Interferon-γ converts human microvascular pericytes into negative regulators of alloimmunity through induction of indoleamine 2,3-dioxygenase 1

Abstract

Early acute rejection of human allografts is mediated by circulating alloreactive host effector memory T cells (TEM). TEM infiltration typically occurs across graft postcapillary venules and involves sequential interactions with graft-derived endothelial cells (ECs) and pericytes (PCs). While the role of ECs in allograft rejection has been extensively studied, contributions of PCs to this process are largely unknown. This study aimed to characterize the effects and mechanisms of interactions between human PCs and allogeneic TEM. We report that unstimulated PCs, like ECs, can directly present alloantigen to TEM, but while IFN-γ-activated ECs (γ-ECs) show increased ability to stimulate alloreactive T cells, IFN-γ-activated PCs (γ-PCs) instead suppress TEM proliferation but not cytokine production or signaling. RNA sequencing analysis of PCs, γ-PCs, ECs, and γ-ECs reveal induction of indoleamine 2,3-dioxygenase 1 (IDO1) in γ-PCs to significantly higher levels than in γ-ECs that correlates with tryptophan depletion in vitro. Consistently, shRNA knockdown of IDO1 markedly reduces γ-PC-mediated immunoregulatory effects. Furthermore, human PCs express IDO1 in a skin allograft rejection humanized mouse model and in human renal allografts with acute T cell-mediated rejection. We conclude that immunosuppressive properties of human PCs are not intrinsic but instead result from IFN-γ-induced IDO1-mediated tryptophan depletion.

Differential functional roles of fibroblasts and pericytes in the formation of tissue-engineered microvascular networks in vitro

“…Finally, we have chosen placental PCs because they are also readily obtained from discarded post-partum tissue and have been characterized both behaviorally and by transcriptomics. 19 We analyzed the interactions of these cells within gels cast in microfluidic chambers because this in vitro setting gives rise to perfusable μVNs, better recapitulating the in vivo setting. The RNA-seq analyses demonstrate that FBs and PCs possess distinct gene expression patterns that likely contribute to the differences in their effects on μVNs within these devices.…”

“…Given the differences that we observed within the microfluidic devices when ECFCs were co-cultured with FBs or PCs, we used RNA-seq to compare bulk gene expression profiles of the FBs and PCs. We prepared and analyzed FB gene expression profiles with RNA-seq using three different donors in the same manner as previously published for PCs 19 (Supplementary Data 1). We identified 1056 genes that were differentially expressed (q-value < 0.05 and log 2 [fold change]≥5; Fig.…”

Differential functional roles of fibroblasts and pericytes in the formation of tissue-engineered microvascular networks in vitro

“…Finally, we have chosen placental PCs because they are also readily obtained from discarded post-partum tissue and have been characterized both behaviorally and by transcriptomics. 21 We analyzed the interactions of these cells within gels cast in microfluidics chambers because this in vitro setting gives rise to perfusable μVNs, better recapitulating the in vivo setting.…”

“…Prep and sequencing was performed as described previously for PC RNA-seq analysis. 21 For purified total RNA collected from FB samples, the three strand-specific sequencing libraries were produced following the Illumina TruSeq stranded protocol. According to Illumina protocol, the libraries underwent 76-bp pairedend sequencing using an Illumina HiSeq 2500, generating an average of 32 million pairedend reads per library.…”

“…Given the differences that we observed within the microfluidic devices when ECFCs were co-cultured with FBs or PCs, we used RNA-seq to compare bulk gene expression profiles of the FBs and PCs. We prepared and analyzed FB gene expression profiles with RNA-seq using three different donors in the same manner as previously published for PCs 21 (Supplemental Table 1). We identified 1056 genes that were differentially expressed (q-value <0.05 and log2[fold change] 5; Fig.…”