Elevated Serum Interleukin-34 Level in Patients with Systemic Lupus Erythematosus Is Associated with Disease Activity
Elevated Serum Interleukin-34 Level in Patients with Systemic Lupus Erythematosus Is Associated with Disease Activity
Huan Huan Xie
Mei Ying Cui
Li Ping Xia
OPEN Published: xx xx xxxx We measured the interleukin-34 (IL-34) level in sera from patients with systemic lupus erythematosus (SLE) and discoid lupus erythematosus (DLE) using an enzyme-linked immunosorbent assay (ELISA). Blood tests, including assays to determine C-reactive protein (CRP), complement (C) 3, C4, immunoglobulin (Ig) A, IgG, IgM, anti-double-stranded DNA antibody (Anti-dsDNA Ab) and hemoglobin (Hb) levels and white blood cell (WBC) and platelet (PLT) counts, were performed using standard methods. Lupus nephritis (LN) was diagnosed according to the American College of Rheumatology (ACR) renal criteria. The SLE disease activity was scored using the SLE Disease Activity Index (SLEDAI). Among the 110 SLE cases, IL-34 could be detected in 79 cases (71.8%). IL-34 was barely detected in the control group. The serum level of IL-34 was significantly higher in the SLE group. No change was observed in the serum IL-34 concentration in the SLE patients regardless of LN status. Correlations were observed between the serum IL-34 level and the disease activity parameters. The SLE patients with detectable IL-34 levels had higher SLEDAI and IgG concentrations and lower C3 and Hb levels than patients with undetectable IL-34 levels. Therefore, IL-34 could be a potential disease activity marker for SLE.
Systemic lupus erythematosus (SLE) is a chronic systemic autoimmune disease characterized by a dysregulated
autoantibody production and complement activation, resulting in multiple system and organ damage,
particularly in the kidneys, blood system and central nervous system1. The treatment of SLE patients with
glucocorticoids and immunosuppressive drugs has a significant impact on the outcome of this disease.
Recently, many cytokines have been shown to play important roles in the pathogenesis of SLE, and several
biological products have been used in SLE patients. In 2011, the US Food and Drug Administration (FDA) approved
belimumab, which is a fully human monoclonal antibody against B lymphocyte stimulator (BLyS), as a new
biological drug for the treatment of SLE2. Several other potential biological targets are currently under investigation3.
However, using the current therapeutic strategies, only a very small proportion of SLE patients achieve long-term
remission4. Thus, more effective and safer drugs are needed to reduce the disease activity and achieve long-term
Interleukin (IL)?34 is a newly discovered cytokine that has no significant amino acid sequence homology to
other cytokines5. Currently, knowledge regarding this cytokine is limited. IL-34 shares a common receptor with
macrophage-colony stimulating factor (M-CSF)6. Because IL-34 is an alternative ligand of the colony-stimulating
factor-1 receptor (CSF-1R), IL-34 binds to CSF-1R and promotes the differentiation and proliferation of
lymphocytes and the expression of cytokines, leading to inflammatory lesions and autoimmunity5,7. The serum level of
IL-34 is elevated in rheumatoid arthritis (RA) patients8,9.
IL-34 can induce the expression of IL-6, interferon ?-inducible protein (IP) 10 and monocyte chemoattractant
protein-1 (MCP-1) in human whole blood7. IL-610,11, IP1012 and MCP-113 participate in the pathogenesis of SLE.
The expression of IL-34 was increased in a study investigating kidney tissues from a murine model of lupus
nephritis (LN)14. However, whether IL-34 is released into the circulation in SLE patients and the relationship
between IL-34 and the clinical parameters remain unclear. We hypothesized that IL-34 might play a role in SLE.
Disease duration (yrs) 5.73 ? 5.37
Clinical features of patients with SLE. In total, 110 SLE patients were recruited. Their clinical
characteristics are summarized in Table?1. LN was found in 63 of the 110 patients.
Patients with SLE had elevated IL-34 levels. While IL-34 was nearly undetectable in the serum
from the DLE patients (9.6%) and healthy controls (5.5%), IL-34 was detected in 79 of the 110 SLE patients
(71.8%). The serum IL-34 level was significantly higher in the patients with SLE than that in the healthy controls
(p < 0.001) (Fig.?1A).
The association between IL-34 and LN was explored. LN was found in 63 of the 110 patients. No difference
was observed in the IL-34 level regardless of the renal disease status (Fig.?1B).
