Fluorescence-tagged salivary small extracellular vesicles as a nanotool in early diagnosis of Parkinson's disease

Simran Rastogi; Komal Rani; Sanskriti Rai; Rishabh Singh; Prahalad Singh Bharti; Vaibhav Sharma; Jyoti Sahu; Vrinda Kapoor; Poorvi Vishwakarma; Sumit Garg; Shivajirao Lahu Gholap; Krishna Kishore Inampudi; Gyan Prakash Modi; Neerja Rani; Madhavi Tripathi; Achal Srivastava; Roopa Rajan; Fredrik Nikolajeff; Saroj Kumar

doi:10.1186/s12916-023-03031-1

Fluorescence-tagged salivary small extracellular vesicles as a nanotool in early diagnosis of Parkinson's disease

BMC Med. 2023 Sep 4;21(1):335. doi: 10.1186/s12916-023-03031-1.

Authors

Simran Rastogi^#¹, Komal Rani^#^{1

2}, Sanskriti Rai¹, Rishabh Singh¹, Prahalad Singh Bharti¹, Vaibhav Sharma³, Jyoti Sahu¹, Vrinda Kapoor⁴, Poorvi Vishwakarma¹, Sumit Garg⁵, Shivajirao Lahu Gholap⁶, Krishna Kishore Inampudi¹, Gyan Prakash Modi⁷, Neerja Rani⁸, Madhavi Tripathi⁵, Achal Srivastava⁹, Roopa Rajan⁹, Fredrik Nikolajeff³, Saroj Kumar^{10

11}

Affiliations

¹ Department of Biophysics, All India Institute of Medical Sciences, New Delhi, 110029, India.
² Department of Pathology & Laboratory Medicine, All India Institute of Medical Sciences Bibinagar, Hyderabad, 508126, India.
³ Department of Health, Education, and Technology, Luleå University of Technology, 97187, Luleå, Sweden.
⁴ School of Interdisciplinary Research, Indian Institute of Technology Delhi, New Delhi, 110016, India.
⁵ Department of Nuclear Medicine, All India Institute of Medical Sciences, New Delhi, 110029, India.
⁶ Department of Chemistry, Indian Institute of Technology Delhi, New Delhi, 110016, India.
⁷ Department of Pharmaceutical Engineering & Technology, Indian Institute of Technology BHU, Varanasi, 221005, India.
⁸ Department of Anatomy, All India Institute of Medical Sciences, New Delhi, 110029, India.
⁹ Department of Neurology, All India Institute of Medical Sciences, New Delhi, 110029, India.
¹⁰ Department of Biophysics, All India Institute of Medical Sciences, New Delhi, 110029, India. saroj.kumar@ltu.se.
¹¹ Department of Health, Education, and Technology, Luleå University of Technology, 97187, Luleå, Sweden. saroj.kumar@ltu.se.

^# Contributed equally.

Abstract

Background: Parkinson's disease is generally asymptomatic at earlier stages. At an early stage, there is an extensive progression in the neuropathological hallmarks, although, at this stage, diagnosis is not possible with currently available diagnostic methods. Therefore, the pressing need is for susceptibility risk biomarkers that can aid in better diagnosis and therapeutics as well can objectively serve to measure the endpoint of disease progression. The role of small extracellular vesicles (sEV) in the progression of neurodegenerative diseases could be potent in playing a revolutionary role in biomarker discovery.

Methods: In our study, the salivary sEV were efficiently isolated by chemical precipitation combined with ultrafiltration from subjects (PD = 70, healthy controls = 26, and prodromal PD = 08), followed by antibody-based validation with CD63, CD9, GAPDH, Flotillin-1, and L1CAM. Morphological characterization of the isolated sEV through transmission electron microscopy. The quantification of sEV was achieved by fluorescence (lipid-binding dye-labeled) nanoparticle tracking analysis and antibody-based (CD63 Alexa fluor 488 tagged sEV) nanoparticle tracking analysis. The total alpha-synuclein (α-syn_Total) in salivary sEVs cargo was quantified by ELISA. The disease severity staging confirmation for n = 18 clinically diagnosed Parkinson's disease patients was done by ^99mTc-TRODAT-single-photon emission computed tomography.

Results: We observed a significant increase in total sEVs concentration in PD patients than in the healthy control (HC), where fluorescence lipid-binding dye-tagged sEV were observed to be higher in PD (p = 0.0001) than in the HC using NTA with a sensitivity of 94.34%. In the prodromal PD cases, the fluorescence lipid-binding dye-tagged sEV concentration was found to be higher (p = 0.008) than in HC. This result was validated through anti-CD63 tagged sEV (p = 0.0006) with similar sensitivity of 94.12%. We further validated our findings with the ELISA based on α-syn_Total concentration in sEV, where it was observed to be higher in PD (p = 0.004) with a sensitivity of 88.24%. The caudate binding ratios in ^99mTc-TRODAT-SPECT represent a positive correlation with sEV concentration (r = 0.8117 with p = 0.0112).

Conclusions: In this study, for the first time, we have found that the fluorescence-tagged sEV has the potential to screen the progression of disease with clinically acceptable sensitivity and can be a potent early detection method for PD.

Keywords: Alpha-synuclein; Nanoparticle tracking analysis; Parkinson’s disease; Prodromal; Saliva; Small extracellular vesicles; TRODAT scan.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Antibodies
Early Diagnosis
Extracellular Vesicles*
Fluorescence
Humans
Lipids
Parkinson Disease* / diagnostic imaging

Substances

Antibodies
Lipids