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H Holthöfer

Publications and source records attributed to H Holthöfer.

89 records · Page 5Linked to original sources
PubMed1986

Binding of Escherichia coli S fimbriae to human kidney epithelium.

Purified S fimbriae and an Escherichia coli strain carrying the recombinant plasmid pANN801-4 that encodes S fimbriae were tested for adhesion to frozen sections of human kidney. The fimbriae and the bacteria bound to the same tissue domains, and in both cases the binding was specifically inhibited by the receptor analog of S fimbria, sialyl(alpha 2-3)lactose. S fimbriae bound specifically to the epithelial elements in the kidneys; to the epithelial cells of proximal and distal tubules as well as of the collecting ducts and to the visceral and parietal glomerular epithelium. In addition, they bound to the vascular endothelium of glomeruli and of the renal interstitium. No binding to connective tissue elements was observed. The results suggest that the biological function of S fimbriae is to mediate the adhesion of E. coli to human epithelial and vascular endothelial cells.

Adenocarcinoma↗
PubMed1986

Localization of binding sites for purified Escherichia coli P fimbriae in the human kidney.

Binding sites in the human kidney for purified P fimbriae of pyelonephritogenic Escherichia coli were determined. The purified KS71A (F7(1)) fimbriae bound only to epithelial elements of the kidney, i.e., to the apical aspect of proximal and distal tubular cells, as well as to the apical and cytoplasmic sites of collecting ducts. In addition, binding was seen at the vascular endothelium throughout the kidney and at the parietal epithelium of the glomeruli. The binding was specifically inhibited by the receptor analog of E. coli P fimbriae, globotriose. The binding sites identified suggested a possible pathogenetic mechanism for the invasion of P-fimbriated bacteria into the renal parenchyma, as well as for their subsequent spread into the circulatory system.

Adenocarcinoma↗
PubMed1986

Fluorochrome-coupled lectins reveal distinct cellular domains in human epidermis.

The distribution of saccharide moieties in human interfollicular epidermis was studied with fluorochrome-coupled lectins. In frozen sections Concanavalin A (Con A), Lens culinaris agglutinin (LCA), Ricinus communis agglutinin I (RCAI), and wheat germ agglutinin (WGA) stained intensively both dermis and viable epidermal cell layers, whereas peanut agglutinin (PNA) bound only to living epidermal cell layers. Ulex europaeus agglutinin I (UEAI) bound to dermal endothelial cells and upper cell layers of the epidermis but left the basal cell layer unstained. Dolichos biflorus agglutinin (DBA) bound only to basal epidermal cells, whereas both soybean agglutinin (SBA) and Helix pomatia agglutinin (HPA) showed strong binding to the spinous and granular cell layers. On routinely processed paraffin sections, a distinctly different staining pattern was seen with many lectins, and to reveal the binding of some lectins a pretreatment with protease was required. All keratin-positive cells in human epidermal cell suspensions, obtained with the suction blister method, bound PNA, whereas only a fraction of the keratinocytes bound either DBA or UEAI. Such a difference in lectin binding pattern was also seen in epidermal cell cultures both immediately after attachment and in organized cell colonies. This suggests that in addition to basal cells, more differentiated epidermal cells from the spinous cell layer are also able to adhere and spread in culture conditions. Gel electrophoretic analysis of the lectin-binding glycoproteins in detergent extracts of metabolically labeled primary keratinocyte cultures revealed that the lectins recognized both distinct and shared glycoproteins. A much different lectin binding pattern was seen in embryonic human skin: fetal epidermis did not show any binding of DBA, whereas UEAI showed diffuse binding to all cell layers but gave a bright staining of dermal endothelial cells. This was in contrast to staining results obtained with a monoclonal cytokeratin antibody, which showed the presence of a distinct basal cell layer in fetal epidermis also. The results indicate that expression of saccharide moieties in human epidermal keratinocytes is related to the stage of cellular differentiation, different cell layers expressing different terminal saccharide moieties. The results also suggest that the emergence of a mature cell surface glycoconjugate pattern in human epidermis is preceded by the acquisition of cell layer-specific, differential keratin expression.

Adult↗
PubMed1985

Alterations in glomerular lectin binding sites of human kidney as detected by fluorescence microscopy.

