Search PubMed⌕ Search

Biomedical subjects

D M Smith

Publications and source records attributed to D M Smith.

At least 19 recordsLinked to original sources
PubMed1997

Identification of constitutive and gamma-interferon- and interleukin 4-regulated proteins in the human renal carcinoma cell line ACHN.

The effects of IFN-gamma and interleukin 4 (IL-4) on cell proliferation and two-dimensional gel electrophoretic protein patterns of the human renal carcinoma cell line ACHN were studied. Treatment of the cells with IFN-gamma resulted in a 40-50% decrease in their proliferation. IL-4 treatment resulted in a 30-40% decrease. Treating cells with both cytokines had the same effect as with IFN-gamma alone, thus precluding a synergistic antiproliferative interaction of these two cytokines. To identify IL-4- and IFN-gamma-regulated proteins in ACHN, two-dimensional preparative gel electrophoresis was used, combined with either capillary electrophoresis or high-performance liquid chromatography and either Edman or mass spectrometric sequencing. The following cytokine-induced proteins were identified: tropomyosin, heat shock protein 27, manganese superoxide dismutase, glutathione S-transferase pi, and protein kinase C inhibitor I. Tropomyosin increased 2-fold when cells were treated with IFN-gamma. Levels of heat shock protein 27 increased 2-fold with IL-4, 3-fold with IFN-gamma, and 4-fold when the cytokines were used in combination. Manganese superoxide dismutase increased 3-fold with IFN-gamma but was unaffected by IL-4. Glutathione S-transferase pi increased 3-fold with IFN-gamma. Levels of protein kinase C inhibitor I increased greater than 3-fold with IL-4, 4-fold with IFN-gamma, and 7-fold when both cytokines were used. In addition, the following constitutive ACHN proteins were identified: copper zinc superoxide dismutase, 60S acidic ribosomal protein P2, and a second heat shock protein 27 isoform. These findings help define the biochemical modes of action of IFN-gamma and IL-4 and their potential in the biological therapy of renal cell carcinoma.

Amino Acid Sequence↗
PubMed1997

Anti-human class I MHC antibodies induce apoptosis by a pathway that is distinct from the Fas antigen-mediated pathway.

5H7, an anti-human class I MHC mAb that recognizes a monomorphic determinant of the alpha3 domain, profoundly inhibits T lymphocyte activation. The present study was designed to determine the role of programmed cell death in 5H7-mediated immune suppression. Incubation of PBMC with 5H7 mAb induced a marked reduction in viable cell recovery (VCR) of T cells (<10% VCR), NK cells (<1% VCR), and B cells (<1% VCR). In addition, 5H7 inhibited proliferation and VCR of JY, DBS-521, and BevD tumor lines as well as cells transfected with HLA-B27. Morphologic changes characteristic of apoptosis were induced by 5H7, including cell membrane blebbing, cytoplasmic vacuolization, condensation of nuclear chromatin, and nuclear fragmentation. Furthermore, DNA fragmentation was demonstrated in 5H7-treated PBMC using a TdT-mediated end-labeling (TUNEL) technique. 5H7, but not anti-Fas mAb, induced apoptosis of cell lines derived from patients with Niemann-Pick disease that lack acidic sphingomyelinase activity. In addition, induction of cell death by 5H7 was not inhibited by IL-1beta-converting enzyme (ICE) inhibitors under conditions that fully suppressed Fas-mediated apoptosis. These findings suggest that signal transduction through "classical" human class I MHC molecules induces apoptosis by a Fas-independent pathway that does not require acidic sphingomyelinase, is independent of ICE protease, and may represent a unique pathway of cell death in human lymphocytes.

Adult↗
PubMed1997

Species, muscle type, and formulation influence the residual concentrations of three endpoint temperature indicators in poultry products.

The objectives were to determine 1) the effect of muscle type and temperature on the concentration of lactate dehydrogenase (LDH), IgG, and serum albumin (SA) in turkey and chicken, and 2) the concentrations of these marker proteins in four turkey products processed to the USDA required endpoint temperature (EPT) of 68.3 C. Ground turkey and chicken breast and thigh meat were placed in thermal death time tubes and heated to 65.5, 68.3, 71.1, or 73.8 C. Turkey bologna, pastrami, smoked sausage, and frankfurters were cooked by a local processor to target temperatures between 65.5 and 73.8 C. Proteins were extracted and quantified by an LDH sandwich ELISA or indirect competitive ELISA for SA and IgG. Although all indicators decreased in concentration as EPT were increased, turkey contained higher (P < 0.0001) residual concentrations of the indicators than chicken. In both turkey and chicken, thigh muscle contained higher concentrations of SA (P < 0.0001) and IgG (P < 0.0001) and lower concentrations of LDH (P < 0.0001) than breast muscle at the same EPT. The concentrations of the three indicators decreased as the EPT of the four turkey thigh products were increased. At an EPT of 68.3 C, pastrami contained the lowest concentrations of SA and IgG, whereas bologna contained the lowest concentration of LDH. The LDH ranged from 441.7 ng/g bologna to 1,283.6 ng/g sausage at 68.3 C. Minimum residual concentrations of each indicator need to be established for each product before the ELISA can be used to verify processing to the USDA minimum EPT.

