Tube-like formation assay was performed for 8 h in VCBM

Tube-like formation assay was performed for 8 h in VCBM. factors. Furthermore, CM treatment of HUVEC for 24 h increased tube formation by ~5.5-fold, while leptin increased tube formation by ~ 80% and VEGF by ~60% at 8 h. The mitogenic and angiogenic effects of both CM were blocked by Aca 1, a peptide ObR antagonist, and by SU1498, which Sevelamer hydrochloride inhibits the VEGF receptor. The best anti-angiogenic and cytostatic effects of Aca1 were obtained with 10 nM and 25 nM, respectively, while for SU1498, the best growth and angiogenic inhibition was observed at 5 M. The combination of 5 M SU1498 and Aca1 at 25 nM (growth inhibition) or at 10 nM (reduction of tube formation) produced superior effects compared with single agent treatments. Conclusions Our data provide the first evidence that LN18 and LN 229 Sevelamer hydrochloride human GBM cells express leptin mRNA and might produce biologically active leptin, which can stimulate tube formation and enhance proliferation of endothelial cells. Furthermore, we demonstrate for the first time that a peptide ObR antagonist inhibits proangiogenic and growth effects of leptin on endothelial cells, and that the pharmacological potential of this compound might be combined with drugs Sevelamer hydrochloride targeting the VEGF pathway. Background Leptin is an adipocyte-derived hormone that plays a major role in the regulation of body weight by inhibiting food intake and stimulating energy expenditure via hypothalamic-mediated effects [1,2]. Besides its anorexigenic function, leptin regulates several physiological processes, including angiogenesis [3-5]. Human endothelium and primary cultures of human endothelial cells express the leptin receptor, ObR [6,7]. em In vitro /em studies exhibited that leptin can stimulate growth and survival of endothelial cells as well as induce their migration and organization into capillary-like tubes [6-9]. em In vivo /em , leptin is able to induce complete angiogenesis in the chick choriallantoic membrane assay [6] and disc angiogenesis system [10] as well as promote neovascularization in corneas of normal, but not ObR-deficient Zucker fa/fa, rats [7] or normal mice [11]. In addition to its own effects, leptin synergizes with vascular endothelial growth factor (VEGF) and basic fibroblastic growth factor (bFGF) in the stimulation of blood vessel growth and vascular permeability [11]. Proangiogenic and mitogenic functions of leptin have been implicated in development and progression of different neoplasms. Multiple studies exhibited that leptin is able Mouse monoclonal to GLP to stimulate survival [12-14], proliferation [15-17], migration and invasiveness [18-22] of several cancer cell types. In addition, leptin might also contribute to tumor neoangiogenesis. Exposure of cancer cells to hypoxic conditions and/or elevated concentrations of growth factors, such as insulin, can activate production of endogenous leptin, raising intratumoral levels of this hormone [23-28]. Proangiogenic effects of leptin can be further potentiated by its ability to upregulate the expression of other angiogenic factors, such as VEGF, bFGF, interleukin 1-, and leukemia inhibitory factor in cancer cells [18,29-31]. New evidence suggests leptin can be involved in the development of brain tumors [13,22,32-35]. Initial work documented the presence of leptin and ObR transcripts in various human intracranial tumors [34]. Other reports exhibited that rat glioma tissues and cell lines express leptin mRNA [33,36], and that in rat C6 cells leptin can increase survival [13,32,33] and enhance migration and invasion of these cells [22]. We recently exhibited that both leptin and ObR proteins are overexpressed in human brain tumors relative to normal brain tissue, and that leptin/ObR expression levels positively correlate with the degree of malignancy. The highest levels of leptin and ObR were found in glioblastoma multiforme (GBM), where both proteins were coexpressed.