Red fluorescent Xenopus laevis: a new tool for grafting analysis

Waldner, Christoph; Roose, Magdalena; Ryffel, Gerhart U.

doi:10.1186/1471-213x-9-37

Cited by 24 publications

(19 citation statements)

References 31 publications

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“…4B right; 4C); in 29.5% of Gli1 -injected embryos (52.4% of which formed ITLSs); and in 25.3% of Kras G12D -injected embryos (51.2% of which formed ITLSs) (Table 1). Although a small percentage of embryos displayed depolarized foci, no ITLSs were observed in uninjected controls or in those injected with control mRNAs encoding fluorescent proteins (tdTomato or GFP3) (Zernicka-Goetz et al, 1996; Waldner et al, 2009) (Fig. 4C).…”

Section: Resultsmentioning

confidence: 93%

Transmembrane voltage potential is an essential cellular parameter for the detection and control of tumor development in aXenopusmodel

Chernet

Levin

2013

Disease Models &Amp; Mechanisms

140

177

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SUMMARYUnderstanding mechanisms that orchestrate cell behavior into appropriately patterned tissues and organs within the organism is an essential element of preventing, detecting and treating cancer. Bioelectric signals (resting transmembrane voltage potential gradients in all cells) underlie an important and broadly conserved set of control mechanisms that regulate pattern formation. We tested the role of transmembrane potential in tumorigenesis mediated by canonical oncogenes in Xenopus laevis. Depolarized membrane potential (Vmem) was a characteristic of induced tumor-like structures (ITLSs) generated by overexpression of Gli1, KrasG12D, Xrel3 or p53Trp248. This bioelectric signature was also present in precursor ITLS sites. Vmem is a bioelectric marker that reveals ITLSs before they become histologically and morphologically apparent. Moreover, voltage was functionally important: overexpression of hyperpolarizing ion transporters caused a return to normal Vmem and significantly reduced ITLS formation in vivo. To characterize the molecular mechanism by which Vmem change regulates ITLS phenotypes, we performed a suppression screen. Vmem hyperpolarization was transduced into downstream events via Vmem-regulated activity of SLC5A8, a sodium-butyrate exchanger previously implicated in human cancer. These data indicate that butyrate, a histone deacetylase (HDAC) inhibitor, might be responsible for transcriptional events that mediate suppression of ITLSs by hyperpolarization. Vmem is a convenient cellular parameter by which tumors induced by human oncogenes can be detected in vivo and represents a new diagnostic modality. Moreover, control of resting membrane potential is functionally involved in the process by which oncogene-bearing cells depart from normal morphogenesis programs to form tumors. Modulation of Vmem levels is a novel and promising strategy for tumor normalization.

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Section: Resultsmentioning

confidence: 93%

Transmembrane voltage potential is an essential cellular parameter for the detection and control of tumor development in aXenopusmodel

Chernet

Levin

2013

Disease Models &Amp; Mechanisms

140

177

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“…It is possible that the pattern of intracellular distribution of TdTom may depend on the cell type. Indeed, the localization of TdTom varies according to the cell type [29], and DsRed2, another type of RFP, shows dotted pattern in cells derived from rhabdomyosarcoma [30].…”

Section: Discussionmentioning

confidence: 99%

Stem Cell-Like Differentiation Potentials of Endometrial Side Population Cells as Revealed by a Newly Developed In Vivo Endometrial Stem Cell Assay

et al. 2012

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BackgroundEndometrial stem/progenitor cells contribute to the cyclical regeneration of human endometrium throughout a woman's reproductive life. Although the candidate cell populations have been extensively studied, no consensus exists regarding which endometrial population represents the stem/progenitor cell fraction in terms of in vivo stem cell activity. We have previously reported that human endometrial side population cells (ESP), but not endometrial main population cells (EMP), exhibit stem cell-like properties, including in vivo reconstitution of endometrium-like tissues when xenotransplanted into immunodeficient mice. The reconstitution efficiency, however, was low presumably because ESP cells alone could not provide a sufficient microenvironment (niche) to support their stem cell activity. The objective of this study was to establish a novel in vivo endometrial stem cell assay employing cell tracking and tissue reconstitution systems and to examine the stem cell properties of ESP through use of this assay.Methodology/Principal FindingsESP and EMP cells isolated from whole endometrial cells were infected with lentivirus to express tandem Tomato (TdTom), a red fluorescent protein. They were mixed with unlabeled whole endometrial cells and then transplanted under the kidney capsule of ovariectomized immunodeficient mice. These mice were treated with estradiol and progesterone for eight weeks and nephrectomized. All of the grafts reconstituted endometrium-like tissues under the kidney capsules. Immunofluorescence revealed that TdTom-positive cells were significantly more abundant in the glandular, stromal, and endothelial cells of the reconstituted endometrium in mice transplanted with TdTom-labeled ESP cells than those with TdTom-labeled EMP cells.Conclusions/SignificanceWe have established a novel in vivo endometrial stem cell assay in which multi-potential differentiation can be identified through cell tracking during in vivo endometrial tissue reconstitution. Using this assay, we demonstrated that ESP cells differentiated into multiple endometrial lineages in the niche provided by whole endometrial cells, indicating that ESP cells are genuine endometrial stem/progenitor cells.

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“…Capped, synthetic tdTomato mRNA (Waldner et al, 2009) was dissolved in water and injected into embryos in 3% Ficoll using standard methods (Sive et al, 2000). Injections were made into both cells of two cell embryos to promote ubiquitous expression.…”

Section: Microinjectionmentioning

confidence: 99%

Ectopic eyes outside the head inXenopustadpoles provide sensory data for light-mediated learning

Blackiston

Levin

2013

Journal of Experimental Biology

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SUMMARYA major roadblock in the biomedical treatment of human sensory disorders, including blindness, has been an incomplete understanding of the nervous system and its ability to adapt to changes in sensory modality. Likewise, fundamental insight into the evolvability of complex functional anatomies requires understanding brain plasticity and the interaction between the nervous system and body architecture. While advances have been made in the generation of artificial and biological replacement components, the brainʼs ability to interpret sensory information arising from ectopic locations is not well understood. We report the use of eye primordia grafts to create ectopic eyes along the body axis of Xenopus tadpoles. These eyes are morphologically identical to native eyes and can be induced at caudal locations. Cell labeling studies reveal that eyes created in the tail send projections to the stomach and trunk. To assess function we performed light-mediated learning assays using an automated machine vision and environmental control system. The results demonstrate that ectopic eyes in the tail of Xenopus tadpoles could confer vision to the host. Thus ectopic visual organs were functional even when present at posterior locations. These data and protocols demonstrate the ability of vertebrate brains to interpret sensory input from ectopic structures and incorporate them into adaptive behavioral programs. This tractable new model for understanding the robust plasticity of the central nervous system has significant implications for regenerative medicine and sensory augmentation technology.

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Red fluorescent Xenopus laevis: a new tool for grafting analysis

Cited by 24 publications

References 31 publications

Transmembrane voltage potential is an essential cellular parameter for the detection and control of tumor development in aXenopusmodel

Transmembrane voltage potential is an essential cellular parameter for the detection and control of tumor development in aXenopusmodel

Stem Cell-Like Differentiation Potentials of Endometrial Side Population Cells as Revealed by a Newly Developed In Vivo Endometrial Stem Cell Assay

Ectopic eyes outside the head inXenopustadpoles provide sensory data for light-mediated learning

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