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Biomedical subjects

D J Smith

Publications and source records attributed to D J Smith.

At least 145 records · Page 8Linked to original sources
PubMed1997

Incorporation of nitric oxide-releasing crosslinked polyethyleneimine microspheres into vascular grafts.

Over the years, many attempts have been made to increase the patency of small- to medium-sized prosthetic vascular grafts. However, none of them has greatly affected long-term rates. Recently, nitric oxide (NO) has been shown to inhibit thrombus formation in such grafts, suggesting that local delivery of NO may help to increase graft patency. This study describes the site-specific delivery of NO by entrapping NO-releasing microspheres in the pores of a vascular graft. NO-releasing polyethyleneimine microspheres (PEIX) were developed using a novel water-in-oil emulsion technique involving chemical crosslinking with a bis-epoxide. The PEIX microspheres were then derivatized with NO forming the [N(O)NO]- moiety of the diazeniumdiolates formerly known as NONOates. These polymeric NO-releasing particles were found to spontaneously release 194 nmol NO/mg with a half-life of over 66 h under physiologic conditions. Fluorescein isothiocyanate-labeled microspheres were then embedded into the pores of a 60-micron nonreinforced Gore-tex vascular graft using a simple evacuation technique and evaluated for microsphere placement and NO release. Scanning electron microscopic analysis showed the microspheres entrapped in the pores of the vascular graft releasing 10 nmol NO/mg with a half-life of 51 h. The microspheres remained entrapped in the graft even after immersion and NO release, as confirmed by fluorescence of the medium. These results suggest that NO-releasing particles can be incorporated into the pores of a vascular graft to deliver therapeutic amounts of NO for the prevention of thrombosis in small-diameter prosthetic grafts.

Blood Vessel Prosthesis↗
PubMed1997

The vibrator mutation causes neurodegeneration via reduced expression of PITP alpha: positional complementation cloning and extragenic suppression.

The mouse vibrator mutation causes an early-onset progressive action tremor, degeneration of brain stem and spinal cord neurons, and juvenile death. We cloned the vibrator mutation using an in vivo positional complementation approach and complete resequencing of the resulting 76 kb critical region from vibrator and its parental chromosome. The mutation is an intracisternal A particle retroposon insertion in intron 4 of the phosphatidylinositol transfer protein alpha gene, causing a 5-fold reduction in RNA and protein levels. Expression of neurofilament light chain is also reduced in vibrator, suggesting one signaling pathway that may underlie vibrator pathology. The vibrator phenotype is suppressed in one intercross. We performed a complete genome scan and mapped a major suppressor locus (Mvb-1) to proximal chromosome 19.

Alleles↗
PubMed1997

Functional screening of 2 Mb of human chromosome 21q22.2 in transgenic mice implicates minibrain in learning defects associated with Down syndrome.

Using Down syndrome as a model for complex trait analysis, we sought to identify loci from chromosome 21q22.2 which, when present in an extra dose, contribute to learning abnormalities. We generated low-copy-number transgenic mice, containing four different yeast artificial chromosomes (YACs) that together cover approximately 2 megabases (Mb) of contiguous DNA from 21q22.2. We subjected independent lines derived from each of these YAC transgenes to a series of behavioural and learning assays. Two of the four YACs caused defects in learning and memory in the transgenic animals, while the other two YACs had no effect. The most severe defects were caused by a 570-kb YAC; the interval responsible for these defects was narrowed to a 180-kb critical region as a consequence of YAC fragmentation. This region contains the human homologue of a Drosophila gene, minibrain, and strongly implicates it in learning defects associated with Down syndrome.

Animals↗
PubMed1997

Functional screening and complex traits: human 21q22.2 sequences affecting learning in mice.

