Workflow and Methods of High-Content Time-Lapse Analysis for Quantifying Intracellular Calcium Signals

Calcium ions (Ca2+) play a fundamental role in a variety of physiological functions in many cell types by acting as a secondary messenger. Variation of intracellular Ca2+ concentration ([Ca2+]i) is often observed when the cell is stimulated. However, it is a challenging task to automatically quantify intracellular [Ca2+]i in a population of cells. In this study, we present a workflow including specific algorithms for the automated intracellular calcium signal analysis using high-content, time-lapse cellular images. The experimental validations indicate the effectiveness of the proposed workflow and algorithms. We applied the workflow to analyze the intracellular calcium signals induced by different concentrations of H2O2 in the cell lines transfected by presenilin-1 (PS-1) that is known to be closely related to the familial Alzheimer’s disease (FAD). The analysis results imply an important role of mutant PS-1, but not normal human PS-1 and mutant human amyloid precursor protein (APP), in enhancing intracellular calcium signaling induced by H2O2.

[1]  Joakim Lindblad,et al.  Algorithms for Cytoplasm Segmentation of Fluorescence Labelled Cells , 2002, Analytical cellular pathology : the journal of the European Society for Analytical Cellular Pathology.

[2]  Jeffrey S. Morris,et al.  Improved peak detection and quantification of mass spectrometry data acquired from surface‐enhanced laser desorption and ionization by denoising spectra with the undecimated discrete wavelet transform , 2005, Proteomics.

[3]  Xiaobo Zhou,et al.  Automated segmentation, classification, and tracking of cancer cell nuclei in time-lapse microscopy , 2006, IEEE Transactions on Biomedical Engineering.

[4]  W. Xia,et al.  Role of presenilin in gamma-secretase cleavage of amyloid precursor protein. , 2000, Experimental gerontology.

[5]  Peter Lipp,et al.  Calcium imaging of individual erythrocytes: problems and approaches. , 2006, Cell calcium.

[6]  P.K Sahoo,et al.  A survey of thresholding techniques , 1988, Comput. Vis. Graph. Image Process..

[7]  Carlos Ortiz-de-Solorzano,et al.  High-throughput analysis of multispectral images of breast cancer tissue , 2006, IEEE Transactions on Image Processing.

[8]  N. Otsu.  A threshold selection method from gray level histograms , 1979 .

[9]  Richard O. Lynch,et al.  Introduction to Design and Analysis of Experiments , 1999, Technometrics.

[10]  J. Richardson,et al.  Actions of neurotoxic beta-amyloid on calcium homeostasis and viability of PC12 cells are blocked by antioxidants but not by calcium channel antagonists. , 1996, Journal of neurochemistry.

[11]  Badrinath Roysam,et al.  Hierarchical, model‐based merging of multiple fragments for improved three‐dimensional segmentation of nuclei , 2005, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[12]  Bülent Sankur,et al.  Survey over image thresholding techniques and quantitative performance evaluation , 2004, J. Electronic Imaging.

[13]  Badrinath Roysam,et al.  A hybrid 3D watershed algorithm incorporating gradient cues and object models for automatic segmentation of nuclei in confocal image stacks , 2003, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[14]  R. Black,et al.  Induction of Alzheimer antigens by an uncoupler of oxidative phosphorylation. , 1990, Archives of Neurology.

[15]  J. Hsu.  Multiple Comparisons: Theory and Methods , 1996 .

[16]  Yan Zhou,et al.  Actions of Neurotoxic β‐Amyloid on Calcium Homeostasis and Viability of PC12 Cells Are Blocked by Antioxidants but Not by Calcium Channel Antagonists , 1996 .

[17]  Serge Beucher,et al.  THE WATERSHED TRANSFORMATION APPLIED TO IMAGE SEGMENTATION , 2009 .

[18]  Tim W. Nattkemper,et al.  Automatic segmentation of digital micrographs: A survey , 2004, MedInfo.

[19]  Junaed Sattar.  Snakes , Shapes and Gradient Vector Flow , 2022 .

