Expression of the recombinant human immunoglobulin J chain in Escherichia coli.

Selective transport of polymeric (p) immunoglobulins (Ig) of IgA and IgM isotypes into external secretions by pIg receptor-mediated mechanism depends on the incorporation of joining (J) chain into the polymers. Until now, availability of a free J chain for immunological and biophysical studies has been limited to preparations of denatured J chain forms with moderate yield. Here we report that a recombinant J chain (rJ) can be over-expressed as a soluble fusion protein with thioredoxin using a modified vector pET32 in Escherichia coli.

In vivo biodistribution of 125IPIP and internal dosimetry of 123IPIP radioiodinated agents selective to the muscarinic acetylcholinergic receptor complex.

The development of new radioiodinated ligands for imaging the muscarinic acetylcholinergic complex (mAChR) using single photon emission computed tomography (SPECT) requires the evaluation of human organ doses prior to approval for human use. Animal biodistribution and excretion data were obtained and evaluated for IPIP, a new mAChR agent. Preliminary biodistribution studies were performed on four different stereoisomers of IPIP.

Z-IQNP: a potential radioligand for SPECT imaging of muscarinic acetylcholine receptors in Alzheimer's disease.

Rationale: The density of the M2 subtype of muscarinic acetylcholine receptors (mAChR) has been shown to be reduced in the brain of patients with Alzheimer's disease (AD). It is therefore of interest to develop a brain imaging method for diagnostic purposes. Z-(R,R)-1-azabicyclo[2.

The genome sequence of Drosophila melanogaster.

The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation.

Evaluation of Z-(R,R)-IQNP for the potential imaging of m2 mAChR rich regions of the brain and heart.

Alterations in the function or density of the m2 muscarinic (mAChR) subtype have been postulated to play an important role in various dementias such as Alzheimer's disease. The ability to image and quantify the m2 mAChR subtype is of importance for a better understanding of the m2 subtype function in various dementias. Z-(R)-1-Azabicyclo[2.

A rapid and simple Sep Pak method for purification of radioiodinated IQNP, a high affinity ligand for the muscarinic receptor.

A simplified procedure for the purification of 1-azabicyclo[2.2.2]oct-3-yl alpha-hydroxy-alpha-(1-iodo-1-propen-3-yl)-alpha-phenylacetate (IQNP) stereoisomers utilizing a silica Sep Pak (SSP) is described.

Dual-label studies with [125I]-3(R)/[131I]-3(S)-BMIPP show similar metabolism in rat tissues.

Unlabelled: Biodistribution studies with the radioiodinated 3(R)- and 3(S)-isomers of 15-(p-iodophenyl)-3-methylpentadecanoic acid (BMIPP) in rats have shown that 3(R)-BMIPP has 20%-25% higher heart uptake than 3(S)-BMIPP (15-180 min). In contrast, the 3(S)-isomer has slightly higher liver uptake, and uptake in other tissues examined is similar.

Methods: To evaluate the possible differences in metabolic fate of the two isomers, a mixture of [125I]-3(R)/[131I]-3(S)-BMIPP was administered to fasted female Fisher rats.

Iodine-123 labelled Z-(R,R)-IQNP: a potential radioligand for visualization of M(1 )and M(2) muscarinic acetylcholine receptors in Alzheimer's disease.

Z-(R)-1-Azabicyclo[2.2.2]oct-3-yl (R)-alpha-hydroxy-alpha-(1-iodo-1-propen-3-yl)-alpha-phenylacetate (Z-IQNP) has high affinity to the M(1 )and M(2) muscarinic acetylcholine receptor (mAChR) subtypes according to previous in vitro and in vivo studies in rats.

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Improved radioiodination of 1,2-dipalmitoyl-3-IPPA via a tributyltin intermediate.

