Managing risk. A priority in the health service.
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Biomedical subjects
S Fleming
Publications and source records attributed to S Fleming.
Calcium dependence of A-CAM function in human renal epithelium.
Previously we have used immunocytochemistry to identify A-CAM as a major cell adhesion molecule in human renal tubular epithelium. In this study, we demonstrate the calcium dependence of cell-cell adhesion and A-CAM function in cultured human renal epithelial cells. Separation of adjacent cells was seen within 1 min following sequestration of calcium from the culture medium with EDTA. This loss of intracellular adhesion in low calcium conditions was accompanied by the disappearance of A-CAM immunoreactivity. The changes were reversible and on return to normal calcium concentrations, A-CAM immunoreactivity reappeared and cell adhesion was re-established. The time course of this recovery was slow, taking 2-3 h. These results demonstrate the calcium dependence of cell adhesion in cultured human renal epithelial cells and confirm the adhesive function of the immunoreactive A-CAM molecule.
Antigens, CD↗
A tumour promoter induces alterations in vinculin and actin distribution in human renal epithelium.
Loss of cell-substratum adhesion is an important factor during tumour progression. We have previously described reduced focal contact components and poorly organized cytoskeletal actin in renal cell carcinomas. In this study, we have used the potent tumour promoter TPA on cultured human renal epithelium to mimic neoplastic transformation. The morphological changes induced by TPA were examined by phase contrast and fluorescence microscopy. TPA treatment caused rounding up of cells and loss of adhesion to either fibronectin or laminin substrata. Cytoskeletal actin was redistributed from orientated stress fibre bundles to a perinuclear circumferential arrangement. This was accompanied by a progressive reduction in the number of vinculin-containing contacts with accumulation of vinculin in punctate spots in the perinuclear region. These altered membrane-cytoskeletal interactions induced by TPA are entirely reversible and mimic epigenetic changes which occur during tumour progression.
Actins↗
Differential expression of integrin alpha chains by renal epithelial cells.
We have investigated the expression of integrin chains by human renal epithelial cells during in vivo renal differentiation and in vitro cell culture on different extracellular matrices. Using the immunoperoxidase technique to visualize the binding of monoclonal antibodies to different integrin chains in fetal and adult kidneys, we found a change during development from alpha 1 beta 1, alpha 3 beta 1, and alpha 4 beta 1-positive blastemal cells to alpha 2 beta 1, alpha 3 beta 1, and alpha 6 beta 1-positive epithelial cells. The pattern of integrin expression correlates with the presence in the extracellular matrix of the appropriate ligands. In in vitro cell culture experiments, renal epithelium expressed alpha 3 and alpha 5 integrins on all extracellular matrices. Integrins alpha 2 and alpha 6 were found only in cells grown on a laminin-containing substratum. Fibronectin and alpha 5 integrin co-localized on the ventral surface of cells grown on a laminin substratum and at the periphery of cells on glass coverslips. These results suggest that there is a close relationship between integrin alpha chain usage and the presence of appropriate ligands in the extracellular matrix.
Antibodies, Monoclonal↗
Polycystic kidney disease in the CBA/N immunodeficient mouse.
We describe a polycystic lesion of the kidney in the CBA/N mouse with an X-linked recessive immunodeficient syndrome. There is progressive cystic dilatation affecting all parts of the nephron. The cyst lining is composed of a single layered epithelium with focal nuclear crowding and the formation of micropapillary structures. The cystic epithelial cells show subnuclear vacuolation. Focal basement membrane thickening is also a feature. There is no significant inflammatory infiltrate present within these kidneys. Electron microscopic examination reveals that the subnuclear vacuolation is due to loss of the membrane infoldings at the basal pole of the epithelial cell with fluid accumulation within the extracellular space. The basement membrane thickening is due to expansion of the lamina densa. These changes are not present at birth but develop progressively with age. The finding of a polycystic kidney lesion in these mice offers an opportunity to investigate the relationship between the immune system and renal cyst formation.
Age Factors↗
Isolated basal keratinocytes express pemphigoid gestationis antigen.
