Analysis of lens cell fate and eye morphogenesis in transgenic mice ablated for cells of the lens lineage

Development ◽  
1989 ◽  
Vol 106 (3) ◽  
pp. 457-463 ◽  
Author(s):  
M.L. Breitman ◽  
D.M. Bryce ◽  
E. Giddens ◽  
S. Clapoff ◽  
D. Goring ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Cell Fate ◽  
Vitreous Body ◽  
Cell Types ◽  
Ciliary Epithelium ◽  
Specific Cell ◽  
Promoter Sequences ◽  
Fiber Cells ◽  
Lens Cell ◽  
Diphtheria Toxin A

Transgenic mice carrying the diphtheria toxin A gene driven by mouse gamma 2-crystallin promoter sequences manifest microphthalmia due to ablation of fiber cells in the ocular lens. Here we map ablation events in the lens by crossing animals hemizygous for the ablation construct with transgenic mice homozygous for the in situ lacZ reporter gene driven by identical gamma 2-crystallin promoter sequences. By comparing the spatial distribution of lacZ-expressing cells and the profile of gamma-crystallin gene expression in the lenses of normal and microphthalmic offspring, the contributions of specific cell types to lens development were examined. The results suggest that phenotypically and developmentally distinct populations of lens fiber cells are able to contribute to the lens nucleus during organogenesis. We also show that dosage of the transgene and its site of integration influence the extent of ablation. In those mice homozygous for the transgene and completely lacking cells of the lens lineage, we show that the sclera, cornea, and ciliary epithelium are reduced in size but, otherwise, reasonably well formed. In contrast, the anterior chamber, iris, and vitreous body are not discernible while the sensory retina is highly convoluted and extensively fills the vitreous chamber.

scholarly journals Histone Acetyltransferases and Stem Cell Identity

Cancers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2407
Author(s):  
Ruicen He ◽  
Arthur Dantas ◽  
Karl Riabowol
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Fate ◽  
Epigenetic Modification ◽  
Cell Types ◽  
Histone Acetyltransferases ◽  
Specific Cell ◽  
Cell Fates ◽  
Cell Identity ◽  
Hematopoietic Stem

Acetylation of histones is a key epigenetic modification involved in transcriptional regulation. The addition of acetyl groups to histone tails generally reduces histone-DNA interactions in the nucleosome leading to increased accessibility for transcription factors and core transcriptional machinery to bind their target sequences. There are approximately 30 histone acetyltransferases and their corresponding complexes, each of which affect the expression of a subset of genes. Because cell identity is determined by gene expression profile, it is unsurprising that the HATs responsible for inducing expression of these genes play a crucial role in determining cell fate. Here, we explore the role of HATs in the maintenance and differentiation of various stem cell types. Several HAT complexes have been characterized to play an important role in activating genes that allow stem cells to self-renew. Knockdown or loss of their activity leads to reduced expression and or differentiation while particular HATs drive differentiation towards specific cell fates. In this study we review functions of the HAT complexes active in pluripotent stem cells, hematopoietic stem cells, muscle satellite cells, mesenchymal stem cells, neural stem cells, and cancer stem cells.

scholarly journals The retinoblastoma protein-binding region of simian virus 40 large T antigen alters cell cycle regulation in lenses of transgenic mice.

1994 ◽  
Vol 14 (10) ◽  
pp. 6743-6754 ◽  
Author(s):  
L Fromm ◽  
W Shawlot ◽  
K Gunning ◽  
J S Butel ◽  
P A Overbeek
Keyword(s):  
Cell Cycle ◽  
Transgenic Mice ◽  
Retinoblastoma Protein ◽  
Simian Virus 40 ◽  
Cell Types ◽  
Simian Virus ◽  
T Antigen ◽  
Lens Fiber ◽  
Large T Antigen ◽  
Fiber Cells

