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Biomedical subjects

S A Goldstein

Publications and source records attributed to S A Goldstein.

At least 19 recordsLinked to original sources
PubMed1996

ORK1, a potassium-selective leak channel with two pore domains cloned from Drosophila melanogaster by expression in Saccharomyces cerevisiae.

A K+ channel gene has been cloned from Drosophila melanogaster by complementation in Saccharomyces cerevisiae cells defective for K+ uptake. Naturally expressed in the neuromuscular tissues of adult flies, this gene confers K+ transport capacity on yeast cells when heterologously expressed. In Xenopus laevis oocytes, expression yields an ungated K(+)-selective current whose attributes resemble the "leak" conductance thought to mediate the resting potential of vertebrate myelinated neurons but whose molecular nature has long remained elusive. The predicted protein has two pore (P) domains and four membrane-spanning helices and is a member of a newly recognized K+ channel family. Expression of the channel in flies and yeast cells makes feasible studies of structure and in vivo function using genetic approaches that are not possible in higher animals.

Amino Acid Sequence↗
PubMed1996

Stimulation of new bone formation by direct transfer of osteogenic plasmid genes.

Degradable matrices containing expression plasmid DNA [gene-activated matrices (GAMs)] were implanted into segmental gaps created in the adult rat femur. Implantation of GAMs containing beta-galactosidase or luciferase plasmids led to DNA uptake and functional enzyme expression by repair cells (granulation tissue) growing into the gap. Implantation of a GAM containing either a bone morphogenetic protein-4 plasmid or a plasmid coding for a fragment of parathyroid hormone (amino acids 1-34) resulted in a biological response of new bone filling the gap. Finally, implantation of a two-plasmid GAM encoding bone morphogenetic protein-4 and the parathyroid hormone fragment, which act synergistically in vitro, caused new bone to form faster than with either factor alone. These studies demonstrate for the first time that repair cells (fibroblasts) in bone can be genetically manipulated in vivo. While serving as a useful tool to study the biology of repair fibroblasts and the wound healing response, the GAM technology may also have wide therapeutic utility.

Animals↗
PubMed1996

Type-I collagen mutation compromises the post-yield behavior of Mov13 long bone.

Despite recent advances in our understanding of the molecular basis of skeletal fragility, little is known about how these molecular alterations lead to whole bone brittleness. In the current study, we investigated the relationship between a type-I collagen mutation and post-yield behavior of whole bone in Mov13 transgenic mice by considering tissue-level organizational issues known to be important for normal bone fracture. Mechanical assays revealed that the post-yield deflection of Mov13 femurs was reduced by 61% relative to littermate controls. Fractographic images revealed that lamellar interfaces which were important for dissipating energy during the failure process of control femurs, were not effective in Mov13 mice. Further investigation revealed that a 22% reduction in bone collagen content, a 2-fold increase in tissue porosity, and significant alterations in collagen organization interfered with normal energy dissipation mechanisms of Mov13 microstructure. Collectively, the results provided the first evidence that the reduced ductility associated with a type-I collagen mutation was mediated by alterations in intermediate structures that normally contribute to the post-yield behavior of cortical bone. The results suggest that, to better understand the pathogenesis of skeletal fragility, it is important to consider the effects of molecular alterations on higher-level structures, particularly those structures that contribute to the failure mechanisms in normal bone.

Animals↗
PubMed1996

Local expression of human growth hormone in bone results in impaired mechanical integrity in the skeletal tissue of transgenic mice.

The effect of local production of human growth hormone on murine cortical bone was investigated using a transgenic mouse model. Femora and humeri from human growth hormone transgenic mice and littermate control mice were obtained, and the geometrical, biomechanical, compositional, and histomorphometric properties of all specimens were determined. The goals were to investigate the effects of local expression of human growth hormone on skeletal integrity, including the functional geometry of long bone and its related structural and mechanical behavior, as well as tissue composition and integrity. As expected, local production of human growth hormone by osteoblasts indeed resulted in longer femora with significantly greater mid-diaphyseal cross-sectional geometry in the transgenic mice (16% increase in cross-sectional area and 29% increase in bending moments of inertia). However, the significant increase in geometry was not associated with a proportional increase in bending stiffness and other structural properties, which suggested that the mechanical properties of the cortical bone tissue may have been inferior. Microspecimen bending tests verified this prediction, given that transgenic cortical bone tissue had significantly lower apparent elastic modulus and ultimate strength (52 and 68%, respectively, of control values). These defects in the whole bone structural and tissue mechanical properties of transgenic specimens were associated with a smaller fraction of ash, larger fractions of woven bone and cartilage islands, and greater porosity in the mid-diaphyseal cortices. These results suggest that local production of human growth hormone by osteoblasts is indeed anabolic for bone, but at the expense of bone tissue integrity.

