Unbiased analysis of peripheral blood mononuclear cells reveals CD4 T cell response to RSV matrix protein

Juilee Thakar; Yu Qian; Lauren Benoodt; David Roumanes; Xing Qiu; Nathan Laniewski; ChinYi Chu; Christopher Slaunwhite; Lu Wang; Aishwarya Mandava; Ivan Chang; Ann R Falsey; Mary T Caserta; Thomas J Mariani; Richard H Scheuermann; Edward E Walsh; David J Topham

doi:10.1016/j.jvacx.2020.100065

Unbiased analysis of peripheral blood mononuclear cells reveals CD4 T cell response to RSV matrix protein

Vaccine X. 2020 Apr 21:5:100065. doi: 10.1016/j.jvacx.2020.100065. eCollection 2020 Aug 7.

Authors

Juilee Thakar^{1

2}, Yu Qian³, Lauren Benoodt^{1

4}, David Roumanes^{1

5}, Xing Qiu², Nathan Laniewski^{1

5}, ChinYi Chu⁶, Christopher Slaunwhite⁶, Lu Wang², Aishwarya Mandava³, Ivan Chang³, Ann R Falsey⁷, Mary T Caserta⁸, Thomas J Mariani⁶, Richard H Scheuermann^{3

9}, Edward E Walsh^{7

8}, David J Topham⁵

Affiliations

¹ Department of Microbiology and Immunology, University of Rochester, Rochester, NY, United States.
² Department of Biostatistics and Computational Biology, University of Rochester, Rochester, NY, United States.
³ J. Craig Venter Institute, La Jolla, CA, United States.
⁴ Biophysics and Computational Biology Graduate Program, University of Rochester, Rochester, NY, United States.
⁵ David H. Smith Center for Vaccine Biology and Immunology, University of Rochester, Rochester, NY, United States.
⁶ Division of Neonatology and Pediatric Molecular and Personalized Medicine Program, Department of Pediatrics, University of Rochester Medical Center, Rochester, NY, United States.
⁷ Department of Medicine, Division of Infectious Diseases, University of Rochester Medical Center, Rochester, NY, United States.
⁸ Division of Pediatric Infectious Diseases, Department of Pediatrics, University of Rochester Medical Center, Rochester, NY, United States.
⁹ Department of Pathology, University of California, San Diego, La Jolla, CA, United States.

Abstract

Respiratory syncytial virus (RSV) is the most important cause of respiratory tract illness especially in young infants that develop severe disease requiring hospitalization, and accounting for 74,000-126,000 admissions in the United States (Rezaee et al., 2017; Resch, 2017). Observations of neonatal and infant T cells suggest that they may express different immune markers compared to T-cells from older children. Flow cytometry analysis of cellular responses using "conventional" anti-viral markers (IL2, IFN-γ, TNF, IL10 and IL4) upon RSV-peptide stimulation detected an overall low RSV response in peripheral blood. Therefore we sought an unbiased approach to identify RSV-specific immune markers using RNA-sequencing upon stimulation of infant PBMCs with overlapping peptides representing RSV antigens. To understand the cellular response using transcriptional signatures, transcription factors and cell-type specific signatures were used to investigate breadth of response across peptides. Unexpected from the ICS data, M peptide induced a response equivalent to the F-peptide and was characterized by activation of GATA2, 3, STAT3 and IRF1. This along with upregulation of several unconventional T cell signatures was only observed upon M-peptide stimulation. Moreover, signatures of natural RSV infections were identified from the data available in the public domain to investigate similarities between transcriptional signatures from PBMCs and upon peptide stimulation. This analysis also suggested activation of T cell response upon M-peptide stimulation. Hence, based on transcriptional response, markers were chosen to validate the role of M-peptide in activation of T cells. Indeed, CD4+CXCL9+ cells were identified upon M-peptide stimulation by flow cytometry. Future work using additional markers identified in this study could reveal additional unconventional T cells responding to RSV infections in infants. In conclusion, T cell responses to RSV in infants may not follow the canonical Th1/Th2 patterns of effector responses but include additional functions that may be unique to the neonatal period and correlate with clinical outcomes.

Keywords: PBMC; Peptide stimulation; RSV; Transcriptomics.

Abstract

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