Regression Of Blood Vessels Research Articles

Notch and VEGF pathways play distinct but complementary roles in tumor angiogenesis

BackgroundAnti-angiogenesis is a validated strategy to treat cancer, with efficacy in controlling both primary tumor growth and metastasis. The role of the Notch family of proteins in tumor angiogenesis is still emerging, but recent data suggest that Notch signaling may function in the physiologic response to loss of VEGF signaling, and thus participate in tumor adaptation to VEGF inhibitors.MethodsWe asked whether combining Notch and VEGF blockade would enhance suppression of tumor angiogenesis and growth, using the NGP neuroblastoma model. NGP tumors were engineered to express a Notch1 decoy construct, which restricts Notch signaling, and then treated with either the anti-VEGF antibody bevacizumab or vehicle.ResultsCombining Notch and VEGF blockade led to blood vessel regression, increasing endothelial cell apoptosis and disrupting pericyte coverage of endothelial cells. Combined Notch and VEGF blockade did not affect tumor weight, but did additively reduce tumor viability.ConclusionsOur results indicate that Notch and VEGF pathways play distinct but complementary roles in tumor angiogenesis, and show that concurrent blockade disrupts primary tumor vasculature and viability further than inhibition of either pathway alone.

A viscoelastic model of blood capillary extension and regression: derivation, analysis, and simulation

This work studies a fundamental problem in blood capillary growth: how the cell proliferation or death induces the stress response and the capillary extension or regression. We develop a one-dimensional viscoelastic model of blood capillary extension/regression under nonlinear friction with surroundings, analyze its solution properties, and simulate various growth patterns in angiogenesis. The mathematical model treats the cell density as the growth pressure eliciting a viscoelastic response from the cells, which again induces extension or regression of the capillary. Nonlinear analysis captures two cases when the biologically meaningful solution exists: (1) the cell density decreases from root to tip, which may occur in vessel regression; (2) the cell density is time-independent and is of small variation along the capillary, which may occur in capillary extension without proliferation. The linear analysis with perturbation in cell density due to proliferation or death predicts the global biological solution exists provided the change in cell density is sufficiently slow in time. Examples with blow-ups are captured by numerical approximations and the global solutions are recovered by slow growth processes, which validate the linear analysis theory. Numerical simulations demonstrate this model can reproduce angiogenesis experiments under several biological conditions including blood vessel extension without proliferation and blood vessel regression.

ATOH7 mutations cause autosomal recessive persistent hyperplasia of the primary vitreous

The vertebrate basic helix-loop-helix (bHLH) transcription factor ATOH7 (Math5) is specifically expressed in the embryonic neural retina and is required for the genesis of retinal ganglion cells (RGCs) and optic nerves. In Atoh7 mutant mice, the absence of trophic factors secreted by RGCs prevents the development of the intrinsic retinal vasculature and the regression of fetal blood vessels, causing persistent hyperplasia of the primary vitreous (PHPV). We therefore screened patients with hereditary PHPV, as well as bilateral optic nerve aplasia (ONA) or hypoplasia (ONH), for mutations in ATOH7. We identified a homozygous ATOH7 mutation (N46H) in a large family with an autosomal recessive PHPV disease trait linked to 10q21, and a heterozygous variant (R65G, p.Arg65Gly) in one of five sporadic ONA patients. High-density single-nucleotide polymorphism analysis also revealed a CNTN4 duplication and an OTX2 deletion in the ONA cohort. Functional analysis of ATOH7 bHLH domain substitutions, by electrophoretic mobility shift and luciferase cotransfection assays, revealed that the N46H variant cannot bind DNA or activate transcription, consistent with structural modeling. The N46H variant also failed to rescue RGC development in mouse Atoh7-/- retinal explants. The R65G variant retains all of these activities, similar to wild-type human ATOH7. Our results strongly suggest that autosomal recessive persistent hyperplastic primary vitreous is caused by N46H and is etiologically related to nonsyndromic congenital retinal nonattachment. The R65G allele, however, cannot explain the ONA phenotype. Our study firmly establishes ATOH7 as a retinal disease gene and provides a functional basis to analyze new coding variants.

