Morphological Analysis for Neuron‐Like Cells in the Vomeronasal Organ of Human Fetuses at the Middle of Gestation

The vomeronasal organ (VNO) of 5‐month‐old fetuses was examined immunohistochemically by the use of an antiserum to protein gene product 9.5 (PGP). The purpose was to identify if the human fetal VNO is lined by neuroepithelium. The PGP antiserum labeled abundant cells within the vomeronasal epithelium (VE), nerve fiber bundles in its lamina propria, and cells associated with these bundles. PGP‐immunoreactive (ir) vomeronasal epithelial cells were classified into three subtypes. Type I cells, about 44% of the total cells observed, did not have any processes and tended to be located in the basal layer of the VE. Type II cells, about 37% had a single apical process that projected toward the lumen, ending at the epithelial surface. Type III cells sent a prominent process mainly toward the basement membrane, and occupied about 19% of the total cells observed. In the lamina propria, a considerable number of PGP‐ir cells was observed. Some of them were present in nerve fiber bundles and contained processes parallel to the bundles. In addition, PGP‐ir nerve fiber bundles and cells associated with them were even present in the portion of the nasal septal mucosa that was very close to the brain. The present results strongly suggested that the VE in human fetuses at mid‐gestation is a neuroepithelium and that the VE may produce migrating cells toward the brain. Anat Rec, 299:88–97, 2016. © 2015 Wiley Periodicals, Inc.     

A putative functional vomeronasal system in anuran tadpoles

We investigated the occurrence and anatomy of the vomeronasal system (VNS) in tadpoles of 13 different anuran species. All of the species possessed a morphologically fully developed VNS with a highly conserved anatomical organisation. We found that a bean-shaped vomeronasal organ (VNO) developed early in the tadpoles, during the final embryonic stages, and was located in the anteromedial nasal region. Histology revealed the presence of bipolar chemosensory neurones in the VNO that were immunoreactive for the Gαo protein. Tract-tracing experiments demonstrated that chemosensory neurones from the VNO reach specific areas in the brain, where a discernible accessory olfactory bulb (AOB) could be observed. The AOB was located in the ventrolateral side of the anterior telencephalon, somewhat caudal to the main olfactory bulb. Synaptophysin-like immunodetection revealed that synaptic contacts between VNO and AOB are established during early larval stages. Moreover, using lectin staining, we identified glomerular structures in the AOB in most of the species that we examined. According to our findings, a significant maturation in the VNS is achieved in anuran larvae. Recent published evidence strongly suggests that the VNS appeared early in vertebrate evolution and was already present in the aquatic last common ancestor of lungfish and tetrapods. In this context, tadpoles may be a good model in which to investigate the anatomical, biochemical and functional aspects of the VNS in an aquatic environment.     

Galectin-3 immunohistochemistry in the vomeronasal organ of the domestic pig, Sus scrofa

The immunohistochemical localization of galectin-3, a β-galactoside-binding protein, was studied in the vomeronasal organ (VNO) of fetal, 1-day-old, and 6-month-old pigs.In all age groups, the porcine VNO consisted of vomeronasal sensory epithelium (VSE) located medially and non-sensory vomeronasal respiratory epithelium (VRE) located laterally. In the pig, the VNO epithelium increased in height with postnatal development from fetus to adult. In the VSE of all stages examined, galectin-3 immunostaining was seen in the supporting cells and free border, but not in receptor or basal cells. Galectin-3 immunostaining was seen in all layers of the VRE, and the intensity increased with postnatal development. In the lamina propria, galectin-3 was detected in some ductal epithelial cells and the vomeronasal nerve sheath, but not in the acini of the Jacobson glands in all age groups. In view of these observations, we postulate that galectin-3 plays a role in cell survival and cell adhesion in both the VSE and VRE of porcine VNO in all age groups.     

