IL-13 and IL-13-induced periostin levels are specifically decreased in patients following endoscopic sinus surgery for chronic rhinosinusitis

Abstract

Background: Type 2 (T2) inflammation plays a pathogenic role in chronic rhinosinusitis (CRS). The effects of endoscopic sinus surgery (ESS) on T2 inflammation are unknown.

Objective: The aim of this study was to compare T2 inflammatory biomarkers from middle meatal (MM) mucus for distinguishing patients with CRS from CRS-free patients, identifying major phenotypes (CRS without nasal polyps [CRSsNP] and CRS with nasal polyps [CRSwNP]), assessing endotypic change, and establishing cross-sectional and longitudinal outcomes in patients undergoing ESS.

Methods: MM mucus samples were collected from patients with CRSsNP and patients with CRSwNP before and 6 to 12 months after ESS and compared with samples from CRS-free control patients. T2 biomarkers were evaluated both continuously and using threshold-based definitions of T2 endotype to identify relationships with patient-reported (based on the 22-Item Sinonasal Outcomes Test and Chronic Rhinosinusitis Patient-Reported Outcomes Measure) and clinician-reported (radiographic and endoscopic) severity. Linear mixed models were developed to analyze clinical variables associated with T2 biomarker levels.

Results: A total of 154 patients with CRS (89 with CRSsNP and 65 with CRSwNP) were enrolled, with a mean interval of 9 months between ESS and follow-up. An analysis of pre-ESS MM mucus samples revealed elevated levels of T2 mediators in patients with CRSwNP versus in patients with CRSsNP and CRS-free controls. Temporally stable correlations between levels of IL-13 and IL-5, levels of periostin and complement 5a, and levels of eosinophil cationic protein (ECP) and eotaxin-3 were observed. On this basis and on the basis of pathologic significance, levels of IL-13, periostin and ECP were further analyzed. After ESS, levels of IL-13 and periostin decreased significantly, whereas ECP levels remained unchanged. Across pre- and post-ESS evaluation, the T2 endotype was associated with radiographic severity but did not predict outcomes. CRSwNP status and African American race were associated with higher levels of IL-13 and periostin, whereas ECP level was higher in patients undergoing extensive surgery.

Conclusion: ESS decreased levels of IL-13 and periostin in the middle meatus. T2 inflammation after ESS was correlated with patient- and clinician-reported severity across phenotypes. Pre-ESS T2 inflammation did not predict post-ESS outcomes.

Keywords: CRSsNP; CRSwNP; Chronic rhinosinusitis; biomarker; endotype; nasal polyps; patient-reported outcome measures.

Figure 1:. Study flowchart illustrating the timeline of biomarkers measured and clinical variables collected.

A) Clinical variables and B) biomarker variables were cross-sectionally and longitudinally analyzed to evaluate the relationship between T2 MM inflammation and disease severity. ESS: Endoscopic Sinus Surgery; PROMs: Patient-Reported Outcome Measures; SNOT-22: 22-item Sinonasal Outcomes Test; LM: Lund-Mackay radiographic score; CRS-PRO: 12-item Chronic Rhinosinusitis Patient-Reported measure; BSIT: Brief Smell Identification Test; CCL26: Eotaxin-3; C5a: Complement 5a.

Figure 2:. Comparison of pre-ESS Type-2 inflammation biomarkers in middle meatal secretions of non-CRS control, CRSsNP and CRSwNP patients.

Pre-ESS middle meatal mucus concentrations of the T2 cytokines A) IL-4, B) IL-5, C) IL-13 D) Periostin, E) Complement 5a, F) IgE and G) CCL26 are higher in CRSwNP than in control and CRSsNP secretions. H) Eosinophilic cationic protein (ECP) levels were significantly different between control patients, CRSsNP and CRSwNP patient middle meatal mucus. Results are shown as median with interquartile range and expressed in pg/ml for IL-4, IL-5, IL-13, Periostin, and Complement 5a, and ng/ml for ECP and IgE. The expression levels above the 90^th percentile of the respective biomarker in non-CRS controls was used to designate “high” levels demarcated by dotted line. *P < .05, **P<.01, ***P < .001, ****P < .0001, by Kruskal-Wallis with Dunn’s multiple comparison. For the entire cohort, n=144 for IL-4, IL-5, IL-13, ECP; n=143 for periostin, Complement 5a, IgE For CRSsNP, n=85 for all mediators. For CRSwNP, n=59 for IL-4, IL-5, IL-13, ECP; n=58 for periostin, complement 5a, IgE

Figure 3:. Clustered correlation heatmaps representing correlation between the 8 studied T2 biomarkers pre-, post- and the change (delta).

