Age-related mechanical work expenditure during normal walking: The Baltimore Longitudinal Study of Aging

The aim of this cross-sectional study was to delineate age-associated kinematic and kinetic gait patterns of normal walking, and to test the hypothesis that older adults exhibit gait patterns that reduce generative mechanical work expenditures (MWEs). We studied 52 adult Baltimore Longitudinal Study of Aging participants (means age 72±9, from 60 to 92 years) who could walk 4 m unaided. Three-dimensional kinematic and kinetic parameters assessed during rotation-defined gait periods were used to estimate MWEs for the rotation of lower extremities about the medial–lateral (ML) and anterior–posterior (AP) axes of proximal joints, which represent MWEs in the AP and ML sides, respectively. Relationships between gait parameters and age were examined using regression analysis with adjustments for walking speed, sex, height, and weight. Older age was associated with slower self-selected walking speed (p<0.001), shorter stride length (p<0.001), and greater propensity of landing flat-footed (p=0.003). With older age, hip generative MWE for thigh rotation was lower about the AP axis (hip abduction and adduction) during stance (p=0.010) and higher about the ML axis (hip extension and flexion) during late stance (p<0.001). Knee absorptive MWE for shank rotation about the AP axis (knee abduction and adduction) during early stance was also lower with older age (p<0.003). These age-related gait patterns may represent a compensatory effort to maintain balance and may also reflect mobility limitations.     

A novel muscle biopsy clamp yields accurate in vivo sarcomere length values

The measurement of in vivo muscle sarcomere length facilitates the definition of in vivo muscle functional properties and comparison of muscle designs amongst functional muscle groups. In vivo sarcomere lengths are available for just a handful of human muscles, largely due to the technical challenges associated with their measurement. The purpose of this report was to develop and test a muscle biopsy clamp that can quickly and accurately measure in vivo muscle sarcomere length. To test the device, muscle biopsies (n=23) were removed from the tibialis anterior muscles of New Zealand White rabbits immediately after sarcomere length measurements were made using laser diffraction. The muscle biopsy contained within the clamp was immediately fixed in Formalin for subsequent sarcomere length measurement. Comparisons of clamp-based and diffraction-based sarcomere lengths demonstrated excellent agreement between the two techniques, especially when the biopsy was obtained at relatively long lengths (above 2.6 μm). Given the intraoperative speed and simplicity of this technique and the relatively low-cost of the biopsy clamp, this method of measuring muscle sarcomere length should help investigators generate much-needed in vivo muscle structural and functional data.     

IGFBP-3 Can Either Inhibit or Enhance EGF-mediated Growth of Breast Epithelial Cells Dependent upon the Presence of Fibronectin

Progression of breast cancer is associated with remodeling of the extracellular matrix, often involving a switch from estrogen dependence to a dependence on EGF receptor (EGFR)/HER-2 and is accompanied by increased expression of the main binding protein for insulin-like growth factors (IGFBP-3). We have examined the effects of IGFBP-3 on EGF responses of breast epithelial cells in the context of changes in the extracellular matrix. On plastic and laminin with MCF-10A normal breast epithelial cells, EGF and IGFBP-3 each increased cell growth and together produced a synergistic response, whereas with T47D breast cancer cells IGFBP-3 alone had no effect, but the ability of EGF to increase cell proliferation was markedly inhibited in the presence of IGFBP-3. In contrast on fibronectin with MCF-10A cells, IGFBP-3 alone inhibited cell growth and blocked EGF-induced proliferation. With the cancer cells, IGFBP-3 alone had no effect but enhanced the EGF-induced increase in cell growth. The insulin-like growth factor-independent effects of IGFBP-3 alone on cell proliferation were completely abrogated in the presence of an EGFR, tyrosine kinase inhibitor, Iressa. Although IGFBP-3 did not affect EGFR phosphorylation [Tyr¹⁰⁶⁸], it was found to modulate receptor internalization and was associated with activation of Rho and subsequent changes in MAPK phosphorylation. The levels of fibronectin and IGFBP-3 within breast tumors may determine their dependence on EGFR and their response to therapies targeting this receptor.     