Libraries of the mammalian genome have generally been propagated in single cells and have been used for gene discovery through in vitro analyses. We have expanded upon this concept by the creation of panels of YAC transgenic mice propagating targeted megabase regions of the genome. Such a panel of mice can be called an 'in vivo library' and genes can be identified based on functional screens of members of the library. To test this approach, we created a 2 Mb in vivo library of human chromosome 21q22.2. Analysis of the library has revealed that one 570 kb YAC, in two separate founder lines, was associated with distinct learning deficits compared with the other 21q22 YAC transgenics and non-transgenic control animals. We have localized the gene on the YAC that causes the deficits by taking advantage of fragmentation of the YAC during the process of microinjection. The responsible gene is the human minibrain gene, and the homolog of the gene in Drosophila is also associated with learning defects. These results suggest that altered dosage of minibrain is associated with abnormal neural development in flies and mice and, in humans, may also be involved in the molecular pathology of Down syndrome.

Animals↗
PubMed1997

CD29 expression on CD4+ gingival lymphocytes supports migration of activated memory T lymphocytes to diseased periodontal tissue.

The cell surface phenotypes of CD+ cells extracted from inflammatory periodontal disease tissues were analyzed using two- and three-color immunofluorescence and flow cytometry. Cells extracted from both adult periodontal and localized juvenile periodontitis lesions showed a depressed CD4/CD8 ratio (1.0 +/- 0.1 adult periodontitis and 1.1 +/- 0.1 localized juvenile periodontitis) compared with cells recovered from normal/marginal gingivitis tissue (1.8 +/- 0.2) or with normal peripheral blood cells (2.1 +/- 0.1) or periodontal disease blood cells (2.1 +/- 0.1 and 1.7 +/- 0.1 for adult periodontitis and juvenile periodontitis, respectively). The monoclonal antibodies anti-2H4 and anti-4B4 were used to identify the CD45RA and CD29 antigens respectively on CD4+ T cells from the periodontal disease lesions. In peripheral blood. CD29+ cells accounted for 66-77% of the CD4+ population, and CD45RA+ cells accounted for 22-27% of the CD4+ subset. No differences in expression were found between peripheral blood lymphocytes from normal subjects and from periodontal disease patients. Two-color analyses of lymphocytes from periodontal diseased tissues showed that 87-89% of the CD4+ population were CD29+ and that 70-79% of the CD4+ cells were CD45RA+. Normal tissues contained significantly fewer CD4+CD29+ cells (56 +/- 4%) and CD4+CD45RA+ cells (40 +/- 4%) on average, and few, if any double-labelled cells could be accounted for. These data implied that a significant percentage of the CD4+ cells from the diseased tissues were both CD29+ and CD45RA+ and that these populations are found in quite different proportions in diseased periodontal tissue than in peripheral blood or nondiseased tissue. In further analyses using three-color cytometry the mean percentage of CD4+ CD29+ CD45RA+ lymphocytes extracted from periodontal disease lesions was 43 +/- 9% of the CD4+ population. These results suggest that CD4+ T lymphocytes in periodontal disease not only demonstrate varying levels of maturity but also that the accumulation of CD4+ T cells within the periodontal tissues may be a result of increased adhesion and transendothelial migration.

Adult↗
PubMed1997

Immunogenicity and protective immunity induced by synthetic peptides associated with a catalytic subdomain of mutans group streptococcal glucosyltransferase.

We examined the immunogenicity and induction of protective immunity of two 19-mer sequences (GGY and AND) which overlapped a highly conserved region which has recently been implicated in the enzymatic activity of glucosyltransferases (GTFs) of the mutans group streptococci. These peptides were synthesized as eight-branched constructs on a lysine core. Serum immunoglobulin G (IgG) antibody, induced by subcutaneous (s.c. [salivary gland vicinity]) injection with these peptide constructs, reacted with the inciting antigen, with mutans streptococcal GTFs, and with a 21-mer peptide (CAT) containing an aspartate previously shown to covalently bind sucrose. Several of these antisera also inhibited the ability of Streptococcus sobrinus GTF to synthesize insoluble glucan. Significant levels of salivary IgA antibody were also induced by GGY and AND peptide constructs after s.c. injection. The effect of immunization with the GGY and AND peptide constructs on the cariogenicity of Streptococcus mutans was studied in three experiments by immunization of weanling Sprague-Dawley rats, twice at 7- to 14-day intervals with peptides, S. sobrinus GTF, or phosphate-buffered saline. All rats were then orally infected with S. mutans SJ. After 63-day infection periods, the GGY and AND-injected groups had significant dental caries reductions compared with sham-injected groups in most experiments. These studies support the existence of an additional catalytic subdomain within the sequence defined by the GGY and AND peptides. Furthermore, the epitopes defined in these sequences have significant immunogenicity, can induce immune responses which interfere with GTF-mediated glucan synthesis in vitro, and can protect rats from experimental dental caries.

