Introduction
Methodology
Clinical samples
Total number | Number of males | Number of females | Mean age (Years) | Age range (years) | |
---|---|---|---|---|---|
Total GBC Cases | 30 | 3 | 27 | 54.5 | 32–86 |
Stages | |||||
GBC, Stage II | 5 | 0 | 5 | 56.6 | 35–86 |
GBC, Stage IIIA | 10 | 1 | 9 | 51.3 | 40–74 |
GBC, Stage IIIB | 12 | 1 | 11 | 54.0 | 40–72 |
GBC, Stage IV | 3 | 1 | 2 | 55 | 38–66 |
Histological grade | |||||
Well-differentiated (G1) | 1 | 0 | 1 | –- | –- |
Moderately-differentiated (G2) | 26 | 2 | 24 | –- | –- |
Poorly-differentiated (G3) | 3 | 1 | 2 | –- | –- |
LN status | |||||
LN Negative | 15 | 1 | 14 | –- | –- |
LN Positive | 15 | 2 | 13 | –- | –- |
Controls (GSD) | 15 | 2 | 13 | 40.8 | 22–56 |
Protein extraction
iTRAQ Labeling
LC–MS/MS analysis
Identification and quantification of proteins
Bioinformatic analysis
Western blot analysis
Immunohistochemistry analysis
Results
Identification of differentially expressed proteins in LN positive GBC
LN Negative GBC
|
LN Positive GBC
| |||||||
---|---|---|---|---|---|---|---|---|
Gene Symbol
|
Protein Name
|
GBC1
|
GBC2
|
GBC3
|
GBC4
|
GBC5
|
GBC6
|
GBC7
|
(A)
| ||||||||
A1BG | Alpha-1B-glycoprotein precursor | 1.296 | 0.538 | 0.51 |
0.447
| 0.581 | 0.883 | 0.734 |
ADH1C | Alcohol dehydrogenase 1C | 0.809 | 4.096 | 1.362 | 0.486 | 0.656 | 0.627 |
0.448
|
AGR2 | Anterior gradient protein 2 | 0.99 | 0.53 |
1.998
| 1.582 |
4.208
| 1.685 | 0.712 |
ALDH18A1 | Delta-1-pyrroline-5-carboxylate synthase isoform X1 | 1.126 | 0.867 | 1.471 |
3.227
| 1.416 | 0.927 | 1.246 |
ANP32A | Acidic leucine-rich nuclear phosphoprotein 32 family member A | 0.845 | 0.808 | 1.534 |
2.300
| 1.439 | 1.214 | 1.219 |
ANXA10 | Annexin A10 isoform X1 | 0.886 | 1.017 | 1.623 |
4.130
| 1.087 | 1.283 | 0.889 |
AOC3 | Membrane primary amine oxidase isoform X1 | 0.559 | 0.749 | 0.856 |
0.50
| 0.733 | 0.703 |
0.485
|
APEX1 | DNA-(apurinic or apyrimidinic site) lyase | 1.372 | 0.688 | 1.223 |
2.057
| 1.462 | 1.311 | 1.311 |
APOC3 | Apolipoprotein C-III precursor | 0.68 | 1.408 | 0.466 |
0.165
|
0.158
|
0.109
|
0.245
|
CAPG | Macrophage-capping protein isoform X1 | 1.274 | 0.606 | 0.877 | 1.363 | 1.103 | 1.402 |
2.061
|
CKMT1A | Creatine kinase U-type, mitochondrial isoform 1 precursor | 1.163 | 0.786 | 1.367 |
3.211
| 1.695 | 1.445 | 1.014 |
COL1A2 | Collagen alpha-2(I) chain precurso | 0.677 | 0.654 | 0.623 | 0.520 |
0.486
| 0.563 | 0.621 |
