Table 3. Pathway enrichment analysis of differentially expressed genes.
| Myoblast differentiation | |||||
| Time point | Pathway | P-value | # Genes (observed) | List of observed genes | SOTA cluster |
| 4 h | MOTHERS AGAINST DPP HOMOLOG | 1.16E-05 | 12 | Id3, Foxc2, Aspn, Smad9, Smad6, Id1, Smad7, Id2, Dlx2, Ctgf, Sp7, Serpine1 | C |
| 4 h | TGF BETA | 3.08E-05 | 15 | Fbxo32, Foxc2, Aspn, Itgb8, Dlx1, Smad9, Smad6, Id1, Smad7, Id2, Dlx2, Ctgf, Adamts5, Sp7, Serpine1 | C |
| 4 h | PARATHYROID HORMONE RELATED PROTEIN | 7.05E-04 | 5 | Igfbp5, Ctgf, Sp7, Nr4a2, Jag1 | |
| 4 h | HAIRY AND ENHANCER OF SPLIT 1 | 6.22E-03 | 3 | Dll1, Ctgf, Jag1 | |
| 4 h | RETINOBLASTOMA 1 | 6.42E-03 | 4 | Dlx1, Id2, Dlx2, Ccnd1 | |
| 4 h | VERY LOW DENSITY LIPOPROTEIN RECEPTOR | 7.48E-03 | 2 | Serpinb2, Serpine1 | C |
| 4 h | LYMPHOID ENHANCER BINDING FACTOR 1 (TCF/LEF) | 7.62E-03 | 3 | Dll1, Vdr, Ccnd1 | |
| 4 h | SEMAPHORIN | 9.75E-03 | 3 | Nrp2, Sema6a, Sema5a | A |
| 12 h | MOTHERS AGAINST DPP HOMOLOG | 2.32E-05 | 289 | Atoh8, Aspn, Smad9, Smad7, Id2, Dlx2, Cdkn1c, Timp3, Nedd9, Scn5a, Id3, Foxc2, Hfe2, Id1, Smad6, Ctgf, Sp7, Mir206 | C |
| 12 h | NOTCH | 1.96E-03 | 251 | Sox8, Dll1, Bcl6b, Dlx2, Heyl, Myc, Fabp7, Adam12, Id3, Dlx1, Id1, Mir206, Prox1 | |
| 12 h | SEMAPHORIN | 2.71E-03 | 47 | Sema6d, Sema6a, Vegfa, Sema5a, Sema3c | B |
| 12 h | CADHERIN 2, TYPE 1, N CADHERIN (NEURONAL) | 4.54E-03 | 33 | Fgfr4, Fgf2, Gja1, Itga6 | |
| 12 h | TGF BETA | 5.42E-03 | 490 | Fbxo32, Atoh8, Aspn, Smad9, Smad7, Id2, Dlx2, Adamts5, Timp3, Nedd9, Scn5a, Adam12, Foxc2, Hfe2, Dlx1, Id1, Smad6, Ctgf, Sp7 | C |
| 12 h | FIBROBLAST GROWTH FACTOR | 6.13E-03 | 254 | Bcl6b, Dlx2, Myog, Fgfr4, Fgf2, Vegfa, Ngf, Gja1, Spry1, Dlx1, Mir206, Prox1 | C |
| 12 h | NERVE GROWTH FACTOR | 7.98E-03 | 111 | Fgf2, Ngf, Alcam, Dlx1, Nefm, Id1, Ret | |
| 24 h | CYCLIN A2 | 5.87E-04 | 49 | Ncoa1, Ccnd1, Rb1, Mybl2, Cdt1, Cdkn1a, Chek1, Uhrf1 | |
| 24 h | RYANODINE RECEPTOR | 6.50E-04 | 28 | Ryr1, Casq1, Srl, Ryr3, Casq2, Trdn | B |
| 24 h | E2F TRANSCRIPTION FACTOR 1 | 8.84E-04 | 52 | Cdkn1c, Ccnd1, Myc, Rb1, Mybl2, Dusp4, Cdkn1a, Chek1 | |
| 24 h | CYCLIN E | 1.52E-03 | 44 | Cdkn1c, Ccnd1, Myc, Rb1, Cdkn1a, Chek1, Mcm3 | |