Analysis of Variance↗
PubMed1997

Composition and functional properties of cholesterol reduced egg yolk.

The composition and functional properties of cholesterol reduced egg yolk (CREY) were compared to those of control egg yolk (EY). The CREY was prepared by absorbing cholesterol with beta-cyclodextrin after dilution and dissociation of granules at pH 10.5. The CREY contained less lipid and protein and more carbohydrate and ash than EY. Egg lipids were fractionated into triglycerides, cholesterol esters, free cholesterol, phosphatidyl choline, and phosphatidyl ethanolamine. Free and esterified cholesterol in CREY were reduced by 91.6 and 94.4%, respectively. Triglycerides were the major lipid class in CREY. The CREY contained more oleic acid and less linoleic acid than the control. Protein solubility in 0.1 and 0.6 M NaCl and sponge cake volume did not differ. The composition of proteins soluble in 0.6 M NaCl in both egg preparations were similar as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The electrophoretic profiles of proteins soluble in 0.1 M NaCl were similar, except that lipovitellin form EY was insoluble under these conditions. The CREY was less yellow than EY, as indicated by beta-carotene concentrations and Hunter b values. These results suggest that beta-cyclodextrin can be used to produce a low cholesterol egg product with compositional and functional properties similar to EY.

Animals↗
PubMed1997

Depression and mortality in medically ill older adults.

OBJECTIVE: To determine if major depressive disorder influences the survival of older, medically ill veterans. DESIGN: Thirty month follow-up of an inception cohort. SETTING: An academically-affiliated Veterans Affairs Medical Center. PARTICIPANTS: One hundred veterans, half of whom have a depressive disorder, recruited from inpatient medical and surgical units in 1990-1991. At initial evaluation all participants were older than age 65, cognitively intact, and medically but not terminally ill. MEASUREMENTS: Severity of medical illness, functional status, social support and presence of depression were measured at initial evaluation, presence of depression was measured 4 to 6 months after initial evaluation, and survival status was determined at 30 months. Retrospective medical record review was used to determine if patients received depression treatment and to explore whether depressed persons received life-sustaining medical treatment less often than nondepressed persons. RESULTS: At 30 months, 36 of the 100 subjects had died. Only two factors predicted mortality: severity of medical illness and depression. The survival of subjects who had recovered from depression at 4 to 6 months was intermediate to never depressed and continuously depressed subjects. Somatic treatment of depression did not alter outcome. There was no significant difference between depressed and nondepressed in how often life-sustaining medical treatments were withheld or withdrawn. CONCLUSION: Presence of major depression in medically-ill older hospitalized veterans continues to be a risk factor for death 30 months after diagnosis.

Activities of Daily Living↗
PubMed1997

Neuropeptide Y induced feeding in the rat is mediated by a novel receptor.

There are now six recognized neuropeptide Y (NPY) receptor subtypes (Y1-Y4 and two recently cloned distinct receptors labeled Y5), of which Y1 and one of the Y5's have been suggested could mediate the effect of NPY on feeding. The fragments NPY(2-36) and NPY(3-36), which bind Y1 only poorly, were injected intracerebroventricularly (icv) and found to have similar dose-response relationships to NPY in the stimulation of feeding. However NPY (13-36), which stimulates both Y2 and Y5, caused no increase in food intake, even at high doses. Maximal stimulation with the classical Y1 agonist [Pro34]-NPY produced only 50% of the maximum effect of NPY itself despite fully inhibiting adenylyl cyclase activity in vitro in a Y1 system. The novel fragment [Pro34]-NPY(3-36) is as effective at stimulating food intake as the classical Y1 analogue [Pro34]-NPY but bound to the Y1 receptor with only 1/20th of the affinity of NPY and failed to inhibit adenylyl cyclase through this receptor. [Pro34]-NPY(3-36) is therefore a relatively appetite-selective ligand. Coadministration of high dose NPY(13-36) and [Pro34]NPY did not enhance feeding compared with [Pro34]-NPY alone. In addition, the NPY Y1 receptor antagonist BIBP-3226, which does not bind Y2, Y4, or Y5 receptors, significantly reduced NPY induced feeding. These results indicate that the feeding effect of icv NPY involves a novel receptor and that it is functionally distinct from the recognized receptor subtypes.

