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Biomedical subjects

K A Sullivan

Publications and source records attributed to K A Sullivan.

At least 19 recordsLinked to original sources
PubMed2004

Differences between collagen morphologies, properties and distribution in diabetic and normal biobreeding and Sprague-Dawley rat sciatic nerves.

Both structural and functional differences between normal and diabetic nerve have been observed, in human patients and animal models. We hypothesize that these structural differences are quantifiable, morphologically and mechanically, with the ultimate aim of understanding the contribution of these differences to permanent nerve damage. The outer collagenous epineurial and perineurial tissues of mammalian peripheral nerves mechanically and chemically shield the conducting axons. We have quantified differences in these collagens, using whole-nerve uniaxial testing, and immunohistochemistry of collagens type I, III, and IV in diabetic and normal nerves. We present results of two studies, on normal and diabetic BioBreeding (BB), and normal, diabetic and weight-controlled Sprague-Dawley (SD) rats, respectively. Overall, we measured slightly higher uniaxial moduli (e.g. 5.9 MPa vs. 3.5 MPa, BB; 10.7 MPa vs. 10.0 MPa, SD at 40% strain) in whole nerves as well as higher peak stresses (e.g. 0.99 MPa vs. 0.74 MPa, BB; 2.16 MPa vs. 1.94 MPa, SD at 40% strain) in the diabetics of both animal models. We measured increased concentrations of types III and IV collagens in the diabetics of both models and mixed upregulation results were observed in type I protein levels. We detected small differences in mechanical properties at the tissue scale, though we found significant structural and morphometric differences at the fibril scale. These findings suggest that whole-tissue mechanical testing is not a sufficient assay for collagen glycation, and that fibrillar and molecular scale assays are needed to detect the earliest stages of diabetic protein glycation.

Animals↗
PubMed2004

A preliminary investigation of the effectiveness of a sleep apnea education program.

OBJECTIVES: This study aimed to evaluate a standardised sleep apnea patient education program and develop a study design that may be used to evaluate other such education programs. METHOD: Thirty-four adults diagnosed with obstructive sleep apnea hypopnea syndrome (OSAHS) underwent a standard sleep apnea education program and completed measures of knowledge of and beliefs about sleep apnea before, after, and 3 months following education. Two outcome measures were used: the Apnea Knowledge Test (AKT) and the Apnea Beliefs Scale (ABS). RESULTS: AKT results showed significant knowledge gains posteducation, which were maintained at follow-up. Patients also reported more positive beliefs about their ability to change their behaviour and comply with continuous positive airway pressure (CPAP) treatment recommendations after education. DISCUSSION: Findings from this preliminary investigation suggest that the education program used in this study may improve patients' knowledge of CPAP and promote functional beliefs about OSAHS treatment. This program clearly warrants further research, and ultimately such programs may prove important in improving CPAP compliance.

Attitude to Health↗
PubMed2003

Digital electron microscopic examination of human sural nerve biopsies.

Diabetic peripheral polyneuropathy is characterized by axonal degeneration and regeneration as well as by Schwann cell and microvascular changes. These changes have been described at both the light (LM) and the electron microscopic (EM) levels; however, EM has not been applied to large clinical trials. Our goal was to adapt the rigorous techniques used for quantifying human biopsies with LM image analysis to accommodate ultrastructural analyses. We applied digital image capture and analysis to the ultrastructural examination of axons in sural nerve biopsies from diabetic patients enrolled in a multicenter clinical trial. The selection of sural nerve biopsies was based on the quality of specimen fixation, absence of physical distortion, and nerve fascicle size (> or =100,000; < or =425,000 microm2). Thin sections were collected on formvar-coated slot grids, stabilized with carbon and scanned on a Phillips CM100 transmission electron microscope. Digital images were captured with a Kodak Megaplus 1.6 camera. A montage was constructed using software derived from aerial mapping applications, and this virtual image was viewed by EM readers. Computer-assisted analyses included identification and labeling of individual axons and axons within regenerating clusters. The average density of regenerating myelinated axon clusters per mm2 was 65.8 +/- 5.1, range of 0-412 (n = 193). These techniques increase the number of samples that may be analyzed by EM and extend the use of this technique to clinical trials using tissue biopsies as a primary endpoint.

Axons↗
PubMed2001

Synthesis of novel retinoid X receptor-selective retinoids.

