bluuebunny/crossref_metadata_2025 huggingface dataset:
Efficacy of monitoring long‐bone fracture healing by measurement of either bone stiffness or resonant frequency: Numerical simulation
Authors: Roberts Sydney G. , Steele Charles R.
Published: February 2005

Development of noninvasive mechanical tests to monitor fracture healing has been hindered because relationships between bone geometry, measurement conditions, and fracture callus strength are not well understood. Beam theory was used to analyze the effects of fracture length, fracture location, end conditions, and fracture callus stiffness on mechanical properties (resonant frequency, bending stiffness, and torsional stiffness) of healing bone. Actual bone mineral geometry from a human tibia, quantified every 1 mm, was used in the beam analysis. Geometry of the fracture callus segment was uniformly scaled from the values for intact bone. Experimental tests on multisegmented machined rods were used to verify analytical methods. Mechanical properties of the healing bone initially increased very rapidly to 30–70% of the stiffness of intact bone, depending on the configuration. The increases then tapered off dramatically. Lateral bending stiffness was sensitive to changes in callus properties for a larger portion of the healing process than was either torsional stiffness or resonant frequency. Because callus strength increases at half the rate of callus stiffness, measures of whole‐bone mechanical properties can provide insight into changes in callus strength until a maximum of less than one‐half the strength of intact bone is regained. The analytical method presented is proposed for clinical use to develop individualized models of bone, fracture, and fixation conditions to identify early stages of healing. Because increases in whole‐bone mechanical properties are small in the later stages of fracture healing, however, such measures must be used prudently beyond the initial stages.
Vascular proliferation and blood supply during distraction osteogenesis: A scanning electron microscopic observation
Authors: Choi In Ho , Ahn Jae Hoon , Chung Chin Youb , Cho Tae‐Joon
Published: February 2005

This scanning electron microscopic study examined the spatial and temporal features of proliferating vessels of regenerating bone tissue and blood supply during distraction osteogenesis. A rat model of tibial lengthening was used with a protocol divided into a latency period of 7 days, a distraction period that lasted 14 days with a daily distraction rate of 0.5 mm on two steps, and a consolidation period of 21 days. Vascular casting was done on the hindlimbs before osteotomy and on postoperative days 7, 14, 21, 28, and 42. Scanning electron microscopic findings were correlated with radiological and histological observations. On days 7 and 14, the proliferation of periosteal vessels was pronounced and there was distinct subperiosteal bone formation on the osteotomized surfaces. On day 2, vascular branches from the medullary canal of the host bone formed a vascular network, which gave rise to multiple axial, straight vascular branches, running parallel to the direction of distraction, toward the interzone, in accordance with the progress of mineralization. On day 28, the periosteum provided vascularization to the peripheral side of the interzone whereas the center of the interzone was still relatively avascular. On day 42, the periosteal and medullary vascular channels were completely connected at the distraction site including the interzone, which was occupied by developing and mature bone trabeculae. These results suggest that vascular proliferation occurs actively during the latency and distraction periods and then gradually decreases over time. A close temporal and spatial relationship exists between formation of regenerated bone and vascular proliferation of the periosteum and medullary canal.
Markers of joint tissue turnover in joint fluids from hips with osteonecrosis of the femoral head
Authors: Jingushi Seiya , Lohmander L. Stefan , Shinmei Masayuki , Hoerrner Lori A. , Lark Michael W. , Sugioka Yoichi , Iwamoto Yukihide
Published: February 2005

Osteonecrosis of the femoral head often results in secondary osteoarthritis of the hip joint; however, the pathologic processes underlying the destruction of articular cartilage are not fully understood. Molecular markers in the hip joint fluids were measured to examine the changes in turnover of cartilage and other joint tissues. Marker data were related to clinical, radiological, and histopathological changes in the articular cartilage of the hip. Forty‐five patients (median age: 43 years) were studied. The median time between the onset of symptoms and sampling of hip synovial fluid was 6 months. Aggrecan fragments, C‐propeptide of type‐II collagen, matrix metalloproteinase‐3, and tissue inhibitor of metalloproteinases‐1 levels in joint fluid were determined by immunoassay. Osteonecrosis of the femoral head was graded by radiology as minimal collapse of the femoral head (stage 2: 26 patients), severe collapse (stage 3: 15 patients), or severe collapse with osteoarthritis (stage 4: four patients). Histologica changes of the articular cartilage, consistent with early‐stage osteoarthritis, were evident at stage 3 and were more advanced at stage 4. The average concentrations of proteoglycan fragments and C‐propeptide of type‐II collagen were 207 (SD 182) μg/ml and 19.6 (SD 19.3) ng/ml, respectively. The average concentrations of matrix metalloproteinase‐3 and tissue inhibitor of metalloproteinases‐ 1 were 177 (SD 291) nM and 23.0 (SD 9.9) nM, respectively. Measurable levels for all markers assayed were noted in the earliest stage of the disease, only a few months after the onset of symptoms and well before the appearance of radiological changes. Levels of matrix metalloproteinase‐3 and molar ratios of matrix metalloproteinase‐3/tissue inhibitor of metalloproteinases‐1 were higher in early stage disease than in later stage disease.
Changes in cross‐sectional geometry of the distal femoral metaphysis associated with inflammatory arthritis are reduced by a bisphosphonate (zoledronate)
Authors: Pysklywec Michael W. , Moran Erica L. , Bogoch Earl R.
Published: February 2005