Regulation of the cell cycle is a critical aspect of cellular proliferation, differentiation, and transformation. In many cell types, the differentiation process is accompanied by a loss of proliferative capability, so that terminally differentiated cells become postmitotic and no longer progress through the cell cycle. In the experiments described here, the ocular lens has been used as a system to examine the role of the retinoblastoma protein (pRb) family in regulation of the cell cycle during differentiation. The ocular lens is an ideal system for such studies, since it is composed of just two cell types: epithelial cells, which are capable of proliferation, and fiber cells, which are postmitotic. In order to inactivate pRb in viable mice, genes encoding either a truncated version of simian virus 40 large T antigen or the E7 protein of human papillomavirus were expressed in a lens-specific fashion in transgenic mice. Lens fiber cells in the transgenic mice were found to incorporate bromodeoxyuridine, implying inappropriate entry into the cell cycle. Surprisingly, the lens fiber cells did not proliferate as tumor cells but instead underwent programmed cell death, resulting in lens ablation and microphthalmia. Analogous lens alterations did not occur in mice expressing a modified version of the truncated T antigen that was mutated in the binding domain for the pRb family. These experimental results indicate that the retinoblastoma protein family plays a crucial role in blocking cell cycle progression and maintaining terminal differentiation in lens fiber cells. Apoptotic cell death ensues when fiber cells are induced to remain in or reenter the cell cycle.

Vertebrate retinal ganglion cells are selected from competent progenitors by the action of Notch

Development ◽  
1995 ◽  
Vol 121 (11) ◽  
pp. 3637-3650 ◽  
Author(s):  
C.P. Austin ◽  
D.E. Feldman ◽  
J.A. Ida ◽  
C.L. Cepko
Keyword(s):  
Cell Fate ◽  
Ganglion Cells ◽  
Notch Pathway ◽  
Cell Types ◽  
Projection Neurons ◽  
Specific Cell ◽  
Vitro Assay ◽  
Notch Activity

The first cells generated during development of the vertebrate retina are the ganglion cells, the projection neurons of the retina. Although they are one of the most intensively studied cell types within the central nervous system, little is known of the mechanisms that determine ganglion cell fate. We demonstrate that ganglion cells are selected from a large group of competent progenitors that comprise the majority of the early embryonic retina and that differentiation within this group is regulated by Notch. Notch activity in vivo was diminished using antisense oligonucleotides or augmented using a retrovirally transduced constitutively active allele of Notch. The number of ganglion cells produced was inversely related to the level of Notch activity. In addition, the Notch ligand Delta inhibited retinal progenitors from differentiating as ganglion cells to the same degree as did activated Notch in an in vitro assay. These results suggest a conserved strategy for neurogenesis in the retina and describe a versatile in vitro and in vivo system with which to examine the action of the Notch pathway in a specific cell fate decision in a vertebrate.

scholarly journals Glucocorticoid agonists enhance retinal stem cell self-renewal and proliferation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Kenneth N. Grisé ◽  
Nelson X. Bautista ◽  
Krystal Jacques ◽  
Brenda L. K. Coles ◽  
Derek van der Kooy
Keyword(s):  
Cell Fate ◽  
Progenitor Cell ◽  
Cell Types ◽  
Retinal Cell ◽  
Ciliary Epithelium ◽  
Cell Transplant ◽  
Self Renewal ◽  
Synthetic Glucocorticoid

Abstract Background Adult mammalian retinal stem cells (RSCs) readily proliferate, self-renew, and generate progeny that differentiate into all retinal cell types in vitro. RSC-derived progeny can be induced to differentiate into photoreceptors, making them a potential source for retinal cell transplant therapies. Despite their proliferative propensity in vitro, RSCs in the adult mammalian eye do not proliferate and do not have a regenerative response to injury. Thus, identifying and modulating the mechanisms that regulate RSC proliferation may enhance the capacity to produce RSC-derived progeny in vitro and enable RSC activation in vivo. Methods Here, we used medium-throughput screening to identify small molecules that can expand the number of RSCs and their progeny in culture. In vitro differentiation assays were used to assess the effects of synthetic glucocorticoid agonist dexamethasone on RSC-derived progenitor cell fate. Intravitreal injections of dexamethasone into adult mouse eyes were used to investigate the effects on endogenous RSCs. Results We discovered that high-affinity synthetic glucocorticoid agonists increase RSC self-renewal and increase retinal progenitor proliferation up to 6-fold without influencing their differentiation in vitro. Intravitreal injection of synthetic glucocorticoid agonist dexamethasone induced in vivo proliferation in the ciliary epithelium—the niche in which adult RSCs reside. Conclusions Together, our results identify glucocorticoids as novel regulators of retinal stem and progenitor cell proliferation in culture and provide evidence that GCs may activate endogenous RSCs.

