EN-13-2081 Supplemental Figure 1: IVF experimental strategy. Cumulous-oocyte-complexes were isolated from superovulated C57Bl/6J females and incubated with previously capacitated caudal epididymal sperm. Following fertilization, zygotes were washed and cultured either under suboptimal WM and 20% oxygen conditions (IVF WM, red), or superior KAA medium with 5% oxygen conditions (IVF KAA, yellow). Blastocysts were used for molecular experiments or transferred into pseudo pregnant CF1 recipients, and newborn mice were maintained into adulthood for subsequent physiological and molecular analyses. As a control, C57Bl/6J females were superovulated and mated overnight with C57Bl/6J males. In vivogenerated blastocysts were isolated 96hrs post-fertilization, used directly in experiments, or transferred to recipients (flushed blastocyst/fb group, blue). Supplemental Figure 2: Conception by IVF WM reduces growth and impairs glucose tolerance. (A) Postnatal growth curves depicting average body weight for male FB (blue) and IVF WM (red) mice. (B) Male intraperitoneal glucose tolerance test (IPGTT) sampled at 19wks for clearance of a 1.5mg/g glucose bolus over 120 min, with area under the curve (AUC) calculated from total glucose levels measured during the entire IPGTT course. (C) Postnatal growth of female FB and IVF WM animals, and (D) female IPGTT sampled in tandem with males. For all measurements, FB males n=8, IVF WM males n=12, FB females n=6, IVF WM females n=8. Error bars depict standard error of the mean, *p<0.05. IVF WM mice display altered metabolic and physiological parameters at 40wks of age. (A) Serum insulin, leptin and corticosterone in male FB and IVF WM animals with (B) parallel measurements in females. At sacrifice, selected morphological measurements included (C) body weight, (D) body mass index (BMI), and (E) anogenital distance. (F,G) Organ weights were normalized as percent body weight for (F) male and (G) female FB and IVF WM males and females. Individual sample sizes are shown in (A,B); sample size for (C-G) was: FB males n=8, IVF WM males n=12, FB females n=5, IVF WM females n=8. Error bars depict standard error of the mean, *p<0.05. Supplemental Figure 3: Histological analysis of glomeruli and pancreatic islets. Histological analysis of (A) FB and (B) IVF WM male kidneys at 40wks revealed no significant changes in (C) number of glomeruli (open arrowheads) or (D) glomerular volume. Analysis conducted on n-5 mice per condition. Immunostaining of insulin in (E) FB and (F) IVF WM male pancreata at 40wks showed no changes in (G) percent of insulin positive tissue. Analysis conducted on n=4 mice per condition. Scale bars represent 300mm, error bars depict standard deviation. Supplemental Figure 4: Intraperitoneal glucose tolerance test performed on female FB (blue) and IVF KAA (yellow) animals aged (A) 13wks, (B) 20wks, and (C) 29wks, with area under the curve (AUC) measured from total glucose levels during entire IPGTT course. For all time points, FB n=8 and IVF KAA n=7 females. Error bars depict standard deviation, *p<0.05. Supplemental Figure 5: IVF KAA morphometric parameters at sacrifice. Comparison of morphological parameters between FB (blue) and IVF KAA (yellow) animals at sacrifice (30wks), including (A) weight, (B) body mass index (BMI), (C) crown-rump length, (D) anogenital distance, (E) anogenital index, or (F) biparietal diameter (BPD). (G-H) Organ sizes depicted as relative body weights for (G) male and (H) female FB and IVF KAA animals. For all measurements, sample size was: FB males n=8, IVF KAA males n=8-14, FB females n=5-8, IVF KAA females n=5-8. Error bars depict standard deviation, *p<0.05.
Supplemental Figure 6: Normal in IVF KAA males. (A) Postnatal growth of FB (blue) and IVF KAA (yellow) males, showing periodic intraperitoneal glucose tolerance sampling (IPGTT) and the administration of a high fat diet beginning at 24wks. (B-D) IPGTT of glucose handling at (B) 13wks, (C) 20wks, and (D) 29wks of age, with corresponding area under the curve (AUC) measured form total glucose levels during entire IPGTT course. (E) Average daily food intake monitored over five-day period. (F) Serum insulin measured at 11wks. (G-I) Intermittent DEXA scanning of (G) bone mineral density and (H) fat content, with calculated (I) percent fat gain in FB and IVF KAA males. Sample size indicated on graphs; DEXA sample size was FB males n=8 and IVF KAA males n=14. Error bars depict standard deviation, *p<0.05. Supplemental Figure 7: DNA methylation analysis of the Txnip locus. Bisulfite sequencing of DNA methylation profiles at the Txnip promoter and 5 UTR revealed no changes in CpG methylation between FB (n=3) and IVF KAA (n=3) blastocysts.
Supplemental Figure 1 C57Bl/6J X C57Bl/6J Preimplantation development IVFWM / IVFKAA Embryo culture Culture medium KAA or WM Oxygen tension 5% or 20% Blastocyst transfer In vivo Blastocysts flushed from the uterine horns Molecular experiments Placentation & fetal growth CF1 Postnatal development IVFWM / IVFKAA experimental group Newborn/adult mice Physiological observations Molecular experiments Flushed blastocyst (FB) control group
Supplemental Figure 3 A B C FB D IVFWM E F G FB IVFWM
Supplemental Figure 7 CpG sites: 1 10 20 30 40 50 TATA-box ATG Exon 1-200 -100 TSS +1 100 200 300 400 IVFKAA FB methylated unmethylated
Supplemental Table 1A Males Females Physiological parameter FB (n=8) IVF WM (n=20) FB (n=7) IVF WM (n=13) Birth weight (g) 1.73 ± 0.06 1.57 ± 0.05 1.58 ± 0.06 1.51 ± 0.05 AGD (mm) 1.85 ± 0.11 2.06 ± 0.07 1.26 ± 0.12 1.07 ± 0.06 AGI (mm/g) 1.07 ± 0.04 1.32* ± 0.05 0.79 ± 0.07 0.72 ± 0.04 Head diameter (mm) 7.63 ± 0.09 7.60 ± 0.11 7.70 ± 0.15 7.52 ± 0.12 Abdominal diameter (mm) 9.68 ± 0.30 9.25 ± 0.22 9.23 ± 0.33 8.86 ± 0.17 Crown-rump length (mm) 30.55 ± 0.46 30.17 ± 0.36 29.93 ± 0.59 29.18 ± 0.52 BMI (g/cm 2 ) 0.185 ± 0.005 0.173 ± 0.004 0.177 ± 0.003 0.177 ± 0.004 Supplemental Table 1B FB (n=14) IVF KAA (n=11) Litter size (# pups) 9.50 ± 0.78 3.55* ± 0.58 Survival (%) 58.9 ± 4.7 19.3* ± 2.8 Sex (% males) 54.5 ± 5.2 55.6 ± 8.8 Males Females Physiological parameter FB (n=9) IVF KAA (n=15) FB (n=8) IVF KAA (n=10) Birth weight (g) 1.51 ± 0.04 1.50 ± 0.06 1.65 ± 0.03 1.40* ± 0.06 AGD (mm) 1.84 ± 0.06 2.07 ± 0.09 1.33 ± 0.06 1.14* ± 0.05 AGI (mm/g) 1.23 ± 0.05 1.43 ± 0.11 0.80 ± 0.03 0.84 ± 0.06 Head diameter (mm) 7.64 ± 0.08 7.63 ± 0.09 8.06 ± 0.16 7.55* ± 0.14 Abdominal diameter (mm) 9.92 ± 0.47 9.16 ± 0.29 8.80 ± 0.26 8.46 ± 0.44 Crown-rump length (mm) 28.92 ± 0.42 29.24 ± 0.38 31.10 ± 0.30 29.04* ± 0.48 BMI (g/cm 2 ) 0.182 ± 0.007 0.175 ± 0.005 0.170 ± 0.003 0.165 ± 0.006 Morphometric parameters measured in FB WM, IVF WM, FB KAA and IVF KAA pups at birth. AGD, anogenital distance; AGI, anogenital index; BMI, body mass index. Values are ±SEM, with p<0.05 indicated by * in bold.