COX4I1 | Cytochrome c oxidase subunit 4 isoform 1, mitochondrial isoform X1 | 0.712 | 1.593 | 1.013 |
2.687
| 1.265 | 0.843 | 1.012 |
COX5A | Cytochrome c oxidase subunit 5A, mitochondrial precursor | 0.652 | 1.782 | 1.321 |
2.817
| 1.397 | 0.894 | 1.006 |
COX6B1 | Cytochrome c oxidase subunit 6B1 | 0.891 | 1.431 | 1.406 |
2.289
| 1.047 | 1.140 | 0.977 |
DLST | Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex, mitochondrial isoform 1 precursor | 0.756 | 1.553 | 1.66 |
3.701
|
2.754
| 0.988 | 1.215 |
ECH1 | Delta(3,5)-Delta(2,4)-dienoyl-coa isomerase, mitochondrial isoform X1 | 0.934 | 1.796 | 1.434 | 1.135 |
2.459
| 0.993 | 0.957 |
EEF1A1 | Elongation factor 1-alpha 1 | 0.665 | 1.741 | 1.307 |
2.511
| 1.817 | 0.666 |
3.143
|
FABP5 | Fatty acid-binding protein 5 | 0.895 | 0.545 | 1.464 | 0.827 | 1.438 | 1.035 |
8.795
|
GMDS | GDP-mannose 4,6 dehydratase isoform 1 | 1.033 | 0.937 | 1.398 |
3.061
| 0.816 | 0.937 | 0.853 |
HADHB | Trifunctional enzyme subunit beta, mitochondrial isoform X1 | 0.988 | 1.565 | 1.457 |
2.118
| 1.514 | 0.995 | 1.093 |
HNRNPA1 | Heterogeneous nuclear ribonucleoprotein A1 isoform X1 | 0.618 | 0.813 | 1.613 |
3.609
|
3.282
| 1.135 |
2.160
|
HNRNPA2B1 | Heterogeneous nuclear ribonucleoproteins A2/B1 isoform X1 | 0.523 | 0.831 | 1.256 |
2.594
|
2.939
| 0.807 |
2.714
|
HNRNPA3 | Heterogeneous nuclear ribonucleoprotein A3 isoform a | 0.699 | 0.659 | 1.356 | 1.376 |
3.393
| 0.672 |
2.360
|
HNRNPK | Heterogeneous nuclear ribonucleoprotein K isoform X1 | 0.91 | 0.816 | 1.922 |
2.541
|
2.434
| 1.183 |
2.334
|
HSP90AA1 | Heat shock protein HSP 90-alpha isoform 1 | 0.743 | 0.976 | 1.611 | 1.744 |
2.088
| 1.139 | 1.574 |
HSP90AB1 | Heat shock protein HSP 90-beta isoform X1 | 0.802 | 0.858 | 1.903 |
2.421
| 1.800 | 1.345 |
2.728
|
HSPA8 | Heat shock cognate 71 kda protein isoform X1 | 0.712 | 1.162 | 1.067 | 1.588 | 1.588 | 1.726 |
2.270
|
HSPA9 | Stress-70 protein, mitochondrial precursor | 0.796 | 1.664 | 1.656 |
3.183
|
2.044
| 1.037 |
2.357
|
IDH2 | Isocitrate dehydrogenase [NADP], mitochondrial isoform 1 precursor | 1.011 | 1.722 | 1.231 |
2.148
| 1.310 | 0.744 | 1.181 |
KRT1 | Keratin, type II cytoskeletal 1 | 1.418 | 0.585 | 0.606 |
0.473
| 0.768 | 0.611 | 0.897 |
KRT17 | Keratin, type I cytoskeletal 17 | 1.088 | 0.765 | 1.091 | 0.827 | 1.556 |
4.911
|
7.180
|
KRT19 | Keratin, type I cytoskeletal 19 | 1.002 | 0.749 | 1.788 |
7.241
|
3.853
|
2.390
| 0.930 |
KRT6A | Keratin, type II cytoskeletal 6A | 1.252 | 0.732 | 1.397 | 1.202 | 0.888 | 1.352 |
