Biomedical subjects

M Godfrey

Publications and source records attributed to M Godfrey.

At least 37 records · Page 2Linked to original sources

PubMed1997

Pressure natriuresis and autoregulation of inner medullary blood flow in canine kidney.

We have evaluated the responses to changes in arterial pressure on regional blood flows in the renal medulla and sodium excretion simultaneously in denervated kidneys of six anesthetized sodium-replete dogs. Renal regional blood flow responses were determined using laser-Doppler needle flow probes and whole-kidney blood flow was assessed using an electromagnetic flow probe. The responses to stepwise reductions in renal arterial pressure (140 to 70 mm Hg) were examined first with a laser-Doppler needle probe inserted in the outer medulla and then repeated after advancing the same probe in the inner medulla. There were no differences in the control values of total renal blood flow (4.4 +/- 0.7 to 4.5 +/- 0.5 mL.min-1.g-1), glomerular filtration rate (0.89 +/- 0.7 to 0.94 +/- 0.9 mL.min-1.g-1), sodium excretion (3.6 +/- 0.6 to 3.4 +/- 0.5 mumol.min-1.g-1), and urinary excretion rate of nitric oxide metabolites (nitrate/nitrite, 1.6 +/- 0.2 to 1.5 +/- 0.2 nmol.min-1.g-1) at the start of both experimental periods. During changes in renal arterial pressure, inner medullary blood flow exhibited efficient autoregulation similar to that in outer medullary blood flow. Usual excretory responses to reductions in renal arterial pressure as well as autoregulation of cortical and whole-kidney blood flows and glomerular filtration rate were observed in these dogs. The slopes of the relationship between arterial pressure and sodium excretion (0.046 +/- 0.007 to 0.044 +/- 0.009 mumol.min-1.g-1.mm Hg-1) or nitrate/nitrite excretion (0.014 +/- 0.003 to 0.013 +/- 0.003 nmol.min-1.g-1.mm Hg-1) were similar in both experimental periods. These data indicate that blood flow to the inner medulla is efficiently autoregulated as in outer medulla and cortex of the kidney in anesthetized dogs and demonstrate further that the arterial pressure-induced natriuretic responses do not require associated changes in medullary blood flow.

Animals↗

PubMed1997

Pressure natriuresis and renal medullary blood flow in dogs.

In the present study, we evaluated the effects of changes in arterial pressure on regional renal blood flows and sodium excretion in anesthetized dogs during control conditions and after 5% volume expansion with isotonic saline. Medullary and cortical blood flow responses were determined with laser-Doppler needle flow probes inserted into the midmedullary and midcortical regions, and whole-kidney blood flow was assessed with an electromagnetic flow probe. Volume expansion in six dogs caused marked increases in urine flow (20.2 +/- 5.5 to 82.5 +/- 22.7 microL.min-1.g-1) and sodium excretion (3.2 +/- 0.5 to 11.1 +/- 2.7 mumol.min-1.g-1), with slight increases in glomerular filtration rate (0.92 +/- 0.03 to 1.01 +/- 0.02 mL.min-1.g-1) but no significant changes in total renal blood flow (4.7 +/- 0.3 to 5.2 +/- 0.6 mL.min-1.g-1), medullary blood flow (+6 +/- 9%), or cortical blood flow (+12 +/- 10%). During stepwise reductions in renal arterial pressure (150 to 75 mm Hg) elicited with a renal arterial occluder, both before and after volume expansion, medullary, cortical, and total renal blood flows as well as glomerular filtration rate exhibited efficient autoregulation, with slopes not significantly different from zero over this range of arterial pressure. Ther were marked increases in the slopes of the relationships between arterial pressure and urine flow (0.18 +/- 0.05 to 0.78 +/- 0.27 microL.min-1.g-1.mm Hg-1) as well as sodium excretion (0.03 +/- 0.004 to 0.10 +/- 0.03 mumol.min-1.g-1.mm Hg-1) during volume expansion. These data demonstrate that medullary blood flow is efficiently autoregulated in dogs during control and volume-expanded states and indicate that the mechanism responsible for the arterial pressure-induced changes in sodium excretion does not depend on coincident alterations in medullary blood flow.