| 24 h | PEROXISOME PROLIFERATOR ACTIVATED RECEPTOR DELTA | 3.09E-03 | 26 | Rgs4, Bcl6, Nr4a2, Pla2g4a, Ptgs2 | C |
| 24 h | CYCLIN DEPENDENT KINASE | 3.73E-03 | 210 | Efna5, Ncoa1, Msln, Id2, Cdkn1c, Ccnd1, Myc, Parvb, Rb1, Mybl2, Cdt1, Myog, Cdkn1a, Hist1h1b, Chek1, Nefm, Mcm3 | |
| 24 h | NOTCH | 4.65E-03 | 251 | Sox8, Bcl6b, Heyl, Myc, Fabp7, Asb2, Id3, Adcy8, Dlx1, Id1, Mfap5, Mir206, Dll1, Dlx2, Mir23b, Smpx, Neurl1a, Prox1, Zeb1 | |
| 24 h | HYPOXIA INDUCIBLE FACTOR 1, ALPHA SUBUNIT (BASIC HELIX LOOP HELIX TRANSCRIPTION FACTOR) | 5.87E-03 | 100 | Slc16a3, Ncoa1, Idh1, Ndrg1, Id1, Mir23b, Abcc1, Vegfa, Egln3, Ptgs2 | C |
| 24 h | CYCLIN D2 | 6.79E-03 | 31 | Bcl6, Ccnd1, Myc, Fgf2, Rb1 | |
| 24 h | CYCLIN DEPENDENT KINASE INHIBITOR 2 | 7.01E-03 | 20 | Cdkn1c, Ccnd1, Rb1, Cdkn1a | D |
| 24 h | SEMAPHORIN | 9.91E-03 | 47 | Sema6d, Sema6a, Nrp2, Vegfa, Sema5a, Sema3c | A |
| 24 h | MOTHERS AGAINST DPP HOMOLOG | 9.93E-03 | 289 | Smad9, Smad7, Id2, Cdkn1c, Id3, Foxc2, Smad6, Id1, Mir206, Cilp, Atoh8, Aspn, Dlx2, Hmga2, Mir23b, Scn5a, Hfe2, Sp7, Dcn, Zeb1 | C |
| Myoblast differentiation with TNF-α treatment | |||||
| Time point | Pathway | P-value | # Genes (observed) | List of observed genes | SOTA cluster |
| 4 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 2.92E-10 | 12 | Mmp9, Ccl2, Fas, Nfkbia, Tnip1, Slc40a1, Tnfaip3, Cxcl10, Serpinb2, Nfkbie, Relb, Vcam1 | E |
| 4 h | NF KAPPA B | 5.89E-07 | 11 | Mmp9, Ccl2, Bcl3, Nfkbia, Nfkb2, Tnfaip3, Cxcl10, Nfkbie, Relb, Vcam1, Mcc | E |
| 4 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 1.54E-05 | 5 | Ccl2, Ccl7, Nfkbia, Cxcl10, Vcam1 | E |
| 4 h | INTERLEUKIN 1 | 7.36E-04 | 5 | Mmp9, Ccl2, Nfkbia, Cxcl10, Vcam1 | E |
| 4 h | RECEPTOR ACTIVATOR OF NUCLEAR FACTOR KAPPA B LIGAND | 9.58E-04 | 5 | Fas, Nfkbia, Tnfaip3, Nfkbie, Relb | |
| 4 h | CD40 LIGAND | 1.06E-03 | 3 | Fas, Nfkbia, Tnip1 | |
| 4 h | INTERLEUKIN 18 (INTERFERON GAMMA INDUCING FACTOR) | 1.13E-03 | 3 | Ccl2, Fas, Vcam1 | E |
| 4 h | TNF RECEPTOR ASSOCIATED FACTOR | 2.56E-03 | 4 | Fas, Nfkbia, Nfkb2, Tnfaip3 | E |
