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J W Wells

Publications and source records attributed to J W Wells.

At least 19 recordsLinked to original sources
PubMed2006

Disentangling surface, bulk, and space-charge-layer conductivity in Si(111)-(7 x 7).

A novel approach for extracting genuine surface conductivities is presented and illustrated using the unresolved example of Si(111)-(7 x 7). Its temperature-dependent conductivity was measured with a microscopic four point probe between room temperature and 100 K. At room temperature the measured conductance corresponds to that expected from the bulk doping level. However, as the temperatures is lowered below approximately 200 K, the conductance decreases by several orders of magnitude in a small temperature range and it saturates at a low temperature value of approximately 4 x 10(-8) Omega(-1), irrespective of bulk doping. This abrupt transition is interpreted as the switching from bulk to surface conduction, an interpretation which is supported by a numerical model for the measured four point probe conductance. The value of the surface conductance is considerably lower than that of a good metal.

Journal Article↗
PubMed2005

Influence of interleukin-4 on the phenotype and function of bone marrow-derived murine dendritic cells generated under serum-free conditions.

Murine bone marrow-derived dendritic cells (DC) can be generated by culture in the presence of granulocyte/macrophage colony-stimulating factor (GM-CSF) alone or GM-CSF in conjunction with interleukin-4 (IL-4). However, these two culture methods result in the production of heterogeneous DC populations with distinct phenotypic and stimulatory properties. In this study, we investigated the properties of DC generated under serum-free conditions in the presence or absence of IL-4 and compared their yield and phenotype to that of DC generated in the presence of fetal calf serum (FCS) (+/-IL-4). We did not observe a significant difference in the total cell yield between these four culture conditions, although the proportion of CD11c+ DC in cultures that received FCS was higher than that of their counterparts generated under serum-free conditions. Also, the four culture conditions generated CD11c+ DC with comparable levels of major histocompatibility complex (MHC) class II, CD40, CD80 and CD86 expression, with the exception of cells cultured under serum-free conditions in the absence of IL-4, which displayed suboptimal levels of these markers. Moreover, we compared the functional and stimulatory properties of DC generated under serum-free conditions in the presence or absence of IL-4. DC cultured in the presence of IL-4 were stronger stimulators of allogeneic splenocytes in a primary mixed lymphocyte reaction (MLR) and of naive antigen-specific OT-II transgenic T cells when pulsed with the class II ovalbumin (OVA)323-339 peptide or whole OVA protein than DC cultured in the absence of IL-4. However, both DC populations displayed a similar capacity to take up fluorescein isothiocyanate (FITC)-albumin by macropinocytosis and FITC-Dextran by the mannose receptor and to secrete IL-12 in response to stimulation with lipopolysaccharide (LPS) or an agonistic anti-CD40 monoclonal antibody. Therefore, we conclude that although both DC culture methods result in the production of DC with similar functional abilities, under serum-free conditions, DC cultured in GM-CSF and IL-4 show an increased stimulatory potential over DC cultured in GM-CSF alone. This is an important consideration in the design of experiments where DC are being exploited as immunotherapeutic vaccines.

Animals↗
PubMed2001

Apparent capacity of cardiac muscarinic receptors for different radiolabeled antagonists.

Muscarinic receptors in sarcolemmal membranes, digitonin-solubilized extracts, and purified preparations from porcine atria have revealed a shortfall in the apparent capacity for N-[3H]methylscopolamine, which was only about 75% of that for [3H]quinuclidinylbenzilate. Since binding at near-saturating concentrations of [3H]quinuclidinylbenzilate was inhibited fully at comparatively low concentrations of unlabeled N-methylscopolamine, the data are inconsistent with the notion that [3H]quinuclidinylbenzilate binds selectively to a subclass of distinct, non-interconverting, and mutually independent sites. The discrepancy is resolved by adjusting the specific activity of N-[3H]methylscopolamine to account for unlabeled scopolamine that was identified in some batches of the radioligand. Also, there was no shortfall in capacity when N-[3H]methylscopolamine was devoid of scopolamine, and the predicted effect was obtained when pure N-[3H]methylscopolamine was supplemented with known amounts of scopolamine. A small discrepancy in the levels of scopolamine estimated pharmacologically and by mass spectrometry can be attributed largely to a difference in the efficiency of ionization between scopolamine and N-methylscopolamine. Different capacities for different radioligands are not uncommon with muscarinic and other G protein-coupled receptors, and in some cases the effect may have been due wholly or in part to an unlabeled impurity. Binding data can be mechanistically ambiguous, particularly when acquired only at graded concentrations of the radioligand. The predicted effects of an unlabeled impurity mimic or resemble those of alternative scenarios such as sequestration behind a hydrophobic barrier, a nucleotide-regulated interconversion from one state of affinity to another, and cooperativity between interacting sites.

