An optical method for determining deformation and form changes in polyethylene surfaces of explanted acetabular cups of hip endoprostheses]

AIM: Most methods used for the determination of volumetric wear of polyethylene cups are based on the assumption that the head of the prosthesis penetrates the cup in "cylindrical" fashion. The new accurate optical method is independent of this disputable assumption. METHOD: The articulating surface of the cup is scanned with light and a data set of 60,000 pixels obtained in this way is stored in a computer. Data obtained from used cups were compared with those obtained from unused cups. The volumetric wear was calculated directly by threefold integration. To assess the changes in surface shape, the data are fitted by an ellipsoid whose long axis defines the mean direction of load. A total of 18 retrieved and 3 unused cups of different types were studied. RESULTS: The unused acetabular cups deviated only slightly from ideal hemispheres. The surfaces showed rotational symmetry, and an undulation having an amplitude of 0.1 mm between dome and equator. For all explanted cups, the assumption of cylindrical penetration of the head into the polyethylene was shown not to represent the true situation. The cup expands in all directions, and the volumetric wear is underestimated by 50% with the traditional methods. The data suggest that long-term survival may be jeopardized when the main direction of loading is centered on the dome of the cup. Ceramic heads were associated with smaller rates of volumetric wear. CONCLUSION: The new optical method is characterised by short measuring times, precision and simple application. Analysis of the wear patterns of polyethylene components using this technique may contribute to a further understanding of the complex mechanisms of aseptic loosening.     

Spatial micromovements of uncemented femoral components after torsional loads

A novel method is presented which permits to assess implanted femoral components with regards to location of fixation and initial stability under cyclic torsional loads. The measurement apparatus tracks the spatial movement at several sites of stem and bone, allowing quantitative analysis of the micromotions and twisting of stem and bone, and the location of torque (force) transfer. Four types of prostheses were compared, which revealed striking differences in torque transfer. Our results for synthetic femurs are consistent with in vivo data on the osseointegration and radiolucensies observed for the stem types of this study. The method can be used to quantitatively compare various stem designs and implantation techniques.     

Modeling of the controlled expression of a harmful protein by a three-plasmid harboring system

A genetically structured mathematical model was developed and used to evaluate the influence of molecular parameters involved in the expression of a harmful recombinant protein (SPA::EcoRI). The system consists of the controlled expression of the endonuclease EcoRI cloned in the plasmid pMTC48. The control is exerted by the lambda CI repressor expressed from the plasmid pRK248cIts. The deleterious effect of the activity of the enzyme EcoRI on the host DNA is prevented by the action of the EcoRI methylase that is expressed constitutively from a third plasmid, pEcoR4. The model includes molecular mechanisms involved in the regulation of the expression of these genes and is used to determine cultural conditions that maximize the production of the recombinant protein.     

Bidirectional activity-dependent morphological plasticity in hippocampal neurons

Dendritic spines on pyramidal neurons receive the vast majority of excitatory input and are considered electrobiochemical processing units, integrating and compartmentalizing synaptic input. Following synaptic plasticity, spines can undergo morphological plasticity, which possibly forms the structural basis for long-term changes in neuronal circuitry. Here, we demonstrate that spines on CA1 pyramidal neurons from organotypic slice cultures show bidirectional activity-dependent morphological plasticity. Using two-photon time-lapse microscopy, we observed that low-frequency stimulation induced NMDA receptor-dependent spine retractions, whereas theta burst stimulation led to the formation of new spines. Moreover, without stimulation the number of spine retractions was on the same order of magnitude as the stimulus-induced spine gain or loss. Finally, we found that the ability of neurons to eliminate spines in an activity-dependent manner decreased with developmental age. Taken together, our data show that hippocampal neurons can undergo bidirectional morphological plasticity; spines are formed and eliminated in an activity-dependent way.     

Enzyme purification by immobilized metal ion affinity partitioning--application to D-hydroxyisocaproate dehydrogenase.

Extraction and purification of D-2-hydroxyisocaproate dehydrogenase from Lactobacillus casei has been studied by means of immobilized metal ion affinity partitioning (IMAP) in aqueous two-phase systems. The partition of the enzyme can be influenced strongly by inclusion of iminodiacetic acid as chelating ligand coupled to polyethylene glycol and loaded with Cu2+ ions into the phase system. This applies to polyethylene glycol/dextran as well as polyethylene glycol/salt phase systems. An increase in enzyme partition coefficient of up to about 1000-fold was observed. Based on the mathematic model presented recently by Suh and Arnold (1990) approximately 6.4 histidine residues were calculated to be involved in the enzyme-metal chelate complex. Direct extraction of the enzyme from both cell homogenate and cell debris supernatant proved unsatisfactory due to disturbances caused by the presence of cell debris and low molecular weight cell components. A combination with a preceding prepurification by a fractional precipitation with polyethylene glycol resulted in a strong affinity effect accompanied by an efficient purification during IMAP (purification factor of 11 with a yield of approximately 90%). Based on this step, an efficient downstream process can be designed for D-hydroxyisocaproate dehydrogenase.     