An increased risk of fracture is a feature of rheumatoid arthritis and of animal models of inflammatory arthritis. We examined geometrical changes in the metaphyseal cortex of the distal femur in an animal model of inflammatory arthritis. Additionally, we examined the effect of a bisphosphonate in preventing these changes. Five groups of rabbits were studies: normal controls, those with inflammatory arthritis, and three groups with arthritis treated with bisphosphonate. To determine geometrical properties, image analysis was performed on digitized cross sections of the femoral metaphyseal cortices. The results demonstrated that the posterior cortical wall was significantly less thick in rabbits with arthrits than in normal rabbits and in the rabbits in the three bisphosphonate treatment groups (p < 0.05). Moment of inertia about the lateral‐medial axis was reduced inrabbits with arthritis compared with normal rabbits (p < 0.05). Cross‐sectional area was not significantly different between groups. The changes suggest a mechanism of weakening of bone in arthritis; when the results are coupled with results of previous porosity studies, severe directional weakness is apparent. Bisphosphonate was effective in preserving bone integrity in inflammatory arthritis.
Quantitative assessment of the effect of 0.05% chlorhexidine on rat articular cartilage metabolismin vitroandIn vivo
Authors: Reading A. D. , Rooney P. , Taylor G. J. S.
Published: February 2005

Wound infection remains a problem. Syringe and needle jet lavage of chlorhexidine gluconate 0.05% removed or killed 99.8% of contaminating bacteria within 1 minute in a wound model. In clinical use, however, possibletoxicity to articular cartilage is a concern. In an established intact rat patella modelin vitro, 1 minute of exposure to chlorhexidine 0.05% and chlorhexidine jet lavage did not significantly alter cartilage metabolism. A 1‐hour exposure decreased metabolic activity.In vivo, a 30‐minute exposure with or without rinsing produced no impairment of metabolic activity 6 weeks later, suggesting that cartilage has the potential for biological recovery. However, injecting and leaving chlorhexidine 0.05% in the joints was detrimental to the metabolic activity of the articular cartilage as assessed 6 weeks later. Thus, chlorhexidine gluconate 0.05% could be used on normal articular cartilage. Any potential damage from prolonged exposure can be avoided by rinsing after 1 minute.
Nitric oxide as a mediator of nucleus pulposus‐induced effects on spinal nerve roots
Authors: Brisby Helena , Byröd Gunnar , Olmarker Kjell , Miller Virginia M. , Aoki Yoshihito , Rydevik Björn
Published: February 2005

Nerve root dysfunction and sciatic pain in disc herniation are considered to be caused by mechanical compression and related to the presence of nucleus pulposus in the epidural space. Autologous nucleus pulposus has been shown to induce endoneural edema and to decrease nerve‐conduction velocity in spinal nerve roots in experimental disc herniation models, and inflammatory mediators have been suggested to be involved in these mechanisms. Nitric oxide, a potent inflammatory mediator, is implicated in vasoregulation, neurotransmission, and neuropathic pain. Nitric oxide synthesis can be induced by different cytokines, e.g., tumor necrosis factor‐α, which recently was shown to be of pathophysiological importance in experimental disc herniation. The enzyme nitric oxide synthase mediates the production of nitric oxide. Three series of experiments were performed in rat and pig disc herniation models to (a) investigate nitric oxide synthase activity in spinal nerve roots after exposure to autologous nucleus pulposus and (b) evaluate the effects of systemic treatment with aminoguanidine, a nitric oxide synthase inhibitor, on vascular permeability and nerve‐conduction velocity. In a disc herniation model in the rat, calcium‐independent nitric oxide synthase activity was measured in nerve roots exposed to nucleus pulposus; however, no nitric oxide synthase activity was detected in nerve roots from animals that underwent a sham operation, reflecting increased inducible nitric oxide synthase activity. In nucleus pulposus‐exposed spinal nerve roots in the pig, the edema was less severe after systemic aminoguanidine administration than without aminoguanidine treatment. Aminoguanidine treatment also significantly reduced the negative effect of nucleus pulposus on nerve‐conduction velocity in spinal nerve roots in the pig. These results demonstrate that nucleus pulposus increases inducible nitric oxide synthase activity in spinal nerve roots and that nitric oxide synthase inhibition reduces nucleus pulposus‐induced edema and prevents reduction of nerve‐conduction velocity. Furthermore, the results suggest that nitric oxide is involved in the pathophysiological effects of nucleus pulposus in disc herniation.
Pamidronate prevents bone loss associated with carrageenan arthritis by reducing resorptive activity but not recruitment of osteoclasts
Authors: Moran Erica L. , Fornasier Victor L. , Bogoch Earl R.
Published: February 2005