Antiangiogenic and antivascular effects of a recombinant tumstatin-derived peptide in a corneal neovascularization model

Tumstatin, a cleavage fragment of collagen IV, is a potent endogenous inhibitor of angiogenesis. Tumstatin-derived peptide T8 possesses all angiostatic properties of full-length tumstatin and indirectly suppresses tumor growth. The potential of T8 to block pathological angiogenesis in the eye has not been explored yet. Here we assess antiangiogenic effects of a recombinant T8 peptide in rabbit corneal neovascularization models.The fusion protein consisting of T8 and thioredoxin was synthesized in a highly efficient Escherichia coli expression system, isolated using ion-exchange chromatography and cleaved with TEV (tobacco etch virus) protease. The target peptide was purified on an anion-exchange resin and by reversed phase high-performance liquid chromatography.The recombinant peptide suppressed the proliferation of basic fibroblast growth factor-induced SVEC-4-10 endothelial cells (simian virus 40-immortalized murine endothelial cells) and inhibited tube formation in these cells in a dose-dependent manner. In rabbit corneal neovascularization models T8 demonstrated the ability to prevent pathological angiogenesis (when injected simultaneously with the induction of neovascularization) and, moreover, to promote the regression of newly-formed blood vessels (when injected on day 8 after angiogenesis stimulation).Our results suggest that T8 may have a therapeutic potential in the treatment of ocular neovascular diseases.

MicroRNA-125b inhibits tube formation of blood vessels through translational suppression of VE-cadherin

Angiogenesis is controlled positively or negatively by extrinsic and intrinsic molecular cues in endothelial cells (ECs); in the tumor microenvironment, the action of positive regulators exceeds that of negative regulators. Thus, overinduction of negative regulators may inhibit tumor angiogenesis. MicroRNAs (miRNAs or miRs) are endogenous short noncoding RNAs regulating gene expression either through translational inhibition or destabilization of target mRNA. Here, we show that miR-125b expression is transiently induced in ECs on stimulation with vascular endothelial growth factor or by ischemia. miR-125b inhibits translation of vascular endothelial (VE)-cadherin mRNA and in vitro tube formation by ECs. Injection of miR-125b into the tumor inhibited VE-cadherin expression by ECs and induced nonfunctional blood vessel formation, resulting in inhibition of tumor growth. It has been suggested that pro-angiogenic signals in ECs also upregulate anti-angiogenic molecules simultaneously via negative feedback. Because miR-125b induction in ECs is transient after pro-angiogenic stimulation, prolonged overexpression of miR-125b could result in blood vessel regression. Thus, miR-125b may be useful in cancer therapy by causing the collapse of the lumen of ECs.

Prolactin and blood vessels: A comparative endocrinology perspective

The hormone prolactin (PRL), fundamental for lactation in mammals, is known to exert a wide diversity of actions in the various vertebrate groups. Blood vessels are surfacing as important PRL targets, contributing to these hormonal functions. PRL promotes the growth of new blood vessels (angiogenesis) and is proteolytically cleaved to vasoinhibins, a family of peptides (including 16-kDa PRL) with potent antiangiogenic and blood vessel regression effects. These opposing actions point to the regulation of the proteases responsible for PRL cleavage as an efficient way to balance blood vessel growth and involution. This review briefly summarizes the effects of PRL and vasoinhibins on blood vessels in mammals and discusses whether similar vascular actions could contribute to the effects of PRL on the development, growth, and reproduction of lower vertebrates. A comparative study in diverse species may lead to a better understanding of blood vessels as a driving force for the biological actions of PRL.

Rapamycin Suppresses Self-Renewal and Vasculogenic Potential of Stem Cells Isolated from Infantile Hemangioma