Animals↗

PubMed1996

Weill-Marchesani syndrome--possible linkage of the autosomal dominant form to 15q21.1.

Weill-Marchesani syndrome comprises short stature, brachydactyly, microspherophakia, glaucoma, and ectopia lentis is regarded as an autosomal recessive trait (McKusick 277600). We present two families each with affected individuals in 3 generations demonstrating autosomal dominant inheritance of Weill-Marchesani syndrome. Linkage analysis in these 2 families suggests a gene for Weill-Marchesani syndrome maps to 15q21.1. The dislocated lenses and connective tissue disorder in these families suggests that fibrillin-1 and microfibril-associated protein 1, which both map to 15q21.1, are candidate genes for Weill-Marchesani syndrome. Immunohistochemistry staining of skin sections from family 1 showed an apparent decrease in fibrillin staining compared to control individuals.

Abnormalities, Multiple↗

PubMed1996

Delivery of a hammerhead ribozyme specifically down-regulates the production of fibrillin-1 by cultured dermal fibroblasts.

The hammerhead ribozyme is a small catalytic RNA molecule. Potential hammerhead ribozymes that possess a catalytic domain and flanking sequence complementary to a target mRNA can cleave in trans at a putative cleavage site within the target molecule. We have investigated the potential of hammerhead ribozymes to down-regulate the product of the fibrillin-1 gene (FBN1). Fibrillin is a 347 kDa glycoprotein that is a major constituent of the elastin-associated microfibrils. Mutations in the FBN1 gene are responsible for Marfan syndrome (MFS), a common systemic disorder of the connective tissue. Many FBN1 mutations responsible for MFS appear to act in a dominant-negative fashion, raising the possibility that reduction of the amount of product from the mutant FBN1 allele might be a valid therapeutic approach for MFS. A trans-acting hammerhead ribozyme (FBN1-RZ1) targeted to the 5' end of the human FBN1 mRNA has been designed and synthesized, and shown to cleave its target efficiently in vitro. FBN1-RZ1 cleavage is magnesium dependent and efficient at both 37 and 50 degrees C. Delivery of the FBN1-RZ1 ribozyme into cultured dermal fibroblasts, by receptor-mediated endocytosis of a ribozyme-transferrin-polylysine complex, specifically reduces both cellular FBN1 mRNA and the deposition of fibrillin in the extracellular matrix. These results suggest that the use of hammerhead ribozymes is a valid approach to the study of fibrillin gene expression and possibly to the development of a therapeutic approach to MFS.

Base Sequence↗

PubMed1996

An accurate method for comparing transcript levels of two alleles or highly homologous genes: application to fibrillin transcripts in Marfan patients' fibroblasts.

We introduce here a novel and generally applicable, solid-phase minisequencing-based approach for rapid estimation of relative levels of transcripts with high sequence homology. This study was undertaken to screen for the consequences of different fibrillin-1 mutations on the transcript levels in patients with the Marfan syndrome (MFS). This dominantly inherited, connective tissue disorder is characterized by pleiotrophic symptoms in cardiovascular, skeletal, and ocular systems. A spectrum of disease mutations in the gene encoding fibrillin-1 (FBN1), a glycoprotein component of extracellular matrix microfibrils, has been identified in MFS patients, but the mechanisms by which mutations result in different phenotypic manifestations are still unknown to a large extent. Our data from the quantitation of FBN1 transcripts provide support for the hypothesis that mutations causing premature stop codons result in a milder phenotype than classical MFS by reducing the stability of the mutant transcript and, consequently, decreasing the interference of mutant polypeptide in the formation of fibrillin fibers. We also applied this mRNA quantitation method to determine the relative ratio between transcripts from the genes coding for two highly homologous microfibrillar components, FBN1 and FBN2, in control fibroblast cultures as well as in fibroblasts from MFS patients. Interestingly, these data show large variations between the levels of the two transcripts in fibroblast cultures, but these variations do not correlate either with the nature of the disease mutation or to the clinical MFS phenotype.