Adaptation, Physiological↗
PubMed1996

Relative effects of wound healing and mechanical stimulus on early bone response to porous-coated implants.

We hypothesized that early bone adaptation to well fixed porous-coated implants is influenced more by wound healing than by mechanical loading. To test this hypothesis, two groups of dogs with identical, hydraulically controlled porous-coated implants interference fit within distal femoral trabecular bone were used. One group had no load: the other had 35 N of load applied to the implants. At 5 weeks after surgery, the resulting adaptation of bone around the implants was quantified on a cellular basis by cytochemical analysis of type-I procollagen synthesis and on a structural basis using three-dimensional micro-computed tomography imaging. The percentage of trabecular surfaces covered by osteoblasts expressing type-I procollagen was significantly increased in bone surrounding the implant in both groups compared with contralateral control bone tissue. There was no difference between the groups with no load or 35 N of load. In addition, measures of trabecular bone structure did not differ significantly between the load and no-load groups. Taken together, these results suggest that wound healing plays a much greater role in the early response of bone to well fixed porous-coated implants than does mechanical stimulus.

Animals↗
PubMed1996

MinK residues line a potassium channel pore.

MinK has neither the P region nor signature sequence that characterizes pore-forming subunits of all known K+ channels. A specific minK region has now been identified that affects external blockade by 2 common probes of K+ channel pores. When mutated to cysteine, residues in this region render minK susceptible to covalent blockade by methanethiosulfonate ethylsulfonate and alter reversible inhibition by tetraethylammonium. The 2 blockers are found to share overlapping binding site determinants and to interact. Since inhibition by external tetraethylammonium is sensitive to voltage and to the internal concentration of permeant ions, we argue that tetraethylammonium blocks by occluding the external end of a water-filled transmembrane pore. These findings support the view that minK is directly involved in forming a K+-selective ion conduction pathway.

Molecular Structure↗
PubMed1996

Oxytetracycline-induced nephrotoxicosis in dogs after intravenous administration for experimental bone labeling.

Tetracyclines have been used as in vivo indicators of new bone formation because they form complexes with mineral at bone-forming surfaces. Four of 12 dogs in a bone-labeling study developed clinical signs of renal disease (vomiting, diarrhea, dehydration, and azotemia) within 1 to 2 days of receiving oxytetracycline at a bone-labeling dose of 25 mg/kg of body weight, once daily for 2 consecutive days. To delineate the relationship between oxytetracycline administration and renal damage, six dogs were given the bone-labeling dose intravenously and were subsequently evaluated by determination of clinical signs, serum biochemical analysis, urinalysis, and histologic examination (experiment 1). Drug administration was modified in the five dogs remaining in the bone-labeling orthopedic study. These dogs received the oxytetracycline dose as a slow intravenous infusion diluted with 250 ml of lactated Ringer's solution (experiment 2). All six dogs of experiment 1 developed persistent isosthenuria within 2 days of receiving the bone-labeling dose of oxytetracycline. Clinical illness (three of six dogs) was associated with azotemia, creatinemia, and hyperphosphatemia. All dogs had multifocal, mild to moderate flattening of renal tubular epithelium, characteristic of nephrosis. None of the dogs of experiment 2 developed any clinical indications of renal disease, and the only biochemical abnormality was isosthenuria in two of the five dogs. Thus the development of clinical signs and biochemical abnormalities associated with the intravenous administration of oxytetracycline was obviated by the slow administration of a dilution of the calculated bone-labeling dose of the antibiotic.

Animals↗
PubMed1995

A new family of outwardly rectifying potassium channel proteins with two pore domains in tandem.

Potassium channels catalyse the permeation of K+ ions across cellular membranes and are identified by a common structural motif, a highly conserved signature sequence of eight amino acids in the P domain of each channel's pore-forming alpha-subunit. Here we describe a novel K+ channel (TOK1) from Saccharomyces cerevisiae that contains two P domains within one continuous polypeptide. Xenopus laevis oocytes expressing the channel exhibit a unique, outwardly rectifying, K(+)-selective current. The channel is permeable to outward flow of ions at membrane potentials above the K+ equilibrium potential; its conduction-voltage relationship is thus sensitive to extracellular K+ ion concentration. In excised membrane patches, external divalent cations block the channel in a voltage-dependent manner, and their removal in this configuration allows inward channel current. These attributes are similar to those described for inwardly rectifying K+ channels, but in the opposite direction, a previously unrecognized channel behaviour. Our results identify a new class of K+ channel which is distinctive in both its primary structure and functional properties. Structural homologues of the channel are present in the genome of Caenorhabditis elegans.