Immunohistochemistry of the canine vomeronasal organ

The canine's olfactory acuity is legendary, but neither its main olfactory system nor its vomeronasal system has been described in much detail. We used immunohistochemistry on paraffin-embedded sections of male and female adult dog vomeronasal organ (VNO) to characterize the expression of proteins known to be expressed in the VNO of several other mammals. Basal cell bodies were more apparent in each section than in rodent VNO and expressed immunoreactivity to anticytokeratin and antiepidermal growth factor receptor antibodies. The thin layer of neurone cell bodies in the sensory epithelium and axon fascicles in the lamina propria expressed immunoreactivity to neurone cell adhesion molecule, neurone-specific beta tubulin and protein gene product 9.5. Some neurones expressed growth-associated protein 43 (GAP43): and a number of those also expressed neurone-specific beta tubulin-immunoreactivity. Some axon fascicles were double labelled for those two proteins. The G-protein alpha subunits Gi and Go, involved in the signal transduction pathway, showed immunoreactivity in the sensory cell layer. Our results demonstrate that the canine vomeronasal organ contains a population of cells that expresses several neuronal markers. Furthermore, GAP43 immunoreactivity suggests that the sensory epithelium is neurogenic in adult dogs.     

Immunohistochemistry of the canine vomeronasal organ

The canine's olfactory acuity is legendary, but neither its main olfactory system nor its vomeronasal system has been described in much detail. We used immunohistochemistry on paraffin-embedded sections of male and female adult dog vomeronasal organ (VNO) to characterize the expression of proteins known to be expressed in the VNO of several other mammals. Basal cell bodies were more apparent in each section than in rodent VNO and expressed immunoreactivity to anticytokeratin and antiepidermal growth factor receptor antibodies. The thin layer of neurone cell bodies in the sensory epithelium and axon fascicles in the lamina propria expressed immunoreactivity to neurone cell adhesion molecule, neurone-specific beta tubulin and protein gene product 9.5. Some neurones expressed growth-associated protein 43 (GAP43): and a number of those also expressed neurone-specific beta tubulin-immunoreactivity. Some axon fascicles were double labelled for those two proteins. The G-protein alpha subunits Gi and Go, involved in the signal transduction pathway, showed immunoreactivity in the sensory cell layer. Our results demonstrate that the canine vomeronasal organ contains a population of cells that expresses several neuronal markers. Furthermore, GAP43 immunoreactivity suggests that the sensory epithelium is neurogenic in adult dogs.     

Epigenetic modification of vomeronasal (V2r) precursor neurons by histone deacetylation

Vomeronasal neurons undergo continuous neurogenesis throughout development and adult life. These neurons originate as stem cells in the apical zone of the lumen of the vomeronasal organ (VNO) and are described as nestin-expressing glia-like progenitor cells (Murdoch and Roskams, 2008). They then migrate horizontally along the basal zone where they differentiate into functional VNO neurons (Kaba et al., 1988). We harvested progenitor cells from the adult VNO and, after 3–6 months of invitro culture, these VNO neurons remained in a stable undifferentiated state expressing nestin, β-tubulin III and vomeronasal type 2 (V2r), but not vomeronasal type 1 (V1r) receptors. Application of histone-deacetylase inhibitors induced development of a neural phenotype that expressed V2r receptors, a down-regulation of nestin expression and no change in any specific genetic markers associated with glial cells. Treatment with valproic acid induced extensive changes in gene expression in the axon guidance pathway. The adult VNO is known to functionally adapt throughout life as a consequence of changes in both a mouse's physiological status and its social environment. These pluripotent cultured neurons may provide valuable insights into how changes in both physiology and environment, exert epigenetic effects on vomeronasal neurons as they undergo continuous neurogenesis and development throughout the life of a mouse.     

