1. Raw data VS. Residual valueB
1898
766 44 C
2169
495 80
Supporting Fig. 1. Neither age nor gender were major drivers of expression differences between the control and NAFLD groups. (A) The cumulative curve of coefficient of variation of gene expression. (B) Overlap between differentially expressed genes using either the residual values from age or the raw data. (C) Overlap between differentially expressed genes using either the residual values from gender or the raw data.

2. Scaled expression values
Age (years)
Supporting Fig. 2. Gene expression in NAFLD. Heat map of top 200 genes variant expressed between control, NAFL and NAFLD samples. Scaled expression values are color-coded according to the legend on the left. The top bar shows the disease status: red, NASH; green, NAFL; black, control. The bottom bars show additional variables for each sample: age and gender (black, female; grey, male; green, missing value). The scale for age is shown on the left.

3. Color key
Supporting Fig. 3. Hierarchical clustering of microarray samples based on inter-array correlation showing distinct clustering of NASH, NAFL and control samples. The heatmap shown the square inter-array correlation matrix(IAC). IAC coefficients were calculated using Pearson correlation. The scale for IAC was shown on the left. The upper bar show age for each sample (green, female; blue, male; white, missing value). The scale for age is shown on the left side(white, missing value).

4. Control NASH A Control NASH B -0.2 0.0 0.2 0.4 Eigengene expression
Eigengene expression
Control
NASH B
Eigengene expression

5. Control
NASH C
Eigengene expression
Supporting Fig. 4. Expression within NASH modules N1, N8 and N14 is shown in the heat-map and summarized with the module. The visualization of these modules was performed using VisANT to plot the 100 strongest connections within each module. Genes that are positively correlated are connected by red lines, whereas genes that are inversely correlated are connected by blue lines. (A) The module N1 is involved in chromosome organization. (B) The module N8 contains genes participate in protein degradation. (C) The module N14 contains genes involved in immune response.

6. NAFL NASH A NAFL NASH B -0.4 0.2 0.0 0.2 Eigengene expression
Eigengene expression
NAFL
NASH B
Eigengene expression

7. NAFL NASH C NAFL NASH D Eigengene expression -0.4 0.2 0.0 0.2
Eigengene expression
NAFL
NASH D
Eigengene expression

8. NAFL NASH E NAFL NASH F -0.2 0.0 0.2 0.4 Eigengene expression
Eigengene expression
NAFL
NASH F
Eigengene expression

9. NAFL
NASH G
Eigengene expression
Supporting Fig. 5. Expression within progression modules P1(A), P2(B), P5(C), P7(D), P9(E), P13(F) and P14(G) is shown in the heat-map and summarized with the module. The visualization of these modules was performed using VisANT to plot the 100 strongest connections within each module. Genes that are positively correlated are connected by red lines, whereas genes that are inversely correlated are connected by blue lines.

10. Supporting Fig. 6. To test the stability of modules, intramodular connectivity in 1000 module gene sets generated by sampling 13 of the 26 array samples were calculated for each module. Then, correlations between the true module gene connectivity values and those from the 1000 randomly selected sets were calculated.