Animals↗
PubMed2001

Nature of the oligomers formed by muscarinic m2 acetylcholine receptors in Sf9 cells.

Wild-type, FLAG-tagged, and c-myc-tagged muscarinic m2 receptors extracted in digitonin-cholate from singly and co-infected Sf9 (Spodoptera frugiperda) cells were indistinguishable in their binding of [3H]quinuclidinylbenzilate, either before or after purification. The FLAG epitope was found to coimmunoprecipitate with the c-myc epitope when co-infected cells were solubilised in digitonin-cholate, n-dodecyl-beta-D-maltoside or Lubrol-PX. The degree of coprecipitation in digitonin-cholate was unaffected by preincubation of the extract for up to 60 min at 30 degrees C, with or without muscarinic receptor ligands; no coimmunoprecipitation occurred in mixed extracts from singly infected cells. As measured by [3H]quinuclidinylbenzilate, the efficiency of immunoprecipitation from co-infected cells was 87% of that from singly infected cells. The amount of receptor immunoprecipitated from the latter, as determined by densitometry, was 2.3-fold that expected from the loss of binding from the extract. The data suggest that at least some of the receptors were trimeric or larger and that oligomers neither formed nor dissociated under the conditions of the experiments. Also, some receptors appear to be non-functional or latent in digitonin-solubilised extracts.

Animals↗
PubMed1997

Constitutive activation of the m5 muscarinic receptor by a series of mutations at the extracellular end of transmembrane 6.

The m5 muscarinic acetylcholine receptor was constitutively activated by a wide range of amino acid substitutions at a residue (serine 465) that is positioned at the junction of the sixth transmembrane domain and the extracellular loop. Of 13 substitutions tested, 11 produced significant increases in constitutive activity. Replacement of serine 465 with large (phenylalanine and valine) or basic residues (arginine and lysine) increased the constitutive activity of the receptor to between 55 and 110% of the maximum response of the wild-type receptor to the agonist carbachol. Other substitutions (e.g., cysteine and leucine) increased the constitutive activity to an intermediate level (30%), while small and acidic residues (glycine, aspartate, and glutamate) caused small or insignificant increases. The increase in the constitutive activity of each mutant receptor correlated with an increase in the potency of carbachol in both binding and functional assays, with the most constitutively activated receptors showing a 40-fold decrease in the EC50 of carbachol. The negative antagonist atropine bound to and reversed the constitutive activity of all mutant receptors with equal potency. These data were fitted to a two-state model of receptor function. The data are consistent with the primary effect of substitutions to serine 465 being to selectively destabilize the inactive state of the receptor, thus favoring formation of the active state in the absence of agonists. Our data strongly support this two-state model of receptor function and identify a critical role of this domain in the activation of muscarinic receptors.

3T3 Cells↗
PubMed1997

Cardiac muscarinic receptors. Cooperativity as the basis for multiple states of affinity.

Cooperativity has been investigated as the mechanistic basis for effects observed with cardiac muscarinic receptors in washed membranes from Syrian hamsters. Specifically, N-[3H]methylscopolamine labeled only 66-75% of the sites labeled by [3H]quinuclidinylbenzilate at apparently saturating concentrations of each radioligand. Also, receptors labeled by N-[3H]methylscopolamine revealed three states of affinity for agonists, both in native membranes and following irreversible blockade of about 80% of the sites by propylbenzilylcholine mustard; in both preparations, guanylylimidodiphosphate (GMP-PNP) effected an apparent interconversion of sites from higher to lower affinity for agonists and from lower to higher affinity for the antagonist. Excellent and mechanistically consistent descriptions of the data were obtained in terms of a model comprising cooperative and noncooperative forms of the receptor; the former was described by a variant of the Adair equation, and the latter was included to account for low-affinity sites that survived treatment with the mustard. If differences in apparent capacity derive from negative cooperativity in the binding of N-[3H]methylscopolamine, the cooperative form of the receptor was at least trivalent in native membranes; otherwise, constraints imposed by the effects of GMP-PNP at the concentrations of radioligand used in the assays dictate that the cooperative form of the receptor was at least tetravalent. In contrast, a divalent receptor is sufficient with the data from alkylated membranes, in accord with the reduced likelihood of interactions between functional sites within an oligomeric array. A model is presented wherein the receptor interconverts spontaneously between two or more states differing in their cooperative properties. The effects of GMP-PNP can be rationalized as a shift in the equilibrium between the different states.