Serum IL-34 level and disease activity in SLE patients. A significant positive correlation was observed
between the IL-34 levels and the disease activity marker SLEDAI (r = 0.319, p = 0.004) (Fig.?2A) but not with
CRP. In contrast, a statistically significant negative correlation was observed between IL-34 and C3 in the SLE
patients (r = 0.324, p = 0.004) (Fig.?2A). Thus, the IL-34 level could be correlated with the SLE disease activity.
Furthermore, the serum IL-34 level was correlated with IgG and anti-dsDNA antibody production in SLE patients
(r = 0.259, p = 0.021; r = 0.352. p = 0.001, respectively) (Fig.?2B). Thus, IL-34 might be associated with antibody
production in SLE pathogenesis.
Because hematological changes are quite common in SLE patients, the IL-34 level is significantly negatively
associated with Hb and PLT (r = 0.334, p = 0.003; r = 0.236, p = 0.038, respectively) (Fig.?2C). No correlation was
observed between the IL-34 level and the WBC count.
According to a multivariate linear regression analysis, SLEDAI and hemoglobin were independently
correlated with the circulating levels of IL-34 in the SLE patients (? = 0.319, p = 0.008 and ? = ?0.275, p = 0.023)
Differences in SLE clinical indices between the IL-34-positive and -negative groups. Of the 110
SLE patents, IL-34 could be detected in 79 patients. These patients were allocated into 2 groups, i.e., IL-34-positive
group (IL-34+ group, IL-34 detectable SLE patients) and the IL-34-negative group (IL-34? group, IL-34
undetectable SLE patients), based on their IL-34 status.
The levels of SLEDAI and IgG in the IL-34+group were much higher than those in the IL-34?group (SLEDAI:
8.75 ? 0.66 vs 6.00 ? 0.68, p = 0.005; IgG: 16.97 ? 0.72 vs 11.79 ? 0.92 g/L, p < 0.0001) (Fig.?3A), while the
levels of C3 and Hb in the IL-34+ group were much lower than those in the IL-34? group (C3: 0.58 ? 0.04 vs
0.72 ? 0.04 g/L, p = 0.019; Hb: 105.3 ? 2.37 vs 116.3 ? 3.04 g/L, p = 0.006) (Fig.?3B).
No difference was found in PLT between the two groups.
This study is the first to show that the IL-34 level is elevated in SLE patients and correlated with the disease
activity. In this study, we reported that the serum IL-34 level was significantly elevated in the SLE patients compared
to that in the healthy controls. Among the SLE patients with or without LN involvement, no statistically
significant difference was observed in the IL-34 level. However, according to Bethunaickan R, the expression of IL-34
is increased in mice with LN14, and IL-34 might play a role in the pathogenesis of LN. The current data are not
sufficient to reach such a conclusion, and more evidence is required.
Moreover, in our study, the IL-34 level is significantly positively correlated with the disease activity as assessed
by the SLEDAI. The serum C3 level decreases in SLE patients, particularly in patients with an active disease status.
In this study, the serum IL-34 level was correlated with the decreased C3 level. Furthermore, the levels of SLEDAI
in the IL-34+group were much higher than those in the IL-34-group. In contrast, the C3 level in the IL-34+group
was much lower. Altogether, the serum IL-34 level might be associated with the disease activity in SLE.
Furthermore, we showed correlations among IL-34, IgG and anti-dsDNA Ab. Thus, IL-34 might be associated
with antibody production in SLE pathogenesis.
Because SLE is a highly heterogeneous disease, characterizing subgroups of SLE patients with specific disease
phenotypes could help clarify the pathogenesis of SLE. Cytokine measurements might lead to the
characterization of these pathways and the identification of potential therapeutic targets. Hematological involvement is
quite common in patients with SLE. In this study, the IL-34 level was negatively correlated with hemoglobin and
exhibited a weak negative association with platelets. However, in the subsequent analysis of the groups with the
IL-34+ & IL-34? SLE patients, a difference in hemoglobin, but not in platelets, was observed. Thus, IL-34 may be
associated with anemia in SLE patients. However, more studies are needed to clarify whether IL-34 plays a role in
the hematological changes observed in SLE patients.
Elevated concentrations of inflammatory mediators are characteristic of autoimmune diseases with chronic or
recurrent inflammation. IL-34 plays pivotal roles in the proliferation and differentiation of mononuclear phagocyte
lineage cells and osteoclastogenesis and is necessary for the maintenance of Langerhans cells after the resolution of
inflammation14,15. Recently, IL-34 has been shown to play important roles in the pathogenesis of RA8,9. IL-34 is
overexpressed in the inflamed salivary glands of patients with Sjogren?s syndrome (SS)16. Furthermore, IL-34 may play
a role in cancer. IL-34 promotes tumor progression and metastatic processes in osteosarcoma via the induction of
angiogenesis17. However, our study has limitation for not including a disease control group, such as another non-SLE
autoimmune disease. Therefore, elevated levels of IL-34 may be non-specific features of systemic inflammation and
not a specific pathogenic factor of lupus. Further studies on IL-34 and other inflammation diseases are needed.
