2.1. α-Synuclein (α-syn) and its variants
α-syn is detectable in both CSF and plasma and is the most widely researched biomarker of PD (Tsao et al., 2022; Estaun-Panzano et al. 2023; Tofaris 2022). Phosphorylation of the Ser129 site results in phosphorylated α-syn (PS-129), while pro-aggregating forms of α-syn, such as oligomeric α-syn (o-α-syn), are also found in CSF and blood (Ma et al. 2023; Constantinides et al. 2021; Zubelzu et al. 2022; Chen et al. 2022; Chen et al. 2022). Meanwhile, the pathogenic β-sheet seed is the pathological conformation of α-syn and can be detected in serum (Okuzumi et al. 2023).
Several studies and meta-analyses have confirmed that when compared to the control group, the total α-syn (t-α-syn) levels in the CSF are consistently lower in PD, MSA, PSP, CBS, and VP groups, with no significant differences among them (Zubelzu et al. 2022; Constantinides et al. 2017; Førland et al. 2020; Koníčková et al. 2023; Aerts et al. 2012). Therefore, t-α-syn levels cannot differentiate between PD and APS.
Notably, levels of o-α-syn in PD and other Parkinsonian syndromes reportedly do not differ significantly but are elevated compared to that of the control group (Eusebi et al. 2017).
Utilizing a Bead-based Luminex assay (with a sensitivity of 9 pg/mL), researchers measured the concentration of pS129 in the CSF of patients with PD, MSA, and PSP, revealing differences among them. To differentiate between the different diseases, receiver operating characteristic analysis following the discovery phase indicated that pS-129/t-α-syn was superior to pS-129 alone, with a specificity of ≥ 80%. The sensitivity among the three different Parkinsonian disease groups were as follows: PD vs MSA, 40%; PD vs PSP, 72%; and MSA vs. PSP, 63% (Wang et al. 2012).
Aggregates of α-syn, including propagative α-syn seeds, showed high diagnostic performance in differentiating between PD and MSA (Siderowf et al. 2023; Painous et al. 2024; Parnetti et al. 2019; Goolla et al. 2023; Shahnawaz et al. 2020) Amplified seeds maintain disease-specific properties, allowing for the differentiation of samples from individuals with PD and MSA (Painous et al. 2024) Okuzumi et al. (2023) suggested that the rate of negative results of IP/RT-QuIC in patients with MSA was significantly higher than that in patients with PD. Additionally, their study also examined the distinctive morphological features of seeds in various diseases. The fibril morphology of products derived from IP/RT-QuIC of serum α-syn seeds in patients with synucleinopathies could differentiate PD, DLB, and MSA, allowing for further research in this area. A study found that the intensity of the signal in MSA was greater than that in PD when aggregation was performed in a specific buffered solution, indicating that α-syn seed aggregation from various diseases require different conditions for optimal detection (Martinez-Valbuena et al. 2022). These results suggest that our follow-up study can focus on the structural diversity and disease specificity of α-syn seeds.
2.2 DOPA decarboxylase (DDC)
A primary pathological feature of PD is the degeneration of dopaminergic neurons in the substantia nigra (Stoker and Greenland 2018). DDC is a diagnostic marker of dopaminergic dysfunction and can be detected in CS (Painous et al. 2024).
Several studies have attempted to reveal the differences in DDC between PD and APS (Paslawski et al. 2023; Pereira et al. 2023). CSF levels of DDC may potentially be useful in differentiating among degenerative Parkinsonisms (PD vs. APS) (Paslawski et al. 2023).
2.2 MK
MK is predominantly expressed during midgestation in embryogenesis, but its presence in normal adult brains is minimal. However, MK may recently play a role in various adult brain pathologies (Neumaier et al. 2023).
MK has demonstrated significant diagnostic potential as its levels were notably higher in patients with PD compared to those with APS (Paslawski et al. 2023).
