Carmen Lilia Sánchez González

Vasoinhibins: endogenous regulators of angiogenesis and vascular function

Vasoinhibins are a family of peptides derived from prolactin, growth hormone and placental lactogen that act on endothelial cells to suppress vasodilation and angiogenesis and to promote apoptosis-mediated vascular regression. Some of the pathways by which vasoinhibins act have now been defined, and recent developments indicate that endogenous vasoinhibins exert tonic and essential actions on blood vessel growth, dilation and regression in vivo. By studying the pathways that can generate vasoinhibins, and the nature of their receptors and key biological mediators, it should be possible to clarify the role of vasoinhibins in controlling vascular function in health and disease.

Elevated vasoinhibins may contribute to endothelial cell dysfunction and low birth weight in preeclampsia

Vasoconstriction and defective placental angiogenesis are key factors in the etiology of preeclampsia. Prolactin levels are elevated in maternal blood throughout pregnancy and the human decidua produces prolactin that is transported to the amniotic fluid. Prolactin is cleaved to yield vasoinhibins, a family of peptides that inhibit angiogenesis and nitric oxide-dependent vasodilation. Here, we conducted a case–control study to measure vasoinhibins in serum, urine, and amniotic fluid obtained from women with severe preeclampsia. We show that all three biological fluids contained significantly higher levels of vasoinhibins in preeclamptic women than in normal pregnant women. Amniotic fluid from preeclamptic women, but not from normal women, inhibited vascular endothelial growth factor-induced endothelial cell proliferation and nitric oxide synthase activity in cultured endothelial cells, and these actions were reversed by antibodies able to neutralize the effects of vasoinhibins. Furthermore, amniotic fluid does not appear to contain neutral prolactin-cleaving proteases, suggesting that vasoinhibins in amniotic fluid are derived from prolactin cleaved within the placenta. Also, cathepsin-D in placental trophoblasts cleaved prolactin to vasoinhibins, and its activity was higher in placental trophoblasts from preeclamptic women than from normal women. Importantly, birth weight of infants in preeclampsia inversely correlated with the extent to which the corresponding AF inhibited endothelial cell proliferation and with its concentration of prolactin+vasoinhibins. These data demonstrate that vasoinhibins are increased in the circulation, urine, and amniotic fluid of preeclamptic women and suggest that these peptides contribute to the endothelial cell dysfunction and compromised birth weight that characterize this disease.

Tryptic amaranth glutelin digests induce endothelial nitric oxide production through inhibition of ACE: Antihypertensive role of amaranth peptides

Amaranth seed proteins have a better balance of essential amino acids than cereals and legumes. In addition, the tryptic hydrolysis of amaranth proteins generates, among other peptides, angiotensin converting enzyme (ACE) inhibitory (ACEi) peptides. ACE converts angiotensin I (Ang I) into Ang II, but is also responsible for the degradation of bradykinin (BK). In contrast to Ang II, BK stimulates vasodilation modulated through endothelial nitric oxide (NO) production. The aim of the present study was to characterize the ACEi activity of amaranth trypsin-digested glutelins (TDGs) and their ability to induce endothelial NO production. An IC(50) value of 200microgml(-1) was measured for TDG inhibition of ACE. TDGs stimulated endothelial NO production in coronary endothelial cells (CEC) by 52% compared to control. The effects of TDGs were comparable to those of BK and Captopril, both used as positive controls of NO production. Consistent with these effects, TDGs induced, in a dose-dependent manner, endothelial NO-dependent vasodilation in isolated rat aortic rings. These results suggest that TDGs induce endothelial NO production and consequent vasodilation through their ACEi activity. Amaranth TDGs have a high potential as a nutraceutical food in prevention of cardiovascular diseases. Further molecular, cellular and physiological studies are currently under way and the results may contribute to a better understanding and control of cardiovascular disorders.

Vasoinhibins: a family of N-terminal prolactin fragments that inhibit angiogenesis and vascular function.

