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PubMed1984

Vascularization and glomerular ultrastructure in the pig mesonephros.

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.

Animals↗
PubMed1978

Scanning electron microscopy of the renal corpuscle of the mesonephros in the lamprey, Entosphenus japonicus Martens.

The renal corpuscle of the lamprey mesonephros was studied under the scanning electron microscope. Bowman's capsules with individual spaces are chockshaped sacs closely packed together along a medial artery. The lateral walls of the capsules are apposed to those of neighbouring capsules. Glomerular capillaries from the medial artery extend radially between the apposed walls of neighbouring Bowman's capsules. Bulgings of capillaries into the capsular space are associated with mesangial folds of the capsular epithelium. The transitional zone of the visceral layer with podocytes and the parietal layer of squamous epithelium is bounded by linearly arranged rod-shaped epithelial cells. Apertures of the urinary tubule are lined by cells equipped with a fascicle of cilia.

Animals↗
PubMed1993

Immunohistochemical localization of transforming growth factor-alpha and epidermal growth factor-receptor in the mesonephros and metanephros of the chicken.

Transforming growth factor-alpha (TGF-alpha) is a polypeptide related to epidermal growth factor (EGF). Both bind to EGF-receptor (EGF-R) to carry out their function in a variety of tissues and cell lines. Several studies have shown their presence in mammalian kidney, however, nothing has to date been stated concerning their existence in avian kidney. Expression of TGF-alpha and EGF-R is reported here for the first time during the development of the chicken kidney. Using immunohistochemical techniques, we identified a TGF-alpha (but not EGF) in mesonephric distal tubule cells from day 8 to day 20 of embryonic development and in metanephric distal tubule cells from day 14 of embryonic development to the adult. The histochemical characteristics of these cells and their histological localization suggest that they may be the "principal cells" of the distal tubules. Similarly, EGF-R was found in mesonephric proximal tubule cells from day 7 to day 18 of embryonic development and in metanephric proximal tubule cells from day 13 of embryonic development up to adult stages. The coexistence of both TGF-alpha and EGF-R from the onset of development of mesonephros and metanephros supports their possible role in mechanisms of proliferation and differentiation of the cells of these organs.

Animals↗
PubMed1983

The pig mesonephros. I. Enzyme histochemical observations on the segmentation of the nephron.

Cryostat sections from the mesonephros of various pig embryos with a crown-rump-length of between 17 and 95 mm were used for light microscopical assays of acid hydrolases (acid phosphatase, beta-D-N-acetylglucosaminidase, beta-D-glucuronidase), oxidoreductases (succinic dehydrogenase, NADH- and NADPH-tetrazolium reductase) and adenosine triphosphatases (Mg2+- and Na+--K+-ATPase). Our main intention was to distinguish more accurately between the different parts of the pig's nephron, which is exceedingly long and coiled. The proximal tubule, that exhibits a high activity for acid phosphatase but none in beta-D-glucuronidase incubations, shows no subsegmentation apart from a stronger reaction of its initial segment that was apparent in three of our assays. In the distal tubule, a preattachment convolution, an attachment zone, and a postattachment coil can be discriminated by a synopsis of all histiograms. The beginning of the collecting tubule is situated in the middle of the organ and not at its dorsal face as was previously believed. Up to three different segments can be discriminated in the collecting tubule. The distal and the collecting tubule harbor on ouabain-sensitive Na+--K+-ATPase activity which decreases considerably towards the Wolffian duct. The enzymatic maturation of the mesonephric pig nephron is almost completed in 17 mm embryos.

Adenosine Triphosphatases↗
PubMed1982

Development of the early human ovary and role of the mesonephros in the differentiation of the cortex.

