Sprouting strategies and dead ends in anti-angiogenic targeting of NETs.

Neuroendocrine tumors (NETs) are a heterogeneous group of neoplasms that arise from cells of the neuroendocrine system. NETs are characterized by being highly vascularized tumors that produce large amounts of proangiogenic factors. Due to their complexity and heterogeneity, progress in the development of successful therapeutic approaches has been limited. For instance, standard chemotherapy-based therapies have proven to be poorly selective for tumor cells and toxic for normal tissues. Considering the urge to develop an efficient therapy to treat NET patients, vascular targeting has been proposed as a new approach to block tumor growth. This review provides an update of the mechanisms regulating different components of vessels and their contribution to tumor progression in order to develop new therapeutic drugs. Following the description of classical anti-angiogenic therapies that target VEGF pathway, new angiogenic targets such as PDGFs, EGFs, FGFs and semaphorins are further explored. Based on recent research in the field, the combination of therapies that target multiple and different components of vessel formation would be the best approach to specifically target NETs and inhibit tumor growth.

Neuroendocrine humoral and vascular components in the pressor pathway for brain angiotensin II: a new axis in long term blood pressure control.

Central nervous system (CNS) administration of angiotensin II (Ang II) raises blood pressure (BP). The rise in BP reflects increased sympathetic outflow and a slower neuromodulatory pressor mechanism mediated by CNS mineralocorticoid receptors (MR). We investigated the hypothesis that the sustained phase of hypertension is associated also with elevated circulating levels of endogenous ouabain (EO), and chronic stimulation of arterial calcium transport proteins including the sodium-calcium exchanger (NCX1), the type 6 canonical transient receptor potential protein (TRPC6), and the sarcoplasmic reticulum calcium ATPase (SERCA2). Wistar rats received a chronic intra-cerebroventricular infusion of vehicle (C) or Ang II (A, 2.5 ng/min, for 14 days) alone or combined with the MR blocker, eplerenone (A+E, 5 µg/day), or the aldosterone synthase inhibitor, FAD286 (A+F, 25 µg/day). Conscious mean BP increased (P<0.05) in A (123 ± 4 mm Hg) vs all other groups. Blood, pituitary and adrenal samples were taken for EO radioimmunoassay (RIA), and aortas for NCX1, TRPC6 and SERCA2 immunoblotting. Central infusion of Ang II raised plasma EO (0.58 ± 0.08 vs C 0.34 ± 0.07 nM (P<0.05), but not in A + E and A + F groups as confirmed by off-line liquid chromatography (LC)-RIA and LC-multistage mass spectrometry. Two novel isomers of EO were elevated by Ang II; the second less polar isomer increased >50-fold in the A+F group. Central Ang II increased arterial expression of NCX1, TRPC6 and SERCA2 (2.6, 1.75 and 3.7-fold, respectively; P<0.01)) but not when co-infused with E or F. Adrenal and pituitary EO were unchanged. We conclude that brain Ang II activates a CNS-humoral axis involving plasma EO. The elevated EO reprograms peripheral ion transport pathways known to control arterial Na(+) and Ca(2+) homeostasis; this increases contractility and augments sympathetic effects. The new axis likely contributes to the chronic pressor effect of brain Ang II.

Vagal nerve activity as a moderator of brain-immune relationships.

We investigated whether vagal tone, as assessed by heart rate variability (HRV), moderates the neural correlates of immune and physiological responses to acute stress. Participants with low and high baseline HRV underwent a reversal learning task as an acute stressor. Natural killer cells, norepinephrine, and adrenocorticotropic hormone in peripheral blood changed with acute stress in the high HRV group only. Activity in the prefrontal cortex and striatum correlated with the immune and physiological indices in the high HRV group. High vagal tone may reflect more flexible top-down brain regulation of immune and physiological activity.

Different vascular permeability between the sensory and secretory circumventricular organs of adult mouse brain.

