Hutter, Caroline; (2002) Role of β1 integrin in epidermal development and homeostasis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Integrins are a ubiquitously expressed family of cell surface receptors that mediate cell-matrix and in some cases also cell-cell adhesion. In addition to just anchoring the cell to its surroundings, they play a crucial role in tissue morphogenesis and maintenance. In the epidermis β1 integrins have been implicated in regulating keratinocyte proliferation and differentiation. The aim of the work presented in this thesis is to elucidate the role of β1 integrins in the epidermis by exploiting cells that lack the β1 subunit. Since deletion of the β1 integrin gene in mice causes embryonic lethality before the development of the epidermis, I have taken two experimental approaches: Differentiation of β1-null embryonic stem (ES) cells into keratinocytes in vitro and generation of β1-null keratinocytes via the Cre/Lox system. ES cells that are homozygous null for the β1 integrin subunit fail to differentiate into keratinocytes in vitro but do differentiate in teratomas and wild-type/β1-null chimeric mice. I found that the impaired differentiation is due to a reduced sensitivity of β1-null ES cells to soluble growth factors in comparison to their wild-type counterparts. I showed that β1-null ES cells can be partially rescued by factors that are secreted by dermal fibroblasts. I could furthermore demonstrate, that TGFα, FGF10 and KGF had an inductive effect on keratinocyte differentiation in vitro. By isolating keratinocytes from mice with a floxed β1 integrin gene that expressed Cre under the keratin 5 promoter and by expressing EGFP-Cre in fl/fl keratinocytes using retoviral infection, I could study the behaviour of β1-null keratinocytes in vitro. I showed that these cells have a reduced ability to adhere to various extracellular matrices and that they are not able to spread and to migrate due to a failure to organize the actin cytoskeleton and form focal adhesions. In addition, I demonstrated that β1-null keratinocytes have a lower proliferative index and start to express involucrin, a marker for terminal differentiation. Taken together, these data confirm the essential role of integrins in the epidermis and demonstrate that the loss of β1 integrins cannot be compensated for by upregulation of other integrins or signalling pathways.

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