Infantile hemangioma (IH) is a common childhood vascular tumor. Although benign, some hemangiomas cause deformation and destruction of features or endanger life. The current treatments, corticosteroid or propranolol, are administered for several months and can have adverse effects for the infant. We designed a high-throughput screen to identify FDA-approved drugs that could be used to treat this tumor. Rapamycin, an mTOR inhibitor, was identified based on its ability to inhibit proliferation of a hemangioma-derived stem cell population, human vasculogenic cells we had previously discovered. In vitro and in vivo studies show that Rapamycin reduces the self-renewal capacity of the hemangioma stem cells, diminishes differentiation potential, and inhibits the vasculogenic activity of these cells in vivo. Longitudinal in vivo imaging of blood flow through vessels formed with hemangioma stem cells shows that Rapamycin also leads to regression of hemangioma blood vessels, consistent with its known anti-angiogenic activity. Finally, we demonstrate that Rapamycin-induced loss of stemness can work in concert with corticosteroid, the current standard therapy for problematic hemangioma, to block hemangioma formation in vivo. Our studies reveal that Rapamycin targets the self-renewal and vascular differentiation potential in patient-derived hemangioma stem cells and suggests a novel therapeutic strategy to prevent formation of this disfiguring and endangering childhood tumor.

The Dll4/Notch pathway controls postangiogenic blood vessel remodeling and regression by modulating vasoconstriction and blood flow

AbstractBlood vessel remodeling is crucial to the formation of the definitive vasculature, but little is known about the mechanisms controlling this process. We show that Delta-like ligand 4 (Dll4)/Notch pathway regulates vessel regression in normal pathologic conditions. Genetic and pharmacologic inhibition of Dll4/Notch prevented retinal capillary regression in the oxygen-induced retinopathy (OIR) model and during normal development. Deletion of the Notch-regulated ankyrin repeat protein, a negative regulator of the Notch pathway, produced an opposite phenotype. Inhibition of Dll4/Notch reduced vessel occlusion, maintaining blood flow that is essential for survival of microvessels. Dll4/Notch inhibition up-regulated the expression of vasodilators adrenomedullin and suppressed the expression of vasoconstrictor angiotensinogen. Angiotensin II induced rapid nonperfusion and regression of developing retinal capillaries, whereas Ace1 and AT1 inhibitors and adrenomedullin attenuated vasoobliteration in OIR, indicating that both pathways are involved in modulating vessel remodeling. In contrast, inhibition of vascular endothelial growth factor-A (VEGF-A) did not result in a pervasive loss of retinal capillaries, demonstrating that reduced expression of VEGF-A is not the proximate cause of capillary regression in OIR. Modulation of VEGF-A and DII4/Notch signaling produced distinct changes in blood vessel morphology and gene expression, indicating that these pathways can have largely independent functions in vascular remodeling.

Perilimbal Bevacizumab Injection for Interface Neovascularization After Deep Anterior Lamellar Keratoplasty

To evaluate the effect of perilimbal bevacizumab injection on neovascularization at the interface of donor and recipient tissues after deep anterior lamellar keratoplasty (DALK). This interventional case series included 4 patients with interface neovascularization after DALK (3 patients with advanced keratoconus and 1 patient with scarring because of herpetic keratitis). Bevacizumab (2.5 mg/0.1 mL) was injected subconjunctivally near the limbus adjacent to the abnormal blood vessels. Depending on regression of new vessels, injections were repeated. If neovascularization occurred in more than 1 quadrant, injections were administered at 2 sites (2.5 mg for each injection). Total corneal neovascularization, longest neovascular length, and degree of neovascular circumference were evaluated by digital slit lamp corneal photography before injection and 1 week, 1, 3, and 6 months afterward. The mean follow-up period was 14 ± 7.52 months from the first bevacizumab injection. Corneal neovascularization, on average,was 33.70% ± 7.8% (SD) of the total corneal surface before the injections, which decreased to 4.83% ± 3.3% (P = 0.005, t test) at the last follow-up examination. Mean degree of neovascularized circumference decreased from 165 degrees before injection to 77.5 degrees (P = 0.023) at the last follow-up examination. Mean longest neovascularized length decreased from 6.714 mm before injection to 1.497 mm (P = 0.021) at the last follow-up examination. At the end of the study, all patients had clear grafts with no sign of graft failure. Mean best-corrected visual acuity before injection was 20/100, which improved to 20/30 at the last examination. This small case series suggests that perilimbal bevacizumab injection is safe and effective for partial regression of new blood vessels at the interface of donor and recipient tissues after DALK and may effectively prevent graft rejection.

