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Journal of Chinese Integrative Medicine ›› 2011, Vol. 9 ›› Issue (10): 1110-1117.doi: 10.3736/jcim20111012

• Original Experimental Research • Previous Articles     Next Articles

Inhibitory effects of osthole, psoralen and aconitine on invasive activities of breast cancer MDA-MB-231BO cell line and the mechanisms

Bao-feng Guo, Sheng Liu(), Yi-yi Ye, Xiang-hui Han   

  1. Pharmacological Laboratory of Traditional Chinese Medicine, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
  • Received:2011-04-15 Accepted:2011-05-09 Online:2011-10-20 Published:2011-10-15
  • Contact: Liu Sheng E-mail:yige823@yahoo.com.cn

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Objective: To explore the inhibitory effects and to investigate the mechanisms of combined treatment of osthole, psoralen with aconitine on human breast cancer cell line MDA-MB-231BO.

Methods: The best inhibitory concentration of osthole, psoralen combined with aconitine on MDA-MB-231BO cells was obtained by stepwise regression analysis after adopting a uniform experiment design. The invasive activities were observed by transwell assays, and expressions of transforming growth factor-β1 (TGF-β1), Smad2, Smad3, Smad4, Smad7, nuclear factor-κB (NF-κB) and receptor activator of NF-κB ligand (RANK) mRNAs were analyzed by real-time quantitative polymerase chain reaction.

Results: The optimal combination concentrations of osthol, psoralen and aconitine were 6.44, 8.89 and 9.44 μg/mL, respectively. Cell invasion was significantly inhibited after 24 hours of treatment using the combination drugs and zoledronic acid. TGF-β1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group and zoledronic acid group were significantly reduced compared with the control group (P<0.01). Furthermore, TGF-β1, Smad2, Smad3, Smad4, Smad7, NF-κB and RANK mRNA expressions of the optimal combination group were significantly lower than those of the weak combination group (P<0.01).

Conclusion: Combination treatment of osthole, psoralen with aconitine can inhibit cancer cell invasion, which is a result of alteration of the TGF-β/Smad signaling pathway and down-regulation of NF-κB and RANK expressions.

Key words: plant extracts, osthole, psoralen, aconitine, breast neoplasm, neoplasm invasiveness

Table 1

Schedule of the uniform design (μg/mL)"


Number
X1 (osthol)
X2 (psoralen)
X3 (aconitine)

1
2.00
7.22
8.33

2
2.89
10.00
6.11

3
3.78
6.67
10.00

4
4.67
9.44
7.78

5
5.56
6.11
5.56

6
6.44
8.89
9.44

7
7.33
5.56
7.22

8
8.22
8.33
5.00

9
9.11
5.00
8.89

10
10.00
7.78
6.67

Table 2

TaqMan primers for gene members"


Gene
Primer sequence (5′→3′)
Product size (bp)
Annealing temperature (℃)

TGF-β1
F: AGCGACTCGCCAGAGTGGTTA

R: GCAGTGTGTTATCCCTGCTGTCA
130
58

Smad2
F: TTAACCGAAATGCCACGGTAGAA

R: GCTCTGCACAAAGATTGCACTATCA
122
62

Smad3
F: AGGCGTGCGGCTCTACTACATC

R: CAGCGAACTCCTGGTTGTTGAA
173
58

Smad4
F: CAGCACTACCACCTGGACTGGA

R: CTGGAATGCAAGCTCATTGTGAA
145
62

Smad7
F: TGCTGTGCAAAGTGTTCAGGTG

R: CCATCGGGTATCTGGAGTAAGGA
177
58

NF-κB
F: GCCTCCACAAGGCAGCAAATA

R: CACCACTGGTCAGAGACTCGGTAA
160
62

RANK
F: TTGTGGCCGCCTAAGTGGA

R: ACCACCTTGATCTGGGTAGCACATA
160
58

GAPDH
F: GCACCGTCAAGGCTGAGAAC

R: TGGTGAAGACGCCAGTGGA
138
62

Table 3

Effects of the uniform design-based combinations on osthole, psoralen and aconitine"


Group
Osthole (μg/mL)
Psoralen (μg/mL)
Aconitine (μg/mL)
Optical density value ($\overline{x}$±s)

1
2.00
7.22
8.33
0.62±0.05

2
2.89
10.00
6.11
0.46±0.03

3
3.78
6.67
10.00
0.51±0.04

4
4.67
9.44
7.78
0.56±0.03

5
5.56
6.11
5.56
0.68±0.04

6
6.44
8.89
9.44
0.43±0.03

7
7.33
5.56
7.22
0.50±0.03

8
8.22
8.33
5.00
0.53±0.04

9
9.11
5.00
8.89
0.57±0.03

10
10.00
7.78
6.67
0.53±0.05

Figure 1

Effects of three components on cell invasion ability after treatment of MDA-MB-231BO cells for 24 h C: Control group; L: Weak combination group; H: Optimal combination group; Y: Zoledronic acid injection group. MDA-MB-231BO cells were observed under a DPFO Olympus microscope by HE staining, ×400). Data were represented as $\overline{x}$±s, n=5. **P<0.01, vs control group; △△P<0.01, vs L group."

Table 4

mRNA expressions in MDA-MB-231BO cells after medicine treatment"


Group
n
Smad7/β-actin
Smad2/β-actin
Smad3/β-actin
Smad4/β-actin

C
5
1
1
1
1

L
5
1.220±0.050
1.385±0.100
0.585±0.040
0.222±0.020

H
5
1.051±0.030**△△
0.500±0.040**△△
0.104±0.010**△△
0.151±0.010**△△

Y
5
1.033±0.020*
0.484±0.040**
0.090±0.005**
0.118±0.010**

Group
n
TGF-β1/β-actin
NF-κB/β-actin
RANK/β-actin

C
5
1
1
1

L
5
0.900±0.050
0.329±0.010
0.627±0.030

H
5
0.475±0.040**△△
0.210±0.010**△△
0.015±0.009**△△

Y
5
0.390±0.030**
0.600±0.020**
0.498±0.030**
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