MicroRNA Expression Pattern in Intraductal Papillary Mucinous Neoplasm

Yun Gyoung Park; Kwang Hyuck Lee; Jong Kyun Lee; Kyu Taek Lee; Dong Wook Choi; Seong Ho Choi; Jin Seok Heo; Kee Taek Jang; Eun Mi Lee; Jung Ok Kim; Shin Young Min; Eun Jung Kim

doi:10.4166/kjg.2011.58.4.190

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Park, Lee, Lee, Lee, Choi, Choi, Heo, Jang, Lee, Kim, Min, and Kim: MicroRNA Expression Pattern in Intraductal Papillary Mucinous Neoplasm

Original Article

Korean J Gastroenterol 2011;58(4):190-200.

Published online: 4 October 2011

DOI: https://doi.org/10.4166/kjg.2011.58.4.190

MicroRNA Expression Pattern in Intraductal Papillary Mucinous Neoplasm

Yun Gyoung Park¹, Kwang Hyuck Lee^1,², Jong Kyun Lee^1,², Kyu Taek Lee^1,², Dong Wook Choi³, Seong Ho Choi³, Jin Seok Heo³, Kee Taek Jang⁴, Eun Mi Lee⁵, Jung Ok Kim¹, Shin Young Min¹, Eun Jung Kim¹

¹Department of Internal Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

²Division of Gastroenterology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

³Department of Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

⁴Department of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

⁵Samsung Biomedical Research Institute, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

Correspondence to: Kwang Hyuck Lee, Department of Gastroenterology, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50, Irwon-dong, Gangnam-gu, Seoul 135-710, Korea. Tel: +82-2-3410-3409, Fax: +82-2-3410-6983, E-mail: lkhyuck@gmail.com

Received 18 May 2011 Accepted 13 July 2011

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background/Aims

Intraductal papillary mucinous neoplasms (IPMN) are precursor lesions of fatal pancreatic cancer. Physiological function of microRNA is to regulate the stability and translation of mRNA. The aberrant microRNA expression is commonly observed in many cancers. The aim of this study was to analyze the expression pattern of microRNA in IPMN and evaluate the role of the microRNA.

Methods

Using two paraffin-embedded IPMN tissues, microRNA expression of normal tissue, IPMN adenoma and carcinoma were compared by cDNA-mediated annealing, selection, extension and ligation microarray assay. Using real time PCR, expression levels of aberrantly up-regulated microRNAs were assessed in another 20 IPMNs, four pancreatic cancer cell lines (Panc1, MiaPaCa-2, XPA-3, BxPC-3) and immortalized pancreatic ductal cell line (HPNE). Effect of suppressing highly over-expressed two microRNAs in pancreatic cancer cell lines with anti-microRNA inhibitors were evaluated using CCK-8 assay.

Results

Among aberrantly expressed 122 microRNAs in IPMN, miR-552, miR-25*, miR-183, miR-1300, miR-196a, miR-182*, and miR-30c–1* were consistently increased more than 3-fold. On average, miR-196a and miR-183 increased 10,824 folds and 26,519 folds in four pancreatic cancer cell lines compared with HPNE. These two microRNAs were also over-expressed in 20 IPMNs compared with HPNE. After applying anti-miRNA inhibitors, cell survival of four pancreatic cancer cell lines decreased by 24.5% with anti-miR-196a and by 14.2% with anti-miR-183 on average.

Conclusions

Aberrant expression of 122 microRNAs was observed in IPMN. Two microRNAs, miR-196a and miR-183-increased in IPMN and pancreatic cancer cell lines compared with immortalized dancreatic ductal cell line. The inhibitions of these microRNAs repressed cell proliferation of pancreatic cancer cell lines.

Keywords: Intraductal papillary mucinous neoplasm, microRNAs, miR-196a, miR-183

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Fig. 1.

MicroRNA-196a (A), microRNA-183 (B), and RNU6 (C) expression in pancreatic cancer cell lines and immortalized pancreatic ductal cells measured by real-time polymerase reaction (a: Panc-1, b: XPA-3, c: BxPC-3, d: MiaPaca-2, e: HPNE).

Fig. 2.

MicroRNA-196a suppression in pancreatic cancer cell lines with RNU6B (a: Control, b: With mock, c: With anti-miR-196). (A) Panc-1, (B) MiaPaCa-2, (C) XPA-3, (D) BxPC-3.

Fig. 3.

MicroRNA-183 suppression in pancreatic cancer cell lines with RNU6B (a: Control, b: With mock, c: With anti-miR-183). (A) Panc-1, (B) MiaPaCa-2, (C) XPA-3, (D) BxPC-3.

