an open data exchange for cell migration research

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TOWARDS AN OPEN DATA EXCHANGE ECOSYSTEM: FORGING A NEW PATH FOR CELL MIGRATION DATA

ANALYSIS AND MINING

18 October 2016 public PhD defense - paola masuzzo

OpenCV; http://www.xinkaiwu.com/index.html

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Introduction to cell migration

Research problems

ResultsCellMissy: an automated tool for cell

migrationA new CellMissy module for single-cell

analysisEngineering features to describe

stochasticityTowards an open data exchange

ecosystem

Conclusions and future perspectives

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Cell migration is necessary for many physiological functions

Basementmembrane Wound

Migrating epithelial cells

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Cell migration is necessary for many physiological functions

Blood vessel

Site of tissue injury

Migrating neutrophil

Basementmembrane Wound

Migrating epithelial cells

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Unfortunately, it is also implicated in many diseases, such as metastatic cancer

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The cytoskeleton is the key structural framework responsible for cell migration

Adapted from Herzog et al., Cell Bio Lab Handbook, 1994

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Different actin filament structures are essential for cell migration

Actin network

Filopodia

Lamellipodium

movementLeading edge

2D migration

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Different actin filament structures are essential for cell migration

Actin network

Filopodia

Lamellipodium

movementLeading edge

2D migration 3D invasion

Basementmembrane

Epithelial cell

Tumor cell

Extracellular matrix

Invadopodia

Protruding bleb

Lamellipodia or pseudopodia

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Cell translocation depends on a cyclic interplay between cell adhesion and de-adhesion

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A typical experimental workflow for a cell migration study is composed of diverse steps

Servier Medical Art, CC-BY 3.0; Cell Image Library, CC-BY 3.0

sample preparation

image acquisition

image processing

data analysis

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Several assays are available for in vitroassessment of cell migration


sample preparation

image acquisition

image processing

data analysis

Wound-healing assay

pipette tip

scratch

Cell-exclusion zone assay

cell-free zone

siliconestopper

Spheroid assay

multicellular spheroid

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Live-cell phase-contrast and fluorescence microscopy generates quantifiable data

Servier Medical Art, CC-BY 3.0; Cell Image Library, Public Domain; Mierke et al., 2011

sample preparation

image acquisition

image processing

data analysis

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Time-lapse video microscopy capturescell migration dynamics

time

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Image processing is a multi-step operation comprising segmentation and tracking

Servier Medical Art, CC-BY 3.0; Cell Image Library, CC-BY 3.0, Harder et al., 2015

imagepre-processing

celltracking

cellsegmentation

sample preparation

image acquisition

image processing

data analysis

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Quantitative parameters are then extracted for cell sheet and single-cell trajectories

Image processed with CELLMIA, UGent (Van Troys M, Ampe C) and DciLabs

area in timeµm²/min

coordinates (x, y, t)µm/min

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Ultimately, data analysis enables interpretation of the experiment

Servier Medical Art, CC-BY 3.0; Cell Image Library, CC-BY 3.0; O’ Brien et al., 2014

sample preparation

image acquisition

image processing

data analysis

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The Ghent platform enables automation of high-throughput cell migration experiments

Phase-contrastlive-cell imaging

time-lapse: 16-48 hinterval: 15-20 min

Adapted from Lynn Huyck, PhD thesis, 2012 (promoter Van Troys M)

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Images are automatically processed

Adapted from Lynn Huyck, PhD thesis, 2012 (promoter Van Troys M); images processed with CELLMIA, UGent and DciLabs

t = 0h t = 24h

t = 0h t = 36h

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Such high-throughput experiments produce complex and rich data sets


sample preparation

image acquisition

image processing

data analysis

• paper laboratory notebooks

• electronic laboratory notebooks

• spreadsheets• text files• protocols• papers...

• raw files• XML files• proprietary

microscope or acquisition software files ND2 for Nikon, LIF for Leica, OIB or OIF for Olympus, LSM or ZVI for Zeiss

• image files with pixel values and metadata

• png, jpeg, tiff, avi• text files

describing processing algorithms

• text files describing extracted features

• graphs, plots• analysis pipelines• text files

describing computational algorithms...

