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D M Smith

Publications and source records attributed to D M Smith.

At least 199 records · Page 11Linked to original sources
PubMed1995

The enzymatic basis for the rat liver 6-hydroxymelatonin sulfotransferase activity.

Understanding 6-hydroxymelatonin (6HM) sulfation is deemed important to explaining normal and oncostatic actions of the pineal gland. Here we identify the enzymatic basis for this sulfation in rats. First, a quantitative assay was designed for measuring hepatic 6HM sulfotransferase (6HMST) activity. The assay was then used to identify a male dominant sexual dimorphism wherein liver from males contains double the 6HMST per g or per 100 g body weight seen in females. Examination of other rat tissues showed that most in vivo 6HM sulfation was likely to occur in liver. In addition, DEAE-Sephadex chromatography of liver cytosol indicated that 80-90% of the 6HMST activity in both sexes was due to an enzyme we named 6HMST II. A minor 6HM sulfotransferase (6HMST I) eluted from the columns prior to the main enzyme. 6HMST II, purified additionally, was shown to convert 6HM to a product that appeared to be 6HM sulfate (6-sulfa-toxymelatonin). The enzyme was inhibited by Na+, K+, Zn2+, and Cd2+. Its pH optimum was 7.80 +/- 0.30. Comparisons are made between 6HMST II, dopamine sulfotransferase II, and aryl sulfotransferase IV.

Animals↗
PubMed1995

Binding sites for islet amyloid polypeptide in mammalian lung: species variation and effects on adenylyl cyclase.

Islet amyloid polypeptide (IAPP) and calcitonin gene related peptide (CGRP) share a 47% sequence homology. IAPP can interact with adenylyl cyclase coupled CGRP receptors. We have examined [125I]IAPP binding in mouse, pig, and guinea pig lung membranes in competition with IAPP, CGRP, and CGRP(8-37). Three types of site were shown by order of potency: (i) mouse, IAPP > CGRP(8-37) >> CGRP; (ii) pig, CGRP > IAPP > CGRP(8-37); and (iii) guinea pig, CGRP = IAPP = CGRP(8-37). Chemical cross-linking of [125I]IAPP and [125I]CGRP binding sites in lung demonstrated that both sites had similar molecular weights in any one species but differed across species, i.e., mouse M(r) = 70,000 and 98,000; pig M(r) = 68,000, 56,000, and 47,000; and guinea pig M(r) = 106,000 and 56,000. Adenylyl cyclase activity was stimulated by forskolin and AlCl3-NaF in rat, mouse, pig, and guinea pig membranes. Only in mouse and pig were CGRP and IAPP able to stimulate adenylyl cyclase activity. In mouse lung CGRP and IAPP stimulated adenylyl cyclase activity with EC50 values of 642 +/- 222 nM (n = 4) and 325 +/- 115 nM (n = 4), respectively. In pig lung membranes EC50 values were 5.7 +/- nM (n = 4) for CGRP and 1230 +/- 1130 nM (n = 4) for IAPP. Thus IAPP either did not stimulate adenylyl cyclase activity in these lung membranes or did so with a low potency.

Adenylyl Cyclases↗
PubMed1995

Glucagon-like peptide-1 is a physiological incretin in rat.

Glucagon-like peptide-1 7-36 amide (GLP-1) has been postulated to be the primary hormonal mediator of the entero-insular axis but evidence has been indirect. The discovery of exendin (9-39), a GLP-1 receptor antagonist, allowed this to be further investigated. The IC50 for GLP-1 receptor binding, using RIN 5AH beta-cell membranes, was found to be 0.36 nmol/l for GLP-1 and 3.44 nmol/l for exendin (9-39). There was no competition by exendin (9-39) at binding sites for glucagon or related peptides. In the anaesthetized fasted rat, insulin release after four doses of GLP-1 (0.1, 0.2, 0.3, and 0.4 nmol/kg) was tested by a 2-min intravenous infusion. Exendin (9-39) (1.5, 3.0, and 4.5 nmol/kg) was administered with GLP-1 0.3 nmol/kg, or saline, and only the highest dose fully inhibited insulin release. Exendin (9-39) at 4.5 nmol/kg had no effect on glucose, arginine, vasoactive intestinal peptide or glucose-dependent insulinotropic peptide stimulated insulin secretion. Postprandial insulin release was studied in conditioned conscious rats after a standard meal. Exendin (9-39) (0.5 nmol/kg) considerably reduced postprandial insulin concentrations, for example by 48% at 15 min (431 +/- 21 pmol/l saline, 224 +/- 32 pmol/l exendin, P < 0.001). Thus, GLP-1 appears to play a major role in the entero-insular axis.