Redefining tissue engineering for nanomedicine in ophthalmology

Working at the nanoscale means to completely rethink how to approach engineering in the body in general and in the eye in particular. In nanomedicine, tissue engineering is the ability to influence an environment either by adding, subtracting or manipulating that environment to allow it to be more conducive for its purpose. The goal is to function at the optimum state, or to return to that optimum state. Additive tissue engineering replaces cells or tissue, or tries to get something to grow that is no longer there. Arrestive tissue engineering tries to stop aberrant growth which, if left uncontrolled, would result in a decrease in function. Nano delivery of therapeutics can perform both additive and arrestive functions influencing the environment either way, depending on the targeting. By manipulating the environment at the nanoscale, the rate and distribution of healing can be controlled. It infers that potential applications of nanomedicine in ophthalmology include procedures, such as corneal endothelial cell transplantation, single retinal ganglion cell repair, check of retinal ganglion cell viability, building of nanofibre scaffolds, such as self-assembling peptides, to create a scaffold-like tissue-bridging structure to provide a framework for axonal regeneration in the case of optic nerve reconnection or eye transplantation, and ocular drug delivery. Examples of potential arrestive therapies include gene-related treatment modalities to inhibit intraocular neovascularization and to block retinal cell apoptosis. Looking towards the future, this review focuses on how nanoscale tissue engineering can be and is being used to influence that local environment.

Novel Discovery of LYVE-1 Expression in the Hyaloid Vascular System

The hyaloid vascular system (HVS) is a transient network nourishing developing eyes and has been widely used as a natural model to study blood vessel regression. Failure of its regression in humans leads to several blinding diseases. Lymphatic vessel endothelial hyaluronic acid receptor (LYVE-1) is a recently defined lymphatic marker that is also expressed by a subpopulation of macrophages. To date, there is no report on its expression in the HVS. This study was conducted to investigate whether LYVE-1 is expressed in the HVS and how it is associated with the vascular structure and macrophage phenotype. Normal C57BL/6 mouse eyeballs were sampled from embryonic day (E) 10.5 to postnatal (P) and adult stages for immunofluorescent microscopic studies with antibodies against LYVE-1, CD31 (panendothelial cell marker), and F4/80 (macrophage marker). Additionally, Angiopoietin-2 (Ang-2) knockout mice with abnormally persistent HVS were examined. The LYVE-1 expression was detected on normal HVS between E12.5 and P14. The LYVE-1(+) cells were F4/80(+) but CD31(-), indicating a macrophage lineage. Additionally, LYVE-1(+) cells bud on CD31(+) vessels and constitute an integral part of the network in both normal developing and Ang-2 knockout mice. This study provides the first evidence that the HVS contains a LYVE-1(+) cellular component in both physiological and pathologic conditions. This novel finding not only provides a new concept in defining the embryogenesis and pathogenesis of the HVS, it also leads to a completely natural model in which to study the functions of the LYVE-1 pathway, an important topic for lymphatic research as well.

Phospholipase Cγ Activation Drives Increased Production of Autotaxin in Endothelial Cells and Lysophosphatidic Acid-Dependent Regression

We previously reported that vascular endothelial growth factor (VEGF)-dependent activation of phospholipase Cgamma1 (PLCgamma) regulated tube stability by competing with phosphoinositide 3-kinase (PI3K) for their common substrate. Here we describe an additional mechanism by which PLCgamma promoted regression of tubes and blood vessels. Namely, it increased the level of autotaxin (ATX), which is a secreted form of lysophospholipase D that produces lysophosphatidic acid (LPA). LPA promoted motility of endothelial cells, leading to disorganization/regression of tubes in vitro. Furthermore, mice that under- or overexpressed members of this intrinsic destabilization pathway showed either delayed or accelerated, respectively, regression of blood vessels. We conclude that endothelial cells can be instructed to engage a PLCgamma-dependent intrinsic destabilization pathway that results in the production of soluble regression factors such as ATX and LPA. These findings are likely to potentiate ongoing efforts to prevent, manage, and eradicate numerous angiogenesis-based diseases such as proliferative diabetic retinopathy and solid tumors.