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Research problems





ecosystem


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The overall objective of this PhD is to advance bioinformatics for cell migration

Cell migration experiments have become de factohigh-throughput, but bioinformatics has lagged behind

Due to lack of automated systems and appropriate algorithms, a big proportion of cell migration data is still not exploited

The heterogeneity of the field hampers open data exchange, impeding advanced data analysis and mining

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Research problems





ecosystem


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CellMissy is our open-source tool for cell migration data management and analysis

0 3h 6h

wound

cells

Experiment

Data Analyzer

Data Loader

Collective cell migration Single-cell migration

Experiment Manager

Masuzzo et al., Bioinformatics, 2013; https://github.com/compomics/cellmissy

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CellMissy guides and captures the experimental setup

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This experimental setup encloses detailed metadata annotation

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CellMissy can automatically import all the data and metadata

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All the data are then stored in a structured way in a relational database

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CellMissy enables efficient data exploration and analysis

time (min)

Area

()

wound areacell-covered area

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A primary focus of data analysis is statistical comparison of samples

analysis report with graphs,tables and results

cell sheet velocity (µm²/min)

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Research problems


migrationA new CellMissy module for single-

cell analysisEngineering features to describe


ecosystem


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Cell migration can occur in both collective and individual fashion

Collective migration

Multicellular streaming

Mesenchymal

Amoeboid (blebs)

Amoeboid (pseudopodia, filopodia)

INDIVIDUAL MIGRATION

COLLECTIVE MIGRATION

Adapted from Friedl et al., J. Exp. Med., 2010

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Many informative parameters can be derived from single-cell trajectories

Masuzzo et al., under review, 2016

x

ysingle cell

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ࢻ

Euclideandistance

Cumulativedistance

x

ysingle cell

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ࢻ

Euclideandistance

Cumulativedistance

x

ysingle cell parameter mathematical description

di: instantaneous displacement of the cell centroid between adjacent time points

si: instantaneous speed between adjacent time points

αi: turning angle between consecutive steps

dtot: cumulative distance, total distance travelled

dnet: Euclidean distance, net distance travelled

ep_dr: end-point directionality ratio (confinement ratio, meandering index)

MD: median displacement

MS: median speed

MTA: median turning angle

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Step-centric parameters are aggregated values of all migration steps

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Trajectory-centric parameters are instead computed on each cell and then averaged for the cell population

...

trajectory 1

trajectory 2

trajectory 3

trajectory 4

trajectory 5

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The new single-cell module allows for both these computations to take place


...

trajectory 1

trajectory 2

trajectory 3

trajectory 4

trajectory 5

trajectory-centric parameters

trajectory displacement (µm)

dens

ity

step displacement (µm)

dens

ity

pool of migration steps

step-centric parameters

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A flexible two-step filtering criterion is implemented for data quality control


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Research problems





ecosystem


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More advanced features are needed to describe the complexity of the phenomenon

Masuzzo et al., in preparation, 2016

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The enclosing circle set is a new way to describe local structure of trajectories


radius: 6 µmnr_circles: 14

directionof motion

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The enclosing circle set is a new way to describe local structure of trajectories



directionof motion


directionof motion

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The fractal dimension is derived from the enclosing circle set

𝐹𝐷 (𝑆 )=lim𝑟→0

¿¿


FD=0.35 FD=0.83

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Research problems





ecosystem


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Data and metadata exchange options are already available in CellMissy

lab A

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lab A lab B

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lab B

This is one file in CellMissy! (≈10 MB)

lab A

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But we can easily extend this conceptto a bigger scale

DataRepository Local Software

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The seed of this idea was planted in the field

Friedl et al., Nature Reviews, 2012

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It only needed some water to grow

Cell migration workshop, Ghent, March 2014; Masuzzo et al., Trends in Cell Biology, 2015

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An open data exchange ecosystem for cell migration research is now on its way

Masuzzo et al., Trends in Cell Biology, 2015

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This open data ecosystem falls into the broader context of open science

Knoth and Pontika, Open Science Taxonomy, figshare, 2015

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Open access is another key factor in the open science equation

Knoth and Pontika, Open Science Taxonomy, figshare, 2015

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The impacts of open access are very broad and affect many areas

Tennant, Masuzzo et al., F1000Research, 2016; Wikimedia Commons, Public Domain

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Publish your work open access can bring you enormous benefits

CC-BY Danny Kingsley & Sarah Brown

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Open access also allows automatic knowledge extraction through text mining

automatically detect a set of core information reported when describing cell migration experiments

check for nomenclature consistency, the use of common terms or ontologies to describe the same concept

construct a knowledge map to describe the state-of-the-art, especially in terms of cell motility-related compounds and cancer cell lines

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Research problems





ecosystem


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This PhD has tackled key bioinformatics challenges in cell migration research

CellMissy is the first free and open-source tool for the management, annotation and storage of cell migration experiments

A new dedicated module, together with novel features, enable detailed and more complex quantification ofsingle-cell migration experiments

International research efforts are currently spent towards the establishment of an open data exchange ecosystem, opening the way to more advanced data analysis and mining strategies

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These results have paved the way to even more exciting opportunities

CellMissy has already been extended with dose-response analysis capabilities, and more development is planned to allow meta-analyses to take place

Further engineering, validation, and selection of single-cell migration features is planned; these features will then be used to automatically detect and classify migratory phenotypes

Joined efforts of MULTIMOT and the CMSO will ultimately enable global data dissemination in the field, allowing data re-use, re-discovery and re-purpose

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an open data exchange for cell migration research

Education