Journal of Traditional Chinese Medicine ›› 2022, Vol. 42 ›› Issue (4): 622-632.DOI: 10.19852/j.cnki.jtcm.20220617.002
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WEI Lexin1, LI Weiyi1, TIAN Ting2, ZHANG Ning1, YANG Shijing1, YANG Dongqing1, LI Guochun1(), YE Fang3()
Received:
2021-04-22
Accepted:
2021-07-18
Online:
2022-08-15
Published:
2022-07-12
Contact:
LI Guochun,YE Fang
About author:
Prof. YE Fang, the Key Research Department of Stasis-heat Pathogenesis of Traditional Chinese Medicine, the First Clinical Medical College, Nanjing University of Chinese Medicine, Nanjing 210023, China. 260958@njucm.edu.cn, Telephone: +86-25-85511926Supported by:
WEI Lexin, LI Weiyi, TIAN Ting, ZHANG Ning, YANG Shijing, YANG Dongqing, LI Guochun, YE Fang. Identification of novel biomarkers and therapeutic target candidates for stasis-heat symptom pattern of acute intracerebral hemorrhage by quantitative plasma proteomics[J]. Journal of Traditional Chinese Medicine, 2022, 42(4): 622-632.
Block | Tissue | Disease or normal | Patient number | Age/ gender | Score of stasis-heat | District and hospital | TMT labeling reagent ID | TMT labeling code |
---|---|---|---|---|---|---|---|---|
1 | 1 | Normal | 1 | 66/F | 0 | Nanjing | 1 | 126 |
1 | 2 | AICH (stasis-heat) | 2 | 61/F | 14 | Nanjing | 1 | 127 |
1 | 4 | AICH (non-stasis-heat) | 3 | 65/F | 7 | Nanjing | 1 | 129 |
Reference | - | - | - | - | 1 | 131 | ||
2 | 6 | Normal | 4 | 60/M | 0 | Nanjing | 2 | 126 |
2 | 7 | AICH (stasis-heat) | 5 | 56/M | 21 | Nanjing | 2 | 127 |
2 | 9 | AICH (non-stasis-heat) | 6 | 58/M | 9 | Nanjing | 2 | 129 |
Reference | - | - | - | - | 2 | 131 | ||
3 | 11 | Normal | 7 | 71/M | 0 | Zhenjiang | 3 | 126 |
3 | 12 | AICH (stasis-heat) | 8 | 69/M | 14 | Zhenjiang | 3 | 127 |
3 | 14 | AICH (non-stasis-heat) | 9 | 66/M | 9 | Zhenjiang | 3 | 129 |
Reference | - | - | - | - | 3 | 131 | ||
4 | 16 | Normal | 10 | 56/F | 0 | Zhenjiang | 4 | 126 |
4 | 17 | AICH (stasis-heat) | 11 | 61/F | 13 | Zhenjiang | 4 | 127 |
4 | 19 | AICH (non-stasis-heat) | 12 | 57/F | 4 | Zhenjiang | 4 | 129 |
Reference | - | - | - | - | 4 | 131 |
Table 1 Experimental protocol and basic characteristics in AICH patients with stasis-heat, AICH patients with non-stasis-heat, and health controls groups (n = 4 per group)
Block | Tissue | Disease or normal | Patient number | Age/ gender | Score of stasis-heat | District and hospital | TMT labeling reagent ID | TMT labeling code |
---|---|---|---|---|---|---|---|---|
1 | 1 | Normal | 1 | 66/F | 0 | Nanjing | 1 | 126 |
1 | 2 | AICH (stasis-heat) | 2 | 61/F | 14 | Nanjing | 1 | 127 |