Alleles↗

PubMed1996

Double mutant fibrillin-1 (FBN1) allele in a patient with neonatal Marfan syndrome.

It is now well established that defects in fibrillin-1 (FBN1) cause the variable and pleiotropic features of Marfan syndrome (MFS) and, at the most severe end of its clinical spectrum, neonatal Marfan syndrome (nMFS). Patients with nMFS have mitral and tricuspid valve involvement and aortic root dilatation, and die of congestive heart failure, often in the first year of life. Although mutations in classical MFS have been observed along the entire length of the FBN1 mRNA, mutations in nMFS appear to cluster in a relatively small region of FBN1, approximately between exons 24 and 34. Here we describe the appearance of two FBN1 mutations in a single allele of an infant with nMFS. The changes were within six bases of each other in exon 26. One was a T3212G transversion resulting in an I1071S amino acid substitution and the second was an A3219T transversion resulting in an E1073D amino acid substitution. This is the first reported double mutant allele in FBN1.

Alleles↗

PubMed1996

Familial occurrence of typical and severe lethal congenital contractural arachnodactyly caused by missplicing of exon 34 of fibrillin-2.

Genetic linkage studies have linked congenital contractural arachnodactyly (CCA), a usually mild heritable connective-tissue disorder, to FBN2, the fibrillin gene on chromosome 5. Recently, FBN2 mutations in two patients with CCA have been described. Here we report an A-->T transversion at the -2 position of the consensus acceptor splice site, resulting in the missplicing of exon 34, a calcium-binding epidermal growth factor-like repeat in fibrillin-2 in a mother and daughter with CCA. Significantly, the mother exhibited a classic CCA phenotype with arachnodactyly, joint contractures, and abnormal pinnae, whereas her daughter exhibited a markedly more severe CCA phenotype, which included cardiovascular and gastrointestinal anomalies that led to death in infancy. Analysis of cloned fibroblasts showed that the mother is a somatic mosaic for the exon 34 missplicing mutation, whereas all the daughter's cells harbored the mutation.

Adult↗

PubMed1995

Prenatal and presymptomatic diagnosis of the Marfan syndrome using fluorescence PCR and an automated sequencer.

The Marfan syndrome (MFS) is a heritable connective tissue disorder characterized by skeletal, ocular, and cardiovascular abnormalities. Defects in fibrillin, an elastin-associated microfibrillar protein, are now known to cause MFS. Since the discovery of fibrillin as the gene responsible for MFS, requests for prenatal and presymptomatic diagnosis have become common-place. Here we report the use of the polymerase chain reaction (PCR), using fluorescence labelled primers and an automated sequencer, to establish linkage data for 'molecular diagnosis'. The mistaken clinical diagnosis of MFS based on the appearance of a common cardiovascular manifestation, mitral valve prolapse, and a positive family history is also discussed.

Autoanalysis↗

PubMed1995

Fibrillin immunofluorescence in pseudoxanthoma elasticum.

BACKGROUND: Pseudoxanthoma elasticum (PXE) is a rare heritable connective tissue disorder manifested by skin, ocular, and cardiovascular anomalies. The basic defect is unknown; however, the microscopic findings are indicative of defects in elastic fibers. Among the components of the elastic fibers are elastin and elastin-associated microfibrils. OBJECTIVE: We assessed the fidelity of this fibrillar system in PXE with the use of antibodies to fibrillin, a major component of elastin-associated microfibrils. METHODS: Using a well-established immunofluorescence assay, we studied fibrillin deposition in dermal fibroblast cultures from 16 patients with PXE. RESULTS: Six of the 16 patients (37%) showed some abnormality of fibrillin deposition in fibroblasts derived from lesional skin. Fibroblasts from nonlesional skin displayed normal fibrillin immunofluorescence. The only sibship studied, however, was discordant for fibrillin immunostaining. CONCLUSION: Unlike the findings in Marfan syndrome, these data are not suggestive of causal fibrillin defects in PXE.

Actin Cytoskeleton↗

PubMed1995

Recurrent mis-splicing of fibrillin exon 32 in two patients with neonatal Marfan syndrome.