Basal keratinocytes were isolated from epidermal cell suspensions prepared by trypsinization of normal human skin. Cells were identified as basal cells by their adherence to collagen and confirmed as basal cells by the presence of pemphigoid antigen. Using an indirect immunofluorescence assay, cells were found to express pemphigoid gestationis-related antigen. Sera from patients with pemphigoid gestationis reacted in one of two immunofluorescence patterns: either polar, in a pattern similar to that observed with bullous pemphigoid serum, or with uniform staining around the cell periphery. Pemphigoid gestationis-related antigen is expressed by isolated basal keratinocytes and is resistant to trypsinization. The heterogeneity of immunofluorescence patterns may correspond to the heterogeneity of antigen recognition by different patients with pemphigoid gestationis.
Antigens↗
MAC387: its non-specificity as a tumour marker or marker of histiocytes.
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Antibodies, Monoclonal↗
Increased energy intake minimizes weight loss in men at high altitude.
The hypothesis that high-altitude weight loss can be prevented by increasing energy intake to meet energy requirement was tested in seven men, 23.7 +/- 4.3 (SD) yr, taken to 4,300 m for 21 days. Energy intake required to maintain body weight at sea level was found to be 3,118 +/- 300 kcal/day, as confirmed by nitrogen balance. Basal metabolic rate (BMR), determined by indirect calorimetry, increased 27% on day 2 at altitude and then decreased and reached a plateau at 17% above the sea level BMR by day 10. Energy expended during strenuous activities was 37% lower at altitude than at sea level. Fecal excretion of energy, nitrogen, total fiber, and total volatile fatty acids was not significantly affected by altitude. Energy intake at altitude was adjusted after 1 wk, on the basis of the increased BMR, to 3,452 +/- 452 kcal/day. Mean nitrogen balance at altitude was negative (-0.25 +/- 0.71 g/day) before energy intake was adjusted but rose significantly thereafter (0.20 +/- 0.71 and 0.44 +/- 0.66 g/day during weeks 2 and 3). Mean body weight decreased 2.1 +/- 1.0 kg over the 3 wk of the study, but the rate of weight loss was significantly diminished after the increase in energy intake (201 +/- 75 vs. 72 +/- 48 g/day). Individual regression lines drawn through 7-day segments of body weight showed that in four of seven subjects the slopes of body weight were not significantly different from zero after the 2nd wk. Thus weight loss ceased in four of seven men in whom increased BMR at altitude was compensated with increased energy intake.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Cytogenetic profile of 109 lipomas.
Cytogenetic analysis of short-term cultures was carried out on 109 lipomas from 92 patients. Clonal chromosomal abnormalities were present in 50% of the tumors analyzed. Based on the results, three main cytogenetic groups were identified and included: (a) tumors with normal karyotypes, (b) tumors with abnormalities involving region q13-15 on chromosome 12, and (c) tumors with other clonal aberrations. Within each of these groups, cytogenetic subgroups could be identified, each characterized by a specific anomaly. Tumors with abnormalities of 12q included specific subgroups with t/ins(1;12)(p32-33;q13-15), t(2;12)(p21-22;q13-14), t(3;12)(q28;q14), t(12;21)(q13;q21), complex, and nonrecurrent aberrations. The group containing heterogeneous clonal aberrations included subgroups with del(13)(q12q22), der(6)(p21-23), der(11)(q13), and nonspecific aberrations. Chromosome bands 1p36, 1p32-33, 2p21-22, 3q27-28, 6p21-23, 11q13, 12q13-15, 13q12, 13q22, 17p13, 17q21, and 21q21-22 were preferentially involved in structural rearrangements in lipomas. The identification of these sites of nonrandom rearrangements may serve to identify genes (at or near the junctions of chromosomal aberrations) involved in normal cellular growth control. Statistical analysis of the data revealed a correlation among karyotypic abnormalities and clinical data, such as age and sex of the patient, and tumor depth, site, and size.
Chromosome Aberrations↗
Morphological evidence that A-CAM is a major intercellular adhesion molecule in human kidney.
We have used immunocytochemistry to identify the major primary adhesion molecule of the cadherin class in human kidney. In frozen sections of kidney, A-CAM was detected using the monoclonal antibody GC4 on the surface of renal tubular epithelial cells. Renal tubular epithelium did not express L-CAM. No cadherin reactivity was found on the glomerular epithelial cells. Cultured renal tubular epithelium was studied by immunofluorescence and immunogold methods. A-CAM was found at the contact points of adjacent epithelial cells, the phenotype of which was confirmed by the demonstration of cytokeratins using the antibody CAM 5.2. The A-CAM molecule in human kidney had an Mr of 130 kD in Western blotting experiments. These results lead us to conclude that A-CAM is the major cadherin of adult human renal epithelium.