2.3 Kallikrein 10
Kallikreins, which is a subgroup of serine proteases, play various physiological roles. Recent research has emphasized their involvement in carcinogenesis, highlighting several kallikreins as promising candidates for novel biomarkers in cancer and other diseases. This supports the potential utility of kallikreins in clinical diagnostics and therapeutic targeting (Wikipedia, nd).
Kallikrein 10 has exhibited specific changes in APS compared to PD and controls; unfortunately, these changes were not elaborated (Paslawski et al. 2023).
2.4 Classic Alzheimer’s disease (AD) biomarkers
Amyloid-beta-Aβ42, tau protein-τT, and phosphorylated tau protein-τP-181 are classical biomarkers of AD (Sung et al. 2023). Notably, their significance in Parkinson’s syndrome has been re-recognized.
When compared to patients with PD, τT/Aβ42 ratio was increased in patients with MSA (Constantinides et al. 2021; Constantinides et al. 2017). An elevated τT/Aβ42 ratio effectively differentiated MSA from PD, with an optimal cut-off value of 0.344 that yielded a sensitivity of 0.71 and specificity of 0.93 (Constantinides et al. 2017).
2.5 Exosomes
Exosomes from peripheral blood and CSF nerve cells have been used to distinguish PD and MSA (Taha and Bogoniewski 2024; Taha 2023; Yan et al. 2024).
Dutta et al.’s (2021) study confirmed that α-syn concentrations in exosomes were markedly lower in the control group and significantly higher in the MSA group compared to the PD group. They created a ratio using α-syn concentrations of putative oligodendroglial exosomes and putative neuronal exosomes with good sensitivity in distinguishing PD and MSA. By incorporating this ratio along with the α-syn and total exosome concentrations, a multinomial logistic model successfully distinguished PD from MSA, with an area under the curve (AUC) of 0.902, sensitivity of 89.8%, and specificity of 86.0% after application to an independent validation cohort.
Meloni et al. (2023) investigated neural-derived extracellular vesicles (NDEVs) isolated from the blood. Analysis of NDEVs revealed a significant increase in o-α-syn levels in PD compared to APS (CBD and PSP). Additionally, levels of Tau aggregates in NDEVs were significantly elevated in APS compared to PD (p < 0.0001). Receiver operating characteristic analysis showed that the concentration of NDEVs of both oligomeric o-α-syn and Tau aggregates exhibited an “excellent” power of classification that effectively distinguished PD from APS. For o-α-syn, the AUC was 0.817 (95% confidence interval (CI): 0.732–0.885; p < 0.0001), sensitivity of 78.6%, and specificity of 77.5%. For Tau aggregates, the AUC was 0.856 (95% CI: 0.776–0.915; p < 0.0001), sensitivity of 90.0%, and specificity of 75.7%.
Taha et al. (2023) was the first to measure pS129-α-syn levels in neuronal extracellular vesicles (nEVs) and oligodendroglial extracellular vesicles (oEVs). They reported that nEV pS129-α-syn concentrations were highest in healthy controls (HC) followed by PD and MSA, but the differences were not statistically significant. Conversely, oEV concentrations of pS129-α-syn were also highest in HC followed by PD and MSA and was significantly higher in both disease groups. Additionally, their study revealed that the oEV/nEV pS129-α-syn ratio increased in the order of HC < PD < MSA. Furthermore, they also measured total tau, pT181-tau (tau phosphorylated at Thr181) in nEVs and oEVs, and/or serum neurofilament light protein (NfL) levels. Due to the detection sensitivity, pT181-tau was detected in very few samples. Other results were similar to experiments involving plasma or CSF.
2.6 NfL
NfL in CSF (cNfL) and plasma (pNfL) is a marker for neuronal damage that may potentially be used to distinguish between clinically similar conditions, such as frontotemporal dementia from AD and PD from APS (Quadalti et al. 2021).
Among Parkinsonian syndromes, the mean cNfL levels were higher in MSA, PSP, and CBS when compared with PD (Wikipedia, nd; Bridel et al. 2019).