Antiangiogenic molecules derived from prolactin (PRL) are not a single entity, but rather a family of peptides with different molecular masses, all containing the N-terminal region of PRL. Cleavage of PRL by cathepsin-D or by matrix metalloproteases generates N-terminal fragments that act on endothelial cells to suppress vasodilation and angiogenesis and promote vascular regression. N-terminal PRL fragments have been identified in cartilage and retina, where angiogenesis is highly restricted. In vivo experiments demonstrate that these PRL fragments exert a tonic and essential suppression of retinal blood vessel growth and dilation. Similar PRL fragments have been detected in the pituitary gland, a highly vascularized organ where the control of vascular growth may differ from that in tissues where angiogenesis is highly restricted. We have previously proposed the name vasoinhibins to describe the collection of N-terminal PRL fragments having blood vessel-blocking activity, and here we discuss their promise as factors to control vascular function in health and disease.

Prolactin and 16K prolactin stimulate release of vasopressin by a direct effect on hypothalamo-neurohypophyseal system

Activity of the magnocellular neurons that synthesize vasopressin and oxytocin in the paraventricular and supraoptic nuclei of the hypothalamus can be modulated by local release of neuromediators within the nuclei. Among the bioactive peptides that may play autocrine or paracrine roles in this system is prolactin (PRL). Paraventricular and supraoptic neurons express PRL mRNA and contain and secrete PRL-like proteins of 23 and 14 kDa. We investigated the localization of PRL receptors in vasopressinergic and oxytocinergic magnocellular neurons using dual-label immunofluorescence. The results demonstrate that both vasopressin- and oxytocin-immunoreactive cells of the paraventricular and supraoptic nuclei contain the PRL receptor. In addition, we investigated the possible regulation of vasopressin secretion by PRL using hypothalamo-neurohypophyseal explants in culture. The results show that PRL and a 16 kDa N-terminal fragment of the hormone that is analogous to the neurohypophyseal 14-kDa PRL fragment stimulate the release of vasopressin. Together, these findings support the hypothesis that vasopressinergic and oxytocinergic neurons of the magnocellular secretory system are regulated directly by various isoforms of PRL via autocrine/paracrine mechanisms.

Vasoinhibins Prevent Bradykinin-Stimulated Endothelial Cell Proliferation by Inactivating eNOS via Reduction of both Intracellular Ca2+ Levels and eNOS Phosphorylation at Ser1179

Vasoinhibins, a family of antiangiogenic peptides derived from prolactin proteolysis, inhibit the vascular effects of several proangiogenic factors, including bradykinin (BK). Here, we report that vasoinhibins block the BK-induced proliferation of bovine umbilical vein endothelial cells. This effect is mediated by the inactivation of endothelial nitric oxide synthase (eNOS), as the NO donor DETA-NONOate reverted vasoinhibin action. It is an experimentally proven fact that the elevation of intracellular Ca2+ levels ([Ca2+]i) upon BK stimulation activates eNOS, and vasoinhibins blocked the BK-mediated activation of phospholipase C and the formation of inositol 1,4,5-triphosphate leading to a reduced release of Ca2+ from intracellular stores. The [Ca2+]i rise evoked by BK also involves the influx of extracellular Ca2+ via canonical transient receptor potential (TRPC) channels. Vasoinhibins likely interfere with TRPC-mediated Ca2+ entry since La3+, which is an enhancer of TRPC4 and TRPC5 channel activity, prevented vasoinhibins from blocking the stimulation by BK of endothelial cell NO production and proliferation, and vasoinhibins reduced the BK-induced increase of TRPC5 mRNA expression. Finally, vasoinhibins prevented the BK-induced phosphorylation of eNOS at Ser1179, a post-translational modification that facilitates Ca2+-calmodulin activation of eNOS. Together, our data show that vasoinhibins, by lowering NO production through the inhibition of both [Ca2+]i mobilization and eNOS phosphorylation, prevent the BK-induced stimulation of endothelial cell proliferation. Thus, vasoinhibins help to regulate BK effects on angiogenesis and vascular homeostasis.

