	Crude P-value	Adjusted OR (95% CI)	Adjusted P-value*
	ALDH2	CYP2E1	HCC	Control	HCC	Control	HCC	Control
	G/G		149	152					1.00	0.469	1.00	0.999
	G/A		121	119					1.04 (0.74–1.46)	0.833	0.99 (0.59–1.65)	0.967
	A/A		30	21					1.46 (0.80–2.66)	0.220	1.00 (0.40–2.48)	0.996
	G/A+A/A		151	140					1.10 (0.80–1.52)	0.561	0.99 (0.61–1.62)	0.972
		C1/C1	203	196					1.00	0.546	1.00	0.381
		C1/C2	87	81					1.04 (0.72–1.49)	0.843	0.67 (0.38–1.19)	0.171
		C2/C2	10	15					0.64 (0.28–1.47)	0.295	0.77 (0.22–2.70)	0.684
		C1/C2+C2/C2	97	96					0.98 (0.69–1.38)	0.888	0.69 (0.40–1.18)	0.170
i	G/G	C1/C1	104	96					1.00	0.397	1.00	0.160
ii	G/G	C1/C2+C2/C2	45	56					0.74 (0.46–1.20)	0.223	0.42 (0.20–0.90)	0.025
iii	G/A+A/A	C1/C1	99	100					0.91 (0.62–1.35)	0.653	0.72 (0.40–1.31)	0.287
iiii	G/A+A/A	C1/C2+C2/C2	52	40					1.20 (0.73–1.97)	0.472	0.83 (0.39–1.77)	0.626
1	G/G	C1/C1			70	79			1.00	0.067	1.00	0.137
2	G/G	C1/C2+C2/C2			32	45			0.80 (0.46–1.40)	0.438	0.45 (0.19–1.06)	0.068
3	G/A+A/A	C1/C1			74	63			1.33 (0.83–2.11)	0.235	1.13 (0.56–2.26)	0.740
4	G/A+A/A	C1/C2+C2/C2			38	23			1.87 (1.01–3.43)	0.045	1.33 (0.54–3.28)	0.538
!	G/G	C1/C1					34	17	1.00	0.043	1.00	0.053
! !	G/G	C1/C2+C2/C2					13	11	0.59 (0.22–1.59)	0.299	0.35 (0.07–1.72)	0.197
! ! !	G/A+A/A	C1/C1					25	37	0.34 (0.16–0.73)	0.006	0.20 (0.06–0.68)	0.010
! ! ! !	G/A+A/A	C1/C2+C2/C2					14	17	0.41 (0.17–1.03)	0.058	0.20 (0.04–0.96)	0.045

Note:

* P adjustment for race and HBsAg.

Abbreviations: HCC, hepatocellular carcinoma; OR, odds ratio; CI, confidence interval.

Table 3 Joint-effects analysis between ALDH2 or CYP2E1 and drinking status on HCC susceptibility

Groups	Drinking status	Genotype	HCC (%) (n=300)	Control (%) (n=292)	Crude OR (95% CI)	Crude P-value	Adjusted OR (95% CI)	Adjusted P-value*
1	−	G/G	47	28	1.00	0.006	1.00	0.022
2	−	G/A+A/A	39	54	0.43 (0.23–0.80)	0.008	0.32 (0.13–0.79)	0.014
3	+/++	G/G	102	124	0.49 (0.29–0.84)	0.009	0.32 (0.15–0.69)	0.004
4	+/++	G/A+A/A	112	86	0.78 (0.45–1.34)	0.362	0.49 (0.23–1.07)	0.075
I	+	G/G	55	77	1.00	0.004	1.00	0.360
II	+	G/A+A/A	69	68	1.42 (0.88–2.30)	0.153	1.75 (0.85–3.58)	0.127
III	++	G/G	47	47	1.40 (0.82–2.38)	0.215	1.53 (0.68–3.42)	0.306
IV	++	G/A+A/A	43	18	3.34 (1.75–6.41)	<0.001	2.00 (0.78–5.12)	0.148
a	−	C1/C1	59	54	1.00	0.982	1.00	0.338
b	−	C1/C2+C2/C2	27	28	0.88 (0.46–1.68)	0.704	0.69 (0.25–1.85)	0.456
c	+/++	C1/C1	144	142	0.93 (0.60–1.44)	0.737	0.71 (0.37–1.37)	0.307
d	+/++	C1/C2+C2/C2	70	68	0.94 (0.57–1.55)	0.815	0.49 (0.23–1.05)	0.067
A	+	C1/C1	88	98	1.00	0.094	1.00	0.534
B	+	C1/C2+C2/C2	36	47	0.85 (0.51–1.44)	0.550	0.61 (0.28–1.34)	0.215
C	++	C1/C1	56	44	1.42 (0.87–2.31)	0.162	1.14 (0.55–2.37)	0.728
D	++	C1/C2+C2/C2	34	21	1.80 (0.97–3.34)	0.060	1.00 (0.39–2.57)	0.995

Notes:

* P adjustment for race and HBsAg. −, Not drinking; +, drinking little; ++, drinking a lot.

