Association of Cerebrospinal Fluid (CSF) Insulin with Cognitive Performance and CSF Biomarkers of Alzheimer’s Disease

INTRODUCTION

Over the last decades, a growing body of evidence from in vitro experiments, animal models, and observations in patients has linked abnormal insulin signaling to Alzheimer’s disease (AD) [1–4]. Multiple pathways may account for this, which have been extensively reviewed elsewhere [1–4]. Insulin is, for example, believed to interfere with the metabolism of amyloid-β (Aβ) and tau [3].

At present, brain insulin signaling cannot be measured directly in humans in vivo. Evidence of deficient insulin signaling in patients with AD therefore stems from postmortem brain studies showing fewer insulin receptors and decreased downstream signaling activity in the brains of individuals with AD as compared to age-matched controls [5]. To overcome the lack of direct measures, previous population-based studies used blood-derived markers of peripheral insulin resistance as a possible indicator of deficient brain signaling. These studies generally showed that hyperinsulinemia [6–8] and insulin resistance [6, 10] are associated with cognitive impairment [8–10] and risk of AD [6–8, 10], although these associations might be limited to, or most prominent in, women [9] and non-carriers of the apolipoprotein E ɛ4 (APOE ɛ4) allele [7, 10]. More recently, blood-derived markers of insulin resistance have also been related to cerebrospinal fluid (CSF) biomarkers of AD in cognitively asymptomatic individuals, but these associations depended on APOE ɛ4 genotype [11, 12].

Evidently, peripheral insulin levels and insulin sensitivity may not accurately reflect the cerebral situation [13]. Transport of insulin across the blood-brain barrier decreases with increasing insulin resistance [14] and there appears to be local production of insulin in the brain [15]. This may explain why there is no clear correlation between plasma and CSF insulin levels [15]. Published data on CSF insulin in relation to AD are, however, scarce, show variable results, and generally (with one exception [16]) did not account for the potential modifying effects of sex and APOE ɛ4 genotype [17–21]. At the same time, an increasing number of clinical trials have shown that intranasal insulin administration enhances memory function (in AD) (e.g., [22–26]), particularly in women and those without APOE ɛ4 [24–26]. Here, we made use of a standardized multicenter memory clinic population to assess the associations of CSF insulin with cognitive performance and CSF biomarkers of AD [i.e., Aβ_1-42, Tau, and phosphorylated Tau (p-Tau)] and explored whether any such association differed by sex or APOE ɛ4 genotype. We hypothesized that higher CSF insulin levels, likely reflecting cerebral insulin resistance, would be associated with ADpathology.

METHODS

RESULTS

Table 1 shows the demographic and clinical characteristics of the study population as a whole, by sex, and by diagnostic group [i.e., SCI (n = 45); aMCI (n = 44), and AD (n = 49)]. Overall, patients were in late adulthood (66±9 years), with a slight male predominance (65.2%). Men had somewhat unhealthier lifestyle variables than women and more frequently reported a history of cardiovascular disease. Cerebrovascular disease was, however, equally distributed between the sexes. Of note, the cardiovascular risk profile was similar across the diagnostic groups, except for the presence of diabetes, which was least common in the SCI group.

DISCUSSION

The present study shows that, among women and non-carriers of the APOE ɛ4 allele, higher levels of CSF insulin were associated with worse cognitive performance and/or higher levels of CSF (p-)Tau. CSF insulin did, however, not differ between diagnostic groups of cognitive impairment (i.e., SCI, aMCI, and AD). These findings add to the growing body of evidence indicating that abnormal insulin signaling is involved in AD [1–4] and also provide an explanation for why women and non-carriers of the APOE ɛ4 allele seem to benefit most from intranasal insulin administration to enhance cognitive performance (in AD) [24–26].

There are few earlier studies on the association between CSF insulin and AD [17–21]. Results on CSF insulin levels in patients with AD were inconsistent and reported CSF insulin to be increased [17], decreased [18, 20], or unchanged [19, 21] in individuals with AD as compared to healthy individuals [17–19, 21] or those with SCI [20]. The most probable explanation for these variable results across studies is the use of different study populations. For example, studies reporting decreased levels of CSF insulin in AD [18, 20] excluded individuals with diabetes, while the present study provides evidence that the presence of diabetes is associated with higher levels of CSF insulin. Other differences between the populations concern the severity of the dementia and the use of different reference groups.

Substantially more research has been conducted on the association between cerebral glucose metabolism, based on fluorodeoxyglucose positron emission tomography (FDG-PET) [36–42] or the CSF to plasma glucose ratio [43], and AD pathology. Collectively, these studies suggest that cerebral hypometabolism is more prominent in individuals with AD [42] and is associated with higher CSF(p-)Tau [36–39] and lower CSF Aβ_1-42 [36, 43] levels. Although it has been suggested that cerebral hypometabolism may reflect cerebral insulin resistance [43], it is important to realize that glucose uptake in the brain is largely independent of insulin levels [44] Even in the hippocampus, one of the brain regions with the highest concentrations of insulin sensitive GLUT-4 receptors, it is believed that glucose update does not necessary require insulin [44]. The link between cerebral hypometabolism and AD does, thus, not directly support a role for insulin resistance in AD pathology. This is particularly the case because glucose itself may have a direct effect on tau metabolism [45] and because it has been suggested that tau-related pathology may precede cerebral hypometabolism rather than being a consequence of it [46].