Fig. 4.

Effect of anti-microRNA-196a and anti-microRNA-183 transfection into Gemcitabine treated pancreatic cancer cell lines. (A) Panc-1, (B) MiaPaCa-2, (C) XPA-3, (D) BxPC-3.

Table 1.

Up and Down Regulated MicroRNA in IPMN Detected by DASL assay

microRNA	Patient 1		Patient 2
Up regulated	Adenoma	Carcinoma	Adenoma	Carcinoma
hsa-miR-552	17.36	3.91	51.52	65.06
hsa-miR-183	14.87	10.16	3.72	3.63
hsa-miR-1300	13.63	4.30	24.37	15.65
hsa-miR-196a	13.21	12.03	9.94	10.54
hsa-miR-182*	9.22	4.22	4.24	3.42
hsa-miR-30c–1*	4.23	7.44	7.72	5.18
hsa-miR-25*	3.84	3.69	8.20	5.13
hsa-miR-183*	8.45	2.20	5.32	2.14
hsa-miR-93*	6.15	2.63	4.70	5.97
hsa-miR-182	5.62	3.48	2.21	2.15
hsa-miR-1268	5.16	2.23	2.92	2.96
hsa-miR-885–3p	3.26	2.52	5.61	4.47
Down regulated	Adenoma	Carcinoma	Adenoma	Carcinoma
hsa-miR-125b–2*	−28.90	−2.65	−15.54	−47.93
hsa-miR-133a	−21.29	−7.51	−3.41	−5.04
hsa-miR-885–5p	−16.55	−3.50	−13.03	−14.75
hsa-miR-30a*	−11.99	−4.20	−9.18	−20.33
hsa-miR-1271	−10.07	−2.45	−4.04	−4.06
hsa-miR-1273	−8.26	−5.24	−4.98	−2.36
HS_182.1	−8.07	−2.02	−9.22	−3.79
hsa-miR-30c–2*	−7.99	−4.24	−4.93	−5.36
hsa-miR-1272	−7.75	−5.29	−4.61	−6.09
hsa-miR-378*	−6.63	−4.13	−2.87	1.39
hsa-miR-483–3p	−6.40	−2.31	−16.25	−4.19
hsa-miR-34c–5p	−6.10	−3.28	−3.60	−3.11
hsa-miR-143*	−5.80	−4.08	−2.21	−2.16
hsa-miR-34b*	−5.53	−3.21	−5.06	−3.41

IPMN, intraductal papillary mucinous neoplasm; DASL assay, cDNA-mediated annealing, selection, extension and ligation assay (Illumina San Diego, CA, USA).

Table 2.

MicroRNA Expression Patterns in Pancreatic Cancer Cell Lines (Panc-1, MiaPaCa-2, XPA-3, BxPC-3) Comparing with Immortalized Pancreatic Ductal Cell Lline (HPNE) by Real-time Polymerase Chain Reaction

Over-expressed microRNA In IPMN	HPNE	Panc-1	MiaPaCa-2	XPA-3	BxPC-3	Mean
hsa-miR-552	1	0	0	0	6.29	1.57
hsa-miR-183	1	5216	13913	22471	1698	10825
hsa-miR-1300	1	0.74	1.51	1.38	0.89	1.13
hsa-miR-196a	1	73385	1966	22397	8327	26519
hsa-miR-182*	1	8.38	31.31	42.24	8.38	22.58
hsa-miR-30c–1*	1	13.33	8.39	6.49	1.54	7.44
hsa-miR-25*	1	9.66	40.62	34.00	6.18	22.61

Table 3.

MicroRNA 196a and MicroRNA-183 Expressions in 20 IPMN Patients (A) Patient's Characteristics (B) MicroRNA-196a and Mic

Table 4.

MicroRNA-196a and MicroRNA-183 Suppression in Pancreatic Cancer Cell Lines

	miR-196a		miR-183
	Reference	With suppression	Reference	With suppression
Panc-1	100	<0.001	100	0.0001
MiaPaca-2	100	0.1	100	0.0001
XPA3	100	0.4	100	0.0001
BxPC3	100	<0.001	100	0.0001

Table 5.

Decrease of Cell Survival by MicroRNA Suppression

	No gemcitabine (%)		Low concentration of gemcitabine (%)
	miR-196a	miR-183	miR-196a	miR-183
XPA-3	25.6	6.6	18.7	4.8
MiaPaCa-2	13.5	−8.2	7.8	−2.7
BxPC-3	38.6	45.4	38.6	45.0
Panc-1	20.3	12.8	12.7	15.1
All	24.5	14.2	19.4	15.5

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