Chinese Journal of Chromatography ›› 2021, Vol. 39 ›› Issue (11): 1191-1202.DOI: 10.3724/SP.J.1123.2021.04009
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YANG Kaige, WANG Weiwei, WANG Yan*(), YAN Chao*(
)
Received:
2021-04-08
Online:
2021-11-08
Published:
2021-10-22
Contact:
WANG Yan,YAN Chao
Supported by:
Fig. 1 Characterization of serum exosomes a. size range of healthy control (HC) serum exosomes measured using nanoparticle tracking analysis (NTA). b. size range of intrahepatic cholangiocarcinoma (iCCA) serum exosomes measured using NTA. c. Western blotting analysis of CD81, CD63 and CD9 in the serum exosomes and serum (HC and iCCA). d. TEM image of HC serum exosomes. e. TEM image of iCCA serum exosomes. White arrow: vesicle.
Fig. 2 Proteome analysis of serum exosomes a. SDS-PAGE analysis of proteins of the serum exosomes and serum (HC and iCCA). b. Venn diagram of the identified proteins of the serum exosomes by LC-MS and ExoCarta database.
Fig. 4 Multidimensional statistical analysis based on the normalized quantitative proteome a. principal components analysis (PCA) based on the normalized quantitative proteome of HC and iCCA serum; R2X[1]=0.24, R2X[2]=0.17. b. PCA based on the normalized quantitative proteome of HC and iCCA serum exosomes; R2X[1]=0.26, R2X[2]=0.17. c. orthogonal partial least-squares discrimination analysis (OPLS-DA) based on the normalized quantitative proteome of HC and iCCA serum; R2X[1]=0.40, R2Xo[1]=0.11. d. OPLS-DA based on the normalized quantitative proteome of HC and iCCA serum exosomes; R2X[1]=0.45, R2Xo[1]=0.10. Ellipse: Hotelling’s T2 (95%).
Sample type | A | R2X(cum) | R2Y(cum) | Q2(cum) |
---|---|---|---|---|
Serum | 1P+1O | 0.509 | 0.999 | 0.969 |
Exosome | 1P+1O | 0.547 | 0.999 | 0.984 |
Table 1 Parameters of OPLS-DA models based on the normalized quantitative proteome of serum and serum exosomes
Sample type | A | R2X(cum) | R2Y(cum) | Q2(cum) |
---|---|---|---|---|
Serum | 1P+1O | 0.509 | 0.999 | 0.969 |
Exosome | 1P+1O | 0.547 | 0.999 | 0.984 |
Serum | Exosome | |||||||
---|---|---|---|---|---|---|---|---|
Accession | Gene symbol | FC(iCCA/HC) | P-value | Accession | Gene symbol | FC(iCCA/HC) | P-value | |
P00450 | CP | 2.2429 | 0.0053 | P04275 | VWF | 2.1460 | 0.0383 | |
P06681 | C2 | 2.2294 | 0.0001 | P01009 | SERPINA1 | 2.4817 | 0.0001 | |
P01833 | PIGR | 12.4548 | 0.0091 | P01877 | IGHA2 | +∞ | 0.0019 | |
Q06033 | ITIH3 | 2.5237 | 0.0086 | P01833 | PIGR | 9.5927 | 0.0048 | |
P36980 | CFHR2 | 3.9305 | 0.0371 | P20742 | PZP | 3.7771 | 0.0034 | |
P00742 | F10 | 2.2807 | 0.0001 | P36980 | CFHR2 | 2.1296 | 0.0109 | |
Q04756 | HGFAC | 2.5523 | 0.0024 | P01011 | SERPINA3 | 2.3901 | 0.0002 | |
P05362 | ICAM1 | +∞ | 0.0004 | P02763 | ORM1 | 2.3329 | 0.0028 | |
Q9BYE9 | CDHR2 | +∞ | 0.0005 | P15144 | ANPEP | 2.0323 | 0.0167 | |
A0A0A0MRZ9 | IGLV5-52 | 2.1752 | 0.0147 | P68133 | ACTA1 | 2.1930 | 0.0357 | |
P19320 | VCAM1 | 3.1375 | 0.0206 | P02775 | PPBP | 2.0735 | 0.0422 | |
P13473 | LAMP2 | 2.1709 | 0.0220 | Q04695 | KRT17 | +∞ | 0.0016 | |