Different lectins were used to study frozen sections of kidney samples showing alterations in routine immunofluorescence studies. Arachis hypogaea agglutinin (peanut lectin, PNA), lacking binding sites in normal glomeruli, bound to the glomeruli in two of the five samples studied, giving a granular fluorescence pattern. Concomitantly with the appearance of PNA-binding, binding sites for wheat germ agglutinin (WGA) appeared to be lost at glomeruli. Furthermore, changes in the expression of glomerular binding sites for Wistaria floribunda (WFA), Helix pomatia (HPA) and Dolichos biflorus (DBA) agglutinins could be seen in the kidneys studied, whereas the binding sites for Ulex europaeus agglutin (UEA I) in vascular endothelia seemed to be unaltered. The results show that kidney specimens presenting changes in routine immunofluorescence studies may also present altered binding for certain lectins. On this basis we propose that certain lectins may aid in characterizing these changes and are thus of potential use in studying diseased kidneys.

Fluorescent Antibody Technique↗
PubMed1984

Lectins as markers for cells infiltrating human renal glomeruli.

This study describes a simple method for detecting mononuclear cells in human renal glomeruli using labeled lectins as probes. The lectins used in this study showed prominent binding to different cell types among the nonresident glomerular cells but not to normal glomerular elements. Monoclonal antibodies against monocytes/macrophages (OKM 1), T-cells (OKT 11), suppressior/cytotoxic T-cells (OKT 8), and anti-lysozyme antibodies were used in double-fluorescence studies with the lectins in an attempt to identify the lectin-positive cells. The results indicate that Bandeiraea simplicifolia I isolectin and Lotus tetragonolobus agglutinin in particular are useful for screening of kidney biopsies for cells of their monocyte/macrophage series and T-cells invading human renal glomeruli.

Antibodies, Monoclonal↗
PubMed1984

Cellular differentiation of basal cell carcinoma studied with fluorescent lectins and cytokeratin antibodies.

The expression of cellular glycoconjugates and cytokeratin polypeptides in 8 basal cell carcinomas (BCC) was studied using fluorochrome-coupled lectins and different keratin-antibodies. Peanut agglutinin and Wistaria floribunda agglutinin, binding to all layers of normal human epidermis, also stained all cells in the basal cell carcinomas. Dolichos biflorus agglutinin, which in normal epidermis binds only to the basal cells, gave a mottled staining pattern in most of the tumors. Instead, Ulex europaeus I agglutinin and soybean agglutinin, which in normal epidermis only bind to the spinous and granular cell layers, did not stain tumor cells in basal cell carcinomas. Rabbit antibodies to human 43-50kD epidermal keratin polypeptides and 2 monoclonal cytokeratin antibodies, PKK1 reacting only with follicular epithelium, and PKK2 reacting also with the basal epidermal cells, brightly stained all cells of the basal cell carcinomas studied, whereas antibodies to human 60-67kD epidermal keratin polypeptides did not bind to the carcinoma cells. The results suggest that the cells in basal cell carcinomas resemble epidermal basal cells both by their glycoconjugate pattern and keratin expression. However, the tumor cells also express cytokeratins, which can be found only in the follicular epithelium, but not in normal interfollicular epidermis.

Adult↗
PubMed1984

Expression of vimentin and cytokeratin types of intermediate filament proteins in developing and adult human kidneys.

Expression of cytokeratins and vimentin in developing and adult human kidney was studied by the indirect immunofluorescence technique using monoclonal and conventional antiintermediate filament antibodies. The undifferentiated cells of the metanephric mesenchyme expressed vimentin but not cytokeratins, whereas only cytokeratins could be visualized in the cells of induced renal vesicles. At the S-shaped body stage of nephrogenesis, the presumptive visceral and parietal cells of the developing glomeruli did not contain either detectable vimentin or cytokeratins, whereas cells of the tubular pole of the S-shaped body were stained within antibodies to cytokeratins. At a later stage the cuboidal visceral epithelial cells of the presumptive glomeruli were brightly positive for vimentin, but not for cytokeratins, and maintained their expression of vimentin during later stages of glomerulogenesis and in adult kidneys. Only some of the cells of the parietal epithelium, but not other glomerular elements, showed cytokeratin-specific staining. In fetal kidneys, the epithelial cells of proximal and distal tubules and of collecting ducts showed a cytoplasmic staining with two monoclonal antibodies to cytokeratins (PKK1 and PKK2), reacting with different cytokeratin polypeptides. In adult kidneys, a basolateral fluorescence in proximal and distal tubules was obtained with PKK1 antibodies, whereas PKK2 stained only the cells of the collecting ducts. Vimentin was not found in tubular epithelial cells at any developmental stage, whereas the cells of collecting ducts showed a transient expression of vimentin in fetal kidneys. Our results show a developmental stage-dependent pattern in the expression of the intermediate filament antigens in the cells of the kidney and show the exceptional expression of only vimentin in the visceral epithelial cells of human glomeruli.