[20]  C. Fowler,et al.  Comparison of the effects of hydrogen peroxide, 4-hydroxy-2-nonenal and β-amyloid (25–35) upon calcium signalling , 1998, Neurochemistry International.

[21]  Gordon L Fain,et al.  Measurement of cytoplasmic calcium concentration in the rods of wild‐type and transducin knock‐out mice , 2002, The Journal of physiology.

[22]  B. Strooper,et al.  Presenilins Form ER Ca2+ Leak Channels, a Function Disrupted by Familial Alzheimer's Disease-Linked Mutations , 2006, Cell.

[23]  Frank M LaFerla,et al.  Calcium dysregulation in Alzheimer's disease: recent advances gained from genetically modified animals. , 2005, Cell calcium.

[24]  Xiaobo Zhou,et al.  High content cellular imaging for drug development , 2006 .

[25]  Tomas Gustavsson,et al.  Time-lapse microscopy and image processing for stem cell research: modeling cell migration , 2003, SPIE Medical Imaging.

[26]  Paweł Pomorski,et al.  Mouse oocytes fertilised by ICSI during in vitro maturation retain the ability to be activated after refertilisation in metaphase II and can generate Ca2+ oscillations , 2007, BMC Developmental Biology.

[27]  F. LaFerla.  Calcium dyshomeostasis and intracellular signalling in alzheimer's disease , 2002, Nature Reviews Neuroscience.

[28]  Joakim Lindblad,et al.  Image analysis for automatic segmentation of cytoplasms and classification of Rac1 activation , 2004, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[29]  C. Colton,et al.  Production of superoxide anions by a CNS macrophage, the microglia , 1987, FEBS letters.

[30]  J. Sethian,et al.  Fronts propagating with curvature-dependent speed: algorithms based on Hamilton-Jacobi formulations , 1988 .

[31]  D. Selkoe,et al.  Enhanced Production and Oligomerization of the 42-residue Amyloid β-Protein by Chinese Hamster Ovary Cells Stably Expressing Mutant Presenilins* , 1997, The Journal of Biological Chemistry.

[32]  J. Hardy,et al.  The Amyloid Hypothesis of Alzheimer ’ s Disease : Progress and Problems on the Road to Therapeutics , 2009 .

[33]  Tony Lindeberg,et al.  Feature Detection with Automatic Scale Selection , 1998, International Journal of Computer Vision.

[34]  W Xia,et al.  Role of presenilin in γ-secretase cleavage of amyloid precursor protein , 2000, Experimental Gerontology.

[35]  Baba C. Vemuri,et al.  Shape Modeling with Front Propagation: A Level Set Approach , 1995, IEEE Trans. Pattern Anal. Mach. Intell..

[36]  Grace E. Stutzmann.  Calcium Dysregulation, IP3 Signaling, and Alzheimer’s Disease , 2005, The Neuroscientist : a review journal bringing neurobiology, neurology and psychiatry.

[37]  Alex M. Andrew,et al.  Level Set Methods and Fast Marching Methods: Evolving Interfaces in Computational Geometry, Fluid Mechanics, Computer Vision, and Materials Science (2nd edition) , 2000 .

[38]  M. Mattson,et al.  A model for beta-amyloid aggregation and neurotoxicity based on free radical generation by the peptide: relevance to Alzheimer disease. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[39]  Guanglei Xiong,et al.  Automated Segmentation of Drosophila RNAi Fluorescence Cellular Images Using Deformable Models , 2006, IEEE Transactions on Circuits and Systems I: Regular Papers.

[40]  Luc Vincent,et al.  Watersheds in Digital Spaces: An Efficient Algorithm Based on Immersion Simulations , 1991, IEEE Trans. Pattern Anal. Mach. Intell..

[41]  S. Osher,et al.  Algorithms Based on Hamilton-Jacobi Formulations , 1988 .

[42]  Hyung-Myung Kim,et al.  Optimization of requantization codebook for vector quantization , 2006, IEEE Transactions on Image Processing.