A morphological study of the vomeronasal organ and the accessory olfactory bulb in the Korean roe deer, Capreolus pygargus

The vomeronasal organ (VNO) and accessory olfactory bulb (AOB) of the Korean roe deer (Capreolus pygargus) were studied histologically to evaluate their morphological characteristics. Grossly, the VNO, encased by cartilage, has a paired tubular structure with a caudal blind end and a rostral connection through incisive ducts on the hard palate. In the VNO, the vomeronasal sensory epithelium (VSE) consists of galectin-3-positive supporting cells, protein gene product (PGP) 9.5-positive receptor cells, and basal cells. The vomeronasal respiratory epithelium (VRE) consists of a pseudostratified epithelium. The AOB strata included a vomeronasal nerve layer (VNL), a glomerular layer (GL), a mitral/tufted cell layer, and a granular cell layer. All lectins used in this study, including Bandeiraea simplicifolia agglutinin isolectin B4 (BSI-B4), soybean agglutinin (SBA), Ulex europaeus agglutinin I (UEA-I), and Triticum vulgaris wheat germ agglutinin (WGA), labeled the VSE with varying intensity. In the AOB, both the VNL and the GL reacted with BSI-B4, SBA, and WGA with varying intensity, but not with UEA-I. This is the first morphological study of the VNO and AOB of the Korean roe deer, which are similar to those of goats.     

The vomeronasal organ of the South American armadillo Chaetophractus villosus (Xenarthra, Mammalia): anatomy, histology and ultrastructure

The vomeronasal organ (VNO) is a chemoreceptive structure that has not been extensively studied in the Xenarthran order. Tissue samples from the VNO of the armadillo Chaetophractus villosus were prepared for light and electron microscopy. The VNO is located in the anterior part of the base of the nasal septum. It is tubular in shape, ～ 18 mm in length and opens in the rostral region of the nasal cavity and with a blind caudal end. Its lumen is lined by sensory (SE) and nonsensory (NSE) epithelium. The SE shows sensory, supporting and basal cells whereas the NSE contains ciliated and nonciliated secretory cells and basal cells. At the ultrastructural level, the sensory cells appear as bipolar neurons with conspicuous microvilli on their free surface. The supporting cells of the SE contain numerous membrane-bound vesicles in their apical regions. A peculiar feature not found in other mammals, is the presence of concentric whorls of RER cisterns frequently observed in their basal expansions. Infiltrating plasma cells can be detected in the SE basal region close to the dorsal junctional area. This region also exhibits an unusual type of basal cell, probably responsible for the generation of new vomeronasal receptor neurons. The ciliated NSE cells exhibit numerous ovoids or irregularly shaped membranous protrusions projecting from the plasma membrane of the cilia. As far as we know, this is the first study reporting the presence of this feature in ciliated NSE cells. The nonciliated cells are characterised by scarce large secretory granules and apical microvilli. The vomeronasal glands are compound-branched tubuloacinar glands with serous acinar cells. Four types of secretory granules are present. The ducts of these glands reach the lumen in the dorsolateral region between the NSE and SE. Hypolemmal nerve terminals were observed contacting secretory cells. Fenestrated and nonfenestrated capillaries constitute the vascular supply to these glands. Plasma cells, intimately associated with acinar cells, were frequently observed.     

Immunolocalization of VEGF, VEGFR-1 and VEGFR-2 in lung tissues after acute hemorrhage in rats

In treatment of hypovolemia it is important to reestablish normal tissue hemodynamics after fluid resuscitation. Vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFR) have been identified as important in many physiological and pathological processes. In this study, we aimed to investigate the histo-physiological effects of VEGF, VEGFR-1 (flt-1) and VEGFR-2 (KDR/flk-1) in resuscitation with different plasma substitutes on lung tissues after acute hemorrhage in rats. Male Sprague-Dawley rats (n=25) were used in this study. The left femoral vein and artery were cannulated for the administration of volume expanders and for direct measurement of mean arterial blood pressure (MAP) (Power-Lab) and heart rate (HR). Fifteen rats were bled (5 ml/10 min) and infused (5 ml/5 min) with one of three randomly selected fluids: (a) dextran-70 (Macrodex); (b) gelatin (Gelofusine); or (c) physiological saline (PS, 0.9% isotonic saline) solutions. Five rats were bled and none were infused (hypovolemia group) and five rats were untreated as the control group. At the end of the experiment, rats were sacrificed and lung tissues were removed for routine processing and paraffin wax embedding. Sections of tissue were stained with hematoxylin and eosin (H&E) and selected blocks were then prepared for indirect immunohistochemical labeling for anti-VEGF, anti-VEGFR-1 and anti-VEGFR-2 primary antibodies. It was observed that both MAP and HR decreased parallel to blood withdrawn in this time interval. The MAP and HR were restored in the following periods. In the control rats, positive immunoreactivity of VEGF and its receptors (VEGFR-1 and VEGFR-2) were detected in respiratory epithelial cells, respiratory and vascular smooth muscle cells, alveolar cells and endothelial cells. While strong immunoreactivities of VEGF and VEGFR-1 were observed in the hypovolemia group, only moderate immunoreactivity of VEGFR-2 was seen in this group. Moderately strong immunolabeling of VEGF and VEGFR-1 were observed in the dextran-70, gelatin and PS resuscitated groups, whereas only weak immunolabeling of VEGFR-2 was observed in these groups. In summary, the vascular protecting effects of these factors were observed with fluid resuscitation, contributing to the pathophysiological changes seen in hypovolemia.     