Q9BT22 | ALG1 | 2.0182 | 0.0281 | Q14766 | LTBP1 | 2.0226 | 0.0274 | |
Q8NFU5 | IPMK | 12.1589 | 0.0006 | Q6UWP8 | SBSN | 4.0805 | 0.0177 | |
A2VCL2 | CCDC162 | 2.4897 | 0.0018 | P07327 | ADH1A | +∞ | 0.0052 | |
P06727 | APOA4 | 0.4316 | 0.0030 | P22792 | CPN2 | 2.8882 | 0.0049 | |
P07996 | THBS1 | 0.4055 | 0.0021 | P35555 | FBN1 | 2.4629 | 0.0282 | |
A0A075B6S6 | IGKV2D-30 | 0.0000 | 0.0288 | P02750 | LRG1 | 2.4234 | 0.0016 | |
P01602 | IGKV1-5 | 0.0000 | 0.0006 | P20930 | FLG | +∞ | 0.0165 | |
P13647 | KRT5 | 0.3018 | 0.0392 | P08571 | CD14 | 3.7509 | 0.0234 | |
Q7Z794 | KRT77 | 0.3075 | 0.0094 | Q9UEW3 | MARCO | 2.4943 | 0.0434 | |
P67936 | TPM4 | 0.0000 | 0.0110 | P02792 | FTL | +∞ | 0.0000 | |
O60229 | KALRN | 0.4881 | 0.0039 | P21333 | FLNA | 2.0853 | 0.0431 | |
P28066 | PSMA5 | +∞ | 0.0000 | |||||
P12814 | ACTN1 | 3.5309 | 0.0364 | |||||
Q9UGM3 | DMBT1 | 2.4154 | 0.0404 | |||||
Q9H4G4 | GLIPR2 | 3.8794 | 0.0397 | |||||
P78509 | RELN | 2.6213 | 0.0149 | |||||
Q15063 | POSTN | +∞ | 0.0000 | |||||
P30101 | PDIA3 | +∞ | 0.0070 | |||||
O14818 | PSMA7 | 2.2647 | 0.0024 | |||||
Q7Z398 | ZNF550 | 2.8163 | 0.0197 | |||||
O95810 | CAVIN2 | +∞ | 0.0001 | |||||
P01861 | IGHG4 | 0.3768 | 0.0009 | |||||
P06396 | GSN | 0.4789 | 0.0032 | |||||
P06727 | APOA4 | 0.2324 | 0.0000 | |||||
Q16610 | ECM1 | 0.2677 | 0.0001 | |||||
P04070 | PROC | 0.2334 | 0.0437 | |||||
P35443 | THBS4 | 0.3062 | 0.0060 | |||||
A0A075B6R2 | IGHV4-4 | 0.3530 | 0.0112 | |||||
P22105 | TNXB | 0.3142 | 0.0009 | |||||
A0A0B4J2H0 | IGHV1-69D | 0.1628 | 0.0016 | |||||
Q04756 | HGFAC | 0.3811 | 0.0005 | |||||
Q14532 | KRT32 | 0.0000 | 0.0007 | |||||
A0A0G2JMI3 | IGHV1-69-2 | 0.3718 | 0.0104 | |||||
A0A087WSZ0 | IGKV1D-8 | 0.0000 | 0.0002 | |||||
P12109 | COL6A1 | 0.4603 | 0.0094 | |||||
Q68EA5 | ZNF57 | 0.4689 | 0.0425 | |||||
Q96MV8 | ZDHHC15 | 0.3387 | 0.0171 | |||||
Q13939 | CCIN | 0.3101 | 0.0093 | |||||
Q9BS31 | ZNF649 | 0.0000 | 0.0021 |
Table 2 Differential proteins between HC and iCCA groups based on the serum and serum exosome samples
Serum | Exosome | |||||||
---|---|---|---|---|---|---|---|---|
Accession | Gene symbol | FC(iCCA/HC) | P-value | Accession | Gene symbol | FC(iCCA/HC) | P-value | |
P00450 | CP | 2.2429 | 0.0053 | P04275 | VWF | 2.1460 | 0.0383 | |
P06681 | C2 | 2.2294 | 0.0001 | P01009 | SERPINA1 | 2.4817 | 0.0001 | |
P01833 | PIGR | 12.4548 | 0.0091 | P01877 | IGHA2 | +∞ | 0.0019 | |
Q06033 | ITIH3 | 2.5237 | 0.0086 | P01833 | PIGR | 9.5927 | 0.0048 | |
P36980 | CFHR2 | 3.9305 | 0.0371 | P20742 | PZP | 3.7771 | 0.0034 | |
P00742 | F10 | 2.2807 | 0.0001 | P36980 | CFHR2 | 2.1296 | 0.0109 | |
Q04756 | HGFAC | 2.5523 | 0.0024 | P01011 | SERPINA3 | 2.3901 | 0.0002 | |
P05362 | ICAM1 | +∞ | 0.0004 | P02763 | ORM1 | 2.3329 | 0.0028 | |
Q9BYE9 | CDHR2 | +∞ | 0.0005 | P15144 | ANPEP | 2.0323 | 0.0167 | |
A0A0A0MRZ9 | IGLV5-52 | 2.1752 | 0.0147 | P68133 | ACTA1 | 2.1930 | 0.0357 | |
P19320 | VCAM1 | 3.1375 | 0.0206 | P02775 | PPBP | 2.0735 | 0.0422 | |
P13473 | LAMP2 | 2.1709 | 0.0220 | Q04695 | KRT17 | +∞ | 0.0016 | |