Carrageenan arthritis is associated with high‐turnover bone loss. We sought to determine whether the bisphosphonate pamidronate can modify this effect of inflammatory arthritis. Sixty mature, New Zealand White rabbits were randomly assigned to five groups: normal with a therapeutic dose of pamidronate (300 μg/kg/day, administered subcutaneously): arthritis (induced in the right tibiofemoral joint with 10 intraarticular carrageenan injections); arthritis with a therapeutic dose of pamidronate (300 μg/kg/day, subcutaneous); and arthritis with a low dose of pamidronate (7.5 μg/kg/day, subcutaneous). All animals received the fluorochrome calcein green (0.5 g/l/day) in drinking water ad libitum from days 21 to 49. Undecalcified, transverse sections of the distal femur were photographed or imaged to determine bone volume; new bone volume; resting, eroded, osteoid, and osteoblast perimeters; and ostcoclast number. Results were evaluated with analysis of variance and pairwise Bonferroni's tests. In trabecular bone adjacent to the joint affected by carrageenan arthritis, resting perimeter was substantially reduced compared with normal joints, and primary indices of osteoblast and osteoclast activity were abnormally high (p < 0.001). Daily treatment with a therapeutic dose of pamidronate (300 μg/kg/day, subcutaneous) during the induction of arthritis significantly decreased new bone volume, osteoid perimeter, and osteoblast perimeter compared with the untreated arthritis group (p < 0.001). Osteoclast number and eroded perimeter remained abnormally high despite treatment with pamidronate. The concomitant increase of bone volume and these osteoclast indices show that pamidronate prevents bone loss in this model of experimental inflammatory arthritis by inhibiting the resorptive activity, but not the formation or recruitment, of osteoclasts. These findings are relevant to the use of bisphosphonates in the treatment of rheumatoid arthritis.
Viscoelastic properties of chondrocytes from normal and osteoarthritic human cartilage
Authors: Trickey Wendy R. , Lee Greta M. , Guilak Farshid
Published: February 2005

The deformation behavior and mechanical properties of articular chondrocytes are believed to play an important role in their response to mechanical loading of the extracellular matrix. This study utilized the micropipette aspiration test to measure the viscoelastic properties of chondrocytes isolated from macroscopically normal or end‐stage osteoarthritic cartilage. A three‐parameter standard linear solid was used to model the viscoelastic behavior of the cells. Significant differences were found between the mechanical properties of chondrocytes isolated from normal and osteoarthritic cartilage. Specifically, osteoarthritic chondrocytes exhibited a significantly higher equilibrium modulus (0.33 ± 0.23 compared with 0.24 ± 0.11 kPa), instantaneous modulus (0.63 ± 0.51 compared with 0.41 ± 0.17 kPa), and apparent viscosity (5.8 ± 6.5 compared with 3.0 ± 1.8 kPa‐s) compared with chondrocytes isolated from macroscopically normal, nonosteoarthritic cartilage. The elastic moduli and relaxation time constant determined experimentally in this study were used to estimate the apparent biphasic properties of the chondrocyte on the basis of the equation for the gel relaxation time of a biphasic material. The differences in viscoelastic properties may reflect alterations in the structure and composition of the chondrocyte cytoskeleton that have previously been associated with osteoarthritic cartilage. Coupled with earlier theoretical models of cell‐matrix interactions in articular cartilage, the increased elastic and viscous properties suggest that the mechanical environment of the chondrocyte may be altered in osteoarthritic cartilage.
Osteoprotegerin inhibits tumor‐induced osteoclastogenesis and bone tumor growth in osteopetrotic mice
Authors: Clohisy Denis R. , Ramnaraine Margaret L. , Scully Sheila , Qi Meiying , Van Gwyneth , Tan Hong Lin , Lacey David L.
Published: February 2005

Osteoprotegerin and osteoprotegerin ligand have recently been identified as novel proteins that inhibit and stimulate, respectively, osteoclast formation. We examined the possibility that osteoprotegerin would inhibit cancer‐induced osteoclastogenesis and cancer growth in bone. An experimental model was used in which osteolytic tumors are known to stimulate osteoclastogenesis and grow in femora of osteoclast‐deficient mice (op/op). Osteoprotegerin treatment decreased the number of osteoclasts by 90% (p < 0.0007) at sites of tumor in a dose‐dependent manner and decreased bone tumor area by greater than 90% (p < 0.003). The mechanisms through which osteoprotegerin decreased osteoclast formation in tumor‐bearing animals included (a) an osteoprotegerin‐mediated, systemic reduction in the number of splenic and bone marrow‐residing osteoclast precursor cells. (b) a decrease in the number of osteoclast precursor cells at sites of tumor as detected by cathepsin K and receptor activator of NFkB mRNA expression, and (c) a decrease in osteoprotegerin ligand mRNA at sites of tumor. These findings suggest that osteoprotegerin treatment, in addition to having direct antagonistic effects on endogenous osteoprotegerin ligand, decreases the number of osteoclast precursors and reduces production of osteoprotegerin ligand at sites of osteolytic tumor.
Reinsertion of stimulated nucleus pulposus cells retards intervertebral disc degeneration: An in vitro and in vivo experimental study
Authors: Okuma Masahiko , Mochida Joji , Nishimura Kazuhiro , Sakabe Kou , Seiki Kanji
Published: February 2005

Reinsertion of autogenous nucleus pulposus, an innovative method to delay further disc degeneration, has been proved with an experimental animal model. This study examined whether coculture of nucleus pulposus cells with annulus fibrosus cells (a) activates annulus fibrosus cells and (b) retards disc degeneration when reinserted into the disc in a rabbit model of disc degeneration. Coculture of the two cell types stimulated proliferation of each, as indicated by increased DNA synthesis measured by increases in DNA polymerase α expression and uptake of 5‐bromo‐2 deoxy‐uridine assessed by an enzyme‐linked immunosorbent assay. In a model of disc degeneration in rabbits, reinsertion of activated nucleus pulposus cells delayed the formation of clusters of chondrocyte‐like cells, the destruction of disc architecture, and the elaboration of type‐II collagen as measured immunohistochemically compared with no treatment. The direct reinsertion of activated nucleus pulposus cells into the disc offers a promising line of investigation for delaying intervertebral disc degeneration, although these results obtained with notochordal cells may not necessarily apply when mature central nucleus pulposus cells are used.
Mechanistic change in the reactivity of substituted phenyl acetates over phenyl thiolacetates toward imidazole in aqueous phase
Authors: Rajarathnam D. , Jeyakumar T. , Nadar P. Ananthakrishna
Published: February 2005

