Many cancer research studies have extensively examined the phosphatase and tensin homolog deleted from chromosome 10 (PTEN) pathway. There are only few reports that suggest that PTEN might affect pain; however, there is still a lack of evidence to show the role of PTEN for modulating pain. Here, we report a role for PTEN in a rodent model of neuropathic pain. We found that chronic constriction injury (CCI) surgery in rats could elicit downregulation of spinal PTEN as well as upregulation of phosphorylated PTEN (phospho-PTEN) and phosphorylated mammalian target of rapamycin (phospho-mTOR). After examining such changes in endogenous PTEN in neuropathic rats, we explored the effects of modulating the spinal PTEN pathway on nociceptive behaviors. The normal rats exhibited mechanical allodynia after intrathecal (i.t.) injection of adenovirus-mediated PTEN antisense oligonucleotide (Ad-antisense PTEN). These data indicate the importance of downregulation of spinal PTEN for nociception. Moreover, upregulation of spinal PTEN by i.t. adenovirus-mediated PTEN (Ad-PTEN) significantly prevented CCI-induced development of nociceptive sensitization, thermal hyperalgesia, mechanical allodynia, cold allodynia, and weight-bearing deficits in neuropathic rats. Furthermore, upregulation of spinal PTEN by i.t. Ad-PTEN significantly attenuated CCI-induced microglia and astrocyte activation, upregulation of tumor necrosis factor-α (TNF-α) and phospho-mTOR, and downregulation of PTEN in neuropathic rats 14 days post injury. These findings demonstrate that PTEN plays a key, beneficial role in a rodent model of neuropathic pain.

α-melanocyte-stimulating hormone (α-MSH) has been characterized as a novel angiogenesis inhibitor. The homeostasis of nitric oxide (NO) plays an important role in neovascularization. However, it remains unclear whether α-MSH mitigates angiogenesis through modulation of NO and its signaling pathway. The present study elucidated the function and mechanism of NO signaling in α-MSH-induced angiogenesis inhibition using cultured human umbilical vein endothelial cells (HUVECs), rat aorta rings, and transgenic zebrafish. By Griess reagent assay, it was found α-MSH dose-dependently reduced the NO release in HUVECs. Immunoblotting and immunofluorescence analysis revealed α-MSH potently suppressed endothelial and inducible nitric oxide synthase (eNOS/iNOS) expression, which was accompanied with inhibition of nuclear factor kappa B (NF-κB) activities. Excessive supply of NO donor l-arginine reversed the α-MSH-induced angiogenesis inhibition in vitro and in vivo. By using antibody neutralization and RNA interference, it was delineated that melanocortin-1 receptor (MC1-R) and melanocortin-2 receptor (MC2-R) participated in α-MSH-induced inhibition of NO production and NF-κB/eNOS/iNOS signaling. This was supported by pharmaceutical inhibition of protein kinase A (PKA), the downstream effector of MC-Rs signaling, using H89 abolished the α-MSH-mediated suppression of NO release and eNOS/iNOS protein level. Therefore, α-MSH exerts anti-angiogenic function by perturbing NO bioavailability and eNOS/iNOS expression in endothelial cells.

Background: WA-25 (dihydroaustrasulfone alcohol, a synthetic derivative of marine compound WE-2) suppresses atherosclerosis in rats by reducing neointima formation. Because angiogenesis plays a critical role in the pathogenesis of atherosclerosis, the present study investigated the angiogenic function and mechanism of WA-25. Methods: The angiogenic effect of WA-25 was evaluated using a rat aortic ring assay and transgenic zebrafish models were established using transgenic Tg(fli-1:EGFP)^y1 and Tg(kdrl:mCherry^ci5-fli1a:negfp^y7) zebrafish embryos. In addition, the effect of WA-25 on distinct angiogenic processes, including matrix metalloproteinase (MMP) expression, endothelial cell proliferation and migration, as well as tube formation, was studied using human umbilical vein endothelial cells (HUVECs). The effect of WA-25 on the endothelial vascular endothelial growth factor (VEGF) signaling pathway was elucidated using qRT-PCR, immunoblot analysis, immunofluorescence and flow cytometric analyses. Results: The application of WA-25 perturbed the development of intersegmental vessels in transgenic zebrafish. Moreover, WA-25 potently suppressed microvessel sprouting in organotypic rat aortic rings. Among cultured endothelial cells, WA-25 significantly and dose-dependently inhibited MMP-2/MMP-9 expression, proliferation, migration and tube formation in HUVECs. Mechanistic studies revealed that WA-25 significantly reduced the VEGF release by reducing VEGF expression at the mRNA and protein levels. In addition, WA-25 reduced surface VEGF receptor 2 (VEGFR2/Flk-1) expression by repressing the VEGFR2 mRNA level. Finally, an exogenous VEGF supply partially rescued the WA-25-induced angiogenesis blockage in vitro and in vivo. Conclusions: WA-25 is a potent angiogenesis inhibitor that acts through the down-regulation of VEGF and VEGFR2 in endothelial cells. General Significance: WA-25 may constitute a novel anti-angiogenic drug that acts by targeting endothelial VEGF/VEGFR2 signaling.

Angiogenesis, the process of neovascularization, plays an important role in physiological and pathological conditions. ST104P is a soluble polysulfated-cyclo-tetrachromotropylene compound with anti-viral and anti-thrombotic activities. However, the functions of ST104P in angiogenesis have never been explored. In this study, we investigated the effects of ST104P in angiogenesis in vitro and in vivo. Application of ST104P potently suppressed the microvessels sprouting in aortic rings ex vivo. Furthermore, ST104P treatment significantly disrupted the vessels’ development in transgenic zebrafish in vivo. Above all, repeated administration of ST104P resulted in delayed tumor growth and prolonged the life span of mice bearing Lewis lung carcinoma. Mechanistic studies revealed that ST104P potently inhibited the migration, tube formation and wound closure of human umbilical endothelial cells (HUVECs). Moreover, ST104P treatment inhibited the secretion and expression of matrix metalloproteinase-2 (MMP-2) in a dose-dependent manner. Together, these results suggest that ST104P is a potent angiogenesis inhibitor and may hold potential for treatment of diseases due to excessive angiogenesis including cancer.