Highly multiplexed proteomic analysis reveals significant tissue and exosomal coagulation pathway derangement in chronic rhinosinusitis with nasal polyps

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Background Oral steroids, traditionally thought of as immunosuppressive agents that are broad in their immunomodulatory effects, are a mainstay of treatment to reduce disease burden in chronic rhinosinusitis with nasal polyps (CRSwNP). The purpose of this study was to determine how differentially expressed proteins in CRSwNP are affected by oral steroid therapy. Methods Matched exosomal proteomic arrays were quantified using aptamer‐based methods in systemic steroid‐naive CRSwNP patients before and after a standardized oral prednisone course (n = 12). Previously identified differentially expressed proteins in CRSwNP patients were compared to determine the effect of steroids on expression. Fisher's exact test and t test were applied to normalized protein expression profiles to determine significance. Results Of 18 proteins previously identified to be highly underexpressed in CRSwNP, 16 (89%) had an average increase after systemic steroid treatment (p < 0.05). Lactoperoxidase, initially present at 9‐fold lower concentrations in CRSwNP subjects, increased by 209% after steroid treatment. A similar trend was observed with other proteins of interest, including platelet factor 4 and C‐C motif ligand 28. The converse of this steroid effect was not true; of the 53 proteins that are highly overexpressed in CRSwNP, only 22 (42%) decreased in quantity with steroid use. Conclusion Proteomic analysis of differentially expressed proteins in CRSwNP demonstrates that systemic steroids cause almost uniform upregulation of transcriptionally decreased proteins, whereas the effects of steroids on transcriptionally increased proteins are more heterogeneous. Thus, proteomic analysis may be an effective tool to understand specific therapeutic benefits of steroid use in polyp disease and to create more targeted treatments.

Section: Discussionsupporting

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Unexpected effects of systemic steroids on the CRSwNP proteome: is protein upregulation more important than inhibition?

Workman

Miyake

Nocera

et al. 2020

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“…However, a greater number of target markers were poorly represented by swab sampling than by cytobrush (including TNF, IFNG, OSM, IL1A, and IL17C). Additionally, recent studies by Mueller et al 51,52 have also identified proteomic assessment of mucus exosome filtrates as a further promising noninvasive ("liquid biopsy") sampling option. Overall, the findings of this study indicate that these less invasive options acceptably reflect tissue patterns for several putative biomarkers of CRS and could thus be used in clinic and research settings as a proxy for tissue processes.…”

Section: Comparisons Of Sampling Methodologiesmentioning

confidence: 99%

Assessing tissue transcription biomarkers of chronic rhinosinusitis: a comparison of sampling methodologies

Hoggard

Douglas

Taylor

et al. 2020

Background Chronic rhinosinusitis (CRS) is a spectrum of complex inflammatory conditions of the sinonasal mucosa. Identification of biomarkers that enable classification and improved delineation among CRS endotypes is of increasing interest. However, the extent to which less invasive sampling methods identify genuine tissue inflammatory patterns is not well understood. The aim of this study was to investigate mucosal swab and cytobrush sampling as less invasive proxies for tissue transcription levels of putative biomarkers of CRS. Methods Expression levels of 21 biomarkers of interest were assessed via custom TaqMan array cards from mucosal biopsy, cytobrush, and swab samples, in 32 patients with CRS. Reported expression levels were compared between each of the 3 sample types within each patient. Results Reported transcription levels from swab samples for IL33, MUC5AC, IL1RN, CXCL8 (IL‐8), TNF, IFNG, IL5, OSM, IL1A, and IL17C, and cytobrush levels for IL33, MUC5AC, IL5RA, IL1RN, CXCL8 (IL‐8), and IL5 were significantly different to tissue levels from matched biopsy samples. Conclusion Reported expression via swab and cytobrush sampling differed from patterns observed in matched tissue for 10 of 21 and 6 of 21 markers, respectively. Non–biopsy‐based studies for these particular markers may therefore not adequately represent tissue inflammatory processes and should be interpreted with caution. Cytobrush samples largely tracked tissue patterns for the remaining target biomarkers. In these cases, cytobrush sampling appears to adequately reflect tissue patterns for several putative biomarkers of CRS, supporting their use in clinical and research settings as a less‐invasive proxy for the assessment of mucosal tissue inflammatory transcription patterns.

“…Pilot studies by our group identified the development of a noninvasive, exosomal biosignature as a promising novel method for studying the etiopathology of CRSwNP while gaining more information about the diagnosis, prognosis, and treatment modalities of CRSwNP . Through this we identified that pappalysin‐A (gene name PAPP‐A) was significantly overexpressed in CRSwNP compared to controls.…”

mentioning

confidence: 99%

Significant polyomic and functional upregulation of the PAPP‐A/IGFBP‐4/5/IGF‐1 axis in chronic rhinosinusitis with nasal polyps

Müller

Nocera

Workman

et al. 2020

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Background Chronic rhinosinusitis with nasal polyps (CRSwNP) is associated with epithelial expansion and polyp survival. However, the molecular mechanism of this aberrant proliferation is unclear. The purpose of this study was to interrogate derangements of the pappalysin‐A/insulin‐like growth factor binding protein/insulin‐like growth factor‐1 (PAPP‐A/IGFBP‐4/5/IGF‐1 axis) as a major contributing factor to polyp growth in CRSwNP. Methods Matched tissue and exosomal proteomic arrays including PAPP‐A, IGFBP‐4, IGFBP‐5, and IGF‐1 were quantified using aptamer‐based methods/Western blots for proteomic analysis and whole‐transcriptome sequencing/quantitative polymerase chain reaction (qPCR) for transcriptomic analysis in CRSwNP and control patients. Functional PAPP‐A assays were then performed in both tissue and exosomes (set 1: n = 20 per group; validation set 2: n = 26 per group). Results Tissue and exosomal PAPP‐A was significantly overexpressed in CRSwNP compared to controls on both a transcriptomic and proteomic level (p < 0.0001). Known inhibitors of PAPP‐A (stanniocalcin‐1/‐2) were significantly downregulated (p < 0.0001) as were PAPP‐A cleavage products (IGFBP‐5 p < 0.0001). PAPP‐A function was shown to be increased 5‐fold to 6‐fold in tissue and exosomes. Conclusion Upregulated tissue and exosomal PAPP‐A signaling is significantly associated with CRSwNP and may be an important factor in the promotion of epithelial proliferation and polyp growth. These data lend further support to the emerging concept of exosomal functional and polyomic analyses as a method to study sinonasal pathology.