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Background . The immunophenotypes and potential excretory function of human mesonephros are not well studied. Methods . Five mesonephros specimens of human embryos from the 6 th to 10 th weeks of gestation were stained with immunohistochemical markers. Results . PAX8 was universally expressed in all renal tubules, while α-methyacyl-CoA racemase (AMACAR) was positive in proximal tubules and GATA3 was positive in distal tubular mesonephric structures. At the 8th weeks of gestation, the mesonephric glomeruli were characterized by opened glomerular capillary loops with Periodic Acid Schiff (PAS)-positive glomerular basement membranes and GATA3-positive mesangial-like cells. By the 8 th week, proximal tubules showed PAS-positive brush borders, indicating reabsorption capacity, and the proximal tubules also demonstrated positivity with kidney injury molecule-1 (KIM-1), representing tubular response to injury. Conclusion . Our overall findings show detailed phenotypes of the glomerular and tubular structures of the mesonephros and indicate that at the 8 th week of gestation, the mesonephros may carry out temporary excretory function before metanephros becomes fully functional.

Controversies regarding the development of the mammalian infrahepatic inferior caval and azygos veins arise from using topography rather than developmental origin as criteria to define venous systems and centre on veins that surround the mesonephros. We compared caudal-vein development in man with that in rodents and pigs (rudimentary and extensive mesonephric development, respectively), and used Amira 3D reconstruction and Cinema 4D-remodelling software for visualisation. The caudal cardinal veins (CCVs) were the only contributors to the inferior caval (IVC) and azygos veins. Development was comparable if temporary vessels that drain the large porcine mesonephros were taken into account. The topography of the CCVs changed concomitant with expansion of adjacent organs (lungs, meso- and metanephroi). The iliac veins arose by gradual extension of the CCVs into the caudal body region. Irrespective of the degree of mesonephric development, the infrarenal part of the IVC developed from the right CCV and the renal part from vascular sprouts of the CCVs in the mesonephros that formed 'subcardinal' veins. The azygos venous system developed from the cranial remnants of the CCVs. Temporary venous collaterals in and around the thoracic sympathetic trunk were interpreted as 'footprints' of the dorsolateral-to-ventromedial change in the local course of the intersegmental and caudal cardinal veins relative to the sympathetic trunk. Interspecies differences in timing of the same events in IVC and azygos-vein development appear to allow for proper joining of conduits for caudal venous return, whereas local changes in topography appear to accommodate efficient venous perfusion. These findings demonstrate that new systems, such as the 'supracardinal' veins, are not necessary to account for changes in the course of the main venous conduits of the embryo., (© 2015 Anatomical Society.)

According to the classical ladder theory, the mesonephric arteries (MAs) have a segmental arrangement and persist after regression of the mesonephros, with some of these vessels becoming definitive renal arteries. To avoid interruption of blood flow, such a vascular switching would require an intermediate stage in which two or more segmental MAs are connected to a definitive renal artery. To examine developmental changes, especially changes in the segmental distribution of MAs, we studied serial paraffin sections of 26 human embryos (approximately 5-7 weeks). At 5-6 weeks, 1-2 pairs of MAs ran anterolaterally or laterally within each of the lower thoracic vertebral segments, while 2-5 pairs of MAs were present in each of the lumbar vertebral segments, but they were usually asymmetrical. The initial metanephros, extending along the aorta from the first lumbar to first sacral vertebra, had no arterial supply despite the presence of multiple MAs running immediately anterior to it. Depending on increased sizes of the adrenal and metanephros, the MAs were reduced in number and restricted in levels from the twelfth thoracic to the second lumbar vertebra. The elimination of MAs first became evident at a level of the major, inferior parts of the metanephros. Therefore, a hypothetical arterial ladder was lost before development of glomeruli in the metanephros. At 7 weeks, after complete elimination of MAs, a pair of symmetrical renal arteries appeared near the superior end of the metanephros. In conclusion, the MAs appear not to persist to become a definitive renal artery., (Copyright © 2015 Elsevier GmbH. All rights reserved.)