Supplemental Table 2. Comprehensive list of metabolites detected by non-targeted mass spectrometry (Metabolon, Inc.) in FB and IVFKAA female serum at 29wks. The Fold IVF KAA Individual IVF values normalized to FB mean Pathway Subpathay Biochemical Platform Change FB p-value IVFKAA 1 IVFKAA 2 IVFKAA 3 IVFKAA 4 Amino acid Glycine, serine glycine GC/MS 0.93 0.6391 0.774999472 0.80681986 1.271633855 0.85653394 and threonine dimethylglycine GC/MS 1.00 0.8820 0.890740492 0.730815751 1.554988129 0.83644671 N-acetylglycine GC/MS 1.78 0.1018 2.063447481 1.52893306 2.165136293 1.35176983 beta-hydroxypyruvate GC/MS 0.93 0.7337 1.182724252 1.184023181 0.602103265 0.73689206 serine GC/MS 0.99 0.8713 0.741475636 1.002831016 1.227909307 1.00142803 threonine LC/MS pos 1.11 0.4234 1.084488344 1.178376082 1.27821146 0.91487441 betaine LC/MS pos 1.08 0.3500 1.206138082 0.960023646 1.166826769 0.97095988 Alanine and aspartate GC/MS 0.72 0.2422 1.45811393 0.453283433 0.613690761 0.36598634 aspartate asparagine GC/MS 1.13 0.7694 0.657265298 0.912298186 1.584612318 1.3565876 3-ureidopropionate LC/MS pos 0.75 0.2362 0.899990636 0.532446858 0.974248525 0.59911977 N-acetyl-beta-alanine LC/MS pos 1.51 0.4048 1.947179946 0.862242614 2.524283796 0.71732319 alanine GC/MS 0.84 0.4228 0.975165877 1.213838863 0.691184834 0.4971564 N-acetylalanine LC/MS neg 0.79 0.2414 0.525454461 0.661025617 0.649216607 1.33367428 N-acetylaspartate (NAA) GC/MS 0.85 0.4543 0.773106293 0.767664355 0.999122268 0.87773194 Glutamate glutamate GC/MS 0.61 0.0601 0.663380996 0.333110232 0.607484028 0.81874049 glutamine LC/MS pos 0.67 0.0279 0.550439093 0.534315036 0.836805627 0.75783066 N-acetylglutamate LC/MS pos 0.70 0.2428 0.841396179 0.482928687 0.860963532 0.63168869 Histidine histidine LC/MS neg 0.94 0.7821 0.905326119 1.046138392 0.921032806 0.86770313 trans-urocanate LC/MS pos 0.95 0.7407 1.865749836 0.867620919 0.403031154 0.68285153 Lysine lysine LC/MS pos 1.13 0.4207 0.999818761 1.107215908 1.324599332 1.0714859 2-aminoadipate LC/MS pos 0.98 0.9966 1.195564046 0.857835143 0.892782256 0.97199571 pipecolate LC/MS pos 1.03 0.7874 1.100281957 0.846071313 1.238915088 0.9448811 glutaroyl carnitine LC/MS pos 1.07 0.8612 2.354291845 0.547067239 0.698569385 0.67188841 Phenylalanine & phenylalanine LC/MS pos 0.97 0.8646 0.975001804 0.948920915 1.01215023 0.94978707 tyrosine phenylacetate LC/MS neg 1.14 0.2185 1.393829188 1.173158142 1 1 p-cresol sulfate LC/MS neg 2.04 0.0232 2.298825551 2.145860785 2.48181037 1.21890576 tyrosine LC/MS pos 1.06 0.6447 1.108043789 1.105716854 1.14823629 0.85906182 3-(4-hydroxyphenyl)lactate LC/MS neg 0.57 0.4374 0.698319513 0.587128495 0.587011081 0.40490203 Tryptophan 4-hydroxyphenylpyruvate LC/MS neg 1.16 0.5954 0.868867183 1.376557764 1.810710677 0.58223869 N-acetylphenylalanine LC/MS pos 1.18 0.4162 1.573649187 0.983427969 1.072017677 1.07468976 N-acetyltyrosine LC/MS neg 1.56 0.2159 1.677850572 1.823042816 1.930646549 0.82271193 phenylacetylglycine LC/MS neg 1.10 0.9329 0.820645408 0.47323566 1.745666954 1.37422197 phenol sulfate LC/MS neg 0.76 0.1918 0.705964595 0.512531612 0.76073771 1.05557497 p-toluic acid GC/MS 0.78 0.1726 0.681150625 0.681150625 0.957203544 0.78436703 kynurenate LC/MS neg 0.83 0.7387 0.799900818 0.509463592 1.164145797 0.82651459 kynurenine LC/MS pos 1.33 0.1571 0.994226102 1.237231876 1.830018688 1.24504198 tryptophan LC/MS pos 1.15 0.1902 1.09153788 1.050578191 1.344320761 1.11985302 indolelactate GC/MS 1.07 0.6274 1.10291918 1.027358263 1.321109771 0.83281062 indolepyruvate LC/MS neg 5.44 0.2386 1.113988605 16.76289892 3.081716403 0.80210367 serotonin (5HT) LC/MS pos 0.35 0.6880 0.502146954 0.15096889 0.553699842 0.19477845 N-acetyltryptophan LC/MS neg 1.71 0.1915 1.785935302 1.516455696 2.256680731 1.29648383 C-glycosyltryptophan* LC/MS pos 0.92 0.5973 1.161118895 0.577419199 1.06739456 0.86506595
Valine, leucine and isoleucine 3-indoxyl sulfate LC/MS neg 1.25 0.3026 1.34768916 1.237221071 1.609108772 0.78985761 indolepropionate LC/MS neg 1.21 0.2252 1.355131025 1.368485202 1.280835166 0.84597652 3-methyl-2-oxobutyrate LC/MS neg 1.72 0.0823 1.988976943 2.076770049 1.154485862 1.65671983 3-methyl-2-oxovalerate LC/MS neg 1.57 0.0341 1.442536675 1.903953687 1.518847444 1.43385304 beta-hydroxyisovalerate LC/MS neg 0.58 0.0415 0.474484256 0.407941678 0.585155886 0.86582907 alpha-hydroxyisocaproate LC/MS neg 0.75 0.2550 0.889709402 0.424022792 1.271430199 0.42402279 isoleucine LC/MS pos 1.03 0.7030 1.053792679 1.082317933 1.074211836 0.91875946 leucine LC/MS pos 0.92 0.7638 1.010805254 0.945014278 0.885619646 0.83563678 N-acetylleucine LC/MS pos 0.76 0.5814 1.091474182 0.665889666 0.701702338 0.56643357 N-acetylisoleucine LC/MS pos 0.98 0.8142 1.203436139 1.11561277 0.935929308 0.64567898 valine LC/MS pos 0.99 0.9091 1.073686261 0.995373302 0.952710271 0.92738518 2-hydroxyisobutyrate GC/MS 0.85 0.3584 0.819157721 0.507254925 1.082458417 0.98364988 4-methyl-2-oxopentanoate LC/MS neg 1.64 0.0592 1.613750877 2.074366988 1.188085975 1.68824542 alpha-hydroxyisovalerate LC/MS neg 1.26 0.2273 0.895881437 1.691778983 