The prolactin family hormones regulate vascular tone through NO and prostacyclin production in isolated rat aortic rings

Aim:Prolactin family hormones include growth hormone, placental lactogen and prolactin, which are able to regulate angiogenesis via NO and prostaglandins. However, their effects on vascular tone are not fully understood. The aim of this study was to evaluate the effects of prolactin family hormones on rat vascular tone in vitro.Methods:Aortic rings were prepared from adult male rats and precontracted with phenylephrine, then treated with the hormones and drugs. The tension was measured with isometric force displacement transducer connected to a polygraph. NO production and prostacyclin release in physiological solution was determined. Cultured rat aortic endothelial cells (RAECs) were treated with the hormones and drugs, and the phosphorylation of eNOS at serine 1177 was assessed using Western bolt analysis.Results:Administration of growth hormone or placental lactogen (0.01–100 nmol/L) induced endothelium-dependent vasodilation. Both the hormones significantly increased the phosphorylation of eNOS in RAECs and NO level in physiological solution. Preincubation with L-NAME blocked growth hormone- or placental lactogen-induced vasodilation and NO production. Preincubation with an antibody against growth hormone receptors blocked growth hormone- and placental lactogen-induced vasodilation. Addition of a single dose of prolactin (0.01 nmol/L) induced sustained vessel relaxation, whereas multiple doses of prolactin induced a biphasic contraction-relaxation effect. The vascular effects of prolactin depended on endothelium. Prolactin significantly increased the level of prostacyclin I2 in physiological solution. Preincubation with indomethacin or an antibody against prolactin receptors blocked prolactin-induced vasodilation.Conclusion:The prolactin family hormones regulate rat vascular tone, selectively promoting either relaxation or contraction of vascular smooth muscle via activation of either growth hormone receptors or prolactin receptors within the endothelium.

Improved biovolume estimation of Microcystis aeruginosa colonies: A statistical approach

The Microcystis aeruginosa complex (MAC) clusters many of the most common freshwater and brackish bloom-forming cyanobacteria. In monitoring protocols, biovolume estimation is a common approach to determine MAC colonies biomass and useful for prediction purposes. Biovolume (μm3 mL-1) is calculated multiplying organism abundance (orgL-1) by colonial volume (μm3org-1). Colonial volume is estimated based on geometric shapes and requires accurate measurements of dimensions using optical microscopy. A trade-off between easy-to-measure but low-accuracy simple shapes (e.g. sphere) and time costly but high-accuracy complex shapes (e.g. ellipsoid) volume estimation is posed. Overestimations effects in ecological studies and management decisions associated to harmful blooms are significant due to the large sizes of MAC colonies. In this work, we aimed to increase the precision of MAC biovolume estimations by developing a statistical model based on two easy-to-measure dimensions. We analyzed field data from a wide environmental gradient (800 km) spanning freshwater to estuarine and seawater. We measured length, width and depth from ca. 5700 colonies under an inverted microscope and estimated colonial volume using three different recommended geometrical shapes (sphere, prolate spheroid and ellipsoid). Because of the non-spherical shape of MAC the ellipsoid resulted in the most accurate approximation, whereas the sphere overestimated colonial volume (3-80) especially for large colonies (MLD higher than 300 μm). Ellipsoid requires measuring three dimensions and is time-consuming. Therefore, we constructed different statistical models to predict organisms depth based on length and width. Splitting the data into training (2/3) and test (1/3) sets, all models resulted in low training (1.41-1.44%) and testing average error (1.3-2.0%). The models were also evaluated using three other independent datasets. The multiple linear model was finally selected to calculate MAC volume as an ellipsoid based on length and width. This work contributes to achieve a better estimation of MAC volume applicable to monitoring programs as well as to ecological research.

Role of single walled carbon nanotubes (SWCNTs) on the airway smooth muscle

Abstract The aim of this work was to evaluate the effects induced by single walled carbon nanotubes (SWCNTs) on the airway smooth muscle tone, using an isolated rat tracheal rings model and monitoring nitric oxide (NO) as a possible mediator involved in the effects. Exposure of tracheal rings to SWCNTs did not modify the smooth muscle tone per se; however,when the ringswere pre-treated with the contractile agent acetylcholine (ACh), all SWCNTs concentrations (0.1-10 μg/mL) induced a transient contractile effect similar to that induced by ACh alone. Interestingly, the NO production was not modified by SWCNTs regardless of the presence of ACh 10 μM. Thus, the data suggest that NO is not involved in the airway smooth muscle contraction induced by SWCNTs. Further investigations are required to understand the effects, mediator(s) and mechanisms of action induced by this type of porous nanomaterial, as well as their fine interactions with the respiratory system structures.