In this exploratory study, the reduced cerebral glucose uptake in type 2 diabetes is corroborated and increased fatty acid uptake (estimated as [18F]FTHA Ki values) in white matter is observed. Another important novel observation is the dependence on APOE-ε4 for fatty acid uptake. Carrying APOE-ε4 results in higher fatty acid uptake in lean controls. Among non-carriers of APOE-ε4, participants with type 2 diabetes had higher [18F]FTHA Ki than lean participants in all investigated brain regions. We also demonstrate lower cerebral perfusion among overweight participants and participants with type 2 diabetes compared with lean participants, and a negative correlation between total tau in CSF and cerebral perfusion. Finally, we report negative correlations between biomarkers of deranged metabolism (HOMA-IR, fasting plasma insulin, HbA1c) and both cerebral perfusion and glucose uptake, and positive correlation between these biomarkers and [18F]FTHA Ki in white matter.

While reduced cerebral perfusion is an established finding in Alzheimer’s disease [25], previous perfusion studies in type 2 diabetes have reported mixed results [19, 20, 26]. Since lower perfusion was also observed in overweight individuals, our findings suggest that alterations in cerebral perfusion may occur before the onset of type 2 diabetes or are related to overweight rather than diabetes itself. The finding that HOMA-IR, fasting insulin levels and HbA1c are inversely associated with cerebral perfusion indicates that the common factor explaining low cerebral perfusion in diabetes and overweight is likely insulin resistance, which is a documented risk factor for Alzheimer’s disease. That cerebral perfusion correlated negatively with total tau levels in the CSF is also of interest, as CSF tau levels are associated with Alzheimer’s disease pathology [27]. Because we did not see any group differences in arterial transit delay times (see ESM 5), we speculate that the altered perfusion primarily reflects changes in microcirculation rather than obstruction of large vessels. We also observe that lean control individuals who carry APOE-ε4 had higher perfusion than those without this variant (ESM 7). This finding is consistent with observations from previous studies [28].

This study also reports lower cerebral glucose uptake among participants with diabetes compared with control participants across all brain regions investigated, except for cerebellum. In addition to being a robust finding in Alzheimer’s disease [29], impaired [18F]FDG uptake in the fasted state has been reported for individuals with obesity, impaired glucose tolerance and type 2 diabetes with cognitive impairment [30,31,32]. That cerebellum is less affected than other brain regions aligns with this structure having been identified as a ‘pathologically preserved’ region in several neurodegenerative disorders (including Alzheimer’s disease) and often serves as a reference region for imaging of amyloid, tau and translocator protein (TSPO) [33,34,35].

A lower rate of cerebral glucose metabolism in participants with diabetes raises the question of to what extent systemic insulin resistance affects cerebral glucose uptake. In the current study, insulin resistance, as measured with HOMA-IR, correlated negatively with MRGlu. Although cerebral insulin signalling is associated with many brain functions, it is not currently considered to be a significant regulator of cerebral glucose metabolism [36], yet its role in Alzheimer’s disease pathology is receiving increased interest [37]. Lastly, it has been reported that individuals with impaired glucose tolerance and obesity show an elevated cerebral glucose uptake during hyperinsulinaemia [30, 31].

We demonstrate that among non-carriers of APOE-ε4, participants with type 2 diabetes had elevated influx rate of the fatty acid PET tracer in several brain regions. This finding aligns with previous studies in obesity and the metabolic syndrome [12, 13]. There are several metabolic pathways for fatty acids in the brain, including biosynthesis pathways (e.g. formation of phospholipids, sphingolipids and triacylglycerols), and fatty acid oxidation for ATP production [38]. Importantly, β-oxidation of fatty acids comes with a higher oxygen demand than oxidation of glucose, and mitochondrial β-oxidation of fatty acids is associated with generation of reactive oxygen species, oxidative stress and mitochondrial dysfunction [39, 40]. It could be speculated that the increased uptake of fatty acids is a compensatory mechanism to account for diminished ATP production following decreased glucose uptake and may eventually result in harmful effects to brain cells.

Among the lean control participants, APOE-ε4 carriers had a higher [18F]FTHA influx rate than non-carriers. This aligns with previous reports showing increased brain uptake of docosahexaenoic acid [41], and increased formation of lipid droplets in astrocytes [42] in APOE-ε4 carriers. Carrying the APOE-ε4 allele has been identified as the single most important genetic risk factor for developing late-onset Alzheimer’s disease [43] and corroborates the idea of altered lipid metabolism in the brain as part of the Alzheimer’s disease pathogenesis. In our data, lean physically active control participants who carried the APOE-ε4 allele displayed similar fatty acid influx rates as overweight participants and participants with diabetes without the allele. This finding indicates that carrying the ε4 isoform and having type 2 diabetes could be two independent risk factors that both contribute to increased cerebral fatty acid uptake, which in turn may provoke negative effects on brain metabolism and contribute to neurodegeneration. Carrying the ε4 isoform has also been observed as a risk factor for neurodegenerative disorders beyond Alzheimer’s disease [44].

