Increased expression of CCL18, CCL19, and CCL17 by dendritic cells from patients with rheumatoid arthritis, and regulation by Fc gamma receptors

Abstract

Background: Dendritic cells (DC) have a role in the regulation of immunity and tolerance, attracting inflammatory cells by the production of various chemokines (CK). Fc gamma receptors (Fc gamma R) may be involved in regulation of the DC function.

Objective: To assess the expression of CK by immature (iDC) and mature DC (mDC) and its regulation by Fc gamma R in patients with RA and healthy donors (HC).

Methods: Expression of CK by DC from patients with RA and from HC was determined by real time quantitative PCR and ELISA. DC were derived from monocytes following standardised protocols. To study the potential regulation by Fc gamma R, iDC were stimulated with immune complexes (IC) during lipopolysaccharide (LPS) induced maturation. The presence of CK was studied in synovial tissue from patients with RA, osteoarthritis, and healthy subjects by RT-PCR and immunohistochemistry.

Results: iDC from patients with RA had markedly increased mRNA levels of the CK CCL18 and CXCL8. Upon maturation with LPS, expression of CCL18, CCL19, CXCL8, CCL3, and CCL17 increased dramatically, reaching significantly higher levels in patients with RA. Monocytes failed to express these CK, except for CXCL8 and CCL3. IC-mediated triggering of the Fc gamma R on DC from patients with highly active RA down regulated all CK, whereas the reverse was seen when DC from patients with low disease activity and healthy donors were stimulated. CCL18 was significantly increased in RA synovial tissue.

Conclusion: Increased CK expression by DC was found in patients with RA. This expression is partly regulated by Fc gamma R triggering and results in an inhibitory DC subtype in RA upon Fc gamma R-mediated triggering.

Figure 1

CK mRNA expression by immature and mature monocyte derived dendritic cells from patients with RA and healthy controls. CK expression of iDC of patients with RA (black bars), healthy subjects (light grey bars), and after full maturation (patients with RA (dark grey bars), and healthy controls (white bars)). The mRNA expression is related to PBGD expression set to level 1. Note the log scale of the y axis. The mean level of mRNA expression is indicated by the number above the bar.

Figure 2

CK expressed by iDC and mDC in relation to RA disease activity. (A) CK mRNA expression by iDC from 10 patients with active RA (black bars), six with inactive disease (grey bars), and 12 healthy controls (white bars). (B) Production of CK by mDC from the same group of patients with active (black bars) and inactive (grey bars) disease, and healthy donors (white bars). *p<0.05. The mRNA expression is related to PBGD expression set to level 1. Note the log scale of the y axis. The mean level of mRNA expression is indicated by the number above the bar.

Figure 3

CK expression by monocytes from patients with RA and healthy controls. Relative expression of CK CXCL8 and CCL3 by monocytes of five patients with active RA (black bars) and five healthy donors (white bars), respectively. *p<0.05. The mRNA expression is related to PBGD expression set to level 1. Note the log scale of the y axis. The mean level of mRNA expression is indicated by the number above the bar.

Figure 4

CK mRNA expression after triggering of FcγR on mature DC. Level of CCL18, CCL22, CCL17, CCL19, CXCL8, and CCL3 mRNA expression in the absence (black bars) or presence (white bars) of IC by mature DC from eight patients with RA and six healthy controls, respectively. _*p<0.05. The mRNA expression is related to PBGD expression set to level 1. Note the log scale of the y axis. The mean level of mRNA expression is indicated by the number above the bar.

Figure 5

Expression of CK in synovial tissue from patients with RA compared with patients with OA and healthy controls. mRNA expression level of the CK CCL18, CCL19, CCL17, and CXCL8 in synovial tissue from five patients with RA (black bars), five patients with OA (grey bars), and five healthy donors (white bars). _*p<0.05. The mRNA expression is related to PBGD expression set to level 1. Note the log scale of the y axis. The mean level of mRNA expression is indicated by the number above the bar.

Figure 6

CCL18 (DC-CK1) production by iDC and mDC of patients with RA and healthy subjects before and after stimulation with anti-IgG complexes (HAGGs). (A) CCL18 protein production in supernatants of iDC and mDC from nine patients with active RA (black bars), five patients with inactive RA (grey bars), and six healthy donors (white bars). (B) CCL18 secretion by iDC after stimulation with LPS alone or LPS+HAGGs in nine patients with active RA (left panel) and six healthy donors (right panel).

Figure 7

Expression of CCL18 (DC-CK1) in the synovial tissue of patients with RA and OA and in healthy donors. The top row represents immunostaining for CCL18 of synovial tissue from a patient with RA, a patient with OA, and a healthy donor, respectively. The bottom row corresponds with immunostaining for DC-LAMP from the same subjects.