Kidney Endothelial Cells Research Articles

Renal Pathology in Fabry Disease

Fabry disease is an X-linked recessive lysosomal storage disease that is caused by deficient activity of the lysosomal enzyme α-galactosidase A (α-Gal A), an enzyme that cleaves terminal α-galactosyl residues. This deficiency results in progressive lysosomal accumulation of glycosphingolipid with

Angiopoietin correlates with glomerular capillary loss in anti-glomerular basement membrane glomerulonephritis

Emerging evidence suggests that endothelial turnover occurs in several glomerular diseases and correlates with resolution or progression of glomerular lesions. We hypothesized that the growth factors modulating embryonic kidney endothelial cell survival and capillary morphogenesis may be implicated in capillary loss that occurs in immune-mediated glomerulonephritis (GN). GN was induced in C57BL/6 mice by intravenous administration of sheep anti-mouse glomerular basement membrane (GBM) globulin and assessed with markers of vascularity in glomerular lesions, correlating these with expression of specific vascular growth factors. As assessed by periodic acid Schiff staining, 14 +/- 4% (mean +/- SD) glomeruli were affected by sclerosis at 14 days after globulin administration, and 33 +/- 5% were affected at 21 days. By 21 days, a significant increase of plasma creatinine and urinary protein occurred. P-selectin expression was increased in glomerular capillaries 14 days after disease induction, and capillary loss, as assessed by immunohistochemistry for platelet-endothelial cell adhesion molecule, vascular endothelial growth factor (VEGF) receptor 2 and the angiopoietin (Ang) receptor Tie-2, was recorded at 14 and 21 days in glomeruli affected by proliferative crescents and/or sclerosis. VEGF-A immunostaining, evident in control glomeruli, was qualitatively diminished in glomeruli with lesions. Ang-1 immunostaining was detected in control glomeruli and was diminished at 14 days after administration of anti-mouse GBM globulin; instead, Ang-1 was immunolocalized to distal tubules. In contrast, Ang-2 immunostaining was barely detectable in control glomeruli but was prominent in disease glomeruli. In GN mice, rare apoptotic glomerular endothelia were detected by electron microscopy and in situ end-labeling, but such cells were not seen in controls. Loss of glomerular capillaries during the course of anti-GBM GN in mice was temporally associated with decreases in endothelial survival molecules VEGF-A and Ang-1, and with up-regulation of Ang-2, an antagonist of Ang-1. A changing balance of these growth factors may contribute to decreased glomerular vascularity in crescentic GN.

Cell surface-associated chondroitin sulfate proteoglycans bind contact phase factor H-kininogen

The kinin system has been recognized as a locally operating hormone system of cardiovascular cells, however, the molecular mechanisms regulating circumscribed kinin release on cell surfaces are not fully understood. In particular, the principal cell docking sites for the kinin precursor, high molecular weight kininogen (HK), are not fully explored. Here we demonstrate by enzymatic digestion, recombinant overexpression, and affinity cross-linking studies that cell surface chondroitin sulfate (CS) chains of proteoglycans (PGs) serve as major HK binding sites on platelet, fibroblast, liver, and endothelial kidney cells. In this way, CS-type PGs may contribute to a local accumulation of kinin precursors on cell surfaces and modulate circumscribed release of short-lived kinin hormones at or next to their site of action.

Effect of cell swelling on membrane and cytoplasmic distribution of pICln.

pICln is found ubiquitously in mammalian cells and is postulated to play a critical role in cell volume regulation. Mutagenesis studies led to the proposal that pICln is a swelling-activated anion channel. However, recent studies in Madin-Darby canine kidney cells and endothelial cells have shown that the protein is localized primarily to the cytoplasm. It has therefore been postulated that activation involves reversible translocation of pICln from the cytoplasm and insertion into the plasma membrane. We tested this hypothesis using several different approaches. Fractionation of C6 glioma cells into plasma membrane- and cytoplasm-containing fractions demonstrated that approximately 90% of the recovered pICln was confined to the cytosol. Swelling had no effect on the relative amount of protein present in the plasma membrane fraction. Immunofluorescence microscopy revealed that pICln is localized primarily, if not exclusively, to the cytoplasm of swollen and nonswollen cells. Similarly, transfection of cells with a green fluorescent protein-labeled pICln construct failed to reveal any membrane localization of the protein. These findings do not support the hypothesis that pICln is a volume regulatory anion channel activated by swelling-induced membrane insertion.

