from DCs that present antigen [12]. In the majority of cases, GC-like lesions in SS were marked by CD21+ follicular dendritic cell networks [48] and signs of proliferation and apoptotic events were described in another study [9, 16]. B-cell fate in this process is believed to depend on the afÞnity of the surface antibody to the antigen and their responsiveness to anti-apoptotic signals [12]. B cells then have to interact with TH cells, which provide Þnal differentiation signals for an adaptive humoral immune response [12].

Interestingly, ectopic GC-like structure formation in SS was indeed paralleled with increased local production of antibodies against Ro and La [16]. Furthermore, patients presenting GC-like structures in the salivary glands presented higher degrees of glandular inßammation and elevated serum titers of RF, anti-Ro, anti-La, and total IgG [48, 96]. The formation of ectopic GC-like follicles in non-lymphoid organs might indeed participate in the pathogenesis of SS and indicate a more severe disease. The occurrence of GClike structures in salivary glands has also been proposed as a potential predictor for the subsequent development of lymphoproliferative disorders [97] (Fig. 15.2). However, GC-like structures were not associated with salivary gland enlargement that is today deÞned as a principal risk factor for lymphoma in SS. In addition, similar signs of ectopic lymphoid tissue formation and lymphoid neogenesis also occur in RA synovia [98], in the thyroid gland of HashimotoÕs thyroiditis [99], and chronic infectious reactions triggered by, for example, Helicobacter pylori in the gut mucosa [100]. To what extent ectopic GC-like structures in SS may overtake processes traditionally assigned to GC in secondary lymphoid organs and what molecules pave the way for their ectopic formation remain to be proven.

cause of hyoposalivation [101] (Figs. 15.2 and 15.3). Hence, loss of secretory capacity, degree of lymphoid inÞltration, and production of speciÞc autoantibodies have been anticipated to correlate with each other and indicate disease state and disease severity [102]. However, several SSrelated observations and substantial experimental evidence could not be accommodated in such a model. Firstly, a lack of correlation between the amount of destroyed glandular tissue and the often disproportional decrease in salivary ßow is often observed [103]. Secondly, glandular tissue isolated from patients with SS retained some functionality although appearing less sensitive to stimuli provided in in vitro systems [104]. Thirdly, administration of muscarinic receptor agonists (pilocarpine hydrochloride or cevimeline) signiÞcantly improved exocrine gland secretion capacity in many patients with SS [105]. A model of SS pathogenesis comprising mechanisms of glandular dysfunction is further supported by the observation that certain murine strains, which naturally [24] or as a result of genetic modiÞcation [70Ð72], retain full secretory function despite severe glandular inßammation. Furthermore, prevention of hyposalivation in NOD mice as a result of heat shock protein (Hsp)60 immunization was not paralleled by a greater decrease in salivary gland inßammation compared to mice in which the onset of hyposalivation could not be prevented [19]. Assuming that glandular destruction could be a result of inhibition of saliva secretion, and not hyposalivation to be the consequence of glandular destruction, led to the preposition of immune system-related processes, which appear to have the potential to mediate glandular dysfunction in SS [106] (Fig. 15.3).

15.2Mechanisms Mediating Salivary Gland Dysfunction

The original reasoning for the impairment in exocrine gland function in SS suggested loss of glandular epithelial cells as a result of lymphocyte-mediated cell death to be the main

15.2.1Acinar Cell Innervation and Humoral Immunity

The secretion of water and electrolytes from acinar cells is directly induced by acetylcholine (ACh) and substance P released by the innervating parasympathic nerve ends [107]. In the salivary glands M1R and M3R are the predominant acetylcholine receptor subtypes, whereas

M3 is predominant in the lacrimal glands [108]. Experiments using knockout strains revealed a predominant role of the M3 subtype in triggering saliva secretion [109, 110]. In contrast to neuromuscular junctions the physical distance between nerve ends and acinar cells is rather long, which leaves the process of neuronal innervation of acinar cells susceptible to events such as degradation of ACh by acetylcholine esterase and antibodies to muscarinic receptors [106].

