Biomedical subjects

Kenneth Ryan

Publications and source records attributed to Kenneth Ryan.

6 recordsLinked to original sources

PubMed2005

Effector and memory CD8+ T cell fate coupled by T-bet and eomesodermin.

Two seemingly unrelated hallmarks of memory CD8(+) T cells are cytokine-driven proliferative renewal after pathogen clearance and a latent effector program in anticipation of rechallenge. Memory CD8(+) T cells and natural killer cells share cytotoxic potential and dependence on the growth factor interleukin 15. We now show that mice with compound mutations of the genes encoding the transcription factors T-bet and eomesodermin were nearly devoid of several lineages dependent on interleukin 15, including memory CD8(+) T cells and mature natural killer cells, and that their cells had defective cytotoxic effector programming. Moreover, T-bet and eomesodermin were responsible for inducing enhanced expression of CD122, the receptor specifying interleukin 15 responsiveness. Therefore, these key transcription factors link the long-term renewal of memory CD8(+) T cells to their characteristic effector potency.

Animals↗

PubMed2005

Eomesodermin is expressed in mouse oocytes and pre-implantation embryos.

T-box genes are a highly conserved family of genes encoding transcription factors, which share a conserved DNA binding domain (the T-box). Appropriate temporal and spatial expression of this gene family is critical for gastrulation and organogenesis in a number of species. The T-box containing gene Eomesodermin was first identified in Xenopus, where it plays a critical role in mesoderm formation. In situ analyses in mice have described the expression patterns of the mouse ortholog of this gene mEomesodermin (mEomes) at the time of implantation and during fetal development. Additional studies involving the disruption of the mEomes gene, have demonstrated an additional role for mEomes in trophoblast formation. However, these analyses did not address the possibility that maternally encoded or pre-blastocyst zygotic transcription of mEomes may also contribute to embryonic development. We show here that mEomes mRNA is present prior to blastocyst formation, and that the protein product of mEomes is associated with nuclear DNA during oocyte development and persistently localizes within all nuclei of the preimplantation embryo until the early blastocyst stage. mEomes protein is associated with the meiotic spindle in the unfertilized egg and with the mitotic spindle at each cell division. Our results are consistent with mEomesodermin having a role in early preimplantation development and inner cell mass formation in addition to its function in the trophoblast lineage.

Animals↗

PubMed2005

Posttranslational control of a cardiac ion channel transgene in vivo: clarithromycin-hMiRP1-Q9E interactions.

The present study investigates a novel gene therapy approach for atrial arrhythmias, using a clarithromycin-responsive ion channel subunit mutation, hMiRP1-Q9E, cloned into an expression plasmid; wild-type expression plasmids encoding human minK-related protein 1 (hMiRP1) were also used as controls. In a series of pig studies, right atrial myocardium was injected at one site with hMiRP1-Q9E plasmid DNA; a separate site in the same right atrium was injected with wild-type plasmid or was sham injected. Two weeks after transfection intravenous clarithromycin administration resulted in a site-specific, dose-dependent prolongation of the repolarization phase of the right atrial epicardial monophasic action potential (MAP) only at the hMiRPQ9E sites, but not at sham or wild-type sites. MAP recordings before clarithromycin administration did not differ between hMiRP1-Q9E and control sites. These studies show that regional control of atrial myocardial repolarization by site-specific transfection with plasmid DNA encoding an antibiotic-responsive ion channel subunit is feasible and, because hMiRP1-Q9E-transfected sites were affected only if clarithromycin was given, provide proof of concept for a posttranslational, controllable gene therapy strategy for atrial arrhythmias.

Action Potentials↗

PubMed2004

Modulation of eomes activity alters the size of the developing heart: implications for in utero cardiac gene therapy.