The morphogenesis of the primary gonadal differentiation, of the sexual differentiation and the growth of the ovarian cortex during its early period have been studied on 10 human embryos between 12 and 95 mm CR-length. Semithin sections of glutaraldehyde-OsO4-fixed and plastic-embedded material were used to demonstrate the structural events on a cellular level. The primary gonadal blastema within the genital ridge is formed by two types of somatic cells; cells segregated from the mesonephros and cells of the proliferating coelomic epithelium. The two types of cells show a tendency to intermingle and they enclose the immigrating primordial germ cells. In the female gonad the indifferent period terminates between day 40 and 42 of ovulation age (20 to 23 mm CR-length). Between day 40 and 50 the blastemal content of the indifferent gonad is remodelled and an ovarian cortex differentiates. Cellular strands extending from the primary blastema and strands from the superficial blastemal layer contribute to the formation of the cortex. Within the newly formed medulla, remnants of the disintegrating primary blastema differentiate into medullary cords. Cells of mesonephric origin which invade the growing cortex via the rete blastema interact with cells deriving from the superficial epithelium, and both exert their opposite influence on the germ cells. While female sexual differentiation is characterized by failure of the dark mesonephric cells to completely penetrate the gonadal blastema, the morphogenetic process resulting in the formation of the ovarian cortex shows a strong invasion of the cortex by the dark mesonephric cells. Dark cells advance at the most superficial layer of the cortex and increase in number at the deeper level of the cortex. Onset of oogonial proliferation and meiotic prophase seems to depend on the numerical proportion between the activating dark and the inhibiting light supporting cells.

Cell Differentiation↗
PubMed1985

The pig mesonephros. III. Distal tubule, collecting tubule, and Wolffian duct: SEM- and TEM-studies.

The ultrastructure of the distal and collecting tubules of mature pig mesonephroi (41st gestational day) was studied in perfusion-fixed embryos. In the distal tubule, the three subsegments postulated on the basis of enzyme histochemistry show only minor differences of their luminal surfaces, mostly of cell size. TEM photographs reveal a single cell type with interdigitating basolateral processes, frequently flattened to 30-120 nm lamellae devoid of organelles. Larger interdigitating processes harbor vertically oriented mitochondria in the form of indented plates. The macula densa cells are small, do not interdigitate, and have distended intercellular spaces. The collecting tubule starts with a dorsal convolution, in which intercalated cells (with apical microfolds and numerous mitochondria) occur in addition to interdigitating cells. Further down this segment, the interdigitating cells are gradually replaced by principal cells characterized by interlocking lateral microvilli, basal infoldings, and relatively few organelles. Intercalated cells extend into the Wolffian duct. Although the pig mesonephros has the most differentiated nephron of the mammals studied so far, with metanephros-like cells, its intrinsic urinary concentrating capacity appears to be low in view of its vascularization pattern and nephron architecture.

Animals↗
PubMed1975

Histochemical localization of Mg2+-Na+-K+-adenosine triphosphatase in different stages of the sheep mesonephros.

Mesonephroi of sheep embryos ranging from 12 to 100 mm C.R. length were examined for the occurrence and localization of transport-ATPase. Native cryostat sections were incubated according to the technique of Guth and Albers for demonstrating the nitrophenylphosphatase activity of Mg2+-Na+-K+-adenosine triphosphatase. The basal cytoplasm of the collecting tubule of the narrow segment of the distal tubule exhibit strong activity, the wide segment of the distal tubule is moderately active. Glomeruli, proximal tubule, and Wolffian duct remain unstained. The basal labyrinths of the reactive nephron segments are believed to be the sites of a Na+-K+ exchange pump. In mature and regressing mesonephroi, the findings fully agree with biochemical data; in maturating mesonephroi, whose basal labyrinth is not yet fully established, the biochemical assay proves to be more sensitive. The specifity of the reaction was ascertained by diverse inhibitors and activating ions. The localization of Mg2+-ATPase is different to the above mentioned reaction pattern, as it shows moderate activity in the proximal tubule, too (mature mesonephros). Mesonephroi of very young embryos exhibit strongest Mg2+-ATPase activity in the proximal tubule; here the distal and collecting tubule stain only moderately.

