S-LDSC Analysis of IBD

I used Stratified Linkage Disequilibrium Score Regression (S-LDSC) to analyze summary statistics from a GWAS of inflammatory bowel disease from Liu et al. For the current analysis, I focused only European participants.

I used the standard reference data sources to associate chromatin regions with cell or tissue types. These reference data sources were:

• The GTEx Project
• The Franke lab dataset
• The Roadmap Epigenetic Project
• The Corces et al. ATAC-seq dataset of 13 blood cell types.
• The ImmGen Project
• The Cahoy Mouse Central Nervous System Dataset

Results

GTEx and Franke lab tissue expression data

The plot below illustrates the pattern of S-LDSC p values across cell types. The table shows the cell types with the lowest p values.

Name	Coefficient	Coefficient_P_value	Reject Null
A02.835.583.443.800.800.Synovial.Fluid	7.21343e-08	5.86657e-07	True
A15.145.Blood	7.55841e-08	6.92179e-07	True
Whole_Blood	5.70557e-08	7.05793e-07	True
A11.118.637.555.567.569.T.Lymphocytes	6.95136e-08	9.21119e-07	True
A15.145.229.Blood.Cells	7.04224e-08	2.83664e-06	True
A11.118.637.Leukocytes	6.44595e-08	1.21546e-05	True
A15.382.490.555.567.Lymphocytes	6.05795e-08	1.25194e-05	True
A15.382.520.604.700.Spleen	5.3496e-08	2.10229e-05	True
A15.145.229.637.555.Leukocytes..Mononuclear	6.53729e-08	2.13756e-05	True
Lung	3.36683e-08	7.06633e-05	True
A11.118.637.555.567.569.200.700.T.Lymphocytes..Regulatory	4.73932e-08	0.000110028	True
A15.378.316.Bone.Marrow.Cells	5.04105e-08	0.000112991	True
Spleen	4.95452e-08	0.000114937	True
A15.382.490.555.567.537.Killer.Cells..Natural	5.06096e-08	0.000275746	True
A10.549.Lymphoid.Tissue	4.5808e-08	0.00027864	True
A15.382.812.260.Dendritic.Cells	4.13252e-08	0.000439731	True
A15.378.316.580.Monocytes	3.77876e-08	0.000972786	False
A10.549.400.Lymph.Nodes	3.91855e-08	0.00113796	False

Interestingly, no gastrointestinal tissue/cell types are significant in this analysis. Instead, the significant tissue/cell types overwhelmingly pertain to the blood or immune system, consistent with the autoimmune nature of IBD.

The significance of lung tissue may reflect sample contamination: as noted by Finucane et al.¹, there was likely blood in the lungs of the post-mortem donors from which the GTEx samples were extracted.

Roadmap Chromatin data

I next applied S-LDSC to the IBD GWAS using reference data generated by Finucane et al.¹ from the Roadmap Epigenetics Project

The results are shown in the graph and table below.