Adult↗
PubMed1983

Altered binding of Ulex europaeus I lectin to psoriatic epidermis.

We have used Ulex europaeus I (UEA I) lectin, specific for alpha-L-fucose-containing glycoconjugates, in fluorescence microscopy to stain cryostat sections of human skin from normal persons and patients with psoriasis and lichen simplex. In normal skin the upper layers of the stratum spinosum and the stratum granulosum were strongly reactive with UEA I, whereas the lower layers of the epidermis did not react. The staining intensity of the upper epidermis was similar to that of the endothelium of dermal blood vessels. Biopsies of the lesional skin of lichen simplex showed an intense UEA I-specific staining throughout the whole epidermis, similar in intensity to that seen in the upper epidermis of normal skin. In psoriatic lesions positive UEA I-specific fluorescence was seen throughout the whole epidermis, but the fluorescence was more faint and often granular. In uninvolved skin of psoriatic patients the whole epidermis showed a diffuse UEA I-specific fluorescence, differing in this respect from normal skin. In normal skin UEA I binds to epidermal cells which are at a certain state of differentiation. The results with psoriatic epidermis confirm that both uninvolved and lesional epidermis have a defect in epidermal maturation, as shown by the altered binding of UEA I lectin.

Epidermis↗
PubMed1983

Lectin binding sites in kidney. A comparative study of 14 animal species.

Six fluorochrome-coupled lectins with different sugar specificities were used to stain frozen tissue sections of kidneys from 14 animal species including mammals, avians, reptiles, and fresh water fish. Each lectin seemed to have a species-, but not strain-, specific binding pattern. Some lectins, however, bound to the same parts of the nephron in all animals studied. Wheat germ agglutinin (WGA) bound prominently to glomeruli in all kidneys. Dolichos biflorus agglutinin (DBA) seemed to bind to only a group of distal tubules in most animals, whereas either proximal or distal tubules were revealed with soybean (SBA) and peanut (PNA) agglutinins. Heterogeneity of basement membranes in different nephron parts was seen in the binding of some lectins. Ulex europeus agglutinin (UEA I), binding specifically to endothelial cells in human tissues, did not react with the endothelium of any other species, but SBA and PNA seemed to prominently stain vascular endothelia of cow and hen vessels, respectively. These results show a species-specific compartmentalization of saccharides to certain parts of the nephron, while there appears to be some common features in saccharide distribution between different animal species as well.

Animals↗
PubMed1983

Cellular origin and differentiation of renal carcinomas. A fluorescence microscopic study with kidney-specific antibodies, antiintermediate filament antibodies, and lectins.

Frozen sections of human renal carcinomas were studied in indirect immunofluorescence using antibodies against intermediate filaments of cytokeratin, desmin and vimentin type, and against proximal tubular brush border and distal tubular Tamm-Horsfall glycoprotein antigens, as well as with fluorochrome-labeled lectins in an attempt to study the origin and stage of differentiation of renal carcinomas. Eighty per cent of the renal carcinomas expressed the brush border antigens, whereas the Tamm-Horsfall glycoprotein could not be found. Antibodies against epidermal cytokeratins reacted only with collecting ducts in normal kidney, whereas antibodies against cytokeratins of Madin-Darby canine kidney epithelial cell line also reacted with glomerular and tubular epithelium. In 93% of the carcinomas tumor cells showed reactivity with both types of antikeratin antibodies. Vimentin, the cytoskeletal protein of mesenchymal cells, was present in the carcinoma cells of 53% of the tumors, although it was not present in normal tubular epithelium. Moreover, vimentin was expressed together with cytokeratin in the carcinoma cells in 57% of the keratin-positive samples as judged by double immunostaining, whereas the muscle type of intermediate filament protein, desmin, was not seen in the malignant cells. Binding sites for Lotus tetragonolobus agglutinin and soybean agglutinin, normally present in the cells of proximal tubules, were lacking or only faintly detectable in the neoplastic cells. Dolichos biflorus agglutinin, normally present in collecting ducts, was not detected in the tumors. The results show that most renal carcinomas express cytokeratin antigens as a sign of their epithelial origin and also show characteristics of proximal tubular cells. On the other hand, the results indicate that lectin-binding sites typical for normal differentiated tubular cells are profoundly modified in renal carcinomas. Ulex europaeus agglutinin did not bind to the malignant cells but decorated the endothelial cells of the tumors.