The kinetics of aminolysis of several substituted phenyl acetates by imidazole is studied in aqueous medium at 20°C and an ionic strength of 0.1 M (KCl). By following the leaving groups spectrophotometrically (λmax = 272–401 nm), under excess free imidazole, pseudo‐first‐order rate constants (kobs) are obtained. For the esters with good nucleofuges, the reaction follows clean second‐order kinetics and the plots of (kobs−kH) against free imidazole concentration are linear at constant pH. The macroscopic nucleophilic substitution rate constants (kN) are obtained as the slopes of these plots and found to be pH independent. For the esters with poor nucleofuges, a rate dependence on more than first power of the free imidazole and a linear dependence of k'2 on free imidazole is observed. Accordingly, the microscopic rate constants for the assisted paths viz. kga and kgb have been disseminated besides for simple bimolecular attack. The Brönsted‐type plots and Hammett plots were constructed whose slope values are consistent with a stepwise mechanism through a bipolar tetrahedral addition intermediate whose formation or breakdown being rate determining for various paths. Comparison of this reaction of oxyesters with the earlier reported works on similar reaction of analogue thiolesters under identical reaction conditions showed remarkable mechanistic differences which are discussed in detail. The discussion is extended to include the details on previously studied ammonolysis of these two types of esters wherein thiolesters showed differed reactivity than that reported in the present study. © 2005 Wiley Periodicals, Inc. Int J Chem Kinet 37: 211–221, 2005
Interactions in Nanosilica‐Polyurethane Composites Evidenced by Plate‐Plate Rheology and DMTA
Authors: Navarro‐Bañón Virtudes , Vega‐Baudrit José , Vázquez Patricia , Martín‐Martínez José Miguel
Published: February 2005

Fumed nanosilicas with different degree of silanization were used to obtain nanosilica‐polyurethane (TPU) composites to modify their rheological and mechanical properties. The higher the silanol content of the nanosilica, the higher the storage modulus and the thermal stability of the silica‐TPU composite. DMTA experiments confirmed that the TPU‐silica interactions increased by increasing the silanol content of the nanosilica.
Effect of the Amount of Calcium Carbonate as Filler on the Rheological and Adhesion Properties of a Water‐Based Polyurethane Dispersion
Authors: Muñoz‐Milán Ana Belén , Pérez‐Limiñana Ma Ángeles , Arán‐Aís Francisca , Torró‐Palau Ana , Orgilés‐Barceló A. César
Published: February 2005

In this study, the properties of water‐based adhesives based on a polyurethane ionomer (PUD) and a micronised CaCO3 as filler was analysed. Different amounts of a micronised CaCO3 (5‐25 wt%) were added to water‐based polyurethane (PUD) adhesive formulations in order to reduce its relatively high cost. The addition of a micronised calcium carbonate filler increased the viscosity, the storage and loss moduli of the PUD adhesives, and imparted pseudoplasticity and thixotropy, more noticeably for the adhesive with the highest calcium carbonate content. The creation of acid‐base interactions seemed to be responsible for the improvement in the rheological properties of the PUD adhesives containing CaCO3 as filler. On the other hand, the addition of CaCO3 filler might deteriorate the adhesion properties to PUD adhesives.
Poly(propylene)/PET/Undecyl Ammonium Montmorillonite Nanocomposites. Synthesis and Characterization
Authors: Velasco José Ignacio , Ardanuy Mónica , Miralles Lourdes , Ortiz Silvia , Maspoch Maria Lluisa , Sánchez‐Soto Miguel , Santana Orlando
Published: February 2005

A natural bentonite rich in calcium montmorillonite (CaMMT) was initially purified and ion‐exchanged to obtain sodium montmorillonite (NaMMT). Both clays were organophillised by cationic exchange reaction with undecyl‐ammonium chloride, and characterized. Isotactic poly(propylene) (PP) was melt‐compounded with both the unmodified and the organophilic montmorillonites. The hybrids produced have been characterized structurally, thermally and mechanically. Maleic anhydride‐grafted PP (MAH‐g‐PP) was used as compatibilizer in some of the formulations. Homologous series of hybrids were also synthesized employing blends of PP/PET and compared with those of the pure PP to investigate possible beneficial effects due to the presence of small amounts of PET on the microstructure and properties of this kind of materials. The analysis of the results indicates some extension of both macromolecules intercalation and clay particles exfoliation in the hybrids prepared with the organophilic montmorillonite. The hybrids prepared with compatibilized PET/PP blends were found to have a better nanostructure.
Structure/Property Relationships for Polyamide 6 / Organoclay Nanocomposites in the Melt and in the Solid State
Authors: Karaman Vitaly M. , Privalko Valery P. , Privalko Eleonora G. , Lehmann Bjorn , Friedrich Klaus
Published: February 2005

Commercial nanocomposites of polyamide 6 prepared by melt compounding with organoclays were characterized by complex viscosities and relaxation time spectra derived from storage and loss shear moduli measured in the melt state at 230°C, and by wide‐ and small‐angle X‐ray scattering, differential scanning calorimetry and stretching calorimetry in the solid state.