Name	Coefficient	Coefficient_P_value	Reject Null
SI-Term-Ileum_ENTEX__H3K27ac	5.88187e-07	7.43564e-11	True
Primary_T_helper_17_cells_PMA-I_stimulated__H3K4me1	6.64904e-07	7.46228e-11	True
Spleen_ENTEX__H3K27ac	3.68515e-07	1.06777e-09	True
Primary_Natural_Killer_cells_from_peripheral_blood__H3K4me1	4.7694e-07	2.18101e-08	True
Primary_T_helper_17_cells_PMA-I_stimulated__H3K27ac	1.11335e-06	2.54249e-08	True
Spleen_ENTEX__H3K4me1	1.33146e-06	3.25759e-08	True
Primary_T_helper_17_cells_PMA-I_stimulated__H3K4me3	1.80868e-06	8.50234e-08	True
Primary_T_helper_memory_cells_from_peripheral_blood_2__H3K4me1	4.64748e-07	1.34853e-07	True
Primary_T_helper_memory_cells_from_peripheral_blood_1__H3K4me1	4.72349e-07	1.51893e-07	True
Sigmoid_Colon__H3K27ac	8.40826e-07	4.7277e-07	True
Primary_T_cells_from_peripheral_blood__H3K4me1	4.22743e-07	5.38612e-07	True
Rectal_Mucosa_Donor_29__H3K9ac	1.76888e-06	6.48073e-07	True
Primary_Natural_Killer_cells_from_peripheral_blood__DNase	1.29915e-06	6.731e-07	True
Primary_T_helper_cells_PMA-I_stimulated__H3K4me1	3.5459e-07	1.09551e-06	True
Spleen_ENTEX__H3K4me3	5.57333e-07	1.22639e-06	True
Primary_T_killer_memory_cells_from_peripheral_blood__H3K4me1	4.83526e-07	1.40831e-06	True
Primary_T_regulatory_cells_from_peripheral_blood__H3K4me1	4.89971e-07	1.86403e-06	True
Primary_T_cells_from_peripheral_blood__DNase	9.94548e-07	2.40567e-06	True
Small_Intestine__H3K27ac	7.45908e-07	2.50416e-06	True
Primary_T_helper_cells_from_peripheral_blood__H3K4me1	3.77567e-07	3.48901e-06	True
Rectal_Mucosa_Donor_29__H3K27ac	6.65295e-07	4.27993e-06	True
Primary_T_cells_effector_memory_enriched_from_peripheral_blood__H3K4me1	6.57463e-07	4.43318e-06	True
Primary_Natural_Killer_cells_from_peripheral_blood__H3K36me3	3.91359e-07	5.88324e-06	True
Primary_B_cells_from_cord_blood__H3K4me1	3.63687e-07	6.66304e-06	True
Primary_T_cells_from_cord_blood__DNase	1.11004e-06	6.88901e-06	True
Primary_T_killer_naive_cells_from_peripheral_blood__H3K4me1	3.26511e-07	8.15872e-06	True
Primary_B_cells_from_peripheral_blood__DNase	6.63583e-07	9.29237e-06	True
Primary_T_cells_from_peripheral_blood__H3K27ac	5.92792e-07	1.11938e-05	True
Colonic_Mucosa__H3K27ac	7.41473e-07	1.45139e-05	True
Primary_T_helper_naive_cells_from_peripheral_blood_1__H3K4me1	3.7948e-07	2.28038e-05	True
Primary_Natural_Killer_cells_from_peripheral_blood__H3K27ac	5.91881e-07	2.33956e-05	True
Primary_T_helper_memory_cells_from_peripheral_blood_1__H3K4me3	1.22213e-06	2.97356e-05	True
Spleen_ENTEX__H3K36me3	1.6328e-07	3.59535e-05	True
Primary_T_helper_cells_PMA-I_stimulated__H3K27ac	4.39121e-07	4.74836e-05	True
Primary_monocytes_from_peripheral_blood__H3K4me1	2.44401e-07	5.10902e-05	True
Stomach_ENTEX__H3K27ac	2.52371e-07	5.36711e-05	True
Primary_monocytes_from_peripheral_blood__DNase	9.03028e-07	6.30244e-05	True
Primary_T_helper_cells_PMA-I_stimulated__H3K4me3	9.27811e-07	7.92555e-05	True
Primary_B_cells_from_peripheral_blood__H3K4me1	2.61357e-07	8.03756e-05	True
Primary_T_killer_naive_cells_from_peripheral_blood__H3K9ac	8.23659e-07	0.000106447	True
Primary_hematopoietic_stem_cells_short_term_culture__H3K4me1	3.07194e-07	0.000117984	True
Primary_hematopoietic_stem_cells__H3K4me1	5.61429e-07	0.000124149	True
SI-Term-Ileum_ENTEX__H3K4me3	9.26661e-07	0.000127295	True
Duodenum_Mucosa__H3K9ac	9.40654e-07	0.000129292	True
Primary_T_helper_17_cells_PMA-I_stimulated__H3K36me3	3.36529e-07	0.000131691	True
Rectal_Mucosa_Donor_31__H3K27ac	4.86019e-07	0.000208176	True
Primary_T_helper_memory_cells_from_peripheral_blood_1__H3K27ac	6.57625e-07	0.000311747	True
Primary_T_regulatory_cells_from_peripheral_blood__H3K27ac	3.87157e-07	0.000400567	True
Stomach_ENTEX__H3K4me3	8.02303e-07	0.000434599	True
SI-Term-Ileum_ENTEX__H3K4me1	5.14523e-07	0.000449404	True
Primary_T_helper_cells_PMA-I_stimulated__H3K36me3	2.40795e-07	0.00044978	True
Primary_mononuclear_cells_from_peripheral_blood__H3K4me1	1.03073e-06	0.000582293	True
Rectal_Mucosa_Donor_31__H3K9ac	9.00682e-07	0.000614263	True
Primary_T_helper_memory_cells_from_peripheral_blood_2__H3K4me3	1.09624e-06	0.000697011	True
Primary_T_cells_from_cord_blood__H3K4me1	4.83073e-07	0.000734568	True
Colonic_Mucosa__H3K9ac	9.28271e-07	0.000768319	True
Sigmoid_Colon__H3K36me3	4.13158e-07	0.000776354	True
Primary_T_cells_effector_memory_enriched_from_peripheral_blood__H3K27ac	5.63262e-07	0.000930381	True
Primary_B_cells_from_peripheral_blood__H3K27ac	3.84286e-07	0.000958017	True
Primary_T_cells_effector_memory_enriched_from_peripheral_blood__H3K4me3	1.00685e-06	0.000989186	True
Sigmoid_Colon__H3K4me3	9.21358e-07	0.0013546	False
Rectal_Mucosa_Donor_29__H3K36me3	3.3545e-07	0.00152473	False
Primary_Natural_Killer_cells_from_peripheral_blood__H3K4me3	1.38162e-06	0.00166122	False
Primary_B_cells_from_cord_blood__H3K4me3	1.27467e-06	0.00178815	False
Duodenum_Mucosa__H3K36me3	2.54483e-07	0.00196143	False
Rectal_Mucosa_Donor_29__H3K4me3	7.34082e-07	0.00200658	False
Primary_B_cells_from_peripheral_blood__H3K36me3	1.49602e-07	0.00208217	False
Primary_neutrophils_from_peripheral_blood__H3K4me3	4.61673e-07	0.0022148	False