Alkylation↗
PubMed1997

Cardiac muscarinic receptors. Relationship between the G protein and multiple states of affinity.

An expanded version of the mobile receptor model has been assessed in studies on the binding of N-[3H]methylscopolamine and [35S]GTPgammaS to cardiac muscarinic receptors and their attendant G proteins in ventricular membranes from hamster. The model comprises two pools of receptor, one of which lacks G proteins, and a heterogeneous population of G proteins that compete for the receptor within the G protein-containing pool. To guide the formulation of the model itself and to define the various parameters, data were combined from assays performed under various conditions with native membranes and following irreversible blockade of about 80% of the receptors with propylbenzilylcholine mustard. Multiple G proteins are indicated primarily by multiple states of affinity evident in the dose-dependent effect of guanyl nucleotides on the binding of carbachol; G protein-free receptors are indicated by sites of low affinity for carbachol that survive treatment with the mustard. The expanded model generally succeeds where more frugal schemes have been inadequate, but it nevertheless fails to yield a mechanistically consistent description of the data. Guanyl nucleotides and partial alkylation do not affect the inhibitory potency of carbachol in a manner consistent with their supposed effect on the equilibrium between uncoupled and G protein-coupled receptors. As inferred from the model, G proteins are lost upon alkylation of the receptor, and their numbers are regulated by guanyl nucleotides. Parameters estimated via N-[3H]methylscopolamine are wholly inconsistent with the same parameters estimated via [35S]GTPgammaS. The failure of the model suggests that multiple states of affinity may not arise from a ligand-regulated equilibrium between free receptors and G proteins on the one hand and one or more RG complexes on the other.

Animals↗
PubMed1997

Evaluation of the temporal evolution of acute spinal cord injury.

RATIONALE AND OBJECTIVES: After receiving a controlled injury to the thoracic cord, five rats were examined on a 1.5-T magnetic resonance (MR) imaging system at regular intervals over 1 month to assess evolution of the injury. METHODS: After the rats received pentobarbital anesthesia, a T10 laminectomy was performed on them, which exposed the dura over the dorsal surface of the spinal cord. With the animal placed in a New York University weight-drop device, a 10-g rod with a flat brass tip was dropped (free-fall) from a height of 50 mm to impact the cord. After injury, the incision was closed with suture material. Each animal was imaged on the day of injury, and at 7, 14, and 28 days after injury. Before contrast injection was administered, sagittal sections were obtained with T2 fast-spin echo and T1-spin echo technique. Each rat then received 0.3-mmol/kg gadoteridol (Gd HP-DO3A or ProHance) intravenously, with the T1 scan repeated. At 28 days, the animals were killed, and the cord was fixed and embedded in paraffin for histologic evaluation. RESULTS: The intensity of cord enhancement in the region of injury, after intravenous (i.v.) contrast injection, was at a maximum on the day of injury, and it decreased in a steady fashion thereafter. The intensity was 11.7 +/- 0.6 on the day of injury, 9.7 +/- 2.6 on day 7, 6.3 +/- 5.3 on day 14, and 0.0 +/- 2.3 on day 28. The results on day 0 and 7 were statistically significant in terms of a difference from that on day 28, with a P value < 0.001. The length of cord injury, assessed postcontrast, also decreased in a steady fashion from the day of injury. The length of injury (in cm) was 1.1 +/- 0.1 on the day of injury, 0.5 +/- 0.2 on day 7, 0.3 +/- 0.1 on day 14, and 0.1 +/- 0.1 on day 28. The results on day 0 and 14 were statistically significant in terms of a difference from those at the next time point, with P values from < 0.01 to < 0.001. Visually, on T2 images, substantial edema was noted on day 0, with progression to focal cord atrophy and gliosis by day 28. CONCLUSIONS: Acute spinal cord injury in a rat model is well visualized on pre- and postcontrast MR scans at 1.5 T. Observation of T2 changes and disruption of the blood-spinal cord barrier provide markers for temporal assessment of spinal cord injury in the rat model.