Adenosine Triphosphatases↗
PubMed1984

Renin immunohistochemistry in the mesonephros and metanephros of the pig embryo.

The occurrence and distribution of renin was investigated in meso- and metanephric kidneys of pig embryos in various gestational stages. The immunohistochemical peroxidase-antiperoxidase-method (PAP) was used on paraffin sections after application of an antiserum against mouse renin which cross reacts with pig renin. Renin immunoreactivity was already found in the mesonephros of 21 day pig embryos (crown-rump(CR)-length 12 mm) with the strongest reaction in the media of the juxtaglomerular afferent arteriole. Efferent vessels, mesonephric arteries, and the aortic wall also contained scattered renin-positive cells. In the definitive kidney, renin was not detected prior to the 25 mm CR-length-stage. In 45 mm embryos, immunocytochemical staining was observed not only in the media of kidney arteries and arterioles, but also in proximal tubules after pinocytic absorption of filtered renin. TEM-studies revealed that the media of both the mesonephric and the developing metanephric arteries and arterioles contains epithelioid cells whose ultrastructure is very similar to that of renin-producing cells in the adult organ. The observed distribution of renin-producing cells along the entire renal arterial tree points to the possibility that the major function of the renin-angiotensin system in the fetal animal is to participate in the stabilization of renal perfusion pressure.

Animals↗
PubMed2005

Early development of the human mesonephros.

The mesonephrogenic cord disintegrates into approximately 35-40 provesicular cell masses which are in close contact with the mesonephric (Wolffian) duct (WD) on their lateral side. Here, the epithelium of the WD is columnar and shares a common basal lamina with the provesicular cell masses. This in turn gives rise to a sickle-shaped pseudostratified epithelium. The concavity of the sickle is filled by spherical cells, the transition of which into the surrounding connective tissue is continuous. The sickle is transformed into a distillation flask and becomes separated from the mesonephric duct while the spherical cells maintain a connection to it by a-for the time being-solid outlet pipe. The columnar epithelium of the mesonephric duct becomes a multilayered cone, whose surface is in contact with the outlet tube. Shortly after, a continuous lumen is formed in the cone and the outlet pipe which is delimited by cells becoming columnar and forming a basal lamina. The epithelial anlage of the nephron is clearly separated from the surrounding mesenchyma by these processes. The flask eventually becomes a corpusculum, the outlet pipe a secretory (proximal) as well as collecting tubule, and the cone of the mesonephric duct a mesoureter. The various sections display differentially differentiated epithelia that are clearly distinct from each other. The mesoureter behaves differently during differentiation of epi- and paragenitale: in the epigenitale, it is short and runs into the collecting tubules of the nephrons at the lateral side of the convolved tubules, whereas a long mesoureter crosses the dorsal side of the convolved tubules and joins the corresponding collecting tubules at the far end of the mesonephros in the paragenitale.

Crown-Rump Length↗
PubMed2006

Stromal cell lines from the aorta-gonado-mesonephros region are potent supporters of murine and human hematopoiesis.

OBJECTIVE: The hematopoietic system is nurtured by a supportive stroma environment allowing maintenance and differentiation of hematopoietic stem cells (HSC). However, only a limited number of these stromal cell clones support hematopoiesis in the absence of cytokine supplementation. So far, only two bone marrow-derived stromal cell lines (OP9 and S17) are capable of inducing hematopoietic differentiation of totipotent murine and human embryonic stem cells (ESC). Here, the potential of more than 100 stromal cell lines developed from the aorta-gonado-mesonephros (AGM) region was investigated in supporting adult and embryonic hematopoiesis. In addition, extensive phenotypic analysis should elucidate possible mechanisms involved in maintenance of hematopoietic stem cell function. METHODS: More than 100 stromal cell clones derived from the AGM region of E10.5 mouse embryos were isolated. Hematopoietic stem cell support was tested for adult murine and human cord blood hematopoietic stem cells and hematopoietic cells derived from murine ESC. Genotypic and phenotypic characterization was performed including gene array analysis. RESULTS: It was demonstrated that multiple clones showed high efficiency in supporting maintenance and expansion of primitive murine and human hematopoietic progenitors. In addition, we demonstrated for the first time that AGM stromal cell lines are also potent inducers of hematopoietic differentiation of murine ESC. Microarray analysis of AGM lines revealed a characteristic genotype with expression of genes involved in regulating hematopoiesis as well as mesodermal and early B cell development. CONCLUSION: These AGM stromal cell lines may be of value in elucidating molecular mechanisms regulating early stem cell development and hematopoietic differentiation from ES-derived mesoderm.