Characterisation of a recombinant P 2Y purinoceptor

We have previously cloned a cDNA encoding a G-protein-coupled P 2 purinoceptor from chick brain and designated this as a P 2Y1 purinoceptor (Webb, T.E., J. Simon, B.J. Krishek, A.N. Bateson, T.G. Smart, B.J. King, G. Burnstock and E.A. Barnard, 1993, FEBS Lett. 324, 219). Here, we describe the further characterisation of this recombinant receptor expressed in both simian kidney endothelial (COS-7) cells and Xenopus oocytes. In transfected COS-7 cell membranes, the recombinant receptor showed a high level of expression ( B max = 7.9 ± 2.2 pmol [ 35 S] dATPα S bound/mg protein ) and affinity ( K d = 6.6 ± 0.3 nM). In these COS-7 cells, the activation of the implanted purinoceptor induced a suramin-sensitive formation of inositol 1,4,5-triphosphate (1,4,5InsP 3). Upon expression in Xenopus oocytes, ATP was the only natural nucleoside triphosphate to elicit a Ca 2+-activated chloride current. The P 2 purinoceptor antagonists suramin and Reactive Blue-2 were both able to inhibit this evoked current. Utilizing both expression systems, the binding affinity profile and the functional pharmacological profile of the agonists, the common series found was: 2-methylthioATP (2-MeSATP) ⩾ ATP > ADP βS > ADP. These two agonist series and the lack of activity of adenosine, α,β-methyleneATP (α,β-meATP), 3′- O-(4- benzoyl)benzoyl-ATP (Bz-ATP) and UTP, together confirmed that this receptor is a specific subtype of the P 2Y purinoceptors.

Postcapillary venule endothelial cells in kidney express a multispecific chemokine receptor that is structurally and functionally identical to the erythroid isoform, which is the Duffy blood group antigen.

The human erythrocyte chemokine receptor has recently been shown to be identical to the Duffy blood group antigen and is expressed in multiple organs, including kidney. Here we have examined the molecular properties of the renal isoform. Immunoblot analysis of erythrocyte and kidney detergent lysates, with a monoclonal antibody (Fy6) to the Duffy antigen, revealed that the renal isoform had a molecular mass of 43-45 kD, which could be distinguished from that observed in erythroid cells (38-47 kD). Chemical cross-linking of kidney membranes to 125I-melanoma growth stimulatory activity (MGSA) indicated that the renal chemokine receptor had a molecular mass of 38-45 kD. Binding of 125I-labeled MGSA to kidney membranes was competitively inhibited by the addition of unlabeled MGSA, IL-8, regulated on activation, normal T expressed and secrted, and monocyte chemotactic protein-1. Scatchard analysis of MGSA binding showed that the chemokine receptor from renal tissues had a binding affinity of 3.5 nM similar to that observed for the erythroid isoform (5-10 nM). The primary structure of the renal chemokine receptor predicted from the nucleotide sequence of cDNA from renal tissues is identical to that reported for the erythroid isoform. Immunocytochemical staining of kidney with Fy6 localized expression to endothelial cells present in postcapillary venules. These studies implicate the Duffy antigen/chemokine receptor in the complex interactions between postcapillary endothelial cells and granulocytes, which are modulated by pro-inflammatory chemokines.

P-selectin and platelet-activating factor mediate initial endotoxin-induced neutropenia.

Polymorphonuclear neutrophil (PMN) accumulation within damaged tissues, a hallmark of acute inflammation, is dependent upon initial adhesion to endothelial cells. In vitro studies suggest that P-selectin and platelet activating factor (PAF) are key molecules in this process by promoting the initial adhesion of PMN to endothelial cells. We report in vivo studies in which intravenous administration of lipopolysaccharide (LPS) to anesthetized rats caused a very rapid onset (< 5 min) of neutropenia, in association with induction of surface expression of P-selectin on microvascular endothelial cells in kidney, liver and lung; analogous induction of P-selectin expression by cultured endothelial cells was observed in response to LPS stimulation in vitro. In addition, treatment with an antibody (Ab) to P-selectin (or use of a PAF antagonist) blocked development of neutropenia in vivo for at least 15 min post-LPS injection, and Ab treatment was shown to block PMN accumulation in tissues. These studies document roles for P-selectin and PAF in the early adhesion of PMN to endothelial cells in vivo.