11. Module-miRNA relationships-1
-0.5
0.5 1
MIR24-2
MIR122
MIR27B
MIR148B
MIR30E
MIRLET7G
MIRLET7D
MIR192
MIR101-2
MIR24-1
MIRLET7F1
MIR29C
MIR23B
MIR105-2
MIR15A
-0.52
(0.006)
0.45
(0.02)
0.3
(0.1)
0.27
(0.2)
0.29
0.38
(0.06)
0.43
(0.03)
0.42
0.33
0.16
(0.4)
0.11
(0.6)
0.19
0.26
-0.04
(0.8)
0.52
0.53
0.51
(0.008)
0.46
0.49
(0.01)
0.2
(0.3)
0.32
0.44
0.41
(0.04)
0.35
(0.08)
0.1
-0.058
-0.1
0.062
-0.21
0.067
(0.7)
-0.019
(0.9)
-0.2
-0.082
-0.18
0.12
0.024
0.21
-0.56
(0.003)
0.73
(2e-05)
0.67
(2e-04)
0.7
(7e-05)
0.59
(0.002)
0.62
(7e-04)
0.36
(0.07)
0.6
(0.001)
0.37
0.47
(0.005)
0.39
(0.05)
0.31
0.17
0.56
0.34
(0.09)
0.078
0.55
-0.087
-0.17
-0.36
-0.072
-0.022
-0.064
0.053
-0.11
-0.43
0.086
-0.55
(0.004)
-0.44
-0.49
-0.57
-0.26
-0.24
-0.32
-0.37
-0.16
-0.27
-0.25
0.65
(3e-04)
-0.64
(5e-04)
-0.48
-0.61
(9e-04)
-0.53
-0.82
(3e-07)
-0.35
-0.23
-0.3
-0.51
(6e-05)
-0.33
-0.19
-0.63
-0.28
-0.061
0.019
-0.071
-0.033
-0.043
0.001
(1)
0.14
(0.5)
0.24
0.22
0.28
0.07
0.23
0.054
0.0075
0.0083
0.049
-0.29
-0.41
0.0033
-0.22
-0.39
-0.079
-0.12
0.048
-0.13
-0.011
-0.31
0.021
0.13
-0.096
0.68
(1e-04)
-0.4
-0.38
(4e-04)
-0.037
-0.34
(0.007)
0.4
-0.14
-0.085
0.02 P1 P2 P3 P4 P5 P6 P7 P8 P9
P10
P11
P12
P13
P14
Supporting Fig. 7. Module-miRNA associations in progression network. Each row corresponds to a module eigengene, column to a miRNA. Each cell contains the corresponding correlation and P-value (bracket). The table is color-coded by correlation according to the color legend.

12. A B C D

13. E F G H

14. I J K L

15. M
Supporting Fig. 8. The individual gene expression of the candidates between control, NAFL and NASH groups were shown. MT1DP(A), MT1X(B), PDIA6(C), SRPRB(D), TCF4(E), RPL8(F), IGHG1(G), NANS(H), VIM(I), PTGS2(J), PAN3(K), YIF1A(L) and SRRM2(M)

16. ** A 37 21 55 ** 16 21 70 B C ** 12 2 D ** 93 19 7 NAFL NASH
GO category/KEGG pathway S1 N5
Sterol biosynthetic process
6.3E-29
3.4E-27
Lipid biosynthetic process
2.8E-20
8.9E-24
Endoplasmic reticulum
3.4E-4
5.8E-6
Microsome
6.2E-4
8.0E-5 ** A 37 21 55
GO category/KEGG pathway S2 N6
Oxidation reduction
9.9E-8
8.6E-15
Microsome
2.1E-13
2.4E-13
Endoplasmic reticulum
1.5E-8
3.8E-8
Drug metabolism
1.4E-20
1.1E-16 ** 16 21 70 B
GO category/KEGG pathway S4 N7
Ion homeostasis
2.6E-2
8.0E-2
Cadmium ion binding
3.5E-16
3.3E-19
Copper ion binding
2.8E-13
4.1E-15
Zinc ion binding
4.1E-4
2.8E-4 C ** 12 2 D **
GO category/KEGG pathway S6
N13
M phase
1.5E-56
1.4E-52
Spindle
3.6E-30
1.4E-26
Chromosome
2.2E-23
6.1E-25
P53 signaling pathway
2.1E-2
1.4E-3 93 19 7

17. GO category/KEGG pathwayS9
N11
RNA processing
2.4E-11
7.8E-4
DNA repair
8.8E-5 NS
Spliceosome
4.1E-6
Ubiquitin mediated proteolysis
7.7E-3 E * 20
892 61
GO category/KEGG pathway
S10
N14
Innate immune response
1.9E-7
9.6E-10
Adaptive immune response
1.9E-5
2.3E-5
MHC class II receptor activity
1.2E-2 NS
Cell adhesion molecules (CAMs)
6.5E-3
1.1E-4 F ** 70 42 43
GO category/KEGG pathway
S13 N8
Negative regulation of protein ubiquitination
2.8E-19
1.0E-11
Positive regulation of protein ubiquitination
8.2E-18
9.4E-12
Regulation of protein modification process
7.3E-7
4.9E-5
Proteasome
6.7E-21
6.3E-16 G ** 77
424
392
Supporting Fig. 9. Modules from NAFL analysis overlap significantly with modules from NASH analysis, as measured by both gene number and gene ontology categories enrichment P value from DAVID. *P<0.001; **P<10-9. P values were obtained using a hypergeometric distribution.