Anesthesia↗
PubMed1995

Pyroglutamyl-phenylalanyl-proline amide attenuates thyrotropin-releasing hormone-stimulated insulin secretion in perifused rat islets and insulin-secreting clonal beta-cell lines.

TRH immunoreactivity has been detected in the pancreas of man and rat and localized to the islets of Langerhans. We studied the effect of synthetic TRH and the related tripeptide pyroglutamyl-phenylalanyl-proline amide (EFP) on isolated perifused rat islets and the glucose-responsive clonal cell lines HIT-T15 and RIN5AH. TRH at 10 nM potentiated [0.5 +/- 0.1 (control) vs. 0.8 +/- 0.1 (TRH) pmol/10(6) cells per 120 min; mean +/- SEM; n = 6; P < 0.001; n = 15], whereas EFP from 1 nM upwards suppressed glucose-stimulated insulin secretion [0.8 +/- 0.1 (control) vs. 0.5 +/- 0.1 (EFP) pmol/10(6) cells per 120 min; P < 0.001; n = 12) in the cell lines. Further, EFP reversed TRH-stimulated insulin release. Similar responses were observed in perifused isolated rat islets at the tested dose of 1 microM. Gel permeation chromatography of rat adult and neonatal whole pancreas, isolated islets, and HIT cell extracts demonstrated the elution of total TRH-like immunoreactivity (t-TRH-LI) in the same position as synthetic TRH. Cation exchange analysis of the t-TRH-LI from rat adult pancreas and HIT cell extracts showed that neutral TRH-like peptides corresponding to synthetic EFP were also present. Reverse-phase fast protein liquid chromatographic analysis of t-TRH-LI in the unbound fraction of these extracts subjected to anion exchange columns, also demonstrated peaks corresponding to synthetic EFP. We conclude that TRH potentiates, whereas EFP inhibits, glucose-stimulated insulin release in isolated perifused islets and the cell lines. In addition, EFP reversed the stimulatory effect of TRH. The presence of EFP-LI in rat adult and neonatal pancreas and HIT cell extracts suggests it may contribute in the modulation of pancreatic endocrine function.

Amino Acid Sequence↗
PubMed1995

An abundant and specific binding site for the novel vasodilator adrenomedullin in the rat.

Rat adrenomedullin is a novel 50-amino acid peptide with structural similarities to the calcitonin family of peptides, calcitonin, calcitonin gene-related peptide (CGRP), and islet amyloid polypeptide (IAPP). Using rat [125I]adrenomedullin, specific binding sites were demonstrated in heart, lung, spleen, liver, soleus, diaphragm, gastrocnemius, and spinal cord membranes. The highest binding was present in heart and lung, which was further characterized. These sites exhibited saturation, dissociation, and competition. In rat lung, only rat (IC50 = 5.8 nM) and human (IC50 = 94 nM) adrenomedullin competed with [125I]adrenomedullin. However, in rat heart, rat (IC50 = 0.2 nM) and human (IC50 = 4.2 nM) adrenomedullin, IAPP (IC50 = 240 nM), and CGRP (IC50 = 1050 nM) all competed with [125I] adrenomedullin. Saturation analysis revealed binding capacities and dissociation constants of 2.8 +/- 0.3 pmol/mg protein and 1.3 +/- 0.3 nM, respectively, in lung and 0.47 +/- 0.11 pmol/mg protein and 0.41 +/- 0.14 nM in heart. Comparison with [125I]CGRP- and [125I]IAPP-binding sites in lung showed that rat adrenomedullin could potently inhibit at these sites (IC50 = 5 and 6 nM, respectively). Chemical cross-linking demonstrated a major band of 83,000 mol wt in lung, diaphragm, spleen, and liver and a band of 94,000 mol wt in heart, soleus, and gastrocnemius. Thus, [125I]adrenomedullin-binding sites in rat lung are abundant and can be differentiated from binding sites in rat heart, both pharmacologically and by mol wt.