Receptor-mediated endocytosis and phagocytosis by rainbow trout head kidney sinusoidal cells

Endocytosis by rainbow trout head kidney sinusoidal cells was studied following intravenous administration of various ligands. Immunohistochemistry was used to demonstrate that endothelial cells took up the soluble ligands ovalbumin and dinitrophenylated human serum albumin (DNP 33HSA). Ovalbumin and DNP 33HSA are endocytosed via the mannose- and the scavenger receptor, respectively. Sinusoidal macrophages took up DNP 33HSA but only occasionally ovalbumin. The particulate ligand, colloidal human serum albumin (cHSA) (0·2-1μm) was taken up by both endothelial cells and sinusoidal macrophages. Endothelial cells were estimated to account for at least 15% of uptake of intravenously injected 125 I-tyramine cellobiose-labelled cHSA ( 125I-TC-cHSA) by head kidney cells. Uptake of 125I-TC-cHSA could be demonstrated in cultured head kidney macrophages. Intracellular transport of in vivo internalized 125I-TC-cHSA to lysosomal compartments occurred at a slower rate compared to ligands endocytosed via the mannose- and scavenger-receptors. Electron microscopic studies of head kidney endothelium that had taken up colloidal carbon, revealed that sinusoidal endothelial cells possess the ultra-structural apparatus characteristic for cells active in endocytosis of blood-borne ligands; namely well defined coated pits along the luminal plasma membrane, numerous intracellular vesicles and large, lysosome-like dense bodies. Thus, the results of this study suggest that both head kidney endothelial cells and sinusoidal macrophages are important 'scavenger' cells in rainbow trout.

Immunohistochemical localization of an inducible form of nitric oxide synthase in various organs of rats treated with Propionibacterium acnes and lipopolysaccharide

Immunohistochemical localization of an endotoxin-inducible form of nitric oxide synthase was examined using rabbit polyclonal antibody against the enzyme purified from rat liver. In rats treated with Propionibacterium acnes and lipopolysaccharide, immunostaining was observed in macrophages, occasional lymphocytes, neutrophils and eosinophils in red pulp of spleen, Kupffer cells, endothelial cells and hepatocytes in liver, alveolar macrophages in lung, macrophages and endothelial cells in adrenal glands, and histiocytes, eosinophils, mast cells and endothelial cells in colon. Immunoreactivity was also evident in the following tissues: histiocytes and endothelial cells in kidney; histiocytes and neutrophils in esophagus; macrophages and eosinophils in duodenum; macrophages, some lymphocytes and mast cells in ileum; histiocytes in thymus; and endothelial cells in heart and aorta. Immunoreactivity was not detected in these organs from untreated rats. Positively staining cells in these rat organs appeared within 2.5 h after lipopolysaccharide administration; their number dramatically increased within the next 2.5 h, remained at high levels for a further 19 h, and then decreased over the following 24 h. The number of positive cells appearing correlated well with the nitric oxide synthase activity biochemically determined in the same organs.

The major site of murine K papovavirus persistence and reactivation is the renal tubular epithelium

