Periodic Reporting for period 1 - TFZN (Understanding the mechanisms that govern organ morphostasis and repair)
Reporting period: 2019-10-01 to 2021-09-30
Understanding organogenesis, organ morphostasis and regeneration is crucial to many areas of biology and medicine, including the generation of “organs-on-a-chip” and controlled in vitro organ engineering for clinical applications. As life expectancy around the world rises, knowledge on organ formation and regeneration is becoming crucial to treat diseases of old age.
This work focused on understanding how tissue patterning and proportions are recovered after damage by the collective behavior of cells interacting simultaneously. For this, the individual researcher used the neuromasts of the superficial lateral line in zebrafish as an optimal experimental system to understand organ regeneration. These small mechanosensory organs enable fishes and amphibians to sense water displacement around their bodies. Neuromasts are formed by a circular pseudo-stratified epithelium of about 70 cells that are classified in three main cell types: mantle cells, sustentacular cells and hair-cells. Mantle cells form an outer rim of the neuromast epithelium. Inner sustentacular cells comprise the majority of cells and behave as stem cells.
Cilliated hair-cells are the mechanosensory elements, and are located in the center of the organ. Neuromast hair cells are homologous to those found in the mammalian inner ear. When fish hair cells are lost due to chemical or mechanical damage, they regenerate an unlimited number of times. By stark contrast, the mammalian ear does not have this capacity. This means that, as we age, hair cells are lost to environmental insults such as loud noises, some medications and certain health conditions. We cannot recover them.
People in old age, therefore, suffer disproportionately from hearing loss and tinnitus (ear ringing). In addition, hair cells in the vestibular system that sense acceleration and gravity are critical for balance. Deficits in this system can cause nausea and falls that can be catastrophic for people.
The scientific objective of this action is to gain understanding of the process of organ patterning during development or regeneration. We have studied the neuromasts of the zebrafish lateral line as a model to disentangle general principles of organogenesis as well as a model of the inner ear in particular.
We narrowed down a list of candidate genes which we then studied in the neuromast by expression and loss-of-function analysis. We then studied how mutations in a subset of genes in the wnt or notch pathways affect the spatial organization of cells within the organ. To achieve it, we used live spinning-disk microscopy. We analyzed the resulting image data with machine learning techniques that allow us to extract a digital representations of the cells in order to compare it with computational models.
Mutations in organogenesis usually result in broad defects in morphology of organs. The processes we have studied, however, are more subtle. We have focused on cell orientation relative to each other, changes in cell-cell contact and their interplay with the cell shape and migration. Because this is seldom studied, specially in the context of a live vertebrate, this work contributes to answer how collectives of cells organize themselves to fulfill all the intricate functions of our body.
This work focused on understanding how tissue patterning and proportions are recovered after damage by the collective behavior of cells interacting simultaneously. For this, the individual researcher used the neuromasts of the superficial lateral line in zebrafish as an optimal experimental system to understand organ regeneration. These small mechanosensory organs enable fishes and amphibians to sense water displacement around their bodies. Neuromasts are formed by a circular pseudo-stratified epithelium of about 70 cells that are classified in three main cell types: mantle cells, sustentacular cells and hair-cells. Mantle cells form an outer rim of the neuromast epithelium. Inner sustentacular cells comprise the majority of cells and behave as stem cells.
Cilliated hair-cells are the mechanosensory elements, and are located in the center of the organ. Neuromast hair cells are homologous to those found in the mammalian inner ear. When fish hair cells are lost due to chemical or mechanical damage, they regenerate an unlimited number of times. By stark contrast, the mammalian ear does not have this capacity. This means that, as we age, hair cells are lost to environmental insults such as loud noises, some medications and certain health conditions. We cannot recover them.
People in old age, therefore, suffer disproportionately from hearing loss and tinnitus (ear ringing). In addition, hair cells in the vestibular system that sense acceleration and gravity are critical for balance. Deficits in this system can cause nausea and falls that can be catastrophic for people.
The scientific objective of this action is to gain understanding of the process of organ patterning during development or regeneration. We have studied the neuromasts of the zebrafish lateral line as a model to disentangle general principles of organogenesis as well as a model of the inner ear in particular.
We narrowed down a list of candidate genes which we then studied in the neuromast by expression and loss-of-function analysis. We then studied how mutations in a subset of genes in the wnt or notch pathways affect the spatial organization of cells within the organ. To achieve it, we used live spinning-disk microscopy. We analyzed the resulting image data with machine learning techniques that allow us to extract a digital representations of the cells in order to compare it with computational models.
Mutations in organogenesis usually result in broad defects in morphology of organs. The processes we have studied, however, are more subtle. We have focused on cell orientation relative to each other, changes in cell-cell contact and their interplay with the cell shape and migration. Because this is seldom studied, specially in the context of a live vertebrate, this work contributes to answer how collectives of cells organize themselves to fulfill all the intricate functions of our body.
For the work carried out in this fellowship, we have studied the dynamics of this process, called planar cell inversion in high detail using live microscopy tracking and computational modelling.