Adrenomedullin↗
PubMed1995

Expression of a novel receptor for the calcitonin peptide family and a salmon calcitonin-like peptide in the alpha-thyrotropin thyrotroph cell line.

We have previously shown an increased incidence of alpha-subunit-producing thyrotroph tumors after salmon calcitonin (sCT) injection into rats. However, it is not clear whether the effects of CT are direct or indirect. Our hypothesis was that for sCT to act directly, it must have a binding site on thyrotrophs. The alpha TSH cell line was used as a model for thyrotrophs. Receptor binding studies using alpha TSH membranes revealed a high affinity binding site for sCT [IC50 = 0.97 +/- 0.18 nM (n = 4); Kd = 5.45 +/- 0.43 nM (n = 3); binding capacity = 6.6 pmol/mg protein (n = 3)]. Rat CT did not compete with binding at this site. Receptor screening for other CT peptide family members revealed high specific binding for CT gene-related peptide (CGRP; IC50 = 0.25 +/- 0.08 nM; n = 3) and islet amyloid polypeptide (IC50 = 4.36 +/- 1.1 nM; n = 3). This together with the absence of rat CT binding excluded a conventional CT-binding site, and we propose a site similar to the CGRP subtype III receptor described in the rat nucleus accumbens. Guanosine 5'O-(3-thiotriphosphate) (GTP gamma S) (20 microM), reduced [125I]CGRP binding to 38% of maximal, indicating that this site is G-protein coupled. Immunocytochemically, all of the cells displayed intense sCT-like immunoreactivity, which was totally abolished by preabsorption of the antibody with sCT. The presence of this receptor supports the hypothesis that sCT mediates tumorigenesis via a direct pituitary action and, together with the coexistence of a sCT-like peptide in these cells, provides evidence for a possible autocrine role of this peptide in the control of thyrotroph function.

Amyloid↗
PubMed1995

Survival of composite chondrocutaneous grafts by vessel implantation: a study in the rabbit ear model.

Composite chondrocutaneous graft reconstruction or reattachment has limited applicability, is traditionally restricted to small segmental losses, and is dependent on the status of the recipient bed and graft periphery for successful revascularization. Surgical enhancement of composite graft survival was experimentally investigated in the rabbit ear model through transposition and appositional placement of an adjacent vascular pedicle. Fluorescein-derived surface-survival determinations, microangiographic vessel-counting methods, and histologic analysis were used to study the effects of vascular augmentation, pedicle design variations, and angiogenic substance in sixty 8-cm2, full-thickness auricular grafts. A statistically significant survival advantage was demonstrated for the implanted grafts, secondary to perivascular angiogenesis from the implanted pedicle.

Angiogenesis Inducing Agents↗
PubMed1995

A 67 kDa protein mediates pituitary adenylate cyclase-activating polypeptide and vasoactive intestinal peptide-stimulated insulin secretion in a hamster clonal beta-cell line.

