Thrombospondin-1 up-regulates expression of cell adhesion molecules and promotes monocyte binding to endothelium.

Expression of cell adhesion molecules (CAM) responsible for leukocyte-endothelium interactions plays a crucial role in inflammation and atherogenesis. Up-regulation of vascular CAM-1 (VCAM-1), intracellular CAM-1 (ICAM-1), and E-selectin expression promotes monocyte recruitment to sites of injury and is considered to be a critical step in atherosclerotic plaque development. Factors that trigger this initial response are not well understood. As platelet activation not only promotes thrombosis but also early stages of atherogenesis, we considered the role of thrombospondin-1 (TSP-1), a matricellular protein released in abundance from activated platelets and accumulated in sites of vascular injury, as a regulator of CAM expression. TSP-1 induced expression of VCAM-1 and ICAM-1 on endothelium of various origins, which in turn, resulted in a significant increase of monocyte attachment. This effect could be mimicked by a peptide derived from the C-terminal domain of TSP-1 and known to interact with CD47 on the cell surface. The essential role of CD47 in the cellular responses to TSP-1 was demonstrated further using inhibitory antibodies and knockdown of CD47 with small interfering RNA. Furthermore, we demonstrated that secretion of endogenous TSP-1 and its interaction with CD47 on the cell surface mediates endothelial response to the major proinflammatory agent, tumor necrosis factor alpha (TNF-alpha). Taken together, this study identifies a novel mechanism regulating CAM expression and subsequent monocyte binding to endothelium, which might influence the development of anti-atherosclerosis therapeutic strategies.

The Toll-Like Receptor 5 Agonist Entolimod Mitigates Lethal Acute Radiation Syndrome in Non-Human Primates.

There are currently no approved medical radiation countermeasures (MRC) to reduce the lethality of high-dose total body ionizing irradiation expected in nuclear emergencies. An ideal MRC would be effective even when administered well after radiation exposure and would counteract the effects of irradiation on the hematopoietic system and gastrointestinal tract that contribute to its lethality. Entolimod is a Toll-like receptor 5 agonist with demonstrated radioprotective/mitigative activity in rodents and radioprotective activity in non-human primates. Here, we report data from several exploratory studies conducted in lethally irradiated non-human primates (rhesus macaques) treated with a single intramuscular injection of entolimod (in the absence of intensive individualized supportive care) administered in a mitigative regimen, 1-48 hours after irradiation. Following exposure to LD50-70/40 of radiation, injection of efficacious doses of entolimod administered as late as 25 hours thereafter reduced the risk of mortality 2-3-fold, providing a statistically significant (P<0.01) absolute survival advantage of 40-60% compared to vehicle treatment. Similar magnitude of survival improvement was also achieved with drug delivered 48 hours after irradiation. Improved survival was accompanied by predominantly significant (P<0.05) effects of entolimod administration on accelerated morphological recovery of hematopoietic and immune system organs, decreased severity and duration of thrombocytopenia, anemia and neutropenia, and increased clonogenic potential of the bone marrow compared to control irradiated animals. Entolimod treatment also led to reduced apoptosis and accelerated crypt regeneration in the gastrointestinal tract. Together, these data indicate that entolimod is a highly promising potential life-saving treatment for victims of radiation disasters.

Molecular pathway for cancer metastasis to bone.

The molecular mechanism leading to the cancer metastasis to bone is poorly understood but yet determines prognosis and therapy. Here, we define a new molecular pathway that may account for the extraordinarily high osteotropism of prostate cancer. By using SPARC (secreted protein, acidic and rich in cysteine)-deficient mice and recombinant SPARC, we demonstrated that SPARC selectively supports the migration of highly metastatic relative to less metastatic prostate cancer cell lines to bone. Increased migration to SPARC can be traced to the activation of integrins alphaVbeta3 and alphaVbeta5 on tumor cells. Such activation is induced by an autocrine vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR)-2 loop on the tumor cells, which also supports the growth and proliferation of prostate cancer cells. A consequence of SPARC recognition by alphaVbeta5 is enhanced VEGF production. Thus, prostate cancer cells expressing VEGF/VEGFR-2 will activate alphaVbeta3 and alphaVbeta5 on their surface and use these integrins to migrate toward SPARC in bone. Within the bone environment, SPARC engagement of these integrins will stimulate growth of the tumor and further production of VEGF to support neoangiogenesis, thereby favoring the development of the metastatic tumor. Supporting this model, activated integrins were found to colocalize with VEGFR-2 in tissue samples of metastatic prostate tumors from patients.

Molecular and functional differences induced in thrombospondin-1 by the single nucleotide polymorphism associated with the risk of premature, familial myocardial infarction.

A serine (Ser-700) amino acid rather than an asparagine (Asn-700) at residue 700 of thrombospondin-1 has been linked to an increased risk for development of premature, familial heart attacks. We now have identified both functional and structural differences between the Ser-700 and Asn-700 thrombospondin-1 variants. The Ser-700 variant increased the rate and extent of platelet aggregation and showed increased surface expression on platelets compared with the Asn-700 variant. These differences could be ascribed to an enhanced interaction of the Ser-700 variant with fibrinogen on the platelet surface and are consistent with a prothrombotic phenotype in Ser-700 individuals. The Ser-700 variant thrombospondin-1 was conformationally more labile than the Asn-700 variant as demonstrated by increased susceptibility to proteolytic digestion and enhanced susceptibility to unfolding by denaturants. These data suggest a potential molecular and cellular basis for a genetic risk factor associated with early onset myocardial infarction.