Holliday junction–recognizing protein promotes cell proliferation and correlates with unfavorable clinical outcome of hepatocellular carcinoma

Figures & data

Figure 1 Images of HJURP immunohistochemistry staining. (A) Low HJURP expression and (B) high HJURP expression. Scale bar: 50 μm.

Abbreviation: HJURP, Holliday junction–recognizing protein.

Table 1 Correlation between HJURP and clinicopatholgical factors

Characters	Number	HJURP		P-value*
		Low	High
Gender				1.000
Female	43	32	11
Male	121	90	31
Age (years)				0.721
<60	77	56	21
≥60	87	66	21
Tumor size (cm)				0.006
≤5	66	57	9
>5	98	65	33
Tumor number				0.584
Single	103	75	28
Multiple	61	47	14
Differentiation				1.000
Well + moderate	98	73	25
Poor	66	49	17
Liver cirrhosis				0.851
Presence	111	83	28
Absence	53	39	14
HBsAg				0.356
Negative	60	42	18
Positive	104	80	24
BCLC stage				0.002
0&A	86	73	13
B&C	78	49	29
TNM stage				0.324
I–II	119	91	28
III–IV	45	31	14

Notes:

* Means calculated by chi-square test. Bold represents statistically significant values (P<0.05).

Abbreviations: BCLC, Barcelona clinic liver cancer; HBsAg, hepatitis B surface antigen; HJURP, Holliday junction–recognizing protein.

Figure 2 Correlation between HJURP expression and the overall survival rate. Survival curves representing low and high HJURP expressions were displayed by Kaplan–Meier method and statistical significance was analyzed by log-rank test (P=0.003).

Abbreviation: HJURP, Holliday junction–recognizing protein.

Table 2 Univariate analysis

Characters	5-Year survival rate	P-value*
Gender		0.302
Female	27.9
Male	36.2
Age (years)		0.850
<60	31.8
≥60	36.1
Tumor size (cm)		0.766
≤5	33.0
>5	35.7
Tumor number		0.016
Single	37.3
Multiple	28.9
Differentiation		0.047
Well + moderate	52.2
Poor	23.8
Liver cirrhosis		0.068
Presence	35.1
Absence	33.0
HBsAg		0.150
Negative	41.7
Positive	28.1
BCLC stage		0.005
0&A	39.8
B&C	26.3
TNM stage		0.004
I–II	37.9
III–IV	24.4
HJURP		0.003
Low	37.8
High	24.5

Notes:

* Means calculated by log-rank test. Bold represents statistically significant values (P<0.05).

Abbreviations: BCLC, Barcelona clinic liver cancer; HBsAg, hepatitis B surface antigen; HJURP, Holliday junction–recognizing protein.

Table 3 Multivariate analysis

Characters	Hazard ratio	95% CI	P-value*
Tumor number			0.001
Single	1
Multiple	2.50	1.42–4.40
Differentiation			0.014
Well + moderate	1
Poor	2.00	1.15–3.49
Liver cirrhosis			0.015
Presence	1
Absence	1.95	1.14–3.33
HJURP			<0.001
Low	1
High	2.75	1.60–4.73

Note:

* Means calculated by Cox regression model.

Abbreviations: BCLC, Barcelona clinic liver cancer; HJURP, Holliday junction–recognizing protein; CI, confidence interval.

Figure 3 HJURP promotes HCC cell line proliferation. (A) HJURP expression in HCC cell line HuH7, SMMC7721, and HepG2 was detected by Western blotting. (B) HJURP mRNA level in HCC tissues was significantly higher than that of the corresponding normal liver tissues (P=0.026). (C) Successful HJURP overexpression in SMMC7721 was verified by Western blotting. HJURP was cloned into pFLAG-CMV2 by double digestion with restriction enzymes and transfected into SMMC7721. HJURP expression 48 h after transfection was detected by Western blotting. (D) Successful HJURP knockdown in SMMC7721 was verified by Western blotting. HJURP knockdown was performed by siRNA. HJURP expression 48 h after siRNA transfection was detected by Western blotting. (E) Cell proliferation was detected by MTT assay in SMMC7721 cell line after regulating HJURP expression. After 24 h of transfection with scrambled RNA, siRNA, control vector, or Flag-HJURP vector, cells were passaged into 96-well plates and starved for 6 h. Then medium containing 10% FBS was added, which was set as time 0. Cell proliferation at time point 24, 48, or 72 h after time 0 was detected. The proliferation ratio of group without any treatment (control group) at 24 h was set as 100% and other groups were standardized with this baseline. Column graph was displayed with ± SEM. *P<0.05 and **P<0.01.

Abbreviations: FBS, fetal bovine serum; HCC, hepatocellular carcinoma; HJURP, Holliday junction–recognizing protein; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium; SEM, standard error of mean; siRNA, small interfering RNA; siHJURP, siRNA against HJURP.