Animals↗
PubMed2006

Promotion of haematopoietic activity in embryonic stem cells by the aorta-gonad-mesonephros microenvironment.

We investigated whether the in vitro differentiation of ES cells into haematopoietic progenitors could be enhanced by exposure to the aorta-gonadal-mesonephros (AGM) microenvironment that is involved in the generation of haematopoietic stem cells (HSC) during embryonic development. We established a co-culture system that combines the requirements for primary organ culture and differentiating ES cells and showed that exposure of differentiating ES cells to the primary AGM region results in a significant increase in the number of ES-derived haematopoietic progenitors. Co-culture of ES cells on the AM20-1B4 stromal cell line derived from the AGM region also increases haematopoietic activity. We conclude that factors promoting the haematopoietic activity of differentiating ES cells present in primary AGM explants are partially retained in the AM20.1B4 stromal cell line and that these factors are likely to be different to those required for adult HSC maintenance.

Animals↗
PubMed2006

Inhibitory effects of homeodomain-interacting protein kinase 2 on the aorta-gonad-mesonephros hematopoiesis.

Definitive hematopoiesis starts in the aorta-gonad-mesonephros (AGM) region of the mouse embryo. Our previous studies revealed that STAT3, a gp130 downstream transcription factor, is required for AGM hematopoiesis and that homeodomain-interacting protein kinase 2 (HIPK2) phosphorylates serine-727 of STAT3. HIPK2 is a serine/threonine kinase known to be involved in transcriptional repression and apoptosis. In the present study, we examined the role of HIPK2 in hematopoiesis in mouse embryo. HIPK2 transcripts were found in fetal hematopoietic tissues such as the mouse AGM region and fetal liver. In cultured AGM cells, HIPK2 protein was detected in adherent cells. Functional analyses of HIPK2 were carried out by introducing wild-type and mutant HIPK2 constructs into AGM cultures. Production of CD45(+) hematopoietic cells was suppressed by forced expression of HIPK2 in AGM cultures. This suppression required the kinase domain and nuclear localization signals of HIPK2, but the kinase activity was dispensable. HIPK2-overexpressing AGM-derived nonadherent cells did not form cobblestone-like colonies in cultures with stromal cells. Furthermore, overexpression of HIPK2 in AGM cultures impeded the expansion of CD45(low)c-Kit(+) cells, which exhibit the immature hematopoietic progenitor phenotype. These data indicate that HIPK2 plays a negative regulatory role in AGM hematopoiesis in the mouse embryo.

Animals↗
PubMed2002

In vivo differentiation of stem cells in the aorta-gonad-mesonephros region of mouse embryo and adult bone marrow.

