To the best of our knowledge, this is the largest in vivo neuropathological study in this field. With a multicenter design, we were able to recruit a large number of patients with PD for various durations and prevent selection bias in a single-center study. Individual matching of controls allowed a good comparison of diagnostic accuracy not only in the entire group but also in the subgroup analyses of the UGI and LGI tracts. The collection of full-depth surgical specimens, excluding biopsied tissue, ensured a large evaluation area and an unbiased comparison between patients and controls. We found that the duration from symptom onset to surgery was an independent predictor of AS accumulation in the GI tract in patients with PD, regardless of the rostrocaudal gradient, age of symptom onset, age at the time of surgery, or sex. This is the first in vivo pathological evidence of the temporally progressive nature of AS accumulation in the GI tract of patients with PD.
Possibility of using LP in the GI tract as a pathological biomarker in patients with PD
Poor sensitivity was a major limitation when using AS accumulation in the GI tract as a pathological biomarker of patients with PD. The sensitivity and specificity of AS accumulation in the GI tract in this study were 75.3% and 91.5%, respectively, which were better than the pooled sensitivity and specificity of 43% and 82%, respectively, reported in the previous meta-analysis. 19 Interestingly, the sensitivity of the mucosal/submucosal evaluation was 46.9% in this study, which was similar to the pooled sensitivity in the meta-analysis. 19 The interpretation and comparison of data from the previous meta-analysis are limited because of the methodological heterogeneity among the included studies. However, the fact that six studies, half of the 12 studies included in the meta-analysis, used biopsied tissue may have a significant impact on the pooled sensitivity. 19 Moreover, the evaluated area of the mucosal/submucosal layer was much larger than that of biopsy, and AS accumulation was mostly found in the submucosal plexuses and large nerve bundles, which are not obtainable in routine endoscopic biopsy procedure. Therefore, this study confirmed the fundamental limit of biopsy as a pathological biomarker of PD, which is consistent with our previous study. 14
This study showed a better diagnostic accuracy than our previous study, which used the same staining methods. 14 Higher sensitivity and specificity would be explained by the use of a more conservative definition of pAS positivity (exclusion of a “diffuse” pattern, which was used in the previous study), 30, 31 avoidance of a selection bias by a large number of participants (97 vs. 33 in the PD patient group), and exclusion of patients with young-onset PD of age at onset ≤ 50, whereas four patients with young-onset PD (12.1%) were included of the previous study. 14 In another study on the submandibular gland, we reported that AS accumulation was present in 56.2% of patients with PD35 but not in three patients with PRKN gene mutation. 36 Therefore, a more stringent selection and evaluation of patients with sporadic PD may contribute to the improvement of diagnostic accuracy.
Disease duration as an independent predictor of LP in the GI tract in patients with PD
Disease duration, which was analyzed as the variable duration from symptom onset to surgery in this study, was found to be a novel predictor of AS accumulation in the GI tract in PD. This also provides important in vivo evidence of the temporally progressive nature of AS accumulation in the GI tract. A recent study reported that the higher the Braak stage in the brain, the higher the positive rate of AS accumulation in the esophagus (Braak stages 0–3 vs. 4–6, 11% vs. 32%).26 However, the study did not evaluate disease duration in the patients. Moreover, the spatial distribution of AS accumulation in the GI tract was not documented because that study only used esophageal specimens. Previous small-number and biopsy-based in vivo studies attempted to evaluate the temporal pattern of AS accumulation in the GI tract but reported no significant relationship between the frequency of AS accumulation and disease duration. 8, 32
The temporally increasing nature of AS accumulation in the GI tract has important implications for future studies in this field. To increase the sensitivity of detecting AS accumulation in biopsied tissue from peripheral organs, alternative methods, such as pathological AS seed amplification37 or confirmation-specific AS antibody, 38 have been used. However, a recent meta-analysis reported that pathological AS seed amplification, which is thought to be a promising method for detecting peripheral synucleinopathy, has a pooled sensitivity of 88% for the diagnosis of PD but 64% for idiopathic RBD, which is a well-known prodromal synucleinopathy. 37 Differences in the pathological progression of AS accumulation according to disease duration may be an under-recognized influencing factor. Therefore, future studies should consider the temporally progressive nature of AS accumulation when developing new methods using peripheral tissues in PD. The possibility of detecting AS accumulation in the peripheral organs during the early or prodromal stages is likely to be low. This is a major limitation and obstacle to overcome during biomarker development using tissue of the GI tract. In addition, this result suggests that the temporal relationship of the frequency of AS accumulation in other tissues, such as the skin or submandibular gland, should be evaluated through a large-scale study.
Limitations
This study has several limitations. First, we could not evaluate and follow up the controls with AS+. We organized and have been following neurologically normal participants who had undergone surgery of the stomach because of an early gastric cancer in a prospective cohort study (EGC-PPD cohort study). 39 Therefore, we investigated whether the controls with AS + participated in the cohort; however, they did not. Second, the acquisition of detailed clinical information, including GI symptoms, RBD, or parkinsonian symptom severity and laterality, was difficult because of the retrospective multicenter design. Therefore, clinicopathological correlation was insufficient in this study. Finally, only 12 patients with PD underwent surgery before the onset of parkinsonian symptoms. Therefore, there was no meaningful interpretation of the even distribution (66.7%) in both UGI and LGI subgroups at this prodromal stage.