Calcification of subcutaneously implanted collagens in relation to cytotoxicity,cellular interactions and crosslinking

In general, calcification of biomaterials occurs through an interaction of host and implanted material factors, but up to now the real origin of pathologic calcification is unknown. In this study we aimed to investigate incidence of calcification of (crosslinked) dermal sheep collagens (DSCs) with respect to their specific properties, during subcutaneous implantation in rats. Three types of DSCs were commercially obtained: non-crosslinked DSC (NDSC), and DSC crosslinked with glutaraldehyde (GDSC) and hexamethylenediisocyanate (HDSC). NDSC, HDSC and GDSC were (enzymatically) tissue culture pretreated to eliminate their cytotoxic products. Beside this, crosslinking methods were modified to optimize mechanical properties and to decrease cytotoxicity, which resulted in HDSC^* and GDSC^*. Furthermore, DSC was crosslinked by activation of the carboxylic groups, i.e. by means of acyl azide and carbodiimide, resulting in AaDSC and CDSC, respectively. After implantation of HDSCs and GDSCs a relation between cytotoxicity and calcification of crosslinked DSC could be made. No relation was found between cellular infiltration of DSCs and calcification. However, from the use of different types and modification of crosslinking methods it might be concluded that calcification is mainly related to stable crosslinks, i.e. to the chemical properties of the obtained material.     

In vitro degradation of dermal sheep collagen cross-linked using a water-soluble carbodiimide

Bacterial collagenase was used to study the susceptibility of dermal sheep collagen (DSC) cross-linked with a mixture of the water-soluble carbodiimide 1-ethyl-3-(3-dimethyl aminopropyl)-carbodiimide hydrochloride and N-hydroxysuccinimide (E/N-DSC) towards enzymatic degradation. Contrary to non-cross-linked DSC (N-DSC), which had a rate of weight-loss of 18.1% per hour upon degradation, no weight loss was observed for E/N-DSC during a 24 h degradation period. The tensile strength of the E/N-DSC samples decreased during this time period, resulting in partially degraded samples having 80% of the initial tensile strength remaining. The susceptibility of E/N-DSC samples towards enzymatic degradation could be controlled by varying the degree of cross-linking of the samples. Ethylene oxide sterilization of E/N-DSC samples made the material more resistant against degradation compared with non-sterilized E/N-DSC samples. This may be explained by a decrease of the adsorption of bacterial collagenase onto the collagen owing to reaction of ethylene oxide with remaining free amine groups in the collagen matrix.     

Position and velocity coupling of postural sway to somatosensory drive

Light touch contact of a fingertip to a stationary surface provides orientation information that enhances control of upright stance. Slight changes in contact force at the fingertip lead to sensory cues about the direction of body sway, allowing attenuation of sway. In the present study, the coupling of postural sway to a moving contact surface was investigated in detail. Head, center of mass, and center of pressure displacement were measured as the contact surface moved rhythmically at 0.1, 0.2, 0.4, 0.6, and 0.8 Hz. Stimulus amplitude decreased with frequency to maintain peak velocity constant across frequency. Head and body sway were highly coherent with contact surface motion at all frequencies except 0.8 Hz, where a drop-off in coherence was observed. Mean frequency of head and body sway matched the driving frequency 相似文献   

The effect of cavity-filling mutations on the thermostability of Bacillus stearothermophilus neutral protease

Cavities in the hydrophobic core of the neutral protease ofBacillus stearothermophilus were analyzed using a threedimensionalmodel that was inferred from the crystal structure of thermolysin,the highly homologous neutral protease of B.thermoproteolyticus(85% sequence identity). Site–directed mutagenesis wasused to fill some of these cavities, thereby improving hydrophobicpacking in the protein interior. The mutations had small effectson the thermostability, even after drastic changes, such asLeu284Trp and Met168Trp. The effects on T50, the temperatureat which 50% of the enzyme is irreversibly inactivated in 30min, ranged from 0.0 to +0.4°C. These results can be explainedby assuming that the mutations have positive and negative structuraleffects of approximately the same magnitude. Alternatively,it could be envisaged that the local unfolding steps, whichrender the enzyme susceptible towards autolysis and which arerate limiting in the process of thermal inactivation, are onlyslightly affected by alterations in the hydrophobic core.     

Single-Site Calcium Initiators for the Controlled Ring-Opening Polymerization of Lactides and Lactones

Summary Single-site calcium initiators containing chelating tmhd (H-tmhd = 2,2,6,6-tetramethylheptane-33-dione) ligands [(THF)Ca(tmhd)_]2[-N(SiMe₃)₂](-tmhd) (2) and [(THF)Ca(tmhd)]₂[-OCH(Me)Ph](-tmhd) (3) have been synthesized and applied for the ring-opening polymerization of L-lactide and -caprolactone. Both 2 and 3 were highly reactive and promoted a fast polymerization of L-lactide and -caprolactone to high monomer conversions under mild conditions (THF as a solvent, room temperature). More importantly, results showed that the ring-opening polymerizations of lactides and lactones initiated by either 3 or 2 in the presence of equivalent 2-propanol are living, to provide polymers and block copolymers of controlled molecular weights and tailored end-groups. The polymerizations were first-order in monomer up to high conversions, in which the in situ initiating system 2/2-propanol revealed no induction period and much faster polymerization kinetics as compared to 3.     

Evidence for non-site-isolation in ZSM-5 from ion-exchange and catalytic studies

Ion exchange of HZSM-5 samples with alkali metal cations, using metal chloride solutions, results in partially exchanged zeolites, MHZSM-5, M = Li, Na, K or Cs. The degree of exchange is found to increase with increasing ionic radius of the cations. The catalytic properties of the alkalized zeolites were evaluated using the reaction conditions under which the catalytic activity of the HZSM-5 samples in terms of n-hexane cracking is proportional to the aluminium content. From the residual catalytic activity exhibited by the Na-, K- and CsHZSM-5 samples it is concluded that each of the larger Na⁺, K⁺ and Cs⁺ ions is influencing more than one AlO ₄ ^– tetrahedron, implying that the aluminium sites in ZSM-5 are not isolated. The ion-exchange results are then interpreted in terms of non-isolated aluminium sites. The ion-exchange and catalytic properties of the zeolites as a function of aluminium content are also discussed.     

Structural and kinetic studies on ligand binding in wild-type and active-site mutants of penicillin acylase

Penicillin acylase catalyses the condensation of Calpha-substituted phenylacetic acids with beta-lactam nucleophiles, producing semi-synthetic beta-lactam antibiotics. For efficient synthesis a low affinity for phenylacetic acid and a high affinity for Calpha-substituted phenylacetic acid derivatives is desirable. We made three active site mutants, alphaF146Y, betaF24A and alphaF146Y/betaF24A, which all had a 2- to 10-fold higher affinity for Calpha-substituted compounds than wild-type enzyme. In addition, betaF24A had a 20-fold reduced affinity for phenylacetic acid. The molecular basis of the improved properties was investigated by X-ray crystallography. These studies showed that the higher affinity of alphaF146Y for (R)-alpha-methylphenylacetic acid can be explained by van der Waals interactions between alphaY146:OH and the Calpha-substituent. The betaF24A mutation causes an opening of the phenylacetic acid binding site. Only (R)-alpha-methylphenylacetic acid, but not phenylacetic acid, induces a conformation with the ligand tightly bound, explaining the weak binding of phenylacetic acid. A comparison of the betaF24A structure with other open conformations of penicillin acylase showed that betaF24 has a fixed position, whereas alphaF146 acts as a flexible lid on the binding site and reorients its position to achieve optimal substrate binding.