Extracellular vesicles (EVs) such as exosomes and microvesicles mediate intercellular communication and regulate a diverse range of crucial biological processes. Host cells that are damaged, infected or transformed release biomarker-containing EVs into the peripheral circulation, where they can be readily accessed for use in diagnostic or prognostic testing. However, current methods of EV isolation from blood plasma are complex and often require relatively large sample volumes, hence are inefficient for widespread use in clinical settings. In work led by collaborator Newman Sze, we report a novel and inexpensive method of rapidly isolating EVs from small volumes of human blood plasma by PRotein Organic Solvent PRecipitation (PROSPR). PROSPR encompasses a rapid three-step protocol to remove soluble proteins from plasma via precipitation in cold acetone, leaving the lipid-encapsulated EVs behind in suspension. This generates higher purity EVs that can then be obtained from filtration or classical ultracentrifugation methods. We foresee that PROSPR-based purification of EVs will significantly accelerate the discovery of new disease biomarkers and the characterization of EVs with potential for clinical applications.

Reference

Gallart-Palau X, Serra A, Wong AS, Sandin S, Lai MK, Chen CP, Kon OL, Sze SK (This paper).

Uncovering novel biomarkers which lead to better diagnosis, prognosis and treatment is undoubtedly an important endeavor, but sometimes, finding what is not a biomarker may be worthwhile as well. Gangliosides are complex ceramide-glycosphingolipids known to play important roles in neuronal development. The GM1 ganglioside binds to β-amyloid and prevents β-sheet conformational changes (Yanagisawa et al., 1995), while GQ1bα and GT1aα are cholinergic neuron-specific gangliosides found to be increased in murine AD models (Ariga et al. 2010; Ariga et al. 2013a). Antibodies to gangliosides have been detected in AD, but the differences compared to controls were too low to be useful for diagnosis (Chapman et al. 1988). Other studies suffered from small patient numbers (n = 4, Hatzinfilippou et al. 2008), or low cut-off values resulting in low specificity (Ariga et al. 2013b).

In this study led by Prof. Nobuhiro Yuki, we measured serum levels of anti-ganglioside antibodies in patients with clinically diagnosed AD (n = 22) and vascular dementia (n = 14) and compared them to sera of patients with Guillain-Barré syndrome (GBS) and multifocal motor neuropathy (MMN), diseases where the antibodies are thought to be pathogenic; as well as to normal controls. We found that titers of IgG and IgM anti-GM1, anti-GQ1bα, and anti-GT1aα antibodies did not differ among AD, vascular dementia, and normal controls, and were remarkably lower than those in GBS and MMN. Our results suggest that the anti-ganglioside antibodies are unlikely to be pathogenic in AD or useful as biomarkers.

References

Miura Y, Miyaji K, Chai YL, Chen CL, Lai MK, Yuki N (This paper).

Ariga T, Yanagisawa M, Wakade C, Ando S, Buccafusco JJ, McDonald MP, Yu RK (2010) Ganglioside metabolism in a transgenic mouse model of Alzheimer’s disease: Expression of Chol-1α antigens in the brain. ASN Neuro 2: e00044.

Ariga T, ItokazuY, McDonald MP, Hirabayashi Y, Ando S, Yu RK (2013a) Brain gangliosides of a transgenic mouse model of Alzheimer’s disease with deficiency in GD3-synthase: Expression of elevated levels of a cholinergic-specific ganglioside, GT1aα. ASN Neuro 5: 141-148.

Ariga T, Kubota M, Nakane M, Oguro K, Yu RK, Ando S (2013b) Anti-Chol-1 antigen, GQ1bα, antibodies are associated with Alzheimer’s disease. PLoS One 8: e63326.

Chapman J, Sela BA, Wertman E, Michaelson DM (1988) Antibodies to ganglioside GM1 in patients with Alzheimer’s disease. Neurosci Lett 86: 235-240.

Hatzifilippou E, Koutsouraki E, Banaki T, Traka M, Costa VG, Baloyannis SJ (2008) Antibodies against GM1 in demented patients. Am J Alzheimers Dis Other Demen 23: 274-279.

Yanagisawa K, Odaka A, Suzuki N, Ihara Y (1995) GM1 ganglioside-bound amyloid beta-protein (Aβ): A possible form of preamyloid in Alzheimer’s disease. Nat Med 1: 1062-1066.

 Despite its importance as the leading cause of vascular dementia in Asia (Chen 2004), the primary pathogenic mechanisms in subcortical ischemic vascular dementia (SIVD) have remained elusive. Because of the lack of approved therapeutic agents for SIVD, there is a pressing need to identify novel therapeutic targets. In work led by collaborators Guanghou Shui and Markus Wenk, we performed comparative lipidomic analyses of SIVD and mixed dementia (i.e., SIVD and Alzheimer’s disease, MixD) which may confer new insights pertaining to the possible interaction between neurodegenerative and vascular mechanisms in the pathogenesis of dementia. Liquid chromatography coupled to mass spectrometry was used to comprehensively analyze the lipidomes of white and gray matter from the temporal cortex of nondemented controls, SIVD, and MixD subjects. Detailed molecular profiles highlighted the pathologic relevance of gray matter sphingolipid fatty acyl chain heterogeneity in dementia. In addition, the levels of sulfatides and lysobisphosphatidic acids were progressively increased in the temporal cortex gray matter from control to SIVD to MixD. White matter phospholipid profiles indicated possible adaptive mechanisms (i.e., increased unsaturation) to chronic ischemia in SIVD and elevated membrane degradation in MixD.

References

Chen CP (2004) Transcultural expression of subcortical vascular disease. J Neurol Sci 226:45-47.

Lam SM, Wang Y, Duan X, Wenk MR, Kalaria RN, Chen CP, Lai MK, Shui G (This paper).

In research led by collaborator Newman Sze, we used an iTRAQ-2D-LC-MS/MS strategy for quantitative analysis of pooled lysates from the neocortex of 21 pathologically confirmed cases of vascular dementia (VaD) and matched non-neurological controls. A total of 144 differentially perturbed proteins out of 2284 confidently identified proteins (false discovery rate=0.3%) were shortlisted for bioinformatics analysis. Western blot analysis of selected proteins using samples from individual patients (n=10 per group) showed significant increases in the abundance of SOD1 and NCAM and reduced ATP5A in VaD. This suggests a state of hypometabolism and vascular insufficiency along with an inflammatory condition during VaD. Elevation of SOD1 and increasing trend for iron-storage proteins (FTL, FTH1) may be indicative of an oxidative imbalance that is accompanied by an aberrant iron metabolism. The synaptic proteins did not exhibit a generalized decrease in abundance (e.g. syntaxin) in the VaD subjects. This reported proteome offers a reference data set for future basic or translational studies on VaD.

Reference

Datta A, Qian J, Chong R, Kalaria RN, Francis P, Lai MK, Chen CP, Sze SK. (This paper).