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D Herbage

Publications and source records attributed to D Herbage.

97 records · Page 6Linked to original sources
PubMedDate not supplied

Mechanical loading regulates protease production by fibroblasts in three-dimensional collagen substrates.

Mechanical loading is important in tissue formation and remodelling, notably in wound repair. The aim of this study was to measure the effects of controlled loading on the release of extracellular matrix protease activities by fibroblasts. Fibroblast populated collagen lattices were subjected to external cyclical loads through a computer controlled unit incorporated into a culture system, a tensioning-Culture Force Monitor. Cyclical loading was compared to untensioned and statically loaded gels (tethered endogenous contraction). Overall changes in a range of protease activities were monitored (chiefly by zymography) as measures of the cyto-mechanical response to these loads. Under static load, 2.5- and 13-fold more matrix metalloproteinase-2 was produced than matrix metalloproteinase-9, at 24 and 48 hours. Total matrix metalloproteinase-9 increased 37 fold on cyclical loading. Total matrix metalloproteinase-3 and urokinase plasminogen activator activities were dramatically reduced on cyclical loading while tissue type plasminogen activator activity was increased. Comparison with cell responses on stiffer substrates (collagen sponges) identified similar matrix metalloproteinase responses to load, but at much reduced levels (4-6 fold matrix metalloproteinase-9 stimulation on loading), showing the importance of matrix compliance to this mechano-response. In conclusion, physiological mechanical loading of fibroblasts in three dimensional collagen lattices elicited complex and substantial changes in matrix modifying proteases. These changes suggest that cells switch between expression of comparable protease activities mainly influencing cell-matrix interactions associated with migration or more generalized extracellular matrix remodelling.

Adult↗
PubMedDate not supplied

Optimization of dynamic culture conditions: effects on biosynthetic activities of chondrocytes grown in collagen sponges.

Application of mechanical stimulation, using dynamic bioreactors, is considered an effective strategy to enhance cellular behavior in load-bearing tissues. In this study, two types of perfusion mode (direct and free flow) are investigated in terms of the biosynthetic activities of chondrocytes grown in collagen sponges by assessment of cell proliferation rate, matrix production, and tissue morphology. Effects of the duration of preculture and dynamic conditioning are further determined. Our results have demonstrated that both bovine and human-derived chondrocytes demonstrate a dose-dependent response to flow rate (0-1 mL/min) in terms of cell number and glycosaminoglycan (GAG) content. This may reflect the weak adhesion of cells to the sponge scaffolds and the immature state of the constructs even after 3 weeks of proliferative culture. Our studies define an optimal flow rate between 0.1 and 0.3 mL/min for direct perfusion and free flow bioreactors. Using fresh bovine chondrocytes and a lower flow rate of 0.1 mL/min, a comparison was made between free flow system and direct perfusion system. In the free flow bioreactor, no cell loss was observed and higher GAG production was measured compared with static cultured controls. However, as with direct perfusion, the enhancement effect of free flow perfusion was strongly dependent on the maturation and organization of the constructs before the stimulation. To address the maturation of the matrix, preculture periods were varied before mechanical conditioning. An increase in culture duration of 18 days before mechanical conditioning resulted in enhanced GAG production compared with controls. Interestingly, additional enhancement was found in specimens that were further subjected to a prolonged duration of perfusion (63% increase after an additional 4 days of perfusion) after prematuration. The free flow system has an advantage over the direct perfusion system, especially when using sponge scaffolds, which have lower mechanical properties; however, mass transfer of nutrients is still more optimal throughout the scaffolds in a direct perfusion system as demonstrated by histological analysis.

Animals↗
PubMedDate not supplied

[Detection of anti-collagen type I and II antibodies by an immunoenzymatic technique (ELISA): results in rheumatoid arthritis, systemic lupus erythematosus and leprosy (author's transl)].

An attempt was made to detect antibodies against type I and/or II collagen in sera from patients with rheumatoid arthritis, systemic lupus erythematosus (SLE) and leprae. This study was performed with an immunoenzymatic technique: ELISA (enzyme-linked immunosorbent assay). The following steps were performed: bovine collagen type I or II was adsorbed on glass beads; possible free sites were saturated by incubating the beads with sheep serum; then, the antibodies specifically bound to collagen were detected by a peroxidase-labelled anti-immunoglobulin; the immune complexes at the surface of the beads were revealed by a substrate specific for peroxidase and of great stability: Trinder's reactive. Using conditions previously shown to be optimal, the prevalence of anti-collagen antibodies was as follows. In patients with lepromatous leprae the percentages of positive sera against collagen type I and II were 40% and 44%, respectively; in patients with tuberculoid leprae the percentages were lower: 10% and 30%, respectively. Ten per cent of the SLE patients had antibodies against collagen type I, half of the prevalence noted for anti-collagen type II antibodies (20%). Finally, 13.6% of the patients with rheumatoid arthritis had antibodies against collagen type I, a percentage very similar to that of the patients with anti-collagen type II antibodies (14.6%).

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