The human vomeronasal organ. Part II: prenatal development

During the 20th century, the human vomeronasal organ (VNO) has been controversial regarding its structure, function, and even identity. Despite reports that provide evidence for its presence throughout prenatal and postnatal ontogeny, some studies and numerous textbooks declare its absence in late fetal and postnatal humans. To that end, the present study was designed to establish firmly whether the human VNO is homologous with that of other mammals and whether it degenerates (partially or completely) or persists throughout prenatal development. Fifty human embryos and fetuses (33 d to 32 wk fertilisation age) and 2 neonates were examined by light microscopy. Four embryonic primates (mouse lemurs) were examined for a comparison of VNO embryogenesis. The presence or absence and structural characteristics of the VNO and supporting tissues are described. The first appearance of the VNO was in the form of bilateral epithelial thickenings of the nasal septum, the vomeronasal primordium. The primordia invaginated between 37 and 43 d of age and formed the tubular VNO. The tubular VNO was located dorsally at a variable distance from, but was always spatially separated from the paraseptal cartilages. The mouse lemurs examined in this study and other reports from the literature indicate that the human VNO resembles that of primates having functional VNOs until just after a tubular VNO is formed. Examination of the VNO and adjacent tissues suggested that the VNO may lose receptor cells and corresponding vomeronasal nerves and become a ciliated, pseudostratified epithelium between ～ 12 and 14 wk of age. Our findings indicate the prenatal human VNO goes through 3 successive stages: early morphogenesis, transformation (of the epithelium), and growth. These observations indicated that (1) all embryonic humans develop a vomeronasal organ which is homologous with the VNOs of other mammals, but which has become displaced and highly variable in relative location during embryogenesis; (2) the human vomeronasal organ does not degenerate prenatally, but very likely loses the functional components of the vomeronasal complex of other mammals; and (3) the remnant of the human VNO persists until birth and beyond.     

Expression pattern of VEGFR-2 (Quek1) during quail development

The growth and maintenance of the blood and lymphatic vascular systems is to a large extent controlled by members of the vascular endothelial growth factor (VEGF) family via the tyrosine kinase receptors (VEGFRs) expressed in angioblastic cells. Here, we analyzed the Quek1 (VEGFR-2) expression pattern by whole mount in situ hybridization during quail development. During early embryogenesis, Quek1 expression was detected at the caudal end of the blastoderm and primitive streak and in the head paraxial mesoderm. In somites, expression was observed from HH-stage 9 onwards in the dorsolateral region of both the forming and recently formed somites. During somite maturation, expression persists in the lateral portion of the somitic compartments, the dermomyotome and the sclerotome. Additionally, a second expression domain in the maturing somite was observed in the medial part of the sclerotome adjacent to the neural tube. This expression domain extended medially and dorsally and surrounded the neural tube during later stages. In the notochord, expression was observed from HH-stage 23 onwards. In the limb bud, expression was initiated in the mesenchyme at HH-stage 17. During organogenesis, expression was detected in the pharyngeal arches and in the anlagen of the esophagus, trachea, stomach, lungs, liver, heart and gut. Expression was also seen in feather buds from day 7 onwards. Our results confirm the angiogenic potential of the mesoderm and suggest that VEGFR-2 expressing cells represent multiple pools of mesodermal precursors of the hematopoietic and angiopoietic lineages.     

