Biomedical subjects

T Boone

Publications and source records attributed to T Boone.

At least 37 records · Page 2Linked to original sources

PubMed1997

GFRalpha-2 and GFRalpha-3 are two new receptors for ligands of the GDNF family.

The receptor for glial cell line-derived neurotrophic factor (GDNF) consists of GFRalpha-1 and Ret. Neurturin is a GDNF-related neurotrophin whose receptor is presently unknown. Here we report that neurturin can bind to either GFRalpha-1 or GFRalpha-2, a novel receptor related to GFRalpha-1. Both GFRalpha-1 and GFRalpha-2 mediate neurturin-induced Ret phosphorylation. GDNF can also bind to either GFRalpha-1 or GFRalpha-2, and activate Ret in the presence of either binding receptor. Although both ligands interact with both receptors, cells expressing GFRalpha-1 bind GDNF more efficiently than neurturin, while cells expressing GFRalpha-2 bind neurturin preferentially. Cross-linking and Ret activation data also suggest that while there is cross-talk, GFRalpha-1 is the primary receptor for GDNF and GFRalpha-2 exhibits a preference for neurturin. We have also cloned a cDNA that apparently codes for a third member of the GFRalpha receptor family. This putative receptor, designated GFRalpha-3, is closely related in amino acid sequence and is nearly identical in the spacing of its cysteine residues to both GFRalpha-1 and GFRalpha-2. Analysis of the tissue distribution of GFRalpha-1, GFRalpha-2, GFRalpha-3, and Ret by Northern blot reveals overlapping but distinct patterns of expression. Consistent with a role in GDNF function, the GFRalphas and Ret are expressed in many of the same tissues, suggesting that GFRalphas mediate the action of GDNF family ligands in vivo.

Amino Acid Sequence↗

PubMed1997

Molecular cloning and functional characterization of a novel CC chemokine, stimulated T cell chemotactic protein (STCP-1) that specifically acts on activated T lymphocytes.

A novel human chemokine STCP-1 (stimulated T cell chemotactic protein) was isolated from an activated macrophage cDNA library. The chemokine has four cysteines positioned in a manner that identifies STCP-1 as a member of the CC chemokine family. The amino acid sequence shows 34% identity with RANTES. The gene consists of 3 exons and 2 introns with the position of intron/exon boundaries similar to that of RANTES. The gene is expressed as a 3.4-kilobase transcript on lymph node, thymus, and Appendix. STCP-1 induces Ca2+ mobilization in a small percentage of primary activated T lymphocytes, but on repeated stimulation the percentage of T lymphocytes that respond to STCP-1 increases. The chemokine STCP-1 does not induce Ca2+ mobilization in monocytes, dendritic cells, neutrophils, eosinophils, lipopolysaccharide-activated B lymphocytes, and freshly isolated resting T lymphocytes. Similarly, STCP-1, while acting as a mild chemoattractant for primary activated T lymphocytes, is a potent chemoattractant for chronically activated T lymphocytes but has no chemoattractant activity for monocytes, neutrophils, eosinophils, and resting T lymphocytes. As STCP-1 acts specifically on activated T lymphocytes, it may play a role in the trafficking of activated/effector T lymphocytes to inflammatory sites and other aspects of activated T lymphocyte physiology.

Amino Acid Sequence↗

PubMed1997

Osteoprotegerin: a novel secreted protein involved in the regulation of bone density.

A novel secreted glycoprotein that regulates bone resorption has been identified. The protein, termed Osteoprotegerin (OPG), is a novel member of the TNF receptor superfamily. In vivo, hepatic expression of OPG in transgenic mice results in a profound yet nonlethal osteopetrosis, coincident with a decrease in later stages of osteoclast differentiation. These same effects are observed upon administration of recombinant OPG into normal mice. In vitro, osteoclast differentiation from precursor cells is blocked in a dose-dependent manner by recombinant OPG. Furthermore, OPG blocks ovariectomy-associated bone loss in rats. These data show that OPG can act as a soluble factor in the regulation of bone mass and imply a utility for OPG in the treatment of osteoporosis associated with increased osteoclast activity.