| 12 h | NF KAPPA B | 1.28E-12 | 18 | Ddx58, Mmp9, Ccl2, Cxcl1, Nfkbia, Nfkb2, Stap2, Ccl5, Cxcl10, Nfkbie, Relb, Vcam1, Cd74, Mcc, Saa3, Bcl3, Tnfaip3, Capn6 | E |
| 12 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 4.29E-12 | 15 | Mmp9, Ccl2, Cxcl1, Nfkbia, Ccl5, Cxcl10, Nfkbie, Relb, Vcam1, Saa3, Fas, Tnip1, Slc40a1, Tnfaip3, Serpinb2 | E |
| 12 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 1.74E-07 | 7 | Ccl7, Ccl2, Cxcl1, Nfkbia, Ccl5, Cxcl10, Vcam1 | C; E |
| 12 h | TOLL LIKE RECEPTOR | 2.08E-05 | 7 | Ddx58, Cxcl1, Stap2, Ccl5, Cxcl10, Saa3, Tnfaip3 | |
| 12 h | TNF RECEPTOR ASSOCIATED FACTOR | 1.02E-04 | 6 | Ddx58, Nfkbia, Nfkb2, Stap2, Fas, Tnfaip3 | E |
| 12 h | INTERLEUKIN 1 | 3.70E-04 | 6 | Mmp9, Ccl2, Cxcl1, Nfkbia, Cxcl10, Vcam1 | E |
| 12 h | MYELOID DIFFERENTIATION PRIMARY RESPONSE GENE (88) | 3.80E-04 | 5 | Ddx58, Cxcl1, Stap2, Ccl5, Cxcl10 | E |
| 12 h | RECEPTOR ACTIVATOR OF NUCLEAR FACTOR KAPPA B LIGAND | 5.05E-04 | 6 | Nfkbia, Ccl5, Nfkbie, Relb, Fas, Tnfaip3 | |
| 12 h | INTERLEUKIN 6 (INTERFERON, BETA 2) | 1.16E-03 | 5 | Ccl2, Cxcl1, Ccl5, Cxcl10, Cp | E |
| 12 h | CD40 LIGAND | 2.40E-03 | 3 | Nfkbia, Fas, Tnip1 | |
| 12 h | INTERLEUKIN 18 (INTERFERON GAMMA INDUCING FACTOR) | 2.55E-03 | 3 | Ccl2, Vcam1, Fas | E |
| 12 h | NUCLEOTIDE OLIGOMERIZATION DOMAIN/CASPASE RECRUITMENT DOMAIN PROTEIN FAMILY | 7.86E-03 | 3 | Ddx58, Ccl5, Tnfaip3 | |
| 12 h | TNFRSF1A ASSOCIATED VIA DEATH DOMAIN | 7.93E-03 | 2 | Stap2, Fas | |
| 24 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 3.25E-10 | 16 | Mmp9, Cxcl1, Ccl5, Nfkbie, Vcam1, Saa3, Lbp, Slc40a1, Tnfaip3, Ccl2, Birc3, Nfkbia, Cxcl10, Relb, Tnip1, Serpinb2 | E |
| 24 h | NF KAPPA B | 5.79E-10 | 19 | Mmp9, Cxcl1, Ccl5, Nfkbie, Vcam1, Cd74, Saa3, Bcl3, Tnfaip3, Ccl2, Birc3, Nfkbia, Nfkb2, Stap2, Cxcl10, Relb, Mcc, Abcb1b, Capn6 | E |
| 24 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 1.03E-08 | 9 | Cxcr4, Cxcl1, Ccl5, Vcam1, Ccl7, Ccl2, Nfkbia, Cxcl10, Abcb1b | C; E |
| 24 h | TOLL LIKE RECEPTOR | 2.92E-04 | 7 | Cxcl1, Ccl5, Saa3, Lbp, Tnfaip3, Stap2, Cxcl10 | E |
| 24 h | INTERLEUKIN 1 | 3.15E-03 | 6 | Mmp9, Cxcl1, Vcam1, Ccl2, Nfkbia, Cxcl10 | E |
| 24 h | MATRIX METALLOPROTEINASE | 4.08E-03 | 6 | Mmp9, Cxcr4, Ccl5, Enpp2, Postn, Adamts5 | C |
