01.12.2017 | Research article | Ausgabe 1/2017 Open Access

CLASP2 is involved in the EMT and early progression after transurethral resection of the bladder tumor
- Zeitschrift:
- BMC Cancer > Ausgabe 1/2017
Background
Methods
Cell lines and cell culture
Western blot analysis
Establishment of stable cell lines
Cell proliferation assay
Clonogenic formation assays
Wound healing assay
Cell invasive assay
Real time semi-quantitative (RT-PCR)
-
CLASP2 (forward: 5’- TTGTCGTCCTCTGTCAGTGC-3’; reverse: 5’- TGCCACGTCTTCTGTCTGTC-3’),
-
E-cadherin (forward: 5’-CGGGAATGCAGTTGAGGATC-3’; reverse: 5’-AGGATGGTGTAAGCGATGGC-3’),
-
Vimentin (forward: 5’-GACCTCTACGAGGAGGAGAT -3’; reverse: 5’-TTGTCAACATCCTGTCTGAA-3’)
-
GAPDH (forward: 5’-ACCACAGTCCATGCCATCAC-3’; reverss: 5’-TCCACCACCCTGTTGCTGTA-3’).
Patients and samples collection
Statistical analysis
Results
Expressions of CLASP2 varied in four BC cell lines
Manipulation of CLASP2 expression changed EMT-related markers
CLASP2 was involved in proliferation and clonogenic formation in BC cells
CLASP2 could boost migration and invasion in BC cells
Patients and baseline characteristics
Variables
|
Patients (
n)
|
Progression within 2 years
|
||
---|---|---|---|---|
No
(
n)
|
Yes
(
n)
|
P
|
||
Total (%)
|
102
|
68
|
34
|
|
Gender
|
||||
Male
|
72
|
52
|
20
|
0.11
|
Female
|
30
|
16
|
14
|
|
Age(years)
|
||||
< 60
|
38
|
27
|
11
|
0.61
|
> 60
|
64
|
41
|
23
|
|
Intravesical instillation
|
||||
Immediate
|
29
|
22
|
7
|
0.31
|
Maintenance
|
73
|
46
|
27
|
|
Tumor stage
|
||||
Ta
|
34
|
25
|
9
|
0.41
|
T1
|
68
|
43
|
25
|
|
Tumor grade
|
||||
G1
|
33
|
26
|
7
|
0.04
|
G2
|
49
|
33
|
16
|
|
G3
|
20
|
9
|
11
|
|
Concomitant CIS
|
||||
Yes
|
3
|
1
|
2
|
0.26
|
No
|
99
|
67
|
32
|
|
Tumor size (cm)
|
||||
< 3
|
59
|
44
|
15
|
0.07
|
> 3
|
43
|
24
|
19
|
|
Multiplicity
|
||||
Single
|
44
|
34
|
10
|
0.06
|
Multiple
|
58
|
34
|
24
|
|
CLASP2 mRNA (tumor)
|
||||
High
|
30
|
6
|
24
|
<0.01
|
Low
|
72
|
62
|
10
|
|
CLASP2 mRNA (urine)
|
||||
High
|
55
|
24
|
31
|
< 0.01
|
Low
|
47
|
44
|
3
|
|
E-cadherin mRNA (tumor)
|
||||
High
|
62
|
52
|
10
|
< 0.01
|
Low
|
40
|
16
|
24
|
|
E-cadherin mRNA (urine)
|
||||
High
|
60
|
54
|
6
|
< 0.01
|
Low
|
42
|
14
|
28
|
|
Vimentin mRNA (tumor)
|
||||
High
|
55
|
31
|
24
|
0.16
|
Low
|
47
|
37
|
10
|
|
Vimentin mRNA (urine)
|
||||
High
|
62
|
36
|
26
|
0.03
|
Low
|
40
|
32
|
8
|
Relationship between variables and progression with 2 years
Univariable analysis
|
|||
RR
a
|
95% CI
b
|
P
|
|
Gender (Male, Female)
|
1.12
|
0.91–1.22
|
0.67
|
Age(<60, >60 years)
|
1.03
|
0.98–1.17
|
0.61
|
Intravesical instillation (immediate, maintenance)
|
0.98
|
0.89–1.78
|
0.27
|
Tumor stage (Ta, T1)
|
1.34
|
1.03–1.56
|
0.67
|
Tumor grade (G1 and G2, G3)
|
2.78
|
1.05–6.93
|
0.01
|
Concomitant CIS (No, Yes)
|
1.52
|
0.21–7.43
|
0.66
|
Tumor size (<3, >3 cm)
|
2.11
|
1.29–4.32
|
0.05
|
Multiplicity (Single, Multiple)
|
1.98
|
0.98–3.92
|
0.07
|
CLASP2 mRNA (low, high in tumor)
|
3.83
|
1.12–12.22
|
0.02
|
CLASP2 mRNA (low, high in urine)
|
2.71
|
1.09–7.77
|
< 0.01
|
E-cadherin mRNA (high, low in tumor)
|
2.97
|
1.17–9.82
|
0.04
|
E-cadherin mRNA (high, low in urine)
|
3.24
|
0.97–10.81
|
0.02
|
Vimentin mRNA (low, high in tumor)
|
1.24
|
0.78–2.87
|
0.32
|
Vimentin mRNA (low, high in urine)
|
1.19
|
0.26–4.98
|
0.09
|
Multivariable analysis
c
|
|||
RR
|
95% CI
|
P
|
|
Tumor stage (Ta, T1)
|
1.14
|
0.91–1.86
|
0.78
|
Tumor grade (G1 and G2, G3)
|
2.19
|
1.01–4.98
|
0.04
|
Tumor size (<3, >3 cm)
|
1.71
|
1.08–3.37
|
0.07
|
Intravesical instillation (immediate, maintenance)
|
1.12
|
0.79–1.45
|
0.52
|
Multiplicity (Single, Multiple)
|
2.32
|
1.18–4.22
|
0.18
|
CLASP2 mRNA (low, high in tumor)
|
2.76
|
1.02–10.11
|
< 0.01
|
CLASP2 mRNA (low, high in urine)
|
3.75
|
1.09–6.12
|
< 0.01
|
E-cadherin mRNA (high, low in tumor)
|
2.78
|
1.17–8.24
|
0.01
|
E-cadherin mRNA (high, low in urine)
|
3.34
|
1.12–7.45
|
0.01
|
Vimentin mRNA (low, high in tumor)
|
1.35
|
0.89–3.43
|
0.51
|
Vimentin mRNA (low, high in urine)
|
1.78
|
0.26–3.48
|
0.35
|
AUC
b
|
95% CI
c
|
P
|
|
---|---|---|---|
Tumor grade
|
0.613
|
0.512–0.708
|
0.02
|
CLASP2 mRNA (in tumor)
|
0.702
|
0.603–0.788
|
<0.01
|
CLASP2 mRNA (in urine)
|
0.716
|
0.619–0.801
|
<0.01
|
E-cadherin mRNA (in tumor)
|
0.692
|
0.593–0.780
|
<0.01
|
E-cadherin mRNA (in urine)
|
0.710
|
0.612–0.795
|
<0.01
|
Vimentin mRNA (in tumor)
|
0.657
|
0.544–0.768
|
0.01
|
Vimentin mRNA (in urine)
|
0.687
|
0.580–0.792
|
<0.01
|
Combination of CLASP2 and E-cadherin mRNA (in urine)
|
0.763
|
0.662–0.864
|
<0.01
|