Dysregulation of IL-33/ST2 signaling and myocardial periarteriolar fibrosis

Abstract

Microvascular dysfunction in the heart and its association with periarteriolar fibrosis may contribute to the diastolic dysfunction seen in heart failure with preserved ejection fraction. Interleukin-33 (IL-33) prevents global myocardial fibrosis in a pressure overloaded left ventricle by acting via its receptor, ST2 (encoded by the gene, Il1rl1); however, whether this cytokine can also modulate periarteriolar fibrosis remains unclear. We utilized two approaches to explore the role of IL-33/ST2 in periarteriolar fibrosis. First, we studied young and old wild type mice to test the hypothesis that IL-33 and ST2 expression change with age. Second, we produced pressure overload in mice deficient in IL-33 or ST2 by transverse aortic constriction (TAC). With age, IL-33 expression increased and ST2 expression decreased. These alterations accompanied increased periarteriolar fibrosis in aged mice. Mice deficient in ST2 but not IL-33 had a significant increase in periarteriolar fibrosis following TAC compared to wild type mice. Thus, loss of ST2 signaling rather than changes in IL-33 expression may contribute to periarteriolar fibrosis during aging or pressure overload, but manipulating this pathway alone may not prevent or reverse fibrosis.

Keywords: Aging; Interleukin-33; Myocardial fibrosis; Periarteriolar fibrosis; Pressure overload; ST2.

Figure 1.

Periarteriolar fibrosis increases with age in WT mice. (A) Representative histology of periarteriolar fibrosis from young (2 months, NIA) and aged (23 months, NIA) WT mice stained with Masson’s trichrome stain. (B) Periarteriolar fibrosis ratio calculated by individual vessel is significantly increased at 23 months of age compared with young (NIA): n= 58 vessels/group (in 6 mice total (3 males and 3 females), old (NIA): n=64 vessels (in 6 mice total (3 males and 3 females)), ****p<0.0001 by Mann-Whitney test. (C) Global myocardial fibrosis calculated as % blue pixels per total heart pixels is significantly increased at 23 months of age compared with 2 months of age, n=6 mice/group (combined genders), **p<0.01 by Mann-Whitney test.

Figure 2.

IL-33 expression in the heart increases with age. (A) IL-33 mRNA expression in the heart at 2, 12, and 24 months of age in WT mice from the NIA. Values are normalized to housekeeping gene TATA-binding protein (TBP) then to young hearts. N=6 (3 males, 3 females) per age. *p<0.05, **p<0.01 by Kruskal-Wallis test followed by Dunn’s multiple comparisons test. (B) Representative image of western analysis of IL-33 protein expression in the heart at 2, 12, and 24 months of age with vinculin used as a loading control protein. Arrows indicate relevant bands, with noticeable absence of the IL-33 band in the Il33^−/− mouse. Identities of the additional bands are not known. (C) Relative band intensity IL-33 by western analysis at 2 and 24 months of age in WT mice from the NIA. *p<0.05 by Mann-Whitney test.

Figure 3.

ST2 expression decreases with age. (A) mRNA expression of total ST2 in the heart at 2, 12, and 24 months of age in WT mice from the NIA. Values normalized first to TBP then to young mice. N=6/group (3 male, 3 female), *p<0.05 by Kruskal-Wallis test followed by Dunn’s multiple comparisons test. (B) mRNA expression of ST2L in the heart, normalized first to TBP then to young mice. N=6/group (3 male, 3 female), *p<0.05 by Kruskal-Wallis test followed by Dunn’s multiple comparisons test. (C) Serum concentrations of sST2 by ELISA at 2, 16, and 24 months of age in male and female WT mice from the NIA.

Figure 4.

Periarteriolar fibrosis increases in mice deficient in either IL-33 or ST2 following pressure overload due to transverse aortic constriction (TAC). (A) Representative photomicrographs of periarteriolar fibrosis in WT, Il33^−/−, and Il1rl1^−/− mice five weeks after sham versus TAC surgery. (B) Calculated periarteriolar fibrosis ratio by individual vessel, n=86-253 vessels/group from a total of 3-11 mice/group, *p<0.05, **p<0.01, ***p<0.001 by Kruskal-Wallis test.