Of note, in contrast the analysis using gene-expression data, in the present epigenetics analysis, both immune-related and gastrointestinal-related cell types are significant, consistent with our understanding of the disease process. This may reflect differences in the specificity of gene expression versus epigenetic reference data.

ImmGen Data

The next step is to use the S-LDSC reference data derived from the ImmGen project. The results are below

Name	Coefficient	Coefficient_P_value	Reject Null
LN.TR.14w.B6	5.88377e-08	1.45162e-06	True
T.4Mem.Sp	5.91919e-08	6.75222e-06	True
T.4Mem44h62l.Sp	5.42097e-08	7.67268e-06	True
NKT.4+.Lv	5.21078e-08	8.54374e-06	True
T.8SP69+.Th	4.98414e-08	8.69673e-06	True
T.4.Pa.BDC	5.27413e-08	1.02889e-05	True
NKT.4-.Lv	5.8454e-08	1.22672e-05	True
CD4Control	5.34947e-08	1.45635e-05	True
NK.b2m-.Sp	4.04081e-08	1.63948e-05	True
NKT.4-.Sp	5.15357e-08	1.71259e-05	True
Tgd.vg2+24alo.Th	4.52212e-08	2.66825e-05	True
T.4Nve.LN	5.42156e-08	2.80202e-05	True
T.4SP24-.Th	4.91772e-08	2.88841e-05	True
T.4.LN.BDC	5.56221e-08	3.43062e-05	True
T.4Nve.Sp	5.44986e-08	5.18253e-05	True
NKT.44+NK1.1+.Th	5.08155e-08	5.95247e-05	True
T.4Mem44h62l.LN	5.41904e-08	6.65735e-05	True
T.4FP3+25+.Sp	4.24075e-08	9.1964e-05	True
T.4Nve.MLN	4.42292e-08	0.000128944	True
T.4Mem.LN	4.91063e-08	0.000130168	True
T.4int8+.Th	4.08383e-08	0.000143531	True
T.4SP69+.Th	4.46602e-08	0.000178913	True
T.4SP24int.Th	4.48964e-08	0.000244919	True
B.MZ.Sp	4.36416e-08	0.00024541	True
ABD.TR.14w.B6	4.15646e-08	0.000284172	True
B.T2.Sp	4.33163e-08	0.000293517	True
B.FrF.BM	4.12463e-08	0.000299895	True
T.8SP24-.Th	4.50035e-08	0.000338027	True
B.Fo.PC	3.803e-08	0.000345648	True
NK.49H-.Sp	4.17266e-08	0.000367327	True
NKT.44-NK1.1-.Th	3.7553e-08	0.000629513	True
B.Fo.LN	3.84346e-08	0.000632388	True
CD4.96h.LN	3.84236e-08	0.000720856	True
T.8Nve.LN	4.29127e-08	0.000723785	True
DC.4+.Sp.ST	3.47431e-08	0.00075239	True
Tgd.vg5-.act.IEL	3.8971e-08	0.00078216	True
T.8SP24int.Th	3.47183e-08	0.000788441	True
CD4.CTR.LN	3.75766e-08	0.00084666	True
DC.8-4-11b+.MLN	3.65043e-08	0.000933421	True
NK.H+.MCMV1.Sp	3.88649e-08	0.000943415	True
T.4.PLN.BDC	4.0467e-08	0.00096887	True
CD8.5h.LN	3.67386e-08	0.000995926	True
T.8Nve.MLN	4.05e-08	0.00113565	True
CD4.5h.LN	4.06854e-08	0.00119258	True
DC.8-4-11b-.SLN	3.63301e-08	0.00125689	True
T.4FP3-.Sp	3.66733e-08	0.00137885	True
B.T1.Sp	3.74316e-08	0.00148824	True
B.T3.Sp	3.41897e-08	0.00172218	False

