We have identified combinations of inflammation molecules that best predict the occurrence of NPS in both CSF (CRP, IP-10, Il-8 and sICAM-1) and serum (Eotaxin-3, IL-6 and CRP). Distinct marker combinations in CSF (TARC only) and serum (sICAM-1, IL-6 and IFN-γ) were associated with the severity of NPS. Individual NPS symptoms were associated with specific neuroinflammatory molecules in both CSF and serum. The identified CSF neuroinflammation molecules were associated with volume changes in specific brain regions. Finally, NPS were associated with more rapid cognitive decline at follow-up and this association was mediated by sICAM-1 CSF levels.
4.1 Specific CSF neuroinflammatory molecules are associated with NPS
Amongst the molecules related to NPS in our study, CRP and sICAM-1 have been previously associated with the presence of AD pathology (22, 33). However, the association of these neuroinflammatory markers with NPS is to our knowledge novel. In this study, sICAM-1 in particular, is strongly associated with NPS. This association could be driven by the large number of participants with AD pathology in the NPI-Q > 0 group. However, accounting for the presence of cerebral core AD pathology does not alter the specific combination of molecules associated with NPS in CSF. Therefore, this association is at least partially independent of AD. In this signature, CRP, IP-10 and sICAM-1 could play different roles as high concentrations of CRP and IP-10 are associated with lower NPI-Q scores, suggesting a “neuroprotective” effect; while a higher concentration of sICAM-1 is associated with the presence of NPS, suggesting a deleterious role.
We have shown this CSF signature is independent of cognitive status suggesting neuroinflammation leading to NPS also occurs in the absence, or before the onset, of cognitive decline. Furthermore, this signature is not dependent on BBB function, suggesting that the related inflammatory process originates within the CNS. We have also shown that it is not only the absolute concentration of these molecules that is associated with NPI-Q score, but rather their relative concentrations between one another and their pattern of expression and interplay. For example, IP-10 does not display a different concentration between participants exhibiting NPS and those who do not, and does not correlate with total NPI-Q score either; it does however strongly associate with the occurrence of NPS when considering changes in CRP and sICAM-1 concentrations. This novel finding is in accordance with the concept that cytokines form a complex network enhancing and/or suppressing the production of each other (34).
4.2 A distinct serum inflammation signature is associated with NPS
We identified a distinct serum inflammatory signature of the occurrence of NPS containing Eotaxin-3, IL-6 and CRP. Unlike the CSF inflammatory marker profile, this signature is related to cognitive status, BBB function and the presence of cerebral core AD pathology. This confirms systemic inflammation may both contribute to predisposing to or enhancing CNS inflammation which may further result in cerebral dysfunction and neuronal injury, and the manifestation of both cognitive impairment and NPS (25). In has been previously shown that systemic inflammation may also reflect cerebral pathology (26) and neuroinflammation related to NPS (22). It is also plausible that following BBB breakdown, caused by AD or neuroinflammation (29), cross-talk between both CNS originating and circulating inflammation occurs (35) as is the case in aging (36). These data suggest a deleterious feedback loop where circulating neuroinflammatory molecules such as IL-6 and IFN-γ can further enhance expression of CNS inflammatory molecules (CRP and IP-10 respectively, (37, 38)) that together can further damage the BBB (39) and activate microglia (40). The result is an escalation of neuroinflammation, further contributing to CNS processes leading to the manifestation of NPS. While we do not elucidate here the origin of these inflammatory processes, since subjects with manifest unstable medical conditions, including inflammation were excluded from the present study, we show these specific markers are particularly relevant for the occurrence of NPS.
4.3 Specific mechanisms relate to the severity of NPS
Distinct molecules, with the exception of sICAM-1 were associated with the severity of NPS both in CSF and serum in our study. This suggests that besides the identified inflammatory processes associated with the appearance of NPS, additional mechanisms may modulate their extent and severity. We therefore suggest that CNS inflammation and BBB breakdown could trigger the appearance of NPS. Following these events, CSF TARC along with circulating, IL-6, IFN-γ, MCP-4 and IL-16, could regulate and enhance existing symptoms or play a regulatory role in the inflammatory response and dictate specific and localized responses.
4.4 Individual NPS symptoms have specific pathological mechanisms
Several individual NPS symptoms were associated to specific neuroinflammatory molecule signatures in CSF and serum. Amongst these, the strongest associations were found with the CSF signatures of Anxiety and Disinhibition and the serum signature of Depression. Symptom specific profiles differed between CSF and serum in all cases. Some of the molecules involved in these signatures (CRP, sICAM-1, IP-10, VEGF) are part of the signature of overall presence of NPS.