Automated recognition of malignancy mentions in biomedical literature

Background  

The rapid proliferation of biomedical text makes it increasingly difficult for researchers to identify, synthesize, and utilize developed knowledge in their fields of interest. Automated information extraction procedures can assist in the acquisition and management of this knowledge. Previous efforts in biomedical text mining have focused primarily upon named entity recognition of well-defined molecular objects such as genes, but less work has been performed to identify disease-related objects and concepts. Furthermore, promise has been tempered by an inability to efficiently scale approaches in ways that minimize manual efforts and still perform with high accuracy. Here, we have applied a machine-learning approach previously successful for identifying molecular entities to a disease concept to determine if the underlying probabilistic model effectively generalizes to unrelated concepts with minimal manual intervention for model retraining.     

Reliability of the proximal skin paddle of the osteocutaneous free fibula flap: a prospective clinical study

The vascularization of the skin paddle of 20 osteocutaneous fibula free flaps in 20 patients was studied. All skin paddles were designed over the proximal and middle third of the fibula. A parallel vascularization of the skin was found in 10 cases. In these cases, an axial (septo)musculocutaneous perforator was found to originate high in the peroneal artery or even in the popliteal artery. This branch runs parallel to the peroneal artery without any further connections with it. In 5 of these 10 cases, no other skin perforators were located within the boundaries of the skin paddle. Harvesting such a flap in the traditional way by blind inclusion of a muscle cuff results in ligation of the supplying vessel of the skin paddle and subsequent loss of the skin. In this series, this would have been the case in 5 of the 20 patients (25 percent). This might explain the bad reputation of the skin paddle of this flap. The high prevalence of the described vascular configuration in a proximally designed skin paddle justifies à vue dissection of all musculocutaneous perforators up to their origin, unless one or more septocutaneous perforators are found within the boundaries of the flap.     

Determination of the triplet state energies of a series of conjugated porphyrin oligomers.

We report a systematic study of the photophysical parameters relevant to photodynamic therapy (PDT) by a new type of sensitizers, conjugated porphyrin oligomers. Due to the strong nonlinear properties of oligomers containing 2, 4 and 8 porphyrin units, these molecules are attractive candidates for PDT via multiphoton excitation. The triplet state energy levels for all molecules have been determined by the triplet quenching method, phosphorescence measurements and DFT calculations. We find that the triplet energies of all the oligomers are sufficient to generate singlet oxygen, >94 kJ mol(-1). However, low singlet oxygen quantum yields are observed for the tetramer and the octamer, as compared to the conjugated dimer and monomeric porphyrin, reflecting the decrease in triplet yield. Thus the conjugated porphyrin dimer is the most promising core structure for PDT applications via multiphoton excitation.     

Detecting protein-protein interactions in the intact cell of Bacillus subtilis (ATCC 6633)

The salt bridge, paired group-specific reagent cyanogen (ethanedinitrile; C(2)N(2)) converts naturally occurring pairs of functional groups into covalently linked products. Cyanogen readily permeates cell walls and membranes. When the paired groups are shared between associated proteins, isolation of the covalently linked proteins allows their identity to be assigned. Examination of organisms of known genome sequence permits identification of the linked proteins by mass spectrometric techniques applied to peptides derived from them. The cyanogen-linked proteins were isolated by polyacrylamide gel electrophoresis. Digestion of the isolated proteins with proteases of known specificity afforded sets of peptides that could be analyzed by mass spectrometry. These data were compared with those derived theoretically from the Swiss Protein Database by computer-based comparisons (Protein Prospector; http://prospector.ucsf.edu). Identification of associated proteins in the ribosome of Bacillus subtilis strain ATCC 6633 showed that there is an association homology with the association patterns of the ribosomal proteins of Haloarcula marismortui and Thermus thermophilus. In addition, other proteins involved in protein biosynthesis were shown to be associated with ribosomal proteins.     