Imagine a dance floor in which a couple, a man and a woman, enter and position themselves in the center of the room. They face each other. New couples enter regularly and they go to dance next to the first couple, after a while, you have all the men looking in the same direction: where the women are, and reciprocally, all women are facing in the same direction, toward the men. Any new couple entering the room can see if they have entered in the "correct" position, if not, they will rotate 180 deg until they join the group and the man is facing towards his partner and all the other women, and vice versa.
Believe it or not, something very similar happens when new hair cells originate in a neuromast. The 180 degree swirl that cells perform is what we called planar cell inversion.
Using a fluorescent microscope, we filmed the process at a resolution that allowed us to measure the velocity, angle, and shape of many cells as they rotate, to characterize the statistics of the process. We then compared the normal rotations to two genetic perturbations known to alter the identity of the hair cells in opposite ways. In the dance floor analogy, this would be like to putting a tuxedo to all participants, or conversely, a dress. To our surprise, these perturbations do very little to alter the dynamics of planar cell inversion. The results of this investigation will be published in a future manuscript.
A mutation in a different gene, called Wnt, affects the orientation of hair cells in a different way. Instead of hair cells being oriented in one direction, or the other, the whole collective forms a swirl, or a spiral pattern. We realized that the formation of the spiral pattern could be explained solely in terms of physical interactions between the cells. The physics involved have been used to describe completely different phenomena in liquid crystals, magnetic systems, and crowds in a hard rock concert. Therefore, the analogy with the dance room is not so misplaced. The physics models describe the formation of complex patterns solely by the interaction of agents with an orientation, where the "agent" can be a hair cell, a spins in a magnet, or a person in a crowd. The results of this study will also be eventually published in a paper on the mechanisms of neuromast patterning.
Along the future papers, and according to the data management plan, we will freely share the data from microscopy images for three reasons: (i) to make our work easier to understand and reproduce. (ii) because microscopy images contain multidimensional data that other researchers/students might use to generate new hypotheses. (iii) the annotated data might be useful to refine machine learning models that extract quantitative features from images of cells.
Imagine a dance floor in which a couple, a man and a woman, enter and position themselves in the center of the room. They face each other. New couples enter regularly and they go to dance next to the first couple, after a while, you have all the men looking in the same direction: where the women are, and reciprocally, all women are facing in the same direction, toward the men. Any new couple entering the room can see if they have entered in the "correct" position, if not, they will rotate 180 deg until they join the group and the man is facing towards his partner and all the other women, and vice versa.
Believe it or not, something very similar happens when new hair cells originate in a neuromast. The 180 degree swirl that cells perform is what we called planar cell inversion.
Using a fluorescent microscope, we filmed the process at a resolution that allowed us to measure the velocity, angle, and shape of many cells as they rotate, to characterize the statistics of the process. We then compared the normal rotations to two genetic perturbations known to alter the identity of the hair cells in opposite ways. In the dance floor analogy, this would be like to putting a tuxedo to all participants, or conversely, a dress. To our surprise, these perturbations do very little to alter the dynamics of planar cell inversion. The results of this investigation will be published in a future manuscript.
A mutation in a different gene, called Wnt, affects the orientation of hair cells in a different way. Instead of hair cells being oriented in one direction, or the other, the whole collective forms a swirl, or a spiral pattern. We realized that the formation of the spiral pattern could be explained solely in terms of physical interactions between the cells. The physics involved have been used to describe completely different phenomena in liquid crystals, magnetic systems, and crowds in a hard rock concert. Therefore, the analogy with the dance room is not so misplaced. The physics models describe the formation of complex patterns solely by the interaction of agents with an orientation, where the "agent" can be a hair cell, a spins in a magnet, or a person in a crowd. The results of this study will also be eventually published in a paper on the mechanisms of neuromast patterning.
Along the future papers, and according to the data management plan, we will freely share the data from microscopy images for three reasons: (i) to make our work easier to understand and reproduce. (ii) because microscopy images contain multidimensional data that other researchers/students might use to generate new hypotheses. (iii) the annotated data might be useful to refine machine learning models that extract quantitative features from images of cells.
The work from this action has pushed the knowledge on general features of multicellular interactions. We have studied the dynamics of how cells position themselves relative to others within a tissue and then how this collective of cells aligns itself with the body axis. This work stems from a modern interdisciplinary approach that combines biology with computer science and physics. As a consequence, the insights provided by our work are likely to be useful for the study of many different tissues, not only the inner ear.
There still remains work to be done in order to translate the results obtained into scientific publications. Even then, published papers seldom have an immediate impact on the wider society except for small academic communities. However, we remain confident that this is an intellectual contribution, that among with many others, will help the development of better treatments. Ultimately, understanding how cells act as a collective during organogenesis will inform the engineering of organs in vitro and the control of tissue repair in vivo.
There still remains work to be done in order to translate the results obtained into scientific publications. Even then, published papers seldom have an immediate impact on the wider society except for small academic communities. However, we remain confident that this is an intellectual contribution, that among with many others, will help the development of better treatments. Ultimately, understanding how cells act as a collective during organogenesis will inform the engineering of organs in vitro and the control of tissue repair in vivo.