Previous evidence suggests neuroinflammation can lead to a variety of NPS (21). We however describe novel and specific associations of the identified molecules, such as sICAM-1 and VEGF-D, with multiple NPI-Q categories suggesting that these symptoms may have common pathogenic mechanisms involving the associated molecule. In particular, higher CSF sICAM-1 levels were positively associated with symptoms of Depression, Anxiety, Apathy and Disinhibition in our cohort and could therefore play a role in the pathogenesis of all these symptom. We also found concentrations of VEGF-D in serum to be associated with the occurrence of Agitation, Depression, and Night-time behavior disorders. Furthermore, serum VEGF-D was associated with the occurrence of NPS only in the presence of cerebral AD. This suggests a role for this molecule in the pathogenesis of these NPS in the specific context of AD.
Contrary to these molecules, some of the identified molecules were associated with only a single symptom category; some of which have previously been reported, such as serum IP-10 with sleep disturbances (41) and serum IL-8 with Anxiety (42). The associations of MCP-4 with Agitation and CSF MDC with Night-time behavior however are novel. Overall, these findings suggest that while neuroinflammation results in a wide spectrum of neuropsychiatric manifestations, specific neuroinflammatory mediators could play a more important role in certain single syndromes.
4.5 Brain regions associated with individual NPS symptoms
The molecules we identified as part of the CSF neuroinflammatory profiles related to NPS have previously been associated with volume changes in specific brain regions (43–45), together suggesting neurodegeneration associated with neuroinflammation within these individual regions. We also observed that the volume of specific brain regions is associated with the concentration of neuroinflammatory markers.
The strong association of sICAM-1 with the volume of the hippocampus and 3rd ventricle (reflecting atrophy of surrounding areas) suggests these regions may play a role in the pathogenesis of NPS, although we cannot exclude sICAM-1 is linked to NPS via mechanisms independent of atrophy. Interestingly, these regions have been associated with the progression of AD pathology and inflammation in previous work (46, 47). Considering our observations and previous reports (13), we infer that decreased volume of the hippocampus and increased 3rd ventricle volume may indicate regional neurodegeneration and neuroinflammation together involved in the pathogenesis of NPS of the symptoms associated with sICAM-1 (i.e. Anxiety, Depression and Disinhibition).
4.5 NPS, neuroinflammation and clinical disease progression
While the occurrence of NPS is associated with the presence of AD pathology, we have shown that the CSF neuroinflammatory signature of NPS is independent of AD. This finding suggests that at CNS level both AD core pathology and neuroinflammatory processes may engage similar pathways leading to NPS. CSF sICAM-1 was previously found to correlate with both tau and pTau181 levels, confirming it is involved in AD-related tau-pathology and neural injury (22). In our models, the interaction between the presence of AD pathology and sICAM-1 was significant, suggesting that sICAM-1 is present in both the AD and neuroinflammation pathways related to NPS.
Higher NPS severity was associated with more rapid cognitive decline. This is in line with previous research describing an association of NPS with more rapid clinical disease progression in AD (4, 10, 9). The sICAM-1 CSF concentration is also associated with cognitive decline and our interaction analysis suggests that the association between NPS and cognitive decline is mediated by increased sICAM-1 in the CNS. A possible underlying mechanism is through altered cerebrovascular reactivity effects with which ICAM molecules are associated (48) which in turn, are known to differ according to cognitive status (49).
4.6 Strengths and limitations
Subjects included in this study had no psychiatric affection or symptom that could interfere with cognition and candidates with more marked neuropsychiatric symptoms were not considered. Consequently, there was a low frequency of some of the single NPS symptoms such as delusions and hallucinations in this cohort which does not allow to address possible relationships between inflammation and these symptoms. Whether the identified signatures may be used in clinical practice as markers of neuroinflammation-related NPS and targets for intervention needs to be further investigated in independent samples. Strengths of this study are the inclusion of elderly subjects with normal cognition or cognitive decline and the assessment of a large panel of inflammatory markers in paired serum and CSF samples. Furthermore, we addressed the relationships of inflammation related NPS with the core cerebral AD pathology, regional brain atrophy, and cognitive decline over time.
4.7 Conclusion
We have identified specific CSF and serum inflammatory signatures associated with NPS that can be considered both contributors to the underlying cerebral pathology and potential biomarkers of NPS. While the CSF signature identified here appears to indicate inflammatory processes that originate within the CNS and interact with the core AD pathology, the serum signature may represent systemic dysregulation of inflammatory activity related to BBB function and impacting CNS processes that lead to NPS. The inflammatory signatures of NPS indicate symptom specific underlying processes opening the perspective of targeted interventions to reduce NPS and their long-term consequences.