Insulin secretion is regulated by neural and neurohormonal factors. The report of nerves releasing pituitary adenylate cyclase-activating polypeptide (PACAP)--a 38 amino acid peptide--in the endocrine pancreas, suggests it may be important in modulating insulin release. We therefore carried out receptor-binding studies on membranes prepared from the glucose-responsive clonal beta-cell line HIT-T15, and also examined the effects of PACAP38, PACAP27--a C-terminal truncated form of the peptide--and vasoactive intestinal peptide (VIP) on insulin and islet amyloid polypeptide (IAPP) release. We showed by chemical cross-linking that PACAP and VIP stimulate secretion from the clonal cells by binding to a receptor with a molecular weight of 67 kDa (n = 4). Binding was saturable when membranes were incubated with 125I-PACAP27 (Kd 1.2 +/- 0.2 nM; Bmax 415.7 +/- 35.3 fmol/mg; n = 4) or 125I-VIP (Kd 1.3 +/- 0.3 nM; Bmax 354.8 +/- 42.8 fmol/mg; n = 4). We also demonstrated an increase in glucose-stimulated insulin (PACAP27, 366.6 +/- 25.8%; PACAP38, 389.9 +/- 13.4%; VIP, 342.6 +/- 16.1% of control; all at 1 microM, P < 0.01 vs control) and IAPP release (PACAP27, 236.9 +/- 26.2%; PACAP38, 226.5 +/- 10.9%; VIP, 242.9 +/- 15.8% of control; all at 1 microM, P < 0.01 vs control). Incubation of the cells with these peptides, for a duration of 12 h, in the presence of 5.5 mM glucose, did not alter the expression of insulin or IAPP. These findings suggest that PACAP and VIP stimulate secretion of insulin and IAPP by binding to a 67 kDa protein on clonal beta-cells and do not alter the transcription of insulin and IAPP under the conditions tested.

Amyloid↗
PubMed1995

The key to scheduling profitably.

Smart scheduling in the dental office can help staff use their time and resources more efficiently and increase the practice's profitability. Communication among staff members is key to achieving this goal.

Appointments and Schedules↗
PubMed1994

The presence of islet amyloid polypeptide/calcitonin gene-related peptide/salmon calcitonin binding sites in the rat nucleus accumbens.

Receptor autoradiographic analysis of binding in rat brain sections for [125I]islet amyloid polypeptide (IAPP), [125I]calcitonin gene-related peptide (CGRP) and [125I]salmon calcitonin indicated dense binding for all three ligands in the nucleus accumbens. Membrane binding studies revealed the existence of high affinity sites for all three peptides. The order of potency of various related peptides at each binding site was investigated and found for [125I]IAPP to be salmon calcitonin > IAPP = alpha CGRP > salmon calcitonin-(8-32); for [125I]CGRP to be alpha CGRP > IAPP > salmon calcitonin; and for [125I]salmon calcitonin to be salmon calcitonin > alpha CGRP > rat calcitonin > salmon calcitonin-(8-32) > IAPP, suggesting that [125I]IAPP targets the CGRP3 receptor subtype. This study confirms the existence of two receptors in the rat nucleus accumbens binding salmon calcitonin, one of which binds alpha CGRP and IAPP with a high affinity.

Amyloid↗
PubMed1994

A case manager intervention to reduce readmissions.

BACKGROUND: Acute hospitalizations represent substantial financial liability to closed health care systems. Among hospitalized patients, those with repeated admissions are high-cost users. Most managed care plans employ case management to control hospital use. This technique attempts to detect and fulfill unmet medical and social needs, intensify postdischarge care, identify and mobilize effective community services, and enhance primary care access. Despite the popularity of case management to control hospital use, few trials have examined its efficacy. METHODS: We conducted a randomized controlled trial of an intervention of case managers at a university-affiliated Veterans Affairs medical center. Six hundred sixty-eight patients aged 45 years or older who were discharged from the general medicine inpatient service, who had access to a telephone, and who received primary care at the hospital's clinics were randomized to the intervention (N = 333) and control (N = 335) groups. Within 24 hours of discharge, case managers mailed educational materials and access information to intervention patients, and within 5 days they called to review and resolve unmet needs, early warning signs, barriers to keeping appointments, and any readmissions. Case managers contacted intervention patients if they made no visits for 30 days. This resulted in a total of 6260 patient-case manager contacts. Control and intervention patients were followed up for 12 months. RESULTS: Intervention patients had more frequent visits per patient per month to the general medicine clinic (0.30 +/- 0.23 vs 0.26 +/- 0.22, P = .008), but we detected no significant differences between groups in nonelective readmissions, readmission days, or total readmissions. CONCLUSIONS: Frequent contacts for education, care, and accessibility by case managers using protocols were ineffective in reducing nonelective readmissions.