Acute Disease↗
PubMed1997

Renal metastasis. Detection and characterization on enhanced magnetic resonance imaging using an animal model.

RATIONALE AND OBJECTIVES: A new animal model was developed in rabbits (renal metastasis using the VX-2 tumor line), and magnetic resonance (MR) imaging performed before and after intravenous gadoteridol injection to assess lesion conspicuity and characterization. METHODS: Six New Zealand White rabbits with renal metastases were studied on a 1.5-tesla Siemens Vision MR unit. Iodinated contrast was given intravenously to the rabbits, before implantation, to visualize the kidneys under fluoroscopy. Using a 5/8-inch 25-gauge needle, 0.1 mL of minced, screened VX-2 tumor was injected percutaneously into each kidney at the corticomedullary junction. Animals were studied on day 7 after implantation. Baseline fast low-angle shot (FLASH) T1-weighted and fast spin echo T2-weighted breathhold scans were first obtained. Then, an additional precontrast turbo-FLASH T1-weighted scan was acquired. After these scans, 0.1 mmol/kg gadoteridol (gadolinium HP-D03A; ProHance) was injected intravenously at a rate of 1.5 mL/second. Dynamic breathhold turbo-FLASH T1 images were then obtained at 0, 6, 12, 19, 25, and 31 seconds after injection and at 1, 2, and 10 minutes after injection. The T1 multislice FLASH two-dimensional scan was repeated at 10 minutes after contrast. Imaging results were analyzed by region of interest measurement and correlated with tissue pathology. RESULTS: On dynamic T1-weighted turbo-FLASH scans, lesion conspicuity, specifically (SI(kidney)-SI(tumor)/noise), increased from 7 +/- 7 signal intensity precontrast to a maximum of 14 +/- 8 at 1 minute after contrast. This increase was statistically significant (P = 0.002). An initial rapid increase in tumor conspicuity occurred within the first 30 seconds after contrast, with the curve flattening thereafter. Lesion conspicuity on the precontrast T2-weighted scans was 9 +/- 10, not statistically different from results with either of the precontrast T1-weighted scan techniques. Using T1-weighted FLASH technique, lesion conspicuity increased from 10 +/- 5 precontrast to 31 +/- 12 postcontrast. As with turbo-FLASH, the improvement in tumor conspicuity after contrast on FLASH scans was statistically significant (P = 0.02). The difference between postcontrast FLASH scans and precontrast T2-weighted scans also was statistically significant, with postcontrast scans superior for lesion conspicuity (P = 0.01). Each tumor was confirmed on pathologic exam. CONCLUSIONS: This research establishes a model of metastasis to the kidney for use in imaging studies. The conspicuity of a small renal metastasis is shown to be improved on early dynamic imaging, as well as at 10 minutes after intravenous injection of 0.1 mmol/kg gadoteridol. Observation of dynamic signal intensity changes provides additional information regarding lesion characterization, supplementing that from precontrast scans.

Animals↗
PubMed1996

Evaluation of gadolinium 2,5-BPA-DO3A, a new macrocyclic hepatobiliary chelate, in normal liver and metastatic disease on high field magnetic resonance imaging.