Technical aspects of separation and simultaneous enzymatic reaction in multiphase enzyme membrane reactors

The technical aspects of the membrane extraction of a compound either from aqueous phase into apolar organic solvent phase or from the apolar phase to the aqueous one and the enzymatic conversion of the solute in a multiphase enzyme membrane reactor are considered. The application possibilities, the selection aspects of membrane material as well as the solvent phase, the water content and its control, the method of the enzyme immobilisation and the operation of the extraction/reaction system are discussed.     

Continuous production of restriction endonucleases: continuous two-stage cultivation with E. coli JM103; continuous cell disintegration and purification by affinity chromatography

The optimization of the production of recombinant DNA-derived proteins in Escherichia coli was investigated. We chose restriction endonucleases EcoRI and EcoRV from E. coli as model proteins, despite the observation that overproduction can result in a toxic effect to the cells. The enzymes were expressed as fusion proteins consisting of protein A from Staphylococcus aureus and the desired enzyme in order to facilitate purification. The expression of the fusion protein was induced by a temperature shift using the p_R promoter of phage lambda regulated by the repressor plasmid pRK248cI. Data from batch fermentations provided the basis for planning a continuous two-stage fermentation. The EcoRI enzyme activity was investigated as a function of the induction time after cell disintegration and allowed an estimation of yield of the continuous culture. Plasmid instability, which was only observed under continuous conditions, could be prevented by adding tetracycline (resistance of the repressor plasmid) to the medium. We established a continuous cell disintegration system and purified the fusion protein semicontinuously by affinity chromatography. The biological activity of the fusion protein was the same as the native endonuclease so there was no need for cleavage of the fusion protein and the product could be used without further processing.Correspondence to: K. Schügerl     

More pronounced salt dependence and higher reactivity for platination of the hairpin r(CGCGUUGUUCGCG) compared with d(CGCGTTGTTCGCG)

The DNA interference pathways exhibited by cisplatin and related anticancer active metal complexes have been extensively studied. Much less is known to what extent RNA interaction pathways may operate in parallel, and perhaps contribute to both antineoplastic activity and toxicity. The present study was designed with the aim of comparing the reactivity of two model systems comprising RNA and DNA hairpins, r(CGCGUUGUUCGCG) and d(CGCGTTGTTCGCG), towards a series of platinum(II) complexes. Three platinum complexes were used as metallation reagents; cis-[PtCl(NH₃)₂(OH₂)]⁺ (1), cis-[PtCl(NH₃)(c-C₆H₁₁NH₂)(OH₂)]⁺ (2), and trans-[PtCl(NH₃)(quinoline)(OH₂)]⁺ (3). The reaction kinetics were studied at pH 6.0, 25 °C, and 1.0 mM ≤ I ≤ 500 mM. For both types of nucleic acid targets, compound 3 was found to react about 1 order of magnitude more rapidly than compounds 1 and 2. Further, all platinum compounds exhibited a more pronounced salt dependence for the interaction with r(CGCGUUGUUCGCG). Chemical and enzymatic cleavage studies revealed similar interaction patterns with r(CGCGUUGUUCGCG) after long exposure times to 1 and 2. A substantial decrease of cleavage intensity was found at residues G4 and G7, indicative of bifunctional adduct formation. Circular dichroism studies showed that platinum adduct formation leads to a structural change of the ribonucleic acid. Thermal denaturation studies revealed platination to cause a decrease of the RNA melting temperatures by 5–10 °C. Our observations therefore suggest that RNA is a kinetically competitive target to DNA. Furthermore, platination causes destabilization of RNA structural elements, which may lead to deleterious intracellular effects on biologically relevant RNA targets.Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Progress in monitoring, modeling and control of bioprocesses during the last 20 years

The paper gives a review on the recent development of bioprocess engineering. It includes monitoring of product formation processes by flow injection analysis, various types of chromatographic and spectroscopic methods as well as by biosensors. The evaluation of mycelial morphology and physiology by digital image analysis is discussed also. It deals with advanced control of indirectly evaluated process variables by means of state estimation/observer, with the use of structured and hybrid models, expert systems and pattern recognition for process optimization and gives a short report on the state of the art of metabolic flux analysis and metabolic engineering.