| Abstract |
The cells of the immune system originate from the bone marrow (BM), where many of them also mature. To better understand the aberrant immune response in systemic lupus erythematosus (SLE), we examined the BM in lupus patients using DNA microarrays and compared it to the peripheral blood.
Patients and Methods: Bone marrow mononuclear cells (BMMCs) from 20 SLE patients (12 with active disease, including nephritis and neuropsychiatric disease; 8 with inactive disease) and peripheral blood mononuclear cells (PBMCs) from 27 patients (16 active/ 11 inactive); BMMCs and PBMCs from 7 healthy individuals and 3 osteoarthritis patients served as controls. Samples were analyzed on genome-scale microarrays with 21,329 genes represented. Multiplex cytokine assay was also performed evaluating the levels of 22 cytokines in the sera of lupus patients relative to controls. These cytokines were IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, CXCL8 (IL-8), IL-10, IL-12 (p40), IL-13, IL-15, IL-17, IL-1Ra, IP-10 interferon (IFN)-α and IFN-γ, TNF-α, granulocyte macrophage colony stimulating factor (GM-CSF), CCL2 [monocyte chemoattractant protein (MCP)-1]/(MCAF), CCL3 [macrophage inflammatory protein (MIP)-1α], CCL4 (MIP-1β) and CCL11 (Eotaxin)
Results: We found 102 differentially expressed genes between patient’s and control’s bone marrow samples (unpaired student t-test). These genes represent several important biologic processes; 54 out of 100 differentially expressed genes are involved in major networks including cell death, cell growth and differentiation cell signaling and cellular growth and proliferation. Comparative analysis of the bone marrow with the peripheral blood of SLE patients identified 88 genes differentially expressed between the two immune compartments; 61 out of 88 participate in various processes that include cell growth and differentiation, cellular movement and morphology, immune response and other hematopoietic cell functions.
Unsurpervised clustering of highly expressed genes revealed two major SLE patient clusters in bone marrow, but not in peripheral blood. The first cluster was comprised of patients with active disease and the second patients in remission. The upregulated genes in the bone marrow of active patients included genes involved in cell death and
4
granulopoiesis. These genes represent components of activated neutrophils as well as positive or negative regulators of cell viability and apoptosis. Linear regression analysis showed that granulopoiesis signature was significantly correlated with SLEDAI in the bone marrow (r = 0.55, p = 0.0259), and the total score was higher in active group of patients versus the inactive (p =0.041).
Serum levels of 22 cytokines were compared between SLE patients and controls. Among the “cellular cytokines”, TFNα ( p=0.0486) was significantly elevated in SLE patients compared to controls, while IL-15 ( p= 0.0097) was also increased in patients. We did not observe any changes in serum “humoral cytokine” levels. Serum chemokine IP10, IL8 and MCP1 levels were also higher in SLE patients. Serum IP10 and IL12 concentration showed a negative association with arthritis. The chemokine MIP1α showed a negative association with neuropsychiatric manifestations of lupus while serum IL2 concentration positively associated with lupus nephritis.
Conclusion: Compared to the peripheral blood, microarray analysis of the bone marrow better differentiated active from inactive lupus patients and patients from controls. These data corroborate previous findings regarding the importance of apoptosis and granulocytes in the pathogenesis of the disease. Furthermore, two cellular cytokines, TNF-alpha and IL15, characteristic of the Th1-mediated immune response, were elevated in patients’ sera relative to controls. Although lupus is considered by many as a Th2 cytokine disease none of the cytokines associated with humoral immunity were found elevated in the patients’ cohort relative to the controls.
|
Search
