2023-NIA-LNG-Seminar-Series

1. @lcolladotor lcolladotor.github.io lcolladotor.github.io/bioc_team_ds Harnessing the power of spatially-resolved transcriptomics one step at a time (part 2/2) Leonardo Collado Torres, Investigator #Visium_SPG_AD 2023 NIA/LNG Neurodegenerative Diseases Seminar Series June 09 2023 Slides available at speakerdeck.com/lcolladotor
2. twitter.com/sanghokwon17/status/1650589385379962881 from 2023-04-24 Sang Ho Kwon @sanghokwon17 DOI: 10.1101/2023.04.20.537710 #Visium_SPG_AD In collaboration with Download data with spatialLIBD::fetch_data() research.libd.org /Visium_SPG_AD
3. Visium spatial clustering works for variables with high % variance explained. But what about other ones? DOI: 10.1038/s41593-020-00787-0
4. Alzheimer’s disease related neuropathology Adopted and modified from B Wang (2018) and the Brain from the Top to Bottom in McGill University 20 um Sang Ho Kwon Amyloid beta pTau
5. Hypothesis Local tissue microenvironments in close proximity to AD-related neuropathology have distinct cellular and molecular signatures. Sang Ho Kwon
6. Visium Spatial Proteogenomics (Visium-SPG) Visium-SPG = Visium SRT + immunofluorescence (using identical tissue samples) Sang Ho Kwon
7. Experimental design & study overview Braak V-VI & CERAD frequent Sang Ho Kwon
8. Donor demographics BrNum Age of death (years old) Sex Ancestry PMI (hours) Dx Best RIN PFC Braak CERAD APOE # Visium Replicates Br3854 65.75 Female European 31.5 Alzheimer 7.0 Stage VI Frequent e3/e4 2 Br3873 88.78 Female European 29.0 Alzheimer 7.2 Stage V Frequent e3/e3 2 Br3880 90.47 Male European 35.0 Alzheimer 7.1 Stage VI Frequent e3/e3 3 Br3874 73.05 Male European 13.5 Control 7.2 Stage IV None e2/e3 3 DOI: 10.1101/2023.04.20.537710
9. AD pathology signal is too small to detect by spatially-resolved gene expression alone research.libd.org/Visium_SPG_AD/
10. Visium * ~5k spots in honeycomb * gene expression per spot * tissue (H&E staining) Immunofluorescence (IF) * multi-channel (6) images * identifies morphological features of interest * large: might be broken in tiles Channel 1 * triangle feature Channel 2 * cloud feature Channel 6 * xyz feature Tissue (bright field image) Visium spot Channel 1 feature Channel 2 feature + Visium Spatial Proteogenomics (Visium-SPG / IF) raw data: 2 types
11. Spot ID Gene 1 Gene 2 Gene X spot0001 0 12 39 spot0002 4 0 27 Spot ID Gene 1 Gene 2 Gene X In Tissue # cells spot0001 0 12 39 true 3 spot0002 4 0 27 false 0 * spaceranger * Loupe Browser * VistoSeg on H&E bright field image Visium Analysis @MadhaviTippani @HeenaDivecha cell VistoSeg DOI: 10.1101/2021.08.04.452489
12. Feature ID X center Y center type intensity feat0001 5 102 triangle 130.4 feat0002 10 30 cloud 99.1 Max (X, Y) Min (X, Y) Area ... SPG / IF Image Analysis * segment each channel * find features Challenges: * morphological features can be quite diverse * images are large * multiple tiles +
13. Spot ID # Triangle # Cloud % triangle % cloud spot0001 0 12 0 17 spot0002 4 0 27 0 Merge Visium & IF IF Spot ID Gene 1 Gene 2 Gene X In Tissue # cells spot0001 0 12 39 true 3 spot0002 4 0 27 false 0 Visium downstream * QC * analyses spatialLIBD DOI: 10.1186/s12864-022-08601-w VistoSeg DOI: 10.1101/2021.08.04.452489 Check version 2!! @MadhaviTippani
14. Estimating pathological burden per spot to generate transcriptome-scale maps of AD pathology 1. both > 2. A𝛽 or pTau > 3. next_both > 4. next_A𝛽 or next_pTau > 5. none
15. AD pathology signal is too small to detect by spatially-resolved gene expression alone research.libd.org/Visium_SPG_AD/
16. Pathology burden is more common in the gray matter BayesSpace with 2 clusters: Cluster 1: gray matter (GM) Cluster 2: white matter (WM) 1 2
17. Identifying transcriptional signatures of AD-related neuropathology Sang Ho Kwon
18. Identifying transcriptional signatures of Abeta-related neuropathology Sang Ho Kwon
19. Validation study design using RNAscope FISH-IF Sang Ho Kwon
20. HALO-based image analysis
21. 6 Abeta-associated DEGs on RNAscope FISH-IF (Kruskal-Wallis test, *p<0.05, &p<0.005, and #p<0.0001) Sang Ho Kwon
22. Visium + RNAscope to provide additional insights into spatial gene expression gradients Sang Ho Kwon
23. Identifying transcriptional signatures of Abeta microenvironment Previously not significant Sang Ho Kwon A𝛽 + next_A𝛽 = A𝛽_env (environment)
24. bioconductor.org/packages/spatialLIBD Pardo et al, 2022 DOI 10.1186/s12864-022-08601-w Maynard, Collado-Torres, 2021 DOI 10.1038/s41593-020-00787-0 Brenda Pardo Abby Spangler @PardoBree @abspangler Louise A. Huuki-Myers @lahuuki
25. Conclusions • Visium-SPG can identify local transcriptional signatures of late-stage AD pathology • RNAscope FISH-IF can further explore spatial gene expression patterns of a few select features • Proof-of-concept study: our work lays the groundwork. Can now use Visium-SPG for other diseases too • Future studies are needed to produce mechanistic insights • Explore our code & data at research.libd.org/Visium_SPG_AD & download it with spatialLIBD::fetch_data() ◦ There’s lots more to explore with this data! Sang Ho Kwon Slide adapted from:
26. twitter.com/sanghokwon17/status/1650589385379962881 from 2023-04-24 Sang Ho Kwon @sanghokwon17 DOI: 10.1101/2023.04.20.537710 #Visium_SPG_AD In collaboration with Download data with spatialLIBD::fetch_data() research.libd.org /Visium_SPG_AD
27. @MadhaviTippani Madhavi Tippani @HeenaDivecha Heena R Divecha @lmwebr Lukas M Weber @stephaniehicks Stephanie C Hicks @abspangler Abby Spangler @martinowk Keri Martinowich @CerceoPage Stephanie C Page @kr_maynard Kristen R Maynard @lcolladotor Leonardo Collado-Torres @Nick-Eagles (GH) Nicholas J Eagles Kelsey D Montgomery Sang Ho Kwon Image Analysis Expression Analysis Data Generation Thomas M Hyde @lahuuki Louise A Huuki-Myers @BoyiGuo Boyi Guo @mattntran Matthew N Tran @sowmyapartybun Sowmya Parthiban Slides available at speakerdeck.com /lcolladotor + Many more LIBD, JHU, and external collaborators @mgrantpeters Melissa Grant-Peters @prashanthi-ravichandran (GH) Prashanthi Ravichandran
28. #GBD23 Thank you for having us over in the UK 󰏅! !Gracias Pilar & Cornelis for the invitation to present at NIA/LNG!
29. lcolladotor.github.io @lcolladotor