Q9BT22 | ALG1 | 2.0182 | 0.0281 | Q14766 | LTBP1 | 2.0226 | 0.0274 | |
Q8NFU5 | IPMK | 12.1589 | 0.0006 | Q6UWP8 | SBSN | 4.0805 | 0.0177 | |
A2VCL2 | CCDC162 | 2.4897 | 0.0018 | P07327 | ADH1A | +∞ | 0.0052 | |
P06727 | APOA4 | 0.4316 | 0.0030 | P22792 | CPN2 | 2.8882 | 0.0049 | |
P07996 | THBS1 | 0.4055 | 0.0021 | P35555 | FBN1 | 2.4629 | 0.0282 | |
A0A075B6S6 | IGKV2D-30 | 0.0000 | 0.0288 | P02750 | LRG1 | 2.4234 | 0.0016 | |
P01602 | IGKV1-5 | 0.0000 | 0.0006 | P20930 | FLG | +∞ | 0.0165 | |
P13647 | KRT5 | 0.3018 | 0.0392 | P08571 | CD14 | 3.7509 | 0.0234 | |
Q7Z794 | KRT77 | 0.3075 | 0.0094 | Q9UEW3 | MARCO | 2.4943 | 0.0434 | |
P67936 | TPM4 | 0.0000 | 0.0110 | P02792 | FTL | +∞ | 0.0000 | |
O60229 | KALRN | 0.4881 | 0.0039 | P21333 | FLNA | 2.0853 | 0.0431 | |
P28066 | PSMA5 | +∞ | 0.0000 | |||||
P12814 | ACTN1 | 3.5309 | 0.0364 | |||||
Q9UGM3 | DMBT1 | 2.4154 | 0.0404 | |||||
Q9H4G4 | GLIPR2 | 3.8794 | 0.0397 | |||||
P78509 | RELN | 2.6213 | 0.0149 | |||||
Q15063 | POSTN | +∞ | 0.0000 | |||||
P30101 | PDIA3 | +∞ | 0.0070 | |||||
O14818 | PSMA7 | 2.2647 | 0.0024 | |||||
Q7Z398 | ZNF550 | 2.8163 | 0.0197 | |||||
O95810 | CAVIN2 | +∞ | 0.0001 | |||||
P01861 | IGHG4 | 0.3768 | 0.0009 | |||||
P06396 | GSN | 0.4789 | 0.0032 | |||||
P06727 | APOA4 | 0.2324 | 0.0000 | |||||
Q16610 | ECM1 | 0.2677 | 0.0001 | |||||
P04070 | PROC | 0.2334 | 0.0437 | |||||
P35443 | THBS4 | 0.3062 | 0.0060 | |||||
A0A075B6R2 | IGHV4-4 | 0.3530 | 0.0112 | |||||
P22105 | TNXB | 0.3142 | 0.0009 | |||||
A0A0B4J2H0 | IGHV1-69D | 0.1628 | 0.0016 | |||||
Q04756 | HGFAC | 0.3811 | 0.0005 | |||||
Q14532 | KRT32 | 0.0000 | 0.0007 | |||||
A0A0G2JMI3 | IGHV1-69-2 | 0.3718 | 0.0104 | |||||
A0A087WSZ0 | IGKV1D-8 | 0.0000 | 0.0002 | |||||
P12109 | COL6A1 | 0.4603 | 0.0094 | |||||
Q68EA5 | ZNF57 | 0.4689 | 0.0425 | |||||
Q96MV8 | ZDHHC15 | 0.3387 | 0.0171 | |||||
Q13939 | CCIN | 0.3101 | 0.0093 | |||||
Q9BS31 | ZNF649 | 0.0000 | 0.0021 |
Fig. 5 Screening of differential proteins between HC and iCCA groups based on the serum and serum exosome samples a. volcano plots of identified proteins in the HC and iCCA groups, based on the serum samples. b. volcano plots of identified proteins in the HC and iCCA groups based on the serum exosome samples. c. Venn diagram of the differential proteins screened between HC and iCCA groups based on the serum exosome samples and ExoCarta database. d. Venn diagram of the differential proteins screened between HC and iCCA groups based on the serum and serum exosome samples.
Fig. 6 Biological information analysis of differential proteins between the HC and iCCA groups based on the serum and serum exosomes samples a. Gene Ontology (GO) analysis of differential proteins based on the serum samples; b. GO analysis of differential proteins based on the serum exosome samples; c. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of differential proteins based on the serum samples; d. KEGG pathways of differential proteins based on the serum exosome samples.
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