Amino Acid Sequence↗

PubMed1996

Refined crystal structure and mutagenesis of human granulocyte-macrophage colony-stimulating factor.

The crystal structure of recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) has been refined against data extending to a resolution of approximately 2.4 A along a* and approximately 1.9 A along b* and c*. Anisotropic scale factors of B11 = -20.8 A2, B22 = 7.4 A2, B33 = 13.3 A2 corrected for the more rapid fall of diffraction in the a* direction. The anisotropy correlates with the weak crystal packing interactions along the a axis. In addition to apolar side chains in the protein core, there are 10 buried hydrogen bonding residues. Those residues involved in intramolecular hydrogen bonding to main chain atoms are better conserved than those hydrogen bonding to other side chain atoms; 24 solvation sites are observed at equivalent positions in the two molecules in the asymmetric unit, and the strongest among these are located in clefts between secondary structural elements. No buried water sites are seen. Two surface clusters of hydrophobic side chains are located near the expected receptor binding regions. Mutagenesis of 11 residues on the helix A/helix C face confirms the importance of Glu-21 and shows that Gly-75 and Gln-86, located on helix C, each cause a greater than fourfold drop in activity. Glu-21 and Gly-75, but not Gln-86, are structurally equivalent to residues involved in the growth hormone binding to its receptor.

Amino Acid Sequence↗

PubMed1996

Cloning and expression of the gene encoding a novel proteinase from Tritirachium album limber.

We have isolated the cDNA and the genomic clones encoding a novel serine proteinase, named proteinase T, from the fungus Tritirachium album Limber. The coding region of the gene is interrupted by two introns. The amino acid sequence of proteinase T as deduced from the nucleotide sequence is about 56% identical to that of proteinase K. Four cysteines are present in the mature proteinase, probably in the form of disulfide bonds. We have also purified the native proteinase from Tritirachium album Limber grown in the presence of 2% skim milk. Proteinase T is extremely stable at 50 degrees C. The thermal stability is not affected in the presence of 1% SDS either at pH 8.0 or 10.0. We have expressed the cDNA of proteinase T in Escherichia coli. The authenticity of the proteinase has been characterized by Western blotting and amino terminal analysis of the recombinant product. High level expression of proteinase T in E. coli as well as the refolding process to generate active proteinase will be discussed in detail.

Amino Acid Sequence↗

PubMed1996

Effects of assistive devices on cardiorespiratory demands in older adults.

BACKGROUND AND PURPOSE: The purpose of this study was to quantify and compare cardiorespiratory demands imposed during unassisted ambulation and ambulation with various assistive devices in older adults. SUBJECTS: Ten volunteers (3 male, 7 female) who were not dependent on assistive devices for ambulation, with a mean age 60.3 years (SD = 8.4, range = 50-74), participated. METHODS: Immediately after a 5-minute steady-state session with each of the assistive devices tested (standard walker, wheeled walker, and single-point cane), subjects ambulated for 2 minutes at a self-selected speed with each device and unassisted while selected cardiorespiratory and metabolic variables were monitored. RESULTS: Ambulation with the use of a standard walker was shown to require 212% more oxygen per meter than unassisted ambulation and 104% more oxygen per meter than ambulation with a wheeled walker. Ambulation with a standard walker elicited 200% and 98% higher heart rate per meter as compared with unassisted ambulation and ambulation with a wheeled walker, respectively. No difference was detected for physiologic demands between unassisted ambulation and ambulation with a cane. CONCLUSION AND DISCUSSION: The decision to prescribe a wheeled walker versus a standard walker may be clinically important with patients who have impaired cardiorespiratory systems.

Aged↗

PubMed1995

Pegylated megakaryocyte growth and development factor abrogates the lethal thrombocytopenia associated with carboplatin and irradiation in mice.