| 24 h | TNF RECEPTOR ASSOCIATED FACTOR | 5.66E-03 | 5 | Tnfaip3, Birc3, Nfkbia, Nfkb2, Stap2 | |
| 24 h | INTERLEUKIN 6 (INTERFERON, BETA 2) | 6.82E-03 | 5 | Cxcl1, Ccl5, Ccl2, Cxcl10, Cp | E |
| Myoblast differentiation with IGF1 treatment | |||||
| Time point | Pathway | P-value | # Genes (observed) | List of observed genes | SOTA cluster |
| 4 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 1.58E-10 | 12 | Mmp9, Ccl2, Fas, Nfkbia, Tnip1, Tnfaip3, Cxcl10, Serpinb2, Il1rn, Nfkbie, Relb, Vcam1 | E |
| 4 h | NF KAPPA B | 2.96E-08 | 12 | Ddx58, Mmp9, Ccl2, Bcl3, Nfkbia, Nfkb2, Tnfaip3, Cxcl10, Nfkbie, Relb, Vcam1, Mcc | E |
| 4 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 1.24E-05 | 5 | Ccl2, Ccl7, Nfkbia, Cxcl10, Vcam1 | C; E |
| 4 h | INTERLEUKIN 18 (INTERFERON GAMMA INDUCING FACTOR) | 4.14E-05 | 4 | Ccl2, Fas, Il1rn, Vcam1 | E |
| 4 h | INTERLEUKIN 1 | 5.62E-05 | 6 | Mmp9, Ccl2, Nfkbia, Cxcl10, Il1rn, Vcam1 | E |
| 4 h | TNF RECEPTOR ASSOCIATED FACTOR | 2.02E-04 | 5 | Ddx58, Fas, Nfkbia, Nfkb2, Tnfaip3 | E |
| 4 h | RECEPTOR ACTIVATOR OF NUCLEAR FACTOR KAPPA B LIGAND | 7.86E-04 | 5 | Fas, Nfkbia, Tnfaip3, Nfkbie, Relb | |
| 4 h | INTERLEUKIN 10 | 8.73E-04 | 4 | Ddx58, Ccl2, Cxcl10, Il1rn | |
| 4 h | CD40 LIGAND | 9.38E-04 | 3 | Fas, Nfkbia, Tnip1 | |
| 12 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 8.98E-12 | 16 | Mmp9, Ccl2, Cxcl1, Nfkbia, Ccl5, Cxcl10, Il1rn, Nfkbie, Relb, Vcam1, Saa3, Fas, Tnip1, Tnfaip3, Slc40a1, Serpinb2 | E |
| 12 h | NF KAPPA B | 8.55E-11 | 18 | Ddx58, Mmp9, Ccl2, Cxcl1, Nfkbia, Nfkb2, Stap2, Ccl5, Cxcl10, Nfkbie, Relb, Vcam1, Cd74, Mcc, Saa3, Bcl3, Tnfaip3, Capn6 | E |
| 12 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 7.07E-07 | 7 | Ccl2, Ccl7, Cxcl1, Nfkbia, Ccl5, Cxcl10, Vcam1 | C; E |
| 12 h | TOLL LIKE RECEPTOR | 7.76E-05 | 7 | Ddx58, Cxcl1, Stap2, Ccl5, Cxcl10, Saa3, Tnfaip3 | |
| 12 h | INTERLEUKIN 1 | 1.53E-04 | 7 | Mmp9, Ccl2, Cxcl1, Nfkbia, Cxcl10, Il1rn, Vcam1 | E |
| 12 h | TNF RECEPTOR ASSOCIATED FACTOR | 3.08E-04 | 6 | Ddx58, Nfkbia, Nfkb2, Stap2, Fas, Tnfaip3 | E |
| 12 h | INTERLEUKIN 18 (INTERFERON GAMMA INDUCING FACTOR) | 3.22E-04 | 4 | Ccl2, Il1rn, Vcam1, Fas | E |