The blood-brain barrier (BBB) prevents free access of circulating molecules to the brain and maintains a specialized brain environment to protect the brain from blood-derived bioactive and toxic molecules; however, the circumventricular organs (CVOs) have fenestrated vasculature. The fenestrated vasculature in the sensory CVOs, including the organum vasculosum of lamina terminalis (OVLT), subfornical organ (SFO) and area postrema (AP), allows neurons and astrocytes to sense a variety of plasma molecules and convey their information into other brain regions and the vasculature in the secretory CVOs, including median eminence (ME) and neurohypophysis (NH), permits neuronal terminals to secrete many peptides into the blood stream. The present study showed that vascular permeability of low-molecular-mass tracers such as fluorescein isothiocyanate (FITC) and Evans Blue was higher in the secretory CVOs and kidney as compared with that in the sensory CVOs. On the other hand, vascular permeability of high-molecular-mass tracers such as FITC-labeled bovine serum albumin and Dextran 70,000 was lower in the CVOs as compared with that in the kidney. Prominent vascular permeability of low- and high-molecular-mass tracers was also observed in the arcuate nucleus. These data demonstrate that vascular permeability for low-molecular-mass molecules is higher in the secretory CVOs as compared with that in the sensory CVOs, possibly for large secretion of peptides to the blood stream. Moreover, vascular permeability for high-molecular-mass tracers in the CVOs is smaller than that of the kidney, indicating that the CVOs are not totally without a BBB.

Aquaporin-1 in blood vessels of rat circumventricular organs.

Although the water channel protein aquaporin-1 (AQP1) is widely observed outside the rat brain in continuous, but not fenestrated, vascular endothelia, it has not previously been observed in any endothelia within the normal rat brain and only to a limited extent in the human brain. In this immunohistochemical study of rat brain, AQP1 has also been found in microvessel endothelia, probably of the fenestrated type, in all circumventricular organs (except the subcommissural organ and the vascular organ of the lamina terminalis): in the median eminence, pineal, subfornical organ, area postrema and choroid plexus. The majority of microvessels in the median eminence, pineal and choroid plexus, known to be exclusively fenestrated, are shown to be AQP1-immunoreactive. In the subfornical organ and area postrema in which many, but not all, microvessels are fenestrated, not all microvessels are AQP1-immunoreactive. In the AQP1-immunoreactive microvessels, the AQP1 probably facilitates water movement between blood and interstitium as one component of the normal fluxes that occur in these specialised sensory and secretory areas. AQP1-immunoreactive endothelia have also been seen in a small population of blood vessels in the cerebral parenchyma outside the circumventricular organs, similar to other observations in human brain. The proposed development of AQP1 modulators to treat various brain pathologies in which AQP1 plays a deleterious role will necessitate further work to determine the effect of such modulators on the normal function of the circumventricular organs.

Evidence for the coexistence of serotonin and calcitonin gene-related peptide at the subcellular level in neuroepithelial bodies in the lung of a marsupial, Isoodon macrourus.

The coexistence of serotonin and calcitonin gene-related peptide (CGRP) in neuroepithelial bodies of the bandicoot, Isoodon macrourus, has been examined using immunocytochemistry at the light- and electron-microscope levels. The avidin-biotin technique of antigen localisation was used initially to identify serotonin-like and CGRP-like immunoreactivity (-LI). Serotonin-LI and CGRP-LI were found in neuroepithelial cells in the lungs of 30-day-old bandicoots. CGRP-LI could also be demonstrated in nerve fibres associated with some neuroepithelial bodies. The protein A-gold technique of antigen localisation was used to label neuroepithelial cells and nerve fibres at the subcellular level. Serotonin-LI and CGRP-LI were observed in the same dense-cored vesicles of most neuroepithelial cells; however, some neuroepithelial cells were shown to possess serotonin-LI without CGRP-LI. Nerve fibres immediately adjacent to neuroepithelial bodies exhibited mainly CGRP-LI. These results show that serotonin-LI and CGRP-LI are present in neuroepithelial cells of the bandicoot in the same secretory vesicles. This pattern of co-localisation may reflect co-ordinated or synergistic actions of these two neuroactive substances.