Aged↗
PubMed1994

Inhibition of T cell activation by a monoclonal antibody reactive against the alpha 3 domain of human MHC class I molecules.

A long term goal of investigators working in our laboratory has been to develop new mAbs for use as immunosuppressive agents. As a means toward achieving this goal, several new mAbs (hybridomas) have been developed by screening fusions for supernatants that possess T cell-inhibitory properties. Of these new mAbs, one mAb, designated 5H7, has been shown to possess both a unique combination of specificity for a monomorphic determinant of the alpha 3 domain of human class I MHC heavy chains and highly potent T cell inhibitory properties. mAb 5H7 profoundly inhibited T cell proliferation in response to anti-CD3 mAb in primary or secondary allogenic MLR or in primary human anti-mouse xenogenic MLR. mAb 5H7 inhibited expression of an early T cell activation marker, Leu23 (CD69); expression of IL-2Rs; and IL-2 production by both CD4+ and CD8+ T cells. mAb 5H7 inhibited IL-2 release by the Jurkat (E6-1) human T cell leukemia line in response to immobilized anti-CD3 mAb, thus, providing further evidence that 5H7 can inhibit activation directly at the level of the T cell. 5H7 profoundly blocked CD3-dependent (anti-CD3, anti-CD3 plus PMA, or anti-CD3 plus anti-CD28) pathways, but only partially blocked a CD3-independent (anti-CD28 plus PMA) pathway of T cell activation. In conclusion, class I MHC molecules that are expressed on the T cell may regulate early TCR/CD3-dependent signaling events. In addition, 5H7 mAb may provide a reagent for suppression of cellular immunity in vivo.

Antibodies, Monoclonal↗
PubMed1994

Enhanced synthesis of tumor necrosis factor-inducible proteins, plasminogen activator inhibitor-2, manganese superoxide dismutase, and protein 28/5.6, is selectively triggered by the 55-kDa tumor necrosis factor receptor in human melanoma cells.

We have demonstrated that A375 melanoma cells express mRNA for both types of tumor necrosis factor (TNF) receptors and receptor proteins on their plasma membranes. Specific agonist and blocking antibodies to either 55-kDa (TNF-R1) or 75-kDa (TNF-R2) TNF receptors combined with two-dimensional gel analysis were employed to determine which receptor type is responsible for mediating the induction of individual melanoma proteins. Our results indicate that the enhanced synthesis of proteins 21/>7 (M(r)/pI), 28/5.6, and 41/5.7 is selectively induced through TNF-R1. TNF induces these proteins; antagonist antibody to TNF-R1 prevents their induction by TNF, and TNF-R1 agonist induces them in the absence of TNF. Identification of these proteins by immunoblot analysis proved that 21/>7 is manganese superoxide dismutase, protein 28/5.6 is unrelated to 27/28-kDa heat shock protein, and protein 41/5.7 is plasminogen activator inhibitor-2. Furthermore, TNF cytotoxicity for A375 cells is also mediated by TNF-R1. These studies indicate that TNF-R1 is a critical signaling receptor for TNF action on A375 cells and demonstrate the potential use of TNF-R1 antibodies to selectively block or enhance specific effects of TNF on melanoma cells.

Base Sequence↗
PubMed1994

Histologic changes in prostate carcinomas treated with leuprolide (luteinizing hormone-releasing hormone effect). Distinction from poor tumor differentiation.

