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In vitro response of retinal pigment epithelial cells exposed to chitosan materials prepared with different cross-linkers.

Lai JY, Li YT, Wang TP - Int J Mol Sci (2010)

Bottom Line: The present study showed that the ARPE-19 cells exposed to GTA cross-linked chitosan membranes had significantly higher cytotoxicity, interleukin-6 levels, and number of TUNEL-positive nuclei than did those exposed to GP treated samples.The findings suggest that while the chitosan molecules bridged by GP are satisfactorily cytocompatible, the counterparts treated by GTA do not seem to be tolerated.In terms of material safety, the GP cross-linked chitosan may be compatible with human RPE cells and may have a potential application as delivery carriers in the treatment of posterior segment diseases.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan, 33302, Taiwan.

ABSTRACT
The interaction between cells and biopolymers is the evaluation indicator of the biocompatibility of materials. The purpose of this work was to examine the responses of retinal pigment epithelial (RPE) cells to genipin (GP) or glutaraldehyde (GTA) cross-linked chitosan by means of cell viability assays, cytokine expression analyses, and apoptosis assays. Evaluations of non-cross-linked chitosan were conducted simultaneously for comparison. Both GP and GTA treated samples with the same extent of cross-linking (around 80%) were prepared by varying cross-linking time. Our results showed that GP cross-linking was carried out by either radical polymerization of the monomers or S(N)2 nucleophilic substitution reaction involving the replacement of the ester group on the monomer with a secondary amide linkage. On the other hand, GTA could react with free amino groups of chitosan, leading to the formation of either the Schiff bases or the Michael-type adducts with terminal aldehydes. The biocompatibility of non-cross-linked chitosan membranes was demonstrated by the absence of any signs of toxicity or inflammation reaction. The present study showed that the ARPE-19 cells exposed to GTA cross-linked chitosan membranes had significantly higher cytotoxicity, interleukin-6 levels, and number of TUNEL-positive nuclei than did those exposed to GP treated samples. In addition, the materials modified with GTA trigger apoptosis at an early stage and may induce toxicity in the RPE cells later. The findings suggest that while the chitosan molecules bridged by GP are satisfactorily cytocompatible, the counterparts treated by GTA do not seem to be tolerated. In terms of material safety, the GP cross-linked chitosan may be compatible with human RPE cells and may have a potential application as delivery carriers in the treatment of posterior segment diseases.

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Related in: MedlinePlus

Gene expression of IL-6 in ARPE-19 cells incubated with various chitosan membranes (5 mg) for 3 days, measured by real-time RT-PCR. Normalization was done using GAPDH. Data in the experimental groups are percentages relative to that of control groups (without materials). An asterisk indicates statistically significant differences (*p < 0.05; n = 3) as compared to controls.
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f6-ijms-11-05256: Gene expression of IL-6 in ARPE-19 cells incubated with various chitosan membranes (5 mg) for 3 days, measured by real-time RT-PCR. Normalization was done using GAPDH. Data in the experimental groups are percentages relative to that of control groups (without materials). An asterisk indicates statistically significant differences (*p < 0.05; n = 3) as compared to controls.

Mentions: IL-6, which appears to be a major cytokine-triggering mediator in the later part of the acute phase, can be used as an indicator of biocompatibility of the materials tested [34]. For evaluation of inflammatory responses to cross-linked chitosan membranes, we utilized real-time RT-PCR and ELISA to quantify the IL-6 expression at both the mRNA and protein levels. As shown in Figure 6, the pro-inflammatory gene expression of ARPE-19 cells after three days of exposure to biomaterials from the Chi, GP-chi, and GTA-chi groups was 117.4 ± 9.5, 113.8 ± 10.2, 860.3 ± 17.9%, respectively. A significant up-regulation of IL-6 mRNA expression was observed in the GTA-chi groups compared with the other groups (p < 0.05). On the other hand, Figure 7 shows the ARPE-19 cell secretion of IL-6 in response to various chitosan samples. The measured concentration of IL-6 in the GTA-chi groups was 1161.7 ± 34.4 pg/mL, which was significantly higher than those in the control (204.8 ± 27.2 pg/mL), Chi (229.3 ± 21.6 pg/mL), and GP-chi (190.5 ± 18.1 pg/mL) groups (p < 0.05).