1,2-Palmitoyl-3-[15-(4-iodophenyl)pentadecan-3-oyl]-rac-glycerol (MIPAG) is a new agent for the clinical evaluation of pancreatic lipase activity and has demonstrated promise in preliminary clinical studies with patients affected with pancreatic insufficiency. Iodine-131-MIPAG was initially prepared via thallium-iodide displacement. Because of the need for a simple method which is amendable for the routine clinical use of MIPAG we have investigated the preparation and radioiodination of MIPAG utilizing the tributyltin precursor, 1,2-palmitoyl-3-[15-(4-tributylstannylphenyl)pentadecan-3-oy l]-rac-glycerol (TBT-MIPAG, 2).

Engineering protein-based machines to emulate key steps of metabolism (biological energy conversion).

Metabolism is the conversion of available energy sources to those energy forms required for sustaining and propagating living organisms; this is simply biological energy conversion. Proteins are the machines of metabolism; they are the engines of motility and the other machines that interconvert energy forms not involving motion. Accordingly, metabolic engineering becomes the use of natural protein-based machines for the good of society.

In vivo targeting of acoustically reflective liposomes for intravascular and transvascular ultrasonic enhancement.

Objectives: The purpose of this study was to target acoustically reflective liposomes to atherosclerotic plaques in vivo for ultrasound image enhancement.

Background: We have previously demonstrated the development of acoustically reflective liposomes that can be conjugated for site-specific acoustic enhancement. This study evaluates the ability of liposomes coupled to antibodies specific for different components of atherosclerotic plaques and thrombi to target and enhance ultrasonic images in vivo.

A 'one-stop' vascular clinic: initial experience of an out-patient immediate-access radioisotope limb blood flow service.

The availability of an 'immediate-access' nuclear medicine service has facilitated the introduction of a 'one-stop' vascular clinic, enabling radioisotope limb blood flow and Doppler ankle:brachial index tests to be performed within 2 h of each other for new patient referrals. The test results are available to assist clinical management decisions during the same clinical session. A patient satisfaction survey was undertaken in 71 patients to assess their perspective of this service provision.

In vivo competition studies of Z-(-,-)-[125I]IQNP against 3-quinuclidinyl 2-(5-bromothienyl)-2-thienylglycolate (BrQNT) demonstrating in vivo m2 muscarinic subtype selectivity for BrQNT.

Alzheimer's disease (AD) involves selective loss of muscarinic m2, but not m1, subtype neuroreceptors in cortical and hippocampal regions of the human brain. Until recently, emission tomographic study of the loss of m2 receptors in AD has been limited by the absence of available m2-selective radioligands that can penetrate the blood-brain barrier. We now demonstrate the in vivo m2 selectivity of an analog of (R)-QNB, 3-quinuclidinyl 2-(5-bromothienyl)-2-thienylglycolate (BrQNT), by dissection and autoradiographic studies of the in vivo inhibition of radioiodinated Z-1-azabicyclo[2.

Elastic protein-based polymers in soft tissue augmentation and generation.

Five elastic protein-based polymers, designed as variations of polymer I, (GVGVP)251, elicited different responses when injected as subcutaneous implants in the guinea pig, a preclinical test used to evaluate materials for soft tissue augmentation and specifically for correction of urinary incontinence. All six polymers, prepared using recombinant DNA technology, expressed at good levels using transformed E. coli fermentation.

In vitro targeting of acoustically reflective immunoliposomes to fibrin under various flow conditions.

We have previously demonstrated the development of acoustically reflective liposomes as a novel ultrasound contrast agent, that can be conjugated to antibodies for site specific acoustic enhancement of pathologically altered vascular tissue. The liposomes are echogenic due to the lipid composition, without gas entrapment, and have a size of less than one micron (Alkan-Onyuksel et al., 1996).

Left ventricular functional improvement after transmyocardial laser revascularization.

Background: Transmyocardial laser revascularization has been used to treat patients with end-stage coronary artery disease that is not amenable to standard revascularization. Although there is evidence of angina relief and quality of life enhancement, there is little information concerning improvement in myocardial contractility. The purpose of this study was to determine whether transmyocardial laser revascularization improves myocardial function in chronically ischemic myocardium.

Expression and purification of Streptococcus pneumoniae hyaluronate lyase from Escherichia coli.