OBJECTIVE: Hematopoietic stem cells (HSCs) are thought to be generated from hemangioblasts, the common precursor cells for blood and endothelial cells, in the aorta-gonad-mesonephros (AGM) region of the mouse embryo. The genetic program of HSCs was recently demonstrated to be plastic, but the potential for AGM-region hemangioblasts to be transplanted and to differentiate in vivo has not been well described. Here we examined the fate of donor cells in mice transplanted with CD45(-) AGM cells, which presumably include hemangioblasts. MATERIALS AND METHODS: CD45(-) cells in the AGM region of embryos at 11.5 days post coitum or CD45(+)CD34(-) side population (SP) of cells in adult bone marrow (BM) derived from enhanced green fluorescent protein transgenic mice were transplanted into the liver of busulfan-treated neonatal mice. Two to 6 months after injection of the cells, the contribution of donor-derived cells in the hematopoietic compartment and in various organs was analyzed by flow cytometry and confocal microscopy. RESULTS: CD45(-) cells from the AGM region not only generated peripheral blood cells but also differentiated into endothelial and other nonhematopoietic cells in liver, kidney, lung, small intestine, and uterus in transplanted mice. A similar engrafting pattern was observed in the small intestine of mice transplanted with BM SP/CD45(+) cells, secondary BM-transplanted mice, and lethally irradiated adult mice that received intravenous injections of BM cells. CONCLUSION: A CD45(-) fraction of the AGM region and CD45(+) BM stem cells share the same in vivo potential to differentiate into hematopoietic, endothelial, smooth muscle, and stroma-like cells when transplanted in mice.

Age Factors↗
PubMedDate not supplied

Hematopoietic cells in cultures of the murine embryonic aorta-gonad-mesonephros region are induced by c-Myb.

Definitive hematopoiesis begins in the para-aortic, splanchnopleural (P-Sp) and aorta-gonad-mesonephros (AGM) regions of mouse embryos and then switches to the fetal liver [1] [2] [3]. Gene-targeted mice lacking the c-Myb transcription factor have severe hematopoietic defects in the fetal liver [4]. The role of c-Myb, if any, in P-Sp/AGM hematopoiesis has not been examined, however. Recently, we reported that oncostatin M can effectively expand both hematopoietic and endothelial-like cells from in vitro cultures of the AGM region [5]. Using this cell culture system, we examined the involvement of c-Myb in definitive hematopoiesis in the P-Sp and AGM regions. When primary cultures from the P-Sp or AGM regions of wild-type mouse embryos were probed with an anti-c-Myb antibody, hematopoietic cells but not endothelial-like cells showed positive staining. In contrast, in the P-Sp/AGM culture from c-myb(-/-) embryos, no hematopoietic cells were generated and endothelial-like cells predominated, indicating that the impairment of hematopoiesis in the liver of c-myb(-/-) embryos is actually preceded by a defect in P-Sp/AGM hematopoiesis. Hematogenic precursor cells were, however, still present in an inert but competent form among the endothelial-like, adherent cell population of c-myb(-/-) P-Sp/AGM cultures. When infected with a retrovirus carrying c-myb cDNA, these cultures gave rise to a significant number of hematopoietic cells. The rescued cells, unlike wild-type hematopoietic cells, were negative for c-Kit (a marker of hematopoietic progenitors), but did express other hematopoietic cell surface markers such as Mac-1, Gr-1 (myeloid markers), CD19, B220, Thy-1.2 (Iymphoid markers), and Ter119 (an erythroid marker). Thus, c-Myb plays a role in the generation of hematopoietic cells in the embryonic P-Sp and AGM regions.

3T3 Cells↗
PubMed1999

Identification of podocalyxin-like protein 1 as a novel cell surface marker for hemangioblasts in the murine aorta-gonad-mesonephros region.

Recent studies with avian embryos and murine embryonic stem cells have suggested that hematopoietic cells are derived from hemangioblasts, the common precursors of hematopoietic and endothelial cells. We molecularly cloned podocalyxin-like protein 1 (PCLP1) as a novel surface marker for endothelial-like cells in the aorta-gonad-mesonephros (AGM) region of mouse embryos, where long-term repopulating hematopoietic stem cells (LTR-HSCs) are known to arise. PCLP1+ CD45 cells in the AGM region incorporated acetylated low-density lipoprotein and produced both hematopoietic and endothelial cells when cocultured with OP9 stromal cells. Moreover, multiple lineages of hematopoietic cells were generated in vivo when PCLP1 +CD45-cells were injected into neonatal liver of busulfan-treated mice. Thus, PCLP1 can be used to separate hemangioblasts that give rise to LTR-HSCs.