Both SLE and RA are systemic autoimmune diseases. Although functional studies are clearly required to
confirm the role of IL-34 in the pathogenesis of SLE, the observed correlations between the elevated levels of
IL-34 and SLEDAI, antibody production and anemia suggest that IL-34 plays a role in the modulation of immune
inflammatory pathways in SLE.
Many cytokines play important roles in SLE18. TNF-? and IL-6 are major inflammatory mediators of
pathology in both SLE and RA18,19. TNF-? also plays a role in anemia. Furthermore, IL-610,11, IP10 12 and MCP-113
participate in the pathogenesis of SLE. IL-34 can induce the expression of IL-6, IP10 and MCP-1 in human whole
blood7. The expression of IL-34 increases with inflammation in patients with inflammatory bowel disease and
experimental colitis and is associated with an elevated TNF? and IL-6 expression20. Altogether, we hypothesize
that IL-34 might play a role in the pathogenesis of SLE by promoting the production of other pro-inflammatory
cytokines, including TNF-?, IL-6, IP10 and MCP-1.
In summary, the serum IL-34 level was significantly elevated in the SLE patients and correlated with the
disease activity and homological changes. IL-34 could be a potential disease activity marker, and this study might
have revealed new insight for the study of SLE disease activity.
Patients. This study was approved by the ethics committee of the 1st Affiliated Hospital of China Medical
University. The experiments were conducted in accordance with the regional Ethics Committee guidelines and
regulations. Informed consent forms were obtained from all patients.
Serum was obtained from 110 patients with SLE and 31 patients with discoid lupus erythematosus (DLE),
which is the most common chronic cutaneous lupus subtype. The diagnosis of SLE was based on the American
College of Rheumatology criteria for SLE21 In addition, 55 healthy age- and gender-matched controls were
recruited. All serum samples were stored at ?70 ?C until analysis. Since drugs can affect the circulating IL-34
levels, the serum was collected only from SLE patients who were not under treatment. Among the 110 SLE
patients, eighty-nine had newly diagnosed SLE, and the other twenty-one were previously diagnosed but had not
taken corticosteroids or immunosuppressive drugs for at least three months before enrollment in the study. All
thirty-one DLE patients were newly diagnosed, and no drugs were used. Blood tests, including assays to
determine C-reactive protein (CRP), complement (C) 3, C4, immunoglobulin (Ig) A, IgG, IgM, anti-double-stranded
DNA antibody (Anti-dsDNA Ab) and hemoglobin (Hb) levels and white blood cell (WBC) and platelet (PLT)
counts, were performed using standard methods. LN was identified if there was evidence of involvement
according to the ACR renal criteria. The disease activity was scored using the SLE Disease Activity Index (SLEDAI)22.
Measurement of IL-34 level in serum samples. The serum IL-34 level was measured using an enzyme-linked
immunosorbent assay (ELISA) according to the manufacturer?s directions (R&D Systems, Minneapolis, MN). Each
sample was measured in triplicate. Since serum antibodies can interfere with the assay for IL-34 and cause false positive
results, a known quantity of recombinant IL-34 was added to the serum that contained the antibodies to determine
whether the measurement was altered. No interference effect was detected (Supplementary Figure).
Statistical analysis. All data are presented as the mean ? SD (parametric) or median (range)
(non-parametric). The t-test and the non-parametric Mann-Whitney test were performed to compare the
variables between groups. Pearson?s or Spearman?s correlation coefficient was used to test the correlations. A
multivariable linear regression analysis was also performed to assess the independent predictive value of IL-34 in
disease activity and hematological changes in the SLE patients. All analyses were performed using SPSS 17.0 and
GraphPad 6 software.
This study was supported by funding from the National Natural Science Foundation of China (No. 81373219),
Scientific Research of the First Hospital of China Medical University (fsfh1007) and the Higher Education
Department of Liaoning Province Research Grant
S.H., X.L.P. and L.J. designed the study and revised the manuscript; X.H.H., Z.L., and C.M.Y. performed the
experiments and drafted the manuscript; and X.H.H. and S.H. analyzed the data.
Supplementary information accompanies this paper at https://doi.org/10.1038/s41598-018-21859-z.
Competing Interests: The authors declare no competing interests.
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