Amino Acid Sequence↗
PubMed1995

Skeletal repair by in situ formation of the mineral phase of bone.

A process has been developed for the in situ formation of the mineral phase of bone. Inorganic calcium and phosphate sources are combined to form a paste that is surgically implanted by injection. Under physiological conditions, the material hardens in minutes concurrent with the formation of dahllite. After 12 hours, dahllite formation was nearly complete, and an ultimate compressive strength of 55 megapascals was achieved. The composition and crystal morphology of the dahllite formed are similar to those of bone. Animal studies provide evidence that the material is remodeled in vivo. A novel approach to skeletal repair is being tested in human trials for various applications; in one of the trials the new biomaterial is being percutaneously placed into acute fractures. After hardening, it serves as internal fixation to maintain proper alignment while healing occurs.

Animals↗
PubMed1995

Static and fatigue failure properties of thoracic and lumbar vertebral bodies and their relation to regional density.

This study investigated (1) whether a characterization of the macroscopic architecture within the vertebral centrum would improve predictions of vertebral strength, (2) if regions in the centrum where least bone loss with age occurs are more predictive of vertebral strength, and (3) whether different patterns of the macroscopic architecture are predictive of static as compared to fatigue strength. To characterize the vertebral macroscopic architecture, a regional bone mineral density (rBMD) technique was used that estimated the cancellous density distribution (in 18 specific regions of the vertebral centrum) for vertebrae T7-L4, from spines of 20 female cadavers. Static and fatigue failure properties of whole vertebrae were obtained, and predictive models of static and fatigue failure properties of whole vertebrae were examined. We found that (1) vertebral failure properties were better predicted by combinations of vertebral regional cancellous density (multiple linear regressions) rather than by any individual region of cancellous density alone (simple linear regressions); (2) models using regions of density that demonstrated minimum decline with age [from the data of Flynn and Cody (Calcif. Tissue Int. 53, S170-S175 (1993))] resulted in better correlations with ex vivo vertebral static failure properties than models using density regions that showed maximum decline with age, and (3) static and fatigue characteristics required different density regions to reach significance. (A comparison of models predictive of static and fatigue failure properties revealed that anterior density regions were most often included in predictive models of the static properties while posterior regions were more predictive of the fatigue properties).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Subunit composition of minK potassium channels.

Expression of minK protein in Xenopus oocytes induces a slowly activating, voltage-dependent, potassium-selective current. Point mutations in minK that alter current gating kinetics, ion selectivity, pharmacology, and response to protein kinase C all support the notion that minK is a structural protein for a channel-type transporter. Yet, minK has just 130 amino acids and a single transmembrane domain. Though larger cloned potassium channels form functional channels through tetrameric subunit association, the subunit composition of minK is unknown. Subunit stoichiometry was determined by coexpression of wild-type minK and a dominant lethal point mutant of minK, which reaches the plasma membrane but passes no current. The results support a model for complete minK potassium channels in which just two minK monomers are present, with other, as yet unidentified, non-minK subunits.

Animals↗
PubMed1994

Three quantitative ultrasound parameters reflect bone structure.

We investigated whether quantitative ultrasound (QUS) parameters are associated with bone structure. In an in vitro study on 20 cubes of trabecular bone, we measured broadband ultrasound attenuation (BUA) and two newly defined parameters--ultrasound velocity through bone (UVB) and ultrasound attenuation in bone (UAB). Bone mineral density (BMD) was measured by dual X-ray absorptiometry (DXA) and bone structure was assessed by microcomputed tomography (microCT) with approximately 80 microns spatial resolution. We found all three QUS parameters to be significantly associated with bone structure independently of BMD. UVB was largely influenced by trabecular separation, UAB by connectivity, and BUA by a combination of both. For a one standard deviation (SD) increase in UVB, a decrease in trabecular separation of 1.2 SD was required compared with a 1.4 SD increase in BMD for the same effect. A 1.0 SD increase in UAB required a reduction in connectivity of 1.4 SD. Multivariate models of QUS versus BMD combined with bone structure parameters showed squared correlation coefficients of r2 = 0.70-0.85 for UVB, r2 = 0.27-0.56 for UAB, and r2 = 0.30-0.68 for BUA compared with r2 = 0.18-0.58 for UVB, r2 < 0.26 for UAB and r2 < 0.13 for BUA for models including BMD alone. QUS thus reflects bone structure, and a combined analysis of QUS and BMD will allow for a more comprehensive assessment of skeletal status than either method alone.