1.335015841 1.13569242 isovalerylglycine LC/MS pos 0.58 0.5885 0.892374854 0.410419092 0.545750873 0.4628638 isobutyrylcarnitine LC/MS pos 0.64 0.2860 0.41486009 0.289867252 1.059765065 0.80893897 2-hydroxy-3-methylvalerate LC/MS neg 0.93 0.6477 0.851768106 0.898575724 1.292710786 0.69670622 2-methylbutyroylcarnitine LC/MS pos 0.63 0.2751 0.442865475 0.474256323 0.911112289 0.70792018 isovalerylcarnitine LC/MS pos 0.35 0.2136 0.271707237 0.248453212 0.438046046 0.44112314 hydroxyisovaleroyl carnitine LC/MS pos 0.85 0.4411 0.875565307 0.624546635 0.915506202 0.99511933 methylglutaroylcarnitine LC/MS pos 0.88 0.4668 2.041149999 0.621140959 0.497258632 0.37738519 isobutyrylglycine LC/MS pos 0.35 0.0987 0.302080951 0.173473588 0.592362223 0.31260005 Cysteine, cysteine GC/MS 0.77 0.2741 1.489313939 0.496254725 0.708738976 0.4039858 methionine, SAM, N-formylmethionine LC/MS neg 0.94 0.8006 0.924506606 0.846971581 1.031610072 0.94956938 taurine taurine GC/MS 1.25 0.5943 3.751029382 0.778892664 0.234019588 0.23401959 methionine LC/MS pos 1.15 0.2974 1.15152836 1.204271781 1.228137089 1.02795183 N-acetylmethionine LC/MS neg 1.10 0.5351 1.245304192 0.979293024 1.250647928 0.92810003 2-hydroxybutyrate (AHB) GC/MS 0.71 0.3705 0.712884213 0.674179738 0.82300766 0.62234339 Urea cycle; arginine-, proline-, Creatine Butanoate Polyamine dimethylarginine (SDMA + LC/MS pos 0.89 0.6471 1.137001898 0.458950032 1.093485136 0.86669197 ADMA) arginine LC/MS pos 1.11 0.9333 0.128773516 0.89229984 2.736765349 0.67288902 ornithine GC/MS 1.31 0.6476 2.589453861 1.109322034 0.770056497 0.78973635 urea GC/MS 1.47 0.0526 1.787648594 1.489823137 1.579356335 1.02116556 proline LC/MS pos 1.20 0.4167 1.330473991 1.483668756 1.130539976 0.83778731 citrulline LC/MS pos 1.08 0.7952 0.956558667 0.731632984 1.565386825 1.07626589 trans-4-hydroxyproline GC/MS 0.92 0.5324 0.850182921 0.760149906 1.305612564 0.76603908 homocitrulline LC/MS pos 1.28 0.2489 1.500988142 1.356225296 1.453310277 0.82905138 creatine LC/MS pos 0.83 0.4286 0.83803478 0.518127766 1.08873888 0.85752604 creatinine LC/MS pos 0.90 0.6268 0.866405229 0.749803922 1.224836601 0.74300654 2-aminobutyrate GC/MS 0.86 0.7839 0.528518879 0.898228405 1.209758573 0.81121306 5-methylthioadenosine (MTA) LC/MS pos 0.66 0.4191 0.503030303 0.503030303 1.101928375 0.5430303
Glutathione S-methylglutathione LC/MS pos 0.33 0.2869 0.30521909 0.529909759 0.280100474 0.20976835 5-oxoproline LC/MS neg 0.78 0.2334 0.820579998 0.427229199 1.052127563 0.82744602 glutathione, oxidized (GSSG) LC/MS pos 0.28 0.0419 0.50334045 0.228154241 0.192339102 0.18922988 cysteine-glutathione disulfide LC/MS pos 0.78 0.2163 1.239521603 0.555112596 0.708199547 0.60398664 Peptide ophthalmate LC/MS pos 0.52 0.0065 0.697873148 0.451196354 0.461071022 0.45119635 Dipeptide alanylalanine GC/MS 1.03 0.8906 0.832898984 0.8078422 1.403371189 1.07667663 gamma-glutamyl gamma-glutamylvaline LC/MS pos 1.18 0.3066 1.566911246 1.07225109 1.1672977 0.91871426 gamma-glutamylleucine LC/MS pos 1.13 0.6700 1.65054426 1.201992196 0.91846375 0.73125899 gamma-glutamylisoleucine* LC/MS pos 1.09 0.6384 1.363247322 1.173128054 1.042465372 0.76979565 Fibrinogen cleavage peptide Carbohydrate Aminosugars Fructose, mannose, galactose, starch, and Glycolysis, sucrose gluconeogenesis, pyruvate Energy Nucleotide sugars, pentose Krebs cycle Oxidative phosphorylation gamma-glutamylmethionine LC/MS pos 2.10 0.3973 0.294256047 2.634626847 3.882799511 1.59375098 gammaglutamylphenylalanine LC/MS pos 1.39 0.2153 1.879791917 1.716740029 1.083740163 0.88088569 gamma-glutamyltyrosine LC/MS pos 1.35 0.1567 1.816124793 1.392834695 1.288040683 0.90383407 gamma-glutamyltryptophan LC/MS pos 1.34 0.1575 1.526114206 1.611420613 1.309308264 0.89821263 gamma-glutamylalanine LC/MS pos 1.16 0.5327 1.379165416 1.418793155 1.134434104 0.71053738 TDTEDKGEFLSEGGGV* LC/MS pos 0.95 0.6535 2.826739074 0.152917852 0.648219981 0.15291785 TDTEDKGEFLSEGGGVR* LC/MS neg 0.55 0.4879 1.728483358 0.027416141 0.4293457 0.02741614 erythronate* GC/MS 0.93 0.5722 0.985690645 0.723547162 1.284726954 0.7373698 fructose GC/MS 1.21 0.3195 1.22109615 1.051936493 1.141626712 1.41575034 mannose GC/MS 1.24 0.0404 1.359471267 1.365520332 1.1181808 1.12221351 sorbitol GC/MS 0.69 0.1305 0.867333333 0.799333333 0.5165 0.57208333 1,5-anhydroglucitol (1,5-AG) GC/MS 1.03 0.8086 1.227247375 0.950893615 1.136164124 0.82508427 glycerate GC/MS 1.39 0.1961 1.831580349 1.372919718 1.034083344 1.31314073 glucose GC/MS 1.06 0.5401 1.257992034 1.096721479 0.969461751 0.92350117 pyruvate GC/MS 1.29 0.0883 1.381842949 1.201577887 1.077049692 1.50398729 lactate GC/MS 0.83 0.0874 0.99015351 0.767409306 0.881201509 0.69404703 glucuronate GC/MS 0.67 0.2991 0.594352339 0.609516107 0.988205958 0.47115514 ribose GC/MS 1.13 0.7237 1.83553191 1.113135583 0.83035736 0.72356242 ribulose GC/MS 0.72 0.5817 0.798542573 0.616434964 1.067883547 0.38940182 arabinose GC/MS 0.78 0.3815 0.688948211 0.688948211 0.688948211 1.03854778 xylonate GC/MS 0.71 0.2721 0.998531123 0.358405982 0.70017741 0.78968352 citrate GC/MS 1.45 0.0227 1.798932641 1.412029424 1.459454781 1.13578537 cis-aconitate LC/MS neg 1.31 0.0183 1.344751109 1.291276491 1.362124199 1.26133563 alpha-ketoglutarate GC/MS 1.76 0.0185 1.21425195 2.215197353 2.018317183 1.595013 succinate GC/MS 0.24 0.0495 0.13937183 0.106913426 0.42405908 0.28774802 succinylcarnitine LC/MS pos 0.70 0.2180 1.169525752 0.688954615 0.479102499 0.48032636 fumarate GC/MS 1.15 0.7516 1.801586473 1.196847928 1.014716627 0.59190064 malate GC/MS 1.23 0.7092 2.307398914 1.131394086 0.79980688 0.67340978 acetylphosphate GC/MS 0.93 0.7193 0.995924239 1.001965955 0.713593862 1.02258451 phosphate GC/MS 0.76 0.1540 0.694971128 0.532135667 0.784707626 1.01320826