K virus, a murine papovavirus, produces a lethal pneumonia in newborn mice. Animals surviving acute illness develop a persistent infection which reactivates under conditions of immunosuppression. The present study was conducted to identify the cell populations which support persistent K virus infection and to determine the cell populations in which this persistent infection is reactivated during immunosuppression. Mice inoculated by the oral route with 100 50% newborn mouse lethal doses (LD 50) of K virus at 14 days of age were followed over a period of 7 months. The distribution of infection was studied by virus assay, immunohistochemistry, and in situ nucleic acid hybridization methods. Viral replication during the acute phase of infection was confined to pulmonary and systemic vascular endothelial cells, as well as to scattered, apparently lymphoid cells within spleens. Beginning 2 months after inoculation, however, specific hybridization for K virus nucleic acids was detected in rare renal tubular epithelial cells, and by 6 months after inoculation renal tubular epithelial cells represented the major site of viral persistence. Positive cells were frequently present in groups of two or more, and a minority of positive cells also expressed viral capsid (V) antigen. Immunosuppression with cyclophosphamide resulted in reactivation of infection, with highest titers of virus being detected in kidneys and with increased numbers of renal tubular epithelial cells expressing viral capsid antigen. Capsid antigen was also detected in rare endothelial cells in kidneys, livers and lungs of these immunosuppressed mice. Although K virus behaves as an endotheliotrope during acute infection, the major site of K virus persistence and reactivation, the renal tubular epithelial cell, is similar to that involved during persistent infection by polyoma virus in mice, SV40 virus in monkeys, and BK and JC viruses in man. The observation that persistently infected renal tubular epithelial cells occur in groups of two or more and occasionally express capsid antigen suggests that virus may persist as a productive infection which is confined by antiviral antibody but maintains itself by cell-to-cell-spread. The present study represents the first instance in which the cell populations which support infection by a member of the polyomavirus subgroup in its natural host have been defined during acute, persistent, and reactivated infection.

Mortality in psittacine birds resembling Pacheco's disease in Spain

The sudden death of five sulphur-crested cockatoos (Cacatua galerita) with no apparent clinical signs was investigated. The birds had hepatitis with intranuclear inclusion bodies (IIBs), as well as IIBs in mononuclear spleen cells, epithelial cells of the kidney tubules and endothelial cells of the glomerular capillaries. Ultrastructurally, icosahedral herpesvirus-like particles, 110 to 115 nm in diameter were found consistent with Pacheco's disease. This is the first time the disease has been reported in cockatoos in Spain.

Structure and function of hepatocyte growth factor

Hepatocyte growth factor (HGF), a potent mitogen for mature hepatocytes in primary culture, was first found in sera of partial hepatectomized rats and seems to be a hepatotrophic factor for liver regeneration which has not been purified over the past 30 years. HGF is composed of the 69 kDa α-subunit and the 34 kDa β-subunit. Molecular cloning reveals that HGF is derived from a single chain precursor of 728 amino acid residues and it contains 4 kringle domains in the α-subunit. HGF gene spans about 70kb and consists of 18 exons and 17 introns. HGF is now thought to be a pleiotropic factor influencing a cell growth and cell motility for various epithelial cells. HGF receptor with K d = 20–30p m is widely distributed in various epithelial cells including hepatocytes. HGF mRNA and HGF activity increase markedly in liver after various liver injuries and in kidney following unilateral nephectomy or acute renal injury. Moreover, HGF mRNA is induced even in the intact lung in response to liver and kidney injury. In situ hybridization reveals that HGF-producing cells are mesenchymal cells such as Kupffer cells and sinusoidal endothelial cells in liver, fenestrated endothelial cells in kidney, and macrophages and endothelial cells in lung. Thus, HGF may play an important role as a paracrine or endocrine mediator through an epithelial-mesenchymal interaction in wound-healing, tissue or organ regeneration, morphogenesis and carcinogenesis.

Localization of [125I]Endothelin-1 in Rat Tissues Observed by Electron-Microscopic Radioautography

[125I]Endothelin-1 (ET-1) was administered intravenously into rats, and binding sites in various tissues were examined by electron-microscopic radioautography. Labeling was observed predominantly on the glomerular fenestrated endothelial cells of kidney, alveolar capillary endothelial cells of lung, fat-storing cells in the sinusoids of liver, and subepithelial myofibroblasts of the small intestine. At first, silver grains were observed mostly on the plasmalemma of these cells and with time translocated into the cytoplasm of these cells. Myofibroblasts of the small intestine were cultured to study the cellular effects of ET. By fura-2 fluorescence method, transient increase and oscillation of intracellular Ca2+ level were observed in the cultured myofibroblasts following the application of ET-1 or endothelin-3 (ET-3). These results suggest that ET induces diverse physiological reactions in specific cells of various tissues.