2.212
|
KRT7 | Keratin, type II cytoskeletal 7 | 0.878 | 0.72 | 0.963 |
2.651
| 1.944 |
2.497
| 0.820 |
LASP1 | LIM and SH3 domain protein 1 isoform a | 1.531 | 1.045 | 1.107 |
2.971
| 1.467 | 0.881 | 1.703 |
LRPPRC | Leucine-rich PPR motif-containing protein, mitochondrial precursor | 1.312 | 0.659 | 1.544 |
3.315
|
2.019
| 0.905 | 1.217 |
MDH2 | Malate dehydrogenase, mitochondrial isoform 1 precursor | 0.797 | 1.519 | 1.651 |
3.556
|
2.404
| 1.465 | 1.694 |
NPM1 | Nucleophosmin isoform 1 | 0.72 | 1.224 | 1.131 |
3.362
| 0.835 | 1.725 |
2.930
|
PGD | 6-phosphogluconate dehydrogenase, decarboxylating isoform 1 | 1.811 | 0.967 | 1.04 | 1.270 | 1.084 | 1.132 |
2.052
|
PKM | Pyruvate kinase PKM isoform c | 0.9 | 0.599 | 1.345 |
2.065
| 1.333 | 1.416 | 1.940 |
PPIA | Peptidyl-prolyl cis–trans isomerase A isoform 1 | 0.733 | 0.908 | 1.3 | 1.778 | 1.747 | 1.446 |
2.435
|
PPIB | Peptidyl-prolyl cis–trans isomerase B precursor | 0.746 | 1.261 | 1.31 | 1.577 | 1.066 | 1.279 |
2.299
|
RPS28 | 40S ribosomal protein S28 | 0.82 | 1.712 | 1.642 |
2.046
| 1.871 | 0.681 |
2.114
|
SERPINH1 | Serpin H1 isoform X1 | 0.988 | 0.881 | 1.269 | 1.046 | 1.450 | 1.061 |
2.587
|
SLC25A3 | Phosphate carrier protein, mitochondrial isoform a precursor | 0.975 | 1.608 | 2.098 |
2.371
| 1.585 | 0.895 | 1.314 |
SLC25A6 | ADP/ATP translocase 3 | 1.247 | 1.422 | 1.776 |
2.021
| 1.701 | 0.950 | 1.430 |
SRI | Sorcin isoform A | 0.868 | 0.753 | 1.011 | 0.828 |
2.380
|
2.394
| 0.941 |
TAGLN2 | Transgelin-2 isoform | 0.566 | 0.535 | 0.634 | 0.965 | 1.624 | 1.629 |
2.082
|
TUFM | Elongation factor Tu, mitochondrial isoform 1 precursor | 0.975 | 0.828 | 1.582 |
3.457
| 1.009 | 0.803 | 1.609 |
TYMP | Thymidine phosphorylase isoform 1 precursor | 1.97 | 1.138 | 1.024 | 0.905 | 1.045 | 1.252 |
2.441
|
VCAN | Versican core protein isoform 1 precursor | 0.626 | 0.725 | 0.895 | 0.690 | 1.059 |
3.567
| 0.807 |
VIM | Vimentin | 0.686 | 0.522 | 0.685 |
0.387
| 0.863 | 1.376 | 1.479 |
(B)
| ||||||||
ADH4 | Alcohol dehydrogenase 4 isoform 1 |
0.50
|
9.87
|
2.15
|
0.49
|
0.50
|
0.47
|
0.43
|
ANXA2 | Annexin A2 isoform 1 | 0.54 |
0.47
| 0.58 | 1.51 | 1.29 |
2.94
|
2.09
|
ANXA5 | Annexin A5 | 0.76 |
0.40
| 0.52 | 1.60 | 1.17 |
2.75
| 1.15 |
RDH16 | Retinol dehydrogenase 16 isoform 1 | 0.53 | 3.57 | 3.54 | 0.68 |
0.49
|
0.39
|
0.50
|
UGT2B7 | UDP-glucuronosyltransferase 2B7 isoform 1 precursor | 0.49 |
3.62
| 1.34 | 0.68 |
0.49
| 0.38 |
0.37
|