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D J Harris

Publications and source records attributed to D J Harris.

At least 37 records · Page 2Linked to original sources
PubMed1998

Novel mutations in the connexin 26 gene (GJB2) that cause autosomal recessive (DFNB1) hearing loss.

Mutations in the connexin 26 (Cx26) gene (GJB2) are associated with the type of autosomal recessive nonsyndromic neurosensory deafness known as "DFNB1." Studies indicate that DFNB1 (13q11-12) causes 20% of all childhood deafness and may have a carrier rate as high as 2. 8%. This study describes the analysis of 58 multiplex families each having at least two affected children diagnosed with autosomal recessive nonsyndromic deafness. Twenty of the 58 families were observed to have mutations in both alleles of Cx26. Thirty-three of 116 chromosomes contained a 30delG allele, for a frequency of .284. This mutation was observed in 2 of 192 control chromosomes, for an estimated gene frequency of .01+/-.007. The homozygous frequency of the 30delG allele is then estimated at .0001, or 1/10,000. Given that the frequency of all childhood hearing impairment is 1/1,000 and that half of that is genetic, the specific mutation 30delG is responsible for 10% of all childhood hearing loss and for 20% of all childhood hereditary hearing loss. Six novel mutations were also observed in the affected population. The deletions detected cause frameshifts that would severely disrupt the protein structure. Three novel missense mutations, Val84Met, Val95Met, and Ser113Pro, were observed. The missense mutation 101T-->C has been reported to be a dominant allele of DFNA3, a dominant nonsyndromic hearing loss. Data further supporting the finding that this mutation does not cause dominant hearing loss are presented. This allele was found in a recessive family segregating independently from the hearing-loss phenotype and in 3 of 192 control chromosomes. These results indicate that 101T-->C is not sufficient to cause hearing loss.

Alleles↗
PubMed1997

Basis of pulmonary toxicity associated with cationic lipid-mediated gene transfer to the mammalian lung.

Studies have indicated that although abundant levels of transgene expression could be achieved in the lungs of mice instilled with cationic lipid:pDNA complexes, the efficiency of gene transfer is low. As a consequence, a relatively large amount of the complex will need to be administered to the human lungs to achieve therapeutic efficacy for indications such as cystic fibrosis. Because all cationic lipids exhibit some level of cytotoxicity in vitro, we assessed the safety profile of one such cationic lipid, GL-67, following administration into the lungs of BALB/c mice. Dose-dependent pulmonary inflammation was observed that was characterized by infiltrates of neutrophils, and, to a lesser extent, macrophages and lymphocytes. The lesions in the lung were multifocal in nature and were manifested primarily at the junction of the terminal bronchioles and alveolar ducts. The degree of inflammation abated with time and there were no apparent permanent fibrotic lesions, even in animals that were treated at the highest doses. Analysis of the individual components of the complex revealed that the pulmonary inflammation was primarily cationic lipid-mediated with a minor contribution from the neutral co-lipid DOPE. Associated with the lesions in the lungs were elevated levels of the pro-inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and interferon-gamma (IFN-gamma) that peaked at days 1-2 post-instillation but resolved to normal limits by day 14. Total cell counts, primarily of neutrophils, were also significantly elevated in the bronchoalveolar lavage fluids of GL-67:pDNA-treated mice between days 1 and 3 but returned to normal limits by day 14. No specific immune responses were detected against the cationic lipid or plasmid DNA in mice that had been either instilled or immunized with the individual components or complex, nor was there any evidence of complement activation. These studies indicate that a significant improvement in the potency of cationic lipid:pDNA formulations is desirable to minimize the toxicity associated with cationic lipids.

Administration, Intranasal↗
PubMed1997

Optimization of formulations and conditions for the aerosol delivery of functional cationic lipid:DNA complexes.

We have examined several variables inherent in aerosolizing cationic lipid:DNA complexes using a jet nebulizer and thereby have optimized the delivery of functional complexes. Maximal aerosol transfer efficiency of cationic lipid:pDNA complexes was quantitated and shown to require the presence of at least 25 mM NaCL as an excipient. This is possibly related to effects on the measured zeta potentials of the complex, which indicate that the complexes are more highly charged in solutions of physiological ionic strength than in solutions of low ionic strength. Inclusion of saline also resulted in retention of the starting lipid to plasmid DNA (pDNA) ratio following nebulization. These data were used to design in vitro aerosolization experiments with tissue culture cells that resulted in the identification of a cationic lipid:pDNA ratio of 0.75:1 (mol:mol) as being optimal for aerosolization. This formulation largely protected pDNA from shear degradation during nebulization and produced a respirable aerosol droplet size (1-3 microns). It was tested further in a mouse model and shown to result in the dose-dependent transfection of mouse lungs, generating the equivalent of several picograms of reporter gene activity per mouse lung. The results of these experiments have provided a set of optimal conditions for nebulizing cationic lipid:pDNA complexes that can be used as a starting point for the further evaluation of aerosol delivery of these nonviral gene delivery vectors in vivo.