Megakaryocyte growth and development factor (MGDF) is a potent inducer of megakaryopoiesis in vitro and thrombopoiesis in vivo. The effects of MGDF appear to be lineage-selective, making this cytokine an ideal candidate for use in alleviating clinically relevant thrombocytopenias. This report describes a murine model of life-threatening thrombocytopenia that results from the combination treatment of carboplatin and sublethal irradiation. Mortality of this regimen is 94% and is associated with widespread internal bleeding. The daily administration of pegylated recombinant human MGDF (PEG-rMGDF) significantly reduced mortality (to < 15%) and ameliorated the depth and duration of thrombocytopenia. The severity of leucopenia and anemia was also reduced, although it was not clear whether these effects were direct. Platelets generated in response to PEG-rMGDF were morphologically indistinguishable from normal platelets. PEG-rMGDF administered in combination with murine granulocyte colony-stimulating factor completely prevented mortality and further reduced leukopenia and thrombocytopenia. These data support the concept that PEG-rMGDF may be useful to treat iatrogenic thrombocytopenias.

Animals↗

PubMed1995

Inhibition of granulocyte-macrophage colony-stimulating factor (GM-CSF) activity by suramin and suramin analogues is correlated to interaction with the GM-CSF nucleotide-binding site.

Suramin and suramin analogues strongly inhibit both nucleotide interaction with the nucleotide-binding site of granulocyte-macrophage colony-stimulating factor (GM-CSF) and bioactivity of the molecule as assessed by competition photoaffinity labeling and cell proliferation assay, respectively. The half-maximal inhibition of cell proliferation by suramin occurs at 68 +/- 2.5 microM; three suramin analogues achieved comparable activity. The degree of competitive inhibition of nucleotide-binding by these compounds and the inhibition of GM-CSF bioactivity are correlated such that the compounds show similar rank-order by both of these methods. The strong interaction of suramin and related compounds with the nucleotide-binding site may mimic nucleotide-mediated inhibition of GM-CSF bioactivity and may be an important mechanism by which suramin acts as a pharmacological anti-growth factor agent.

Binding Sites↗

PubMed1995

Thrombocytopenia in dogs induced by granulocyte-macrophage colony-stimulating factor: increased destruction of circulating platelets.

Administration of recombinant canine granulocyte-macrophage colony-stimulating factor (rcGM-CSF) to normal dogs in previous studies induced an increase in peripheral blood neutrophils and a dose-dependent decrease in platelet counts. In six dogs that received the highest tested dose of rcGM-CSF (50 micrograms/kg/d) for a minimum of 12 days, the mean nadir of the platelet count was 46,000/microL (range, 4,000 to 91,000/microL) on day 9 +/- 1.1 after starting therapy, compared with a mean baseline platelet count of 398,000/microL (range, 240,000 to 555,000/microL). In three dogs, survival of autologous 111In-labeled platelets was reduced from a mean of 4.9 days to 1.3 days during the administration of rcGM-CSF. Biodistribution studies with gamma camera imaging indicated that there was an increase in mean hepatic uptake during the administration of rcGM-CSF, from 15% to 44% of the total injected 111In-labeled platelets at 2 hours, whereas splenic uptake was not significantly changed. In contrast, in two evaluable dogs who were recipients of 111In-labeled platelets from matched allogeneic donors receiving rcGM-CSF, platelet survival was not reduced and no increased hepatic uptake was noted. A third dog became alloimmunized to the matched donor platelets and was not evaluable. Immunohistologic studies of liver and spleen were performed with monoclonal antibodies specific for canine gpIIb/IIIa and P-selectin in dogs treated with rcGM-CSF and compared with untreated controls. On treatment, a marked reduction of platelets in the red pulp of the spleen was evident, and in general, the presence of platelet antigen in the liver was unchanged. Therefore, platelets were not being sequestered, but destroyed in the liver and spleen. The platelet antigens, P-selectin and gpIIb/IIIa, were identified in association with Kupffer cells in the liver, but no difference in the number of distribution of these Kupffer cells was found between controls and rcGM-CSF-treated dogs. In the spleen during rcGM-CSF treatment, most platelet antigens were associated with large mononuclear cells in the marginal zone. During administration of rcGM-CSF, CD1c and CD11c expression was increased on Kupffer cells. Platelet P-selectin expression and binding of leukocytes to circulating platelets were unchanged from baseline studies with rcGM-CSF treatment. In conclusion, during the administration of rcGM-CSF to dogs, a local process in the liver and spleen is induced resulting in thrombocytopenia.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1995