RATIONALE AND OBJECTIVES: A new hepatobiliary gadolinium chelate, gadolinium (Gd) 2,5-BPA-DO3A, was compared in two animal species with Gd HP-DO3A (gadoteridol), a clinically approved extracellular contrast agent, and Gd Cy2-DOTA, a second hepatobiliary chelate in preclinical development. The ligand in Gd 2,5-BPA-DO3A is macrocyclic in nature, as opposed to the linear structure of Gd DTPA. Gadolinium 2,5-BPA-DO3A was evaluated on magnetic resonance imaging at 1.5 T, examining specifically liver parenchymal enhancement and lesion delineation, the latter in metastatic disease. METHODS: Gadolinium 2,5-BPA-DO3A was evaluated in five normal rhesus monkeys and four New Zealand White rabbits with implanted VX-2 liver tumors. These studies were compared with magnetic resonance exams in the same animals using Gd HP-DO3A and Gd Cy2-DOTA. A contrast dose of 0.1 mmol/kg intravenous was employed in each instance, with the sequence of administration (for the three agents) randomized and at least 72 hours between injections. Spin echo breathhold T1-weighted scans were obtained before and at multiple times after contrast administration. Postcontrast scans were acquired from 1 to 60 minutes after injection in the monkeys and from 1 to 240 minutes in the rabbits. RESULTS: Enhancement of normal liver parenchyma was markedly superior with Gd 2,5-BPA-DO3A compared with Gd HP-DO3A and Gd Cy2-DOTA in both monkeys and rabbits. At 2 and 60 minutes after contrast administration, the liver signal intensity in the monkey was 452 +/- 56 and 440 +/- 69 with Gd 2,5-BPA-DO3A compared with 295 +/- 34 and 256 +/- 38 with Gd HP-DO3A. The difference between agents was statistically significant at all postcontrast time points in the rhesus monkey. Excretion of contrast into the gall bladder was consistently observed after Gd 2,5-BPA-DO3A injection in both animal species. Maximum lesion conspicuity occurred in the rabbit at 45 minutes after Gd 2,5-BPA-DO3A administration. At 45 minutes postinjection, liver-lesion contrast was 0.60 +/- 0.15 with Gd 2,5-BPA-DO3A, 0.35 +/- 0.11 with Gd Cy2-DOTA, and 0.12 +/- 0.04 with Gd HP-DO3A, with the differences between agents being statistically significant. CONCLUSIONS: Gadolinium 2,5-BPA-DO3A is superior to both Gd Cy2-DOTA and Gd HP-DO3A in the degree of enhancement of normal liver parenchyma achieved after intravenous injection. This leads to improved liver lesion delineation with Gd 2,5-BPA-DO3A on delayed postcontrast magnetic resonance scans.

Animals↗
PubMed1996

The use of gadolinium-BOPTA on magnetic resonance imaging in brain infection.

RATIONALE AND OBJECTIVES: The use of gadolinium (Gd)-BOPTA as a magnetic resonance contrast agent for central nervous system disease was studied in a canine brain abscess model. METHODS: A Streptococcus faecalis brain abscess was evaluated in five dogs at 1.5T. Imaging was performed during the late cerebritis stage, at 5 to 7 days after surgery. Magnetic resonance scans were acquired before and at 1, 5, 15, 30, 45, and 60 minutes after contrast administration, using a dose of 0.1 mmol/kg. Scans also were acquired both before and after contrast injection with the implementation of magnetization transfer. RESULTS: Lesion enhancement, quantified by region-of-interest measurement, peaked at 5 minutes after contrast injection. Both the increase in lesion enhancement from 1 to 5 minutes after injection and the decrease from 5 to 15 minutes after injection, although small, were statistically significant (P < 0.004 and P < 0.03, respectively). The application of magnetization transfer improved lesion enhancement, as measured by signal difference/noise, by 39%. This result also was statistically significant (P < 0.001). CONCLUSIONS: In intraparenchymal brain infection, Gd-BOPTA provides effective lesion enhancement when used at a dose of 0.1 mmol/kg. Further research is needed to compare the magnitude of enhancement achieved with Gd-BOPTA, which has weak protein binding and both hepatobiliary and renal excretion, with that with Gd chelates, which have pure renal excretion.

Animals↗
PubMed1996

Choice of metal ion and formulation concentration for first-pass brain perfusion studies with magnetic resonance imaging at 1.5 tesla.