Administration, Intranasal↗
PubMed1997

A deletion polymorphism due to Alu-Alu recombination in intron 2 of the retinoblastoma gene: association with human gliomas.

The retinoblastoma gene (RB) encodes a tumor suppressor that is inactivated in a number of different types of cancer. We searched for gross alterations of this gene in tumors of the central nervous system by using Southern blot hybridization. A common alteration was found in several tumors and was mapped to the region around exon 2. Nucleotide sequencing showed that the alteration was caused by a 799-bp deletion in intron 2 of the RB gene and was probably due to homologous recombination between two Alu repeats. Deletions of this type have not been found previously in the RB gene. The deletion turned out to be a polymorphism with an allele frequency estimated at 2.2% in 185 patients without cancer. The deletion was found in five of 48 patients with brain tumors (allele frequency of 5.2%). This difference is not statistically significant (P = 0.149, Fisher's exact test). Confining the analysis only to glioma brain tumors revealed a statistically significant difference compared with the cancer-free patient controls (P = 0.027, Fisher's exact test). Further study is needed to determine if the deletion is a weak brain cancer-predisposing mutation or a harmless polymorphism. Finding this mutation in a tumor and the germline DNA of a retinoblastoma patient could lead to incorrect estimation of the heritability of a tumor.

Base Sequence↗
PubMed1997

Reciprocal translocation 4;11 with both adjacent-1 segregants viable within a family.

We describe a family carrying a balanced 4;11 translocation in which both adjacent-1 segregants are viable. The proband had an unbalanced karyotype: 46,XY,der(11)t(4;11)(q34.3;q23.1)mat. At 8.5 years of age he showed trigonocephaly, hypertelorism, epicanthal folds, down-slanting palpebral fissures, low-set ears, anteverted nares, down-turned carp-shaped mouth, and bilateral fifth finger clinodactyly. His maternal aunt was also dysmorphic with high-arched palate, short philtrum and mild developmental delay. Her karyotype was 46,XX,der(4)t(4;11)(q34.3;q23.1)pat. Other relatives who likely carried a chromosomally unbalanced segregant were identified from photographs and medical records. We compare the clinical findings in our family with descriptions of other similar karyotypic abnormalities from previous case reports.

Abnormalities, Multiple↗
PubMed1996

Detailed analysis of structures and formulations of cationic lipids for efficient gene transfer to the lung.

Cationic lipid-mediated gene transfer of cystic fibrosis transmembrane conductance regulator (CFTR) cDNA represents a promising approach for treatment of cystic fibrosis (CF). Here, we report on the structures of several novel cationic lipids that are effective for gene delivery to the lungs of mice. An amphiphile (#67) consisting of a cholesterol anchor linked to a spermine headgroup in a "T-shape" configuration was shown to be particularly efficacious. An optimized formulation of #67 and plasmid vector encoding chloramphenicol acetyl-transferase (CAT) was capable of generating up to 1 microgram of CAT enzyme/lung following intranasal instillation into BALB/c mice. This represents a 1,000-fold increase in expression above that obtained in animals instilled with naked pDNA alone and is greater than 100-fold more active than cationic lipids used previously for CFTR gene expression. When directly compared with adenovirus-based vectors containing similar transcription units, the number of molecules of gene product expressed using lipid-mediated transfer was equivalent to vector administration at multiplicities of infection ranging from 1 to 20. The level of transgene expression in the lungs of BALB/c mice peaked between days 1 and 4 post-instillation, followed by a rapid decline to approximately 20% of the maximal value by day 7. Undiminished levels of transgene expression in the lung could be obtained following repeated intranasal administration of #67:DOPE:pCF1-CAT in nude mice. Transfection of cells with formulations of #67:DOPE:pCF1-CFTR generated cAMP-stimulated CFTR chloride channel and fluid transport activities, two well-characterized defects associated with CF cells. Taken together, the data demonstrate that cationic lipid-mediated gene delivery and expression of CFTR in CF lungs is a viable and promising approach for treatment of the disease.

Adenoviruses, Human↗
PubMed1995

The characteristics and causes of sheep losses in the Victorian Mallee.

The extent and causes of sheep losses in the semi-arid Mallee region of north-western Victoria were assessed by interviewing the owners of 79 randomly selected farms running 241 flocks in 1987/88 and 245 flocks in 1988/89. Mean annual losses were higher in ram flocks (21%) than in ewe flocks (7%), in flocks of non-Merino sheep (rams 24%, ewes 11%, weaners 5%) than in Merino (rams 11%, ewes 6%, wethers 4%, weaners 4%) and in ewe flocks 3 or more years old (10%) than in young ewe flocks (3.5%). In flocks where losses exceeded 5%, the causes most often reported by farmers were blowfly strike (especially in Merino sheep and weaners), ewe losses in autumn close to lambing, and heliotrope (Heliotropium europaeum) poisoning. Heliotrope poisoning was considered by the authors to be the main reason for the higher losses in old ewes than in young ewes and in non-Merino sheep than in Merino sheep. Losses of ewes associated with pregnancy and lambing were considered by the authors to be often predisposed by liver damage caused by heliotrope poisoning, and high losses in non-Merino ram flocks were attributed to both heliotrope poisoning and their ability to escape through boundary fences. Reasons for continuing high losses due to enterotoxaemia are discussed. Losses due to gastro-intestinal parasites, footrot and foot abscess were low.