Amino Acid Sequence↗
PubMed1997

Maxillary sinusitis caused by Pleurophomopsis lignicola.

An immunocompetent 59-year-old man developed sinusitis over a 6- to 8-month period after cutting down a rotted maple tree (Acer sp.). A polypoid obstruction with a bloody drainage was evident in his right nasal cavity. A computed tomographic scan showed an opacification of the maxillary sinus. Surgery was performed to remove a fungus ball that had extended into the patient's medial sinus cavity. Sections of the sinonasal mucosa revealed marked acute and chronic sinusitis with inflammation, congestion, and hemorrhage. Sections from the pasty brown to black debrided material revealed a fungus ball consisting of an extensive network of brown-pigmented, septate, profusely branched hyphae. When grown on oat agar, the phaeoid fungus produced pycnidia and was identified as Pleurophomopsis lignicola. The genus Pleurophomopsis includes seven species, which are all known from plant material. This report documents for the first time a coelomycetous fungus, P. lignicola, causing sinusitis in an immunocompetent patient.

Fungi↗
PubMed1997

Intratracheal instillation of a novel NO/nucleophile adduct selectively reduces pulmonary hypertension.

We examined the pulmonary and systemic hemodynamic effects of administering soluble nitric oxide (NO) donor compounds (NO/nucleophile adducts, i.e., NONOates) directly into the trachea of animals with experimentally induced pulmonary hypertension. Steady-state pulmonary hypertension was created by using the thromboxane agonist U-46619. Yorkshire pigs were randomly assigned to one of four groups: group 1, intratracheal saline (control; n = 8); group 2, intratracheal sodium nitroprusside (n = 6); group 3, intratracheal ethylputreanine NONOate (n = 6); and group 4, intratracheal 2-(dimethylamino)-ethylputreanine NONOate (DMAEP/NO; n = 6). Pulmonary and systemic hemodynamics were monitored after drug instillation. Group 4 had significant reductions in pulmonary vascular resistance index (PVRI) at all time points compared with steady state and compared with group 1 (P < 0.05), whereas systemic vascular resistance index did not change. The mean change in mean pulmonary arterial pressure in group 4 was -33.1 +/- 1.2% compared with +6.4 +/- 1.3% in group 1 (P < 0.001), and the mean change in mean arterial pressure was -9.3 +/- 0.7% compared with a control value of -0.9 +/- 0.5% (P < 0.05). Groups 2 and 3 had significant decreases in both PVRI and systemic vascular resistance index compared with steady state and with group 1. In conclusion, intratracheal instillation of a polar-charged tertiary amine NONOate DMAEP/NO results in the selective reduction of PVRI. Intermittent intratracheal instillation of selective NONOates may be an alternative to continuously inhaled NO in the treatment of pulmonary hypertension.

Amino Acids, Diamino↗
PubMed1997

Mini-exon epitope tagging for analysis of the protein coding potential of genomic sequence.

A novel approach to gene discovery and analysis is described. A small exon flanked by consensus 3' and 5' splice sites was synthesized. The exon contains open reading frames encoding 43 amino acid peptides. There are no stop codons in any of the three reading frames, and each reading frame contains an epitope recognized by the same monoclonal antibody. The exon can be inserted into the introns of genes, and the resulting small peptide will be incorporated into the protein encoded by the host gene, regardless of the class of intron. The protein can then be recognized by the antibody, permitting functional studies.