Effects of the obese gene product on body weight regulation in ob/ob mice.

C57BL/6J mice with a mutation in the obese (ob) gene are obese, diabetic, and exhibit reduced activity, metabolism, and body temperature. Daily intraperitoneal injection of these mice with recombinant OB protein lowered their body weight, percent body fat, food intake, and serum concentrations of glucose and insulin. In addition, metabolic rate, body temperature, and activity levels were increased by this treatment. None of these parameters was altered beyond the level observed in lean controls, suggesting that the OB protein normalized the metabolic status of the ob/ob mice. Lean animals injected with OB protein maintained a smaller weight loss throughout the 28-day study and showed no changes in any of the metabolic parameters. These data suggest that the OB protein regulates body weight and fat deposition through effects on metabolism and appetite.

Adipose Tissue↗

PubMed1995

Recombinant human megakaryocyte growth and development factor stimulates thrombocytopoiesis in normal nonhuman primates.

Megakaryocyte growth and development factor (MGDF) is a novel cytokine that binds to the c-mpl receptor and stimulates megakaryocyte development in vitro and in vivo. This report describes the ability of recombinant human (r-Hu) MGDF to affect megakaryocytopoiesis in normal nonhuman primates. r-HuMGDF was administered subcutaneously to normal, male rhesus monkeys once per day for 10 consecutive days at dosages of 2.5, 25, or 250 micrograms/kg of body weight. Bone marrow and peripheral blood were assayed for clonogenic activity and peripheral blood counts were monitored. Circulating platelet counts increased significantly (P < .05) for all doses within 6 days of r-HuMGDF administration and reached maximal levels between day 12 and day 14 postcytokine administration. The 2.5, 25.0, and 250.0 micrograms/kg/d doses elicited peak mean platelet counts that were 592%, 670%, and 449% of baseline, respectively. Bone marrow-derived clonogenic data showed significant increases in the concentration of megakaryocyte (MEG)-colony-forming unit (CFU) and granulocyte-erythroid-macrophage-megakaryocyte (GEMM)-CFU, whereas that of granulocyte-macrophage (GM)-CFU and burst-forming unit-erythroid (BFU-e) remained unchanged during the administration of r-HuMGDF. These data show that r-HuMGDF is a potent stimulator of thrombocytopoiesis in the normal nonhuman primate.

Animals↗

PubMed1995

Identification of N-terminus peptide of human granulocyte/macrophage colony stimulating factor as the site of nucleotide interaction.

The interaction of nucleotides with recombinant human granulocyte/macrophage colony stimulating factor (rhGM-CSF) has been investigated. Utilizing nucleotide photoaffinity probes [gamma 32P]-8N3ATP and [beta 32P]-8N3Ap4A, an analog of alarmone, the specificity of interaction was demonstrated by saturation of photoinsertion by these analogs and protection of photoinsertion by these analogs in the presence of natural nucleotide. The site of photoinsertion was tentatively determined to be Ser9. The photolabeled cytokine has lost most of its biological activity in a cellular proliferation assay, indicating a possible physiological role for this interaction.

Adenosine Triphosphate↗

PubMed1995

Influence of a 36-h fast on the central and peripheral components of VO2 during submaximal exercise and peak oxygen uptake.