The Marfan syndrome (MFS) is an autosomal dominant heritable disorder of connective tissue. Variable and pleiotropic clinical features are observed in the skeletal, ocular, and cardiovascular systems. The most severe end of the phenotypic spectrum of this disorder comprises a group of patients usually diagnosed at birth, who have a life expectancy of little more than a year. To distinguish this group of patients from those with classical MFS, we refer to them as neonatal Marfan syndrome (nMFS). These infants usually die of congestive heart failure rather than aortic aneurysmal disease, the most frequent cause of morbidity and mortality in classical MFS. Defects in fibrillin, an elastin-associated microfibrillar glycoprotein, are now known to cause both the classical and neonatal forms of MFS. Here we report the recurrent mis-splicing of fibrillin (FBN1) exon 32, a precursor EGF-like calcium binding domain, in two unrelated infants with nMFS. The mis-splicing, in one patient, was due to an A-->T transversion at the -2 position of the consensus acceptor splice site; while that in the second patient was caused by a G-->A transition at the +1 position of the donor splice site. Characterization of FBN1 mutations in individuals at the most severe end of the Marfan syndrome spectrum should provide greater understanding of the multiple domains and regions of fibrillin.

Alleles↗

PubMed1995

Quantitation of fibrillin immunofluorescence in fibroblast cultures in the Marfan syndrome.

The Marfan syndrome (MFS) is a heritable connective tissue disorder manifested by defects in the skeletal, ocular, and cardiovascular systems. Diagnosis of MFS is based on clinical findings. At present there are no laboratory tests for specific determination of this disorder. Defects in fibrillin, an elastin-associated microfibrillar glycoprotein, are now known to cause the variable and pleiotropic manifestations of MFS. Immunofluorescence studies of skin sections and dermal fibroblast cultures were the first to show this association. Most unequivocal cases of the Marfan syndrome exhibited an apparent reduction in fibrillin immunofluorescence. The prospect of examining patients whose clinical findings suggest a possible diagnosis of the Marfan syndrome has stimulated us to attempt quantitation of immunofluorescence. In the study described here we used computer-enhanced image analysis to establish "normal" and "abnormal" (Marfan) parameters of fibrillin immunofluorescence in dermal fibroblast cultures. Quantitation of fluorescence from control individuals showed a median of 21%, while the median fibrillin fluorescence in MFS patients was 6% with a confidence interval of < or = 15%. These findings were statistically significant to p < 0.01. It is hoped that these analyses may become a useful adjunct in the clinical diagnosis of MFS.

Cells, Cultured↗

PubMed1995

Low dose aspirin for the treatment of fetal growth restriction: a randomized controlled trial.

The purpose of this study was to investigate the hypothesis that maternal administration of 100mg aspirin each day will improve birth-weight and other measures of neonatal size when given as a treatment to pregnancies complicated by fetal growth restriction and umbilical-placental insufficiency. A randomized, double-blind, placebo controlled study design was employed; 51 pregnant women were enrolled. The entry criteria were a fetal abdominal circumference < 10th per centile together with an umbilical artery Doppler systolic/diastolic ratio > 95th per centile between 28 and 36 weeks' gestation. Compliance was assessed by serial measurement of maternal serum thromboxane B2 levels. The mean gestational age at enrolment was 32 weeks and at delivery was 36 weeks. There were no differences between the 2 groups in gestational age at birth; birth-weight or birth-weight ratio; circumferences of the head, chest or abdomen; skin fold thicknesses; or neonatal morbidity. Low dose aspirin therapy did not alter Doppler systolic/diastolic ratios. After 14 days therapy, mean thromboxane B2 levels fell more than 80% from baseline values; 10.5% of women did not demonstrate biochemical confirmation of aspirin ingestion, despite verbal confirmation of compliance. We conclude that low dose aspirin therapy is not of benefit in the treatment of pregnancies complicated by fetal growth restriction and umbilical-placental insufficiency between 28 and 36 weeks' gestation.

Adult↗

PubMed1995

Relation between pressure natriuresis and urinary excretion of nitrate/nitrite in anesthetized dogs.