Chloride Channels in Basolateral TAL Membranes. XVIII. Phenylglyoxal Induces Functional mcClC-Ka Activity in Basolateral MTAL Membranes

Cultured mouse MTAL cells contain more mRNA encoding the Cl^– channel mcClC-Ka, which mediates CTAL Cl^– absorption, than mRNA encoding the Cl^– channel mmClC-Ka, which mediates MTAL Cl^– absorption. mmClC-Ka and mcClC-Ka have three functional differences: 1) mmClC-Ka open time probability, P _o, increases with increasing cytosolic Cl^–, but variations in cytosolic Cl^– do not affect P _o in mcClC-Ka; 2) mmClC-Ka is gated by (ATP + PKA), while (ATP + PKA) have no effect on P _o in mcClC-Ka; and 3) mmClC-Ka channels have single-ion occupancy, while mcClC-Ka channels have multi-ion occupancy. Using basolateral vesicles from MTAL cells fused into bilayers, we evaluated the effects of 1 mM cytosolic phenylglyoxal (PGO), which binds covalently to lysine or arginine, on Cl^– channels. With PGO pretreatment, Cl^– channels were uniformly not gated either with increases in cytosolic-face Cl^– or with (ATP + PKA) at 2 mm cytosolic-face Cl^–; and they exhibited multi-ion occupancy kinetics typical for mcClC-Ka channels. Thus, in basolateral MTAL membranes, blockade of Cl^– access to arginine or lysine residues on mmClC-Ka by PGO results in Cl^– channels having the functional characteristics of mcClC-Ka channels.     

Plasmid DNA purification by selective calcium silicate adsorption of closely related impurities

The selective adsorption of supercoiled plasmid, open-circular plasmid, and genomic DNA to gyrolite, a compound from the class of crystalline calcium silicate hydrates, is investigated and exploited for purification purposes. Genomic DNA and open-circular plasmid bind to gyrolite adsorbents with greater affinity than the more conformationally constrained supercoiled plasmid. As such, the gyrolite adsorbents are an economical and scaleable alternative to chromatographic purification for the removal of DNA impurities from solutions containing supercoiled plasmid. The advantage of gyrolite adsorbents is their lower unit price and ability to selectively adsorb DNA impurities without binding supercoiled plasmid under certain conditions. The effects of ionic strength, temperature, chelating agent, divalent cation, and lyotropic salts on adsorption of highly purified plasmid are studied to understand the forces that bind DNA to gyrolite, a structure with hydrophilic and hydrophobic characteristics. The results indicate that DNA binding is governed by hydrogen bonding, electrostatic bridging with divalent cations, shielding of electrostatic repulsion, hydrophobic adsorption, and disruption of integral surface water layer on gyrolite. On the basis of results from a range of Hofmeister series salts, strongly hydrated anions may enhance DNA adsorption by promoting hydrophobic interactions between DNA and gyrolite. Conversely, the very weakly hydrated chaotrope I(-) may enhance adsorption by strongly associating with hydrophobic siloxanes of gyrolite, thereby disrupting an integral water layer, which competes for hydrogen bonding sites.     

Identification of amino acid residues participating in intermolecular salt bridges between self-associating proteins

Salt bridges between self-associating hen egg white (HEW) lysozyme and bovine insulin molecules were converted to covalent links by ethanedinitrile (cyanogen) and identified using mass spectrometry. Peptides resulting from cyanogen-mediated intermolecular cross-linking of HEW lysozyme were detected using in-gel digestion and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Sequence data from electrospray ionization-quadrupole-time of flight mass spectrometry (ESI-Q-TOF MS) revealed that one of the peptides has a covalent bond between Asp 66 and Arg 14. The self-assembly of bovine insulin was also investigated using cyanogen. Using high-performance liquid chromatography (HPLC) coupled with ESI-Q-TOF MS, an intermolecular salt bridge association was identified by covalently linking the B chain C-terminal carboxyl group of Ala 30 and the charged imidazole of His 5 (B chain). A method was developed incorporating cyanogen, enzymatic digestion, one-dimensional gel electrophoresis, MALDI-TOF MS, and HPLC ESI-Q-TOF MS to identify amino acid residues participating in salt bridge formations at protein-protein interfaces. The novelty of this approach is the ease with which cyanogen can be administered to a protein sample and the apparent lack of nonspecific cross-linking side reactions interfering with the analysis.