1 | 4 | AICH (non-stasis-heat) | 3 | 65/F | 7 | Nanjing | 1 | 129 |
Reference | - | - | - | - | 1 | 131 | ||
2 | 6 | Normal | 4 | 60/M | 0 | Nanjing | 2 | 126 |
2 | 7 | AICH (stasis-heat) | 5 | 56/M | 21 | Nanjing | 2 | 127 |
2 | 9 | AICH (non-stasis-heat) | 6 | 58/M | 9 | Nanjing | 2 | 129 |
Reference | - | - | - | - | 2 | 131 | ||
3 | 11 | Normal | 7 | 71/M | 0 | Zhenjiang | 3 | 126 |
3 | 12 | AICH (stasis-heat) | 8 | 69/M | 14 | Zhenjiang | 3 | 127 |
3 | 14 | AICH (non-stasis-heat) | 9 | 66/M | 9 | Zhenjiang | 3 | 129 |
Reference | - | - | - | - | 3 | 131 | ||
4 | 16 | Normal | 10 | 56/F | 0 | Zhenjiang | 4 | 126 |
4 | 17 | AICH (stasis-heat) | 11 | 61/F | 13 | Zhenjiang | 4 | 127 |
4 | 19 | AICH (non-stasis-heat) | 12 | 57/F | 4 | Zhenjiang | 4 | 129 |
Reference | - | - | - | - | 4 | 131 |
Figure 1 Experimental profile AICH: acute intracerebral hemorrhage; SH: stasis-heat; NSH: non-stasis-heat; HC: healthy controls; TMT: tandem mass tags; LC-MS/MS: liquid chromatography tandem-mass spectrometry.
No. | UniProt accession | Symbol | Entrez gene name | Abundance in SHP | Abundance in HG | Fold change | 95%CI for fold change | P value | |
---|---|---|---|---|---|---|---|---|---|
1 | V9HWD8 | A1BG | Alpha-1-B glycoprotein | 1.678±0.102 | 1.127±0.359 | 1.490±0.091 | 1.490 (1.346, 1.634) | 0.034 | |
2 | A0A024R694 | ACTN1 | Actinin, alpha 1 | 1.408±0.086 | 0.680±0.238 | 2.069±0.126 | 2.069 (0.939, 3.200) | 0.051 | |
3 | P06727 | APOA4 | Apolipoprotein A-IV | 0.772±0.078 | 1.418±0.334 | -1.851±0.186 | -1.851 (-2.147, -1.554) | 0.004 | |
4 | V9HWE3 | CA1 | Carbonic anhydrase I | 1.015±0.413 | 0.667±0.292 | 1.002±1.586 | 1.002 (-1.521, 3.525) | 0.016 | |
5 | V9HW21 | CA2 | Carbonic anhydrase II | 1.366±0.138 | 0.901±0.071 | 1.516±0.153 | 1.516 (1.136, 1.896) | 0.022 | |
6 | A0A087WXI2 | FCGBP | Fc fragment of igg binding protein | 1.076±0.095 | 0.801±0.180 | 1.343±0.118 | 1.343 (1.155, 1.531) | 0.047 | |
7 | P02671 | FGA | Fibrinogen alpha chain | 1.151±0.144 | 0.694±0.114 | 1.658±0.208 | 1.658 (1.327, 1.989) | 0.043 | |
8 | V9HVY1 | FGB | Fibrinogen beta chain | 1.069±0.042 | 0.719±0.145 | 1.487±0.058 | 1.487 (1.394, 1.580) | 0.042 | |
9 | V9HWI6 | GC | Group-specific component (vitamin D binding protein) | 0.943±0.155 | 1.553±0.497 | -1.688±0.327 | -1.688 (-2.209, -1.167) | 0.065 | |
10 | A8K335 | GGH | Gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) | 1.093±0.232 | 0.723±0.138 | 1.513±0.321 | 1.513 (0.716, 2.310) | 0.054 | |
11 | P06396 | GSN | Gelsolin | 0.974±0.092 | 1.367±0.326 | -1.414±0.134 | -1.414 (-1.628, -1.200) | 0.049 | |