RATIONALE AND OBJECTIVES: Gadolinium (Gd) and dysprosium (Dy) analogues, chelated with HP-DO3A, were compared at both 0.5- and 1.0-mol/L concentrations for efficacy in first-pass brain studies on magnetic resonance (MR) imaging at 1.5 tesla (T). METHODS: Ten healthy cats were examined with a dose of 0.3 mmol/kg, using two concentrations of each agent (0.5 mol/L and 1.0 mol/L, 20 examinations). Gadolinium-HP-DO3A (gadoteridol or ProHance) or Dy-HP-DO3A was injected at 9 mL/second, with acquisition of 64 sequential steady-state free precession (SSFP) images at a rate of one each 0.6 second. RESULTS: The change in white matter signal intensity, at the peak of the first pass of the contrast agent bolus in the brain, was -231 +/- 68 for the 0.5-mol/L formulation of Gd-HP-DO3A, compared with -267 +/- 57 for the 1.0-mol/L formulation. Using the 0.5-mol/L formulation of Dy-HP-DO3A, the change at peak was -318 +/- 42, a result statistically improved compared with both the 0.5-mol/L (P < 0.02) and 1.0-mol/L (P < 0.04) Gd-HP-DO3A formulations. A further improvement was observed with the 1.0-mol/L Dy-HP-DO3A formulation, with the change being -368 +/- 33. CONCLUSIONS: First-pass brain MR studies at 1.5 T are improved by use of higher concentration Gd chelate formulations (1.0 versus 0.5 mol/L) and by substitution of the Dy ion for the Gd ion in the chelate. Injection of higher-concentration formulations results in higher initial arterial metal ion concentration. Incomplete blood mixing on transit during first pass causes the higher initial concentration, which then results in a greater susceptibility effect on imaging. The superiority of the Dy formulation compared with the Gd formulation is anticipated because of the higher magnetic moment of Dy. The curves for tissue signal intensity versus time during first pass return artifactually to near baseline after Gd chelate injection (when SSFP imaging techniques are used), a differentiating feature from results with the Dy chelate. This difference can be explained by a substantial T1 effect of the Gd chelate, despite acquisition of images that are predominantly susceptibility weighted.

Animals↗
PubMed1996

Early dynamic magnetic resonance imaging of liver metastases with 0.3 and 0.6 mmol/kg gadoteridol injection.

RATIONALE AND OBJECTIVES: The potential for improvement in liver-lesion conspicuity on early dynamic scans after bolus intravenous gadolinium (Gd) chelate administration was evaluated using gadoteridol (Gd-HP-DO3A; Prohance) at doses of 0.3 and 0.6 mmol/kg. METHODS: Five New Zealand white rabbits with focal VX-2 adenocarcinoma liver metastases were studied on a 1.5-tesla Siemens Vision scanner. Each rabbit was imaged twice (on two separate days), after injections of 0.3 mmol/kg and 0.6 mmol/ kg Gd-HP-DO3A. The contrast dose (0.3 or 0.6 mmol/kg) was given as a single intravenous injection. The order of injection for the two doses was randomized, with the two studies (in any one rabbit) separated by 24 hours to allow for clearance. Contrast was administered using an autoinjector at a rate of 1.5 mL/second. Turbo-fast low-angle shot scans were obtained before and at 6, 12, 19, 25, 31, 60, 120, 180, 240, 300, and 600 seconds after contrast injection. The lesions were confirmed, after killing the rabbit, by gross and microscopic examination. RESULTS: The enhancement of normal liver parenchyma, assessed by (SIt-SIo)/SIo.100, (SI = signal intensity) peaked at 32% +/- 4% 19 seconds after injection of 0.3 mmol/kg and at 38% +/- 5% 31 seconds after injection of 0.6 mmol/kg. The difference in maximum parenchymal enhancement achieved, comparing the 0.3 and 0.6 mmol/kg doses, was statistically significant (P < 0.03). Lesion conspicuity, specifically (SIliver-SIlesion/noise), increased from 4.5 +/- 2.3 precontrast to a maximum of 6.8 +/- 1.2 at 19 seconds postcontrast using a dose of 0.3 mmol/kg, with the difference statistically significant (P < 0.03). The increase with a dose of 0.6 mmol/kg was from 4.2 +/- 0.7 to 6.5 +/- 1.9 with this difference also statistically significant (P < 0.02). There was no statistically significant difference in lesion conspicuity between the doses of 0.3 and 0.6 mmol/kg. CONCLUSIONS: Conspicuity of liver metastases can be improved substantially with dynamic magnetic resonance imaging and rapid intravenous bolus contrast injection with a dose of 0.3 mmol/kg. No further improvement is noted at a dose of 0.6 mmol/kg, despite greater positive contrast enhancement of normal liver parenchyma.

