Biomedical subjects

J D Geiger

Publications and source records attributed to J D Geiger.

At least 73 records · Page 4Linked to original sources

PubMed1996

Regulation of ryanodine receptor calcium release channels by diadenosine polyphosphates.

[3H]Ryanodine binding to, as well as functions of, ryanodine receptor intracellular Ca2+ release channel complexes are modulated by several adenosine-based compounds. In this study, we determined the effects of endogenous compounds termed diadenosine polyphosphates (ApnAs; n = 2-6 phosphate groups) on [3Hlryanodine binding to membranes prepared from rat brain and skeletal and cardiac muscle. Under low ionic strength buffer conditions, [3H]ryanodine binding to brain membranes was significantly increased by 171% with 333 microM P1,P5-di(adenosine-5') pentaphosphate (Ap5A) and by 209% with the same concentration of the metabolism-resistant ATP analogue betagamma-methyleneadenosine 5'-triphosphate (AMP-PCP) compared with control values for [3H]ryanodine binding of 9.6 +/- 1.8 fmol/mg of protein. Dose-related increases in [3H]ryanodine binding were observed for all five ApnAs tested [P1,P2-di(adenosine-5') pyrophosphate (Ap2A), P1,P3-di(adenosine-5') triphosphate (Ap3A), P1,P4-di(adenosine-5') tetraphosphate (Ap4A), Ap5A, and P1,P6-di(adenosine-5') hexaphosphate (Ap6A)] as well as AMP-PCP; oxidized salts of ApnAs stimulated [3H]ryanodine binding to a greater degree than did nonoxidized APnAs. The apparent rank order for the capacity of these agents to increase [3H]-ryanodine binding was oxidized Ap4A = oxidized Ap5A > oxidized Ap3A > Ap6A > AMP-PCP > Ap5A > AP2A. Addition of the approximate EC50 dose of oxidized Ap4A (37 microM) increased the affinity (KD) of ryanodine receptors from 34 +/- 7 to 12 +/- 2 nM, the apparent binding site density (Bmax) was not significantly different from control values of 107 +/- 33 fmol/mg of protein. Increases in [3H]-ryanodine binding by either oxidized Ap4A or nonoxidized Ap5A were not further enhanced by coincubation with AMP-PCP, which suggests a similar site of action for the ApnAs and AMP-PCP. [3H]Ryanodine binding to skeletal and cardiac muscle membranes was enhanced by addition of oxidized AP4A Ap5A, and AMP-PCP. Oxidized AP4A increased the specific binding by ninefold in skeletal muscle and by threefold in cardiac muscle. These results suggest that ApnAs, at physiologically relevant concentrations, may serve as endogenous modulators of ryanodine receptor-gated Ca2+ release channels.

Adenosine Triphosphate↗

PubMed1996

Characterization of inhibitor-sensitive and -resistant adenosine transporters in cultured human fetal astrocytes.

The kinetic characteristics of [3H]adenosine uptake, the extent to which accumulated [3H]adenosine was metabolized, the effects such metabolism had on measurements of apparent Michaelis-Menten kinetic values of KT and Vmax, and the sensitivities with which nucleoside transport inhibitors blocked [3H] adenosine accumulations were determined in cultured human fetal astrocytes. KT and Vmax values for accumulations of [3H]-labeled purines using 15-s incubations in the absence of the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) and the adenosine kinase inhibitor 5'-iodotubercidin (ITU) were 6.2 microM and 0.15 nmol/min/mg of protein for the high-affinity and 2.6 mM and 21 nmol/min/mg of protein for the low-affinity components, respectively. In the presence of EHNA and ITU, where < 4% of accumulated [3H] adenosine was metabolized, transport per se was measured, and kinetic values for KT and Vmax were 179 microM and 5.2 nmol/min/mg of protein, respectively. In the absence of EHNA and ITU, accumulated [3H]adenosine was rapidly metabolized to AMP, ADP, and ATP, and caused an appearance of "concentrative" uptake in that the intracellular levels of [3H]-labeled purines (adenosine plus its metabolites) were 1.4-fold higher than in the medium. No apparent concentrative accumulations of [3H]adenosine were found when assays were conducted using short incubation times in the absence or presence of EHNA and ITU. The nucleoside transport inhibitors dipyridamole (DPR), nitrobenzylthioinosine (NBI), and dilazep biphasically inhibited [3H]-adenosine transport; for the inhibitor-sensitive components the IC50 values were 0.7 nM for NBI, 1.3 nM for DPR, and 3.3 nM for dilazep, and for the inhibitor-resistant component the IC50 values were 2.5 microM for NBI, 5.1 microM for dilazep, and 39.0 microM for DPR. These findings, in cultured human fetal astrocytes, represent the first demonstration of inhibitor-sensitive and -resistant adenosine transporters in nontransformed human cells.

