Conceptualization, P.L., B.A.K., E.E. and V.L.; data curation, B.A.K., E.E., A.Y., N.L., J.L., M.S., I.E.V. and S.F.G.; formal analysis, P.L. and I.B.; funding acquisition, J.A.P., P.B., L.B. and V.L.; investigation, P.B., L.B., S.F.G. and V.L.; methodology, P.L. and V.L.; project administration, V.L.; resources, R.W.; supervision, J.A.P., P.B., L.B., S.F.G. and V.L.; visualization, P.L.; writing—original draft, P.L., E.E. and V.L.; writing—review and editing, P.L., E.E., R.W., P.B., L.B. and S.F.G. All authors have read and agreed to the published version of the manuscript.
This work was supported by a Farmer Family Foundation grant award to P.B., with L.B., J.A.P., and V.L. as co-investigators. V.L. is supported by grants from the Department of Defense (W81XWH1810512), the National Institute of Neurological Disorders and Stroke (1R21NS112614-01, 1R01NS113894-01A1, 1R01NS114409-01A1), and a Gibby & Friends vs. Parky Award. S.F.G. is supported by grants from the National Institute of Neurological Disorders and Stroke (1R01NS110838-01A1), the National Institutes on Aging (1R21AG067083-01), the Alzheimer’s Association (AARG-17-530020) and the Michael J. Fox Foundation (MJFF16201).
The Parkinson’s disease (PD) appendix exhibits shifts in the active microbiota that affect lipid metabolism. Metatranscriptomic analysis was used to determine the changes in the functional microbiota in the PD appendix (n = 12 PD, 16 controls). (a) Microbiota changes in the PD appendix. Metatranscriptome data were analyzed by MetaPhlAn2 and a zero-inflated Gaussian mixture model in metagenomeSeq, adjusting for age, sex, RNA integrity number (RIN), and post-mortem interval. Results are displayed using GraPhlAn, showing the taxonomic tree with the kingdom in the center, and branching outwards to phylum, class, order, family, and genus. Microbial taxa highlighted in red are increased in PD, and blue are decreased in PD (q < 0.05, metagenomeSeq). (b) Microbiota metabolic processes are altered in the PD appendix. Top microbial pathways are altered in PD as identified by HumanN2. Red dashed line denotes q < 0.1 pathways as determined by metagenomeSeq.
Proteomic analysis identifies the altered lipid metabolism pathways in the PD appendix and ileum. Pathway enrichment analysis of proteomic changes in the PD appendix relative to controls (n = 3 PD, 3 controls) (a) and in PD ileum relative to controls (n = 4 PD, 4 controls) (b). Pathway analysis of quantitative proteomic data was performed using g:Profiler. Nodes are pathways altered in the PD appendix that were clustered into functionally similar networks by EnrichmentMap (nodes are q < 0.05 pathways, hypergeometric test). Node size represents the number of genes in the pathway gene set, and the edges connect pathways with similar gene sets (0.7 similarity cutoff). The lipid metabolism pathway network is highlighted in peach. (c) Top 10 proteins that were most consistently altered in the PD appendix and ileum. Heatmap showing the proteins ranked as the most consistently disrupted in the PD appendix and ileum, as determined by a robust ranking algorithm. Log fold change is shown, and red signifies greater disruption in PD.
Increase in the microbiota-derived secondary bile acids in the appendix of PD patients. Bile acid analysis was performed by liquid chromatography–mass spectrometry in the PD and control appendix (n = 15 PD, 12 controls) and ileum (n = 20 PD, 20 controls). Bile acid changes were determined by robust linear regression, controlled for age, sex, and postmortem interval. (a) Illustration of the bile acid changes identified in this study and the bile acid pathway. Primary bile acids are generated in the liver and secondary bile acids are produced by microbiota in the intestine. In the secondary bile acid section of the image, boxes highlight the DCA and LCA groups (DCA, LCA and their respective conjugates). Bile acids increased in the PD appendix or PD ileum, relative to controls, are marked by a blue and green arrow, respectively. The flame symbol denotes hydrophobic bile acids that have proinflammatory effects when elevated. (b) Secondary bile acid changes in the PD appendix and PD ileum. The boxplot center line represents the mean, the lower and upper limits are the first and third quartiles (25th and 75th percentiles), and the whiskers are 1.5× the interquartile range. * p < 0.05, robust linear regression.
Dysfunctional cholesterol and lipid metabolism in the PD ileum. Transcript levels of genes in the ileum (a) and liver (b) affecting the abundance of cholesterol and bile in the enterohepatic circulation. Transcript levels of genes affecting cholesterol and bile acid homeostasis (NR1H2, NR1H3, NR1H4, GPBAR1) and their transport and reabsorption into the enterohepatic circulation (NPC1L1, ABCG5, ABCG8, ASBT, OSTα, OSTβ, FABP6) were examined in the ileum. In the liver, the transcript levels of these genes or the equivalent bile acid transporters (NTCP, FABP1) were examined, as well as the rate-limiting enzymes for bile acid production (CYP7A1, CYP27A1). Transcript levels were analyzed by qPCR and normalized to housekeeping genes (villin1, β-actin). The relative expression ± s.e.m in the ileum (n = 8 PD, 6 controls) and liver (n = 6 PD, 6 controls). * p < 0.05, one-way ANOVA.