12 | D9YZU5 | HBB | Hemoglobin, beta | 1.201±0.741 | 0.511±0.205 | 1.782±2.468 | 1.782 (-2.145, 5.709) | 0.056 | |
13 | Q7Z379 | IGH | Immunoglobulin heavy locus | 0.866±0.424 | 1.889±0.614 | -2.504±0.998 | -2.504 (-4.982, -0.026) | 0.067 | |
14 | P18428 | LBP | Lipopolysaccharide binding protein | 1.108±0.258 | 0.610±0.167 | 1.583±0.369 | 1.583 (0.996, 2.170) | 0.067 | |
15 | V9HWF6 | ORM1 | Orosomucoid 1 | 0.837±0.097 | 0.472±0.121 | 1.774±0.206 | 1.774 (1.446, 2.102) | 0.031 | |
16 | P07737 | PFN1 | Profilin 1 | 1.276±0.326 | 0.758±0.238 | 1.682±0.410 | 1.682 (0.998, 2.366) | 0.022 | |
17 | D3JV41 | PPBP | Pro-platelet basic protein (chemokine (C-X-C motif) ligand 7) | 1.433±0.508 | 0.627±0.095 | 2.284±0.810 | 2.284 (0.995, 3.574) | 0.041 | |
18 | B4DF70 | PRDX2 | Peroxiredoxin 2 | 1.423±0.209 | 0.735±0.111 | 1.937±0.285 | 1.937 (1.230, 2.645) | 0.009 | |
19 | A8K2W3 | SDPR | Serum deprivation response | 1.066±0.332 | 0.449±0.068 | 2.377±0.741 | 2.377 (0.538, 4.217) | 0.053 | |
20 | A2VCK8 | TMSB10/TMSB4X | Thymosin beta 10 | 1.762±0.221 | 0.634±0.262 | 2.778±0.349 | 2.778 (2.223, 3.333) | 0.029 | |
21 | A0A024QZN4 | VCL | Vinculin | 1.369±0.125 | 0.852±0.141 | 1.607±0.146 | 1.607 (1.375, 1.840) | 0.015 |
Table 2 List of proteins with different abundance in stasis-heat patients compared with health group ($\bar{x}±s$)
No. | UniProt accession | Symbol | Entrez gene name | Abundance in SHP | Abundance in HG | Fold change | 95%CI for fold change | P value | |
---|---|---|---|---|---|---|---|---|---|
1 | V9HWD8 | A1BG | Alpha-1-B glycoprotein | 1.678±0.102 | 1.127±0.359 | 1.490±0.091 | 1.490 (1.346, 1.634) | 0.034 | |
2 | A0A024R694 | ACTN1 | Actinin, alpha 1 | 1.408±0.086 | 0.680±0.238 | 2.069±0.126 | 2.069 (0.939, 3.200) | 0.051 | |
3 | P06727 | APOA4 | Apolipoprotein A-IV | 0.772±0.078 | 1.418±0.334 | -1.851±0.186 | -1.851 (-2.147, -1.554) | 0.004 | |
4 | V9HWE3 | CA1 | Carbonic anhydrase I | 1.015±0.413 | 0.667±0.292 | 1.002±1.586 | 1.002 (-1.521, 3.525) | 0.016 | |
5 | V9HW21 | CA2 | Carbonic anhydrase II | 1.366±0.138 | 0.901±0.071 | 1.516±0.153 | 1.516 (1.136, 1.896) | 0.022 | |
6 | A0A087WXI2 | FCGBP | Fc fragment of igg binding protein | 1.076±0.095 | 0.801±0.180 | 1.343±0.118 | 1.343 (1.155, 1.531) | 0.047 | |
7 | P02671 | FGA | Fibrinogen alpha chain | 1.151±0.144 | 0.694±0.114 | 1.658±0.208 | 1.658 (1.327, 1.989) | 0.043 | |
8 | V9HVY1 | FGB | Fibrinogen beta chain | 1.069±0.042 | 0.719±0.145 | 1.487±0.058 | 1.487 (1.394, 1.580) | 0.042 | |
9 | V9HWI6 | GC | Group-specific component (vitamin D binding protein) | 0.943±0.155 | 1.553±0.497 | -1.688±0.327 | -1.688 (-2.209, -1.167) | 0.065 | |