Adenine↗

PubMed1996

Surgery for hepatoblastoma in children.

The survival of children with hepatoblastoma, the most common malignant tumor of the liver in children, has improved dramatically over the past 20 years. This progress has been made with advances in surgical technique and improved chemotherapy, primarily with adriamycin and cisplatin. Despite these advances, those patients who present with advanced disease have unsatisfactory survival. Continued refinement of liver surgery and chemotherapy, as well as the use of new techniques such as chemoembolization, immunotherapy, and molecular biology, should lead to improved survival in patients with advanced disease.

Antineoplastic Agents↗

PubMed1996

Identification of a human immunodeficiency virus type 1 Tat epitope that is neuroexcitatory and neurotoxic.

Tat is an 86- to 104-amino-acid viral protein that activates human immunodeficiency virus type 1 expression, modifies several cellular functions, and causes neurotoxicity. Here, we determined the extent to which peptide fragments of human immunodeficiency virus type 1 BRU Tat1-86 produced neurotoxicity, increased levels of intracellular calcium ([Ca2+]i), and affected neuronal excitability. Tat31-61 but not Tat48-85 dose dependently increased cytotoxicity and levels of [Ca2+]i in cultured human fetal brain cells. Similarly, Tat31-61 but not Tat48-85 depolarized rat hippocampal CA1 neurons in slices of rat brain. The neurotoxicity and increases in [Ca2+]i could be significantly inhibited by non-N-methyl-D-aspartate excitatory amino acid receptor antagonists. Shorter 15-mer peptides which overlapped by 10 amino acids each and which represented the entire sequence of Tat1-86 failed to produce any measurable neurotoxicity. Although it remains to be determined if Tat acts directly on neurons and/or indirectly via glial cells, these findings do suggest that Tat neurotoxicity is conformationally dependent, that the active site resides within the first exon of Tat between residues 31 to 61, and that these effects are mediated at least in part by excitatory amino acid receptors.

Animals↗

PubMed1996

Effects of iodotubercidin on adenosine kinase activity and nucleoside transport in DDT1 MF-2 smooth muscle cells.

Iodotubercidin is an adenosine kinase inhibitor that through its ability to increase levels of endogenous adenosine can enhance adenosine's receptor-mediated effects. We investigated whether iodotubercidin can inhibit [3H]adenosine accumulation by inhibiting transport processes in addition to inhibition of intracellular trapping of labeled adenine nucleotides. Under conditions in which extensive metabolism of intracellular adenosine was present, [3H]adenosine accumulation by DDT1 MF-2 cells was almost completely inhibited by iodotubercidin and the adenosine deaminase inhibitor erythro-9-(2-hydroxy-3-nonyl)-adenine or by the nucleoside transport inhibitor nitrobenzylthioinosine. By using similar conditions, [3H]adenosine accumulation was significantly greater in Na+ buffer than in buffer containing N-methyl-D-glucamine in place of Na+; however, this effect may be explained by an observed 40% inhibition of adenosine kinase activity by N-methyl-D-glucamine. By using uptake intervals of 14 sec to represent the transport component of uptake, iodotubercidin decreased the affinity for adenosine, by about 3-fold, but had no effect on maximum velocity of transport. That these effects of iodotubercidin were due to direct interactions with nucleoside transporters was supported by findings that iodotubercidin inhibited [3H]nitrobenzylthioinosine binding to nucleoside transporters with a Ki value of 4 microM and inhibited [3H]uridine and [3H]formycin B uptake with IC50 values of 7 and 15 microM, respectively. These data suggest that iodotubercidin, at pharmacologically relevant concentrations, inhibits nucleoside transport independently of its well characterized inhibition of adenosine kinase and that N-methyl-D-glucamine must be used with caution in experiments to determine the possible presence of Na+ gradient-dependent concentrative nucleoside transporters.