To determine if a 36-h fast affects the central and peripheral components of VO2 during submaximal exercise and peak oxygen uptake (VO2 peak), 10 physically healthy males exercised first for 10 min at approximately 60% of their VO2 peak and then to fatigue. Their cardiovascular responses were compared with those of 10 physically healthy controls (males) who were individually matched by VO2 peak values. Fasting had no significant effect on the central (heart rate, HR; stroke volume, SV; cardiac output, Q) and peripheral (arteriovenous oxygen content difference, a-vO2 diff.) components of VO2 during submaximal exercise. The respiratory exchange ratio (RER) during submaximal exercise was significantly decreased in the fasted subjects. Based on the RER, fasting altered substrate utilization of the muscle and thereby increased the contribution of fat during submaximal exercise. Fasting had no significant effect on VO2 peak, HR or O2 pulse, but RER was significantly lower during incremental exercise to fatigue. These results demonstrate that VO2 during submaximal exercise in fasted subjects is achieved by similar central and peripheral responses as found in non-fasted subjects, and that fasting does not reduce VO2 peak.

Adult↗

PubMed1995

The effect of massage on oxygen consumption at rest.

This study determined the effect of massage on oxygen consumption at rest. Ten healthy, adult males (mean age = 28 years) volunteered to serve as subjects. During the Control Session, each subject was placed in the supine position on a massage table to remain motionless for 30 minutes. During the Treatment Session, each subject received a 30-minute sports massage of the lower extremities. Oxygen consumption was determined via the Beckman Metabolic Measurement Cart, which was upgraded to estimate cardiac output using the CO2 rebreathing (equilibrium) method. Paired t-tests were used for all tests of statistical significance. There was no significant difference in the subjects' oxygen consumption with the massage. Also, there were no significant differences in heart rate, stroke volume, cardiac output, and arteriovenous oxygen difference during the massage. These findings indicate (1) that massaging the lower extremities results in neither an increase nor a decrease in the subjects' expenditure of energy at rest and (2) that the energy cost of metabolism at rest is determined by the same central and/or peripheral adjustments.

Adult↗

PubMed1995

Effects of sexual intercourse on maximal aerobic power, oxygen pulse, and double product in male sedentary subjects.

It is well known that athletes in the United States are told to abstain from sexual intercourse prior to athletic competition. The rationale for such a policy appears to be related to the hypothesis that sexual intercourse decreases the athletes' ability to perform efficiently and/or maximally. But the effect that sexual intercourse may have on exercise performance has not been examined widely. Very likely, the restrictions placed on athletes have little to do with the athletes' physiological ability to substain a particular exercise intensity and/or perform maximally. The purpose of this study was to determine the effects of sexual intercourse 12 hours prior to maximal treadmill exercise on aerobic power, oxygen pulse, and double product (i.e., an index of relative cardiac work). Eleven male subjects were tested on the treadmill with and without prior sexual intercourse. The results from the maximal exercise tests showed that aerobic power, oxygen pulse, and double product were not different. Therefore, the data suggest that it is justified to dismiss the point of view that sexual intercourse decreases maximal exercise performance.

Adult↗

PubMed1994

Effect of recombinant canine granulocyte-macrophage colony-stimulating factor on hematopoietic recovery after otherwise lethal total body irradiation.

