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      Tension-Compression Loading with Chemical Stimulation Results in Additive Increases to Functional Properties of Anatomic Meniscal Constructs

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      PLoS ONE
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          Abstract

          Objective

          This study aimed to improve the functional properties of anatomically-shaped meniscus constructs through simultaneous tension and compression mechanical stimulation in conjunction with chemical stimulation.

          Methods

          Scaffoldless meniscal constructs were subjected to simultaneous tension and compressive stimulation and chemical stimulation. The temporal aspect of mechanical loadingwas studied by employing two separate five day stimulation periods. Chemical stimulation consisted of the application of a catabolic GAG-depleting enzyme, chondroitinase ABC (C-ABC), and an anabolic growth factor, TGF-β1. Mechanical and chemical stimulation combinations were studied through a full-factorial experimental design and assessed for histological, biochemical, and biomechanical properties following 4 wks of culture.

          Results

          Mechanical loading applied from days 10–14 resulted in significant increases in compressive, tensile, and biochemical properties of meniscal constructs. When mechanical and chemical stimuliwere combined significant additive increases in collagen per wet weight (4-fold), compressive instantaneous (3-fold) and relaxation (2-fold) moduli, and tensile moduli in the circumferential (4-fold) and radial (6-fold) directions were obtained.

          Conclusions

          This study demonstrates that a stimulation regimen of simultaneous tension and compression mechanical stimulation, C-ABC, and TGF-β1 is able to create anatomic meniscus constructs replicating the compressive mechanical properties, and collagen and GAG content of native tissue. In addition, this study significantly advances meniscus tissue engineering by being the first to apply simultaneous tension and compression mechanical stimulation and observe enhancement of tensile and compressive properties following mechanical stimulation.

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          Most cited references57

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          The differential adhesion hypothesis: a direct evaluation.

          The differential adhesion hypothesis (DAH), advanced in the 1960s, proposed that the liquid-like tissue-spreading and cell segregation phenomena of development arise from tissue surface tensions that in turn arise from differences in intercellular adhesiveness. Our earlier measurements of liquid-like cell aggregate surface tensions have shown that, without exception, a cell aggregate of lower surface tension tends to envelop one of higher surface tension to which it adheres. We here measure the surface tensions of L cell aggregates transfected to express N-, P- or E-cadherin in varied, measured amounts. We report that in these aggregates, in which cadherins are essentially the only cell-cell adhesion molecules, the aggregate surface tensions are a direct, linear function of cadherin expression level. Taken together with our earlier results, the conclusion follows that the liquid-like morphogenetic cell and tissue rearrangements of cell sorting, tissue spreading and segregation represent self-assembly processes guided by the diminution of adhesive-free energy as cells tend to maximize their mutual binding. This conclusion relates to the physics governing these morphogenetic phenomena and applies independently of issues such as the specificities of intercellular adhesives.
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            The statistics of synergism.

            Biological scientists often want to determine whether two agents or events, for example, extracellular stimuli and/or intracellular signaling pathways, act synergistically when eliciting a biological response. When setting out to study whether two experimental treatments act synergistically, most biologists design the correct experiment--they administer four treatment combinations consisting of (1) the first treatment alone, (2) the second treatment alone, (3) both treatments together, and (4) neither treatment (i.e. the control). Many biologists are less clear about the correct statistical approach to determining whether the data collected in such an experimental design support a conclusion regarding synergism, or lack thereof. The non-additivity of two experimental treatments that is central to the definition of synergism leads to an algebraic formulation corresponding to the statistical null hypothesis appropriate for testing whether or not there is synergism. The resulting complex contrast among the four treatment group means is identical to the interaction effect tested in a two-way analysis of variance (ANOVA). This should not be surprising, because synergism, by definition, occurs when two treatments interact, rather than act independently, to influence a biological response. Hence, in the most readily implemented approach, the correct statistical analysis of a question of synergism is based on testing the interaction effect in a two-way ANOVA. This review presents the rationale for this correct approach to analysing data when the question is of synergism, and applies this approach to a recent published example. In addition, a common incorrect approach to analysing data with regards to synergism is presented. Finally, several associated statistical issues with regard to correctly implementing a two-way ANOVA are discussed.
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              Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose culture.

              This study focuses on the effect of static and dynamic mechanical compression on the biosynthetic activity of chondrocytes cultured within agarose gel. Chondrocyte/agarose disks (3 mm diameter) were placed between impermeable platens and subjected to uniaxial unconfined compression at various times in culture (2-43 days). [35S]sulfate and [3H]proline radiolabel incorporation were used as measures of proteoglycan and protein synthesis, respectively. Graded levels of static compression (up to 50%) produced little or no change in biosynthesis at very early times, but resulted in significant decreases in synthesis with increasing compression amplitude at later times in culture; the latter observation was qualitatively similar to that seen in intact cartilage explants. Dynamic compression of approximately 3% dynamic strain amplitude (approximately equal to 30 microns displacement amplitude) at 0.01-1.0 Hz, superimposed on a static offset compression, stimulated radiolabel incorporation by an amount that increased with time in culture prior to loading as more matrix was deposited around and near the cells. This stimulation was also similar to that observed in cartilage explants. The presence of greater matrix content at later times in culture also created differences in biosynthetic response at the center versus near the periphery of the 3 mm chondrocyte/agarose disks. The fact that chondrocyte response to static compression was significantly affected by the presence or absence of matrix, as were the physical properties of the disks, suggested that cell-matrix interactions (e.g. mechanical and/or receptor mediated) and extracellular physicochemical effects (increased [Na+], reduced pH) may be more important than matrix-independent cell deformation and transport limitations in determining the biosynthetic response to static compression. For dynamic compression, fluid flow, streaming potentials, and cell-matrix interactions appeared to be more significant as stimuli than the small increase in fluid pressure, altered molecular transport, and matrix-independent cell deformation. The qualitative similarity in the biosynthetic response to mechanical compression of chondrocytes cultured in agarose gel and chondrocytes in intact cartilage further indicates that gel culture preserves certain physiological features of chondrocyte behavior and can be used to investigate chondrocyte response to physical and chemical stimuli in a controlled manner.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2011
                16 November 2011
                : 6
                : 11
                : e27857
                Affiliations
                [1]Department of Biomedical Engineering, University of California Davis, Davis, California, United States of America
                University of Pittsburgh, United States of America
                Author notes

                Conceived and designed the experiments: DJH KAA. Performed the experiments: DJH. Analyzed the data: DJH KAA. Contributed reagents/materials/analysis tools: DJH KAA. Wrote the paper: DJH KAA.

                Article
                PONE-D-11-10874
                10.1371/journal.pone.0027857
                3218070
                22114714
                91a8e32a-2aaf-48ae-a15f-abd692031461
                Huey, Athanasiou. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                History
                : 15 June 2011
                : 26 October 2011
                Page count
                Pages: 9
                Categories
                Research Article
                Biology
                Biotechnology
                Bioengineering
                Biomedical Engineering
                Biomaterials
                Tissue Engineering
                Engineering
                Bioengineering
                Biomedical Engineering
                Materials Science
                Biomaterials

                Uncategorized
                Uncategorized

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