Absorptiometry, Photon↗
PubMed1994

The relationship between the structural and orthogonal compressive properties of trabecular bone.

In this study, cubes of trabecular bone with a wide range of structural properties were scanned on a micro-computed tomography system to produce complete three-dimensional digitizations from which morphological and architectural parameters could be measured in a nondestructive manner. The cubes were then mechanically tested in uniaxial compression in three orthogonal directions and to failure in one direction to find the orthogonal tangent elastic moduli and ultimate strengths. After testing, the cubes were weighed and ashed to determine the apparent and ash densities. A high correlation between the basic stereologic measurements was found, indicating that there is a relationship between the amount of bone and number of trabeculae in cancellous bone. Regression analysis was used to estimate the modulus and ultimate strength; these regressions accounted for 68-90% of the variance in these measures. These relationships were dependent on the metaphyseal type and donor, with the modulus also dependent on the direction of testing. This indicates that the properties of the individual trabeculae, as well as their amount and organization, may be important in predicting the mechanical properties of cancellous bone.

Adult↗
PubMed1994

Displacements of the menisci under joint load: an in vitro study in human knees.

The purpose of this study was to test whether the menisci displace under joint compression combined with internal-external torques and anterior-posterior forces at fixed flexion angles. We further determined differences in displacements between the medial and lateral menisci. Loads were applied to the joint, and joint load and displacements were measured. Meniscal displacements were measured radiographically. With a joint compressive load of 1 kN, internal and external joint rotations caused the lateral meniscus to displace, on average, 0.37 mm deg-1 in the anterior-posterior direction, while the medial meniscus displaced 0.19 mm deg-1. Anterior and posterior joint translation, performed under 1 kN joint compression, caused the lateral meniscus to displace, on average, 0.66 mm mm-1 in the anterior-posterior direction, while the medial meniscus displaced 0.43 mm mm-1. Greater meniscal displacements were found at 15 and 30 degrees flexion than at 0 degrees for the lateral meniscus in internal rotation. Lateral meniscal displacements were larger than those of the medial with posterior tibial translation at full extension and with internal rotation.

Femur↗
PubMed1994

The charybdotoxin receptor of a Shaker K+ channel: peptide and channel residues mediating molecular recognition.

Charybdotoxin (CTX) is a peptide of known structure that inhibits Shaker K+ channels by a pore-blocking mechanism. Point mutagenesis of all 30 solvent-exposed residues identified the part of the CTX molecular surface making contact with the receptor in the K+ channel. All close-contact residues are clustered in a well-defined interaction surface; the shape of this surface implies that the outer opening of the Shaker channel conduction pore abruptly widens to a 25 x 35 A plateau. A mutagenic scan of the S5-S6 linker sequence of the Shaker K+ channel identified those channel residues influencing CTX binding affinity. The Shaker residues making the strongest contribution to toxin binding are located close to the pore-lining sequence, and more distant residues on both sides of this region influence CTX binding weakly, probably by an electrostatic mechanism. Complementary mutagenesis of both CTX and Shaker suggests that Shaker-F425 contacts a specific area near T8 and T9 on the CTX molecular surface. This contact point constrains Shaker-F425 to be located at a 20 A radial distance from the pore axis and 10-15 A above the "floor" of the CTX receptor.

Amino Acid Sequence↗
PubMed1994

Cannulated hip screws: a study of fixation integrity, cut-out resistance, and high-cycle bending fatigue performance.

One major advancement in the treatment of femoral neck fractures has been the development of cannulated screws. This study investigates the integrity of five commercially available cannulated screw systems. Transcervical fractures were created in 25 adult femora and repaired by three cannulated hip screws of a randomly assigned design. The repaired specimens were subjected to a triangular loading pattern for 1,000 cycles, then loaded to ultimate failure. Secondly, screws of each type were inserted into femoral heads to a point 5-mm from the chondral surface, then ramp loaded to determine the push-through stiffness and maximum load. Finally, individual screws were tested in high-cycle four-point bending fatigue in a custom fixture. In push-through, the Orthomet screws could withstand a significantly greater maximum load than the Synthes screws. The fatigue life of the Ace screws was significantly longer than the Howmedica, Synthes, and Orthomet screws, whereas the Orthomet screws performed significantly worse than all other screw types. Although cannulated screw systems uniquely address the problems of push-through resistance and high-cycle fatigue failure, certain designs are more susceptible to failure.

Aged↗