[Intraspinal organ in man]. / Intraspinal'nyi organ cheloveka.

In the ependymal zone of the spinal cord at the LI-SIII level an ependymal glandular organ has been described. Its highest secretory activity coincides with the period of the greatest functional activity of the human reproductive system functioning. Certain stages in development of the organ, its cell composition, blood supply, afferent and efferent innervation have been studied. In secretory cells of the organ peptide with cardio- and vasotonic properties has been identified immunochemically. The action of the organ is considered in connection with role, which the ependymal glands of the brain play in regulation of the organism's function.

Quantitative fine structure of capillaries in subregions of the rat subfornical organ.

The differentiated cytology across subregions of the rat subfornical organ (SFO) prompted our hypothesis that ultrastructural features of capillary endothelial cells would vary topographically and quantitatively within this small nucleus. We used electron microscopic and computer-based morphometric methods to assess fine structural dimensions of the capillary endothelium in four distinct subregions of the SFO from Long-Evans and homozygous Brattleboro rats. Three types of capillary were present. Type III capillaries (resembling those of endocrine glands) had an average wall thickness of 0.17 microns, 54% thinner than those of Type I and II capillaries. Pericapillary spaces around Type III capillaries measured 56 microns2, 100% larger than for Type I vessels (resembling those of skeletal muscle). Only Type III capillaries contained fenestrations (9 per microns2 of endothelial cell) and were the predominant type of capillary in central and caudal subregions of the SFO. Type I capillaries, prevalent in the transitional subregion between the central and rostral parts of the SFO, had 10 cytoplasmic vesicles per micron2 of endothelial cell area, a number not different from that of Type III capillaries but 3x the frequency found in Type II vessels. Type II capillaries (those typical of "blood-brain barrier" endothelium) had low vesicular density (3 per microns2), no fenestrations, and no pericapillary spaces. Luminal diameters and the densities of mitochondria and intercellular junctions were not different among capillary types or subregions in the SFO. Furthermore, there were no morphometric differences for any capillary dimensions between Long-Evans and Brattleboro rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Are close contacts between astrocytes and endothelial cells a prerequisite condition of a blood-brain barrier? The rat subfornical organ as an example.

The microvessels of the rat subfornical organ (SFO) are heterogeneous: those of the caudal part lack a blood-brain barrier (BBB) unlike those of the rostral part. The astroglial environment of these microvessels has been studied by combining an immunocytochemical technique employing an anti-GFAP (glial fibrillary acidic protein) antiserum with the morphological detection of a barrier to the protein-silver complex. All the SFO microvessels are surrounded by astrocytes characterized by a tumescent aspect; however, the relative proximity between the astrocytic feet and the endothelial cells varies considerably. The capillaries provided with a barrier (rostral SFO) are contiguous with the astrocytes from which they are only separated by a basement membrane. The capillaries devoid of BBB (caudal SFO) are surrounded by a pericapillary space that keeps the astrocytes at a short distance (capillaries with a very rich vesicular endothelium) or at a long distance (capillaries with a fenestrated endothelium). The astrocytes are absent in the choroid plexus where all microvessels are fenestrated and lack a barrier. These data suggest that the astrocytes release one or more signals which in their vicinity inhibit the expression of endothelial morphological characteristics (fenestrations, vesicles) responsible for the leakage of plasmatic proteins from the blood to the cerebral parenchyma of the circumventricular organs.

Topography and morphometry of capillaries in the rat subfornical organ.