Olfactory epithelia differentially express neuronal markers

All three olfactory epithelia, the olfactory epithelium proper (OE), the septal organ of Masera (SO), and the vomeronasal organ of Jacobson (VNO) originate from the olfactory placode. Here, their diverse neurochemical phenotypes were analyzed using the immunohistochemical expression pattern of different neuronal markers. The olfactory bulb (OB) served as neuronal control. Neuronal Nuclei Marker (NeuN) is neither expressed in sensory neurons in any of the three olfactory epithelia, nor in relay neurons (mitral/tufted cells) of the OB. However, OB interneurons (periglomerular/granule cells) labeled, as did supranuclear structures of VNO supporting cells and VNO glands. Protein Gene Product 9.5 (PGP9.5 = C-terminal ubiquitin hydrolase L1 = UCHL1) expression is exactly the opposite: all olfactory sensory neurons express PGP9.5 as do OB mitral/tufted cells but not interneurons. Neuron Specific Enolase (NSE) expression is highest in the most apically located OE and SO sensory neurons and patchy in VNO. In contrast, the cytoplasm of the most basally located neurons of OE and SO immunoreacted for Growth Associated Protein 43 (GAP-43/B50). In VNO neurons GAP-43 labeling is also nuclear. In the cytoplasm, Olfactory Marker Protein (OMP) is most intensely expressed in SO, followed by OE and least in VNO neurons; further, OMP is also expressed in the nucleus of basally located VNO neurons. OB mitral/tufted cells express OMP at low levels. Neurons closer to respiratory epithelium often expressed a higher level of neuronal markers, suggesting a role of those markers for neuronal protection against take-over. Within the VNO the neurons show clear apical–basal expression diversity, as they do for factors of the signal transduction cascade. Overall, expression patterns of the investigated neuronal markers suggest that OE and SO are more similar to each other than to VNO.     

Voltage-gated channel properties of epithelial cells in porcine vomeronasal organ

Bipolar vomeronasal sensory neurons (VSNs) in the vomeronasal organ (VNO) are believed to detect pheromones in most mammals. The vomeronasal sensory epithelium (VSE) is composed of VSNs and supporting cells. There are morphological differences in VNOs between species. Many electrophysiological experiments have been performed on rodent VSEs but few on other mammals. We therefore investigated voltage-gated channel properties of cells in the porcine VSE using slice whole-cell voltage-clamp techniques. In immunohistochemical study of the porcine VSE, most PGP9.5-immunoreactive cells were found between the middle and basal region, and negative cells were distributed in the apical to middle region. Depolarizing pulses to epithelial cells from −90 mV produced transient inward Na⁺ channel currents and sustained outward K⁺ channel currents with various amplitudes. The distribution of cells having high and low Na⁺ current densities was mostly consistent with the histological distribution of VSNs and supporting cells, respectively. The half-inactivation voltage of voltage-gated Na⁺ channels in supporting cells was 26 mV more negative than that in VSNs. Voltage-gated K⁺ channel currents in both cell types were suppressed by tetraethylammonium to the same extent. VSNs possessed TTX-sensitive voltage-gated Na⁺ channels and Ni²⁺-sensitive T-type Ca²⁺ channels. These results suggest that the histological distribution of porcine vomeronasal epithelial cells is more similar to the dog and goat than to rodents, and that the electrophysiological characteristics of porcine vomeronasal epithelial cells are similar to those of rodents. It is also suggested that porcine VSNs detecting pheromones generate action potentials through these channels.     