Blood flow promotes emergence of definitive hematopoietic stem cells (HSCs) in the developing embryo, yet the signals generated by hemodynamic forces that influence hematopoietic potential remain poorly defined. Here we show that fluid shear stress endows long-term multilineage engraftment potential upon early hematopoietic tissues at embryonic day 9.5, an embryonic stage not previously described to harbor HSCs. Effects on hematopoiesis are mediated in part by a cascade downstream of wall shear stress that involves calcium efflux and stimulation of the prostaglandin E2 (PGE2)-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signaling axis. Blockade of the PGE2-cAMP-PKA pathway in the aorta-gonad-mesonephros (AGM) abolished enhancement in hematopoietic activity. Furthermore, Ncx1 heartbeat mutants, as well as static cultures of AGM, exhibit lower levels of expression of prostaglandin synthases and reduced phosphorylation of the cAMP response element-binding protein (CREB). Similar to flow-exposed cultures, transient treatment of AGM with the synthetic analogue 16,16-dimethyl-PGE2 stimulates more robust engraftment of adult recipients and greater lymphoid reconstitution. These data provide one mechanism by which biomechanical forces induced by blood flow modulate hematopoietic potential., (© 2015 Diaz et al.)

In a previous study, we underlined the functional role of the TPO receptor, Mpl, in the establishment of definitive mouse hematopoiesis, by demonstrating that the lack of Mpl led to a delayed production of definitive hematopoietic cells in the aorta-gonad-mesonephros (AGM) region, and resulted in the production of hematopoietic stem cells (HSCs) with an impaired activity at E11.5. In order to more accurately estimate the role of Mpl during generation of HSCs in the aorta, we performed an analysis of these AGMs at the time of the first HSC emergence (E10.5). Our results indicated that while Mpl-/- AGMs were found to contain more hematopoietic cells (HC) than C57Bl6 AGMs at E10.5, a defect in the expansion process of the HC/HSCs was detected in explant cultures of these AGMs, likely due to an increased apoptosis of these cells. To determine the molecular mechanisms by which invalidation of Mpl receptor affects the temporal distribution and expansion of HC/HSCs in the AGM, a study of the transcription level of of Mpl target genes was conducted. Expression of Runx1, a master transcription factor for the formation of hematopoietic progenitor (HP) cells and HSCs from the vasculature, as well as expression of Meis1 and HoxB4, known to play a role in self-renewal and expansion of HSCs, were found to be down regulated in E10.5 Mpl-/- AGMs. Our data indicate that Mpl is an active player during the first steps of definitive hematopoiesis establishment through direct regulation of the expression of transcription factors or genes important for the self-renewal, proliferation and apoptosis of HSCs.

The E11.5 aorta-gonad-mesonephros (AGM) region is a site of hematopoietic stem cell (HSC) development prior to colonisation of the embryonic liver. The generation of HSCs in the embryo starting from E11 is very rapid. Here, we have assessed hematopoietic development in the AGM region during E11 at precise somitic ages. Although the numbers of committed hematopoietic precursors fluctuate throughout the day, the repopulation activity in the AGM region noticeably increases from mid (44 s.p.) to end (48 s.p.) day 11 of gestation. While prior to mid day 11 two thirds of AGM regions contain no definitive HSCs, shortly prior to liver colonisation, all older day 11 embryos contain definitive HSC. Nevertheless, all E11 AGM regions even at early somitic stages have the capacity to expand numbers of definitive HSCs ex vivo. Quantitative anatomical analysis confirmed preferential localization of intra-aortic clusters (IACs) to the ventral domain of the dorsal aorta during entire day 11 of development. No clear correlation was established between IAC numbers and the presence of definitive HSCs.

The spatial and temporal pattern of the appearance of the fibroblast growth factor proteins (FGF-8 and FGF-10), the bone morphogenetic proteins (BMP-2/4 subfamily and BMP-7) and the vascular endothelial growth factor protein (VEGF) was investigated in the human mesonephros and metanephros of the 5-9 week-old conceptuses. In the mesonephros, both FGF's and BMP's were found in all structures and their expression slightly decreased in the early fetal period. VEGF positivity appeared in all mesonephric structures, and increased in the fetal period coincidently with formation of the mesonephric blood vessel network. In the metanephros, FGF-8 first appeared only in the metanephric mesenchyme, but from the 7th week on, its reactivity increased and spread to other metanephric structures. FGF-10 positive cells appeared in all metanephric structures already in the 5th week, and slightly intensified with progression of development. Cell survival and nephrogenesis in the permanent kidney might be associated with the appearance of both growth factors. Both BMP-2/4 and BMP-7 displayed a similar pattern of reactivity in all metanephric structures, and their reactivity intensified with advancing development. Alterations in their pattern of appearance might lead to the formation of small and dysplastic kidneys. Already in the earliest developmental stages, VEGF protein appeared in all metanephric structures. At later stages, VEGF showed more intense reaction in the collecting system than in the differentiating nephrons and interstitium. Due to VEGF involvement in vasculogenesis and angiogenesis, abnormal VEGF appearance might lead to impaired formation of the blood vessel network in the human permanent kidney.

