Brief Articles
Copyright ©2009 The WJG Press and Baishideng.
World J Gastroenterol. Jun 7, 2009; 15(21): 2595-2601
Published online Jun 7, 2009. doi: 10.3748/wjg.15.2595
Table 1 Methylation-specific PCR, real-time PCR primer, probe sequence and annealing temperature
GenePrimer purposePrimer sequence (5'--3')Annealing temperature (°C)Product size (bp)
DKK-3MSP-MF: GGGGCGGGCGGCGGGGC58120
R: ACATCTCCGCTCTACGCCCG
MSP-UF: TTAGGGGTGGGTGGTGGGGT56126
R: CTACATCTCCACTCTACACCCA
Real time RT-PCRF: GTAAGTTTCCCCTCTGGCTTG6090
R: AAGCACCAGACTGTGAAGCCT
Probe: FAM+AGGTGTTGTGCATTTGTTCAGCTCCC+TAMRA
WIF-1MSP-MF: CGTTTTATTGGGCGTATCGT57145
R: ACTAACGCGAACGAAATACGA
MSP-UF: GGGTGTTTTATTGGGTGTATTGT52154
R: AAAAAAACTAACACAAACAAAATACAAAC
Real time RT-PCRF: TCCAAACACCTCAAAATGCTATC60119
R: GAACCCATCAGGACACTCGC
Probe: FAM+ACAAGCTGAGTGCCCAGGCGG+TAMRA
GAPDHReal time RT-PCRF: CCACTCCTCCACCTTTGAC60102
R: ACCCTGTTGCTGTAGCCA
Probe: FAM+TTGCCCTCAACGACCACTTTGTC+TAMRA
Table 2 Methylation level of DKK-3 and WIF-1 in two tissues (mean ± SE, %)
Methylation levelDKK-3WIF-1
HCC tissues37.61 ± 4.2629.70 ± 3.24
Adjacent non-cancerous tissues15.96 ± 3.9118.34 ± 4.02
P < 0.05P < 0.05
Table 3 Methylation and clinical data of DKK-3 genes in HCC and adjacent non-cancerous tissues
Clinical dataMethylation in HCC
Methylation in adjacent non-cancerous tissues
NegativePositivePNegativePositiveP
Sex
Male12151.0002161.000
Female3351
Age (yr)
< 601370.0111730.393
≥ 6021194
Child
A12170.3082361.000
B3131
AFP (ng/mL)
< 207100.3081530.674
≥ 2088114
Number of tumor
Single11131.0002040.358
Multiple4563
Tumor diameter (cm)
< 3231.000321.000
≥ 31315235
HBsAg
(+)15140.1082361.000
(-)0431
Hepatocirrhosis
(+)15100.0042051.000
(-)0862
False capsule
(+)14140.3462350.282
(-)1432
Tumor thrombus
(+)121.000301.000
(-)1416237
Edmonson stage
I/II13151.0002261.000
III2341
Table 4 Methylation and clinical data of WIF-1 genes in HCC and adjacent non-cancerous tissues
Clinical dataMethylation in HCC
Methylation in adjacent non-cancerous tissues
NegativePositivePNegativePositiveP
Sex
Male17100.6592341.000
Female3351
Age (yr)
< 60751.0001731.000
≥ 60138112
Child
A17121.0002451.000
B3140
AFP (ng/mL)
< 201181.0001631.000
≥ 2095122
Number of tumor
Single14101.0002130.597
Multiple6372
Tumor diameter (cm)
< 3231.000500.569
≥ 31810235
HBsAg
(+)2090.0172451.000
(-)0440
Hepatocirrhosis
(+)1870.0352141.000
(-)2671
False capsule
(+)18100.3602441.000
(-)2341
Tumor thrombus
(+)120.547210.400
(-)1911264
Edmonson stage
I/II16120.6252350.569
III4150