Adenosine Kinase↗

PubMed1996

Thyroid tumors in children.

Thyroid tumors in children continue to generate tremendous interest and controversy. Despite a frequent presentation with metastatic disease, these differentiated tumors have an excellent prognosis. Because most pediatric patients have prolonged survival, long-term follow-up is mandatory. The extent of surgery for this disease continues to be controversial. Genetic testing is changing dramatically the management of patients with multiple endocrine neoplasia and will lead to further understanding of this intriguing syndrome. Over the next decade, classification of differentiated thyroid cancers by means of more clinically useful biologic markers may affect the clinical management of children with thyroid cancer.

Adolescent↗

PubMed1995

Enhancement of NMDA-induced increases in levels of endogenous adenosine by adenosine deaminase and adenosine transport inhibition in rat striatum.

Unilateral microinjection of N-methyl-D-aspartate (NMDA) into striatum of rats subsequently killed by high-energy focused microwave irradiation significantly increased in vivo levels of endogenous adenosine. At a dose of 25 nmol NMDA, levels of adenosine in injected striata were 263% of levels in uninjected contralateral striata. An inhibitor of adenosine deaminase (deoxycoformycin, DCF) in combination with an inhibitor of adenosine transport (dilazep, DLZP) at a dose that did not affect levels of endogenous adenosine, potentiated NMDA-induced increases in adenosine levels to 426% of contralateral striata. In the presence of DCF and DLZP, NMDA dose-dependently increased levels of adenosine (% of contralateral striatum) from 166% at 10 nmol to 622% at 100 nmol. NMDA-induced increases in levels of endogenous adenosine were completely blocked by prior administration of the NMDA receptor antagonist MK 801 (dizocilpine). Inhibitors of adenosine metabolism and transport may provide therapeutic benefit by potentiating excitatory amino acid-induced increases in levels of endogenous adenosine in vivo.

Adenosine↗

PubMed1995

HIV-1 coat protein gp120-induced increases in levels of intrasynaptosomal calcium.

The effects of human immunodeficiency virus type-1 (HIV-1) coat protein gp120 on levels of intrasynaptosomal calcium ([Ca2+]i) were determined in rat cortical synaptosomes. gp120 at concentrations of > or = 400 pM, significantly (P < 0.05) increased levels of [Ca2+]i. Treatment with 20 mM KCl, reduced the concentrations of gp120 necessary to produce significant (P < 0.001) increases in [Ca2+]i. gp120-evoked increases in [Ca2+]i were prevented either by treatment with dantrolene or by removal of extracellular calcium with BAPTA. The peak levels of gp120-induced increases in [Ca2+]i were not affected by calcium channel blockers lanthanum and nicardipine, by glutamate receptor antagonists MK-801 and NBQX, or by removal of endogenous glutamate with glutamate dehydrogenase. gp120-induced [Ca2+]i increases in presynaptic terminals may play a role in HIV-mediated effects in the central nervous system.

Animals↗

PubMed1995

Human immunodeficiency virus type 1 tat activates non-N-methyl-D-aspartate excitatory amino acid receptors and causes neurotoxicity.

The human immunodeficiency virus type 1 (HIV-1) protein Tat is known to be released from HIV-1-infected cells. We show that micromolar concentrations of Tat depolarized young rat and adult human neurons. In addition, Tat, at similar concentrations, was toxic to human fetal neurons in culture. Tat-induced responses were insensitive to the Na+ channel blocker tetrodotoxin, suggesting a direct effect of Tat on neurons. Tat-induced depolarizations and cytotoxicity were blocked by the excitatory amino acid antagonist kynurenate. The N-methyl-D-aspartate receptor antagonist D-2-amino-5-phosphonovalerate had little effect on Tat-induced depolarizations but did provide protection from Tat neurotoxicity. These results suggest that Tat, released from HIV-1-infected cells, may be an important mediator of neurotoxicity observed in HIV-1 encephalopathy.