Lipid pyrophosphate (PPi) GC/MS 1.09 0.9890 1.130285178 1.311297638 0.373598874 1.54861582 Essential fatty acid linoleate (18:2n6) LC/MS neg 1.20 0.2154 1.447426618 1.356755127 0.993264978 0.98974668 linolenate [alpha or gamma; LC/MS neg 1.18 0.3378 1.487266055 1.339400612 0.902262997 0.9951315 (18:3n3 or 6)] dihomo-linolenate (20:3n3 or LC/MS neg 1.15 0.4264 1.446670461 1.134092203 0.857598179 1.14251565 n6) eicosapentaenoate (EPA; 20:5n3) LC/MS neg 1.00 0.8603 0.921879208 0.974465689 1.05133792 1.04663745 docosapentaenoate (n3 DPA; LC/MS neg 1.09 0.9924 1.8904859 1.081613368 0.703237678 0.66563883 22:5n3) docosapentaenoate (n6 DPA; LC/MS neg 0.77 0.0986 0.976170987 0.791905412 0.550159163 0.74815825 22:5n6) docosahexaenoate (DHA; 22:6n3) LC/MS neg 0.91 0.2567 0.865388768 0.847923614 0.950315476 0.99551376 Medium chain fatty acid Long chain fatty acid caproate (6:0) LC/MS neg 0.78 0.4293 0.368083129 0.505280054 1.226746642 1.00540678 heptanoate (7:0) LC/MS neg 0.70 0.4091 0.272654993 0.395291562 1.213644385 0.90856556 caprylate (8:0) LC/MS neg 0.70 0.4272 0.203603604 0.336576577 1.353441441 0.92533333 pelargonate (9:0) LC/MS neg 0.76 0.5017 0.225526739 0.409856005 1.404323773 1.00623847 caprate (10:0) LC/MS neg 0.83 0.4976 0.25872604 0.400853771 1.600318071 1.04904997 undecanoate (11:0) LC/MS neg 0.80 0.4424 0.373598894 0.631643598 1.261181425 0.92987343 10-undecenoate (11:1n1) LC/MS neg 1.24 0.6190 0.850627972 0.667333252 1.887672235 1.5578588 laurate (12:0) LC/MS neg 0.86 0.4439 0.489324337 0.658656068 1.264164895 1.02920788 myristate (14:0) LC/MS neg 0.88 0.1224 0.934706953 0.823711079 0.909078338 0.8441771 myristoleate (14:1n5) LC/MS neg 0.93 0.5922 0.693563557 0.919188753 1.130537536 0.97687318 palmitate (16:0) LC/MS neg 0.94 0.2281 1.032690297 0.889005037 0.941081441 0.90478001 palmitoleate (16:1n7) LC/MS neg 1.03 0.5439 0.952190212 1.085913829 1.055923587 1.04349612 margarate (17:0) LC/MS neg 0.97 0.9169 0.92082389 0.758038405 1.11907549 1.07255112 10-heptadecenoate (17:1n7) LC/MS neg 1.05 0.9398 1.486560063 1.164635919 0.760354169 0.77642395 stearate (18:0) LC/MS neg 0.86 0.0392 0.944306132 0.775656985 0.837344398 0.872568 oleate (18:1n9) GC/MS 1.25 0.2335 1.34312909 1.488637716 1.18179655 0.97596669 cis-vaccenate (18:1n7) GC/MS 1.31 0.1731 1.478772854 1.52272023 1.155815984 1.0978956 stearidonate (18:4n3) LC/MS neg 1.05 0.8320 1.271459118 1.367681499 0.691477446 0.86549231 nonadecanoate (19:0) LC/MS neg 1.03 0.9056 1.309090909 1.145929724 0.852119545 0.8120045 10-nonadecenoate (19:1n9) LC/MS neg 1.08 0.5720 1.275301937 1.226694442 0.980383404 0.84973836 eicosenoate (20:1n9 or 11) LC/MS neg 1.42 0.2287 1.856176205 1.909322521 1.01514944 0.90304906 dihomo-linoleate (20:2n6) LC/MS neg 1.12 0.3891 1.336885326 1.205103138 0.917293968 1.03783361 mead acid (20:3n9) LC/MS neg 0.74 0.1306 0.945331683 0.710786336 0.570766299 0.73310154 arachidonate (20:4n6) LC/MS neg 0.75 0.0798 0.689328277 0.538912965 0.794149512 0.96433731 docosadienoate (22:2n6) LC/MS neg 1.07 0.5941 1.149528813 0.854974564 1.202012621 1.07436133 adrenate (22:4n6) LC/MS neg 1.10 0.3568 1.236145091 1.140304223 1.004999468 1.02276354 Fatty acid, methyl palmitate, methyl ester GC/MS 0.93 0.7580 0.605973303 0.614424277 1.415538269 1.07288966 ester Fatty acid, ester n-butyl Oleate GC/MS 0.84 0.6853 0.547961075 0.505946864 1.08819895 1.20250232 Fatty acid, 4-hydroxybutyrate (GHB) GC/MS 0.30 0.0156 0.228929046 0.125436862 0.447833979 0.41564845 monohydroxy
monohydroxy 16-hydroxypalmitate LC/MS neg 0.91 0.7469 0.847967346 0.64069061 1.132468507 1.02868939 2-hydroxystearate LC/MS neg 0.99 0.6640 0.470100341 0.311938413 1.555973973 1.63043116 2-hydroxypalmitate LC/MS neg 0.86 0.4355 0.582391836 0.421304733 1.30067163 1.13250377 13-HODE + 9-HODE LC/MS neg 0.76 0.6780 1.109763026 0.825394498 0.535747668 0.57452853 Fatty acid, 2-hydroxyglutarate GC/MS 0.67 0.0951 0.802963935 0.537753507 0.671979623 0.6552303 dicarboxylate sebacate (decanedioate) LC/MS neg 1.16 0.1233 1.347873018 1.107082561 1.16110819 1.03456865 hexadecanedioate LC/MS neg 0.78 0.2584 0.695213055 0.535369976 1.051291682 0.84161763 octadecanedioate LC/MS neg 0.99 0.9786 1.312612752 0.519251585 1.17581568 0.96355858 2-hydroxyadipate GC/MS 0.76 0.5881 0.896130663 0.654792922 1.088625316 0.4066887 Fatty acid, amide stearamide GC/MS 1.14 0.4028 0.966248353 1.185376196 1.208755945 1.19798292 Fatty acid, branched 15-methylpalmitate (isobar LC/MS neg 0.96 