Antigens↗
PubMed1982

Ulex europaeus I lectin as a marker for vascular endothelium in human tissues.

Ulex europaeus I agglutinin, a lectin specific for some alpha-L-fucose-containing glycocompounds, was used in fluorescence microscopy to stain cryostat sections of human tissues. The endothelium of vessels of all sizes was stained ubiquitously in all tissues studied as judged by double staining with a known endothelial marker, antibodies against human clotting factor VIII. Cultured human umbilical vein endothelial cells, but not fibroblasts, also bound Ulex lectin. The staining was not affected by the blood group type of the tissue donor. In some tissues Ulex lectin presented additional binding to epithelial structures. Also, this was independent on the blood group or the ability of the tissue donor to secrete soluble blood group substances. Lotus tetragonolobus agglutinin, another lectin specific for some alpha-L-fucose-containing moieties failed to react with endothelial cells. Our results suggest that Ulex europaeus I agglutinin is a good histologic marker for endothelium in human tissues.

Blood Vessels↗
PubMedDate not supplied

Diabetes mellitus associated changes in glomerular glycocompounds: a fluorescence microscopical study.

Renal biopsy specimens showing histological alterations typical of advanced diabetic glomerulopathy were studied for changes in glomerular glycoconjugates, using fluorochrome-coupled lectins as probes. All samples studied showed a marked reduction in the binding of Triticum vulgaris (WGA) lectin in the glomerular basement membranes. On the other hand, 'new' glomerular binding sites for the lectins of Dolichos biflorus (DBA), Helix pomatia (HPA) and Arachis hypogaea (PNA), recognizing galactosyl moieties of glycoconjugates and giving no reaction in normal glomeruli, were seen in all samples studied. In addition, Wistaria floribunda lectin (WFA), recognizing galactosyl and, N-acetylgalactosaminyl configurations in glycoconjugates, gave a typical linear binding along the glomerular basement membranes, differing markedly from its reaction with normal kidney. Ulex europaeus (UEA I) showed reduced binding in the glomeruli of diabetic nephropathy. The results show that changes in glomerular glycoconjugates may appear in diabetic nephropathy, suggesting a disturbance in the turnover of the non-reducing terminal saccharide residues. In addition, the results show that lectins are useful probes for studying these changes further.

Adult↗
PubMedDate not supplied

mRNA differential display analysis of nephrotic kidney glomeruli.

BACKGROUND: Differential display RT-PCR (DDRT-PCR) is a new powerful technique for identification and characterization of altered gene expression in eukaryotic cells and tissues. We studied here changes in kidney glomerular gene expression in patients with congenital nephrotic syndrome of the Finnish type (CNF), an inherited kidney disease with heavy proteinuria already in utero. METHODS: Using the DDRT-PCR approach and isolated glomeruli from removed human kidneys, we compared the gene expression patterns of normal human and CNF glomeruli. Differential expression of candidate genes was verified by Northern blotting, and the corresponding PCR fragments were sequenced and compared to known sequences in databanks. RESULTS: We found several genes and sequence tags with altered expression in nephrotic glomeruli including fragments with close homologies to cytochrome c oxidase subunit I, integrin-linked kinase, insulin-like growth factor II receptor and eotaxin, and also clones resembling anchyrin and cadherin-like consensus sequences. CONCLUSION: All the sequences identified are of interest in respect to pathogenesis of proteinuria. Furthermore, this study reveals potentially new members to known gene families with tissue and cell type-specific expression.

Adult↗
PubMedDate not supplied

Nephrin mRNA regulation by protein kinase C.