A comprehensive stereological analysis was performed to define capillary dimensions in individual subregions of the subfornical organ in Long-Evans, homozygous Brattleboro, and Sprague-Dawley rats. Capillary density, volume fraction, length, surface area, and diameter were assessed in four regions in the sagittal plane (rostral, "transitional," central, and caudal) and two zones in the coronal plane (dorsal and ventromedial). The ventromedial zones in the central and caudal regions correspond to areas of dense perikarya and neuropil containing neural afferent inputs to the subfornical organ (e.g., putative fiber terminals for angiotensin II), whereas the dorsal zones of these regions are apparently the predominant sites of perikarya having efferent projections directed outside of the organ. The morphometric analysis revealed heterogeneous capillary density across subregions of the subfornical organ (range of 132 to 931 capillaries/mm2 in the three rat groups). Capillaries in the ventromedial zones of the central and caudal regions had significantly greater density, volume fraction, and surface area, but smaller diameters, than those in the adjacent dorsal zones and more rostral regions. Across all subregions within the dorsal zone, there was generally a consistent morphometric pattern in the three rat groups. No differences in capillary dimensions in any part of the subfornical organ were found between the Long-Evans and Brattleboro rats. A qualitative electron microscopic investigation of endothelial cells in each subregion of the subfornical organ in Long-Evans rats revealed at least three types of capillary oriented according to region: in the rostral region were capillaries having no endothelial fenestrations or pericapillary spaces, and few vesicles, in the "transitional" region between the rostral and central regions, capillaries having no endothelial fenestrations, substantial numbers of vesicles, and narrow but perceptible pericapillary spaces were found, and in the central and caudal regions, capillaries having abundant endothelial fenestrations and vesicles, expansive pericapillary labyrinths, and relatively thin walls were present. These findings from light microscopic morphometry and electron microscopy in rats indicate a heterogeneity of capillary organization that shows topographical correspondence to the cytology and putative functions of the subfornical organ.

Angioarchitecture of the CNS, pituitary gland, and intracerebral grafts revealed with peroxidase cytochemistry.

Blood vessels of the fetal, neonatal, and adult subprimate and primate CNS, including circumventricular organs (e.g., median eminence, pituitary gland, etc.), and of solid CNS and nonneural (anterior pituitary gland) allografts placed within brains of adult mammalian hosts were visualized with peroxidase cytochemistry applied in three ways: to tissues from animals injected systemically with native horseradish peroxidase (HRP) or peroxidase conjugated to the lectin wheat germ agglutinin (WGA) prior to perfusion fixation; to tissues from animals infused with native HRP into the aorta subsequent to perfusion fixation; and to tissues from animals fixed by immersion and incubated for endogenous peroxidase activity in red cells retained within blood vessels. In neonatal and adult animals receiving native HRP intravascularly, non-fenestrated vessels contributing to a blood-brain barrier were outlined with HRP reaction product when tetramethylbenzidine (TMB) as opposed to diaminobenzidine (DAB) was used as the chromogen; fenestrated vessels of circumventricular organs were not discernible due to the density of extravascular reaction product. Fenestrated and non-fenestrated cerebral and extracerebral blood vessels exposed to bloodborne WGA-HRP were visible when incubated in TMB and DAB solutions. Native HRP infused into the aorta of fixed animals likewise labeled non- fenestrated vessels throughout the brain upon exposure to TMB or DAB but obscured fenestrated vessels of the circumventricular organs. Endogenous peroxidase activity of red cells, seen equally well with TMB and DAB, outlined blood vessels throughout the cerebral gray and white matter and all circumventricular organs in fetal, neonatal, and adult animals. Application of the three peroxidase cytochemical approaches to study the development or absence of a blood-brain barrier in intracerebral allografts demonstrated that the vascularization of day 16-19 fetal/1 day neonatal CNS allografts is not well defined prior to 7 days following intracerebral placement of the grafts. CNS allografts secured from donor sites expected to possess a blood-brain barrier exhibited blood vessels that were not leaky to HRP injected intravenously in the host. Fenestrated blood vessels associated with anterior pituitary allografts were evident prior to 3 days posttransplantation within the host brain and permitted blood-borne HRP in the host to enter the graft and surrounding host brain parenchyma.

Vascular and leptomeningeal projections of the subcommissural organ in reptiles. Lectin-histochemical, immunocytochemical, and ultrastructural studies.

