Elevated levels of soluble CD40 ligand (sCD40L) in serum of patients with systemic autoimmune diseases

Abstract

The CD40–CD40L costimulatory pathway is involved in the evolution of many autoimmune diseases including systemic lupus erythematosus (SLE), rheumatoid arthritis (RA) and Sjögren's syndrome (SS). Increased levels of sCD40L in the serum have been associated with disease activity in SLE. The aim of this study was to investigate the role of sCD40L in the development of lupus nephritis and examine its possible association with cryoglobulinemia in Sjögren's syndrome. We used a 2-site sandwich ELISA to measure the levels of sCD40L in sera, from 64 patients with SLE, RA and SS and 17 healthy blood donors. Biological specimens from the affected tissues such as urine from patients with lupus nephritis and saliva from patients with SS were also tested. In this regard, paired sera and first morning urine samples from 6 SLE patients (3 with active lupus nephritis and 3 with inactive lupus nephritis) were tested with the sCD40L ELISA protocol as well as paired sera and salivary samples from 5 patients with SS and cryoglobulinemia, 5 patients with SS and anti-Ro or anti-La autoantibodies and 5 age-matched healthy control donors. We also examined possible correlations of sCD40L levels with several laboratory and clinical parameters in SS and SLE. We found that sera from SLE and SS patients had significantly higher levels of sCD40L compared to sera from healthy control donors. No sCD40L was detected, in urine samples of patients with either active or inactive nephritis and in salivary samples from SS patients or normal subjects. Soluble CD40L is elevated in sera of SS and SLE patients but further investigation is needed to determine its possible role in SLE nephritis and Sjögren's syndrome.

Introduction

T lymphocytes regulate the specificity of immune response to foreign antigens and control the function of other immune cells such as B lymphocytes, monocytes and dendritic cells. Activation of costimulatory pathways such as CD40–CD40L and CD28–B7.1/B7.2 is needed to achieve an efficient adaptive immune response that leads to the activation, proliferation and differentiation of the specific T-cell clone.

In the past years the CD40–CD40L pathway has been studied extensively. CD40 molecule, a member of the tumor necrosis factor (TNF)-receptor family, is a 39 kDa transmembranic protein expressed on the surface of antigen-presenting cells including B cells, activated macrophages, dendritic cells, monocytes and follicular dendritic cells [1], [2] as well as a number of other cell types such as mesangial, endothelial and epithelial distal cells [3]. CD40L is a type II membrane protein of 33 kDa which belongs to the TNF gene family and is normally expressed on Th1, Th2 cells and activated platelets [4], [5]. The CD40–CD40L ligation provides the signals required for B-cell activation and differentiation, immunoglobulin class switching and further activation of T cells. In addition, CD40 ligation on DCs leads these cells to maturation in order to express an antigen presenting phenotype. CD40L has been also found to be expressed on a variety of other cell types such as platelets, B cells, mast cells, basophils and NK cells in many diseases [1]. CD40–CD40L interaction is involved in many autoimmune diseases such as systemic lupus erythematosus (SLE) [6], rheumatoid arthritis (RA) [7], experimental allergic encephalitis (EAE) [8], multiple sclerosis (MS) [9], inflammatory bowel diseases (IBD) [10], chronic urticaria (CU) [11] and in allograft rejection [12]. CD40L circulates in a soluble form of 18–20 kDa in the serum of patients with SLE [13], [14]. The soluble CD40L has been associated with disease activity and severity [14]. The major sources of sCD40L are activated T lymphocytes and activated platelets. Soluble CD40L circulates in monomer, dimmer or trimer form and is considered functional and capable of inducing a CD40–CD40 ligation with CD40 bearing cells [13].

The involvement of CD40–CD40L pathway in the pathogenesis of lupus nephritis has been documented by several studies in humans [6], [15], [16], [17], [18], [19], [20] and murine models [21], [22], [23], [24], [25]. In addition, the blockade of the pathway by anti-CD40L antibodies has ameliorated the clinical and laboratory parameters of nephritis in both mice [26], [27], [28], [29] and humans [30]. We assumed that the sCD40L is able to cross the glomerular barrier and potentially interact with CD40 bearing cells of renal tissue at the initial phase. Therefore, one of the aims of our study was to detect sCD40L in urine samples from SLE patients with active nephritis, considering that urine is originated directly from the injured tissue.

Similarly, few studies are referring to the role of the CD40–CD40L pathway in the pathogenesis of SS [31], [32], [33], [34]. Recently, it has been shown that salivary gland epithelial cells from pSS patients express high levels of CD40 molecule [35]. We speculated that sCD40L might interact with the CD40 bearing epithelial cells of the injured glandular tissue and create a further induction of their antigen-presenting phenotype. In the present study, we measured the levels of sCD40L in sera and salivary samples from patients with pSS and evaluated possible correlations with clinical manifestations such as cryoglobulinemia, glomerulonephritis and peripheral neuropathy, as well as serologic findings such as anti-Ro/anti-La autoantibodies and RF.