Significant correlations of upregulated MPO expression with cytokine imbalance in ankylosing spondylitis patients and the inhibitory effect mediated by mesenchymal stem cells

Abstract

Background: Little is known regarding both the role of myeloperoxidase (MPO) and the impact of mesenchymal stem cells (MSCs) on inflammatory and immune responses in ankylosing spondylitis (AS). This study is aimed to explore the role of MPO and the regulatory effect of umbilical cord-derived MSCs on MPO expression in monocytes in AS.

Methods: MPO mRNA expression in the peripheral blood mononuclear cells (PBMCs) was detected by Real-time PCR. Cytokines including IL-2, IFN-γ, IL-17 A, IL-4, IL-10, IL-6 and TNF-α were determined by flow cytometry. A co-culture system was established by culturing THP-1 cells with MSCs at a ratio of 5:1.

Results: Increased mRNA expression of MPO was observed in PBMCs of AS patients compared to healthy controls (P < 0.05). The mRNA expression of MPO was positively associated with C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) (P < 0.05) in AS. Besides, the levels of IL-2, IL-10, IFN-γ, IL-17 A, IL-4, IL-6, TNF-α in plasma were notably increased in AS (P < 0.05). Positive correlations between MPO expression and IL-2, IFN -γ, IL-4, TNF-α as well as IL-6 were demonstrated in AS (P < 0.05). Furthermore, MSCs remarkably suppressed the mRNA expression of MPO along with the secretion of IL-17 A and TNF-α, but promoted IL-10 generation in monocytes.

Conclusion: MPO expression is significantly upregulated and correlates with cytokine imbalance in AS. It may serve as a valuable immunotherapeutic target for AS. MSCs can significantly inhibit monocyte-mediated inflammatory response potentially by downregulating MPO in monocytes.

Keywords: Ankylosing spondylitis; Cytokines; Mesenchymal stem cells; Myeloperoxidase.

Fig. 1

MPO mRNA expression in PBMCs from experimental group and controls. (AS, ankylosing spondylitis; NC, normal controls; Mann-Whitney U test, ****, P < 0.0001)

Fig. 2

Correlation of MPO mRNA expression level with AS clinical indexes. (A) The MPO mRNA level in AS was positively associated with the consistence of CRP (n = 18, P = 0.0021); (B) The MPO mRNA level in AS was positively linked with consistence of ESR (n = 25, P = 0. 0373); (C) The MPO mRNA level in AS had no significant correlation with age (n = 26, P = 0. 9508); (D) The mRNA level of MPO in AS was not statistically associated with peripheral blood platelet count (n = 25, P = 0. 0665)

Fig. 3

Plasma cytokines difference between AS patients and controls. (A) The concentration of plasma IL-2; (B) The level of plasma IL-4; (C) The level of plasma IL-6; (D) The level of plasma IL-10; (E) The level of plasma TNF-α; (F) The level of plasma IFN-γ; (G) The level of plasma IL-17 A. (All comparisions were analyzed by Mann-Whitney U test, ****, P < 0.0001)

Fig. 4

Correlation analysis between MPO mRNA expression levels and plasma cytokine levels in AS patients. (A) MPO mRNA expression level in PBMCs was positively correlated with plasma IL-2 of AS patients, n = 24, P = 0.0049; (B) MPO mRNA expression level in PBMCs was positively correlated with plasma IL-4 of AS patients, n = 24, P = 0.0321; (C) MPO mRNA expression level in PBMCs was positively correlated with plasma IL-6 of AS patients, n = 25, P < 0.0001;(D) MPO mRNA expression level in PBMCs had no significant correlation with plasma IL-10 of AS patients, n = 25, P = 0.1894; (E) MPO mRNA expression level in PBMCs was positively correlated with plasma TNF-α of AS patients, n = 22, P = 0.0099; (F) MPO mRNA expression level in PBMCs was positively correlated with plasma IFN -γ of AS patients, n = 24, P = 0.0164; (G) MPO mRNA expression level in PBMCs had no significant correlation with plasma IL-17 A of AS patients, n = 25, P = 0.2357

Fig. 5

MSCs inhibit the MPO mRNA expression in THP-1 cells stimulated by LPS. (*, P < 0.05; ***, P < 0.001; n = 4)

Fig. 6

Altering effects of umbilical cord-derived MSCs on cytokines production of THP-1 cells stimulated by LPS. (A) The concentration of IL-2 in the supernatant of the co-cultured system: t-test, P > 0.05, n = 3; (B) The level of IL-4 in the supernatant of the co-cultured system; (C) The level of IL-6 in the supernatant of the co-cultured system; (D) The level of IL-10 in the supernatant of the co-cultured system; (E) The level of TNF-α in the supernatant of the co-cultured system; (F) The level of IFN-γ in the supernatant of the co-cultured system; (G) The level of IL-17 A in the supernatant of the co-cultured system (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; n = 3)