In the melt state, the decrease of ∼ 25 % (compared to the pristine sample) in Newtonian viscosity at the lowest clay loading (2.5 %) suggested a lower equilibrium elasticity modulus of an entangled melt, as the small amounts of organoclay nanoparticles acted as specific “diluents” for the initial entanglement network. However, by increasing the clay loading this effect disappeared due to the importance of strong interactions at the nanoparticle/melt interface, leading to the formation of a fairly thick boundary interphase (BI) around the nanoparticles and ending up in the build‐up of an “infinite cluster” of clay nanoparticles coated with BI at the highest (albeit still unusually low) clay loading (7.5 %).

In the solid state, organoclay nanoparticles proved to induce the crystallographic α → γ transformation of PA6 , while the matrix crystallinity in nanocomposites remained essentially unchanged. In the range of elastic (reversible) behavior below the apparent yield strains ϵ* , the highest Young's moduli E and the lowest linear thermal expansion coefficients αL were observed for dried nanocomposites, while the lowest E and the highest αL corresponded to the moisturized, pristine PA6. The endothermal process of shape distortion of the lamellar crystals was assumed to precede the onset of the exothermal process of lamellar fragmentation in the range of inelastic (irreversible) behaviour of PA6 above ϵ* . The energy balance of the inelastic behaviour of nanocomposites was dominated by the endothermal process of lamellar shape distortion.
Structure Analysis of PVC Nanocomposites
Authors: Kovarova Lucie , Kalendova Alena , Gerard Jean‐Francois , Malac Jiri , Simonik Josef , Weiss Zdenek
Published: February 2005

In this paper poly(vinyl chloride)/clay nanocomposites were prepared by melt intercalation using a single screw extruder. Problems with thermal stability of these nanocomposites during compounding were largely eliminated by pre‐treatment of the organoclay with plasticizer (dioctyl phthalate), which created a barrier between polymer and quaternary amine. These nanocomposite materials were analyzed with respect to their morphology. The intercalation, exfoliation, nano‐phase dispersion and orientation were investigated using Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM) and X‐ray diffraction (XRD). Moreover, different types of sample preparation for these techniques were tested as well. It was found that partially intercalated and disordered structure arose in poly (vinyl chloride) composites containing sodium type of montmorillonite, while a fine dispersion of partial to nearly full exfoliation of individual montmorillonite layers in poly (vinyl chloride) matrix was observed when this clay was organically modified. Finally, the influence of different mixing time (in extruder) on nano‐phase morphology was surveyed.
Preparation, Characterisation and Properties of Nanocomposites Based on Epoxy Resins – An Overview
Authors: Hartwig Andreas , Sebald Monika , Pütz Dirk , Aberle Lisa
Published: February 2005

An overview about the use of nanoparticles in epoxy resins is given. Deaggregated fumed silica, sol gel materials and other spheroidic nanoparticles improved the abrasion resistance and the mechanical properties of filled epoxides, and several properties can be improved by specific particles. Combustion properties, strength and permeation can be improved by organically modified layered silicates. The proper characterisation of the nanocomposites is still an art. TEM analysis of the cured materials and light scattering with 3D cross‐correlation for the liquid samples were sufficient methods for the characterisation of filled epoxides.
Predictive Models of Ethylene‐Vinyl Acetate (EVA) Copolymers with Powdered Zn Fillers
Authors: Balart Rafael , Parres Francisco , López Juan , García David
Published: February 2005

Metallic Zn filler was used to increase the electrical properties of ethylene‐vinyl acetate copolymer (EVA). Other properties were also modified such as mechanical and thermal behavior. It is possible to predict the behaviour of EVA‐Zn system by using different models.
Dynamic Mechanical Behavior of PP/PET/MAPP Blends Filled with Glass Beads
Authors: Rodríguez‐Pérez M.A. , Velasco J.I. , González‐Peña J.I. , Ruiz‐Herrero J.L. , Arencon D. , de Saja J.A.
Published: February 2005

PP/PET/MAPP blends have been filled with 50 wt% of glass beads. The orientation of the PP crystalline phase, the crystallization behavior and the dynamic mechanical response of these materials have been analyzed. The dynamic mechanical response is strongly affected by the presence of the glass beads, being possible to detect the effect of PET and MAPP on the storage modulus and loss factor values. Moreover, the alpha relaxation of the composites is visibly affected by thermal treatments.
Rainbow trout has two genes for growth hormone
Authors: Agellon L. B. , Davies S. L. , Lin C.‐M. , Chen T. T. , Powers D. A.
Published: February 2005

We report the primary structures of two mRNA species (GH1 and GH2), each predicted from the cloned cDNA and genomic gene sequences, that encode growth hormone in rainbow trout (Salmo gairdneri). Both GH1 and GH2 mRNA contain open reading frames comprising 630 nucleotides and encode 210 amino acid residues, of which 11 are variant. The translated regions of mRNA are flanked by a short 5′‐untranslated sequence, which is highly conserved, and a relatively long 3′‐untranslated sequence, which is highly divergent. The differences at the 3′‐untranslated regions suggest that the GH1 and GH2 mRNA originate from different loci. RNA blot analysis of trout pituitary RNA using an oligonucleotide probe specific for the GH2 sequence indicates that the cloned gene is expressed. The GH1 and GH2 mRNA likely are transcribed from two distinct loci, which were duplicated during tetraploidization of the salmonid genome between 50 and 100 million years ago.
Protein phosphorylation in meiotically competent and incompetent mouse oocytes
Authors: Bornslaeger Elayne A. , Mattei Peter M. , Schultz Richard M.
Published: February 2005