Retinoids 1-5 have been identified as potent RXR agonists for evaluation in the treatment of non-insulin-dependent (type II) diabetes mellitus (NIDDM). Highly convergent syntheses of 1-5 have been developed. The core tetrahydronaphthalene 7, employed in the synthesis of 1 and 2, was prepared in 98% yield using an AlCl(3)-catalyzed (0.03 equiv) Friedel-Crafts alkylation of toluene with 2,5-dichloro-2,5-dimethylhexane 6. A nitromethane-mediated Fridel-Crafts acylation of 7 with chloromethylnicotinate 9 was developed to prepare ketone 10 in 68% yield. Chelate-controlled addition of MeMgCl to 10 followed by dehydration afforded olefin 11 in 65% yield. Cyclopropanation of 11 with trimethylsulfoxonium ylide, followed by saponification, completed a five-step synthesis of 1 in 33% yield. FeCl(3)-catalyzed (0.05 equiv) Friedel-Crafts acylation of 7 with chloromethylterephthalate 14 afforded ketone 15 in 81% yield. Saponification of 15 and reaction with 50% aqueous NH(2)OH in AcOH afforded a 9:1 mixture of cis and trans oximes, from which the desired cis-oxime 2 was isolated in 43% yield. The core bromo-dihydronaphthalene 29 required for the synthesis of 3-5 was prepared by a Shapiro reaction. Transmetalation of 29 and reaction with Weinreb amides 30b or 36 afforded ketones 32 and 37, which were converted into 3-5 using chemistry comparable to the tetrahydronaphthylene series. Suzuki coupling of boronic acids 41 and 42 with vinyl triflate 43 provided an alternative approach to the synthesis of this class of compounds.

Receptors, Retinoic Acid↗
PubMed2001

Depletion of taurine in experimental diabetic neuropathy: implications for nerve metabolic, vascular, and functional deficits.

In diabetes, increased oxidative stress, disruption of signal transduction pathways, and endothelial dysfunction have been critically implicated in the pathogenesis of experimental diabetic neuropathy (EDN). The development of nerve conduction slowing in diabetes is accompanied by depletion of the beta-amino acid taurine. Since taurine functions as an antioxidant, calcium modulator, and vasodilator, taurine depletion may provide a pathogenetic link between nerve metabolic, vascular, and functional deficits complicating diabetes. The mechanism(s) of nerve taurine depletion, the localization of critical taurine deficits, and its pathophysiological significance in EDN are however unknown. This study explored the pathophysiological effects of selective nerve taurine replacement in streptozotocin-diabetic (STZ-D) rats. A polyclonal human taurine transporter (TT) antibody was also generated in order to determine potential loci of critical taurine depletion. Two weeks of STZ-D reduced sciatic motor nerve conduction velocity (NCV) by 23% (P < 0.01), decreased composite nerve blood flow by 38% (P < 0.01), and reduced nerve taurine content by 29% (P < 0.05). In STZ-D rats, a 1% taurine diet corrected nerve taurine depletion, prevented motor NCV slowing, and partially attenuated composite nerve blood flow deficits. After 6 weeks of STZ-D, a 1% taurine diet ameliorated motor NCV slowing and endoneurial nutritive blood flow deficits, prevented digital sensory NCV slowing, and reduced ouabain-sensitive nerve (Na,K)-ATPase activity. Immunohistochemical studies localized taurine and the TT to the vascular endothelium and Schwann cells of the sciatic nerve. In conclusion, taurine depletion in the vascular endothelium and Schwann cells of the sciatic nerve may contribute to the neurovascular and metabolic deficits in EDN.

Animals↗
PubMed2001

Treatment of tardive dyskinesia with donepezil: a pilot study.

BACKGROUND: Tardive dyskinesia (TD) remains a significant clinical problem for which there is no uniformly effective treatment. Earlier trials with acetylcholine precursors may have been disappointing because of underlying damage to striatal cholinergic neurons in patients with TD. In contrast, new cholinesterase inhibitors, developed for the treatment of dementia, may improve TD by directly increasing cholinergic synaptic transmission. METHOD: We conducted an 8-week open-label trial of donepezil in the treatment of TD. Ten patients with schizophrenia or schizoaffective disorder who received stable doses of antipsychotics and met DSM-IV criteria for TD were treated with donepezil, 5 to 10 mg/day, for 6 weeks after a 2-week baseline period. Changes in total Abnormal Involuntary Movement Scale (AIMS) scores measured every 2 weeks were assessed for significance. Patients were also assessed using the Brief Psychiatric Rating Scale, the Mini-Mental State Examination, the Barnes Akathisia Scale, and the Simpson-Angus Scale. RESULTS: Total AIMS scores decreased significantly (p = .0009), with no changes in other measures. Nine patients showed a positive response. Improvement was greatest in orofacial and upper extremity movements. No significant interactions were noted between the total AIMS scores and age (p > .29), duration of TD (p > .38), or duration of antipsychotic treatment (p > .14). CONCLUSION: Donepezil appeared to be effective in suppressing TD in this pilot study. However, placebo-controlled, double-blind studies are necessary before donepezil can be recommended as a treatment for TD.