scholarly journals Single-Cell Genomics: Catalyst for Cell Fate Engineering

2021 ◽  
Vol 9 ◽  
Author(s):  
Boxun Li ◽  
Gary C. Hon
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Cellular Heterogeneity ◽  
State Transitions ◽  
Small Scale ◽  
Specific Cell ◽  
Direct Reprogramming ◽  
Cell State

As we near a complete catalog of mammalian cell types, the capability to engineer specific cell types on demand would transform biomedical research and regenerative medicine. However, the current pace of discovering new cell types far outstrips our ability to engineer them. One attractive strategy for cellular engineering is direct reprogramming, where induction of specific transcription factor (TF) cocktails orchestrates cell state transitions. Here, we review the foundational studies of TF-mediated reprogramming in the context of a general framework for cell fate engineering, which consists of: discovering new reprogramming cocktails, assessing engineered cells, and revealing molecular mechanisms. Traditional bulk reprogramming methods established a strong foundation for TF-mediated reprogramming, but were limited by their small scale and difficulty resolving cellular heterogeneity. Recently, single-cell technologies have overcome these challenges to rapidly accelerate progress in cell fate engineering. In the next decade, we anticipate that these tools will enable unprecedented control of cell state.

scholarly journals A multiscale model of epigenetic heterogeneity reveals the kinetic routes of pathological cell fate reprogramming

2018 ◽  
Author(s):  
Núria Folguera-Blasco ◽  
Rubén Pérez-Carrasco ◽  
Elisabet Cuyás ◽  
Javier A. Menendez ◽  
Tomás Alarcón
Keyword(s):  
Stochastic Model ◽  
Cell Fate ◽  
Multiple Scales ◽  
Coupled System ◽  
Multiscale Model ◽  
Cell Types ◽  
Specific Cell ◽  
New Therapeutic Approaches ◽  
Correct Path

AbstractThe inherent capacity of somatic cells to switch their phenotypic status in response to damage stimuli in vivo might have a pivotal role in ageing and cancer. However, how the entry-exit mechanisms of phenotype reprogramming are established remains poorly understood. In an attempt to elucidate such mechanisms, we herein introduce a stochastic model of combined epigenetic regulation (ER)-gene regulatory network (GRN) to study the plastic phenotypic behaviours driven by ER heterogeneity. Furthermore, based on the existence of multiple scales, we formulate a method for stochastic model reduction, from which we derive an efficient hybrid simulation scheme that allows us to deal with such complex systems. Our analysis of the coupled system reveals a regime of tristability in which pluripotent stem-like and differentiated steady-states coexist with a third indecisive state. Crucially, ER heterogeneity of differentiation genes is for the most part responsible for conferring abnormal robustness to pluripotent stem-like states. We then formulate epigenetic heterogeneity-based strategies capable of unlocking and facilitating the transit from differentiation-refractory (pluripotent stem-like) to differentiation-primed epistates. The application of the hybrid numerical method validated the likelihood of such switching involving solely kinetic changes in epigenetic factors. Our results suggest that epigenetic heterogeneity regulates the mechanisms and kinetics of phenotypic robustness of cell fate reprogramming. The occurrence of tunable switches capable of modifying the nature of cell fate reprogramming from pathological to physiological might pave the way for new therapeutic strategies to regulate reparative reprogramming in ageing and cancer.Author summaryCertain modifications of the structure and functioning of the protein/DNA complex called chromatin can allow adult, fully differentiated cells to adopt a stem cell-like pluripotent state in a purely epigenetic manner, not involving changes in the underlying DNA sequence. Such reprogramming-like phenomena may constitute an innate reparative route through which human tissues respond to injury and could also serve as a novel regenerative strategy in human pathological situations in which tissue or organ repair is impaired. However, it should be noted that in vivo reprogramming would be capable of maintaining tissue homeostasis provided the acquisition of pluripotency features is strictly transient and accompanied by an accurate replenishment of the specific cell types being lost. Crucially, an excessive reprogramming to pluripotency in the absence of controlled re-differentiation would impair the repair or the replacement of damaged cells, thereby promoting pathological alterations of cell fate. A mechanistic understanding of how the degree of chromatin plasticity dictates the reparative versus pathological behaviour of in vivo reprogramming to rejuvenate aged tissues while preventing tumorigenesis is urgently needed, including especially the intrinsic epigenetic heterogeneity of the tissue resident cells being reprogrammed. We here introduce a novel method that mathematically captures how epigenetic heterogeneity is actually the driving force that governs the routes and kinetics to entry into and exit from a pathological pluripotent-like state. Moreover, our approach computationally validates the likelihood of unlocking chronic, unrestrained pluripotent states and drive their differentiation down the correct path by solely manipulating the intensity and direction of few epigenetic control switches. Our approach could inspire new therapeutic approaches based on in vivo cell reprogramming for efficient tissue regeneration and rejuvenation and cancer treatment.