Antibodies, Monoclonal↗
C. L. Oakley Lecture (1991). Cell adhesion and epithelial differentiation.
The differentiated function of epithelial tissues is dependent on the molecules which mediate the cohesion and polarization of the epithelium. Separate molecular families are responsible for the adhesion between adjacent cells and the adhesion between cells and the substratum. Using the kidney as a model of epithelial differentiation it has been shown that the assembly of a basal lamina containing the glycoprotein laminin and cadherin mediated intercellular adhesion are key events. The Wilms' tumour anti-oncogene plays a regulatory role in renal epithelial differentiation. Tissue modeling and tumour formation are both accompanied by reduced cell adhesion molecule function.
Cell Adhesion↗
Composition and organization of cell-substratum contacts in normal and neoplastic renal epithelium.
We have investigated the molecular basis of the organization of cell-substratum contact in normal and neoplastic renal epithelium. The several components of focal contacts and non-collagenous basement membrane glycoproteins have been identified by antibodies, detected by immunofluorescence and viewed by laser scanning microscopy. Tubular epithelial cells grown on glass coverslips expressed laminin at the cell periphery. Co-localized with laminin were receptors of the beta 1 integrin class, and the cytoplasmic plaque proteins vinculin and talin. When basement membrane glycoproteins laminin or fibronectin were exogenously supplied by growing cells on coated coverslips, the vinculin-, talin- and integrin-containing contacts became organized into linear arrays with intervening free cell membranes. Under these conditions the actin cytoskeleton became highly organized. By contrast some tumour cells showed a reduction in focal contact molecules and a failure to organize these structures in response to laminin or fibronectin. This loss of cell matrix interaction may be important during the progression of renal tumours.
Basement Membrane↗
The effect of inhibitors of platelet aggregation on the metabolism of platelet-activating factor (PAF) in washed rabbit platelets.
The rabbit platelet metabolizes platelet-activating factor (PAF) intracellularly. PAF is deacetylated to produce lysoPAF which, in turn, can be acylated to produce 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine (alkylacyl GPC). Some PAF receptor antagonists have been shown to inhibit this metabolic conversion. In the present study we examined whether the PAF receptor antagonists SRI 63-441 and WEB 2086 would inhibit the metabolism of PAF by intact rabbit platelets. In addition, we examined whether iloprost, a stable analogue of prostaglandin I2 (PGI2), and a potent inhibitor of platelet activation induced by a range of agonists, would also inhibit PAF metabolism. We found that SRI 63-441 and WEB 2086 caused an almost complete inhibition of the conversion of PAF to alkylacyl GPC. Iloprost caused up to a 50% inhibition of PAF metabolism compared to antagonist-free controls. Iloprost (and PGI2) is thought to inhibit platelet response by elevation of cAMP, while receptor antagonists act by blocking PAF binding to its receptor. Since iloprost caused partial inhibition of PAF metabolism, the results of this study suggest that inhibition of PAF metabolism does not occur solely due to competitive inhibition of PAF binding to its receptor.
Animals↗
Morphological evidence for calcium-dependent association of calgranulin with the epidermal cytoskeleton in inflammatory dermatoses.
The association of calgranulins, intracellular calcium-binding proteins, with the keratinocyte cytoskeleton has been studied. These molecules are expressed in various inflammatory dermatoses and in organ-culture explants. Triton X-100 extraction in the presence of calcium or EDTA suggested that calgranulins are detergent insoluble in the presence of calcium. The molecules were localized in a plaque-like structure at the cell periphery in lesional skin and in organ-culture explants. Following induction of calgranulins in vitro there was a redistribution of the intermediate filament cytoskeleton into a perinuclear halo, although desmosomes remained intact. These various features suggest that these members of the S-100 protein family have a role in cytoskeletal changes seen in various skin diseases.
Blotting, Western↗
11 beta-Hydroxysteroid dehydrogenase mRNA expression in rat kidney.
11 beta-Hydroxysteroid dehydrogenase (11 beta-OHSD) protects nonspecific renal mineralocorticoid receptors from exposure to circulating glucocorticoid in vivo by catalyzing the conversion of corticosterone to inactive 11-dehydrocorticosterone. Although 11 beta-OHSD bioactivity and aldosterone binding sites are found in distal tubular cells, mineralocorticoid receptor and 11 beta-OHSD immunoreactivities are not colocalized. However, there are several kidney isoforms of 11 beta-OHSD, not all of which may be immunoreactive, whereas only a single mRNA species has been described. Using in situ hybridization we found 11 beta-OHSD mRNA is highly expressed in all renal tubular epithelia in the rat. It is therefore likely that 11 beta-OHSD is colocalized with mineralocorticoid receptors in distal tubular cells.