Adult↗
PubMed2000

The CXCR4 agonist ligand stromal derived factor-1 maintains high affinity for receptors in both Galpha(i)-coupled and uncoupled states.

The alpha chemokine receptor CXCR4 and its only characterized chemokine ligand, stromal cell-derived factor-1 (SDF-1), are postulated to be important in the development of the B-cell arm of the immune system. In addition, CXCR4 is a critical coreceptor in support of viral entry by T-cell line tropic strains (X4) of the Human Immunodeficiency Virus Type 1 (HIV-1), viral variants which predominate in some infected individuals in end stage disease. SDF-1 can block X4-tropic HIV-1 infection of CD4+ target cells in vitro, and allelic variants of the human gene encoding SDF-1 in vivo correlate with delayed disease progression. Therefore, CXCR4 may be an appropriate target for therapeutic intervention in acquired immunodeficiency syndrome (AIDS), and knowledge of the pharmacology of SDF-1 binding to its cognate receptor will be important in the interpretation of these experiments. We report here a Kd derived using a competition binding assay of 4.5 nM for CXCR4 endogenously expressed on peripheral blood monocytes and T-cells. This affinity is similar to that which SDF-1 exhibits when binding to endogenous CXCR4 on an established immortal Jurkat T-cell line as well as recombinant CXCR4 transfected into Chinese Hamster Ovary (CHO) cells. We also demonstrate that the determined affinity of SDF-1 for CXCR4 is reflective of its ability to induce a CXCR4-mediated signal transduction in these different cell types. Furthermore, using Bordetella pertussis toxin, we observe that high affinity binding of SDF-1 to CXCR4 is independent of the G-protein coupled state of the receptor, as uncoupling of G-protein did not lead to the appearance of measurable low affinity SDF-1 binding sites. Moreover, binding affinity and receptor number were unaffected by uncoupling for both recombinant and endogenously expressed CXCR4. Thus, SDF-1 is novel among agonist ligands of G protein-coupled receptors in that it appears to have equal affinity for both the G protein-coupled and uncoupled states of CXCR4.

Animals↗
PubMed2000

Treatment of antibody-mediated accelerated rejection using plasmapheresis.

Accelerated antibody-mediated rejection is believed to be due to an anamnestic response of an allograft recipient to donor antigens. Few reports have demonstrated successful reversal of this type of rejection, and no consensus exists for either diagnosis or treatment. Accelerated antibody-mediated rejection was suspected on the basis of clinical findings and confirmed by cytotoxic and flow crossmatches, and leukocyte antibody screens. Serial crossmatches and antibody screens were performed through post-transplant day 112. Plasmapheresis was performed on post-transplant days 1, 2, 4, 6, 12, 14, 20, and 28. The duration of treatment was determined by the cytotoxic crossmatch results. We present a case of successfully treated accelerated antibody-mediated rejection using plasmapheresis and aggressive immunosuppression. Serial crossmatch and leukocyte antibody screen results are presented that confirm the production of anti-donor antibody and demonstrate the effectiveness of the treatment protocol in eliminating detectable levels of the anti-donor antibody. At 6 months post-transplant, the patient has a serum creatinine of 1.1 and has not had any additional rejection episodes or infectious complications. The protocol suggested in this paper allows for rapid diagnosis, institution of treatment, and monitoring the efficacy of treatment, providing the basis for follow-up clinical trials.

Antibodies↗
PubMed1999

Neurons undergo apoptosis in animal and cell culture models of diabetes.