Specific changes in the two dimensional gel electrophoretic pattern of mouse oocyte phosphoproteins precede germinal vesicle breakdown (GVBD). We report that changes in the relative abundance of phosphoamino acids occurred prior to GVBD. We also report data that further strengthen the close association of the changes in phosphoprotein patterns with resumption of meiosis. The calmodulin antagonist W7, which transiently inhibits GVBD, inhibited partially at least two of the maturation‐associated phosphoprotein changes, the dephosphorylation of a 60,000 Mr 24,000 and 28,000 occurred; all these changes occurred, however, in oocytes from juvenile mice that were competent to resume meiosis. The microinjection of the heat‐stable inhibitor of cyclic adenosine monophosphate (cAMP)‐dependent protein kinase (PKI), which induces GVBD in fully grown oocytes, did not induce GVBD in meiotically incompetent oocytes. Microinjected PKI did not induce the increased protein phosphorylations associated with maturation, but it did induce the dephosphorylation of the 60,000 Mr phosphoprotein. These results provide molecular markers for commitment to resume meiosis in GV‐intact oocytes and indicate a potential basis for meiotic incompetence.
Changes in polyadenylation of lactate dehydrogenase‐X mRNA during spermatogenesis in mice
Authors: Fujimoto Hirokazu , Erickson Robert P. , Toné Shigenobu
Published: February 2005

The expression of the mRNA for mouse testicular lactate dehydrogenase (LDH‐X) was examined by Northern analyses of meiotic and postmeiotic spermatogenic cell populations. Silver grains accumulated in cells inside the second layer from the periphery of the seminiferous tubule, confirming previous findings that LDH‐X mRNA first appears in the spermatocyte and continues to accumulate until the late spermatid stage. Northern analyses showed that meiotic and postmeiotic cells contained 1.2 and 1.3 kb classes of hybridizing mRNA, respectively. RNase H digestion of oligo(dT)‐hybridized RNA and poly(U)‐Sepharose column chromatography with differential elution by formamide revealed that the difference in size of the two classes of mRNAs was due to the poly(A) tail length of the LDH‐X mRNA. When the distribution of the LDH‐X mRNA was examined across polysome gradients, both mRNAs were partially associated with polysomes. These results suggest that the changes in the polyadenylation of LDH‐X mRNA were associated with the meiotic division during spermatogenesis in the mouse. They raise the possibility that the stable accumulation of the LDH‐X mRNAs in the postmeiotic cells is enhanced by poly(A) tails of increased length.
Sexing the human fetus and identification of polyploid nuclei by DNA–DNA in situ hybridisation in interphase nuclei
Authors: West John D. , Gosden Christine M. , Gosden John R. , West Katrine M. , Davidson Zoe , Davidson Catherine , Nicolaides Kypros H.
Published: February 2005

Samples of human adult lymphocytes, fetal lymphocytes, amniotic fluid cells, and chorionic villus cells were sexed independently by cytogenetics and DNA–DNA in situ hybridisation to a tritiated Y probe. For the in situ hybridisation analysis, the presence of Y bodies (hybridisation bodies) in 100 interphase nuclei were scored after autoradiography. In all, 82/83 samples were sexed in this way (one technical failure) and 78/82 were sexed by both in situ hybridisation and cytogenetics. There was complete agreement between the two methods. There was a considerable variation (40–100%) in the percentage of interphase nuclei with a hybridisation body among the male samples, but very few nuclei from female samples showed significant hybridisation. In situ hybridisation could be used to sex the conceptus when males but not females are at risk for various X‐linked genetic disorders and may also be useful for detecting 45,X/46,XY mosaicism or polyploid/diploid mosaicism. This would be particularly useful for direct preparations of chorionic villus samples, which often prove difficult to analyse cytogenetically but offer the best means of avoiding maternal contamination. Some interphase nuclei had more than one hybridisation body, and this was most commonly found among amniotic fluid cells. Comparison of sizes of nuclei with one or two hybridisation bodies strongly suggested that most of the amniotic fluid cell nuclei with two hybridisation bodies were tetraploid.
The use of DNA probes in preimplantation and prenatal diagnosis
Authors: West John D.
Published: February 2005

DNA probes are now widely used for prenatal diagnosis, but the prospect of preimplantation diagnosis of genetic disorders requires the development of sensitive genetic tests that can be performed on small numbers of cells removed from a preimplantation‐stage pre‐embryo. The sensitivity of molecular tests can now be increased by specifically amplifying the target DNA with the polymerase chain reaction. In situ hybridisation with chromosome‐specific DNA probes to repeated sequences also permits the detection of particular numerical chromosome aberrations or the distinction of male and female pre‐embryos when only a few interphase nuclei are available. We have used in situ hybridisation to a Y chromosome‐specific DNA probe to sex preimplantation‐stage pre‐embryos and to sex fetuses from samples of chorionic villus cells, amniotic fluid cells, and fetal blood. These two approaches (amplification of target DNA and in situ hybridisation) provide suitable tests for improving prenatal diagnosis particularly when few cells are available and they offer the possibility of tests suitable for preimplantation diagnosis.
Expression of two actin genes during larval development in the sea urchin Strongylocentrotus purpuratus
Authors: Cameron R. Andrew , Britten Roy J. , Davidson Eric H.
Published: February 2005