Hemangioblast, a precursor possessing hematopoietic and endothelial potential, is identified as the blast colony-forming cell in the murine gastrulating embryos (E7.0-E7.5). Whether hemangioblast exists in the somite-stage embryos is unknown, even though hemogenic endothelium is regarded as the precursor of definitive hematopoiesis in the aorta-gonad-mesonephros (AGM) region. To address the issue, we developed a unique three-step assay of high proliferative potential (HPP) precursors. The AGM region contained a kind of HPP precursor that displayed hematopoietic self-renewal capacity and was able to differentiate into functional endothelial cells in vitro (i.e., incorporating DiI-acetylated low-density lipoprotein, expressing von Willebrand factors, and forming network structures in Matrigel). The clonal nature was verified by cell mixing assay. However, the bilineage precursor with high proliferative potential-the HPP-hemangioblast (HA)-was not readily detected in the yolk sac (E8.25-E12.5), embryonic circulation (E10.5), placenta (E10.5-E11.5), fetal liver (E11.5-E12.5), and even umbilical artery (E11.5), reflective of its strictly spatial-regulated ontogeny. Expression of CD45, a panhematopoietic marker, distinguished hematopoietic-restricted HPP-colony-forming cell from the bipotential HPP-HA. Finally, we revealed that basic fibroblast growth factor, other than vascular endothelial growth factor or transforming growth factor-beta1, was a positive modulator of the HPP-HA proliferation. Taken together, the HPP-HA represents a novel model for definitive hemangioblast in the mouse AGM region and will shed light on molecular mechanisms underlying the hemangioblast development. Disclosure of potential conflicts of interest is found at the end of this article.

beta-Catenin is a structural component of adherens junctions, a regulator of the Wnt signalling pathway and a transcriptional co-activator with a key role in vascular patterning. The avian mesonephros is a transitory embryonic kidney that is used in the study of vascular development and degeneration. Here we examine beta-catenin expression in this model during vascular development and degeneration. Quail embryos with developing or degenerating mesonephros were studied, on day 6 (30HH) or day 11 of incubation (40HH), respectively. QH1 whole mounts of developing mesonephros revealed numerous angioblast-like cells situated in the paramesonephric duct that seem to invade the mesonephros. Although these cells did not express beta-catenin, the surrounding periductal mesenchymal cells translocated high levels of beta-catenin into the nucleus. In contrast, degenerating mesonephros were devoid of angioblast-like cells and beta-catenin was lower than in the developing mesonephros. beta-Catenin was significantly reduced in the glomerular capillary tuffs, indicating that it was particularly down-regulated in the vascular system. No sex-related differences in beta-catenin expression were observed in degenerating mesonephros. Furthermore, two special populations of glomerular and peritubular endothelial cells were observed in degenerating mesonephros: one translocating beta-catenin into the nucleus and the other in apoptosis that did not translocate it. In conclusion, our results indicate that the paramesonephric duct is a potential new vasculogenetic pathway, and suggest that beta-catenin plays a role in the fate of mesonephric endothelial cells.

The blood vessels of the bovine mesonephros - a corrosion cast study Using scanning electron microscopy plastoid corrosion cast specimens of the mesonephros of 30-45-day-old bovine foetuses were investigated. The studies showed that the blood vessels of the mesonephros are organized in two separate networks similar to these of the teleosts and amphibians. The glomerula mesonephrica originate from the aorta implying that they belong to the high pressure system. The tubuli mesonephrici are surrounded by a filamentous independent capillary network which is shown for the first time in these studies. This capillary network originates from the caudal cardinal veins and represents a mesonephric portal system with a relatively low blood pressure. The blood from the two capillary networks diverts into terminal veins and into the subcardinal veins. The questions of haemodynamics in the mesonephros are discussed.

Embryonic tissues, in common with other tissues, including tumours, tend to develop a substantial vasculature when transplanted onto the chorioallantoic membrane (CAM). Studies conducted to date have not examined in any detail the identity of vessels that supply these grafts, although it is known that the survival of transplanted tissues depends on their ability to connect with CAM vessels supplying oxygen and nutrients. We grafted the mesonephros, a challenging model for studies in vascular development, when it was fully developed (HH35). We used reciprocal chick-quail transplantations in order to study the arterial and venous connections and to analyse the cell invasion from the CAM to the organ, whose degeneration in normal conditions is rapid. The revascularization of the grafted mesonephros was produced by the formation of peripheral anastomoses between the graft and previous host vasculatures. The assembly of graft and CAM blood vessels occurred between relatively large arteries or veins, resulting in chimeric vessels of varying morphology depending on their arterial or venous status. Grafts showed an increased angiogenesis from their original vasculature, suggesting that the normal vascular degeneration of the mesonephros was partially inhibited. Three types of isolated host haemangioblast were identified in the mesonephros: migrating angioblast-like cells, indicating vasculogenesis, undifferentiated haematopoietic cells and macrophages, which might have been involved in the angiogenesis. Tomato lectin was found to bind activated macrophages in avian embryos.