2-Amino-5-phosphonovalerate↗

PubMed1995

Involvement of bidirectional adenosine transporters in the release of L-[3H]adenosine from rat brain synaptosomal preparations.

Adenosine transport inhibitors as enhancers of extracellular levels of endogenous adenosine would, presumably, only be effective if, for example, (1) the inhibitors block influx to a greater degree than efflux (release) of intracellular adenosine or (2) the inhibitors block equally well the influx and efflux of adenosine, but significant amounts of adenosine are formed as a result of dephosphorylation of released adenine nucleotides. Limited information is available regarding the directional symmetry of adenosine transporters in neural cells. Using rat brain crude P2 synaptosomal preparations preloaded with L-[3H]adenosine, our objectives here were to determine (1) if L-[3H]adenosine, a substrate for adenosine transporters that is more metabolically stable than physiological D-adenosine, was being released from synaptosomal preparations, (2) the optimal conditions necessary to observe the release, and (3) the degree to which this release was mediated by efflux through bidirectional nucleoside transporters. L-[3H]Adenosine release was found to be concentration and time dependent, temperature sensitive, and linear with synaptosomal protein. L-[3H]Adenosine release was inhibited dose-dependently by dipyridamole, nitrobenzylthioinosine, and dilazep; at concentrations of 100 microM inhibition was at least 40% for dipyridamole, 52% for nitrobenzylthioinosine, and 49% for dilazep. After loading with L-[3H]adenosine alone or I-[3H]adenosine plus unlabeled L-adenosine, D-adenosine, or uridine, L-[3H]adenosine release was inhibited 42% by L-adenosine, 69% by uridine, and 81% by D-adenosine.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine↗

PubMed1995

A phase II trial of interleukin-2 and interferon-alpha in the treatment of metastatic colorectal carcinoma.

A total of 29 patients with stage IV colorectal cancer were entered into a phase II trial of bolus interleukin-2 (IL-2) and interferon-alpha (IFN alpha) (3 x 10(6) U/m2 of each cytokine given i.v. q8h x 15 doses and repeated in 2 weeks). Immunologic parameters measured on isolated peripheral blood lymphocytes revealed increased activated T cells with upregulated natural killer and lymphokine-activated killer activity. Among 24 evaluable patients, there were 4 partial responses (17%) of short duration ( < or = 6 months). Three of the responding patients had been refractory to prior chemotherapy. Overall median survival in the 24 evaluable patients was 18.5 months. Therapy necessitated an inpatient setting, with the most common toxicities being hypotension, hepatic insufficiency, fever, hypocalcemia, nausea/vomiting, and renal insufficiency. There were two treatment-related deaths. Because neither IL-2 nor IFN alpha alone has significant activity against colorectal cancer, the responses observed in this study suggest a potential synergistic effect between the two cytokines. However, the toxicity and short duration of response without survival benefit do not support the routine use of this regimen as a therapeutic modality for this tumor histology.

Adult↗

PubMed1994

Effects of diadenosine polyphosphates on sodium nitroprusside-induced soluble guanylate cyclase activity in rat cerebellum.

Diadenosine polyphosphates (ApnAs,n = 1-6 phosphates) have been shown to interact with heme containing proteins and in the present study we determined for guanylate cyclase (GC), a heme containing enzyme whose activity is modulated by adenine nucleotides and nitric oxide, the degree to which ApnAs affect sodium nitroprusside (SNP, a nitric oxide donor)-induced GC activity using cytosolic fractions of rat cerebellum and radioimmunoassays for cGMP. SNP-stimulated GC activity was inhibited by up to 60% by ApnAs; rank order of potency was Ap6A > Ap5A > Ap4A > Ap3A = Ap2A. At concentrations ranging from 0.01 to 100 microM, only Ap6A potentiated in a concentration-dependent manner (apparent EC50 value 1 microM), by up to two-fold, SNP-induced GC activity. The effects of Ap6A on GC activity were neither to changes in SNP degradation nor due to metabolites of Ap6A. These results suggest that ApnAs through its effects on GC activity may play a role in regulating a variety of CNS functions.