We report the first measurements of cell number, total RNA, and transcript accumulations for two actin genes during larval development of the sea urchin Strongylocentrotus purpuratus. At 5 weeks of feeding, when development of laboratory‐raised larvae is completed, the cell number has increased about 100‐fold with respect to the pluteus‐stage embryo to about 150,000 ± 50,000, and the total RNA has increased 46‐fold to about 130 ng per larva. The transcripts of the Cylla cytoskeletal actin gene, which is expressed in adult tissues, continue to accumulate throughout larval development. A contrasting pattern of transcript accumulation is observed for Cyllla, a different cytoskeletal actin gene that in the embryo is expressed only in aboral ectoderm. These transcripts increase in number early in larval development, when the larval epidermis is differentiating, and then decline in quantity. It is known that at metamorphosis the larval epidermis is largely histolyzed and that the Cyllla gene is not expressed in the juvenile or adult.
Expression and struture of the CyIIIb actin gene of the sea urchin Strongylocentrotus purpuratus
Authors: Flytzanis Constantin N. , Bogosian Elizabeth A. , Niemeyer Christina C.
Published: February 2005

The developmental pattern of expression of the Strongylocentrotus purpuratus Cylllb actin gene was determined by RNA blot hybridizations carried out with a gene‐specific probe and total embryonic RNA isolated from various stages of development. The results indicate that the Cylllb mRNA is not detected in the maternal pool, and, although the gene is activated at the early stages (about 10 hr postfertilization), considerable amounts of mRNA do not accumulate until well into the pluteus stage 3 days later. These results suggest either a post‐transcriptional regulatory mechanism that governs early embryonic expression of the Cylllb actin or a late embryonic transcriptional enhancement of this gene. We present here the complete nucleotide sequence of the Cylllb gene, which lies within the 10,361 base pairs of the sequenced region. The entire transcription unit is 7,455 nt long and shares structural similarities with the other cytoskeletal‐type actin genes from this sea urchi. Sequence comparisons of Cylllb to the Cyllla actin gene, to which it is linked, reveals extensive homology even in the introns. The deduced amino acid sequence of the Cylllb actin shows five amino acid substitutions compared with the Cyllla actin and nine when compared with the Cyl, the endodermal embryonic cytoskeletal‐type actin. Five out of these nine amino acid differences occur within a small peptide (position 257 to 267). The 5′ flanking sequence of the Cylllb gene shows a remarkable homology (∼75–80%) with the Cyllla upstream region up to the position −200 and a lack of any obvious similarity further upstream. This observation suggests that the two genes possibly share some common regulatory factors.
Differential regulation of protein biosynthesis at translational level in the fat body cells of adult Locusta migratoria
Authors: Glinka A. V. , Triseleva T. A.
Published: February 2005

Vitellogenin (Vg) and lipophorin (Lp) are synthesized by the fat body of adult locust (Locusta migratoria) females. We have shown by an immunohistochemical technique that both proteins are produced in the same cells of the fat body. The rate of Vg synthesis was measured with the use of double immunoprecipitation of labeled proteins at oviposition and 24 h later. It was found that the rate of Vg synthesis declined significantly by the time of oviposition; however, 24 h later, it was raised to the highest possible level. The rate of Lp synthesis remained constant at both indicated points. The similar postlaying increase in the Vg synthesis rate was observed in the fat bodies of females treated by α‐amanitin immediately after oviposition. The data provide evidence that Vg biosynthesis in L. migratoria is regulated by selective periodical repression and derepression of Vg mRNAs in the fat body cells but not by total inhibition and stimulation of protein‐synthesizing machinery.
Flow cytometric analysis of rodent epididymal spermatozoal chromatin condensation and loss of free sulfhydryl groups
Authors: Evenson D. P. , Baer R. K. , Jost L. K.
Published: February 2005

Flow cytometric measurements were made on acridine orange (AO) and 7‐diethylamino‐3‐(4′‐maleimidylphenyl)‐4‐methylcoumarin (CPM)‐stained epididymal‐ and vas deferens‐derived spermatozoal nuclei to follow the course of chromatin condensation and oxidation of free sulfhydryl groups, respectively, during passage through mouse and rat posttesticular reproductive tracts. Alterations of mouse and rat spermatozoal chromatin during transition from a testicular elongated spermatids to epididymal caput spermatozoc resulted in a threefold loss of DNA stainability with AO. Passage of spermatozoa from the caput to corpus epididymis was accompanied by an approximate 15% loss of DNA stainability, which was maintained at that level throughout passage into the vas deferens. AO stainability of epididymal spermatozoal nuclei was generally independent of –SH group stainability. CPM stanability of rat spermatozoal nuclei free –SH groups was 83%, 18%, and 11% of caput spermatozoal values for corpus, cauda epididymis, and vas deferens, respectively. Comparable values for mice were 69%, 20%, and 18%. CPM stainability was relatively homogeneous for these mouse and rat reproductive tract regions, except mouse corpus epididymis spermatozoal nuclei stained very heterogeneously. Rat spermatozoa detained by ligature up to 7 days in the caput, corpus, and cauda epididymi had CPM staining values equal to or below those of normal vas spermatozoa, indicating that disulfide (S‐S) bonding is intrinsic to the spermatozoa and is independent of the epididymal environment. These data suggest that chromatin condensation and loss of spermatozoal DNA stainability during passage from the testis to the vas deferens are independent of S‐S bonding. Furthermore, the results are in agreement with previous findings suggesting that autoxidation of SH groups occurs independently of movement through the epididymis.
Different cytoskeletal organization in two maturation stages of Discoglossus pictus (anura) oocytes: Thickness and stability of actin microfilaments and tropomyosin immunolocalization
Authors: Campanella Chiara , Chaponnier Christine , Quaglia Lucia , Gualtieri Roberto , Gabbiani Giulio
Published: February 2005

