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Towards a dermal papilla specific expression system in mice

Eriksson, Daniel (2013) MOBM15 20131
Degree Projects in Molecular Biology
Abstract
Abstract

Stem cells hold great promise for regenerative medicine. In order to realize the potential of stem cell-based therapies, it is highly required to understand the basic mechanisms of stem cell regulation and transform such knowledge into medical applications. However, understanding stem cell regulations is still challenging due to technical difficulties in studying stem cell biology. The hair follicle is a continuously regenerating mini-organ, in which keratinocyte and melanocyte stem cells cooperate to achieve periodical cycles of pigmented hair formation. The mouse hair follicle provides an ideal model by which to investigate molecular basis of stem cell regulation as defective molecular regulation results in abnormal hair... (More)
Abstract

Stem cells hold great promise for regenerative medicine. In order to realize the potential of stem cell-based therapies, it is highly required to understand the basic mechanisms of stem cell regulation and transform such knowledge into medical applications. However, understanding stem cell regulations is still challenging due to technical difficulties in studying stem cell biology. The hair follicle is a continuously regenerating mini-organ, in which keratinocyte and melanocyte stem cells cooperate to achieve periodical cycles of pigmented hair formation. The mouse hair follicle provides an ideal model by which to investigate molecular basis of stem cell regulation as defective molecular regulation results in abnormal hair formation, a phenotype easily recognized by appearance. It has been widely appreciated that the regulation of these stem cells is mediated through the epithelial-mesenchymal interactions where a cluster of mesenchymal cells, known as the dermal papilla, plays a key role. Here we aim to establish a system manipulating genes in dermal papilla to identify genes involved in the stem cell regulation. To this end, the purpose of this study is to identify a promoter element permitting stable and specific dermal papilla transgene expression in mice. In this report, by integrating BAC transgenesis and transposon-meditated gene delivery systems, we establish a simple pipeline to identify a gene expression element enabling dermal papilla-specific genetic modifications. (Less)
Abstract
Popular science summary:

Healing: what we can learn from white mice

Stem cells are a cell type responsible for building the body. They are the ancestor cells from which all the cells making up the body’s tissues are created. Stem cells play an important role in maintaining the body in shape by repairing damages as they occur during life.
We believe that if we had a better understanding of how these stem cells signal and talk within the body we could persuade them to heal more damages than they normally do. This could have enormous potential to help many people suffering from degenerative diseases such as Parkinson disease, Alzheimer disease, or muscular dystrophy to return to a normal life.

The stem cells building mouse fur
... (More)
Popular science summary:

Healing: what we can learn from white mice

Stem cells are a cell type responsible for building the body. They are the ancestor cells from which all the cells making up the body’s tissues are created. Stem cells play an important role in maintaining the body in shape by repairing damages as they occur during life.
We believe that if we had a better understanding of how these stem cells signal and talk within the body we could persuade them to heal more damages than they normally do. This could have enormous potential to help many people suffering from degenerative diseases such as Parkinson disease, Alzheimer disease, or muscular dystrophy to return to a normal life.

The stem cells building mouse fur
In mice the fur is built by two types of stem cells inside the mouse hair follicle. They are called keratinocytes and melanocytes. The keratinocytes build the hair and the melanocytes provide it with color. These stem cells take orders from other key operator cells called the dermal papilla, located inside the follicle, which instructs the stem cells when to make and when to shed the hairs. Thus, the dermal papilla controls the hair growth and color.
This gives potential to the dermal papilla to become an attractive model system to investigate which molecules tell stem cells to do what. If one important molecule is missing, the stem cells would no longer hear that signal and stop working. If such a molecule would be mutated in mice they would become either hairless or white which is easily recognized by strange hair coat appearance. Such characteristics of the dermal papilla can be utilized to quickly screen many molecules to identify important ones for the stem cell regulation.
To do this we need to find a DNA element which is only used in the dermal papilla so that we can remove signaling molecules without changing a lot of other things around the mouse. To find it we made a huge DNA construct called bacterial artificial chromosome (BAC) carrying a fluorescent protein gene called GFP under the control of a promoter we believe to be used only inside the dermal papilla. Cells using this promoter would then glow in green.
To make the construct work, it needs to be inserted into the host genome. The GFP alone is very small and is very easily inserted into the host genome. However, the larger construct that we constructed needed a little help getting into the genome. Therefore we used a special protein called transposase which is an enzyme that cuts DNA from one place and pastes it in another. To test if this approach worked, we confirmed that our GFP construct could be inserted into the genome of human cells. Next we need to insert it into the mouse genome. This would allow us to see what parts of the mouse are glowing and if only the dermal papilla is green we could use our system to learn more about stem cell signalling.

