Blood tau-PT217 contributes to the anesthesia/surgery-induced delirium-like behavior in aged mice

Abstract

Introduction: Blood phosphorylated tau at threonine 217 (tau-PT217) is a newly established biomarker for Alzheimer's disease and postoperative delirium in patients. However, the mechanisms and consequences of acute changes in blood tau-PT217 remain largely unknown.

Methods: We investigated the effects of anesthesia/surgery on blood tau-PT217 in aged mice, and evaluated the associated changes in B cell populations, neuronal excitability in anterior cingulate cortex, and delirium-like behavior using positron emission tomography imaging, nanoneedle technology, flow cytometry, electrophysiology, and behavioral tests.

Results: Anesthesia/surgery induced acute increases in blood tau-PT217 via enhanced generation in the lungs and release from B cells. Tau-PT217 might cross the blood-brain barrier, increasing neuronal excitability and inducing delirium-like behavior. B cell transfer and WS635, a mitochondrial function enhancer, mitigated the anesthesia/surgery-induced changes.

Discussion: Acute increases in blood tau-PT217 may contribute to brain dysfunction and postoperative delirium. Targeting B cells or mitochondrial function may have therapeutic potential for preventing or treating these conditions.

Keywords: anesthesia; delirium; phosphorylated tau at threonine 217; surgery; tau; tau phosphorylation.

Fig. 1.. PET study of A/S-induced increases in Tau-PT217 amounts in lungs, blood, and brain of mice.

a. Representative PET images of [¹⁸F]Flortaucipir focus on the whole body of the mouse for up to 120 minutes. Time-activity curves of [¹⁸F]Flortaucipir in the lungs (b and c) and blood (d and e) show significant differences in the distribution of [¹⁸F]Flortaucipir in lungs and blood between the time before A/S (baseline) and the time after A/S in the mice. f. Representative PET images of [18F]Flortaucipir focus on the mouse brain tissues (0-120 minutes; axial, sagittal, and coronal). Time-activity curves of [¹⁸F]Flortaucipir in the whole brain (g and h) between baseline and A/S in the mice. Standardized uptake value ratio (SUVR) of [¹⁸F]Flortaucipir in the cortex (i and j) and hippocampus (k and l), showing a significant difference of [¹⁸F]Flortaucipir brain uptake between baseline and A/S in the mice. Cerebellar gray matter was used as a reference region to yield SUVR for [¹⁸F]Flortaucipir. N = 4 mice in each group. Student’s t-test was used to analyze the data presented in c, e, h, j, and i. The P values refer to the difference of [¹⁸F]Flortaucipir uptake between the baseline and the A/S in the mice. Error bar indicates standard deviation. PET, positron emission tomography; A/S, anesthesia/surgery; Tau-PT217, Tau phosphorylated at threonine 217; TACs, time activity curves; AUC, area under each TAC; SUV, standardized uptake value; SUVR, standardized uptake value ratio.

Fig. 2.. A/S increased Tau-PT217 amounts in the lungs, blood, and brain of mice.

a. Classical immunoassay monitors absorbance signal due to the increased amount of chromogen as the analyte concentration increases. b. Nanoneedle technology detecting spectrum shift from individual nanoneedles originated from additional mass deposition on each nanoneedle, forming an antibody-antigen sandwich complex. c. Low concentrations of plasma Tau-PT217 were detected with nanoneedle technology. Significant increases in Tau-PT217 amount were observed 3 and 6 h after the A/S compared to the control condition in mice. Quantitative western blot showing A/S increased Tau-PT217 amount in the cortex (d and e) and hippocampus (f and g) compared to control condition in the mice. Comparisons of A/S-induced increases in Tau-PT217 amounts between lungs and brain tissues (cortex) 30 minutes following A/S (h, i and j), demonstrating that the A/S caused more increases of Tau-PT217 and lesser decreases of total Tau in lungs compared to the cortex of mice. Comparisons of A/S-induced increases in Tau-PT217 amounts between lungs and brain tissues (cortex) 3 hours following A/S (k, l, and m), demonstrating that the A/S caused more increases of Tau-PT217 in the cortex compared to lungs of mice. N = 4 mice in each group, as demonstrated in each panel of the figure. One-way ANOVA and the post-hoc analysis with Bonferroni were used to analyze the data presented in c. Student’s t-test was used to analyze the data presented in e and g. Two-way ANOVA and the post-hoc analysis with Bonferroni were used to analyze the data presented in i, j. l and m. The P values refer to the differences of Tau-PT217 between the control condition and A/S in the mice. Error bar indicates standard deviation. A/S, anesthesia/surgery; Tau-PT217, Tau phosphorylated at threonine 217.

Fig. 3.. A/S decreased the percentage of blood B cells in aged mice.

Flow cytometry showed that A/S decreased the percentage of blood B cells (live CD45+ CD3− LY6G− CD11b− CD19+) compared to the control condition (a and b). Flow cytometry also showed that A/S decreased mitochondrial membrane potential (represented by TRAM amounts) of B cells in the blood of mice (c and d). e. Confocal microscope images showed that B cells (upper panel), but not T cells (middle panel), could bind to the 14 amino acid peptide containing Tau-PT217, but not the 14 amino acid peptide without containing Tau-PT217 (bottom panel). N = 5 - 7 mice in each group, as demonstrated in each panel of the figure. The Student's t-test was used to analyze the data presented in b and d. The P values refer to the differences in the numbers of B cells or mitochondrial membrane potential between the control condition and A/S in the mice. Error bar indicates standard deviation. Tau-PT217, Tau phosphorylated at threonine 217; A/S, anesthesia, and surgery; TMRM, tetramethylrhodamine methyl ester; DAPI, 4',6-diamidino-2-phenylindole.