The importance of the latter was illustrated when studying NOD Igμnull mice, which lack

functional B cells. Although T-cell-dominated

focal inßammation was found in the salivary glands of NOD Igμnull mice, they retained sali-

vary ßow rates comparable to control strains [111]. Reversible induction of hyposalivation through serum transfer from old NOD mice and IgG fractions obtained from patients with SS in these mice [111] further supports the notion of a contribution of serum components to the pathogenesis of SS. Functional analyses demonstrated that IgG fractions from patients with primary SS reduced the carbachol-evoked increase in Ca2+ in murine and human acinar cells [112] as well as they had the potential to disturb muscarinic receptor-associated contraction of colon- [113] and bladder muscle cells [114]. Due to the laborious method, unfortunately, both tests described above are inadequate to screen large patient cohorts. The use of M3R transfectants in combination with ßow cytometry [115], or the application of newly developed enzyme-linked immunosorbant assays (ELISAs) [116], may represent an alternative to further investigate the role and diagnostic value of anti-M3R antibodies in SS. However, studies also highlighted the apparent challenges when attempting detection of antimuscarinic receptor antibodies by conventional immunochemical methods [117, 118]. Such difÞculties continue to hamper the identiÞcation of the true incidence of anti-M3R antibodies in SS. In addition, antibodies inhibiting the function of M3R have also been shown to be present in some patients with scleroderma [113].

ACh ligation with M3R in vitro has been shown to mediate protection from apoptotic cell death [119]. Hence, inhibition of anti-apoptotic

effects mediated through M3R signaling might potentially link functional quiescence with cellular destruction. However, chronic stimulation of membrane-bound M3R might also result in receptor desensitization over time [120].

In addition, it should be taken into account that mononuclear cells express muscarinic receptors, secrete ACh, and encode the synthesizing choline acetyltransferase and the degrading acetylcholine esterase [121], what adds to the complexity of how immune cells might affect the signaling mechanisms of saliva secretion [106].

15.2.2 Inflammatory Mediators

Results obtained in vitro have suggested that cytokines such as IL-1α and IL-1β may affect the process of saliva secretion by inhibiting the release of ACh from cholinergic nerves [122, 123]. Evidence against interferences of such kind was, however, presented when no relation between concanavalin A-induced cytokine production and Ach-evoked Ca2+ mobilization [124] was found.

Arguing against one aspect of the hypothesis involving MMP in the pathogenesis of SS [46, 125], namely, shedding M3R from the extracellular surface, is that, in contrast with murine models of SS, acini in the human labial salivary glands of patients with SS express signiÞcantly higher numbers of M3R compared to control individuals [126].

15.2.3Fluid Movement in the Salivary Glands and Aquaporins

Obviously, the process of saliva secretion requires movement of ßuid from the vasculature into the interstitial space of the salivary glands and ultimately into the ducts [26]. Water movement across the water permeable acini can occur either paracellularly or transcellularly [26]. SpeciÞc molecules conducting water in and out of cells, while preventing the passage of ions and other solutes critically, contribute to the process described above [127]. Termed

aquaporins (AQPs), these transmembrane proteins form channels in the cell membrane and increase the water permeability of the lipid bilayer by up to a 100-fold [127]. Different isoforms exist and show speciÞc cellular and subcellular distributions in salivary and lacrimal glands [128]. AQP-5 is located in the apical membrane of acinar cells, AQP-3 is present within the basal membrane of acinar cells and AQP-1 is expressed on myoepithelial cells [128]. Monoclonal antiM3R antibodies had the potential to prevent translocation of AQPs to the plasma membrane [129]. Since then, AQPs have been suspected to contribute to the loss of exocrine gland secretory function in SS (Fig. 15.3). Mice deÞcient of AQP- 5 exhibit an approximate decrease of 65% in salivary secretion capacity compared to wild-type mice [130] and AQP-5 has also been suggested to contribute to reduced saliva secretion observed in NOD mice [131]. However, analysis of AQP- 5 distribution within the glandular tissue from patients with SS revealed controversial results [132, 133]. AQP-1 expression was decreased by 38% in salivary glands of patients with SS suggesting a possible insufÞcient AQP-1-dependent water ßow across myoepithelial cells [134].

developing a certain autoimmune disease [138, 139]. A genetic predisposition for SS also seems to exist; a notion supported by reports about the accumulation of SS in certain families and among twins [140]. Subsequently, genes coding for immunorelevant molecules have been studied to identify certain genes, genotypes, and single-nucleotide polymorphisms, which might be associated with SS. A variant of the minor histocompatibility antigen HA-1 has been associated with reduced risk of primary SS [141], whereas another study indicated a signiÞcant association between primary SS and a speciÞc interferon regulatory factor (IRF)5 allele [142]. In contrast, no association of SS with polymorphisms in Fas and FasL [143], IL-10, TNF-α, interleukin-1 receptor antagonist (IL-1RA) [144], CTLA-4 [145], TNF- α receptor 2 [146], or CCR7 [147] genes were observed. For more comprehensive discussion of genes and SS please see Chapter 8.