直肠腺癌VEGF-C及VEGFR-3的表达与淋巴转移的关系

目的为探讨癌细胞淋巴管转移机理,观察血管内皮生长因子-C(VEGF-C)和血管内皮生长因子受体-3(VEGFR-3)在直肠腺癌组织及淋巴管的表达。方法取人直肠腺癌手术材料30例,用免疫组化方法检测癌区和癌周正常区VEGF-C和VEGFR-3的表达情况。结果VEGF-C主要表达在直肠腺癌的癌细胞胞浆,VEGFR-3主要在淋巴管内皮细胞有阳性表达,两者在癌区的表达率均高于正常区直肠组织;癌区淋巴管的平均面密度(33.81±5.67)高于正常区平均面密度(20.13±3.27)。结论VEGF-C和VEGFR-3在人直肠腺癌中的过表达,可能与淋巴管增生和扩张,促进癌细胞的淋巴转移有关。     

Pseudocowpox virus encodes a homolog of vascular endothelial growth factor

We have identified a gene encoding a homolog of vascular endothelial growth factor (VEGF) in the Pseudocowpox virus (PCPV) genome. The predicted protein shows 27% amino acid identity to human VEGF-A. It also shows 41 and 61% amino acid identity to VEGFs encoded by orf virus (ORFV) strains NZ2 and NZ7, respectively. Assays of the expressed VEGF-like protein of PCPV (PCPV(VR634)VEGF) demonstrated that PCPV(VR634)VEGF is mitogenic for endothelial cells and is capable of inducing vascular permeability. PCPV(VR634)VEGF bound VEGF receptor-2 (VEGFR-2) but did not bind VEGFR-1 or VEGFR-3. These results indicate that PCPV(VR634)VEGF is a biologically active member of the VEGF family which shares with the ORFV-encoded VEGFs a receptor binding profile that differs from those of all cellular members of the VEGF family. It seems likely that the biological activities of PCPV(VR634)VEGF contribute to the proliferative and highly vascularized nature of PCPV lesions.     

Immunohistochemical expression and distribution of VEGFR-3 in malignant mesothelioma

Homogeneity of mesothelial and lymphatic endothelial cells, express some markers that are presumed to be exclusive of the endothelium was recently reported. This similarity is important to improve the diagnosis and prognosis of malignant mesothelioma (MM). Additionally, some of these markers provide the rationale for specific molecular-targeted novel therapies aimed at MM, an aggressive malignant neoplasm, with an usually dismal prognosis. The goal of our study was to determine the prevalence and expression pattern of VEGF receptor-3 (VEGFR-3) immunoreactivity in MM and whether this immunoreactivity occurs in different phenotypes of this neoplasm. Formalin-fixed and paraffin-embedded samples from 29 MM cases and 5 metastatic carcinomas were immuno-stained for VEGFR-3 according to the streptavidin-biotin-peroxidase complex technique using a primary antibody (Zymed Laboratories, CA, USA) diluted at 1:200. Lymphatic vessels (LV) were outlined mainly in the peripheral area surrounding the neoplasms. Blood vessels were only rarely positive for VEGFR-3 in a pattern easily distinguishable from LV. In 25 out of 29 cases (86.2%) LV were strongly positive for VEGFR-3: 14 cases (48.2%) exhibited positive VEGFR-3 reactivity in malignant cells. Epitheliod MM showed a moderate to intense VEGFR-3 positive reaction in LV from 8 out of 19 cases. Among the other histological subtypes, a positive VEGFR-3 reaction was noted in malignant cells from two cases of transitional and one case of pleomorphic MM. Malignant cells from two out of three biphasic and one out of three sarcomatoid MM were also positive for VEGFR-3. Interestingly, one case of the multicystic subtype was negative for VEGFR-3 in malignant cells and faintly positive in an occasional LV. All cases of metastatic carcinoma were negative for VEGFR-3 in the neoplastic cells. In conclusion, VEGFR-3 was expressed in malignant cells from different subtypes of MM, reinforcing the putative role of this marker as a potential therapeutic target in this group of neoplasia.     

