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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

Cross-linking index of GP and GTA treated chitosan membranes as a function of cross-linking time. An asterisk indicates statistically significant differences (*p < 0.05; n = 5) for the mean value of cross-linking index compared with the value at the previous time point.
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f2-ijms-11-05256: Cross-linking index of GP and GTA treated chitosan membranes as a function of cross-linking time. An asterisk indicates statistically significant differences (*p < 0.05; n = 5) for the mean value of cross-linking index compared with the value at the previous time point.

Mentions: The ninhydrin assay has been shown to be applicable for determining the extent of cross-linking in amino group containing biomaterials [4,13]. In this study, the cross-linking index of GP and GTA treated chitosan membranes was investigated as a function of cross-linking time (Figure 2). For GP-chi groups, the cross-linking degree significantly increased with increasing reaction time from 1 to 24 h (p < 0.05). A similar trend was found for the GTA-chi groups. At each time point, the cross-linking index of chitosan samples treated with GTA was significantly higher than their counterparts with GP (p < 0.05), which indicated that the GTA demonstrates a higher reactivity during chemical cross-linking. These findings are in accordance with previous results showing that the GTA produces stronger cross-linking to gelatin molecules than using GP [25]. It is known that the cross-linker concentration, pH, temperature, and time greatly affect the cross-linking degree of biomaterials [26], which may further determine their compatibility with cells and tissues. Here, the chitosan membranes were modified with 10 mM of either GP or GTA at 25 °C, pH 7.4 for different time periods (0–24 h). To examine the influence of chemical cross-linkers on the biocompatibility of chitosan materials, the GP and GTA treated samples with the same extent of cross-linking (i.e., 78.5 ± 2.8% for GP-chi at 8 h and 76.8 ± 2.6% for GTA-chi at 3 h) were characterized by various in vitro assays.


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)

Cross-linking index of GP and GTA treated chitosan membranes as a function of cross-linking time. An asterisk indicates statistically significant differences (*p < 0.05; n = 5) for the mean value of cross-linking index compared with the value at the previous time point.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f2-ijms-11-05256: Cross-linking index of GP and GTA treated chitosan membranes as a function of cross-linking time. An asterisk indicates statistically significant differences (*p < 0.05; n = 5) for the mean value of cross-linking index compared with the value at the previous time point.
Mentions: The ninhydrin assay has been shown to be applicable for determining the extent of cross-linking in amino group containing biomaterials [4,13]. In this study, the cross-linking index of GP and GTA treated chitosan membranes was investigated as a function of cross-linking time (Figure 2). For GP-chi groups, the cross-linking degree significantly increased with increasing reaction time from 1 to 24 h (p < 0.05). A similar trend was found for the GTA-chi groups. At each time point, the cross-linking index of chitosan samples treated with GTA was significantly higher than their counterparts with GP (p < 0.05), which indicated that the GTA demonstrates a higher reactivity during chemical cross-linking. These findings are in accordance with previous results showing that the GTA produces stronger cross-linking to gelatin molecules than using GP [25]. It is known that the cross-linker concentration, pH, temperature, and time greatly affect the cross-linking degree of biomaterials [26], which may further determine their compatibility with cells and tissues. Here, the chitosan membranes were modified with 10 mM of either GP or GTA at 25 °C, pH 7.4 for different time periods (0–24 h). To examine the influence of chemical cross-linkers on the biocompatibility of chitosan materials, the GP and GTA treated samples with the same extent of cross-linking (i.e., 78.5 ± 2.8% for GP-chi at 8 h and 76.8 ± 2.6% for GTA-chi at 3 h) were characterized by various in vitro assays.

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