Abbreviations: HCC, hepatocellular carcinoma; OR, odds ratio; CI, confidence interval.

Figure 1 Lymphocytes and ALDH2/CYP2E1 mRNA expression profiles. (A) Lymphocytes stained by the H&E method. (B) Curve of lymphocyte survival rate–culture time. (C) Curve of lymphocyte inhibitory rate–ethanol concentration. (D) Curves of lymphocyte inhibitory rate–culture time at different ethanol concentrations. (E and F) ALDH2, CYP2E1, and GAPDH mRNA expression levels at different ethanol concentrations.

Figure 2 mRNA expression levels of ALDH2 and CYP2E1 in different groups. (A and B) ALDH2 mRNA expression levels in HCC and control groups. (C and D) CYP2E1 mRNA expression levels in HCC and control groups. (E and F) ALDH2 mRNA expression levels of the GG and GA+AA genotypes. (G and H) CYP2E1 mRNA expression levels of the C1/C1 and C1/C2+C2/C2 genotypes.

Note: * P<0.05; ** P<0.01; ^# P>0.05.

Abbreviation: HCC, hepatocellular carcinoma.

Table 4 Changes in ALDH2 and CYP2E1 mRNA expression levels after ethanol exposures

Gene	Group	Genotype	Subject (n=30)	Ethanol concentration (mmol/L)
				0	50	100	200
ALDH2	HCC		20	0.37±0.20	0.55±0.24*	0.44±0.28	0.35±0.15
	Control		10	0.34±0.14	0.51±0.21	0.35±0.28	0.26±0.12
CYP2E1	HCC		20	0.09±0.03	0.14±0.08*	0.09±0.08	0.05±0.04
	Control		10	0.04±0.01	0.055±0.02**	0.02±0.01**	0.01±0.00**
ALDH2		G/G	18	0.39±0.19	0.57±0.19**	0.48±0.29	0.33±0.15
		G/A+A/A	12	0.12±0.11	0.49±0.29**	0.36±0.17**	0.26±0.13*
CYP2E1		C1/C1	20	0.03±0.02	0.04±0.04	0.03±0.03	0.03±0.02
		C1/C2+C2/C2	10	0.11±0.07	0.24±0.16*	0.15±0.08	0.13±0.08

Note:

* P<0.05,

** P<0.01; comparisons were done between before ethanol intervention (0 mmol/L) and different ethanol intervention concentrations (50, 100, 200 mmol/L); data are presented as the mean±standard deviation.

Abbreviation: HCC, hepatocellular carcinoma.

Figure 3 Expression levels of ALDH2 and CYP2E1 in different tissues. (A) ALDH2 expression levels in different tissues. (B) CYP2E1 expression levels in different tissues.

Figure 4 Gene expression levels and protein interactions. (A and B) ALDH2 and CYP2E1 gene expression levels in normal and primary tumor tissues. (C) Interactions of ALDH2 and CYP2E1 with other genes. (D) Interactions of ALDH2 and CYP2E1 with other proteins.

Figure 4 Gene expression levels and protein interactions. (A and B) ALDH2 and CYP2E1 gene expression levels in normal and primary tumor tissues. (C) Interactions of ALDH2 and CYP2E1 with other genes. (D) Interactions of ALDH2 and CYP2E1 with other proteins.

Table 5 Functional information of ALDH2 rs671 and CYP2E1 rs2031920 genetic polymorphisms

Gene site	FS score	Functional category	Prediction tool	Prediction result
ALDH2	0.856	Protein coding	PolyPhen	Probably damaging
rs671			SIFT	Damaging
			SNP effect	Benign
			LS-SNP	Benign
			SNPs3D	Deleterious
			Ensembl-NS	Nonsynonymous
		Splicing regulation	ESEfinder	Changed
			ESRSearchi	Changed
			PESX	Changed
			RESCUE_ESE	Changed
		Post translation	OGPET	Not exist
			Sulfinator	Not exist
CYP2E1	0.398	Transcriptional regulation	Ensembl-NS	Frameshift coding
			GoldenPath	Not exist
			Ensembl-TR	Regulatory region

Abbreviation: FS, functional significance.