0.7693 0.808031436 0.755702511 1.179988499 1.1142419 with 2-methylpalmitate) 17-methylstearate LC/MS neg 0.92 0.4949 1.175906014 0.907349978 0.748353215 0.83853589 Eicosanoid 12-HETE LC/MS neg 0.55 0.3850 0.688228468 0.657281733 0.362480448 0.49245673 Fatty acid propionylcarnitine LC/MS pos 0.90 0.7547 0.631878732 0.468810289 1.284244373 1.20551217 (also butyrylcarnitine LC/MS pos 0.25 0.0385 0.09929501 0.069774872 0.427040958 0.412004 BCAA Fatty acid ) valerylcarnitine LC/MS pos 0.57 0.2549 0.499445311 0.499445311 0.694245703 0.59198997 valerylglycine LC/MS pos 0.74 0.4627 0.565130764 0.526718242 0.834736408 1.03979465 isovalerate LC/MS neg 0.88 0.8604 0.548524244 0.880194638 1.090462264 0.98301836 hexanoylglycine LC/MS pos 0.40 0.0133 0.213707469 0.354600482 0.500110998 0.52959197 Carnitine deoxycarnitine LC/MS pos 0.96 0.6807 0.693330687 0.679436284 1.332175466 1.13914252 carnitine LC/MS pos 0.77 0.0301 0.710189952 0.722794248 0.724835789 0.92401917 3-dehydrocarnitine* LC/MS pos 1.21 0.4300 0.917301751 0.87507724 1.279093718 1.77013388 acetylcarnitine LC/MS pos 1.09 0.4982 1.00724246 0.948870392 1.199005513 1.20116744 hexanoylcarnitine LC/MS pos 0.23 0.0346 0.103907492 0.117498767 0.402214063 0.30470762 octanoylcarnitine LC/MS pos 0.61 0.1590 0.548902852 0.548902852 0.709676172 0.62778154 palmitoylcarnitine LC/MS pos 0.73 0.1930 0.648668135 0.668776287 0.808812337 0.77676747 stearoylcarnitine LC/MS pos 0.98 0.8186 1.099573773 0.989843112 0.905141924 0.90858801 oleoylcarnitine LC/MS pos 0.86 0.9019 0.758921751 0.850522027 1.221579541 0.61020772 Bile acid cholate LC/MS neg 2.58 0.2098 1.747106355 2.196112688 6.116269928 0.26992793 12-dehydrocholate LC/MS neg 3.44 0.1343 2.43136833 1.871557659 8.437392427 1 taurocholate LC/MS neg 0.37 0.9487 0.108886723 1.329606231 0.037025842 0.00742474 taurochenodeoxycholate LC/MS neg 0.21 0.9442 0.038348797 0.797539715 0.006186859 0.00150918 deoxycholate LC/MS neg 2.62 0.0172 2 2.103094532 4.598897838 1.79703264 taurodeoxycholate LC/MS neg 0.22 0.9898 0.091092939 0.70653853 0.061051752 0.01336119 beta-muricholate LC/MS neg 1.42 0.3165 0.996075994 1.304168771 2.817669684 0.55798594 alpha-muricholate LC/MS neg 1.07 0.8482 0.783910129 1.35046204 2.052908136 0.07450625 tauroursodeoxycholate LC/MS neg 0.26 0.9324 0.043284352 0.95517826 0.021608661 0.00492386 Glycerolipid glycerol GC/MS 0.71 0.0338 0.739021956 0.567032601 0.674567532 0.86610113 glycerol 3-phosphate (G3P) GC/MS 0.82 0.4713 0.374628625 0.318604103 1.388823391 1.20396133 Inositol glycerophosphorylcholine LC/MS pos 0.67 0.2370 0.596899674 0.288565858 0.646474006 1.16769947 (GPC) myo-inositol GC/MS 1.16 0.6511 1.603419402 1.397055515 0.943834829 0.68198065 inositol phosphate (I1P) GC/MS 0.83 0.3974 0.723294063 0.547486294 0.891076636 1.14144407
Ketone bodies 3-hydroxybutyrate (BHBA) GC/MS 2.28 0.0400 2.138928687 1.872533161 3.291953598 1.82483708 Lysolipid palmitoylglycerophosphoetha nolamine 2- palmitoylglycerophosphoetha nolamine* LC/MS neg 0.97 0.7040 0.712645868 0.657966182 1.427387473 1.06263396 LC/MS neg 1.11 0.8943 1.007094563 2.542484023 0.474798555 0.40707604 LC/MS neg 1.06 0.3585 1.001126267 0.962983356 1.126967839 1.15610061 stearoylglycerophosphoethan olamine oleoylglycerophosphoethanol amine 2- oleoylglycerophosphoethanol amine* LC/MS neg 0.79 0.9214 1.067814205 0.901874186 0.582148991 0.60804409 LC/MS neg 1.14 0.4333 1.431028512 1.047096849 0.845665474 1.22140136 LC/MS neg 1.21 0.3760 1.241048357 1.380460194 1.000861326 1.21988434 linoleoylglycerophosphoethan olamine* 2- LC/MS neg 1.14 0.5161 1.415200875 1.297188636 0.793094274 1.06778609 linoleoylglycerophosphoethan olamine* arachidonoylglycerophospho ethanolamine* 2- arachidonoylglycerophospho ethanolamine* LC/MS neg 0.81 0.3975 0.924197889 0.896625957 0.697578141 0.73053198 LC/MS neg 0.80 0.7533 0.965810479 1.083009606 0.525076825 0.61382656 2- LC/MS pos 0.67 0.0466 0.804080969 0.821803105 0.529713031 0.51524266 docosahexaenoylglycerophos phoethanolamine* LC/MS pos 0.46 0.0038 0.617649181 0.340645068 0.493253481 0.38119633 myristoylglycerophosphocholi ne LC/MS pos 0.40 0.0235 0.513846038 0.328728482 0.421967283 0.35446864 palmitoylglycerophosphocholi ne 2- LC/MS pos 0.56 < 0.001 0.642932378 0.489572361 0.557059961 0.53109141 palmitoylglycerophosphocholi ne* palmitoleoylglycerophosphoc holine* LC/MS pos 0.67 0.0043 0.662109672 0.603259336 0.809929909 0.62009852
