Double-network acrylamide hydrogel compositions adapted to achieve cartilage-like dynamic stiffness

Since articular cartilage has a limited potential for spontaneous healing, various techniques are employed to repair cartilage lesions. Acrylate-based double-network (DN) hydrogels containing ~90% water have shown promising properties as repair materials for skeletal system soft tissues. Although their mechanical properties approach those of native cartilage, the critical factor—stiffness—of DN-gels does not equal the stiffness of articular cartilage. This study investigated whether revised PAMPS/PAAm compositions with lower water content result in stiffness parameters closer to cartilage. DN-gels containing 61, 86 and 90% water were evaluated using two non-destructive, mm-scale indentation test modes: fast-impact (FI) and slow-sinusoidal (SS) deformation. Deformation resistance (dynamic modulus) and energy handling (loss angle) were determined. The dynamic modulus increased with decreasing water content in both testing modes. In the 61% water DN-gel, the modulus resembled that of cartilage (FI-mode: DN-gel = 12, cartilage = 17; SS-mode: DN-gel = 4, cartilage = 1.7MPa). Loss angle increased with decreasing water content in fast-impact, but not in slow-sinusoidal deformation. However, loss angle was still much lower than cartilage (FI: DN-gel = 5, cartilage = 11; SS: DN-gel = 10, cartilage = 32°), indicating somewhat less ability to dissipate energy. Overall, results show that it is possible to adapt DN-gel composition to produce dynamic stiffness properties close to normal articular cartilag

A comparison of healthy human and swine articular cartilage dynamic indentation mechanics

Articular cartilage is a multicomponent, poroviscoelastic tissue with nonlinear mechanical properties vital to its function. A consequent goal of repair or replacement of injured cartilage is to achieve mechanical properties in the repair tissue similar to healthy native cartilage. Since fresh healthy human articular cartilage (HC) is not readily available, we tested whether swine cartilage (SC) could serve as a suitable substitute for mechanical comparisons. To a first approximation, cartilage tissue and surgical substitutes can be evaluated mechanically as viscoelastic materials. Stiffness measurements (dynamic modulus, loss angle) are vital to function and are also a non-destructive means of evaluation. Since viscoelastic material stiffness is strongly strain rate dependent, stiffness was tested under different loading conditions related to function. Stiffness of healthy HC and SC specimens was determined and compared using two non-destructive, mm-scale indentation test modes: fast impact and slow sinusoidal deformation. Deformation resistance (dynamic modulus) and energy handling (loss angle) were determined. For equivalent anatomic locations, there was no difference in dynamic modulus. However, the HC loss angle was ~35% lower in fast impact and ~12% higher in slow sinusoidal mode. Differences seem attributable to age (young SC, older HC) but also to species anatomy and biology. Test mode-related differences in human-swine loss angle support use of multiple function-related test modes. Keeping loss angle differences in mind, swine specimens could serve as a standard of comparison for mechanical evaluation of e.g. engineered cartilage or synthetic repair material

Mechanics of the exceptional anuran ear

The anuran ear is frequently used for studying fundamental properties of vertebrate auditory systems. This is due to its unique anatomical features, most prominently the lack of a basilar membrane and the presence of two dedicated acoustic end organs, the basilar papilla and the amphibian papilla. Our current anatomical and functional knowledge implies that three distinct regions can be identified within these two organs. The basilar papilla functions as a single auditory filter. The low-frequency portion of the amphibian papilla is an electrically tuned, tonotopically organized auditory end organ. The high-frequency portion of the amphibian papilla is mechanically tuned and tonotopically organized, and it emits spontaneous otoacoustic emissions. This high-frequency portion of the amphibian papilla shows a remarkable, functional resemblance to the mammalian cochlea

Postsynaptic nigrostriatal dopamine receptors and their role in movement regulation

The article presents the hypothesis that nigrostriatal dopamine may regulate movement by modulation of tone and contraction in skeletal muscles through a concentration-dependent influence on the postsynaptic D1 and D2 receptors on the follow manner: nigrostriatal axons innervate both receptor types within the striatal locus somatotopically responsible for motor control in agonist/antagonist muscle pair around a given joint. D1 receptors interact with lower and D2 receptors with higher dopamine concentrations. Synaptic dopamine concentration increases immediately before movement starts. We hypothesize that increasing dopamine concentrations stimulate first the D1 receptors and reduce muscle tone in the antagonist muscle and than stimulate D2 receptors and induce contraction in the agonist muscle. The preceded muscle tone reduction in the antagonist muscle eases the efficient contraction of the agonist. Our hypothesis is applicable for an explanation of physiological movement regulation, different forms of movement pathology and therapeutic drug effects. Further, this hypothesis provides a theoretical basis for experimental investigation of dopaminergic motor control and development of new strategies for treatment of movement disorders