An important consideration in this study is the potential impact of perfusion changes on PET tracer delivery to the brain. The lower MRGlu in participants with type 2 diabetes compared with both overweight and lean control individuals is unlikely to be only due to perfusion changes, as type 2 diabetes and overweight groups had similar perfusion. Higher [18F]FTHA in white matter in type 2 diabetes or type 2 diabetes APOE-ε4 non-carriers was observed despite reduced perfusion in participants with type 2 diabetes. Moreover, there was no correlation between [18F]FTHA Ki values and perfusion (r=0.06, p=0.72 in white matter, ESM 8). Therefore, it is likely that the increased influx of [18F]FTHA is mainly explained at the cellular level. However, the higher [18F]FTHA influx in lean APOE-ε4 carriers might be perfusion-related, as they also showed increased perfusion. We acknowledge that the full impact of perfusion on PET tracer uptake is not completely assessed in our analyses.

Our study used lean, physically active participants as a control group, contrasting with sedentary lifestyles in the other groups. This choice aimed to represent an ‘optimal’ metabolic state that could be targeted in potential intervention trials, and to maximise the difference in insulin resistance between groups, emphasising its potential connection to brain metabolism. However, this design introduces a limitation: by including physical activity as an additional variable, it does not allow for isolating obesity effects independent of activity levels. This constraint should be considered when interpreting the results.

Further, the potential influence of glucose-lowering medications on our results warrants consideration. While SGLT2 inhibitors, insulin and GLP1 receptor antagonists were excluded, participants with type 2 diabetes were treated with DPP4 inhibitors, statins and metformin. Few participants (2/10) were on DPP4 inhibitors, while a majority were on metformin (9/10) and therefore we cannot exclude the possibility that metformin impacted the results. However, previous studies suggest that the effect of metformin on [18F]FDG uptake is small and localised [45, 46]. With regards to statins, a subgroup analysis (not shown) revealed no significant differences in [18F]FDG uptake, [18F]FTHA uptake or perfusion between statin users (5/10 participants) and non-users. Therefore, we believe that the included medications have not substantially impacted the interpretation of our data, though this limitation should be noted.

For [18F]FTHA, we only report the rate constant, Ki, which represents the net influx rate of the tracer into brain tissue [23]. We interpret this parameter to be indicative of cerebral fatty acid uptake, although the outcome measure technically only reflects the influx of the tracer itself. Some studies have reported ‘fatty acid uptake’ [12, 13] analogous to MRGlu for [18F]FDG. Fatty acid uptake is calculated by multiplying tracer influx rate by plasma substrate concentration and correcting for transport and phosphorylation rate differences. This conversion is relevant for [18F]FDG due to glucose-tracer competition at blood–brain barrier transporters. However, such competition between [18F]FTHA and circulating NEFA has not been demonstrated to be rate-limiting for brain fatty acid uptake. In our study, circulating NEFA measurements were unreliable due to unexpected variance between pre- and post-PET scans, attributed to methodological issues rather than true NEFA variability. Consequently, we omitted fatty acid uptake estimation. Notably, obese individuals and those with type 2 diabetes typically have, if anything, elevated serum NEFA [47]. Therefore, we can conclude the following: (1) that if fatty acid–[18F]FTHA competition is rate-limiting for brain fatty acid exposure, the elevated Ki values among participants with type 2 diabetes may be underestimated; and (2) that with higher Ki values and serum NEFA, the difference in cerebral fatty acid uptake between control participants and participants with type 2 diabetes is likely larger than reported here.

We acknowledge that the current study includes a small and slightly unbalanced sample size, where participants with diabetes were marginally older and had a different gender distribution compared with other groups. This is particularly important to consider when interpreting the [18F]FTHA results, as, in contrast to both perfusion and glucose uptake, this is the first study to report an APOE-ε4-dependent increase in fatty acid uptake. Due to the small sample size, correlational analyses were performed on the entire cohort, possibly resulting in inflated associations, Additionally, the sample size prevented a meaningful gender analysis, limiting the generalisability of our findings across genders. Finally, because correction for multiple comparisons has not been performed, the findings should be considered exploratory and need to be replicated in larger trials.

In summary, the results from this study show that lower cerebral perfusion, reduced brain glucose and increased fatty acid uptake occur in individuals with type 2 diabetes. They further demonstrate that APOE-ε4 carriers, who have increased risk for developing late-onset Alzheimer’s disease, also have increased fatty acid uptake in the brain. We propose that these metabolic changes are key to understanding the mechanisms that link together type 2 diabetes and neurodegenerative disorders.