T2 biomarkers were correlated and clustered for each of the two observed time points as the change (delta) observed between the two timepoints. In particular, IL-5 and IL-13; ECP and CCl26; and periostin and Complement 5a were stably co-clustered cross-sectionally and longitudinally. *P < .05, **P<.01, ***P < .001, ****P < .0001, For the entire cohort, n=144 for IL-4, IL-5, IL-13, ECP; n=143 for periostin, Complement 5a, IgE For CRSsNP, n=85 for all mediators. For CRSwNP, n=59 for IL-4, IL-5, IL-13, ECP; n=58 for periostin, Complement 5a, IgE

Figure 4:. IL-13, Periostin but not ECP decreased significantly following ESS.

IL-13 and Periostin levels but not ECP were significantly decreased in middle meatal mucus for the A: Entire CRS cohort and C: CRSwNP patients. In B: CRSsNP patients, only IL-13 decreased significantly. Lines denote median biomarker levels. *P < .05, **P<.01, ***P < .001, by Wilcoxon matched pairs analysis. Benjamini-Hochberg adjusted p-values are reported.

Figure 5:. Sankey plots illustrating changes in endotype when characterized by IL-13, periostin, and ECP in CRSsNP and CRSwNP patients.

Visualization of the changes in T2 endotype between pre-ESS and post-ESS observations in CRSsNP and CRSwNP patients when endotype is defined by A) IL-13, B) periostin and C) ECP. Averages of Lund-Mackay (LM) score, modified Lund-Kennedy score, Chronic Rhinosinusitis Patient-Reported Outcomes (CRS-PRO), 22-item Sinonasal Outcomes Test (SNOT-22), and the Brief Smell Identification Test (BSIT) are reported in the respectively defined T2 high and low groups. T2 high and low groups were constructed using the 90^th percentile value among the non-CRS controls for IL-13, periostin and ECP. T-tests were conducted to compare high and low groups separately for pre-ESS and post-ESS. *P < .05, **P<.01, ***P < .001, by t-test analysis. Benjamini-Hochberg adjusted p-values are reported.

Figure 6:. Correlations between Il-13, Periostin and ECP with radiographic, endoscopic and patient-reported measures of severity pre-ESS, post-ESS and the delta for the entire cohort of patients.

For the entire cohort, A) Pre-ESS IL-13, periostin and ECP had weak to moderate and highly significant correlation with LM radiographic severity and patient-reported severity measured by the CRS-PRO. B) Post-ESS, all three T2 biomarkers had weak but highly significant correlation with LM radiographic and endoscopic MLK endpoints and patient reported severity assessed by the CRS-PRO. C) In analysis of the changes in radiographic and patient reported severity and the changes in T2 biomarkers, once again found that changes in LM radiographic severity were best correlated with changes in IL-13, periostin and ECP. The intensity of red and blue increase as strength of correlation becomes positive or negative, respectively as represented in the heatmap. Spearman R values are shown in the center of heat-map and significance in the bottom-right corner. *P < .05, **P <.01, ***P < .001, ****P < .0001. Benjamini-Hochberg adjusted p-values are reported. Pre-ESS CRS-PRO and SNOT-22 information was available on n=97 and n=108 patients respectively LM, Lund-Mackay. CRS-PRO, Chronic Rhinosinusitis Patient Reported Outcome measure. SNOT-22, 22-Item Sino-Nasal Outcomes Test.

Figure 7:. Linear Mixed Models Forest Plot

Results of adjusted linear mixed models depicting the effect of each considered variable on levels of IL-13, periostin and ECP. The results for IL-13 and periostin are expressed in pg/ml whereas those for ECP are ng/ml. *P<.05, **P<.01, ***P<.001, ****P<0.0001.