10 | A8K335 | GGH | Gamma-glutamyl hydrolase (conjugase, folylpolygammaglutamyl hydrolase) | 1.093±0.232 | 0.723±0.138 | 1.513±0.321 | 1.513 (0.716, 2.310) | 0.054 | |
11 | P06396 | GSN | Gelsolin | 0.974±0.092 | 1.367±0.326 | -1.414±0.134 | -1.414 (-1.628, -1.200) | 0.049 | |
12 | D9YZU5 | HBB | Hemoglobin, beta | 1.201±0.741 | 0.511±0.205 | 1.782±2.468 | 1.782 (-2.145, 5.709) | 0.056 | |
13 | Q7Z379 | IGH | Immunoglobulin heavy locus | 0.866±0.424 | 1.889±0.614 | -2.504±0.998 | -2.504 (-4.982, -0.026) | 0.067 | |
14 | P18428 | LBP | Lipopolysaccharide binding protein | 1.108±0.258 | 0.610±0.167 | 1.583±0.369 | 1.583 (0.996, 2.170) | 0.067 | |
15 | V9HWF6 | ORM1 | Orosomucoid 1 | 0.837±0.097 | 0.472±0.121 | 1.774±0.206 | 1.774 (1.446, 2.102) | 0.031 | |
16 | P07737 | PFN1 | Profilin 1 | 1.276±0.326 | 0.758±0.238 | 1.682±0.410 | 1.682 (0.998, 2.366) | 0.022 | |
17 | D3JV41 | PPBP | Pro-platelet basic protein (chemokine (C-X-C motif) ligand 7) | 1.433±0.508 | 0.627±0.095 | 2.284±0.810 | 2.284 (0.995, 3.574) | 0.041 | |
18 | B4DF70 | PRDX2 | Peroxiredoxin 2 | 1.423±0.209 | 0.735±0.111 | 1.937±0.285 | 1.937 (1.230, 2.645) | 0.009 | |
19 | A8K2W3 | SDPR | Serum deprivation response | 1.066±0.332 | 0.449±0.068 | 2.377±0.741 | 2.377 (0.538, 4.217) | 0.053 | |
20 | A2VCK8 | TMSB10/TMSB4X | Thymosin beta 10 | 1.762±0.221 | 0.634±0.262 | 2.778±0.349 | 2.778 (2.223, 3.333) | 0.029 | |
21 | A0A024QZN4 | VCL | Vinculin | 1.369±0.125 | 0.852±0.141 | 1.607±0.146 | 1.607 (1.375, 1.840) | 0.015 |
No. | UniProt accession | Symbol | Entrez gene name | Abundance in nonSHP | Abundance in HG | Fold change | 95%CI for fold change | P value |
---|---|---|---|---|---|---|---|---|
1 | P06727 | APOA4 | Apolipoprotein A-IV | 0.820±0.152 | 1.418±0.334 | –1.781±0.386 | –1.77(–2.39,–1.16) | 0.006 |
2 | V9HW21 | CA2 | Carbonic anhydrase II | 1.225±0.267 | 0.901±0.070 | 1.360±0.296 | 1.35(0.62,2.09) | 0.073 |
3 | H0YCV9 | CD44 | CD44 molecule (Indian blood group) | 1.183±0.641 | 0.944±0.671 | 1.252±0.679 | 1.25(0.17,2.33) | 0.003 |
4 | L8EB32 | COL14A1 | Collagen, type XIV, alpha 1 | 0.994±0.138 | 2.573±0.453 | –2.614±0.365 | –2.61(–5.89,0.66) | 0.087 |
5 | Q53S54 | CUL3 | Cullin 3 | 1.406±0.472 | 0.496±0.144 | 2.831±0.950 | 2.82(0.46,5.19) | 0.063 |
6 | V9HWI6 | GC | Group-specific component (vitamin D binding protein) | 0.980±0.300 | 1.552±0.496 | –1.679±0.422 | –1.67(–2.35,–1.00) | 0.083 |
7 | P06396 | GSN | Gelsolin | 0.919±0.105 | 1.367±0.325 | –1.501±0.176 | –1.50(–1.78,–1.22) | 0.029 |
