Biomedical subjects

Louise E Reynolds

Publications and source records attributed to Louise E Reynolds.

7 recordsLinked to original sources

PubMed2006

Primary mouse endothelial cell culture for assays of angiogenesis.

The growth of new blood vessels, known as angiogenesis, is a dynamic but highly regulated process involving many different regulatory pathways. Endothelial cell migration and proliferation is also essential for this process to occur. Studying the behavior of endothelial cells and how they repond to the factors involved in angiogenesis is important in understanding how the process is controlled. The ability to generatic primary endothelial cells allows for such detailed studies. In this chapter, we present the method for endothelial cell isolation that we have found to be successful and that we routinely employ to isolate and culture primary endothelial cells.

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PubMed2005

Bone marrow cells engraft within the epidermis and proliferate in vivo with no evidence of cell fusion.

In adults, bone marrow-derived cells (BMDC) can contribute to the structure of various non-haematopoietic tissues, including skin. However, the physiological importance of these cells is unclear. This study establishes that bone marrow-derived epidermal cells are proliferative and, moreover, demonstrates for the first time that BMDC can localize to a known stem cell niche: the CD34-positive bulge region of mouse hair follicles. In addition, engraftment of bone marrow cells into the epidermis is significantly increased in wounded skin, bone marrow-derived keratinocytes can form colonies in the regenerating epidermis in vivo, and the colony-forming capacity of these cells can be recapitulated in vitro. In some tissues this apparent plasticity is attributed to differentiation, and in others to cell fusion. Evidence is also provided that bone marrow cells form epidermal keratinocytes without undergoing cell fusion. These data suggest a functional role for bone marrow cells in epidermal regeneration, entering known epidermal stem cell niches without heterokaryon formation.

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PubMed2004

Elevated Flk1 (vascular endothelial growth factor receptor 2) signaling mediates enhanced angiogenesis in beta3-integrin-deficient mice.

Tumor growth, tumor angiogenesis, and vascular endothelial growth factor (VEGF)-specific angiogenesis are all enhanced in beta(3)-integrin-null mice. Furthermore, endothelial cells isolated from beta(3)-null mice show elevated levels of Flk1 (VEGF receptor 2) expression, suggesting that beta(3)-integrin can control the amplitude of VEGF responses by controlling Flk1 levels or activity. We now show that Flk1 signaling is required for the enhanced tumor growth and angiogenesis seen in beta(3)-null mice. Moreover, beta(3)-null endothelial cells exhibit enhanced migration and proliferation in response to VEGF in vitro, and this phenotype requires Flk1 signaling. Upon VEGF stimulation, beta(3)-null endothelial cells exhibit higher levels of phosphorylated Flk1 and extracellular-related kinases 1 and 2 than wild-type endothelial cells. Furthermore, signaling via ERK1/2 is required to mediate the elevated responses to VEGF observed in beta(3)-null endothelial cells and aortic rings in vitro. These data confirm that VEGF signaling via Flk1 is enhanced in beta(3)-integrin-deficient mice and suggests that this increase may mediate the enhanced angiogenesis and tumor growth observed in these mice in vivo.

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PubMed2004

Beta3-integrin regulates vascular endothelial growth factor-A-dependent permeability.

OBJECTIVE: Beta3-integrin deficiency has been implicated in increasing levels of Flk-1 expression on endothelial cells and enhancing vascular endothelial growth factor (VEGF)-induced angiogenesis. We determined the role of beta3-integrin in mediating VEGF-A-induced blood vessel permeability through Flk-1. METHODS AND RESULTS: Using the Miles assay, we demonstrated that VEGF-A-induced plasma leakage was enhanced in beta3-null mice when compared with wild-type controls. This was not caused by any changes in blood vessel structure (as detected by light or electron microscopy) or by changes in endothelial cell-cell adhesion proteins (as determined by Western blot analysis, flow cytometry, and immunofluorescence). Circulating levels of VEGF, baseline blood vessel leakage, and leakage in response to an acute inflammatory stimulus were identical in wild-type and beta3-null mice. However, VEGF-A-induced leakage was abolished in beta3-null mice by the inhibition of Flk-1, indicating that the elevated levels of Flk-1 on beta3-null endothelial cells enhance VEGF-A-induced permeability. CONCLUSIONS: beta3-integrin-deficiency increases the sensitivity of endothelial cells to VEGF-A by elevating Flk-1 expression and, as a consequence, enhances VEGF-A-mediated permeability.

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PubMed2003

Integrins in angiogenesis: multitalented molecules in a balancing act.

Over the last 10-15 years the varied roles of cell adhesion molecules in the development of new blood vessels have received extensive attention. To date, more than 500 publications have been dedicated specifically to the role of a single family of adhesion molecules, namely integrins, in the process of angiogenesis. Although one can now appreciate the involvement of integrins in this process, and indeed antagonists of integrins are presently being tested as anti-angiogenic treatments, the precise regulation and exact action of integrins is still unclear. Here we will clarify the varied role of integrins and aim to elucidate and simplify the combined functions of these molecules in angiogenesis.

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PubMed2003

Desmosomal proteins, including desmoglein 3, serve as novel negative markers for epidermal stem cell-containing population of keratinocytes.

No single method has been universally adopted for identifying and isolating epidermal stem/progenitor cells, and the emergence of new markers of stem cell populations is worth exploring. Here we report, for the first time, that clusters of basal keratinocytes at the tips of the rete ridges in human palm, previously recognised as a major repository of stem cells, had very low levels of desmoplakin protein and mRNA expression, compared with cells at the sides of the ridges or above the dermal papillae. We found that in populations of palm keratinocytes, selected by their ability to adhere rapidly to type IV collagen, there were significantly reduced levels of desmoplakin and other major desmosome proteins. We then showed that a low desmoglein 3 (Dsg3) expression on the cell surface could be used to enrich for a cell population with high clonogenecity, colony forming efficiency and enhanced proliferative potential, but with a low ability to form the abortive clones, compared with populations with a higher Dsg3 expression. Moreover, stringent sorting of populations showing both beta1 integrin-bright and Dsg3-dull expression enabled even further enrichment of a population containing the putative epidermal stem cells. These findings provide the basis for a new strategy for epidermal stem/progenitor cell enrichment, and encourage further study of the role of desmosomes in stem cell biology.

Cadherins↗

PubMed2002

Enhanced pathological angiogenesis in mice lacking beta3 integrin or beta3 and beta5 integrins.

Inhibition of alphavbeta3 or alphavbeta5 integrin function has been reported to suppress neovascularization and tumor growth, suggesting that these integrins are critical modulators of angiogenesis. Here we report that mice lacking beta3 integrins or both beta3 and beta5 integrins not only support tumorigenesis, but have enhanced tumor growth as well. Moreover, the tumors in these integrin-deficient mice display enhanced angiogenesis, strongly suggesting that neither beta3 nor beta5 integrins are essential for neovascularization. We also observed that angiogenic responses to hypoxia and vascular endothelial growth factor (VEGF) are augmented significantly in the absence of beta3 integrins. We found no evidence that the expression or functions of other integrins were altered as a consequence of the beta3 deficiency, but we did observe elevated levels of VEGF receptor-2 (also called Flk-1) in beta3-null endothelial cells. These data indicate that alphavbeta3 and alphavbeta5 integrins are not essential for vascular development or pathological angiogenesis and highlight the need for further evaluation of the mechanisms of action of alphav-integrin antagonists in anti-angiogenic therapeutics.

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