Alterations in intrarenal nitric oxide (NO) formation during changes in renal arterial pressure (RAP) have been suggested as a mechanism mediating pressure natriuresis. To test this hypothesis further, we examined the relation between RAP and the urinary excretion rate of nitrate/nitrite (NO3-/NO2-; NO metabolites) in anesthetized sodium-replete dogs before (n = 9) and during (n = 6) intrarenal infusion of the NO synthesis inhibitor nitro-L-arginine (NLA; 50 micrograms.kg-1.min-1). Urinary NO3-/NO2- concentrations were measured with the Griess reaction and spectrophotometry methods after enzymatic reduction of NO3- to NO2- in the samples. During control conditions, there were decreases in the urinary NO3-/NO2- excretion rate in response to reductions in RAP (150 to 75 mm Hg; slope, 0.04 +/- 0.01 nmol.min-1.g-1.mm Hg-1) in association with decreases in urinary sodium excretion (UNaV). There was a positive correlation between changes in NO3-/NO2- excretion rate and changes in RAP (r = .48; P < .005) or UNaV (r = .59; P < .001). NLA infusion resulted in decreases in NO3-/NO2- excretion rate (4.8 +/- 1.4 to 1.0 +/- 0.3 nmol.min-1.g-1) in association with reductions in UNaV (4.3 +/- 0.3 to 0.7 +/- 0.2 microL.min-1.g-1), fractional excretion of sodium (2.9 +/- 0.2% to 0.5 +/- 0.1%), and renal blood flow (4.8 +/- 0.3 to 3.3 +/- 0.2 mL.min-1.g-1), without changes in glomerular filtration rate. Furthermore, there was a marked attenuation of the NO3-/NO2- and sodium excretory responses to alterations in RAP during NO synthesis inhibition.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Oxidoreductases↗

PubMed1995

Abnormal morphology of fibrillin microfibrils in fibroblast cultures from patients with neonatal Marfan syndrome.

The Marfan syndrome (MFS) is a connective tissue disorder manifested by variable and pleiotropic features in the skeletal, ocular, and cardiovascular systems. The average life span in MFS is about 35 years. A group with much more severe cardiovascular disease and a mean life span of approximately 1 year also exists. We refer to this latter group as "neonatal Marfan syndrome" (nMFS). Fibrillin defects are now known to be the cause of MFS and nMFS. Immunofluorescence studies were the first to demonstrate this association. Here we describe immunofluorescence studies in a series of 10 neonates and summarize their salient clinical features. In vitro accumulation of fibrillin reactive fibers was assayed using monoclonal antibodies to fibrillin in hyperconfluent fibroblast cultures. As was previously observed in MFS, fibroblast cultures from nMFS patients showed an apparent decrease in accumulation of immunostainable fibrillin. Significantly, however, the morphology of the immunostained fibrils in the nMFS cultures were abnormal and differed not only from control cultures, but also from those seen in cultures of MFS fibroblasts. The nMFS fibrils appeared short, fragmented, and frayed, characteristics that are not seen in MFS. Both the clinical and fibrillin morphology data provide evidence to suggest a useful subclassification of nMFS in the spectrum of MFS.

Cells, Cultured↗

PubMed1994

Phase angle changes of photically entrained circadian rhythms following a single nonphotic stimulus.

Syrian hamsters entrained to a light-dark (LD) cycle of 14:10 h were given the opportunity to run in novel wheels for 3 h in the middle of the light phase. This manipulation transiently altered the phase angle of entrainment to the LD cycle: activity onset was significantly advanced (by about 0.5 h) on the day after the pulse and gradually drifted back toward its prepulse time. When animals were held in LD 11.5:12.5 h, a photoperiod in which onset time occurs later relative to the time of lights-off, they again advanced about 0.5 h in response to the pulse of wheel running, but many animals retained an advanced phase angle for at least 7 days, and some for more than 21 days. Individual changes in phase angle were highly correlated with the prepulse phase angle: the more negative the phase angle, the greater the advance subsequent to the novel wheel pulse. These results show that a single, short-duration, nonphotic manipulation can produce long-lasting alterations in the phase angle of entrainment to a LD cycle.