Recombinant canine granulocyte-macrophage colony-stimulating factor (rcGM-CSF) was studied in normal dogs and in dogs receiving otherwise lethal total body irradiation (TBI) without marrow transplant. Five normal dogs receiving 25 micrograms/kg of rcGM-CSF by subcutaneous (SC) injection twice daily (BID) for 14 days showed increases in peripheral blood neutrophil counts of three to five times the baseline. Platelet counts decreased during administration of rcGM-CSF to a mean nadir of 52,800. Ten dogs received 400 cGy TBI at 10 cGy/min from two opposing 60Co sources and no marrow graft. Within 2 hours of TBI, rcGM-CSF was begun at a dose of 50 micrograms/kg SC BID for 5 doses and then continued at 25 micrograms/kg SC BID for 21 days. Only 1 of the 10 dogs receiving rcGM-CSF survived with complete and sustained recovery of hematopoiesis. One of 13 historical control dogs survived after 400 cGy with no hematopoietic growth factor or marrow infusion. Results with rcGM-CSF were compared with previous and concurrent data with G-CSF studied in the same model. Of 10 dogs receiving G-CSF, 8 survived with complete and sustained hematopoietic recovery, a significantly better survival than that seen with rcGM-CSF (P = .006). Neutrophil counts were sustained at higher levels after TBI for the first 18 days in the G-CSF group (P < .016) and the neutrophil nadirs were higher. No differences in neutrophil nadirs were noted between the rcGM-CSF and control groups. Dogs treated with rcGM-CSF experienced a more rapid decline of platelet counts than G-CSF-treated or control dogs over the first 18 days (P < .001). The nadir of the platelet count was higher in the control group than in either the G-CSF or rcGM-CSF group and no significant difference was observed between the G-CSF and rcGM-CSF groups. After otherwise lethal TBI (400 cGy) in dogs, rcGM-CSF was not effective in promoting hematopoietic recovery or improving survival.

Animals↗

PubMed1994

Phase I trial of intravenous and intraperitoneal administration of granulocyte-macrophage colony-stimulating factor.

To assess the toxicity, pharmacokinetics, and local and systemic effects of the intraperitoneal (i.p.) administration of granulocyte-macrophage colony-stimulating factor (GM-CSF) at various dosages, 13 patients with predominantly i.p. malignancies refractory to standard chemotherapy were studied. GM-CSF was administered intravenously (i.v.) for 5 consecutive days; 21 days later the same dosage of GM-CSF was administered i.p. for 5 consecutive days. Four dosage levels were studied: 1, 2, 4, and 8 micrograms/kg/day. GM-CSF was well tolerated after i.v. and i.p. administration at doses up to 8 micrograms/kg/day. A transient fall followed by an elevation of circulating white cells was observed over a 24-h period after both i.v. and i.p. GM-CSF administration (mean minimum +/- SE as % baseline): 38 +/- 8% at 30 min after i.v. administration, 21 +/- 5% at 60 min after i.p. administration; mean maximum: 220 +/- 41% at 6 h after i.v. administration, 202 +/- 39% at 12 h after i.p. administration). The magnitude and time course of these changes were very similar for the two routes despite an up to 400-fold difference in serum GM-CSF levels at the same time points. Changes in leukocyte count and differential and neutrophil function were also similar over the 3-week period after both i.v. and i.p. administration. In the only patient who had i.p. GM-CSF levels assayed, i.p. administration achieved high levels of GM-CSF in peritoneal fluid (Cmax 343 ng/ml) with maintenance of high concentrations over 24 h (C24h 128 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1993

Crystallization and preliminary X-ray analysis of leukemia inhibitory factor.

Leukemia inhibitory factor (LIF) is a polyfunctional molecule with significant and diverse biological activities. LIF is a glycoprotein secreted by a number of different cell types in vitro. It is induced in fibroblasts, lymphocytes, monocytes and astrocytes by various inducers such as serum, TNF, interleukin-IP and EGF. Due to extensive and variable glycosylation the molecular weight can range from 38 to 67 kDA. The biological functions of LIF are mediated through a receptor and a signal transducer, gp130, which is also used by factors like interleukin-6 (IL-6), cilliary neurotropic factor (CNTF), and oncostatin M (OSM). Here, we report the crystallization of the non-glycosylated human-like LIF expressed in E. coli. The present crystals diffract to 2.0 A using synchrotron radiation. They belong to the monoclinic space group C2, and the cell dimensions are a = 61.5 A, b = 45.3 A, c = 77.7 A and beta = 112.3 degrees.

Cloning, Molecular↗