Overall these results are rather non-specific: there are significant cell types in all the categories. However, the lowest p values are associated with T and natural-killer cell types (and NK cells have broadly similar gene expression patterns to T cells). This finding is consistent with the view of inflammatory bowel disease as primarily T-cell driven

Corces et al. ATAC-seq data

Looking at the Corces et al. ATAC-seq dataset, we again see broad significance with T and NK cell attaining the lowest p values:

Name	Coefficient	Coefficient_P_value	Reject Null
CD8	8.38248e-07	3.26601e-07	True
CD4	7.77934e-07	5.8346e-06	True
NK	7.14325e-07	1.23519e-05	True
Bcell	5.69139e-07	3.08321e-05	True
Mono	8.27562e-07	4.07921e-05	True
HSC	2.69509e-07	0.00922097	False
MPP	2.37462e-07	0.0106805	False
LMPP	2.71358e-07	0.0182358	False

Cahoy and GTEx-Brain data

As expected, there are no significant cell types in these datasets, consistent with IBD being a non-neural disease.

Name	Coefficient	Coefficient_P_value	Reject Null
Oligodendrocyte	3.8018e-09	0.350595	False
Neuron	-1.63365e-09	0.57887	False
Astrocyte	-4.33476e-09	0.678332	False

Name	Coefficient	Coefficient_P_value	Reject Null
Brain_Spinal_cord_(cervical_c-1)	5.27018e-09	0.255	False
Brain_Cerebellum	3.8986e-09	0.332546	False
Brain_Amygdala	2.29904e-09	0.364457	False
Brain_Putamen_(basal_ganglia)	2.49498e-09	0.365589	False
Brain_Frontal_Cortex_(BA9)	2.41676e-09	0.372215	False
Brain_Cortex	2.0825e-09	0.392953	False
Brain_Anterior_cingulate_cortex_(BA24)	1.6171e-09	0.408329	False
Brain_Cerebellar_Hemisphere	-1.74109e-09	0.582644	False

Reproducing

To reproduce, run the IBD analysis script here.

---

1. Hilary K Finucane, Yakir A Reshef, Verneri Anttila, Kamil Slowikowski, Alexander Gusev, Andrea Byrnes, Steven Gazal, Po-Ru Loh, Caleb Lareau, Noam Shoresh, and others. Heritability enrichment of specifically expressed genes identifies disease-relevant tissues and cell types. Nature Genetics, 50(4):621–629, 2018. URL: https://pmc.ncbi.nlm.nih.gov/articles/PMC5896795/. ↩↩