Chondroitin Sulfate in Palatal Wound Healing
X.H. Zou1,
W.C. Foong1,
T. Cao1,
B.H. Bay2,
H.W. Ouyang3, and
G.W. Yip2,*
1 Faculty of Dentistry, National University of Singapore, Singapore;
2 Department of Anatomy, Faculty of Medicine, National University of Singapore, 4 Medical Drive, Block MD 10, Singapore 117597, Singapore; and
3 Division of Bioengineering, Faculty of Engineering, National University of Singapore, Singapore;
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Figure 1. Chondroitin sulfate regulates palatal fibroblast adhesion and proliferation. (A) Degradation of chondroitin sulfate by the addition of 0.1 unit/mL chondroitinase ABC to the culture medium for 8 hrs results in a decrease in the number of adherent cells (Student’s t test; p = 0.0076). (B) Conversely, supplementation of the culture medium with chondroitin-6-sulfate for the same time period leads to a dose-dependent increase in cell adhesion (one-way ANOVA; p < 0.0001). (C) In the cell proliferation assay, statistical comparison by one-way ANOVA shows that palatal fibroblasts cultured in the presence of chondroitin-6-sulfate over a seven-day period show a dose-dependent increase in cell numbers (p < 0.0001). Values represent mean ± SEM of 3 experiments.
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Figure 2. The sulfate group is required for biological effects of chondroitin sulfate. Palatal fibroblasts were cultured for 8 hrs in normal culture medium, medium supplemented with 30 mmol/L chlorate, or medium with 30 mmol/L chlorate plus (A) 10 mmol/L sulfate or (B) 100 ng/mL chondroitin-6-sulfate (C6S). The number of adherent cells varied among treatment groups (one-way ANOVA; p < 0.0001), with fewer adherent cells in the chlorate-alone-treated group compared with the control group (Tukey’s test; p < 0.001 in both panels). Fibroblasts treated with chlorate plus exogenous sulfate or chondroitin-6-sulfate showed significantly greater adhesion than those treated with chlorate alone (p < 0.001 in each case). (C) In the cell proliferation assay, continuous chlorate treatment also led to reduced cell numbers after a seven-day culture period (p < 0.001). This reduction was significantly blocked by the addition of 100 ng/mL chondroitin-6-sulfate to the culture medium (p < 0.001). Values represent mean ± SEM of 3 experiments.
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Figure 3. Chondroitin sulfate regulates cell migration and wound closure in vitro. (A) Scanning electron micrograph of palatal fibroblasts cultured for 18 hrs in the presence of 30 mmol/L chlorate, after the making of a linear wound as described in MATERIALS & METHODS. The distance between the wound edges is indicated (*). (B) Statistical comparison of the distance between wound edges 18 hrs after wounding occurred shows a significant difference (one-way ANOVA; p < 0.001) among cells cultured in normal medium, medium supplemented with 30 mmol/L chlorate, or medium with 30 mmol/L chlorate plus 100 ng/mL chondroitin-6-sulfate (C6S). The wound gap in the chlorate-alone-treated group was significantly wider compared with that in the control group (Tukey’s test; p < 0.001) or with the group treated with chlorate plus chondroitin-6-sulfate (p < 0.001). Values represent mean ± SEM of 9 wounds per group.
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Figure 4. Biological effects of chondroitin-4-sulfate. (A) Palatal fibroblasts cultured for 8 hrs in medium supplemented with chondroitin-4-sulfate showed a dose-dependent reduction in cell adhesion (one-way ANOVA; p = 0.0049). (B) In the cell proliferation assay, statistical comparison shows that continuous supplementation of the culture medium with chondroitin-4-sulfate led to a dose-dependent increase in cell numbers after a seven-day culture period (p = 0.0074). Values represent mean ± SEM of 3 experiments.
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