Amino Acid Sequence↗
PubMed1998

In vitro expansion of murine multipotential hematopoietic progenitors from the embryonic aorta-gonad-mesonephros region.

The origin of hematopoietic stem cells (HSCs) and their growth factor requirement are poorly understood. Here we describe a new in vitro culture system of the aorta-gonad-mesonephros (AGM) region, where long-term repopulating HSCs first arise. We demonstrate that oncostatin M (OSM) is expressed in the AGM and is absolutely required for the expansion of multipotential hematopoietic progenitors in vitro. In addition, OSM enhances the formation of endothelial cell clusters. Thus, OSM appears to be a key cytokine for the development of multipotential hematopoietic progenitors in the AGM, possibly acting on common precursor cells between HSCs and endothelial cells. By using the AGM culture derived from macrophage colony-stimulating factor (M-CSF)-deficient op/op mutant embryos, we also show a pivotal role for M-CSF in fetal myelopoiesis.

Animals↗
PubMed1997

Ontogeny of angiotensin II receptors, types 1 and 2, in ovine mesonephros and metanephros.

By RNAse protection assay, hybridization histochemistry, and in vitro autoradiography it was shown that both mRNA and protein for AT1 and AT2 receptors were present in ovine fetal meso- and metanephroi at 40 days of gestation (term approximately 150 days). AT1 mRNA was localized to presumptive mesangial cells of glomeruli at 40-, 75-, 131-gestational-day-old fetuses and two-day-old lambs, in addition to being widely present in interstitial cells of the cortex and medulla, once these zones formed (60 days). By two days after birth the medullary AT1 distribution was confined to the inner stripe of the outer medulla. AT2 mRNA was present in peripheral interstitial/tissue of the mesonephros, and interstitial tissue surrounding developing glomeruli, but not the outermost nephrogenic mesenchyme in the metanephros from 40 to approximately 131 days (the period of active nephrogenesis). In addition, AT2 mRNA was localized to epithelial cells of the macula densa in metanephroi (40 to 131 gestational days) during, but not after completion, of nephrogenesis. These studies suggest that angiotensin II (Ang II) could have differentiating effects, via AT1 receptors, from very early in development. The unique epithelial site of AT2 expression in the macula densa raises the possibility that Ang II may play a role in the invariant positioning of the macula densa at the pole of its glomerulus, via this receptor.

Aging↗
PubMed2004

Evidence for a fruit fly hemangioblast and similarities between lymph-gland hematopoiesis in fruit fly and mammal aorta-gonadal-mesonephros mesoderm.

The Drosophila melanogaster lymph gland is a hematopoietic organ and, together with prospective vascular cells (cardioblasts) and excretory cells (pericardial nephrocytes), arises from the cardiogenic mesoderm. Clonal analysis provided evidence for a hemangioblast that can give rise to two daughter cells: one that differentiates into heart or aorta and another that differentiates into blood. In addition, the GATA factor gene pannier (pnr) and the homeobox gene tinman (tin), which are controlled by the convergence of Decapentaplegic (Dpp), fibroblast growth factor (FGF), Wingless (Wg) and Notch signaling, are required for the development of all cardiogenic mesoderm, including the lymph gland. Here we show that an essential genetic switch that differentiates between the blood or nephrocyte and vascular lineages involves the Notch pathway. Further specification occurs through specific expression of the GATA factor Serpent (Srp) in the lymph-gland primordium. Our findings suggest that there is a close parallel between the molecular mechanisms functioning in the D. melanogaster cardiogenic mesoderm and those functioning in the mammalian aorta-gonadal-mesonephros mesoderm.

Animals↗