Adenylyl Cyclases↗
PubMed1997

Protein requirements of sheep in late pregnancy: partitioning of nitrogen between gravid uterus and maternal tissues.

The objective of this study was to quantify effects of maternal protein nutrition on N accretion or loss in conceptus and maternal tissues of ewes during late pregnancy. Ewes, pregnant with twins, were fed low (LP, 79 g CP/kg DM), medium (MP, 116 g CP/kg DM), or high (HP, 157 g CP/kg DM) protein diets, each with an estimated ME concentration of 2.7 Mcal/kg DM, between d 111 and 140 of pregnancy; all ewes had been fed the same diet (2.7 Mcal ME, 120 g CP/kg DM) for the previous 30 d (d 80 to 110). Dry matter intakes were varied (LP = 1.0, MP = 1.2, and HP = 1.4 kg/d) according to predicted energy costs of protein deposition for each diet. Nitrogen accretion was estimated by comparative slaughter (d 140 minus d 110) and by collection of excreta between d 120 and 130. Fresh weights of maternal and gravid uterine tissues were measured at slaughter, before proximate analysis of these components. Whole-body N retention was directly and linearly related to N intake, but efficiency of deposition of apparently absorbed N decreased linearly with increasing N intake (LP, .79; MP, .70; HP, .62). Nitrogen accretion in the gravid uterus, maternal viscera, and mammary gland was significantly less in LP than in MP or HP ewes. Nitrogen balance in maternal carcass tissues was linearly related to N intake, ranging from a negative value in LP ewes to a positive value in HP ewes (LP, -63 g; MP -39 g; HP, 55 g). These data provide the basis for estimating N requirements for protein accretion in the conceptus and in maternal tissues during late pregnancy. They also highlight the capacity of maternal carcass tissues to mobilize or deposit amino acids in response to variations in dietary protein supply.

Animals↗
PubMed1996

A rat skeletal muscle cell line (L6) expresses specific adrenomedullin binding sites but activates adenylate cyclase via calcitonin gene-related peptide receptors.

We have previously demonstrated specific binding sites for adrenomedullin, a novel member of the calcitonin family of peptides, in rat muscles. It is unclear whether these receptors are vascular or muscular. Receptors for the structurally similar calcitonin gene-related peptide (CGRP) are present on myocytes and might be involved in the regulation of myocyte glucose metabolism and control by motor neurons. We investigated whether adrenomedullin binding sites were present on L6 myocytes. Specific [125I]adrenomedullin binding sites were demonstrated where adrenomedullin competed with an IC50 of 0.22 +/- 0.04 nM (mean +/- S.E.M.) and a concentration of binding sites (Bmax) of 0.95 +/- 0.19 pmol/mg of protein (mean +/- S.E.M.). CGRP and the specific CGRP receptor antagonist CGRP(8-37) competed weakly at this site (IC50 > 10 and 601 +/- 298 nM respectively). Binding studies with [125I]CGRP revealed a binding site for CGRP (IC50 = 0.13 +/- 0.01 nM; Bmax = 0.83 +/- 0.10 pmol/mg of protein) where both CGRP(8-37) and adrenomedullin competed with [125I]CGRP with IC50 values of 1.15 +/- 0.12 and 8.68 +/- 0.98 nM respectively. Chemical cross-linking showed the CGRP and adrenomedullin binding site-ligand complexes to have approximate molecular masses of 82 and 76 kDa respectively. Both CGRP and adrenomedullin increased adenylate cyclase activity with similar potencies. In both cases adenylate cyclase activation was blocked by CGRP(8-37). Stimulation with 10 nM adrenomedullin or CGRP caused an increase in the percentage of total activated cellular cAMP-dependent protein kinase from 38% in resting cells to 100% and 98% respectively. Therefore in L6 cells adrenomedullin can bind to CGRP receptors, activating adenylate cyclase and cAMP-dependent protein kinase.

Adenylyl Cyclases↗
PubMed1996

Specific adrenomedullin binding sites and hypotension in the rat systemic vascular bed.