Adenine Nucleotides↗

PubMed1994

Ionic strength dependence of calcium, adenine nucleotide, magnesium, and caffeine actions on ryanodine receptors in rat brain.

[3H]Ryanodine binding studies of ryanodine receptors in brain membrane preparations typically require the presence of high salt concentrations in assay incubations to yield optimal levels of binding. Here, radioligand binding measurements on rat cerebral cortical tissues were conducted under high (1.0 M KCl) and low (200 mM KCl) salt buffer conditions to determine the effects of ionic strength on receptor binding properties as well as on modulation of ligand binding by Ca2+, Mg2+, beta, gamma-methylene-adenosine 5'-triphosphate (AMP-PCP), and caffeine. In 1.0 M KCl buffer, labeled titration/equilibrium analyses yielded two classes of binding sites with apparent KD (nM) and Bmax (fmol/mg of protein) values of 2.4 and 34, respectively, for the high-affinity site and 19.9 and 157, respectively, for the low-affinity site. Unlabeled titration/equilibrium measurements gave a single high-affinity site with a KD value of 1.9 nM and a Bmax value of 95 fmol/mg of protein. The apparent KD value derived from association and dissociation studies was 20 pM. Equilibrium binding was activated by Ca2+ (KD/Ca2+ = 14 nM), inhibited by Mg2+ (IC50 = 5.0 mM), and unaffected by AMP-PCP or caffeine. In 200 mM KCl buffer conditions, labeled titration analyses gave only a single site with a KD value similar to and a Bmax value 1.8-fold greater than those obtained for the low-affinity site in 1.0 M KCl buffer. In unlabeled titration measurements, the KD value was fivefold lower, whereas the Bmax value was unaffected. The KD value derived from association and dissociation analysis was 2.4-fold greater in 200 mM KCl compared with 1.0 M KCl buffer conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗

PubMed1994

Characterization of a novel binding site for the human immunodeficiency virus type 1 envelope protein gp120 on human fetal astrocytes.

125I-labeled recombinant gp120 bound to primary cultures of human fetal astrocytes with a single class of 260-kDa binding sites, with a Kd of 26 nM and maximal number of binding sites of 29.9 fmol/4 x 10(4) cells. Neither CD4 nor galactocerebroside was detectable on astrocytes, and 125I-labeled recombinant gp120 binding to astrocytes was not blocked by antibodies against galactocerebroside or the gp120 binding domain of CD4.

Antibodies, Monoclonal↗

PubMed1994

Chlorotyrosine exerts renal effects and antagonizes renal and gastric responses to atrial natriuretic peptide.

3-Chloro-L-tyrosine (3CT) is an inhibitor of tyrosine hydroxylase, the rate-limiting enzyme for catecholamine synthesis. In vivo inhibition of tyrosine hydroxylase results in lower catecholamine levels. 3CT (0.5 mg/kg), administered as a bolus i.v. to anesthetized uninephrectomized rats, elicited increases of 72% and 44% in urinary sodium concentration and volume, respectively, whereas a dose of 1 mg/kg caused increases of 27% and 29%. 3CT, 1 mg/kg, resulted in a 2-fold increase in plasma aldosterone (ALD); 0.5 mg/kg was without significant effect. At a dose of 1 mg/kg 3CT significantly antagonized the renal effects of atrial natriuretic peptide (ANP) (1.5 micrograms kg-1 min-1 by intrarenal infusion), expressed as an enhanced excretion of urine volume (102 +/- 14 vs. 70 +/- 11 microliters/min) and sodium (16.1 +/- 1.8 vs. 11.5 +/- 1.7 microEq/min) and increased osmolar clearance (171 +/- 12 vs. 144 +/- 13 microliters/min). A dose of 0.5 mg/kg of 3CT did not produce these same responses to ANP. The increased urine flow caused by 3CT may reflect reduced norepinephrine synthesis. The inverse dose-effect relationship of 3CT on urine flow rate may result from concomitant depletion of dopamine (DA) and elevated circulating ALD. The antagonism of 3CT on responses to ANP is not at the receptor level, because 3CT did not compete for [125I] ANP binding or inhibit ANP-stimulated guanylate cyclase in kidney cell membranes. It was proposed that the reduced basal sympathetic and renal DA tone, together with the elevated ALD level, account for this antagonism.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Failure of 2'-deoxycoformycin to protect against transient forebrain ischemia in rat.