Amino Acid Sequence↗
PubMed1997

Distribution, elimination, and residues of [14C]clenbuterol HCl in Holstein calves.

Clenbuterol HCl is a beta-adrenergic agonist that has been used illegally in Europe and the United States by some livestock producers to increase carcass leanness. The objectives of this study were to determine the metabolic disposition, distribution of radioactivity, and the concentrations of parent clenbuterol in tissues after a single oral dose of [14C] clenbuterol HCl in calves. [14C]Clenbuterol HCl (1.59 microCi/mg, 3 mg/kg BW) was administered to a 74- and a 96-kg Holstein bull calf as a single oral dose. Blood samples were taken at 1, 3, 6, 12, 24, 36, and 48 h after dosing; urine and feces were collected separately and placed into respective pools from 0 to 6, 6 to 12, 12 to 24, 24 to 36, and 36 to 48 h after dosing. At 48 h after dosing, calves were anesthetized and exsanguinated, and visceral organs, bile, eyes, brain, skeletal muscle, skin, adipose tissue, and the remainder of the carcass were collected. Tissues were processed by coarse grinding through a Hobart grinder, followed by homogenization on dry ice. Eyes were dissected and the various tissues and excreta were assayed for total radiocarbon content by combustion analysis and(or) liquid scintillation counting. Parent clenbuterol was measured in selected tissues by HPLC after solvent extraction. Urinary, fecal, and carcass radioactivity averaged 41.5 +/- 8.1, 2.4 +/- .4, and 52.3 +/- 8.7% of the dose, respectively (mean +/- SD.). Average total recovery of radiocarbon was 96.2 +/- .4%. Radioactive residues present in carcasses averaged (ppm; mean +/- SD.): blood, .6 +/- .2; heart, 1.4 +/- .0; lungs, 8.4 +/- 1.7; spleen, 2.6 +/- .3; liver, 5.0 +/- .4; kidney, 5.9 +/- .0; brain, 1.9 +/- .4; adipose tissue, 1.1 +/- .2; rumen, reticulum, omasum, abomasum, 2.3 +/- .4; small intestine, 3.2 +/- .3; large intestine, 4.0 +/- .4; skeletal muscle, 1.0 +/- .2; bile, 12.5 +/- 4.0; white skin, .7 +/- .1; black skin, 4.0 +/- .7; remainder of the carcass, 1.0 +/- .2. Ocular residues were as follows: aqueous humor, 6.3 +/- 1.2; cornea, 13.5 +/- 8.6; iris, 255.8 +/- 167.0; lens, 2.3 +/- 1.5; vitreous humor, 2.2 +/- .4; retina/choroid, 84.5 +/- 34.1; sclera, 11.1 +/- 2.1. Mean concentrations of parent clenbuterol in tissues were (ppm; mean +/- SD): lung, 6.8 +/- .9; liver, 2.2 +/- .5; kidney, 3.7 +/- .5; and heart, .9 +/- .1. Parent clenbuterol represented from 43.9% of the total residue in liver to 81.2% of the total residue in lung.

Adipose Tissue↗
PubMed1997

Characteristics of the new long-stay population in an inner Melbourne acute psychiatric hospital.