The potent vasodilator peptide, adrenomedullin, has been shown to be present in plasma, suggesting a physiological role in cardiovascular control. Here we investigated the hypotensive action of adrenomedullin in vivo, using the anaesthetised rat as the bioassay model, and adrenomedullin binding sites using ligand binding assays on rat blood vessel membranes. Rat alpha CGRP and both human and rat adrenomedullins induced dose-dependent, powerful and long-lasting hypotensive effects. At peptide doses used in this study (0.02-2 nmol/kg), the efficacy of both human and rat adrenomedullins was lower than that of rat alpha CGRP. The CGRP1-receptor antagonist, human CGRP(8-37) (200 nmol/kg) was able to completely inhibit the hypotensive effect of rat alpha CGRP (0.2 nmol/kg) but not that of rat adrenomedullin (2 nmol/kg), implying that the adrenomedullin action is independent of CGRP1-receptors. Ligand binding assays confirmed the presence of both CGRP and adrenomedullin binding sites in rat blood vessels. The 125I-rat adrenomedullin binding site has a Kd = 0.32 +/- 0.12 nM (n = 4) for rat adrenomedullin but has a Ki > 10(-6) M for rat alpha CGRP. Chemical cross-linking and SDS-PAGE analysis revealed theadrenomedullin binding protein to have a M(r) of 83000 with a minor band of M(r) = 99000. The results suggest that the hypotensive effect of adrenomedullin may be mediated via specific adrenomedullin binding sites, in vivo.

Adrenomedullin↗
PubMed1996

Some techniques for the analysis of work sampling data.

Work sampling is an observational technique that produces counts representing the number of times that an individual has been observed performing each of several tasks. These data are collected using either systematic or random times of observation, and typically exhibit correlation between repeated observations on the same individual, with the degree of correlation being a function of the amount of time elapsed between measurements. Using several recently developed statistical techniques, we illustrate how it is possible to carry out analyses of these nominal outcomes that account for the correlation between repeated outcomes. We use description of a work sampling study to motivate the techniques and we compare empirically results from analyses based on several different underlying assumptions.

Adult↗
PubMed1996

A role for glucagon-like peptide-1 in the central regulation of feeding.

The sequence of glucagon-like peptide-1 (7-36) amide (GLP-1) is completely conserved in all mammalian species studied, implying that it plays a critical physiological role. We have shown that GLP-1 and its specific receptors are present in the hypothalamus. No physiological role for central GLP-1 has been established. We report here that intracerebroventricular (ICV) GLP-1 powerfully inhibits feeding in fasted rats. ICV injection of the specific GLP-1-receptor antagonist, exendin (9-39), blocked the inhibitory effect of GLP-1 on food intake. Exendin (9-39) alone had no influence on fast-induced feeding but more than doubled food intake in satiated rats, and augmented the feeding response to the appetite stimulant, neuropeptide Y. Induction of c-fos is a marker of neuronal activation. Following ICV GLP-1 injection, c-fos appeared exclusively in the paraventricular nucleus of the hypothalamus and central nucleus of the amygdala, and this was inhibited by prior administration of exendin (9-39). Both of these regions of the brain are of primary importance in the regulation of feeding. These findings suggest that central GLP-1 is a new physiological mediator of satiety.

Animals↗
PubMed1996

Adrenomedullin stimulates DNA synthesis and cell proliferation via elevation of cAMP in Swiss 3T3 cells.

Our results demonstrate that the novel vasoactive regulatory peptide adrenomedullin is a potent mitogen for Swiss 3T3 cells. Acting via a specific adrenomedullin receptor, it stimulates a dose-dependent increase in DNA synthesis in synergy with insulin. Additionally, adrenomedullin stimulates further progression through the cell cycle resulting in cell proliferation, an effect that was further enhanced by the presence of insulin. Adrenomedullin rapidly induces accumulation of intracellular cAMP but does not stimulate an increase in intracellular Ca2+, activation of protein kinase C, or tyrosine phosphorylation of intracellular substrates. Adrenomedullin-stimulated mitogenesis is markedly enhanced in Swiss 3T3 cells stably transfected with a constitutively activated Gs alpha, which are highly sensitive to agents that elevate cAMP, and is inhibited by the PKA inhibitor H-89. Adrenomedullin is, thus, identified as a novel mitogenic regulatory peptide acting via cAMP.

3T3 Cells↗
PubMed1996

Lability of the presumptive pectoral and pelvic girdle territories compared using celomic grafts.

The concept of "limbness" usually implies "wingness" or "legness". We present evidence in this study that in the case of the proximal forelimb, limbness may exist independent of wingness. Presumptive wing and leg regions were grafted into the opposite limb (wing into leg or leg into wing) of stage 18 hosts. Several stage 10+ to 11+ presumptive wing regions grafted into host legs developed pelvic girdles associated with distal wing elements. This phenomenon was not seen in presumptive wing region grafts older than stage 12-. Presumptive leg regions grafted into host wings developed leg structures only, including pelvic girdle and leg digits. The pectoral girdle appears to be labile in early stages, whereas the pelvic girdle is not labile even during early stages.