Abstract 1387: Development of novel antiangiogenic biologics: multifunctional VEGF traps

Abstract Current anti-VEGF biologics have been successfully used as therapeutic agents for cancer and age-related macular degeneration (AMD). Since these strategies target VEGF systemically, side effects, long-term toxicities, such as proteinuria and thromboembolic events, and need for frequent eye injections in AMD treatment, prevail. Therefore, the aim of this study was to generate novel anti-VEGF biologics that inhibit VEGF activity specifically at the desired target site. Two classes of antibody-based recombinant proteins were engineered that simultaneously bind VEGF and either: 1) the extracellular matrix (ECM) or 2) target-site specific antigens. The first subgroup of proteins, “shV-H traps”, is comprised of a Fc domain shortened form of VEGF-trap linked to a sequence of hydrophobic amino acids, with various affinities for heparin sulfate proteoglycans of the ECM, designed to have a short systemic half-life. The second subgroup of molecules, “V-lassos”, is composed of a C-terminus positioned form of VEGF-trap linked to single-chain variable domain antibodies specific for either HER-2 or fibronectin extra domain B (EDB), expressed on tumour cell surfaces or in the vascular bed of solid tumours, respectively. Recombinant proteins were expressed in transgenic cancer cell lines in a doxycycline inducible manner and were shown to inhibit VEGF activity and also retain the native function of their constituent domains. Specifically, the shV-H traps adhered to the ECM and the HER-2 V-lasso inhibited the proliferation of HER-2 positive cancer cell lines. Xenograft studies were performed using the aforementioned transgenic cancer cell lines in order to determine the efficacy of shV-H traps and Vlasso. shV-H traps were able to inhibit or delay tumour growth of A-673, Pc-3 and HT-29 xenografts. In contrast to soluble VEGF-trap, sh-VH traps were retained at the tumour site and were undetectable in the circulation. Moreover, sh-VH traps did not cause any of the side-effects observed with soluble VEGF-trap, such as delay of wound healing and regression of trachea blood vessels. Both Vlassos were able to inhibit or delay the tumour growth of SKOV-3 and A-673 xenografts. Furthermore, the transgenic approach for inducible expression indicated that HER-2 Vlasso is more effective compared to anti-HER-2 Ab and VEGF-trap used alone or in combination. Preliminary results suggest that high recombinant protein levels can be generated via large-scale production in 293F cells, indicating that the new multifunctional VEGF-traps can be produced in large amounts for clinical studies. These novel classes of anti-angiogenic molecules could potentially be advantageous in a clinical setting. Furthermore, they can be useful in order to further study and understand the mechanisms involved in VEGF inhibition. Finally, similar dual function proteins can be designed for inhibition of other molecules with disease relevance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1387.

The inflammatory mediator oncostatin M induces angiopoietin 2 expression in endothelial cells in vitro and in vivo.

Members of the glycoprotein 130 (gp130) receptor-gp130 ligand family play a role in angiogenesis in different tissues. We tested the effect of this cytokine family on the angiopoietin (Ang)-Tie system, which is involved in blood vessel maturation, stabilization, and regression. Oncostatin M (OSM) increased Ang2 expression in human umbilical vein endothelial cells via Janus kinase/signal transducer and activator of transcription (JAK/STAT) and mitogen-activated protein (MAP) kinase activation. Furthermore, OSM induced Ang2 expression in macrovascular endothelial cells isolated from the human aorta and in microvascular endothelial cells isolated from human heart. Our in vivo experiments revealed that mRNA expression of Ang2 in hearts of mice injected with OSM increased significantly, and levels of OSM mRNA significantly correlated with mRNA levels of Ang2 in human hearts. In addition, OSM increased the expression of its own receptors, gp130 and OSM receptor, in endothelial cells in vitro and in mice in vivo, and levels of OSM mRNA significantly correlated with mRNA levels of gp130 and OSM receptor in human hearts. Our data, showing the effects of OSM on the Ang-Tie system in endothelial cells, in hearts of mice, and in human heart tissue, provide yet another link between inflammation and angiogenesis.