Recent clinical trials indicate that the severity of diabetic neuropathy is correlated with the level of patient glycemic control. In the current study, hyperglycemia induces apoptotic changes in dorsal root ganglion neurons and Schwann cells in vivo both in streptozotocin-treated diabetic rats and in rats made acutely hyperglycemic with infused glucose. Typical apoptotic nuclear and cytoplasmic changes are observed. In addition mitochondrial changes recently reported to occur as part of the apoptotic cascade, such as ballooning of mitochondria and disruption of the internal cristae, are seen in diabetic dorsal root ganglion neurons and Schwann cells. Similar changes have been reported in neurons in the presence of oxidative stress. In order to study the neurotoxic effects of high glucose we developed an in vitro model using rat dorsal root ganglion neurons. In dorsal root ganglion cultured in defined medium, addition of moderate glucose levels results in neurite degeneration and apoptosis. These changes are coupled with activation of caspase-3, dependent on the concentration of glucose. The apoptotic changes observed in vitro are similar to those observed in vivo. In contrast, addition of IGF-I, even at physiological concentrations, prevents activation of caspase-3 and neuronal apoptosis in vitro. We suggest that oxidative stress may promote the mitochondrial changes in diabetic animals and lead to activation of programmed cell death caspase pathways. These results imply a new pathogenetic mechanism for diabetic sensory neuropathy.

Animals↗
PubMed1999

Generation of mice expressing the human glucagon receptor with a direct replacement vector.

The process of evaluating the in vivo efficacy of non-peptidyl receptor antagonists in animal models is frequently complicated by failure of compounds displaying high affinity against the human receptors to show measurable affinity at the corresponding rodent receptors. In order to generate a suitable animal model in which to evaluate the in vivo activity of non-peptidyl glucagon receptor antagonists, we have utilized a direct targeting approach to replace the murine glucagon receptor with the human glucagon receptor gene by homologous recombination. Specific expression of the human glucagon receptor (GR) in the livers of transgenic mice was confirmed with an RNase protection assay, and the pharmacology of the human GRs expressed in the livers of these mice parallels that of human GR in a recombinant CHO cell line with respect to both binding of 125I-glucagon and the ability of glucagon to stimulate cAMP production. L-168,049, a non-peptidyl GR antagonist selective for the human GR shows a 3.5 fold higher affinity for liver membrane preparations of human GR expressing mice (IC50 = 172 +/- 98 nM) in the presence of MgCl2 in marked contrast to the measured affinity of the murine receptor (IC50 = 611 +/- 197 nM) for this non-peptidyl antagonist. The human receptors expressed are functional as measured by the ability of glucagon to stimulate cAMP production and the selectivity of this antagonist for the human receptor is further verified by its ability to block glucagon-stimulated cyclase activity with 5 fold higher potency (IC50 = 97.2 +/- 13.9 nM) than for the murine receptor (IC50 = 504 +/- 247 nM). Thus we have developed a novel animal model for evaluating GR antagonists in vivo. These mice offer the advantage that the regulatory sequences which direct tissue specific and temporal expression of the GR have been unaltered and thus expression of the human gene in these mice remains in the normal chromosomal context.

Animals↗
PubMed1998

Molecular characterization and expression of cloned human galanin receptors GALR2 and GALR3.

Galanin is a 29- or 30-amino acid peptide with wide-ranging effects on hormone release, feeding behavior, smooth muscle contractility, and somatosensory neuronal function. Three distinct galanin receptor (GALR) subtypes, designated GALR1, 2, and 3, have been cloned from the rat. We report here the cloning of the human GALR2 and GALR3 genes, an initial characterization of their pharmacology with respect to radioligand binding and signal transduction pathways, and a profile of their expression in brain and peripheral tissues. Human GALR2 and GALR3 show, respectively, 92 and 89% amino acid sequence identity with their rat homologues. Radioligand binding studies with 125I-galanin show that recombinant human GALR2 binds with high affinity to human galanin (K(D) = 0.3 nM). Human GALR3 binds galanin with less affinity (IC50 of 12 nM for porcine galanin and 75 nM for human galanin). Human GALR2 was shown to couple to phospholipase C and elevation of intracellular calcium levels as assessed by aequorin luminescence in HEK-293 cells and by Xenopus melanophore pigment aggregation and dispersion assays, in contrast to human GALR1 and human GALR3, which signal predominantly through inhibition of adenylate cyclase. GALR2 mRNA shows a wide distribution in the brain (mammillary nuclei, dentate gyrus, cingulate gyrus, and posterior hypothalamic, supraoptic, and arcuate nuclei), and restricted peripheral tissue distribution with highest mRNA levels detected in human small intestine. In comparison, whereas GALR3 mRNA was expressed in many areas of the rat brain, there was abundant expression in the primary olfactory cortex, olfactory tubercle, the islands of Calleja, the hippocampal CA regions of Ammon's horn, and the dentate gyrus. GALR3 mRNA was highly expressed in human testis and was detectable in adrenal gland and pancreas. The genes for human GALR2 and 3 were localized to chromosomes 17q25 and 22q12.2-13.1, respectively.