11-beta-Hydroxysteroid Dehydrogenases↗
Changes in rates of unscheduled hospital readmissions and changes in efficiency following the introduction of the Medicare prospective payment system. An analysis using risk-adjusted data.
The purpose of this study was to analyze changes in rates of unscheduled readmissions and changes in technical efficiency following the introduction of the Medicare Prospective Payment System (PPS). We developed the Risk-Adjusted Readmissions Index (RARI), which allowed us to make comparisons in rates of unanticipated readmissions across hospitals and over time. Data envelopment analysis (DEA), a linear programming technique, was used to measure changes in technical efficiency by comparing the inputs used and the outputs produced across a cohort of hospitals, while adjusting for changes over time in case mix and case complexity. Rates of unscheduled readmissions and efficiency scores were computed for a sample of 245 hospitals for each year. Although both readmission rates and efficiency scores increased for most hospitals, there was no evidence that those hospitals that experienced the greatest increases in efficiency had the largest increases in their rates of unscheduled readmissions.
Abstracting and Indexing↗
Cell types expressing the Wilms' tumour gene (WT1) in Wilms' tumours: implications for tumour histogenesis.
Wilms' tumour (nephroblastoma), a childhood embryonal kidney tumour, is believed to arise from malignant transformation of abnormally persistent metanephric blastemal cells. At a histological level, tumours show a remarkable mimicry of the normal nephrogenic pathway. There is histological and epidemiological evidence for at least two pathogenetic groupings within Wilms' tumour which may reflect different timings of the tumorigenic insult in this pathway and/or involvement of different genes. Tumorigenesis is thought to result from loss of function of a so-called tumour-suppressor gene which has an essential role in control of normal genitourinary development. Such a candidate, Wilms' tumour gene (WT1) mapping to chromosome 11p13, has been isolated and is known to be mutated in some tumours. We have examined the cell types expressing this gene in 32 Wilms' tumours and in nephroblastomatosis by in situ mRNA hybridization. Our results show that WT1 is expressed only in neoplastic structures whose normal counterparts also express the gene and that abnormally persistent high levels of expression are common in both these lesions. Thus, WT1 expression is a good marker for tumour differentiation and reveals how the normal pattern of differentiation is disrupted in Wilms' tumours. We postulate that mutation of the WT1 gene at the 11p13 locus results in Wilms' tumours associated with intralobar nephrogenic rests, which frequently show stromal-predominant histology. We have used our results and ideas to reinterpret current theories on tumour histogenesis and propose a model which explains how patterns of epithelial differentiation are disrupted in Wilms' tumour and how malignant stroma can result from mutation in WT1.
Antibodies, Monoclonal↗
The candidate Wilms' tumour gene is involved in genitourinary development.
Wilms' tumour is an embryonic kidney tumour thought to arise through aberrant mesenchymal stem cell differentiation and to result from loss of function of a 'tumour suppressor' gene(s). Both sporadic and syndrome-associated Wilms' tumours are accompanied by an increased frequency of abnormalities of the urinary tract and genitalia. Deletional analysis of individuals with the WAGR syndrome (for, Wilms' tumour, aniridia, genitourinary abnormalities and mental retardation) showed that a Wilms' tumour gene lies at chromosomal position 11p13. This led to the isolation of a candidate Wilms' tumour gene, encoding a zinc-finger protein which is likely to be a transcription factor. To gain insight into the role of this candidate gene in normal development and tumorigenesis, we have now performed in situ messenger RNA hybridization on sections of human embryos and Wilms' tumours. The candidate Wilms' tumour gene is expressed specifically in the condensed mesenchyme, renal vesicle and glomerular epithelium of the developing kidney, in the related mesonephric glomeruli and in cells approximating these structures in tumours. The other main sites of expression are the genital ridge, fetal gonad and mesothelium. These data suggest that (1) this candidate is indeed a Wilms' tumour gene, (2) the associated genital abnormalities are pleiotropic effects of mutation in the Wilms' tumour gene itself, in support of recent genetic analysis, and (3) this gene has a specific role in kidney development and a wider role in mesenchymal-epithelial transitions.
Blotting, Northern↗