Growth Hormone-Releasing Hormone Modulation of Neuronal Exosome Biomarkers in Mild Cognitive Impairment

Abstract

Age-related changes in cognition are linked to decreased expression of somatotropins, GHRH and IGF-1. Mild cognitive impairment (MCI) and Alzheimer's disease (AD) are heterogeneous conditions. The loss of GHRH signaling in the brain may be mechanistically involved in AD pathogenesis. The consequent need to identify AD at an early and perhaps more treatable stage has fueled research into blood-based, exosome biomarkers. Plasma exosomes from participants enrolled in a randomized, double-blind, placebo-controlled 20-week trial of GHRH administration, were isolated, precipitated, and enriched by immuno-absorption with anti-L1CAM antibody (neural adhesion protein) from adults with MCI and age-matched, cognitively normal controls (CNC). Extracted protein cargo from neuronally-derived exosomes (NDEs) were assessed by ELISAs for protein levels implicated in AD neuropathology and for synaptic proteins altered by AD. Plasma NDE concentrations of Aβ1-42 were significantly increased while plasma NDE concentrations of NRGN, synaptophysin, synaptotagmin, and synaptopodin were significantly decreased in patients with MCI, independent of GHRH treatment. Plasma NDE concentrations of ptau-S396 and GAP43 were not affected by cognitive status (CNC versus MCI) or by GHRH treatment. Aβ1-42, neurogranin (NRGN), synaptophysin, synaptotagmin, and synaptopodin demonstrated the highest diagnostic accuracy for distinguishing between CNC and MCI patients, while synaptophysin and synaptotagmin demonstrated moderate accuracy in distinguishing between placebo-treated and GHRH-treated, MCI patients.

Keywords: Amyloid; GHRH; biomarker; growth hormone; mild cognitive impairment; neuronal exosomes; synapse; tau.

Fig. 1.

Plasma NDE concentrations for Aβ_1-42, NRGN, synaptophysin, synaptotagmin, and synaptopodin accurately identify MCI patients based on protein cargo content. Plasma NDE concentrations for AD-related and synaptic-related proteins were quantified using ELISA. Plasma NDE concentrations for CD81 were not statistically different between CNC and MCI patients (A). Plasma NDE concentrations for Aβ_1-42 (B) were significantly higher while plasma NDE concentrations for NRGN (D), synaptophysin (E), synaptotagmin (F), and synaptopodin (G), were significantly lower in MCI patients as compared to CNC patients. We found no significant difference in plasma NDE concentrations for ptau-S396 (C) and GAP43 (H) between CNC and MCI patients. Unpaired t-test: **p < 0.01 versus CNC, ***p < 0.001 versus CNC, ***p < 0.0001 versus CNC.

Fig. 2.

Plasma NDE concentrations of Aβ_1-42, NRGN, synaptophysin, synaptotagmin, and synaptopodin are not modulated with GHRH treatment while plasma NDE concentrations of p-tauS396 and GAP43 are not modulated by cognitive status or GHRH treatment. Plasma NDE concentrations for Aβ_1-42 (A) were increased while plasma NDE concentrations for NRGN (B), synaptophysin (C), synaptotagmin (D), and synaptopodin (E) were significantly reduced in MCI patients as compared to CNC patients, independent of GHRH treatment. Plasma NDE concentrations for ptau-S396 (F) and GAP43 (G) were not significantly different between both patient groups (CNC versus MCI), independent of GHRH treatment. There was no significant difference in plasma NDE concentrations for all biomarkers (A-G) in placebo-treated, MCI patients as compared to GHRH-treated, MCI patients. 1-way ANOVA with Tukey’s multiple comparisons post-hoc test; *p < 0.05 versus placebo-treated, CNC, **p < 0.01 versus GHRH-treated, CNC.

Fig. 3.

Plasma NDE concentrations of Aβ_1-42, NRGN, synaptophysin, synaptotagmin, and synaptopodin demonstrated the highest diagnostic accuracy for distinguishing between MCI and CNC patients as measured by ROC analysis. Representative ROC curves for plasma NDE concentrations for Aβ_1-42, NRGN, synaptophysin, synaptotagmin, and synaptopodin for placebo-treated, MCI patients as compared to placebo-treated, CNC patients (A) and GHRH-treated, MCI patients as compared to GHRH-treated, CNC patients (B). Aβ_1-42, NRGN, synaptophysin, synaptotagmin, and synaptopodin demonstrate the highest diagnostic accuracy for distinguishing CNC and MCI patients, independent of GHRH treatment. Plasma NDE concentrations of Aβ_1-42, ptau-S396, NRGN, synaptopodin, and GAP43 (C) demonstrated the lowest diagnostic accuracy for distinguishing between placebo-treated and GHRH-treated, MCI patients. Plasma NDE concentrations for a synaptophysin and synaptotagmin demonstrated moderate diagnostic accuracy for distinguishing between placebo-treated and GHRH-treated, MCI patients (D).