Abscess↗
PubMed1995

Posterior keratoconus. Case report with scanning electron microscopy.

We report the clinical and scanning electron microscopic findings in an older adult patient with unilateral posterior keratoconus. Clinically, the cornea showed typical posterior concavity with focal underlying anterior ectasia and an epithelial iron ring. Scanning electron microscopy of the cornea removed at penetrating keratoplasty revealed a pericentral posterior concavity. The endothelial layer was intact, with moderate endothelial cell pleomorphism. There were no excrescences of Descemet's membrane or endothelial tags. The lack of discrete irregularities on the posterior corneal surface in this case is different from previous descriptions of the pathological appearance of posterior keratoconus, which have been used to support a relationship of this condition to central corneal mesenchymal dysgenesis. This is the first report of scanning electron microscopic findings in posterior keratoconus.

Aged↗
PubMed1995

Evidence for a distinct region causing a cat-like cry in patients with 5p deletions.

The cri-du-chat syndrome is a contiguous gene syndrome that results from a deletion of the short arm of chromosome 5 (5p). Patients present with a cat-like cry at birth, which is usually considered diagnostic of this syndrome. Additional features of the syndrome include failure to thrive, microcephaly, hypertelorism, epicanthal folds, hypotonia, and severe mental retardation. We report on four families in which patients with 5p deletions have only the characteristic cat-like cry, with normal to mildly delayed development. The precise locations of the deletions in each family were determined by FISH using lambda phage and cosmid clones. All of the deletion breakpoints map distal to a chromosomal region that is implicated with the facial features and severe mental and developmental delay in the cri-du-chat syndrome. DNA clones mapping in the chromosomal region associated with the cat-like cry feature will be useful diagnostic tools. They will allow for the distinction between 5p deletions that will result in the severe delay observed in most cri-du-chat syndrome patients and those deletions that result in the isolated cat-like cry feature, which is associated with a better prognosis.

Adult↗
PubMed1994

Retinal signal transmission in Duchenne muscular dystrophy: evidence for dysfunction in the photoreceptor/depolarizing bipolar cell pathway.

There have been reports of abnormal retinal neurotransmission determined by electroretinography in boys with Duchenne and Becker muscular dystrophy. Dystrophin may play a role in transmitting signals between photoreceptors and the excitatory synapse of the ON-bipolar cell. These electroretinographic changes appeared to be limited to the rod ON-pathway but we felt there was also similar abnormality in the cone ON-pathway. We used long-duration stimuli to separate ON-(depolarizing bipolar cell) and OFF (hyperpolarizing bipolar cell) contributions to the cone-dominated ERG to better understand how the retina functions in boys with Duchenne muscular dystrophy. We recorded the electroretinograms of 11 boys with Duchenne muscular dystrophy and found abnormal signal transmission at the level of the photoreceptor and ON-bipolar cell in both the rod and cone generated responses. The OFF-bipolar cell that responds to the offset of the stimulus continues to function normally. The results support our hypothesis that retinal dystrophin plays a role in receptor function or controlling ion channels at the level of the photoreceptor and depolarizing bipolar cell.

Adolescent↗
PubMed1994

Disposition of lupanine and 13-hydroxylupanine in man.

1. The in vivo disposition of lupanine and 13-hydroxylupanine was studied in subjects identified as poor metabolizers (PM, n = 4) and extensive metabolizers (EM, n = 7) phenotypes for cytochrome P4502D6 (CYP2D6). 2. After oral administration (40.26 mumol), the half-life (t1/2) of lupanine determined from urinary excretion rate studies in EM subjects was 6.2 +/- 0.5 h (mean +/- SEM) with 95.5 +/- 6.0% of the dose recovered unchanged within 72 h. Similarly, in PM subjects t1/2 = 6.5 +/- 0.9 h and recovery 89.9 +/- 4.5%. 3. For orally administered 13-hydroxylupanine (37.83 mumol) the t1/2 in EM subjects was 6.8 +/- 1.0 h with a recovery of 100.5 +/- 5.3%, and in PM subjects t1/2 = 5.9 +/- 1.6 h with a recovery of 102.5 +/- 4.8%. 4. The t1/2s of both lupanine and 13-hydroxylupanine respectively did not differ significantly between EM and PM phenotypes. In addition, total recovery of dose for both alkaloids was similar between phenotypes. 5. In most subjects, > 76% of lupanine and > 85% of 13-hydroxylupanine was recovered as the unchanged compound. Significant apparent partial dehydroxylation of 13-hydroxy-lupanine was observed in one EM (14% of dose) and one PM (34% of dose) subject. 6. Overall, the finding of a high urinary recovery of unchanged lupanine or 13-hydroxylupanine together with similar t1/2s for both alkaloids in EM and PM CYP2D6 phenotypes suggests that clinical toxicity is unlikely to result from the use of lupin seed in footstuffs.

Alkaloids↗