Adenocarcinoma↗
PubMed1996

Hepatic abscesses. Magnetic resonance imaging findings using gadolinium-BOPTA.

RATIONALE AND OBJECTIVES: Gadolinium (Gd)-BOPTA was evaluated in a rabbit liver abscess model and compared with Gd-HP-DO3A, examining lesion conspicuity and characterization. METHODS: Five New Zealand White rabbits with a liver abscess were studied on a 1.5-tesla Siemens Vision magnetic resonance unit. The disease model was created by surgically implanting a gel capsule filled with fusobacterium into the central or left lobe of the liver. For imaging, the animals were ventilated using a Harvard pump. Pancuronium bromide (0.12 mg/kg) was administered to allow acquisition of breath-hold scans. Magnetic resonance scans were obtained in each animal on days 2 and 3 after surgery. Every animal was studied twice, once after intravenous injection of 0.3 mmol/kg Gd-HP-DO3A (gadoteridol; ProHance) and once after intravenous injection of 0.1 mmol/kg Gd-BOPTA (gadobenate dimeglumine; MultiHance). The order of injection for the two agents was randomized with the two studies in each animal, separated by 24 hours to permit clearance. Image acquisition was performed in each instance with respiration suspended. Baseline two-dimensional spin-echo T1-weighted and fast spin-echo T2-weighted breath-hold scans were obtained first. The voxel dimensions were 5 x 0.8 x 0.8 mm3. Imaging times were 23 seconds for the T1-weighted scan and 26 seconds for the T2-weighted scan. Postcontrast scans, using spin-echo T1-weighted technique, were obtained at 1, 3, 5, and 15 minutes after contrast injection, whether Gd-HP-DO3A or Gd-BOPTA was used. Additional scans were obtained at 30, 45, and 60 minutes after Gd-BOPTA administration. At the completion of imaging on day 3, each animal was killed and the liver was removed and taken to a veterinary pathologist at the University's animal disease diagnostic lab for gross and histologic examination. RESULTS: The enhancement of normal liver parenchyma, assessed by region of interest measurement and specifically as (SI(t) - SI0)/SI0 x 100, peaked at 119 +/- 37% 1 minute after injection of 0.3 mmol/kg Gd-HP-DO3A and at 126 +/- 30% 30 minutes after injection of 0.1 mmol/kg Gd-BOPTA. The difference in enhancement achieved, comparing results at each time point, was statistically significant only at 1 and 3 minutes postcontrast (P = 0.003 and 0.03). Lesion conspicuity, specifically (SIliver - SIlesion/noise), increased from 272 +/- 29 precontrast to a maximum of 639 +/- 73 at 30 minutes postcontrast using a dose of 0.1 mmol/kg Gd-BOPTA, with the improvement statistically significant (P = 0.0003). Lesion conspicuity on the T2-weighted scan was 137 +/- , with the Gd-BOPTA scan markedly superior (P = 0.00004). On scans at 45 and 60 minutes after Gd-BOPTA administration, a progressive increase in signal intensity in the central necrotic portion of the lesion was observed. This was most consistent with gradual diffusion of the agent from the adjacent liver into the lesion. Using Gd-HP-DO3A at 0.3 mmol/kg (three times the dose for Gd-BOPTA), lesion conspicuity increase from 305 +/- 37 precontrast to a maximum of 701 +/- 92 at 1 minute postcontrast, with this difference also statistically significant (P = 0.0004). The abscess rim exhibited moderate contrast enhancement, greater than that of normal liver parenchyma, on early postcontrast images with Gd-HP-DO3A. CONCLUSIONS: The conspicuity of an early liver abscess is improved markedly on delayed imaging after administration of 0.1 mmol/kg Gd-BOPTA. Although a similar magnitude of parenchymal enhancement can be obtained after the administration of an extracellular agent, such as Gd-HP-DO3A, high-contrast dose (0.3 mmol/kg) and early dynamic imaging are required. The appearance of a liver abscess on late scans (45 to 60 minutes) after Gd-BOPTA injection is distinct from that of nonnecrotic metastases, with diffusion of the agent into the lesion noted.

Animals↗
PubMed1995

Cooperativity manifest in the binding properties of purified cardiac muscarinic receptors.