Morphological symptoms of mesonephric kidney damage were analysed in chick embryos treated with nephrotoxic agents--CDDP or DBE. The drugs were administered intraamniotically on ED 3 at doses 0.03 and 0.3 microg CDDP or 100 and 300 microg DBE per embryo. Body weight and absolute and relative measures of the mesonephroi (length, weight and form) were evaluated on ED 10. The higher doses of both agents affected the mass of this organ significantly. Simultaneously, a dose-dependent increase of renal malformations was detected in treated embryos, while the incidence of gross and cardiovascular defects was low (DBE) or absent (CDDP). Together with less pronounced effects on the total body growth, the results gave evidence for a higher sensitivity of the mesonephros to toxic insult when compared to the whole organism. A direct cytotoxic effect multiplied by concomitant injury of blood supply seemed to be the main cause of CDDP nephrotoxicity. In the case of DBE, damage to the mesonephros was probably associated with a primary impairment of the vascular network. The chick embryo in ovo provides a promising system for the assessment of nephrotoxic effects induced by prospective therapeutic agents and environmental contaminants during the prenatal period.

Vasculogenesis and angiogenesis are involved in a coordinated program for the development of the mesonephric subcardinal venous plexus of quail embryo. Vasculogenesis occurs between days 3 and 4 of incubation, while angiogenesis takes place from day 5 to day 7. Examination of vascular corrosion casts and whole mounts, and tissue sections labelled with specific markers to hemangioblast lineage (QH1, LEP100 and AcPase activity), allowed us to distinguish six phases in the formation of subcardinal plexus. (1) Appearance of isolated angioblast-like cells where the subcardinal plexus will form. (2) Alignment of angioblast-like cells into cellular strands. (3) Formation of compact vascular cords by association of angioblast-like strands. (4) Polygonal interconnection of vascular cords to constitute the primary subcardinal plexus. In this stage, isolated angioblast-like cells were present inside inter-vascular spaces. (5) The splitting of primary inter-vascular spaces by angiogenic sprouts to form secondary subcardinal plexus (outward angiogenesis). Isolated angioblast-like cells were not present in this stage. (6) Expansion of the secondary subcardinal plexus by insertion of slender transcapillary tissue pillars (inward angiogenesis) and angiogenic sprouts. We also describe three morphogenetic gradients during the development of the subcardinal plexus: ventral-to-dorsal, cranial-to-caudal and lateral-to-medial.

Background: In the chick embryo, both mesonephros and metanephros have a renal portal system. The classical literature gives uncertain answers about the development and degeneration of the meso- and metanephric portal venous system. Some mesonephric vessels present angiogenic processes to colonize the metanephros, while others show signs of degeneration and disappear together with the mesonephros. The adult avian kidney has a conspicuously placed valve, the renal portal valve. The development of this functionally important renal portal valve has not yet been studied in detail., Methods: Scanning electron microscopy of vascular corrosion casts has been used in this study. Strong mesonephric degeneration as well as metanephric growth and maturation occur in the developmental stages selected for this investigation (7.5, 9, 11, 14, and 21 days of incubation)., Results: The mesonephric afferent venous system in the chick embryo is supplied by two vessels, the posterior and the anterior mesonephric portal veins. The posterior mesonephric portal veins show a similar pattern to the anuran (amphibian) kidney. The anterior mesonephric portal vein has not previously been described. Constrictions were found in this vessel, a probable sign of subsequent degeneration. The metanephric afferent venous system is also supplied by two vessels: the caudal and cranial metanephric portal veins. The caudal metanephric portal vein is derived from the postcardinal vein. The cranial metanephric portal vein grows independently throughout the development of the mesonephric vascular system. It is connected to the vertebral venous sinus already at the beginning of its development. The renal portal valve first appears as a capillary network that communicates with the developing afferent and efferent metanephric venous systems. This capillary network later develops to a venous valve. The metanephric afferent venous system shows typical angiogenic signs in corrosion cast, such as nodular protrusions, holes, and enlarged vessels., Conclusions: The postcardinal vein first supplies only the mesonephric tissue as a portal vessel. Then it becomes a common source for both kidney generations. Finally it supplies only the metanephric tissue with venous blood. However, two independent vessels were found to supply the cranial renal regions: the anterior mesonephric portal vein and the cranial metanephric portal vein.