2- palmitoleoylglycerophosphoc holine* heptadecanoylglycerophosph ocholine LC/MS pos 0.49 0.0065 0.689611502 0.354527508 0.45457593 0.4630685 LC/MS pos 0.60 0.0031 0.704885993 0.490515424 0.653458517 0.55182985 LC/MS pos 0.84 0.4970 0.700695078 0.950642421 1.069250392 0.64078261 stearoylglycerophosphocholin e 2- LC/MS pos 0.57 0.0101 0.620549128 0.511552409 0.675945433 0.46437576 stearoylglycerophosphocholin e* oleoylglycerophosphocholine 2- oleoylglycerophosphocholine * linoleoylglycerophosphocholi ne 2- linoleoylglycerophosphocholi ne* arachidoylglycerophosphoch oline LC/MS pos 0.35 < 0.001 0.434235297 0.280423726 0.400011239 0.29936216 LC/MS pos 0.22 0.0021 0.357300857 0.234969554 0.128892755 0.14837828 LC/MS pos 0.98 0.7162 0.957653936 1.048964901 0.999106215 0.91513878 LC/MS pos 1.03 0.8583 1.16572024 1.195207085 1.007554051 0.74915343 LC/MS pos 0.50 0.1922 0.487186634 0.532425812 0.586759766 0.38086576 LC/MS pos 0.67 0.0313 0.802058943 0.57642665 0.698337752 0.59090117 eicosadienoylglycerophospho choline* LC/MS pos 0.34 < 0.001 0.462485184 0.26758025 0.294143435 0.31700466 eicosatrienoylglycerophospho choline* arachidonoylglycerophospho choline* 2- arachidonoylglycerophospho choline* docosapentaenoylglyceropho sphocholine* LC/MS pos 0.87 0.1213 0.828283288 0.894939331 0.996471418 0.77756277 LC/MS pos 0.56 0.0116 0.662811467 0.525792703 0.610041205 0.43878544 LC/MS pos 0.65 0.0631 0.638681117 0.611738666 0.774538238 0.55533506 LC/MS pos 0.75 0.0788 0.684424417 0.758796404 0.925733297 0.61552715 docosahexaenoylglycerophos phocholine*
Nucleotide Monoacylglycerol Sphingolipid Sterol/Steroid Purine, (hypo)xanthine/ino Purine, adenine containing Purine, guanine containing 2- LC/MS pos 0.51 0.0069 0.611607372 0.499202743 0.595353622 0.34660952 docosahexaenoylglycerophos phocholine* stearoylglycerophosphoinosit ol linoleoylglycerophosphoinosit ol* arachidonoylglycerophosphoi nositol* 2- arachidonoylglycerophosphoi nositol* LC/MS neg 0.87 0.9937 0.9711532 0.953222011 0.839896896 0.70182674 LC/MS neg 1.39 0.0725 1.685166618 1.125921557 1.493128871 1.24352698 LC/MS neg 0.84 0.3085 0.78980241 0.555016369 1.03489749 0.99096798 LC/MS neg 0.71 0.1357 0.660578905 0.517800087 0.703004178 0.96905073 LC/MS neg 1.14 0.3677 1.181719086 0.875654694 1.09632308 1.4017494 palmitoylplasmenylethanolam ine* palmitoylglycerol ( GC/MS 1.53 0.1595 1.666366197 2.198197183 1.177690141 1.07583099 monopalmitin) stearoylglycerol ( GC/MS 1.11 0.5239 0.884523216 1.449687997 1.091513038 1.00685695 monostearin) oleoylglycerol ( LC/MS pos 1.31 0.7417 1.152963141 2.918384338 0.865353079 0.29992179 monoolein) linoleoylglycerol ( LC/MS neg 2.52 0.1183 3.277799726 4.072329433 0.804497797 1.93952287 monolinolein) sphinganine LC/MS pos 0.92 0.9977 1.18141902 0.505937531 0.949106876 1.04670193 palmitoyl sphingomyelin GC/MS 0.90 0.7123 0.799049091 0.821870914 1.03810769 0.9572804 stearoyl sphingomyelin GC/MS 0.90 0.9369 0.960454384 0.816566087 0.932812685 0.87478405 cholesterol GC/MS 0.97 0.9116 0.92356855 0.984236759 1.090978466 0.87129468 dihydrocholesterol GC/MS 1.06 0.5302 1.062787136 1.049479854 1.121191319 1.02402704 corticosterone LC/MS pos 3.79 0.0570 5.212312333 4.060464797 0.885171728 5.00582711 7-alpha-hydroxy-3-oxo-4- cholestenoate (7-Hoca) LC/MS neg 0.73 0.1427 0.723050063 0.511754436 0.795160177 0.8721426 xanthine GC/MS 0.20 0.4618 0.117475918 0.117475918 0.354804946 0.2299136 xanthosine LC/MS pos 0.24 0.0934 0.199848012 0.199848012 0.362507478 0.19984801 adenosine LC/MS pos 1.03 0.9670 1.55475344 0.716910468 0.716910468 1.11835428 Nmethyladenosine LC/MS pos 0.93 0.6065 0.888787209 0.490505605 1.155901268 1.18904904 Nmethylguanosine LC/MS pos 0.88 0.5727 0.802150789 1.063237873 0.710789012 0.94111046 N6- carbamoylthreonyladenosine LC/MS pos 0.88 0.3350 0.86120682 0.791589363 1.058927467 0.81827017 Purine, urate Pyrimidine, cytidine containing urate LC/MS neg 0.33 0.0801 0.366055339 0.196764527 0.457744231 0.31691569 allantoin GC/MS 1.72 0.0543 2.021170983 1.497518091 2.313139166 1.06022367 cytidine LC/MS pos 0.63 0.1196 1.050728095 0.600649719 0.296962675 0.56711874 2'-deoxycytidine LC/MS pos 1.31 0.2851 0.939019262 1.105447613 2.070821768 1.13752208
Cofactors and vitamins Xenobiotics Pyrimidine, orotate Pyrimidine containing, thymine Pyrimidine containing, uracil containing Purine and pyrimidine Ascorbate and aldarate Hemoglobin and porphyrin Nicotinate and nicotinamide Pantothenate and CoA Pyridoxal Riboflavin Tocopherol Vitamin B6 Benzoate orotate GC/MS 2.35 0.1308 1.742738149 1.861686606 4.833063639 0.97779664 thymidine LC/MS neg 2.09 0.0795 1.375857003 1.211067581 3.777424094 2.00097943 uracil GC/MS 0.69 0.5369 1.162459257 0.509682367 0.545599795 0.53761104 5,6-dihydrouracil GC/MS 1.06 0.6694 1.349107358 0.915285993 1.225984315 0.76893017 uridine LC/MS neg 0.54 0.1157 0.541964444 0.439371567 0.596597956 0.58466718 pseudouridine LC/MS neg 1.23 0.1474 1.233019967 1.01888326 1.509143616 1.14849232 2'-deoxyuridine LC/MS neg 1.64 0.2246 1.022771909 0.951749031 2.925845321 1.64615956 methylphosphate GC/MS 1.01 0.8353 1.210685009 1.040987468 0.903420334 0.90040516 gulono-1,4-lactone GC/MS 1.00 0.7859 1.897048219 0.959735759 0.726012421 0.42464702 ascorbate (Vitamin C) GC/MS 0.87 0.5383 0.859584965 0.663969013 1.095276221 0.87152966 threonate GC/MS 0.58 0.0757 0.862376398 0.341420363 0.596963311 0.50294483 glucurono-6,3-lactone LC/MS neg 4.59 0.0422 7.114599011 7.653264531 2.052318498 1.54088726 heme* LC/MS pos 0.29 0.0532 0.690615982 0.072956797 0.119756052 0.2892683 L-urobilin LC/MS pos 1.00 1 1 1 1 bilirubin (E,E)* LC/MS pos 1.38 0.4426 0.560788213 1.589321763 1.876525355 1.47844165 biliverdin LC/MS pos 0.57 0.4053 0.496103058 0.9451661 0.471875913 0.35867243 nicotinamide GC/MS 0.92 0.6294 1.681843464 0.789829162 0.662932062 0.55216528 