Accounting for the thickness effect in dynamic spherical indentation of a viscoelastic layer: Application to non-destructive testing of articular cartilage

In recent years, dynamic indentation tests have been shown to be useful both in identification of mechanical properties of biological tissues (such as articular cartilage) and assessing their viability. We consider frictionless flat-ended and spherical sinusoidally-driven indentation tests utilizing displacement-controlled loading protocol. Articular cartilage tissue is modeled as a viscoelastic material with a time-independent Poisson's ratio. We study the dynamic indentation stiffness with the aim of formulating criteria for evaluation the quality of articular cartilage in order to be able to discriminate its degenerative state. In particular, evaluating the dynamic indentation stiffness at the turning point of the flat-ended indentation test, we introduce the so-called incomplete storage modulus. Considering the time difference between the time moments when the dynamic stiffness vanishes (contact force reaches its maximum) and the dynamic stiffness becomes infinite (indenter displacement reaches its maximum), we introduce the so-called incomplete loss angle. Analogous quantities can be introduced in the spherical sinusoidally-driven indentation test, however, to account for the thickness effect, a special approach is required. We apply an asymptotic modeling approach for analyzing and interpreting the results of the dynamic spherical indentation test in terms of the geometrical parameter of the indenter and viscoelastic characteristics of the material. Some implications to non-destructive indentation diagnostics of cartilage degeneration are discussed.Comment: 29 pages, 7 Figure

Mutational analysis of the potential phosphorylation sites for protein kinase C on the CCK(A) receptor

1. Many G protein-coupled receptors contain potential phosphorylation sites for protein kinase C (PKC), the exact role of which is poorly understood. In the present study, a mutant cholecystokinin(A) (CCK(A)) receptor was generated in which the four consensus sites for PKC action were changed in an alanine. Both the wild-type (CCK(A)WT) and mutant (CCK(A)MT) receptor were stably expressed in Chinese hamster ovary (CHO) cells. 2. Binding of [(3)H]-cholecystokinin-(26-33)-peptide amide (CCK-8) to membranes prepared from CHO-CCK(A)WT cells and CHO-CCK(A)MT cells revealed no difference in binding affinity (K(d) values of 0.72 nM and 0.86 nM CCK-8, respectively). 3. The dose-response curves for CCK-8-induced cyclic AMP accumulation and inositol 1,4,5-trisphosphate (Ins(1,4,5)P(3)) formation were shifted to the left in CHO-CCK(A)MT cells. This leftward shift was mimicked by the potent inhibitor of protein kinase activity, staurosporine. However, the effect of staurosporine was restricted to CHO-CCK(A)WT cells. This demonstrates that attenuation of CCK-8-induced activation of adenylyl cyclase and phospholipase C-β involves a staurosporine-sensitive kinase, which acts directly at the potential sites of PKC action on the CCK(A) receptor in CCK-8-stimulated CHO-CCK(A)WT cells. 4. The potent PKC activator, 12-O-tetradecanoylphorbol 13-acetate (TPA), evoked a rightward shift of the dose-response curve for CCK-8-induced cyclic AMP accumulation in CHO-CCK(A)WT cells but not CHO-CCK(A)MT cells. This is in agreement with the idea that PKC acts directly at the CCK(A) receptor to attenuate adenylyl cyclase activation. 5. In contrast, TPA evoked a rightward shift of the dose-response curve for CCK-8-induced Ins(1,4,5)P(3) formation in both cell lines. This demonstrates that high-level PKC activation inhibits CCK-8-induced Ins(1,4,5)P(3) formation also at a post-receptor site. 6. TPA inhibition of agonist-induced Ca(2+) mobilization was only partly reversed in CHO-CCK(A)MT cells. TPA also inhibited Ca(2+) mobilization in response to the G protein activator, Mas-7. These findings are in agreement with the idea that partial reversal of agonist-induced Ca(2+) mobilization is due to the presence of an additional site of PKC inhibition downstream of the receptor and that the mutant receptor itself is not inhibited by the action of PKC. 7. The data presented demonstrate that the predicted sites for PKC action on the CCK(A) receptor are the only sites involved in TPA-induced uncoupling of the receptor from its G proteins. In addition, the present study unveils a post-receptor site of PKC action, the physiological relevance of which may be that it provides a means for the cell to inhibit phospholipase C-β activation by receptors that are not phosphorylated by PKC