8 | Q6ZW64 | IGH | Immunoglobulin heavy locus | 1.216±0.404 | 0.528±0.238 | 2.304±0.765 | 2.30(1.08,3.52) | 0.064 |
9 | A0A087X0P6 | IGKV2D-29 | Immunoglobulin kappa variable 2D-29 | 0.659±0.188 | 1.133±0.032 | –1.791±0.512 | –1.79(–6.39,2.81) | 0.081 |
10 | P18428 | LBP | Lipopolysaccharide binding protein | 1.252±0.377 | 0.699±0.167 | 1.789±0.539 | 1.78(0.93,2.64) | 0.018 |
11 | V9HWF6 | ORM1 | Orosomucoid 1 | 0.985±0.323 | 0.471±0.120 | 2.090±0.686 | 2.08(0.99,3.18) | 0.006 |
Table 3 List of proteins with different abundance in non-stasis heat patients compared with health group ($\bar{x}±s$)
No. | UniProt accession | Symbol | Entrez gene name | Abundance in nonSHP | Abundance in HG | Fold change | 95%CI for fold change | P value |
---|---|---|---|---|---|---|---|---|
1 | P06727 | APOA4 | Apolipoprotein A-IV | 0.820±0.152 | 1.418±0.334 | –1.781±0.386 | –1.77(–2.39,–1.16) | 0.006 |
2 | V9HW21 | CA2 | Carbonic anhydrase II | 1.225±0.267 | 0.901±0.070 | 1.360±0.296 | 1.35(0.62,2.09) | 0.073 |
3 | H0YCV9 | CD44 | CD44 molecule (Indian blood group) | 1.183±0.641 | 0.944±0.671 | 1.252±0.679 | 1.25(0.17,2.33) | 0.003 |
4 | L8EB32 | COL14A1 | Collagen, type XIV, alpha 1 | 0.994±0.138 | 2.573±0.453 | –2.614±0.365 | –2.61(–5.89,0.66) | 0.087 |
5 | Q53S54 | CUL3 | Cullin 3 | 1.406±0.472 | 0.496±0.144 | 2.831±0.950 | 2.82(0.46,5.19) | 0.063 |
6 | V9HWI6 | GC | Group-specific component (vitamin D binding protein) | 0.980±0.300 | 1.552±0.496 | –1.679±0.422 | –1.67(–2.35,–1.00) | 0.083 |
7 | P06396 | GSN | Gelsolin | 0.919±0.105 | 1.367±0.325 | –1.501±0.176 | –1.50(–1.78,–1.22) | 0.029 |
8 | Q6ZW64 | IGH | Immunoglobulin heavy locus | 1.216±0.404 | 0.528±0.238 | 2.304±0.765 | 2.30(1.08,3.52) | 0.064 |
9 | A0A087X0P6 | IGKV2D-29 | Immunoglobulin kappa variable 2D-29 | 0.659±0.188 | 1.133±0.032 | –1.791±0.512 | –1.79(–6.39,2.81) | 0.081 |
10 | P18428 | LBP | Lipopolysaccharide binding protein | 1.252±0.377 | 0.699±0.167 | 1.789±0.539 | 1.78(0.93,2.64) | 0.018 |
11 | V9HWF6 | ORM1 | Orosomucoid 1 | 0.985±0.323 | 0.471±0.120 | 2.090±0.686 | 2.08(0.99,3.18) | 0.006 |
No. | UniProt accession | Symbol | Entrez gene name | Abundance in SHP | Abundance in nonSHP | Fold change | 95%CI for fold change | P value | |
---|---|---|---|---|---|---|---|---|---|
1 | V9HWD8 | A1BG | Alpha-1-B glycoprotein | 1.678±0.102 | 1.017±0.130 | 1.648±0.100 | 1.64(1.48,1.80) | 0.015 | |
2 | A0A024R694 | ACTN1 | Actinin, alpha 1 | 1.407±0.085 | 0.750±0.060 | 1.876±0.114 | 1.87(0.85,2.90) | 0.062 | |
3 | V9HWE3 | CA1 | Carbonic anhydrase I | 1.015±0.412 | 0.761±0.232 | 1.333±0.542 | 1.33(0.47,2.19) | 0.061 | |