Pterygia: Pathogenesis and the Role of Subconjunctival Bevacizumab in Treatment

A pterygium is a very common conjunctival degenerative condition. It has been well established that there are different factors that are interrelated and involved in the growth of pterygia. Historically described more as a degenerative process, inflammation and fibrovascular proliferation have proven to be very important factors. Many studies have shown VEGF to be increased in the pathogenesis of pterygia. There are a variety of options for the management of pterygium. The over expression of VEGF in pterygium tissue led us to develop anti-angiogenic/anti-VEGF therapy which could induce regression of blood vessels and hence retard progression of pterygium. The role of angiogenesis and of VEGF in ocular pathology is established including in corneal neovascularization, specifically in pterygial tissue. Evidence suggests that local bevacizumab may be effective in treatment of ocular surface neovascularization. Currently, data in the use of subconjunctival bevacizumab in the treatment of pterygia are not conclusive. New anti-angiogenic therapies will hopefully focus more on facilitating delivery into tissue, increasing the duration of effect while continuing to minimize adverse side effects.

Ovarian Administration of Angiopoietin-1 Ab Increases Follicular Apoptosis Mediated in part by an Imbalance in the Ratio of Androgens/Estrogens and a Decrease in the Granulosa and Theca Cell Proliferation.

The angiopoietin (ANGPT)-receptor (TEK) system plays a crucial role in blood vessel development and regression. Previously, we showed that the inhibition of ANGPT1 caused an increase in the number of atretic follicles and a decrease in the number of both antral follicles (AF) and preovulatory follicles in gonadotropin-treated rat ovaries. The aim of this study was to evaluate the effect of local administration of anti-ANGPT1 antibody (ANGPT1-Ab) on steroidogenesis, apoptosis, cellular proliferation and vascular development, in ovarian follicles from prepubertal eCG-treated rats. Female immature rats superovulated with eCG (25 IU/rat) were injected with an antibody for ANGPT 1 (10ng/ovary) in 5 μl of saline under the bursa of one ovary (ANGPT1 Ab ovary). The contralateral ovary was injected with the same volume and concentration of normal goat IgG (Control ovary). Rats were killed 48 hours after surgery by CO2 asphyxiation. The ovaries were removed, cleaned off adhering tissue in culture medium and AF (250- 400 μm) were dissected under a stereoscopic microscope and used for steroid extractions. Treatment with ANGPT1 Ab resulted in a significant increase in follicular androsterone levels (C: 59.93±7.5 vs ANGPT1 Ab: 85.65±6.7 ng/ovary, p <0.05, n=7), whereas a significant decrease in estradiol was observed (C: 12.13±2.3 vs ANGPT1 Ab: 8.7±2.3 ng/ovary, p<0.05, n=7). However, follicular progesterone levels were unchanged. As granulosa cells (gCs) of the follicle die by apoptosis during the process of follicular atresia, the TUNEL technique was performed on histological ovarian slides from ovaries obtained 48 h after injection of ANGPT1 Ab. Specific staining was observed in gCs and ANGPT1 Ab treatment caused a significant increase in the number of apoptotic cells in AF (C: 2.29±0.36 vs ANGPT1 Ab: 6.08±0.84 apoptotic cells/field, p<0.05). To identify proliferating cells, slides were incubated with anti-PCNA Ab. A proliferation index (PCNA-positive cells expressed as a percentage of the total number of cells) was calculated in the theca and granulosa compartments for each antral follicle. PCNA protein expression decreased significantly in gCs (C: 63.98±3.03 vs ANGPT1 Ab: 46.81±4.02; p<0.01) as well as cTs (C: 59.64±3.7 vs ANGPT1 Ab: 41.37±2.46; p<0.01) from AF in ANGPT1 Ab-treated ovaries. In order to evaluate the endothelial cell (eC) density, histological ovarian slides were immunostained using an antibody raised against the Von Willebrand Factor. No significant differences were found in the % of endothelial cell area between control and ANGPT1 Ab-treated ovaries In conclusion, intraovarian neutralizing antibody against ANGPT1 interferes with the rat follicular development associated with an increase in ovarian apoptosis mediated in part by an imbalance in the ratio of androgen/estrogen steroids and a decrease in granulosa and theca cell proliferation. Therefore, ANGPT1 would play a critical intraovarian survival role for gonadotropin-dependent folliculogenesis. Supported by: UBA (EX237), CONICET (PIP 5471), ANPCYT (PICT 5-26047) and Roemmers Foundation.

Angiogenesis in the human corpus luteum.