BACKGROUND: Mutations in the recently cloned NPHS1 gene result in congenital nephrotic syndrome of the Finnish type (CNF). The protein product of NPHS1, nephrin, is expressed uniquely in kidney glomerular podocytes, and is the first true component of the interpodocyte slit membrane. The precise functions of nephrin remain unknown, but the presence of several tyrosine residues in the intracellular domain suggest a role in signalling. We searched for nephrin expressing cell line for use in signal transduction studies and also characterized the main features of calcium signalling in nephrin-deficient cultural glomerular epithelial cells. METHODS: We used A293 cell line, found to naturally express nephrin, as well as cultured CNF glomerular cells using reverse-transcription PCR, immunocytochemistry and intracellular Ca2+ measurements. RESULTS: Phorbol-12-myristate-13-acetate significantly upregulated the nephrin expression in A293 cells, while no change was found after treatment with additional stimulants for other main signalling pathways, e.g. okadaic acid, lysophosphatidic acid, bradykinin, angiotensin II (ANG II) and arginine vasopressin (AVP). No changes in basal or ANG II- or AVP-stimulated intracellular Ca2+ fluxes in CNF glomerular cells were observed. CONCLUSIONS: Protein kinase C may be the key intracellular signalling system in the regulation of nephrin.

Analysis of Variance↗
PubMedDate not supplied

Noncollagenous matrix components of glomeruli in congenital nephrotic syndrome of the Finnish type: evidence of abnormal splitting of nidogen?

The congenital nephrotic syndrome of the Finnish type (CNF) is a rare autosomal recessive disease with proteinuria starting already in utero, prematurity and nephrotic syndrome developing within the first weeks of life. The basic defect of this disease is unknown but has been suggested to be restricted to the kidney glomeruli and especially to the glomerular basement membrane (GBM). The location of the major matrix components in the glomeruli of CNF patient kidneys has previously been reported. Using indirect immunofluorescence microscopy we here describe the more recently characterized components of the glomerular extracellular matrix, including nidogen, tenascin, vitronectin and chondroitin sulfate proteoglycan in CNF and control kidney glomeruli. The accumulation of tenascin and chondroitin sulfate in the renal interstitium as well as a more granular deposition pattern of vitronectin in the mesangium of CNF glomeruli as compared to the control kidneys were observed. These changes were considered secondary to the massive proteinuria, reflected also by the presence of glomerular sclerosis and interstitial fibrosis in the CNF kidney samples. Additionally, analysis of GBM components by immunoblotting revealed either increased or decreased proportionate amounts of fibronectin and laminin in the GBM of CNF kidneys, respectively. Interestingly, different proportionate amounts of proteolytic fragments of nidogen were found in CNF glomeruli as compared to controls. Equal levels of nidogen mRNA were found in the cortical tissue of CNF and control glomeruli. Since nidogen is crucial for the supramolecular organization of basement membranes, these results suggest that an unusual fragmentation of nidogen, due to abnormal assembly, degradation or reorganization of glomerular extracellular matrix, may be associated with the basic defect of CNF.

Basement Membrane↗
PubMedDate not supplied

Expression of nine developmental stage-specific genes in human kidney and cultured renal cells.

BACKGROUND: Several genes transiently expressed during the maturation of the metanephrogenic mesenchyme have been reported in recent years while there is accumulating evidence of a reverted developmental pathway during tissue damage and loss of function. METHODS: Here we studied the expression of nine genes associated with kidney maturation from samples of normal human fetal, juvenile and adult kidneys and cultured glomerular cells using Northern blotting analysis. Subsequently, kidneys from patients with congenital nephrotic syndrome of the Finnish type (CNF), presenting with heavy proteinuria, and Wilms' tumor tissue were studied for the corresponding expression pattern for evidence of dedifferentiation/persistence of a fetal expression pattern. RESULTS: All the genes studied had their distinct expression pattern within the tissues and cells tested. No conclusive evidence of dedifferentiation could be obtained in the samples from CNF kidneys, whereas, as expected, the gene expression pattern of Wilms' tumor tissue was highly similar to that of fetal kidney tissue. CONCLUSION: Some genes thought to be only transiently expressed during kidney maturation were, however, constantly found in the samples from fetal to newborn and adult kidneys.

Adolescent↗