| 12 h | MYELOID DIFFERENTIATION PRIMARY RESPONSE GENE (88) | 9.46E-04 | 5 | Ddx58, Cxcl1, Stap2, Ccl5, Cxcl10 | E |
| 12 h | RECEPTOR ACTIVATOR OF NUCLEAR FACTOR KAPPA B LIGAND | 1.46E-03 | 6 | Nfkbia, Ccl5, Nfkbie, Relb, Fas, Tnfaip3 | |
| 12 h | INTERLEUKIN 6 (INTERFERON, BETA 2) | 2.81E-03 | 5 | Ccl2, Cxcl1, Ccl5, Cxcl10, Cp | E |
| 12 h | CD40 LIGAND | 4.18E-03 | 3 | Nfkbia, Fas, Tnip1 | |
| 12 h | INTERLEUKIN 10 | 5.99E-03 | 4 | Ddx58, Ccl2, Cxcl10, Il1rn | |
| 24 h | TUMOR NECROSIS FACTOR (TNF SUPERFAMILY, MEMBER 2) | 3.25E-10 | 16 | Mmp9, Cxcl1, Ccl5, Nfkbie, Vcam1, Saa3, Lbp, Slc40a1, Tnfaip3, Ccl2, Birc3, Nfkbia, Cxcl10, Relb, Tnip1, Serpinb2 | E |
| 24 h | NF KAPPA B | 5.79E-10 | 19 | Mmp9, Cxcl1, Ccl5, Nfkbie, Vcam1, Cd74, Saa3, Bcl3, Tnfaip3, Ccl2, Birc3, Nfkbia, Nfkb2, Stap2, Cxcl10, Relb, Mcc, Abcb1b, Capn6 | E |
| 24 h | CHEMOKINE (C C MOTIF) LIGAND 2 | 1.03E-08 | 9 | Cxcr4, Cxcl1, Ccl5, Vcam1, Ccl7, Ccl2, Nfkbia, Cxcl10, Abcb1b | C; E |
| 24 h | TOLL LIKE RECEPTOR | 2.92E-04 | 7 | Cxcl1, Ccl5, Saa3, Lbp, Tnfaip3, Stap2, Cxcl10 | |
| 24 h | INTERLEUKIN 1 | 3.15E-03 | 6 | Mmp9, Cxcl1, Vcam1, Ccl2, Nfkbia, Cxcl10 | E |
| 24 h | MATRIX METALLOPROTEINASE | 4.08E-03 | 6 | Mmp9, Cxcr4, Ccl5, Enpp2, Postn, Adamts5 | C |
| 24 h | TNF RECEPTOR ASSOCIATED FACTOR | 5.66E-03 | 5 | Tnfaip3, Birc3, Nfkbia, Nfkb2, Stap2 | E |
| 24 h | INTERLEUKIN 6 (INTERFERON, BETA 2) | 6.82E-03 | 5 | Cxcl1, Ccl5, Ccl2, Cxcl10, Cp | E |
Signal transduction pathway associations, which were enriched after 4 h (“induction of differentiation”/immediate response), and 12 h (very early differentiation) of treatment, and 24 h (early differentiation) treatment, are depicted. The effects of differentiation without or with TNF-α or with IGF1 compared with TNF-α treatment are shown. Pathway enrichment was based on cocitation with a p value cutoff of <0.01. Genes within significantly enriched pathways are listed. In addition, it is indicated in which SOTA cluster a pathway is enriched. Pathways highlighted in bold are retrieved in enrichment analyses of genes identified by principal component analysis which are shown in Table 1.