Animals↗

PubMed1994

Melanin as an adsorbent for drug residues.

The binding of seven veterinary drugs (clenbuterol, chlorpromazine, diethylstilbestrol, 19-nortestosterone, salbutamol, salicylic acid and trenbolone) to melanin from Sepia officinalis was investigated. Basic and hydrophobic drugs were the most strongly bound. Desorption by ethanol was complete for neutral drugs but only partial for the basic drugs, which suggests that binding of the latter involves an ionic component. A method of synthesizing melanin in an immobilized form (melanin-PS) on the surface of porous silica was developed. When the drug binding properties of melanin-PS were investigated, its capacity to bind the basic drug clenbuterol was found to be higher (5.9 nmol mg-1) than that for the neutral hydrophobic drug 19-nortestosterone (0.56 nmol mg-1); for both drugs the attainment of binding equilibrium with melanin-PS was relatively rapid (< 5 min). By virtue of its binding kinetics, high capacity and mechanical robustness, melanin-PS offers potential for use in chromatography or solid-phase extraction and may additionally enable modelling of drug-melanin interactions.

Adsorption↗

PubMed1994

From fluorescence to the gene: the skin in the Marfan syndrome.

The Marfan syndrome (MFS) is a heritable connective tissue disorder manifested by defects in the skeletal, ocular, and cardiovascular systems. It is inherited as an autosomal dominant trait, with an incidence of nearly one per 10,000 population without gender or ethnic predilection. About 30% of MFS cases are sporadic, representing new dominant mutations. Although diagnostic criteria may be met, intra- and inter-familial variability in syndromic expression can be quite marked. Life expectancy is significantly reduced, and more than 85% of patients die of cardiovascular complications. Elastin-associated microfibrils are classically identified by a 10-12 nm cross-sectional diameter and close association with amorphous elastin. Characteristically, these microfibrils are seen as a fringe surrounding elastin cores. Despite their name, these fibrils are frequently localized in tissues not associated with elastin. One of the major and best characterized microfibrillar proteins is the glycoprotein, fibrillin. Fibrillin is rich in cysteine residues and thus, extensively intrachain disulfide bonded. Defects in fibrillin are now known to cause the variable and pleiotropic manifestations of MFS. Immunofluorescence studies of skin sections and dermal fibroblast cultures were the first to show this association. The cloning and sequencing of the entire fibrillin coding region has enabled the identification of the myriad of mutations that cause MFS. Identification of the molecular lesions covering the gamut of MFS clinical variability should allow the construction of genotype/phenotype correlations. It is hoped that once available, they may become of prognostic value in the clinical management of MFS.

Actin Cytoskeleton↗

PubMed1993

Decreased extracellular deposition of fibrillin and decorin in neonatal Marfan syndrome fibroblasts.

Abnormalities of the microfibrillar protein fibrillin (Fib) have been reported in Marfan syndrome (MFS). The so-called neonatal Marfan syndrome (nMFS) is a lethal phenotype displaying features that are not seen in classical MFS. We have therefore studied the biosynthesis and extracellular deposition of Fib and decorin in fibroblasts from a patient with nMFS and controls. Immunofluorescence of the patient's cell cultures showed an almost complete absence of Fib and a marked reduction of decorin in the extracellular matrix (ECM). The nMFS skin revealed Fib on subbasal microfibrillar bundles in the papillary dermis, and Fib associated with elastic fibers in the reticular dermis; the bundles and fibers were fragmented and thinner than normal. Pulse-chase labeling of cells with [35S]Met/Cys revealed moderately reduced secretion, but a diminished deposition of Fib in the ECM; this was more apparent at a longer chase time. Fib mRNA and synthesis appeared to be normal, whereas both decorin mRNA and biosynthesis were reduced. We therefore assume a structural Fib defect in this patient causing reduced deposition into and/or enhanced removal from the ECM, whereas the reduced decorin biosynthesis may be a secondary regulatory phenomenon. The clinical relevance of this remains unclear. Our findings imply that Fib defects may be responsible for the severe, complex phenotype of nMFS.

Blotting, Northern↗