Animals↗
PubMed1996

Mesonephros has a role in limb development and is related to thalidomide embryopathy.

Recent studies have demonstrated a link between limb reduction defects and mesonephros removal [Geduspan and Solursh, 1992) Dev. Biol., 151:242-250]. However, there is some question as to whether the limb-reduction defects seen in that study resulted from the removal of mesonephros or from the formation of scar tissue medial to the limb territory. The current study was conducted to test the hypothesis that elimination of the mesonephros without producing scar tissue adjacent to the limb will adversely affect limb morphogenesis. The hypothesis was tested by the insertion of tantalum foil barriers into various levels of the intermediate mesoderm of developing chick embryos to prevent the caudal elongation of the mesonephros. Limb reduction defects were obtained when the mesonephros was prevented from forming caudal to somite 14. No limb defects were seen when a foil barrier was placed into the intermediate mesoderm at the level of somite 21 or 25. Our results support the notion that a signal from the mesonephros is necessary for normal limb development. In addition, it appears that a craniocaudal factor emanating from the mesonephros plays a role in limb development. The limb reduction defects obtained in this study were also compared to the pattern of thalidomide embryopathy in humans. There is a close correspondence between the types of limb reduction anomalies seen with thalidomide and mesonephric blocks and between the severity of defects vs. the timing of thalidomide intake or mesonephric blockage. A model for possible thalidomide embryopathy is presented.

Animals↗
PubMed1996

Risk factors for nonelective hospital readmissions.

We previously reported a predictive model that identified potentially modifiable risk factors for nonelective readmission to a county hospital. The objectives of this study were to determine if those risk factors were generalizable to a different population. We found that the previously reported risk factors were generalizable, and other potentially modifiable risk factors were identified in this population of veterans. However, further research is needed to establish whether or not the risk factors can be modified and whether or not modification improves outcomes.

Aged↗
PubMed1996

Reduced NPY induced feeding in diabetic but not steroid-treated rats: lack of evidence for changes in receptor number or affinity.

Concentrations of the potent hypothalamic appetite stimulating peptide neuropeptide Y (NPY), and its mRNA, are increased in rats with experimental diabetes, suggesting a role in the hyperphagia of this disorder. The 2-h feeding responses to intracerebroventricular (i.c.v.) injection of neuropeptide Y (NPY) (5, 10, and 15 mu g doses) were measured in male Wistar rats treated with streptozotocin (55 mg/kg) to induce diabetes. Streptozotocin-diabetic rats given i.c.v. NPY exhibited reduced feeding responses compared to controls (P < 0.05). Dexamethasone treated rats exhibit similar changes in NPY content and mRNA in the hypothalamus to those seen in diabetes, but are not hyperphagic. Feeding responses were also measured in this model, to assess whether high levels of endogenous NPY might account for the reduced response in diabetes. In contrast, the feeding response to NPY in comparison to controls was unaltered in dexamethasone treated rats. To investigate whether altered NPY receptor number or affinity, was the underlying mechanism for these divergent responses, receptor binding experiments were performed using (125)I-PYY and membranes prepared from rat hypothalamus. No significant difference was found in receptor number or affinity between the 2 groups (B(max): 114.7 +/- 18.9 vs 127.4 +/- 27.1 fmol/mg protein, K(d): 99.6 +/- 28.2 vs 135.1 +/- 32.4 pM). Similarly no difference was found between hypothalamic membranes prepared from dexamethasone-treated and control animals. NPY receptor subtypes in the hypothalamus were compared with that of cortex (predominantly Y1) and hippocampus (predominantly Y2) using the Y1-specific ligand [Leu(31)Pro(34)] NPY. These studies showed that the binding profile in the hypothalamus most closely matched that in the hippocampus, suggesting that the majority of hypothalamic receptors were of the Y2 subtype. Receptor autoradiography revealed low binding in the hypothalamus, and particularly in the paraventricular nucleus of the hypothalamus. Competition with [Leu(31)Pro(34)] NPY confirmed that only a low density of binding to Y1 like receptors was present in the hypothalamus. No difference was observed between control and streptozotocin treated animals. The feeding response to exogenous NPY is reduced in experimental diabetes, but not in dexamethasone treated rats. These differing responses do not appear to be due to altered NPY receptor number or affinity in the hypothalamus.

Animals↗