Pneumococcal hyaluronate lyase enzyme breaks down hyaluronan of the extracellular matrix of tissues and possibly contributes to the invasion of host tissue and to the penetration of host defenses by this bacterial pathogen. In light of the emergence of increasing numbers of antibiotic-resistant strains, the understanding of the mechanism of action of hyaluronate lyase enzyme may lead to a better understanding of interactions between a host and bacterial pathogens and may contribute to more efficient treatment of bacterial infections. The native Streptococcus pneumoniae hyaluronate lyase enzyme has a molecular mass of 107 kDa but undergoes conversion to smaller enzymatically active forms.

Evaluation and metabolite studies of 125I- and 123I-labelled E-(R,R)-IQNP: potential radioligands for visualization of M1 muscarinic acetylcholine receptors in brain.

A new ligand for the M1 muscarinic receptor subtype, E-(R,R)-1-azabicyclo[2.2.2]oct-3-yl alpha-hydroxy-alpha-(1-iodo-1-propen-3-yl)-alpha-phenylacetate (E-IQNP), was labelled with 125I and 123I for autoradiographic studies on human whole-brain cryosections and SPET studies, respectively, in Cynomolgus monkey.

A hypothesis regarding vascular acoustic emission accompanying arterial injury induced by balloon angioplasty.

Stress-induced structural damage is often accompanied by sound release. This behavior is known as acoustic emission (AE). We hypothesize that vascular injury such as that produced by balloon angioplasty is associated with AE.

Effects of configuration on the myocardial uptake of radioiodinated 3(R)-BMIPP and 3(S)-BMIPP in rats.

Unlabelled: Radioiodinated 3(R)-(+)- and 3(S)-(-)-15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid (BMIPP) were prepared and evaluated in rats to investigate the effects of absolute configuration of the 3(beta)-methyl group on myocardial uptake and release kinetics.

Methods: The 3(R)-(+)-BMIPP analog was synthesized by initial acylation of a thiophene template with the acid chloride of ethyl 3(R)-methylglutarate. 3(S)-(-)-BMIPP was obtained by separation from the mixture of diastereomeric amides prepared from reaction of the acid chloride of racemic BMIPP with the S-(-)-alpha-methylbenzylamine.

Optical imaging of neuronal activity in tissue labeled by retrograde transport of Calcium Green Dextran.

In many neurophysiological studies it is desirable to simultaneously record the activity of a large number of neurons. This is particularly true in the study of vertebrate motor systems that generate rhythmic behaviors, such as the pattern generator for locomotion in vertebrate spinal cord. Optical imaging of neurons labeled with appropriate fluorescent dyes, in which fluorescence is activity-dependent, provides a means to record the activity of many neurons at the same time, while also providing fine spatial resolution of the position and morphology of active neurons.

Radiolabeled ligands for imaging the muscarinic-cholinergic receptors of the heart and brain.

Interest in the potential use of cerebral SPECT and PET imaging for determination of the density and activity of muscarinic-cholinergic receptors (mAChR) has been stimulated by the changes in these receptors which occur in many neurological diseases. In addition, the important involvement of mAChR in modulating negative inotropic cardiac activity suggests that such receptor ligands may have important applications in evaluation of changes which may occur in cardiac disease. In this paper, the properties of several key muscarinic receptor ligands being developed or which have been used for clinical SPECT and PET are discussed.

Positron emission tomographic investigations of central muscarinic cholinergic receptors with three isomers of [76Br]BrQNP.

We studied the potential of three radiobrominated isomers of BrQNP, (Z(-,-)-[76Br]BrQNP, E(-,-)-[76Br]BrQNP and E(-,+)-[76Br]BrQNP), as suitable radioligands for imaging of central muscarinic cholinergic receptors in the human brain. These radioligands were stereospecifically prepared by electrophilic radiobromodestannylation of the respective tributylstannyl precursors using no-carrier-added [76Br]BrNH4 and peracetic acid. Preliminary pharmacological characterizations were determined by biodistribution, autoradiography, competition, displacement and metabolite studies in rats.