Investigation of novel autoantibodies in Sjogren’s syndrome utilizing Sera from the Sjogren’s international collaborative clinical alliance cohort

Studies utilizing the IL-14α transgenic (IL14aTG) mouse model of SS demonstrated that loss of salivary gland function occurs before infiltration of the submandibular and lacrimal glands with lymphocytes [8,9]. Furthermore, anti-SP1 and anti-CA6 were demonstrated in the sera of the mice at 6 months of age, a time in which salivary gland function was lost but no significant lymphocytic infiltration was noted in the glands [5]. We were therefore interested in studying patients with dry eyes and dry mouth lacking lymphocytic infiltration of the salivary glands. The SICCA cohort contained 9 such patients with lip biopsies described as showing sclerosing chronic sialoadenitis but no evidence of lymphocytic infiltration. The group contained 4 males and had a mean age of 49.8 years. At the same time, we studied 40 sera from patients with mild disease (focus score < 1 per 4 mm²) and 40 patients with moderate disease (focus score > 3 per 4 mm²). The group with mild disease contained 3 males and had a mean age of 49.9 years. The group with moderate disease contained 2 males and had a mean age of 51 years. There were 50 normal controls that included 5 males with a mean age of 37.5 years. Figure 1 demonstrates that in the patients with focus score = 0, more patients expressed antibodies anti-SP1, anti-CA6 and anti-PSP than anti-Ro and anti-La (6 vs 2). In patients with mild (F <1) or moderate disease (F >3), the majority of the patients expressed anti-Ro or anti-La while fewer numbers of patient sera contained anti-SP1, anti-CA6 and anti-PSP. In each of the study groups, F = 0, F <1 and F > 3, the expression of the autoantibodies, anti-CA6, anti-PSP, anti-SP1, anti-Ro and anti-La, was statistically significant when compared to the normal controls (in all cases p < .0005). The increased expression of anti- CA6 in the F0 group compared to the F <1 group (p = .032) or the F > 3 group (p = .006) were statistically significant. Neither anti-PSP nor anti-SP1 reached statistical significance because of the small numbers in the F0 group, although there was a trend for their expression to be higher in the F0 group. On the other hand, the expression of anti-Ro was significantly reduced in the F0 group compared to the F <1 (p = .0021) and F > 3 (p = .0003) groups. The reduced expression of anti-La in the F0 group compared to the F <1 and F > 3 groups did not quite reach statistical significance.

In the focus score = 0 group, there were 7 patients lacking anti-Ro or anti-La. In the F <1 group 3 patients lacked anti-Ro or anti-La while in the F > 3 group 6 patients lacked these autoantibodies. Figure 2 demonstrates that in the anti-Ro or anti-La negative patients, most patients expressed anti-SP1, anti-CA6 or anti-PSP, although there were a few patients who expressed none of these autoantibodies in the focus score = 0 and the moderate groups. Because of the small numbers, these observations do not reach statistical significance. More patients need to be studied to confirm these trends.

Our understanding of the pathophysiology of SS is currently in flux. While early models of SS had suggested a disease largely driven by the adaptive immune system, more recent models have implicated the role of the innate immune system in the initiation of the disease [10]. Of note, in the IL14aTG model for SS the innate immune system is responsible for the majority of the destruction of the salivary and lacrimal glands [11,12]. Lymphocytes are found in the salivary and lacrimal glands only after significant loss of salivary gland function [12]. In this model, antibodies anti-SP1, anti-CA6 and anti-PSP occur during this early stage of disease, while antibodies anti-Ro and anti-La occur later in the disease course [5].

The data presented in this manuscript suggest that patients with low FS on salivary gland biopsies express anti-SP1, anti-CA6 and anti-PSP more frequently than anti-Ro and anti-La, just like the IL14aTG mice. The FS = 0 group, however, does not meet full criteria for SS, so they would not officially be given that designation. There are no data available to determine whether this group ever will meet full SS criteria. Previously published studies have shown frequent expression of anti-SP1 in patients with idiopathic dry eyes and dry mouth for less than 2 years [5]. What is unclear from this work is how many of these patients in time will meet full criteria for SS and how many of these patients will not progress further.

Interestingly, a recent study from Jonsson et al. demonstrated the existence of anti-Ro or anti-La in the sera of patients often before they developed clinical evidence of SS [13]. Similar finding were made in a previous study in SLE [14]. Many patients who developed SS, however, did not express anti-Ro or anti-La. Antibodies anti-SP1, anti-CA6 and anti-PSP were not evaluated in these studies.

One of the novel aspects of the antigens SP1, CA6 and PSP compared to Ro and La is that they are found selectively in the salivary and lacrimal glands. Ro and La are found virtually in every cell. It is unclear why antibodies anti-Ro and anti-La should be specific for SS, and in fact they are found in SLE and various other autoimmune diseases. Perhaps certain types of cellular injury are necessary before antibodies anti-Ro and anti-La appear. It has been postulated that SP1, CA6 and PSP may have roles in the adherence and/or clearance of various infections [15]. If this were in fact the case, it would be logical to suspect that a hapten-carrier system established with an infectious agent could lead to the development of antibodies to these antigens. If we postulate that SS is initiated by infections in the salivary and lacrimal glands, production of antibodies anti-SP1, anti-CA6 and anti-PSP early in the course of disease would make sense.

The role of autoantibodies in SS is currently poorly understood. While vaccination of mice with Ro results in salivary gland injury and anti-Ro antibodies, it is unclear whether the anti-Ro antibodies participate in the salivary gland injury, are part of a reparative mechanism for the injury and/or are merely a marker for disease [16]. Attempts have been made to correlate autoantibodies with particular disease manifestations in SS, such as anti-carbonic anhydrase 2 antibodies with renal tubular acidosis, however no direct link has been made between the autoantibodies and tissue dysfunction [17]. Similarly, the anti-SP1, anti-CA6 and anti-PSP antibodies are currently evaluated only as markers of disease. Future studies will need to investigate their significance.

While these studies suggest that anti-SP1, anti-CA6 and anti-PSP are early markers for SS, future studies will have to evaluate these autoantibodies in other cohorts of patients. It will be necessary to determine the long-term consequences of expression of these autoantibodies in particular patients and “normal controls”. These autoantibodies may denote particular stages of SS and/or particular forms of SS.