Auphen and dibutyryl cAMP suppress growth of hepatocellular carcinoma by regulating expression of aquaporins 3 and 9 in vivo

Table 1 Clinicopathological data of the hepatocellular carcinoma cohort

Clinicopathologic parameters	Frequency	%
All cases	30
Gender
Male	23	76.7
Female	7	23.3
Age (yr)
< 50	5	16.7
> 50	25	83.3
Tumor size (cm)
< 5	8	26.7
> 5	22	73.3
Serum HBsAg
Positive	24	80.0
Negative	6	20.0
Serum AFP (ng/mL)
< 25	4	13.3
> 25	26	86.7
Cirrhosis
Presence	27	90.0
Absence	3	10.0
UICC stage
I + II	13	43.3
III + IV	17	56.7
Metastasis/Recurrence
Yes	18	60.0
No	12	40.0
Edmondson grade
Low (I/II)	10	33.3
High (III/IV)	20	66.7

Table 2 Primer sequences used for the amplification of different genes by quantitative PCR

Gene	Forward primer sequence	Reverse primer sequence
AQP3	5'-CACAGCCGGCATCTTTGCTA-3'	5'-TGGCCAGCACACACACGATA-3'
AQP9	5'-CTTAACAATTCACAAGGCACTT-3'	5'-TCTCAGCCAGCTACTGATCTTC-3'
AFP	5'-ACCCTGGTGTTGGCCAGTGC-3'	5'-GCAGCGCTACACCCTGAGCT-3'
β-actin	5'-TCACCCACACTGTGCCCATCTACGA-3'	5'-CAGCGGAACCGCTCATTGCCAATGG-3'

Table 3 Correlation of AQP3 and AQP9 expression levels (low and high) to clinico-pathological data

Factors	AQP3		OR (95%CI)	P value	AQP9		OR (95%CI)	P value
	Low	High			Low	High
Gender
Male	4	19	0.526 (0.074-3.748)	0.603	16	1	7.111 (0.686-73.76)	0.138
Female	2	5			9	4
Age (yr)
< 50	1	4	1.000 (0.091-11.03)	1.000	3	2	0.205 (0.024-1.771)	0.183
> 50	5	20			22	3
Tumor size (cm)
< 5	3	5	3.800 (0.580-24.90)	0.300	6	2	0.474 (0.063-3.540)	0.589
> 5	3	19			19	3
Serum HBsAg
Positive	5	19	1.316 (0.124-13.98)	1.000	21	3	1.185 (0.166-8.475)	1.000
Negative	1	5			4	2
Serum AFP (ng/mL)
< 25	3	2	11.00 (1.271-95.23)	0.041	4	0	2.302 (0.107-49.54)	1.000
> 25	3	22			21	5
Cirrhosis
Presence	6	21	2.116 (0.096-46.56)	1.000	11	2	6.000 (0.780-46.17)	0.102
Absence	0	3			14	3
UICC stage
I + II	4	4	10.00 (1.341-74.55)	0.029	2	4	0.022 (0.002-0.300)	0.003
III + IV	2	20			23	1
Metastasis/recurrence
Yes	1	17	0.082 (0.008-0.839)	0.026	17	1	12.67 (1.177-136.4)	0.031
No	5	7			8	4
Edmondson grade
Low (I/II)	3	3	14.00 (1.741-112.6)	0.016	7	4	0.097 (0.009-1.029)	0.047
High (III/IV)	3	21			18	1

Table 4 Cox regression analysis of patients with hepatocellular carcinoma

Variables	Univariate		P value
	HR	95%CI
AQP3 expression (1 = low, 2 = high)	4.948	1.037-23.605	0.045
AQP9 expression (1 = low, 2 = high)	10.835	1.488-78.877	0.019
Gender (1 = Male, 2 = Female)	2.681	0.852-8.436	0.092
Age (1 < 50, 2 ≥ 50)	2.579	0.332-20.049	0.365
Tumor size (1 < 5 cm, 2 ≥ 5)	1.261	0.334-4.759	0.732
Serum HBsAg (1 = Positive, 2= Negative)	0.575	0.153-2.154	0.411
Serum AFP (1 < 25 ng/mL, 2 ≥ 25 ng/mL)	1.102	0.293-4.146	0.886
Cirrhosis (1 = Presence, 2 = Absence)	1.401	0.373-5.268	0.618
UICC stage (1 = I + II, 2 = III + IV)	0.115	0.027-0.485	0.003
Metastasis/Recurrence (1 = Yes, 2 = No)	0.228	0.065-0.804	0.021
Edmondson grade [1 = Low (I/II), 2 = High (III/IV)]	0.560	0.175-1.791	0.328