Genetic ablation in transgenic mice with an attenuated diphtheria toxin A gene

1990 ◽  
Vol 10 (2) ◽  
pp. 474-479
Author(s):  
M L Breitman ◽  
H Rombola ◽  
I H Maxwell ◽  
G K Klintworth ◽  
A Bernstein
Keyword(s):  
Transgenic Mice ◽  
Diphtheria Toxin ◽  
Transgenic Animals ◽  
Regulatory Sequences ◽  
Animal Cells ◽  
Toxin A ◽  
Tissue Specific ◽  
Genetic Ablation ◽  
Fiber Cells ◽  
Diphtheria Toxin A

We have previously generated microphthalmic mice lacking lens fiber cells by targeting the expression of the diphtheria toxin A (DT-A) gene in transgenic mice with regulatory sequences associated with the mouse gamma 2-crystallin gene. Because of the extreme toxicity of DT to animal cells and the potential leakiness of many tissue-specific regulatory regions, we investigated whether there might be an experimental advantage in using a mutant, attenuated form of the DT-A gene (tox-176) fused to gamma 2-crystallin regulatory sequences to ablate fiber cells in the ocular lens. In contrast to the microphthalmia observed in transgenic animals carrying the native DT-A gene, independent lines of mice transgenic for the gamma 2tox176 construct displayed predominantly cataracts or clinical anophthalmia. These contrasting phenotypes were transmitted within each pedigree, although for some lines some phenotypic heterogeneity among offspring was noted. The difference in phenotype between cataractous and clinically anophthalmic transgenic lines could not be ascribed to differences in the transgene copy number. Instead, the results suggest that transgene expression and hence the extent of genetic ablation are modulated by the site of chromosomal integration and, to a lesser extent, by epigenetic events. They also suggest that the attenuated gamma 2tox176 construct can integrate into chromosomal regions that are particularly favorable for expression without compromising embryological development and therefore that the tox-176 gene may be more versatile and effective than the wild-type DT-A gene for achieving genetic ablation with a broad range of cell- or tissue-specific regulatory sequences.

Modifications of cell fate specification in equal-cleaving nemertean embryos: alternate patterns of spiralian development

Development ◽  
1995 ◽  
Vol 121 (10) ◽  
pp. 3175-3185 ◽  
Author(s):  
M.Q. Martindale ◽  
J.Q. Henry
Keyword(s):  
Cell Fate ◽  
Cell Lineage ◽  
Cell Types ◽  
Embryonic Cell ◽  
Specific Cell ◽  
Cell Fate Specification ◽  
Cell Fates ◽  
Developmental Potential ◽  
Fate Specification ◽  
Symmetry Properties

