Quantitative proteomics analysis based on tandem mass tag labeling coupled with labeling coupled with liquid chromatography-tandem mass spectrometry discovers the effect of silibinin on non-alcoholic fatty liver disease in mice

Figures & data

Figure 1. Comparison of body weight (a), liver weight (b), food intake (c), fasting blood glucose (d), fasting insulin (e), IPGTT (f) and AUC (g) of mice in each group.

Figure 2. Comparison of liver TG content in mice of each group (a) and the effects of SIL on liver lipid deposition in HFD mice by H&E and oil red O staining (×400) (b).

Table 1. Summary of MS/MS spectrum database search analysis

Total spectrum	Matched spectrum	Peptides	Unique peptides	Identified proteins	Quantifiable proteins
324,969	53,238 (16.38%)	29,758	28,765	4565	4198

Figure 3. Experimental strategies for quantitative proteome analysis and quality control validation. (a) Pair wise Pearson’s correlation coefficients; (b) Average peptide mass error; (c) Length distribution of all identified peptides.

Table 2. List of proteins involved in sugar and lipid metabolism with at least 1.2-fold altered expression due to HFD and/or HFD+SIL

Protein accession^(a)	Protein description^(b)	MW [kDa]^(c)	Score(d)	Gene name^(e)	Coverage [%]^(f)	Peptides^(g)	Unique peptides^(h)	HFD/ND Ratio^(i)	HFD+SIL/HFD Ratio^(i)
P43883	Perilipin-2	46.646	189.700	Plin2	34.6	13	13	1.947	0.787
Q91ZP3	Phosphatidate phosphatase LPIN1	102.000	4.573	Lpin1	1.9	2	2	1.377	0.767
Q920E5	Farnesyl pyrophosphate synthase	40.581	37.954	Fdps	29.5	10	10	0.56	1.222
P13745	Glutathione S-transferase A1	25.608	5.449	Gsta1	33.6	7	4	0.476	0.738

1. Protein description, protein functional description

2. Gene name, indicates the name of the gene that code for the protein sequence

3. MW [kDa], protein molecular weight,unit [kDa]

4. Score, a simple rule to be used to judge whether a result is significant or not

5. Coverage [%], percent of identified peptdie sequence covering in protein sequence

6. Peptides, number of peptides whitch spectrum hit

7. Unique peptides, number of identified peptides that only come from this protein groups

8. The protein relative abundance ratios, HFD/ND and HFD+SIL/HFD, were calculated from the average of at least three biological replicates from each sample group; HFD/ND and HFD+SIL/HFD ratios are shown for a better comparison of the correction effect of SIL.

Figure 4. Sub-cellular functional annotations and GO analysis of identified proteins. (a) Sub-cellular localization of identified proteins; (b) GO annotation in terms of cellular component; (c) GO annotation in terms of molecular function; (d) GO annotation in terms of biological process.

Figure 5. KEGG enrichment analysis of DEPs. (a) KEGG pathway enrichment; (b) Significance rankings of KEGG pathway enrichment in HFD versus ND; The pathway of up-regulated DEPs in HFD versus ND; The pathway of down-regulated DEPs in HFD versus ND; (c) Significance rankings of KEGG pathway enrichment in HFD+SIL versus HFD; The pathway of up-regulated DEPs in HFD+SIL versus HFD; The pathway of down-regulated DEPs in HFD+SIL versus HFD.

Figure 6. The relative protein expression of PLIN2, LPIN1, FDPS and GSTA1 in liver. *P < 0.05 and **P < 0.01 vs ND, #P < 0.05 and ##P < 0.01 vs HFD.

Data availability statement

All data are available in the article.