OBJECTIVE: There has been an accumulation of so-called 'new long-stay' (NLS) patients in psychiatric hospitals. To date, no Australian studies have characterised this patient group. We aimed to study the demography, and clinical and social functioning of the NLS population at Royal Park Hospital (RPH) together with patients' attitudes to treatment and views on future placement. METHOD: All 30 NLS patients at RPH were identified. Twenty-seven consenting patients were assessed using the following standardised instruments: Manchester Scale for psychopathology, Life Skills Profile (LSP), Physical Health Index and Patient Attitude Questionnaire. Information on past psychiatric history, past treatment and current treatment was collected. Insight and compliance were assessed. RESULTS: The majority of patients were single men with a diagnosis of schizophrenia. Forty-one percent were detained in hospital involuntarily and 56% were considered dangerous to themselves or others. The patients were characterised by high levels of positive and negative symptoms. They were most impaired with respect to 'social contact' relative to the other subscales of the LSP. While 10 (48%) patients expressed a desire to leave hospital, only one patient considered that anyone would cohabit with them. Over two-thirds considered they had been unwell and that medication had helped. Staff rated one-third as having major problems with compliance. About two-thirds of patients had disability secondary to comorbid physical illnesses. CONCLUSION: Like other NLS patients studied in the United Kingdom and Ireland, this group had significant handicaps secondary to psychiatric illness, concomitant physical illness and disability and behaviour unacceptable in community settings. They were also characterised by significant social isolation. These factors may be important determinants of rehabilitation failure and need to be addressed in the process of de-institutionalisation as well as in longitudinal studies examining these and other factors predicting NLS status.

Activities of Daily Living↗
PubMed1997

Dose-dependent pain-facilitatory and -inhibitory actions of neurotensin are revealed by SR 48692, a nonpeptide neurotensin antagonist: influence on the antinociceptive effect of morphine.

Neurotensin has bipolar (facilitatory and inhibitory) effects on pain modulation that may physiologically exist in homeostasis. Facilitation predominates at low (picomolar) doses of neurotensin injected into the rostroventral medial medulla (RVM), whereas higher doses (nanomolar) produce antinociception. SR 48692, a neurotensin receptor antagonist, discriminates between receptors mediating these responses. Consistent with its promotion of pain facilitation, the minimal antinociceptive responses to a 30-pmol dose of neurotensin microinjected into the RVM were markedly enhanced by prior injection of SR 48692 into the site (detected using the tail-flick test in awake rats). SR 48692 had a triphasic effect on the antinociception from a 10-nmol dose of neurotensin. Antinociception was attenuated by femtomolar doses, attenuation was reversed by low picomolar doses (corresponded to those blocking the pain-facilitatory effect of neurotensin) and the response was again blocked, but incompletely, by higher doses. The existence of multiple neurotensin receptor subtypes may explain these data. Physiologically, pain facilitation appears to be a prominent role for neurotensin because the microinjection of SR 48692 alone causes some antinociception. Furthermore, pain-facilitatory (i.e., antianalgesic) neurotensin mechanisms dominate in the pharmacology of opioids; the response to morphine administered either into the PAG or systemically was potentiated only by the RVM or systemic injection of SR 48692. On the other hand, reversal of the enhancement of antinociception occurred under certain circumstances with SR 48692, particularly after its systemic administration.

Analgesics, Opioid↗
PubMed1996

Nitric oxide-releasing polymers containing the [N(O)NO]- group.

Ions of structure X[N(O)NO]- display broad-spectrum pharmacological activity that correlates with the rate and extent of their spontaneous, first-order decomposition to nitric oxide when dissolved. We report incorporation of this functional group into polymeric matrices that can be used for altering the time course of nitric oxide release and/or targeting it to tissues with which the polymers are in physical contact. Structural types prepared include those in which the [N(O)NO]- group is attached to heteroatoms in low molecular weight species that are noncovalently distributed throughout the polymeric matrix, in groupings pendant to the polymer backbone, and in the polymer backbone itself. They range in physical form from films that can be coated onto other surfaces to microspheres, gels, powders, and moldable resins. Chemiluminescence measurements confirm that polymers to which the [N(O)NO]- group is attached can serve as localized sources of nitric oxide, with one prototype providing sustained NO release for 5 weeks in pH 7.4 buffer at 37 degrees C. The latter composition, a cross-linked poly-(ethylenimine) that had been exposed to NO, inhibited the in vitro proliferation of rat aorta smooth muscle cells when added as a powder to the culture medium and showed potent antiplatelet activity when coated on a normally thrombogenic vascular graft situated in an arteriovenous shunt in a baboon's circulatory system. The results suggest that polymers containing the [N(O)NO]- functional group may hold considerable promise for a variety of biomedical applications in which local delivery of NO is desired.

Animals↗