In the snake, Natrix maura, and the turtle, Mauremys caspica, the basal processes of the ependymal cells of the subcommissural organ project toward the local blood vessels and the leptomeninges. These processes and their endings were studied using aldehyde-fuchsin (AF), periodic-acid Schiff (PAS), periodic-acid silver-methenamine (PA-SM), concanavalin A (ConA), wheat germ agglutinin (WGA), immunoperoxidase staining (employing an antiserum against bovine Reissner's fiber; AFRU), and conventional transmission electron microscopy. For the purposes of comparison, the ventricular cell pole was also analyzed. The secretory material located in the ventricular cell pole and that present in ependymal endings had only a few staining properties in common, i.e., affinity for AF, ConA, and AFRU at a dilution of 1:1000. On the other hand, PAS, PA-SM, WGA, and AFRU at a dilution of 1:200,000 stained the apical (ventricular) secretory material but not the secretory material of the ependymal processes. The histochemical features of the secretory material located in the terminals of ependymal processes, as well as the presence at these sites of numerous rough-endoplasmic-reticulum cisternae and secretory granules, suggest that secretory material may be synthesized in these terminals. The probable fate of this material, i.e., release to the perivascular and leptomeningeal spaces or transport to the ventricular cell pole, is discussed.

The microcirculation of rat circumventricular organs and pituitary gland.

Blood volume, blood flow, and blood-to-tissue transfer of an amino acid in circumventricular organs, such as the median eminence and subfornical organ, and the pituitary gland of conscious rats were measured by using quantitative autoradiographic techniques and computer-assisted processing of the tissue images. Retained erythrocyte and plasma volumes observed in circumventricular organs and the anterior and neural lobes of the pituitary gland were dissimilar but in all cases greater by several times than those in cerebral grey matter; these findings suggest the presence of a dense network of high-resistance microvessels in circumventricular organs. The rate of capillary blood flow in the subfornical organ and median eminence was similar to that of grey matter, whereas blood flow in the pituitary neural lobe was several times higher than in grey matter. Thus the apparent velocity of intracapillary blood flow is much higher in the neural lobe than in the subfornical organ. Blood-to-tissue transfer of a small neutral amino acid, alpha-aminoisobutyric acid, was 200 to 700 times more rapid in circumventricular organs and pituitary neural lobe than in the inferior colliculus and caudate nucleus, structures having a blood-brain barrier (BBB). Morphometric analyses indicated that capillary volume and surface area were two times larger in the neural lobe than in the subfornical organ. Moreover, capillaries of the neural lobe and subfornical organ had numerous endothelial fenestrations and cytoplasmic pits or vesicles, whereas capillaries of the inferior colliculus had no fenestrations and fewer vesicles. These studies demonstrate quantitative differences in the microcirculatory systems not only between circumventricular organs and BBB structures but also among circumventricular organs.

Is the avian glycogen body a secretory organ?

In the present study we have observed with the electron microscope the glycogen body of the chick, Gallus domesticus, which constitutes a spinal cord circumventricular organ specific of birds. We have detected in young chickens and embryos the presence of considerable amounts of glycogen particles in the cerebrospinal and vascular compartments, coming from the glycogen body cells, which are able to discharge part of their glycogen into either the central canal or the capillary lumen, via ependymal and endothelial cells respectively. If this secretion is a physiological condition, we propose that the glycogen body would play a role in the maintenance of the hydric and glucose haemostasia in the central nervous system, as well as in the osmoregulation.

Ultrastructural changes in the urophysis of Mollienesia sphenops following adaptation to seawater.

The urophysis or neurohemal contact site of the caudal neurosecretory system of Mollienesia sphenops, the black molly, was studied in animals adapted to an artificial seawater environment. This species of fish was chosen for these studies because of its known ability to osmoregulate and its adaptability to the laboratory aquarium. The urophysis of freshwater acclimated mollys contained an abundance of neurosecretory granules. However, in fish subjected to a seawater environment for one week the number of neurosecretory granules was significantly decreased. In addition, there was an increase in blood cell infiltration of the urophysis.