Expression and localization of placenta growth factor and PlGF receptors in human meningiomas

It has previously been suggested that in human brain tumours, endothelial cell proliferation during angiogenesis is regulated by a paracrine mechanism involving vascular endothelial growth factor (VEGF) and its receptors (VEGF receptor 1 and VEGF receptor 2). The mechanism of growth factor up-regulation is based on hypoxic activation of mRNA expression and mRNA stabilization and genetic events, leading to an increase of growth factor gene expression. The role of the other newly discovered VEGF family members with a high specificity for endothelial cells in the pathogenesis of glial neoplasms is unknown. To investigate which other members of the VEGF family are overexpressed in human brain tumours, the mRNA levels of placenta growth factor (PlGF), VEGF-A, and VEGF-B genes were determined by northern blot analysis in surgically obtained human meningiomas. In the 16 meningiomas examined, the mRNA for PlGF was highly expressed in four tumours and VEGF-A mRNA was highly abundant in three tumour samples. There was no close correlation between PlGF mRNA levels and VEGF-A expression levels. VEGF-B mRNA was abundantly expressed in all tumour samples at uniform levels. In a PlGF-positive tumour sample, immunoreactive VEGFR-1 and VEGFR-2 were detected in endothelial cells of the blood vessels. PlGF protein was detectable in most but not all capillaries of the tumour. PlGF is thus highly up-regulated in a subset of human meningiomas and may therefore have functions, in some tumour vessels, connected to endothelial cell maturation and tube formation. These findings suggest that PlGF, in addition to VEGF-A, may be another positive factor in tumour angiogenesis in human meningiomas. Copyright © 1999 John Wiley & Sons, Ltd.     

Immunohistochemical Expression of Vascular Endothelial Growth Factor and Vascular Endothelial Growth Factor Receptor in Canine Cutaneous Fibrosarcomas

The expression of five markers associated with tumour angiogenesis, proliferation and apoptosis was studied in 24 canine cutaneous fibrosarcomas. Tumours were assigned histological grades and were immunohistochemically evaluated for the expression of vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptor-2 (VEGFR-2). Additionally, intra-tumour microvessel density (iMVD) was assessed by immunohistochemical labelling for expression of von Willebrand factor (vWf) and tumour proliferation index (PI) was measured following labelling of Ki-67 antigen. Finally, tumour apoptotic index (AI) was determined by application of the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP end-labelling method (TUNEL). VEGF and VEGFR-2 expression were detected in 22/24 (92%) and 24/24 (100%) of fibrosarcomas, respectively. There was correlation between VEGF and VEGFR-2 expression (r = 0.51) and between histological grade and PI (r = 0.82). A significant difference in PI between tumours of different histological grade was found (P < 0.05). The median PI in grade 2 and 3 tumours (30.6 and 54.7, respectively) was significantly higher than in grade 1 tumours (6.4). Therefore, only PI correlates significantly with the histological grade of canine cutaneous fibrosarcomas. The potential for autocrine activity for VEGF exists in canine cutaneous fibrosarcomas, as VEGF and VEGFR-2 expression was found in most tumours.     

Vascular endothelial growth factor receptor 3 directly regulates murine neurogenesis

Neural stem cells (NSCs) are slowly dividing astrocytes that are intimately associated with capillary endothelial cells in the subventricular zone (SVZ) of the brain. Functionally, members of the vascular endothelial growth factor (VEGF) family can stimulate neurogenesis as well as angiogenesis, but it has been unclear whether they act directly via VEGF receptors (VEGFRs) expressed by neural cells, or indirectly via the release of growth factors from angiogenic capillaries. Here, we show that VEGFR-3, a receptor required for lymphangiogenesis, is expressed by NSCs and is directly required for neurogenesis. Vegfr3:YFP reporter mice show VEGFR-3 expression in multipotent NSCs, which are capable of self-renewal and are activated by the VEGFR-3 ligand VEGF-C in vitro. Overexpression of VEGF-C stimulates VEGFR-3-expressing NSCs and neurogenesis in the SVZ without affecting angiogenesis. Conversely, conditional deletion of Vegfr3 in neural cells, inducible deletion in subventricular astrocytes, and blocking of VEGFR-3 signaling with antibodies reduce SVZ neurogenesis. Therefore, VEGF-C/VEGFR-3 signaling acts directly on NSCs and regulates adult neurogenesis, opening potential approaches for treatment of neurodegenerative diseases.