Congenital heart disease is the most prevalent cause of infant morbidity and mortality in developed countries. The mechanisms responsible for many specific types of congenital cardiac malformations are strongly associated with gene abnormalities. However, at this time no strategies for gene therapy of the various congenital heart malformations have been investigated. In the present studies we focus on Eomesodermin (Eomes), a T-box transcription factor expressed in developing vertebrate mesoderm. Although Eomes is required for early mesodermal patterning and differentiation, the role of Eomes in cardiac development is unknown. In the present studies we demonstrate that Eomes is expressed in the developing heart, with a pronounced myocardial distribution in the Xenopus ventricle during late cardiac development. Using either a conditional dominant-interfering approach (GR-Eomes--engrailed) or an Eomes-activating approach (GR-Eomes-VP16) we demonstrate that manipulating Eomes activity during late cardiac development can either suppress ventricular development (GR-Eomes-enR) or increase ventricular myocardial size (GR-Eomes-VP16). Thus, a potential gene therapy approach for treating both congenital ventricular hypoplasia (e.g., the hypoplastic left heart syndrome) and hypertrophic cardiomyopathy is hypothetically implicit from the present results.

Animals↗

PubMed2003

PTP alpha regulates integrin-stimulated FAK autophosphorylation and cytoskeletal rearrangement in cell spreading and migration.

We investigated the molecular and cellular actions of receptor protein tyrosine phosphatase (PTP) alpha in integrin signaling using immortalized fibroblasts derived from wild-type and PTP alpha-deficient mouse embryos. Defects in PTP alpha-/- migration in a wound healing assay were associated with altered cell shape and focal adhesion kinase (FAK) phosphorylation. The reduced haptotaxis to fibronectin (FN) of PTP alpha-/- cells was increased by expression of active (but not inactive) PTP alpha. Integrin-mediated formation of src-FAK and fyn-FAK complexes was reduced or abolished in PTP alpha-/- cells on FN, concomitant with markedly reduced phosphorylation of FAK at Tyr397. Reintroduction of active (but not inactive) PTP alpha restored FAK Tyr-397 phosphorylation. FN-induced cytoskeletal rearrangement was retarded in PTP alpha-/- cells, with delayed filamentous actin stress fiber assembly and focal adhesion formation. This mimicked the effects of treating wild-type fibroblasts with the src family protein tyrosine kinase (Src-PTK) inhibitor PP2. These results, together with the reduced src/fyn tyrosine kinase activity in PTP alpha-/- fibroblasts (Ponniah et al., 1999; Su et al., 1999), suggest that PTP alpha functions in integrin signaling and cell migration as an Src-PTK activator. Our paper establishes that PTP alpha is required for early integrin-proximal events, acting upstream of FAK to affect the timely and efficient phosphorylation of FAK Tyr-397.

Adenoviridae↗

PubMed2003

T-box genes and cardiac development.

BACKGROUND: T-box genes play roles in vertebrate gastrulation and in later organogenesis. Their existence in all metazoans examined so far indicates that this is an evolutionarily ancient gene family. Drosophila melanogaster has eight T-box genes, whereas Caenorhabditis elegans has 22. Mammals appear to have at least 18 T-box genes, comprising five subfamilies. METHODS: A full range of cytological, developmental, molecular and genetic methodologies have recently been applied to the study of T-box genes. RESULTS: Over the last 5 years, mutations in TBX1 and TBX5 have been implicated in two human disorders with haplo-insufficient cardiovascular phenotypes, DiGeorge/velocardiofacial syndrome and Holt-Oram ("heart-hand") syndrome. Interestingly, the number of T-box gene family members discovered to have cardiac or pharyngeal arch expression domains during vertebrate embryonic development has steadily grown. In addition, various Tbx5 loss-of-function models in organisms as distant as the mouse and zebrafish do indeed phenocopy Holt-Oram syndrome. Finally, the intriguing discovery earlier this year that a T-box gene is expressed in a subset of cardioblasts in D. melanogaster suggests that members of this gene family may have fundamental, conserved roles in cardiovascular pattern formation. CONCLUSIONS: These developments prompted us to review the current understanding of the contribution of T-box genes to cardiovascular morphogenesis.

Abnormalities, Multiple↗