BACKGROUND: Treatment of patients with prostate carcinoma using drugs with antiandrogenic effects is increasingly common. One of the most popular agents is leuprolide (with or without flutamide), an agonist of luteinizing hormone-releasing hormone (LH-RH). Prostates exposed to antiandrogenic agents develop distinctive pathologic changes. The ability of a group of 18 pathologists to recognize these LH-RH effects on prostate cancers and to distinguish them from poor tumor differentiation was tested using a Delphi technique. METHODS: The Delphi method maximizes the opportunity for individual contributions by eliminating opportunities for group dynamics and confrontational interaction. In this study, participants were provided with a set of duplicate transparencies and a list of written definitions of both poor tumor differentiation and LH-RH effect. They were instructed to examine the cases privately using the definitions provided and to distinguish poor tumor differentiation from LH-RH effect. Responses of the participants were shared anonymously, and each was asked to reexamine the cases in light of this knowledge. RESULTS: After a learning experience accomplished by one round of examination and feedback, LH-RH effect could be correctly distinguished from poor tumor differentiation in greater than 90% of cases by 83% of the participants. A small percentage of cases confounded complete consensus, although every case was correctly identified by most participants. CONCLUSIONS: The pathologic effects associated with androgen deprivation with leuprolide (plus flutamide) are sufficiently distinctive to be recognized by knowledgeable pathologists. In most cases, the tissue shrinkage representing a favorable response to the drug can be distinguished from solid or infiltrating patterns of poorly differentiated carcinoma.

Cell Differentiation↗
PubMed1994

Exercise classes in rest homes: effect on physical function.

AIM: To determine if regular exercise classes, planned and supervised by a physiotherapist, improved physical function in a sample of frail institutionalised elderly. METHOD: Weekly one hour and twice weekly 10 minute exercise classes were given by a physiotherapist, to a group of 34 residents who consented from randomly selected rest homes. The study took place over a period of one year. At entry, 4, 9 and 12 months the physical function was compared with the matched control group who received regular rest home activities, without physiotherapy input. The study used a sit stand test to measure change in physical function. This was indicated by recording (a) the time taken and (b) the hand assistance required to stand up. RESULTS: Compared with the control group, the study group showed statistically significant improvement (p < 0.05) in physical function as measured by sit stand time. The sit stand hand assistance test was found to be too insensitive to measure improvement or to differentiate between the two groups. The programme was enjoyable and had no health complications. CONCLUSION: Regular exercise classes planned and supervised by a physiotherapist are safe and improve the physical functioning and daily activity levels of the frail, institutionalised elderly. They are a cost effective use of the physiotherapist's limited time.

Aged↗
PubMed1994

Molecular genetic changes associated with ovarian cancer.

Multiple specific chromosomal deletions can be found in human epithelial ovarian cancer by cytogenetic analysis or molecular techniques. Somatic allelic deletion or loss of heterozygosity (LOH) in a tumor is considered circumstantial evidence for the location of tumor suppressor genes. We have examined 27 primary epithelial ovarian tumors for the presence of LOH at 19 polymorphic markers on chromosomes 1, 5, 6, 9, 11, 13, and 17. Markers near the adenomatous polyposis coli (APC) gene at 5q21 showed LOH in 50% (10/20) of informative cases. LOH was seen in 53% (8/15) at the IFNA locus on 9p, another region implicated in other tumors, but not previously associated with ovarian cancer. We observed LOH for markers on 11p15 in 50% (12/24) of ovarian cancer DNAs from informative cases, while only 25% (4/16) at 11q13 and 29% (5/17) at 11q24 showed LOH. Only a portion of distal 11p was deleted in six cases. The incidence of LOH (50%) at HGH (17q22-q24) was greater than that at D17S579 (39%; 17q21), a locus tightly linked to BRCA1. Sixty-four percent (7/11) showed allelic loss at 17p11. LOH was infrequently observed at markers on chromosomes 1, 6, and 13q. Most cases showing LOH were stage III or IV, and most showed LOH at more than one locus. These studies support the concept that multiple genetic loci are involved in ovarian tumorigenesis. Two additional regions thought to harbor genes important in other cancers, 5q21 and 9p21, can now be added to the growing spectrum of molecular alterations seen in ovarian cancer.

Adult↗