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Promoter methylation analysis of DKK2 may be a potential biomarker for early detection of cervical cancer Cover

Promoter methylation analysis of DKK2 may be a potential biomarker for early detection of cervical cancer

Asian Biomedicine
Volume 16 (2022): Issue 4 (August 2022)
By: Xian Zhang,  Aihua Li,  Jie Wu,  Yu Wu,  Xiaoping Ma,  Yanjun Liu,  Qingfa Chen and  Yan Zhang  
Open Access
|Aug 2022

Figures & Tables

Figure 1

DKK2 mRNA expression in cervical cells and tissues. (A–E) RT-PCR analysis of DKK2 mRNA expression in cervical normal and cancer tissues. **P < 0.01 compared with normal tissues. (F) RT-PCR analysis of DKK2 mRNA expression in normal cervical epithelial cell lines and in 9 human cervical cancer cell lines. *P < 0.05 compared with normal cervical epithelial cell lines. (G) RT-PCR analysis of DKK2 mRNA relative expression in normal cervical epithelial cell lines and 4 human cervical cancer cell lines (Hela, Caski, HT-3, and C33A cell lines). **P < 0.01 compared with normal cervical epithelial cell lines. (H) DKK2 mRNA expression in LSIL, HSIL, tumor, and normal cervical tissues. The PCR products were separated in 2% agarose gels using a DL1000 marker set (catalog No. 3591A; Takara) marker set; the band of interest is between markers 100 bp and 200 bp at 169 bp. DKK2, Dickkopf 2; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; H, HSIL; L, LSIL; N, normal; RT-PCR, reverse transcription-polymerase chain reaction; T, tumor; TCGA, The Cancer Genome Atlas; bp, base pairs

Figure 2

DKK2 methylation level in cervical cancer cell lines and tissues. (A) MSP analysis of DKK2 hypermethylation in Hela, Caski, HT-3, and C33A cervical cancer cell lines. (B) DKK2 expression after treatment with 5-Aza-dC for 4 d in cervical cancer cell lines. GAPDH was used as a housekeeping control gene. (C) DKK2 methylation levels in TCGA samples analyzed by data mining. **P < 0.01 compared with normal tissues. (D) MSP analysis of DKK2 CGIs methylation frequency in primary cervical cancer tissues. CGIs, CpG islands; DKK2, Dickkopf 2; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; M or MSP, methylation-specific polymerase chain reaction (PCR); TCGA, The Cancer Genome Atlas; U, unmethylated-specific PCR.

DKK2 mRNA expression and methylation in cervical neoplasms

DiagnosisnmRNA expression (%)PMethylation (%)P
Normal2929/29 (100) 0/29 (0)
LSIL2525/25 (100)0.000031/25 (4.0)0.005
HSIL3821/38 (55.2)0.0019/38 (23.7)0.037
Cervical cancer7925/79 (31.6)0.01441/79 (51.9)0.004

Relationship between HR-HPV infection type and DKK2 methylation rate

GroupHR-HPV (+)nDKK2 methylationP
n%
Cervical cancer 0.015
HPV16/18(+)483266.7
HPV16/18(−)25936
Precancerous lesions 0.48
HPV16/18(+)29724.1
HPV16/18(−)23315

DKK2 mRNA expression, DKK2 promoter methylation and clinicopathological characteristics of cervical cancer

Clinical statusnDKK2 mRNA expressionDKK2 methylation
+ P+ P
Clinical stage
Stage IA22813 (46.4%)15 (53.6%)0.09912 (42.9%)16 (57.1%)0.32
Stage IB1+IB2419 (22.0%)32 (78.0%) 22 (53.7%)19 (46.3%)
Stage IIA103 (30.0%)7 (70.0%) 7 (70.0%)3 (30.0%)
Histology
Squamous cell carcinoma7021 (30.0%)49 (70.0%)0.3836 (51.4%)34 (48.6%)0.55
Adenocarcinoma94 (44.4%)5 (55.6%) 5 (55.6%)4 (44.4%)
Tumor grade
Grade 1155 (33.3%)10 (66.7%)0.816 (40.0%)9 (60.0%)0.17
Grade 24212 (28.6%)30 (71.4%) 20 (47.6%)22 (52.4%)
Grade 3228 (36.4%)14 (63.6%) 15 (68.2%)7 (31.8%)
Lymph node metastasis
Negative6223 (37.1%)39 (62.9%)0.04728 (45.2%)34 (54.8%)0.022
Positive172 (11.8%)15 (88.2%) 13 (76.5%)4 (23.5%)

Primer pairs used for mRNA expression and the size of PCR products

GenePrimer information (5′-3′)Product size
DKK25′-GTACCAAGGACTGGCATTCG-3′ (F)5′-ATCTCGGTGGCAGCGCTTCT-3′ (R)169 bp
GAPDH5′-CCAGCAAGAGCACAAGAGGAA-3′ (F)5′-CAAGGGGTCTACATGGCAACT-3′ (R)114 bp
DKK2 (M)5′-AGAGTTAAATCGTCGAGATTTC-3′ (F)5′-CTAAAAACAATCAAATACGAAACG-3′ (R)146 bp
DKK2 (U)5′-GGAGAGTTAAATTGTTGAGATTTT-3′ (F)5′-ACTAAAAACAATCAAATACAAAACA-3′ (R)149 bp
DOI: https://doi.org/10.2478/abm-2022-0022 | Journal eISSN: 1875-855X | Journal ISSN: 1905-7415
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Language: English
Page range: 181 - 189
Published on: Aug 31, 2022
Published by: Chulalongkorn University
In partnership with: Paradigm Publishing Services
Publication frequency: 6 issues per year
Keywords:
biomarkers,
data mining,
DKK2 protein,
human,
DNA methylation,
human papillomavirus,
uterine cervical neoplasms
Related subjects:
Medicine,
Assistive professions, nursing,
Basic medical science,
Basic medical science, other,
Clinical medicine,
Clinical medicine, other

© 2022 Xian Zhang, Aihua Li, Jie Wu, Yu Wu, Xiaoping Ma, Yanjun Liu, Qingfa Chen, Yan Zhang, published by Chulalongkorn University
This work is licensed under the Creative Commons Attribution 4.0 License.

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