Angiogenesis is important for the formation and development of the corpus luteum and for maintenance of luteal function. Blood vessel regression is an important physiological phenomenon in the corpus luteum, which is associated with tissue involution during structural luteolysis. Angiogenesis actively occurs during the early luteal phase and is completed by the mid-luteal phase. Perivascular cells (pericytes) increase in number from the early luteal phase to the mid-luteal phase, suggesting that blood vessels are gradually stabilized until the mid-luteal phase. In the corpus luteum undergoing luteolysis, blood vessels and pericytes decrease in number, which is related to structural involution. In the corpus luteum of early pregnancy, the number of blood vessels with pericytes increases, suggesting that angiogenesis occurs again, accompanied by blood vessel stabilization. These changes in vasculature of the corpus luteum are regulated by the collaboration with vascular endothelial growth factor, which is involved in proliferation of vascular endothelial cells, and angiopoietins, which are involved in stabilization of blood vessels. This review focuses on angiogenesis, blood vessel stabilization and blood vessel regression during the divergent phases of luteal formation, luteal regression and luteal rescue by pregnancy. (Reprod Med Biol 2008; 7: 91-103).

Role of Prolactin and Vasoinhibins in the Regulation of Vascular Function in Mammary Gland

The formation of new blood vessels has become a major focus of mammary gland research stimulated by the therapeutic opportunities of controlling angiogenesis in breast cancer. Normal growth and involution of the mammary gland are profoundly affected by the expansion and regression of blood vessels, whereas dysregulation of angiogenesis is characteristic of breast cancer growth and metastasis. Prolactin stimulates the growth and differentiation of the mammary gland under normal conditions, but its role in breast cancer is controversial. Its action is complicated by the fact that prolactin itself is angiogenic, but proteases cleave prolactin to generate vasoinhibins, a family of peptides that act on endothelial cells to suppress angiogenesis and vasodilation and to promote apoptosis-mediated vascular regression. This review summarizes our current knowledge about the vascular effects of prolactin and the generation and action of vasoinhibins, and discusses their possible contribution to the regulation of blood vessels in the normal and malignant mammary gland.

Intrabursal Administration of the Antiangiopoietin 1 Antibody Produces a Delay in Rat Follicular Development Associated with an Increase in Ovarian Apoptosis Mediated by Changes in the Expression of BCL2 Related Genes1

The angiopoietin (ANGPT) receptor (TEK) system plays a crucial role in blood vessel development and regression. To date, no reports have addressed the actions of the anti-ANGPT1 antibody on gonadotropin-stimulated follicular development and atresia in the ovary. Therefore, in this study we specifically investigated whether ANGPT1 plays a critical intraovarian survival role for gonadotropin-dependent folliculogenesis. In particular, we examined the effect of local administration of anti-ANGPT1 antibody on follicular development, apoptosis, and expression of BCL2 protein family members (BAX, BCL2, and BCL2L1), TNFRSF6, and FASLG in ovarian follicles from prepubertal eCG-treated rats. The inhibition of ANGPT1 caused an increase in the number of atretic follicles and a decrease in the number of both antral follicles (AFs) and preovulatory follicles in gonadotropin-treated rat ovaries. Taking into account that follicular atresia is mediated by apoptosis, we analyzed the effect of the antibody against ANGPT1 on programmed cell death. The inhibition of the action of ANGPT1 caused an increase both in the number of apoptotic granulosa cells in AFs and in the spontaneous DNA fragmentation of AFs cultured in serum-free medium. Besides, AFs obtained from rats treated with intraovarian antibodies against ANGPT1 showed both a decrease in BCL2 and an increase in BAX protein levels. Moreover, a reduction in the BCL2L1(L)/BCL2L1(S) ratio was observed in this group, with a reduction of BCL2L1(L) greater than that of BCL2L1(S), thus showing that the expression of these antiapoptotic proteins is lower in follicles from treated rats than in those from untreated ones. Our findings suggest that the inhibition of ANGPT1 activity causes an increase in the number of atretic follicles mediated by ovarian apoptosis through an imbalance in the ratio of antiapoptotic to proapoptotic proteins. This could take place through a paracrine effect on granulosa cells mediated by the TEK receptor in theca cells. Therefore, these data clearly indicate that ANGPT1 is necessary for follicular development induced by gonadotropins.

Cellular actions of angiogenesis inhibitors on blood vessels

Cellular actions of angiogenesis inhibitors on blood vessels