Background: The general morphology of the mesonephric and metanephric arteries in chicken embryos has already been described previously. Moreover, the general basis of glomerulogenesis has also been established. However, the degeneration of the mesonephric vascular system, and especially glomerular degeneration, have not been well established yet. Also the morphology of the metanephric angiogenic buds has not been studied yet., Methods: Scanning electron microscopy of vascular corrosion casts and critical point dried specimens as well as light microscopy of serially sectioned material has been used in this study. Mesonephric degeneration coincides in time with metanephros growth and maturation in the developmental stages of chicken embryos chosen for this investigation (7.5, 9, 11, and 14 days of incubation)., Results: The arterial system of the mesonephros in embryonic chicken is similar to that of the anuran kidney, as described in the literature. The morphology of the degenerating mesonephric glomeruli shows that the glomerular capillaries are more thick, tortuous, and numerous than those in normal glomeruli. The podocytes also show degeneration. The arterial system of the metanephros grows directly from the aorta and from the mesonephric arterial system. During these stages of rapid growth, the metanephros shows angiogenic buds. These angiogenic buds can be either pointed or round blind endings., Conclusions: The distribution and topography of the mesonephric and metanephric arteries is in general accordance with the literature. The process of glomerular degeneration in the mesonephros seems similar to glomerular senescence in man but is different from that of the aged rat glomeruli. The round angiogenic buds observed in the metanephros resemble tumoral angiogenic buds in some aspects. However, both angiogenesis and the degenerative phenomena are part of the normal developmental process. Consequently, the involved mechanisms are probably under sole genetic control. The system studied here offers therefore the possibility to study vascular growth and degeneration on the same model in physiological conditions without application of vasoactive or pathological agents.

The purpose of the present study was to describe geometry of branching, topography and sources of blood vessels supplying the mesonephros of three different species of amphibia and to infere from the results their common and species-specific features. The detailed information on arrangement of portal and arterial blood supply to mesonephros is necessary for better understanding of the conditions under which these kidneys function. For this reason the blood vessels of the mesonephros of adult Hynobius Keyserlingi, Triturus vulgaris and Rana temporaria were injected with stained gelatin. The urodelian kidneys were shown to be supplied by numerous segmental arteries. Three types of arterial supply to glomeruli were distinguished: those mediated by short, medium and long branches, which create different physical conditions for the glomerular filtration in pertinent nephrons--as the blood pressure necessarily decreases with the increasing distance from the aorta. The tributaries to the renal portal vein were identified in all three species as the veins draining hind limbs, tail, pelvis and body wall. It is being suggested that the purpose of this large drainage area is to secure a sufficient volume of blood to the peritubular network.

The paper describes an unusual angiographic pattern within the context of stenosis of the pyeloureteral union and giant hydronephrosis of the adult, with a presumed congenital origin. From the embryonic point of view we discuss the remote possibility of this being a real "agenesis" of the renal artery with no agenesis of the kidney, irrigated by multiple segmental arteries following the characteristic metamerism of the mesonephros. On the other hand, it seems more consistent to explain such vascular pattern as an image of severe atrophy, in correlation with the hydronephrosis advanced stage, and not as a primary congenital defect. However, the interaction of both factors (congenital-malformative and acquired-involutive) could be more overlapped than what initially appears to be.

Vascularization of the pig mesonephros was investigated in embryos 5-8 cm in length. Vascular injections with microfil were cleared and dissected; corrosion casts were studied under the scanning electron microscope (SEM). Perfusion-fixed tissue was used for SEM and transmission electron microscope (TEM) studies, including freeze-fracture specimens. The branches of one mesonephric artery carry up to 15 glomeruli. Several glomeruli occupy the same arterial branch, with very short afferent arterioles proper. The efferent vessels, frequently 2-5, leave the extensive vascular pole opposite the entering arteriole and split into peritubular capillaries radiating towards the superficial veins. These capillaries form vascular regions in the shape of flattened pyramids. Along its course, one nephron is supplied by vessels derived from 4-7 glomeruli. The nephrons have less vascular contact than in the definitive kidney. The ultrastructure of the single mesonephric vessels matches the metanephric counterparts. Epithelioid cells with renin granules are common in afferent arterioles, larger arteries, and efferent vessels. The lobulated glomeruli are up to 750 micron long and flattened, showing the usual features of podocytes, mesangial cells, and an attenuated endothelium with fenestrations between 50 and 250 nm. It partially retains its own basement membrane. There is no proximal mesangium.