Supervisor: Masatake Osawa
Master´s Degree Project - Cell and Molecular Biology 30 credits 2013
Department of Biology, Lund University,
Department of Molecular Design and Synthesis, Regeneration Technology,
Regeneration and Advanced Medical Sciences, Graduate School of Medicine Gifu Universit (Less)
Please use this url to cite or link to this publication:
author
Eriksson, Daniel
supervisor
organization
course
MOBM15 20131
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
4255033
date added to LUP
2014-02-03 13:33:42
date last changed
2014-02-03 13:33:42
@misc{4255033,
  abstract     = {Popular science summary:

Healing: what we can learn from white mice

Stem cells are a cell type responsible for building the body. They are the ancestor cells from which all the cells making up the body’s tissues are created. Stem cells play an important role in maintaining the body in shape by repairing damages as they occur during life. 
 We believe that if we had a better understanding of how these stem cells signal and talk within the body we could persuade them to heal more damages than they normally do. This could have enormous potential to help many people suffering from degenerative diseases such as Parkinson disease, Alzheimer disease, or muscular dystrophy to return to a normal life. 

The stem cells building mouse fur
 In mice the fur is built by two types of stem cells inside the mouse hair follicle. They are called keratinocytes and melanocytes. The keratinocytes build the hair and the melanocytes provide it with color. These stem cells take orders from other key operator cells called the dermal papilla, located inside the follicle, which instructs the stem cells when to make and when to shed the hairs. Thus, the dermal papilla controls the hair growth and color. 
 This gives potential to the dermal papilla to become an attractive model system to investigate which molecules tell stem cells to do what. If one important molecule is missing, the stem cells would no longer hear that signal and stop working. If such a molecule would be mutated in mice they would become either hairless or white which is easily recognized by strange hair coat appearance. Such characteristics of the dermal papilla can be utilized to quickly screen many molecules to identify important ones for the stem cell regulation. 
 To do this we need to find a DNA element which is only used in the dermal papilla so that we can remove signaling molecules without changing a lot of other things around the mouse. To find it we made a huge DNA construct called bacterial artificial chromosome (BAC) carrying a fluorescent protein gene called GFP under the control of a promoter we believe to be used only inside the dermal papilla. Cells using this promoter would then glow in green.
 To make the construct work, it needs to be inserted into the host genome. The GFP alone is very small and is very easily inserted into the host genome. However, the larger construct that we constructed needed a little help getting into the genome. Therefore we used a special protein called transposase which is an enzyme that cuts DNA from one place and pastes it in another. To test if this approach worked, we confirmed that our GFP construct could be inserted into the genome of human cells. Next we need to insert it into the mouse genome. This would allow us to see what parts of the mouse are glowing and if only the dermal papilla is green we could use our system to learn more about stem cell signalling.

Supervisor: Masatake Osawa
Master´s Degree Project - Cell and Molecular Biology 30 credits 2013
Department of Biology, Lund University,
Department of Molecular Design and Synthesis, Regeneration Technology,
Regeneration and Advanced Medical Sciences, Graduate School of Medicine Gifu Universit},
  author       = {Eriksson, Daniel},
  language     = {eng},
  note         = {Student Paper},
  title        = {Towards a dermal papilla specific expression system in mice},
  year         = {2013},
}