Common patterns of gene regulation associated with Cesarean section and the development of islet autoimmunity - indications of immune cell activation

Abstract

Birth by Cesarean section increases the risk of developing type 1 diabetes later in life. We aimed to elucidate common regulatory processes observed after Cesarean section and the development of islet autoimmunity, which precedes type 1 diabetes, by investigating the transcriptome of blood cells in the developing immune system. To investigate Cesarean section effects, we analyzed longitudinal gene expression profiles from peripheral blood mononuclear cells taken at several time points from children with increased familial and genetic risk for type 1 diabetes. For islet autoimmunity, we compared gene expression differences between children after initiation of islet autoimmunity and age-matched children who did not develop islet autoantibodies. Finally, we compared both results to identify common regulatory patterns. We identified the pentose phosphate pathway and pyrimidine metabolism - both involved in nucleotide synthesis and cell proliferation - to be differentially expressed in children born by Cesarean section and after islet autoimmunity. Comparison of global gene expression signatures showed that transcriptomic changes were systematically and significantly correlated between Cesarean section and islet autoimmunity. Moreover, signatures of both Cesarean section and islet autoimmunity correlated with transcriptional changes observed during activation of isolated CD4+ T lymphocytes. In conclusion, we identified shared molecular changes relating to immune cell activation in children born by Cesarean section and children who developed autoimmunity. Our results serve as a starting point for further investigations on how a type 1 diabetes risk factor impacts the young immune system at a molecular level.

Trial registration: ClinicalTrials.gov NCT01115621.

Figure 1

Overview. (a) Study workflow: Parallel analyses were performed to detect differential gene expression and pathway enrichment for Cesarean section and islet autoimmunity. The results were then compared in a combined analysis and related to expression patterns of lymphocyte activation. (b) Schematic overview of the longitudinal study design for Cesarean section and islet autoimmunity analyses. (c) Schematic illustration of the generalized additive mixed effect model (GAMM) to analyze the longitudinal dataset, including intercept, a time effect, a random effect for multiple measurements, and the investigated Cesarean section vs. vaginal delivery effect. Abbreviations: CS = Cesarean section, VD = Vaginal delivery, Ab+ = Islet autoantibody-positive, Ab− = Islet autoantibody-negative.

Figure 2

Cesarean section analysis. (a) Histogram of unadjusted p-values for CS association per gene. (b) Histogram of p-values after multiple testing adjustment by controlling the false discovery rate. (c) Sorted −log10(p-values) of pathway enrichment. Dashed line indicates the multiple testing threshold at an FDR of 0.1. Abbreviations: CS = Cesarean section, VD = Vaginal delivery.

Figure 3

Ab+ analysis. (a) Histogram of unadjusted p-values from single gene analysis of islet autoimmunity positive vs. age-matched children who did not develop islet autoimmunity. (b) Histogram of p-values after multiple testing, controlling the false discovery rate. (c) Sorted −log10(p-values) of pathway enrichment for islet autoimmunity status. Dashed line indicates the multiple testing threshold at an FDR of 0.1. Abbreviations: Ab+ = Islet autoantibody-positive, Ab− = Islet autoantibody negative.

Figure 4

Comparison of Cesarean section and islet autoimmunity signatures. (a) The pyrimidine metabolism pathway is shown as an example, with two-sided node coloring according to directed p-values (−log10(p) * sign(t-statistic)). Pathway image from KEGG. (b) T-statistics per gene from both analyses; the x-axis indicates results from islet autoimmunity status and the y-axis the results from Cesarean section vs. vaginal delivery. (c) Empirical distribution of correlation coefficients between Cesarean section and permuted class labels of islet autoimmunity status for 5,000 permutations. The red line indicates the ‘true’ correlation between the results from the analysis of Cesarean section and islet autoimmunity status. Black lines indicate the correlation between Cesarean section and multiple first-degree relatives (multiple FDR), maternal diabetes (MD), and gender. (d) Correlation of pathway directed p-values in Cesarean section analysis and islet autoimmunity status analysis. (e) Empirical distribution of correlation coefficients between Cesarean section and permuted class labels of islet autoimmunity status at pathway level for 1,000 permutations. The red line indicates the ‘true’ correlation between Cesarean section pathways and islet autoimmunity status pathways. Abbreviations: CS = Cesarean section, VD = Vaginal delivery, Ab+ = Islet autoantibody-positive, Ab− = Islet autoantibody negative.

Figure 5

Signatures of immune cell activation. (a) Correlation between single gene effects in Cesarean section (CS) compared to the association between naïve and activated CD4+ cells. (b) Histogram of correlation of permuted class labels of CD4 T cells and Cesarean section at the single gene level and the “true” correlation effect. (c) Correlation between single gene effects in islet autoimmunity status (Ab) compared to the association between naïve and activated CD4+ cells. (d) Histogram of correlation of permuted class labels of CD4 T cells and islet autoimmunity status at the single gene level and the “true” effect. (e) Correlation between pathway effects in Cesarean section compared to the association between naïve and activated pathways for CD4+ cells. (f) Histogram of correlation of permuted class labels of CD4 T cells and Cesarean section at the pathway level and the “true” effect. (g) Correlation between pathway effects in islet autoimmunity status compared to the association between naïve and activated pathways for CD4+ cells. (h) Histogram of correlation of permuted class labels of CD4 T cells and islet autoimmunity status at the pathway level and the “true” effect.