15.3.1 Environmental Factors

Environmental factors, most often sialotropic viruses, have been suspected to trigger the subsequent etiopathogenesis of SS through mechanisms of molecular mimicry in genetically sus-

Immunity and Etiology of Autoimmune Diseases: A View on Sjögren’s Syndrome

According to the paradigm of self versus nonself discrimination, the immune system, early in life, is taught to ignore or tolerate self, whereas it is supposed to recognize and attack foreign or non-self [12, 135, 136]. Ever since researchers studied the etiology of autoimmune diseases, the process of clonal deletion and spontaneous mutations of immune cells have been subjects of investigation in order to reveal at what level and by what process self versus non-self discrimination might have been compromised [12, 135, 137] (Fig. 15.2).

To a variable degree, depending on the autoimmune condition studied, an individualÕs genetic background deÞnes a personÕs susceptibility for

mimicry is thereby deÞned as the possibility that sequence similarities and structural homology between foreign and self-peptides are sufÞcient to result in the cross-activation of autoreactive T or B cells by pathogen-derived peptides to mount pathogenic effector responses against self [149].

The infectious agents which have received most attention in the Þeld of SS are EpsteinÐBarr virus [150], human T-cell leukemia virus-1 [151], and hepatitis C virus (HCV) [152]. Sialadenitis, now considered as an extrahepatic manifestation of chronic HCV infection, received attention in the newly proposed AmericanÐEuropean Consensus Group criteria, in which HCV infection is listed as an illegibility criterion in clinical studies investigating SS [1].

To elaborate a possible viral etiology of

Sialadenitis was observed as a result and in one particular strain anti-Ro and anti-La antibodies were observed [153]. Apoptotic cells, however, were solely detected during the acute, but not during the chronic phase of the inßammation and Fas and TNF-receptor I-mediated apoptosis seemed to be critical in clearing MCMV-infected cells from salivary glands [154]. As suggested by the authors, such defects may lead to a postinfectious, chronic inßammation resembling the histopathology of SS [154]. SigniÞcant reduction in both the number of inßammatory foci and extent of tissue destruction in salivary glands were observed as a result of local FasL gene transfer [154]. A more recent study identiÞed a coxsackie virus as a potential agent involved in the induction or maintenance of SS in a Greek population [155]. However, the results could not be validated in a French patient cohort [156].

Although, associations between SS and all viral candidates yet investigated are rather weak, microarray-based investigation of the salivary gland transcriptome in patients with SS [3] and in congenic mouse models for SS [157, 158] indicated an activated type-I and type-II IFN system, which might originally have been triggerred by a viral infection. Critically promoting host defense against viruses, IFNs do directly affect multiple cell types and processes involved at an early stage of the immune reaction such as activation of NK cells and macrophages and may also facilitate antigen presentation to lymphocytes by pDC [159]. For more comprehensive discussion of viruses, genes, and SS please see Chapter 8.

and to T cells through APCs (co-stimulation) (Fig. 15.2). APCs in turn receive secondary signals via pattern recognition receptors (PRRs) such as TLRs [12]. PRRs are germline-encoded and received much of attention as they may

ary distance between a host and the invading microorganism [34]. Due to their role in either supressing or activating APCs, TLRs have more recently received considerable attention in the Þeld of autoimmunity [35]. Research has increasingly demonstrated that many of the same PRRs also recognize self-epitopes that either are released from dying or distressed cells or are present at the surface of apoptotic cells or apoptotic bodies [35].

Activated TLR3 and TLR7 pathways, prior to manifestation of the disease, have been described in the salivary glands in an experimental model of SS [158]. Furthermore, epithelial cells and macrophages within the salivary glands from patients with SS express TLRs [32, 33]. An interesting concept, involving the evolution of TLRs, revolves around the question of how the immune system is able to also take into consideration tissue-speciÞc properties when shaping an efÞcient immune response [160, 161]. Considering recognition of endogenous danger signals to be recognized by PRR would attribute to the distressed tissue and its mediators a distinct role in the hostÕs decision on immunological tolerance, immune activation, and qualitative aspects of the immune reaction [160, 161] (Fig. 15.2).

15.3.2 Secondary Signals

Mounting an adaptive effector response requires the co-operation of at least two different cell types in each distinct phase of the immune response, e.g., APC/T-cell interaction and T-cell help delivery to B cells [12] (Fig. 15.2). TodayÕs concept of immunity, besides the recognition of an assumingly foreign peptide, acknowledges the importance of secondary signals in the hostÕs decision making process of self versus non-self discrimination [12]. These secondary signals are provided to B cells by TH cells (T-cell help)

15.3.3Apoptosis, Autoantigens, and Potential Danger Signals in the Salivary Glands

Asking the question why the salivary glands are often the primary target organ of a distinct pathogenic immune reaction, it has been speculated that due to their anatomical localization and their short excretory ducts exocrine glands should be considered as a locus minoris resistentiae, where apoptotic acinar cells cannot in the usual way be scavenged as they disintegrate