We determined whether 2'-deoxycoformycin (DCF), a potent highly specific inhibitor of adenosine deaminase (ADA), protected against transient forebrain ischemic neuronal injury in rat. Anesthetized male Sprague-Dawley rats received i.p. injections of either saline, 0.5 mg/kg or 5 mg/kg DCF 2 h before undergoing a 10-min forebrain ischemic insult induced by bilateral carotid artery occlusion with concomitant hypotension. Rat brain sections taken 7 days post-ischemia showed damage mostly in the CA1 region of the hippocampus. Quantification of neuronal injury showed no significant differences between saline- or DCF-treated rats. These results indicate that, contrary to previous reports, DCF does not protect against the neuronal damage that follows forebrain ischemia in rat.

Animals↗

PubMed1993

Adoptive cellular therapy of malignancy.

The realization that human cancers can be responsive to the manipulation of the immune system has only recently been documented. The immune approaches to the treatment of malignancy can be broadly classified into either active or passive immunotherapies. With active immunotherapy, treatment relies on the in vivo stimulation of the endogenous host immune system to react against tumors with the administration of biological response-modifying agents (ie, bacterial adjuvants, cytokines, tumor vaccines). With passive immunotherapy, treatment involves the delivery of biologic reagents with established tumor-immune reactivity (ie, antibodies or cells) that can directly or indirectly mediate antitumor effects and does not necessarily depend on an intact host immune system. Cellular therapy of malignancy has become more feasible with increased understanding of the interactions between immune cells and tumors. This article will review our current understanding of the principles underlying these interactions.

Adjuvants, Immunologic↗

PubMed1993

Generation of T-cells reactive to the poorly immunogenic B16-BL6 melanoma with efficacy in the treatment of spontaneous metastases.

The B16-BL6 (BL6) melanoma is a poorly immunogenic murine tumor that is highly invasive and spontaneously metastasizes from the primary site. Utilizing an established anti-CD3/interleukin-2 (IL-2) culture procedure, we have previously reported that lymph nodes (LNs) draining immunogenic murine sarcomas contained preeffector cells that could be activated to differentiate into therapeutic effector cells for adoptive immunotherapy. By contrast, LNs draining the poorly immunogenic BL6 melanoma were found not to be a reliable source of preeffector cells. Instead, sensitization of preeffector cells reactive to BL6 required the subcutaneous inoculation of tumor admixed with Corynebacterium parvum. LN cells draining these vaccination sites demonstrated therapeutic efficacy only after subsequent anti-CD3/IL-2 activation. The sensitization of preeffector cells was dependent on the presence of tumor antigen and an optimal dose of C. parvum (< or = 50 micrograms). Furthermore, kinetic analysis revealed that the preeffector response was transient after tumor vaccination. The therapeutic efficacy of anti-CD3/IL-2 activated LN cells was further evaluated in the treatment of spontaneous macroscopic BL6 visceral metastases. Spontaneous visceral metastases were induced in animals by inoculation with BL6 tumor in the footpad followed by amputation of the primary tumor 3 weeks later. The systemic transfer of 10(8) anti-CD3/IL-2 activated T-cells and the concomitant intraperitoneal administration of subtherapeutic doses of IL-2 1 week after amputation cured 50% of the animals and prolonged median survival time (MST) to > 140 days. All mice except one that received no treatment or was treated with IL-2 alone succumbed to visceral metastases with an MST of approximately 23 days. This study characterizes a model whereby the weak immune response to the BL6 melanoma can be positively or negatively modulated for the generation of antitumor reactive T-cells useful in adoptive immunotherapy.

Animals↗