pantothenate LC/MS pos 0.98 0.8113 1.023812988 0.910519424 1.148649304 0.8419419 pyridoxal LC/MS pos 1.36 0.0549 1.559093395 1.029106155 1.588624901 1.26122623 flavin adenine dinucleotide LC/MS neg 1.11 0.4676 1.081953162 1.306511382 1.113010159 0.93070156 (FAD) riboflavin (Vitamin B2) LC/MS pos 0.87 0.5881 0.860225141 0.845386321 1.037352891 0.73588607 alpha-tocopherol GC/MS 1.16 0.2227 1.122384381 1.19700832 1.227373316 1.07801339 pyridoxate LC/MS neg 0.68 0.2457 1.00885391 0.419366172 0.617314314 0.69446982 hippurate LC/MS neg 0.76 0.7304 0.918879891 0.558493512 0.744892185 0.81623573 catechol sulfate LC/MS neg 1.28 0.6572 1.703544237 2.039170386 0.814071935 0.5737989 benzoate GC/MS 0.72 0.3812 0.169814381 0.187240748 1.254698399 1.28612147 4-hydroxybenzoate GC/MS 0.45 0.1833 0.518444049 0.337792901 0.174155131 0.77401593 methyl-4-hydroxybenzoate GC/MS 0.69 0.2096 0.355751562 0.603802781 0.412309716 1.37766688 Chemical Food component/plant Sugar, sugar substitute, starch glycolate (hydroxyacetate) GC/MS 0.99 0.8554 0.980037588 0.697617717 1.147559303 1.11799665 glycerol 2-phosphate GC/MS 0.93 0.6855 0.47136254 0.543207395 1.426145498 1.29732717 trizma acetate GC/MS 1.75 0.9139 0.218988817 6.524830534 0.165312379 0.08220716 2-ethylhexanoate GC/MS 0.67 0.3921 0.120926261 0.164921626 0.969524468 1.42476671 tartarate GC/MS 0.80 0.6076 1.629809199 0.520365301 0.706944473 0.3457399 ergothioneine LC/MS pos 1.34 0.5570 2.314023048 1.512883594 0.777625275 0.74188787 erythritol GC/MS 0.93 0.5943 1.006975082 0.618440835 1.220318931 0.88423336
Supplemental Table 3. Evidence of metabolic perturbation in IVFKAA female liver, muscle, gonadal fat, and pancreatic islets at 29wks of age. The table shows canonical pathways significantly altered in IVFKAA tissues relative to FB controls as determined by Ingenuity Pathway Analysis (p<0.05, Fisher s exact test). Pathway p-value # Genes Gonadal Fat LXR/RXR Activation 0.0001 10 ALB,MLXIPL,APOB,MSR1,APOH,APOC3,APOF,SERPINA1,FGA,GC Atherosclerosis Signaling 0.0004 9 ALB,APOB,MSR1,SELP,APOC3,APOF,PLA2G5,SERPINA1,PDGFB Ephrin Receptor Signaling 0.0006 11 ROCK2,EPHB4,AKT2,SDCBP,GNA11,GNB5,ABL1,GNB1L,RASA1,PDGFB,GNG7 Nicotine Degradation III 0.0006 5 CYP2D6,CYP1A2,CYP4B1,AOX1,CYP2C18 Estrogen Biosynthesis 0.0006 5 CYP2D6,CYP1A2,CYP4B1,CYP2C18,AKR1C4 Bupropion Degradation 0.0010 4 CYP2D6,CYP1A2,CYP4B1,CYP2C18 Acetone Degradation I (to Methylglyoxal) 0.0012 4 CYP2D6,CYP1A2,CYP4B1,CYP2C18 Nicotine Degradation II 0.0015 5 CYP2D6,CYP1A2,CYP4B1,AOX1,CYP2C18 RhoGDI Signaling 0.0020 10 ROCK2,CDH5,GNA11,GNB5,CD44,MYL4,ARHGEF1,ARHGDIA,GNB1L,GNG7 Tyrosine Degradation I 0.0023 2 HPD,FAH Superpathway of Methionine Degradation 0.0029 4 BHMT,SUV39H2,MAT1A,PCCB Ephrin B Signaling 0.0030 6 ROCK2,EPHB4,GNA11,GNB5,GNB1L,GNG7 G Protein Signaling Mediated by Tubby 0.0033 4 GNB5,ABL1,GNB1L,GNG7 Extrinsic Prothrombin Activation Pathway 0.0037 3 FGB,FGA,FGG TREM1 Signaling 0.0041 5 AKT2,TYROBP,CASP1,LAT2,CD83 Coagulation System 0.0046 4 SERPINA1,FGB,FGA,FGG Melatonin Degradation I 0.0051 4 CYP2D6,CYP1A2,CYP4B1,CYP2C18 Superpathway of Melatonin Degradation 0.0075 4 CYP2D6,CYP1A2,CYP4B1,CYP2C18 IL-12 Signaling and Production in Macrophages 0.0084 7 ALB,AKT2,APOB,APOC3,APOF,SERPINA1,REL CXCR4 Signaling 0.0089 8 ROCK2,AKT2,ADCY4,GNA11,GNB5,MYL4,GNB1L,GNG7 TCA Cycle II (Eukaryotic) 0.0093 3 CS,ACO2,MDH2 Genes
Gluconeogenesis I 0.0105 3 GPI,ALDOB,MDH2 Thrombin Signaling 0.0109 9 ROCK2,AKT2,ADCY4,GNA11,GNB5,MYL4,ARHGEF1,GNB1L,GNG7 Telomerase Signaling 0.0110 6 ETS1,AKT2,ABL1,ETS2,ELK3,HDAC5 Cardiac β-adrenergic Signaling 0.0130 7 PDE2A,ADCY4,GNB5,GNB1L,PPP1CA,GNG7,AKAP11 Intrinsic Prothrombin Activation Pathway 0.0148 3 FGB,FGA,FGG Relaxin Signaling 0.0153 7 PDE2A,AKT2,ADCY4,GNA11,GNB5,GNB1L,GNG7 Sphingosine-phosphate Signaling 0.0168 6 S1PR3,AKT2,ADCY4,CASP1,PDGFB,ASAH1 EPHB4,AKT2,GNA11,GNB5,ABL1,GIT1,PDGFB,GNG7,ROCK2,FZD4,SDCBP,MYL4,CHMP1A,GN Axonal Guidance Signaling 0.0194 15 B1L,RASA1 Production of Nitric Oxide and Reactive Oxygen Species in Macrophages 0.0201 8 ALB,AKT2,APOB,APOC3,APOF,CAT,SERPINA1,PPP1CA α-adrenergic Signaling 0.0233 5 GYS1,ADCY4,GNB5,GNB1L,GNG7 Gαq Signaling 0.0244 7 ROCK2,AKT2,GYS1,GNA11,GNB5,GNB1L,GNG7 FXR/RXR Activation 0.0245 5 AKT2,MLXIPL,APOB,APOC3,FETUB Huntington's Disease Signaling 0.0247 9 AKT2,ATP5B,GNA11,CASP1,GNB5,GNB1L,RASA1,GNG7,HDAC5 NAD biosynthesis II (from tryptophan) 0.0270 2 TDO2,ABL1 Chondroitin Sulfate Degradation (Metazoa) 0.0270 2 CD44,ARSB Gα12/13 Signaling 0.0273 6 ROCK2,AKT2,CDH5,MYL4,ARHGEF1,RASA1 G Beta Gamma Signaling 0.0280 5 AKT2,GNA11,GNB5,GNB1L,GNG7 Gαi Signaling 0.0294 6 S1PR3,APLNR,ADCY4,GNB5,GNB1L,GNG7 CCR5 Signaling in Macrophages 0.0305 4 CCR5,GNB5,GNB1L,GNG7 Leukocyte Extravasation Signaling 0.0306 8 ROCK2,F11R,CDH5,CD44,ABL1,VCL,MMP24,MMP12 Breast Cancer Regulation by Stathmin1 0.0306 8 ROCK2,CCNE2,ADCY4,GNB5,ARHGEF1,GNB1L,PPP1CA,GNG7 Retinol Biosynthesis 0.0326 3 Ces1b/Ces1c,LIPE,PNPLA2 IL-1 Signaling 0.0333 5 ADCY4,GNA11,GNB5,GNB1L,GNG7 Methionine Degradation I (to Homocysteine) 0.0355 2 SUV39H2,MAT1A