Table 5 mRNA expression and clinical data of DKK-3 in HCC and adjacent non-cancerous tissues (mean ± SE)
Clinical dataCaseThe mRNA expression of DKK-3 in HCCtPThe mRNA expression of DKK-3 in adjacent non-cancerous tissuestP
Sex
Male270.776 ± 0.3460.1530.8800.779 ± 0.3190.8880.404
Female60.761 ± 0.1680.913 ± 0.338
Age (yr)
< 60200.756 ± 0.3560.3340.7410.819 ± 0.3410.1950.847
≥ 60130.795 ± 0.2850.840 ± 0.274
Child
A290.761 ± 0.3310.7890.4660.829 ± 0.3290.1710.869
B40.863 ± 0.2250.813 ± 0.155
AFP (ng/mL)
< 20170.716 ± 0.2871.0690.2940.840 ± 0.3540.1240.902
≥ 20160.835 ± 0.3840.826 ± 0.281
Number of tumor
Single240.740 ± 0.3150.9710.3480.823 ± 0.2980.2620.797
Multiple90.860 ± 0.3290.860 ± 0.377
Tumor diameter (cm)
< 340.908 ± 0.3000.9430.3990.792 ± 0.3520.2900.780
≥ 3290.755 ± 0.3220.841 ± 0.315
HBsAg
(+)290.660 ± 0.2330.9840.3720.817 ± 0.3040.6390.563
(-)40.789 ± 0.3290.955 ± 0.418
Hepatocirrhosis
(+)250.780 ± 0.3190.0460.9640.761 ± 0.2682.1570.057
(-)80.774 ± 0.3651.060 ± 0.361
False capsule
(+)280.757 ± 0.3261.7500.1320.819 ± 0.3070.7900.466
(-)51.014 ± 0.2990.966 ± 0.395
Tumor thrombus
(+)30.770 ± 0.3300.7380.5190.811 ± 0.3071.0880.379
(-)300.890 ± 0.2601.057 ± 0.379
Edmonson stage
I/II280.807 ± 0.3353.3800.0200.820 ± 0.3170.5640.596
III50.586 ± 0.3500.910 ± 0.333
Table 6 mRNA expression and clinical data of WIF-1 in HCC and adjacent non-cancerous tissues (mean ± SE)
Clinical dataCaseThe mRNA expression of WIF-1 in HCCtPThe mRNA expression of WIF-1 in adjacent non-cancerous tissuestP
Sex
Male270.882 ± 0.5420.9420.3650.828 ± 0.3200.2860.783
Female60.720 ± 0.3360.787 ± 0.318
Age (yr)
< 60120.767 ± 0.3971.0920.2890.793 ± 0.2470.5510.589
≥ 60210.985 ± 0.6430.862 ± 0.407
Child
A290.854 ± 0.5430.0670.9480.793 ± 0.3091.2430.286
B40.845 ± 0.1711.015 ± 0.337
AFP (ng/mL)
< 20190.873 ± 0.5160.2310.8190.828 ± 0.3770.1480.883
≥ 20140.831 ± 0.5190.812 ± 0.247
Number of tumor
Single240.927 ± 0.5321.2920.2120.801 ± 0.2880.4960.630
Multiple90.704 ± 0.4080.872 ± 0.393
Tumor diameter (cm)
< 351.002 ± 0.7500.5070.6360.796 ± 0.4150.1470.889
≥ 3280.826 ± 0.4690.825 ± 0.304
HBsAg
(+)290.875 ± 0.5270.1550.8860.838 ± 0.3120.7540.497
(-)40.820 ± 0.6810.695 ± 0.360
Hepatocirrhosis
(+)250.840 ± 0.5230.2600.7970.810 ± 0.3220.3310.746
(-)80.893 ± 0.4860.853 ± 0.314
False capsule
(+)280.860 ± 0.4900.1520.8860.816 ± 0.2960.1440.892
(-)50.812 ± 0.6760.846 ± 0.454
Tumor thrombus
(+)30.815 ± 0.5051.3920.2760.806 ± 0.3190.8400.474
(-)301.223 ± 0.4810.960 ± 0.300
Edmonson stage
I/II280.910 ± 0.5171.0050.0830.821 ± 0.3080.1410.894
III50.532 ± 0.3600.848 ± 0.401