This study is the first to show the clinical significance of CSF anti-Sm. Thus, the results in the present study have disclosed that CSF anti-Sm is elevated in ACS, a severe form of diffuse NPSLE. By contrast, there were no significant differences in CSF anti-RNP among various subtypes of NPSLE, although both CSF anti-Sm and anti-RNP were increased in NPSLE compared with non-SLE control. It has been pointed out that anti-Sm might be involved in the pathogenesis of NPSLE [9],[10]. Especially, serum anti-Sm was associated with organic brain syndrome, consisting mainly of ACS of diffuse NPSLE [11]. Consistently, serum anti-Sm appeared to be elevated in ACS in the present study as well, despite the lack of statistical significance.
Previous studies demonstrated that CSF anti-RNP was associated with NPSLE and CNS manifestations in MCTD [12]. However, the elevation of CSF anti-RNP was not specific in ACS or diffuse NPSLE, since CSF anti-RNP was also elevated in patients with neurologic syndromes, including aseptic meningitis, headache, demyelinating disorder or movement disorder [12]. Accordingly, in the present study, CSF anti-RNP was also elevated in focal NPSLE comparably to diffuse NPSLE, confirming the observation in the previous studies [12]. How anti-RNP is involved in the development of NPSLE is currently unknown. Although the possibility of induction of proinflammatory cytokines by anti-RNP was suggested [12], further studies are required to confirm this point.
Autoantibodies to NMDA receptors, a subgroup of the glutamate receptor family, have recently attracted increasing attention [5]-[7],[21],[22]. Thus, DeGiorgio et al. showed that injection of anti-NR2 glutamate receptor binding antibodies (purified antibodies from the sera or CSF from NPSLE patients) into mice brain resulted in apoptosis of the neuronal cells without signs of inflammation [6]. Of note, Kowal et al. have demonstrated that mice induced to express anti-NR2 in systemic circulation have no neuronal damage unless breakdown of the BBB takes place [7]. Importantly, we have demonstrated that monoclonal anti-Sm binds to the surface of SK-N-MC cells and Neuro2a cells, indicating that anti-Sm reacts with neurons. Moreover, the presence of greater amounts of anti-Sm in CSF was associated with ACS in the present study. Notably, the effect of anti-NR2 antibodies on neurons has been shown to be dose dependent [22]. Thus, at low concentrations they alter synaptic function, whereas at higher concentrations they can cause neuronal cell death by apoptosis [22]. It is therefore suggested that the presence of higher concentrations of anti-Sm within the CNS might cause more extensive neuronal damage, resulting in the development of ACS. The influences of anti-Sm on the function and survival of neurons are currently undetermined and need to be explored in further studies.
It has been well recognized that intrathecal production of immunoglobulin is increased in NPSLE irrespective of focal NPSLE or diffuse NPSLE [18],[19]. Notably, previous studies revealed that CSF anti-RNP index was elevated in NPSLE [12]. In the present study, CSF anti-RNP index as well as CSF anti-Sm index was elevated in NPSLE compared with non-SLE control. However, there were no significant differences in CSF anti-Sm or anti-RNP index among various subsets of NPSLE. Therefore, the elevations of CSF anti-Sm levels in ACS compared with non-ACS diffuse NPSLE or with focal NPSLE cannot be accounted for by the increased intrathecal synthesis of anti-Sm.
BBB dysfunction results in the elevation of CSF immunoglobulin through the increased transudation from the systemic circulation into the CNS. In the present study, Q albumin values were not significantly elevated in NPSLE compared with non-SLE control, consistently with the previous studies [18],[19]. However, among various types of NPSLE, Q albumin was significantly elevated in ACS compared with non-ACS diffuse NPSLE or with focal NPSLE in the present study, as is consistent with our recent studies [20]. On the other hand, CSF anti-Sm was significantly correlated with Q albumin, and more closely with serum anti-Sm. However, there were no significant differences in serum anti-Sm among various subsets of NPSLE and non-NPSLE, although it appeared to be higher in ACS. Taken together, these data indicate that the damage in BBB rather than the elevation of serum anti-Sm is more crucial for the elevation of CSF anti-Sm in ACS.
We have recently demonstrated that CSF anti-NR2 levels and Q albumin were significantly higher in ACS than in non-ACS diffuse NPSLE, indicating that the severity of BBB damage plays a crucial role in the development of ACS through the accelerated entry of larger amounts of anti-NR2 into the CNS [20]. The data in the present study have further disclosed that the elevation of CSF anti-Sm due to the damaged BBB is also involved in the development of ACS. In addition, CSF anti-Sm was significantly correlated with CSF anti-NR2 in NPSLE in the present study. Since there was no significant correlation between serum anti-Sm and anti-NR2, the positive correlation between CSF anti-Sm and anti-NR2 might be due to the BBB damage. More importantly, the data in the present study indicate that the elevation of both anti-Sm and anti-NR2 in CSF plays a crucial role in the development of ACS. Further studies to explore the effects of coexistence of anti-Sm and anti-NR2 on neuronal cells would be important to delineate the precise mechanism for the development of ACS.
The mechanism of damage in BBB in ACS has not been determined at present. In this regard, it is likely that several autoantibodies, such as anti-ribosomal P protein antibodies and anti-NR2 antibodies, might result in BBB damage, since they react with endothelial cells [23]-[25]. Notably, recent studies have disclosed that an anti-Sm autoantibody synergized with hemoglobin to enhance the secretion of proinflammatory cytokines while eliciting the increased production of monocyte migratory signals from endothelial cells [26]. It is therefore also possible that anti-Sm also might result in BBB damage. Further studies are required to explore the roles of a variety of autoantibodies in BBB damage.
In the present study, there were no differences in CSF anti-Sm or CSF anti-RNP between non-ACS diffuse NPSLE and focal NPSLE. Since a number of autoantibodies have been reported to be reactive to neurons, including anti-ribosomal P protein antibodies [27], anti-Ro antibodies [28], some anti-cardiolipin antibodies [29] and anti-NR2 antibodies [6]-[8], it is possible that the patterns of expression of several antibodies or their combination in CSF might be different between non-ACS diffuse NPSLE and focal NPSLE. Moreover, it is also possible that anti-ribosomal P protein antibodies, anti-Ro antibodies and some anti-cardiolipin antibodies might be also involved in the development of ACS in combination with anti-Sm and anti-NR2. Further studies are required to delineate the whole spectrum of neuron-reactive autoantibodies in CSF in order to understand the variability of manifestations of NPSLE.
A limitation of our study is that it is cross-sectional and the observations are associations and not fully causal, though the binding of anti-Sm to neuroblastoma cell lines suggests plausibility. Another limitation is the possibility that anti-Sm might have preferential properties to penetrate the CNS, leading to its higher CSF levels just as a reflection of the peripheral milieu. Therefore, it would be ideal to have an additional control of SLE patients with anti-Sm and anti-RNP in sera without CNS symptoms and study CSF anti-Sm and anti-RNP in such patients. In this regard, 5 of the 23 patients with focal NPSLE showed elevation of serum anti-Sm over 50.0 U/ml without psychiatric manifestations. When compared with 5 patients with ACS diffuse NPSLE who showed almost the same values for serum anti-Sm, the 5 patients with focal NPSLE showed significantly lower CSF anti-Sm values (data not shown). Therefore, it is strongly suggested that elevation of CSF anti-Sm might be causal for ACS diffuse NPSLE.