Docosahexaenoic Acid (DHA) Signaling 0.0376 3 AKT2,BCL2A1,PNPLA2 Acute Phase Response Signaling 0.0414 7 ALB,AKT2,APOH,SERPINA1,FGB,FGA,FGG Insulin Receptor Signaling 0.0414 6 AKT2,GYS1,LIPE,IRS2,VAMP2,PPP1CA Ethanol Degradation IV 0.0448 2 CAT,ACSS2 Cysteine Biosynthesis III (mammalia) 0.0448 2 SUV39H2,MAT1A DNA damage-induced 14-3-3σ Signaling 0.0497 2 CCNE2,AKT2 Pathway p-value # Genes Liver Genes Nur77 Signaling in T Lymphocytes 0.0004 8 CASP9,HDAC2,CYCS,MEF2D,CHP1,FCER1G,NFATC1,PPP3CC Antigen Presentation Pathway 0.0006 6 HLA-C,HLA-B,CD74,TAP1,TAPBP,MR1 Complement System 0.0007 6 CFD,SERPING1,MASP2,C4B (includes others),c1qc,c3ar1 Spermine Biosynthesis 0.0016 2 SMS,AMD1 Protein Ubiquitination Pathway 0.0042 19 UCHL3,DNAJB4,UBE2L3,CUL1,DNAJC19,DNAJB9,DNAJB14,TAP1,DNAJC21,PAN2,PSMC6,CUL 2,HLA-C,PSMD12,USP16,HLA-B,USP46,SUGT1,DNAJB1 Spermidine Biosynthesis I 0.0047 2 SMS,AMD1 Role of PKR in Interferon Induction and Antiviral Response 0.0049 6 IKBKB,CASP9,CYCS,TAB2,NFKB2,IRF1 LXR/RXR Activation 0.0054 11 KNG1,PON1,ITIH4,SAA1,VTN,MYLIP,CLU,C4B (includes others),pltp,nfkb2,il1rap Assembly of RNA Polymerase II Complex 0.0088 6 TAF9,CCNH,CDK7,DR1,TAF9B,POLR2K Bile Acid Biosynthesis, Neutral Pathway 0.0108 3 BAAT,CYP27A1,AKR1C4 Methylglyoxal Degradation III 0.0108 3 Akr1b10,AKR1C4,AKR1B1 Melatonin Degradation I 0.0123 5 Cyp2c40 (includes others),ugt2b28,ugt2b15,cyp2c18,sult1d1 icos-icosl Signaling in T Helper Cells 0.0170 9 GAB2,SHC1,IKBKB,CHP1,FCER1G,NFATC1,PPP3CC,NFKB2,INPP5D
Type I Diabetes Mellitus Signaling 0.0180 9 IKBKB,CASP9,CYCS,HLA-C,HLA-B,FCER1G,NFKB2,IL1RAP,IRF1 Superpathway of Melatonin Degradation 0.0192 5 Cyp2c40 (includes others),ugt2b28,ugt2b15,cyp2c18,sult1d1 Thyroid Hormone Metabolism II (via Conjugation and/or Degradation) 0.0205 3 UGT2B28,UGT2B15,Sult1d1 Systemic Lupus Erythematosus Signaling 0.0236 12 KNG1,RNU12,SNRPE,HLA-C,RNU2-1,CREM,HLA-B,FCER1G,ZCRB1,NFATC1,HNRNPC,INPP5D Tryptophan Degradation to 2-amino- 3-carboxymuconate Semialdehyde 0.0296 2 TDO2,HAAO Mitochondrial Dysfunction 0.0321 10 NDUFA5,COX7A2,CASP9,CYCS,COX6C,NDUFB4,NDUFB6,UQCRB,GPX4,NDUFS4 Role of JAK2 in Hormone-like Cytokine Signaling 0.0381 4 SH2B3,SHC1,EPOR,SIRPA Cytotoxic T Lymphocyte-mediated Apoptosis of Target Cells 0.0395 5 CASP9,CYCS,HLA-C,HLA-B,FCER1G Asparagine Degradation I 0.0402 1 ASPG L-cysteine Degradation II 0.0402 1 CTH TWEAK Signaling 0.0421 4 IKBKB,CASP9,CYCS,NFKB2 Mineralocorticoid Biosynthesis 0.0482 2 HSD3B2,Cyp2d22 UDP-N-acetyl-D-galactosamine Biosynthesis II 0.0482 2 GNPNAT1,GNPDA1 Pathway p-value # Genes Skeletal Muscle Hypoxia Signaling in the Cardiovascular System 0.0098 6 UBE2A,SUMO1,UBE2V2,HIF1A,NOS3,UBE2I Selenocysteine Biosynthesis II (Archaea and Eukaryotes) 0.0125 2 SEPHS2,PSTK UDP-N-acetyl-D-galactosamine Biosynthesis II 0.0282 2 GNPNAT1,GNPDA2 Genes
Folate Transformations I 0.0282 2 MTHFS,MTHFR RhoGDI Signaling 0.0295 10 ROCK2,ARHGAP5,MYL9,GNAI3,CDH4,RHOB,ARPC5,GDI2,RHOJ,GNG5 Glutamate Removal from Folates 0.0301 1 GGH Xanthine and Xanthosine Salvage 0.0301 1 PNP CXCR4 Signaling 0.0310 9 ROCK2,MYL9,GNAI3,FOS,RHOB,ADCY4,MAPK8,RHOJ,GNG5 Glucocorticoid Receptor Signaling 0.0333 13 CCNH,SGK1,MAPK8,BGLAP,TBP,GTF2E2,TAF7,CD163,FOS,DUSP1,SUMO1,CHUK,UBE2I Sphingosine-phosphate Signaling 0.0340 7 PLCD3,GNAI3,CASP12,RHOB,ADCY4,RHOJ,CASP8 Cell Cycle Control of Chromosomal Replication 0.0420 3 ORC2,RPA3,ORC4 Assembly of RNA Polymerase II Complex 0.0456 4 CCNH,TBP,GTF2E2,TAF7 Pathway p-value # Genes D-myo-inositol (1,4,5,6)- Tetrakisphosphate Biosynthesis 0.0001 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 D-myo-inositol (3,4,5,6)- tetrakisphosphate Biosynthesis 0.0001 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 D-myo-inositol-5-phosphate Metabolism 0.0002 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 3-phosphoinositide Degradation 0.0002 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 3-phosphoinositide Biosynthesis 0.0003 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 Superpathway of Inositol Phosphate Compounds 0.0009 6 TNS3,NUDT11,PPP1R12A,ILKAP,PTEN,NUDT1 Lipoate Biosynthesis and Incorporation II 0.0129 1 LIAS Role of Hypercytokinemia/hyperchemokine mia in the Pathogenesis of Influenza 0.0131 2 IL1B,IFNA1/IFNA13 Pancreatic Islets Genes
Systemic Lupus Erythematosus Signaling 0.0184 4 AKT2,SNRPB2,IL1B,IFNA1/IFNA13 Oxidized GTP and dgtp Detoxification 0.0193 1 NUDT1 L-serine Degradation 0.0193 1 SDSL EIF2 Signaling 0.0196 4 AKT2,RPL30,RPL12,RPSA Role of Cytokines in Mediating Communication between Immune Cells 0.0214 2 IL1B,IFNA1/IFNA13 Docosahexaenoic Acid (DHA) Signaling 0.0238 2 AKT2,IL1B ILK Signaling 0.0249 4 AKT2,PPP1R12A,ILKAP,PTEN Melanoma Signaling 0.0301 2 AKT2,PTEN Role of Tissue Factor in Cancer 0.0313 3 AKT2,IL1B,PTEN Integrin Signaling 0.0353 4 AKT2,PPP1R12A,ILKAP,PTEN Role of PI3K/AKT Signaling in the Pathogenesis of Influenza 0.0415 2 AKT2,IFNA1/IFNA13 Endometrial Cancer Signaling 0.0445 2 AKT2,PTEN TREM1 Signaling 0.0461 2 AKT2,IL1B PI3K Signaling in B Lymphocytes 0.0464 3 CD81,AKT2,PTEN Cardiac β-adrenergic Signaling 0.0473 3 PPP1R12A,GNG4,AKAP11 Insulin Receptor Signaling 0.0482 3 AKT2,PPP1R12A,PTEN