In Discoglossus pictus oocytes, the germinative area (GA) contains long and irregular microvilli where actin microfilaments are located. In the egg, the funnel‐shaped dimple that originates by invagination of the GA is present. In the dimple both microvilli and microfilament bundles have a very orderly appearance. This report extends previous observations (Campanella and Gabbiani, Gamete Res 3:99–114, 1980) and shows that GA microfilaments are thinner (36 Å average) than dimple microfilaments (60 Å average), as measured in ultrathin section. Moreover, the interfilament distance is smaller in GA bundles than in the dimple bundles. To get an insight into actin‐depolymerizing factor (ADF) in which cryostat sections were incubated prior to immunofluorescent staining with antiactin antibodies. The microfilaments of the GA microvilli and partially of the oocyte cortex are resistant to ADF when compared to those in the dimple and the rest of the egg cortex. We also investigated immunocytochemically the presence of tropomyosin and found that this protein is localized in the dimple and in the cortex of oocytes and eggs but is absent in the GA.
Bovine oviductal fluid components and their potential role in sperm cholesterol efflux
Authors: Ehrenwald Eduardo , Foote Robert H. , Parks John E.
Published: February 2005

Bovine oviductal fluid (OF) was collected and analyzed throughout the estrous cycle, and the capacity of the protein and lipoprotein components to support cholesterol efflux from bovine sperm was evaluated. Blood was collected and assayed for progesterone (P4) to monitor the estrous cycle. Protein and lipoprotein separation was achieved by density gradient centrifugation. Two major bands were identified. The first (1.056 < ∂20 < 1.140 g/ml) corresponded to bovine and rabbit plasma high‐density lipoprotein (HDL) based on distribution in the density gradient and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. The second band (1.235 < ∂20<1.243 g/ml) consisted predominantly of oviductal fluid albumin (OFA). Oviductal fluid protein concentration increased as serum P4 decreased around the time of estrus. Mean OF protein concentration was 21.3 mg/ml when serum P4 was lower than 0.5 ng/ml and 6.9 mg/ml when serum P4 was greater than 0.5 ng/ml. An inverse log relationship was found between HDL protein concentration and serum P4. Unesterified cholesterol (UC), cholesteryl ester, and phospholipid (PL) content of HDL for HDL protein concentrations of 3–56.1 μg/ml were 1.35–46.2 μg/ml, 1.91–44.48 μg/ml, and 1.69–59.8 μg/ml, respectively. Phosphatidylcholine and ‐ethanolamine were the major PLs present in the HDL fraction and their molar ratio (4:1 mol/mol) was relatively constant through the estrous cycle. The OFA fraction of the same samples accounted for more than 90% of total protein and for most of the variation in OF protein. To determine the ability of OF components to serve as sperm cholesterol acceptors, OF samples were incubated 1:1 (v/v) with and without 4 × 108 bovine sperm in 1.0 ml of modified Tyrode's solution and OF for 2 hr at 39°C. After incubation, HDL and OFA fractions were isolated and analyzed for changes in protein and lipid content. After OF, samples were incubated with sperm, an increase in UC was found in the HDL fractions. UC in HDL increased by 12.1 ± 1.0 μg/ml (X ± SE) when serum P4 was ≤0.5 ng/ml. For samples corresponding to higher serum P4, the increase in UC was 3.60 ± 0.89 μg/ml. Values for UC in HDL were corrected for the contribution of UC from OFA of OF samples. Cholesterol efflux from sperm has been implicated in the process of sperm capacitation. These results indicate that HDL from OF is elevated during the follicular phase of the estrous cycle and can serve as an acceptor for bovine sperm cholesterol.
Molecular cloning of an acrosomal sperm antigen gene and the production of its recombinant protein for immunocontraceptive vaccine
Authors: Liu Ming‐Sun , Chan Kaimin , Lau Yun‐Fai , Lee Chi‐Yu Gregory
Published: February 2005

A monoclonal antibody, HS‐63, which reacts specifically with a highly conserved sperm acrosome antigen, was shown to inhibit in vitro fertilization of mouse and human. The corresponding sperm antigen designated as MSA‐63 was purified to homogeneity from mouse testes and used as an immunogen to generate polyclonal antisera in rabbits. The cDNA fragments of MSA‐63 gene were cloned from mouse testis cDNA library by an immunoscreening method using polyclonal antisera specific for MSA‐63. Using the established cDNA clone as a probe, the gene encoding for MSA‐63 protein was found to be conserved among different mammalian species. Only one specific mRNA 1.5 kb in size was identified from the adult mouse testis among different mouse tissues. The recombinant fusion protein containing MSA‐63 protein fragment was produced in Escherichia coli and used to immunize female mice. Similar to the original HS‐63 monoclonal antibody, the antisera thus produced reacted only with the sperm acrosome and revealed significant inhibition to the in vitro fertilization of mouse oocytes. The results of this preliminary study suggest that it is feasible to mass produce sperm‐specific antigens or their antigenic fragments by recombinant DNA technology for the development of sperm antigen‐based immunocontraceptive vaccines.