Despite the numerous reviews that have been published on the mechanisms through which cerebral insulin might interfere with Aβ and tau metabolism (e.g., [4, 47]), only one previous study has actually evaluated CSF insulin in relation to CSF Aβ_1-42 and (p-)Tau levels [20]. The overall null results of that study are in line with the null associations we observed in our overall study population. We, however, now show that higher levels of CSF insulin were associated with higher levels of CSF (p-)Tau in women and non-carriers of the APOE ɛ4 allele, which reinforces data from experimental studies linking insulin to tau metabolism. From a pathophysiological viewpoint, higher levels of CSF insulin, potentially reflecting cerebral insulin resistance and thus deficient brain insulin signaling, may result in reduced activity of the phosphoinositide 3-kinase-protein kinase B pathway. This, in turn, leads to over activation of glycogen synthase kinase-3, and consequently, hyperphosphorylation of tau [47]. That the association between CSF insulin and tau differs with APOE ɛ4 genotype could be explained by a higher degree of insulin resistance among non-carriers relative to carriers of the APOE ɛ4 allele, as illustrated by differences in insulin-mediated glucose disposal rates [16]. Alternatively, a potential association between CSF insulin and tau in APOE ɛ4 carriers might have been simply overshadowed by the high AD risk associated with APOE ɛ4 carriership.

The sex differences observed in our study are in line with experimental data and clinical studies showing that the brains of men and women are differentially sensitive to the effects of insulin [23, 49]. More specifically, previous studies have shown that the anorexigenic effects of intranasally administered insulin are more prominent in men, while its memory enhancing effects are more prominent women. It has been suggested [24] that these sex differences can be explained by the cognitive effects of estrogen [50]. A more recent study, however, showed that young and postmenopausal women are comparable sensitivity to the memory enhancing effects of intranasal insulin [51], which reiterates that estrogen explains the observed sex differences. What explains these differences should be explored in future studies. Perhaps it may relate to other physiological or anatomical factors that differ between sexes, for example the density of hippocampal insulin receptors.

Notably, we did not observe any association between CSF levels of insulin and Aβ, while a previous study noted that a single dose of intranasal insulin increased plasma levels of the short form of Aβ (i.e., Aβ₄₀) [52]. A potential explanation for this finding are the complex and dual effects insulin (signaling) may have on Aβ clearance [4]. On the one hand, insulin may promote Aβ clearance by accelerating its intraneuronal transport and increasing protein levels of the insulin degrading enzyme (IDE), a metalloprotease that is able to degrade Aβ [4]. On the other hand, IDE also catabolizes insulin, for which it has a higher affinity, which means that insulin at the same time can inhibit Aβ degradation [4]. An alternative explanation is that cerebral insulin is involved in AD via pathways other than Aβ metabolism (reviewedin [33]).

The present study has limitations. First and foremost, CSF samples were not necessarily taken in the fasting state, nor was lumbar puncture performed at a fixed time of the day. The time of day at which lumbar puncture was performed was also not recorded in the database. Hence, we were unable to adjust our analyses for potential biorhythm effects, while the production and secretion of at least insulin [53] and Aβ [54] seem to follow a circadian rhythm. Although this may have led us to underestimate the association of CSF insulin with AD pathology, it is unlikely to explain the observed modulating effects of either sex or APOE ɛ4 genotype. A second limitation that may have further affected our estimates of the association between CSF insulin and CSF p-Tau concerns the use of an antibody for p-Tau that was specific for phosphorylation at threonine 181, while deficient insulin signaling leads to a widespread phosphorylation of tau at multiple sites. Next, it can be questioned whether higher CSF insulin levels reflect cerebral insulin resistance the way hyperinsulinemia in the plasma reflects peripheral insulin resistance. Note, however, that individuals with diabetes, who are the most likely to have cerebral insulin resistance despite the fact that cerebral and peripheral resistance not necessarily concur [13], had higher levels of CSF insulin. Fourth, multiple analyses were performed without correction for multiple testing, though stratified analyses were only performed in case of statistically significant interaction with a clear a priori hypothesis. Fifth, in the current study, no blood samples were analyzed. Such data would have provided valuable information on how peripheral and cerebral insulin metabolism may interact in relation to cognitive performance and ADpathology, although compartmentalization is also possible. Finally, it can be questioned whether the composition of the CSF is directly comparable to that of the brain’s interstitial fluid [51]. Undoubtedly, however, CSF insulin is a better indicator of deficient brain signaling than blood-derived markers.

In conclusion, our results indicate that higher CSF insulin levels are related to impairment in cognitive performance and biomarkers of AD among women and non-carriers of the APOE ɛ4 allele. As such, our data suggest that the potential involvement of cerebral insulin (resistance) in the pathogenesis of AD is not uniform across individuals. Future studies are needed to determine whether endogenous high CSF insulin levels indeed reflect cerebral insulin resistance, as this might have implications for selection of patients who may benefit from the intranasal use of insulin to improve or preserve cognitive performance. Longitudinal studies are required to strengthen the pathophysiological relevance of ourfindings.