Fig. 4.. Treatment with B cells mitigated the A/S-induced delirium-like behaviors in mice.

a. Experimental scheme: B cells (6 x 10⁶) were negatively sorted from donor mice (4 months old female mice) using EasySep^™ and injected (2 x 10⁶) into recipient-aged mice, followed by control condition and A/S, and behavioral tests. b. Representative flow cytograms of B cells in the blood of recipient mice. c. B cell amounts in the mice with control condition plus vehicle treatment and the mice with A/S plus B cell treatment. d. B cell treatment attenuated the A/S-induced increases in blood Tau-PT217 amounts in the aged mice. Quantitative western blot showing that B cell treatment attenuated the A/S-induced increases in cortex Tau-PT217 in the mice (e and f). Treatment with B cells mitigated the A/S-induced delirium-like behaviors in mice demonstrated in individual tests (g) and composite Z score (h). N = 10 - 12 mice in each group of behavioral test, and N = 4 - 6 mice in each flow cytometry, nanoneedle, and western blot study as demonstrated in each panel of the figure. The Student's t-test was used to analyze the data presented in c. Two-way ANOVA and the post-hoc analysis with Bonferroni were used to analyze the data presented in d, f, and h. The P values refer to the difference in B cell amounts, Tau-PT217 amounts, and Z score between the control condition and A/S in the mice. Error bar indicates standard deviation. Tau-PT217, Tau phosphorylated at threonine 217; A/S, anesthesia and surgery.

Fig. 5.. WS635 mitigated the A/S-induced changes in B cells, Tau-PT217 amounts, and delirium-like behaviors in mice.

Flow cytometry showed that WS635 attenuated the A/S-induced reductions in blood B cells (a and b) and mitochondrial membrane potential (represented by TRAM) (c and d). Nanoneedle analysis showed that WS635 attenuated the A/S-induced increases in blood Tau-PT217 amounts in the mice (e). Quantitative western blot showing that WS635 attenuated the A/S-induced increases in cortex Tau-PT217 amounts in the mice (f and g). WS635 rescued the A/S-induced delirium-like behaviors in mice, demonstrated in individual test (h) and composite Z score (i) in the mice. N = 10 - 12 mice in each group of behavioral test, and N = 4 - 6 mice in each group in flow cytometry, nanoneedle, and western blot study as demonstrated in each panel of the figure. Two-way ANOVA and the post-hoc analysis with Bonferroni were used to analyze the data presented in b, d, e, g, and i. The P values refer to the differences in B cells, Tau-PT217, and Z scores between the control condition and A/S in the mice. Error bar indicates standard deviation. Tau-PT217, Tau phosphorylated at threonine 217; A/S, anesthesia, and surgery; TMRM, tetramethylrhodamine methyl ester.

Fig. 6.. WS635 mitigated the A/S-induced ACC neuronal excitability of the aged mice.

The whole cell patch-clamp recording sample trace of neurons action potential firing rate with 150 pA of current injection in ACC slice harvested from the aged mice following the control condition (a) and the aged mice following the A/S (b). c. Action potential firing rates of ACC neurons with different amounts of current injection were higher in mice that underwent the A/S compared to control condition (mixed two-way ANOVA, F = 9.23, p < 0.01, control: n = 16, 3 mice, A/S: n = 16, 4 mice). The whole cell patch-clamp recording sample trace of neurons action potential firing rate with 150 pA of current injection in ACC slice harvested from the aged mice following the control condition (d) and the aged mice following the A/S (e) pretreated with the vehicle of WS635. f. The action potential firing rates of ACC neurons still increased after the A/S in the ACC slice of the aged mice pretreated with vehicle (mixed two-way ANOVA, F = 16.9, p < 0.01, control + vehicle: n = 40, 5 mice, A/S + vehicle: n = 37, 5 mice). The whole cell patch-clamp recording sample trace of neurons action potential firing rate with 150 pA of current injection in ACC slice harvested from the aged mice following the control condition (g) and the aged mice following the A/S (h) pretreated with WS635. i. The action potential firing rates of ACC neurons did not increase after the A/S compared to control condition in the aged mice pretreated with WS635 (mixed two-way ANOVA, F = 0.72, p = 0.32, control + WS635: n = 19, 3 mice, A/S + WS635: n = 21, 3 mice). Tau-PT217, Tau phosphorylated at threonine 217; A/S, anesthesia, and surgery; ACC, anterior cingulate cortex.

Fig. 7.. The hypothesized pathway of the acutely increased blood Tau-PT217 and postoperative delirium-like behavior in aged mice.

The anesthesia/surgery increased the generation of Tau-PT217 potentially in the lungs of aged mice, leading to increased blood Tau-PT217 amounts in the mice. The mitochondrial function in B cells was impaired after the A/S, leading to fewer B cells in the blood. Moreover, B cells bind to Tau-PT217. Thus, the reductions in blood B cells result in the release of Tau-PT217, leading to more Tau-PT217 in the blood of mice. The acutely increased blood Tau-PT217 may then enter the brain, leading to increased Tau-217 amounts in the cortex and hippocampus, finally leading to an increased action potential firing rate in ACC of aged mice and postoperative delirium-like behavior in the mice. Tau-PT217, Tau phosphorylated at threonine 217; A/S, anesthesia, and surgery; ACC, anterior cingulate cortex.