4 | H0YCV9 | CD44 | CD44 molecule (Indian blood group) | 1.062±0.615 | 1.183±0.641 | –1.149±1.375 | –1.15(–3.33,1.03) | 0.074 | |
5 | A5PL27 | CP | Ceruloplasmin (ferroxidase) | 1.308±0.115 | 0.941±0.080 | 1.390±0.123 | 1.38(1.19,1.58) | 0.046 | |
6 | B4DF70 | PRDX2 | Peroxiredoxin 2 | 1.422±0.209 | 0.921±0.136 | 1.543±0.226 | 1.54(0.98,2.10) | 0.029 | |
7 | A0A024QZN4 | VCL | Vinculin | 1.369±0.124 | 1.025±0.242 | 1.334±0.121 | 1.33(1.14,1.52) | 0.087 |
Table 4 List of proteins with different abundance in stasis-heat patients compared with non-stasis-heat patients ($\bar{x}±s$)
No. | UniProt accession | Symbol | Entrez gene name | Abundance in SHP | Abundance in nonSHP | Fold change | 95%CI for fold change | P value | |
---|---|---|---|---|---|---|---|---|---|
1 | V9HWD8 | A1BG | Alpha-1-B glycoprotein | 1.678±0.102 | 1.017±0.130 | 1.648±0.100 | 1.64(1.48,1.80) | 0.015 | |
2 | A0A024R694 | ACTN1 | Actinin, alpha 1 | 1.407±0.085 | 0.750±0.060 | 1.876±0.114 | 1.87(0.85,2.90) | 0.062 | |
3 | V9HWE3 | CA1 | Carbonic anhydrase I | 1.015±0.412 | 0.761±0.232 | 1.333±0.542 | 1.33(0.47,2.19) | 0.061 | |
4 | H0YCV9 | CD44 | CD44 molecule (Indian blood group) | 1.062±0.615 | 1.183±0.641 | –1.149±1.375 | –1.15(–3.33,1.03) | 0.074 | |
5 | A5PL27 | CP | Ceruloplasmin (ferroxidase) | 1.308±0.115 | 0.941±0.080 | 1.390±0.123 | 1.38(1.19,1.58) | 0.046 | |
6 | B4DF70 | PRDX2 | Peroxiredoxin 2 | 1.422±0.209 | 0.921±0.136 | 1.543±0.226 | 1.54(0.98,2.10) | 0.029 | |
7 | A0A024QZN4 | VCL | Vinculin | 1.369±0.124 | 1.025±0.242 | 1.334±0.121 | 1.33(1.14,1.52) | 0.087 |
Figure 2 Comparison of differential proteins among stasis-heat patients, non-stasis-heat patients, and health groups A: venn diagram showing the differential proteins among the three groups; B: bar graph showing the expression levels of proteins between stasis-heat and non-stasis-heat group. A1BG: Alpha-1-B glycoprotein; CP: copper-protein; ACTN1: alpha 1; CA1: carbonic anhydrase I; PRDX2: peroxiredoxin 2; VCL: vinculin; SH: stasis-heat; NSH: non-stasis-heat; HC: health control.
Figure 3 Western blotting analysis of A1BG, ACTN1, CA1, CP, PRDX1, VCL1, and CD44 from healthy controls, stasis-heat group, and non-stasis-heat group A: Western blotting analysis of the seven proteins; B: the intensity of each band was measured using imaging analysis. aP ≤ 0.05, comparison of non-stasis-heat group and healthy group with stasis-heat group separately. A1BG: Alpha-1-B glycoprotein; CP: copper-protein; ACTN1: alpha 1; CA1: carbonic anhydrase I; PRDX2: peroxiredoxin 2; VCL: vinculin.
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