The nemerteans belong to a phylum of coelomate worms that display a highly conserved pattern of cell divisions referred to as spiral cleavage. It has recently been shown that the fates of the four embryonic cell quadrants in two species of nemerteans are not homologous to those in other spiralian embryos, such as the annelids and molluscs (Henry, J. Q. and Martindale, M. Q. (1994a) Develop. Genetics 15, 64–78). Equal-cleaving molluscs utilize inductive interactions to establish quadrant-specific cell fates and embryonic symmetry properties following fifth cleavage. In order to elucidate the manner in which cell fates are established in nemertean embryos, we have conducted cell isolation and deletion experiments to examine the developmental potential of the early cleavage blastomeres of two equal-cleaving nemerteans, Nemertopsis bivittata and Cerebratulus lacteus. These two species display different modes of development: N. bivittata develops directly via a non-feeding larvae, while C. lacteus develops to form a feeding pilidium larva which undergoes a radical metamorphosis to give rise to the juvenile worm. By examining the development of certain structures and cell types characteristic of quadrant-specific fates for each of these species, we have shown that isolated blastomeres of the indirect-developing nemertean, C. lacteus, are capable of generating cell fates that are not a consequence of that cell's normal developmental program. For instance, dorsal blastomeres can form muscle fibers when cultured in isolation. In contrast, isolated blastomeres of the direct-developing species, N. bivittata do not regulate their development to the same extent. Some cell fates are specified in a precocious manner in this species, such as those that give rise to the eyes. Thus, these findings indicate that equal-cleaving spiralian embryos can utilize different mechanisms of cell fate and axis specification. The implications of these patterns of nemertean development are discussed in relation to experimental work in other spiralian embryos, and a model is presented that accounts for possible evolutionary changes in cell lineage and the process of cell fate specification amongst these protostome phyla.

scholarly journals Controlling embryonic stem cell proliferation and pluripotency: the role of PI3K- and GSK-3-dependent signalling

2011 ◽  
Vol 39 (2) ◽  
pp. 674-678 ◽  
Author(s):  
Melanie J. Welham ◽  
Emmajayne Kingham ◽  
Yolanda Sanchez-Ripoll ◽  
Benjamin Kumpfmueller ◽  
Michael Storm ◽  
...  
Keyword(s):  
Cell Fate ◽  
Molecular Mechanisms ◽  
Inner Cell Mass ◽  
Glycogen Synthase ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Cell Types ◽  
Specific Cell ◽  
Human Escs ◽  
Phosphoinositide 3 Kinase

ESCs (embryonic stem cells) are derived from the inner cell mass of pre-implantation embryos and are pluripotent, meaning they can differentiate into all of the cells that make up the adult organism. This property of pluripotency makes ESCs attractive as a model system for studying early development and for the generation of specific cell types for use in regenerative medicine and drug screening. In order to harness their potential, the molecular mechanisms regulating ESC pluripotency, proliferation and differentiation (i.e. cell fate) need to be understood so that pluripotency can be maintained during expansion, while differentiation to specific lineages can be induced accurately when required. The present review focuses on the potential roles that PI3K (phosphoinositide 3-kinase) and GSK-3 (glycogen synthase kinase 3)-dependent signalling play in the co-ordination and integration of mouse ESC pluripotency and proliferation and contrast this with our understanding of their functions in human ESCs.

scholarly journals Developing defined substrates for stem cell culture and differentiation

2018 ◽  
Vol 373 (1750) ◽  
pp. 20170230 ◽  
Author(s):  
Louise Hagbard ◽  
Katherine Cameron ◽  
Paul August ◽  
Christopher Penton ◽  
Malin Parmar ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Stem Cell ◽  
Cell Fate ◽  
Cell Types ◽  
Specific Cell ◽  
Biologically Relevant ◽  
Stem Cell Maintenance ◽  
Signalling Molecules ◽  
Cell Matrix

Over the past few decades, a variety of different reagents for stem cell maintenance and differentiation have been commercialized. These reagents share a common goal in facilitating the manufacture of products suitable for cell therapy while reducing the amount of non-defined components. Lessons from developmental biology have identified signalling molecules that can guide the differentiation process in vitro , but less attention has been paid to the extracellular matrix used. With the introduction of more biologically relevant and defined matrices, that better mimic specific cell niches, researchers now have powerful resources to fine-tune their in vitro differentiation systems, which may allow the manufacture of therapeutically relevant cell types. In this review article, we revisit the basics of the extracellular matrix, and explore the important role of the cell–matrix interaction. We focus on laminin proteins because they help to maintain pluripotency and drive cell fate specification. This article is part of the theme issue ‘Designer human tissue: coming to a lab near you’.

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