Muscarinic receptors were solubilized from porcine atria in digitonin-cholate and were purified by chromatography on DEAE-Sepharose and 3-(2'-aminobenzhydryloxy)tropane-Sepharose. The product identified on Western blots migrated with an apparent molecular mass of 60-75 kDa, with additional bands indicative of homotrimers (190 kDa) and homotetramers (240 kDa). Receptor eluted from the affinity column was accompanied by a mixture of guanyl nucleotide-binding proteins (G-proteins) identified on Western blots as Gi1/2, G(o), Gq/11, and Gs (preparation M2G); the G-proteins were largely removed by further processing on hydroxyapatite (preparation M2). Solubilized purified receptors bound muscarinic ligands in an apparently cooperative manner. In studies at equilibrium, the antagonists [3H]AF-DX 384, N-[3H]methylscopolamine (NMS), and [3H]quinuclidinylbenzilate (QNB) revealed Hill coefficients between about 0.8 and 1.2. Also, the apparent capacity for [3H]QNB exceeded that for [3H]AF-DX 384 and [3H]NMS by about 1.5-fold in M2 and by 2-fold in M2G. Binding to M2G at high concentrations of [3H]QNB was fully inhibited by unlabeled NMS, which therefore affected sites not labeled at similar concentrations of [3H]NMS. Oxotremorine-M displayed a biphasic inhibitory effect on the binding of [3H]AF-DX 384 in M2 and M2G, suggesting that multiple states of affinity are intrinsic to the receptor; 5'-guanylylimidodiphosphate was without appreciable effect in M2 but resulted in a bell-shaped binding profile for the agonist in M2G. All of the data can be described in terms of cooperative interactions within a receptor that is at least tetravalent and presumably an oligomer. In the context of the model, copurifying G-proteins and guanyl nucleotides serve to regulate the degree of cooperativity between successive equivalents of muscarinic ligands.

Allosteric Regulation↗
PubMed1995

Comparison of gadolinium Cy2DOTA, a new hepatobiliary agent, and gadolinium HP-DO3A, an extracellular agent, in healthy liver and metastatic disease.

RATIONALE AND OBJECTIVES: A new gadolinium (Gd) chelate with preferential hepatobiliary uptake, Gd Cy2DOTA, was compared in two animal species with Gd HP-DO3A (gadoteridol), a clinically approved contrast agent with extracellular distribution. Liver enhancement was evaluated for these two contrast agents using magnetic resonance imaging, whereas an experimental model of metastatic disease was used to evaluate the agents' efficacy for liver-lesion delineation. METHODS: The two agents were compared in four healthy Rhesus monkeys (eight studies) and five New Zealand White rabbits with implanted VX-2 liver tumors (ten studies). The contrast dose was 0.1 mmol/kg, with the agents given in random order and at least 72 hours between contrast injections. Breathhold T1-weighted spin echo scans were obtained at 1.5 tesla (T) before and after contrast was administered. Postcontrast scans were obtained 1 to 90 minutes after injection in the monkeys and 1 to 240 minutes after injection in the rabbits. RESULTS: Prolonged hepatic enhancement, superior in degree to that with Gd HP-DO3A, was noted in both monkeys and rabbits after injection of Gd Cy2DOTA. Two minutes after contrast, liver SI was 1.94 +/- 0.05 with Gd Cy2DOTA compared with 1.51 +/- 0.05 with Gd HP-DO3A in monkeys. Sixty minutes after contrast, liver SI was 1.60 +/- 0.09 compared with 1.20 +/- 0.02. The difference between agents was significant at all times from 2 to 60 minutes after contrast injection (P < 0.01). Excretion of contrast into the gall bladder was observed in both animal species with Gd Cy2DOTA but not with Gd HP-DO3A. The maximum improvement in lesion conspicuity (rabbit) occurred 45 minutes after injection of Gd Cy2DOTA and 5 minutes after injection of Gd HP-DO3A. Sixty minutes after injection, liver-lesion contrast was 246 +/- 61 with Gd Cy2DOTA and 106 +/- 28 with Gd HP-DO3A, with a significant difference (P < 0.02). CONCLUSIONS: Gd Cy2DOTA provides greater enhancement of the liver parenchyma on immediate and delayed magnetic resonance scans than does Gd HP-DO3A. On delayed scans, Gd Cy2DOTA provides superior delineation of metastatic liver lesions.

Adenocarcinoma↗