In vitro response of retinal pigment epithelial cells exposed to chitosan materials prepared with different cross-linkers.

Lai JY, Li YT, Wang TP - Int J Mol Sci (2010)

Gene expression of IL-6 in ARPE-19 cells incubated with various chitosan membranes (5 mg) for 3 days, measured by real-time RT-PCR. Normalization was done using GAPDH. Data in the experimental groups are percentages relative to that of control groups (without materials). An asterisk indicates statistically significant differences (*p < 0.05; n = 3) as compared to controls.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3100825&req=5

f6-ijms-11-05256: Gene expression of IL-6 in ARPE-19 cells incubated with various chitosan membranes (5 mg) for 3 days, measured by real-time RT-PCR. Normalization was done using GAPDH. Data in the experimental groups are percentages relative to that of control groups (without materials). An asterisk indicates statistically significant differences (*p < 0.05; n = 3) as compared to controls.
Mentions: IL-6, which appears to be a major cytokine-triggering mediator in the later part of the acute phase, can be used as an indicator of biocompatibility of the materials tested [34]. For evaluation of inflammatory responses to cross-linked chitosan membranes, we utilized real-time RT-PCR and ELISA to quantify the IL-6 expression at both the mRNA and protein levels. As shown in Figure 6, the pro-inflammatory gene expression of ARPE-19 cells after three days of exposure to biomaterials from the Chi, GP-chi, and GTA-chi groups was 117.4 ± 9.5, 113.8 ± 10.2, 860.3 ± 17.9%, respectively. A significant up-regulation of IL-6 mRNA expression was observed in the GTA-chi groups compared with the other groups (p < 0.05). On the other hand, Figure 7 shows the ARPE-19 cell secretion of IL-6 in response to various chitosan samples. The measured concentration of IL-6 in the GTA-chi groups was 1161.7 ± 34.4 pg/mL, which was significantly higher than those in the control (204.8 ± 27.2 pg/mL), Chi (229.3 ± 21.6 pg/mL), and GP-chi (190.5 ± 18.1 pg/mL) groups (p < 0.05).

Bottom Line: The present study showed that the ARPE-19 cells exposed to GTA cross-linked chitosan membranes had significantly higher cytotoxicity, interleukin-6 levels, and number of TUNEL-positive nuclei than did those exposed to GP treated samples.The findings suggest that while the chitosan molecules bridged by GP are satisfactorily cytocompatible, the counterparts treated by GTA do not seem to be tolerated.In terms of material safety, the GP cross-linked chitosan may be compatible with human RPE cells and may have a potential application as delivery carriers in the treatment of posterior segment diseases.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan, 33302, Taiwan.

ABSTRACT
The interaction between cells and biopolymers is the evaluation indicator of the biocompatibility of materials. The purpose of this work was to examine the responses of retinal pigment epithelial (RPE) cells to genipin (GP) or glutaraldehyde (GTA) cross-linked chitosan by means of cell viability assays, cytokine expression analyses, and apoptosis assays. Evaluations of non-cross-linked chitosan were conducted simultaneously for comparison. Both GP and GTA treated samples with the same extent of cross-linking (around 80%) were prepared by varying cross-linking time. Our results showed that GP cross-linking was carried out by either radical polymerization of the monomers or S(N)2 nucleophilic substitution reaction involving the replacement of the ester group on the monomer with a secondary amide linkage. On the other hand, GTA could react with free amino groups of chitosan, leading to the formation of either the Schiff bases or the Michael-type adducts with terminal aldehydes. The biocompatibility of non-cross-linked chitosan membranes was demonstrated by the absence of any signs of toxicity or inflammation reaction. The present study showed that the ARPE-19 cells exposed to GTA cross-linked chitosan membranes had significantly higher cytotoxicity, interleukin-6 levels, and number of TUNEL-positive nuclei than did those exposed to GP treated samples. In addition, the materials modified with GTA trigger apoptosis at an early stage and may induce toxicity in the RPE cells later. The findings suggest that while the chitosan molecules bridged by GP are satisfactorily cytocompatible, the counterparts treated by GTA do not seem to be tolerated. In terms of material safety, the GP cross-linked chitosan may be compatible with human RPE cells and may have a potential application as delivery carriers in the treatment of posterior segment diseases.

Show MeSH
Related in: MedlinePlus