Amino Acid Sequence↗
PubMed1997

First report of recombination between the HLA-DR and HLA-DQ loci within a family.

Although unusual associations of HLA-DR and HLA-DQ alleles seen in ancestral haplotypes have indicated that recombination between these genes occurred in the past, an actual crossover event between DR and DQ has never been shown within a family. In a study of families with Graves' disease we have identified an individual from a three generation family who inherited a maternal haplotype that is the result of a recombinational event between the HLA-DR and the HLA-DQ loci on her chromosomes. Family members were typed for HLA class I by the lymphocyte microcytotoxicity test and for HLA class II by polymerase chain reaction (PCR) with sequence-specific primers or with sequence-specific oligonucleotide probes after PCR. Based on linkage disequilibrium it is likely that the recombinant haplotype is present in the proband rather than his brother. This haplotype was subsequently inherited by one of the proband's sons. The data presented support the conclusion that the recombinant haplotype resulted from a crossover event between the mother's DRB1 and DQA1 genes. Thus, recombination between the HLA-DR and HLA-DQ genes has been demonstrated within this family; a recombination event not previously described.

Adult↗
PubMed1997

Pharmacological characterization and tissue distribution of the human and rat GALR1 receptors.

The diverse biological functions of galanin are mediated via membrane bound high-affinity receptors. In order to identify and characterize potential galanin receptor subtypes, we have examined the specific 125I-galanin binding to the CHP-212 human neuroblastoma cell line. The galanin receptors expressed in CHP-212 cells, like GALR1 have high affinity for galanin (Kd = 0.07 nM) and the potency for inhibition of 125I-galanin binding by galanin peptides parallels that of hGALR1 expressed in a stable CHO cell line. We confirmed that GALR1 is expressed in these cells by RT-PCR. We further determined the tissue expression patterns of hGALR1 which is expressed in a variety of human tissues at a very low level, with the highest levels seen in heart, small intestine and prostate. A species of approximately 70 kDa is recognized by antisera specific for hGALR1 by Western blot analysis and should allow future measurements of receptor protein expression.

Amino Acid Sequence↗
PubMed1997

Molecular cloning and characterization of a new receptor for galanin.

Galanin (GAL) is a widely distributed neuropeptide with diverse biological effects including modulation of hormone release, antinociception and modification of feeding behavior. Its effects are mediated through G-protein-coupled receptors (GPCR) for which only a single type has been cloned, GAL receptor 1 (GALR1). We describe the cloning of a second galanin receptor type, GALR2, from rat hypothalamus. The GALR2 amino acid sequence is 38% identical to GALR1 and is pharmacologically similar to GALR1 when expressed in COS-7 cells. GALR2 is encoded by a single gene containing at least one intron and expressed in a diverse range of tissues.

Amino Acid Sequence↗
PubMed1997

Suramin disrupts insulin-like growth factor-II (IGF-II) mediated autocrine growth in human SH-SY5Y neuroblastoma cells.

Suramin, traditionally used in the treatment of trypanosomiasis, is under investigation in the treatment of cancer. One side effect that limits its use is the onset of a sensorimotor polyneuropathy. In order to investigate the mechanism by which suramin induces polyneuropathy, we examined its effects on SH-SY5Y human neuroblastoma cells, an in vitro model of neuronal growth and differentiation. Addition of 50-400 micrograms/ml suramin to SH-SY5Y cells grown in 0.6% CS inhibited [3H]thymidine ([3H]TdR) incorporation and cell growth. Upon removal of suramin, [3H]TdR incorporation increased, demonstrating that levels of suramin used were cytostatic and not cytotoxic. Analysis of suramin-treated SH-SY5Y cells by flow cytometry revealed growth arrest in the G1/G0 phase of the cell cycle. IGF-II-induced SH-SY5Y growth is mediated by the type I IGF receptor (IGF-IR). Therefore, we examined its effect on IGF-IR tyrosine phosphorylation. Suramin prevented IGF-II-stimulated IGF-IR tyrosine phosphorylation. These results indicate that in SH-SY5Y cells, suramin acts as a cytostatic agent and can block IGF-II-dependent cell growth by preventing IGF-IR activation. Thus, suramin toxicity in the peripheral nervous system may be due, in part, to preventing IGF and other growth factors from activating their receptors.

Brain Neoplasms↗