Evidence for the occurrence of human erythrocyte membrane sialoglycoproteins in human kidney endothelial cells.

Several mouse monoclonal antibodies recognising different epitopes on human erythrocyte sialoglycoproteins alpha and beta (syn: glycophorins A and C, respectively) react with human renal endothelium. Those monoclonal anti-blood group M and anti-blood group N antibodies which recognised the M/N epitopes on sialidase-treated human erythrocytes also reacted with human renal endothelium whilst those which recognise the M/N epitopes on native, but not on sialidase-treated human erythrocytes were not reactive with human renal endothelium. These results provide evidence for the occurrence of alpha and beta molecules in renal endothelium and suggest that alpha in renal endothelium is incompletely sialylated.

Evidence for the Occurrence of Human Erythrocyte Membrane Sialoglycoproteins in Human Kidney Endothelial Cells

Evidence for the Occurrence of Human Erythrocyte Membrane Sialoglycoproteins in Human Kidney Endothelial Cells

The response of cultured human kidney capillary endothelium to immunologic stimuli

We have developed in culture for the first time one of the actual target cells in kidney rejection by establishing long-term cultures of human kidney capillary endothelium and have begun to explore their response to immunologic stimuli. Microvascular endothelial cells were obtained from the human kidney cortex by mechanical separation, enzyme digestion, and density gradient centrifugation. The cells were exposed to 1% PHA, a PHA-stimulated PBL supernatant, human and mouse IL-1 and human γ-interferon. They were stained with FITC-conjugated monoclonalantibodies specific for β 2-microglobulin, HLA-DR, and DQ and analyzed by flow cytometry (FACS-II). HKCE cells express HLA-ABC antigens during growth and confluent phases but do not express DR or DQ antigens. Only the PHA-stimulated PBL supernatant and γ-interferon induced class II antigen expression. A change in cellular morphology was noted after 72 hr of incubation in the presence of lectin-stimulated PBL supernatant. Our findings show that microvascular endothelial cells express selected Ia antigens only after immunologic induction. This is an agreement with studies using umbilical and aortic endothelial cells. These data suggest that a dynamic interaction takes place between activated lymphocytes whose products induce expression of class II antigens and human kidney endothelial cells.

Cationic dyes reveal proteoglycans on the surface of epithelial and endothelial kidney cells.

The glomerular epithelial cells of the rat kidney fixed by vascular perfusion with an aldehyde solution containing either safranine O or alcian blue (and 0.3 M MgCl2) display filaments which are located close to the outer surface of the plasma membrane. These filaments are similar to those revealed by the same methods in the laminae rarae of the glomerular basement membrane. Alcian blue (and MgCl2) further demonstrates the presence of anionic sites inside the endothelial cell pores of the glomerular and peritubular capillaries, on the luminal surface of endothelial cells of large renal vessels and along the basolateral surface of the epithelial cells of the Bowman capsule and of the proximal convoluted tubule.

Self-assessment

The distribution of Ia-like antigens in human kidney has been investigated by indirect immunofluorescence staining of frozen sections with monoclonal antibodies to framework determinants of Ia-like antigens. These antigens are localized by patchy and interrupted linear fluorescence along the endothelium of glomerular efferent arterioles and capillaries, intertubular cortical and medullary capillaries as well as the vasa recta. Ia-like antigens are undetectable, however, on the endothelia of arcuate and interlobular arteries and on tubular epithelial cells. The expression of Ia-like antigens on kidney endothelial cells may account for the role of Ia-like antigens in kidney allograft rejection.

Expression of Ia-like antigens on the vasculature of human kidney

The distribution of Ia-like antigens in human kidney has been investigated by indirect immunofluorescence staining of frozen sections with monoclonal antibodies to framework determinants of Ia-like antigens. These antigens are localized by patchy and interrupted linear fluorescence along the endothelium of glomerular efferent arterioles and capillaries, intertubular cortical and medullary capillaries as well as the vasa recta. Ia-like antigens are undetectable, however, on the endothelia of arcuate and interlobular arteries and on tubular epithelial cells. The expression of Ia-like antigens on kidney endothelial cells may account for the role of Ia-like antigens in kidney allograft rejection.