| Objective: To construct two nano-sized bionic-function scaffolds, i.e."nano-silver/chitosan film","nano-gold/chitosan film"bionic function scaffold; and to investigate the efficacy, bio-safety , attached ration and growth of keratinocytes on bionic-function scaffold.Method:"Nano-silver(gold)/chitosan film"bionic-function scaffold were fabricated by self-assembly technology."Nano-silver /chitosan film"bionic-function scaffold was used as nano-silver bionic wound dressing due to distinct antimicrobial properties of nano-silver. The sterility, pyrogen, skin irritation, subcutaneous, systemic acute toxicity tests were performed to evaluate the bio-safety of this nano-silver wound dressing. The efficacy of this wound dressing was tested in vivo experiments on deep partial-thickness wound created on Sprague Dawley rats (SD rats). Therapeutic effect was evaluated through macroscopic observation, calculating healing ration and pathological section. In addition, the concentration of silver in SD rats'blood and other tissues (liver, brain, kidney) was determined by flame atomic absorption spectrophotometry (FAAS). And the results were compared with that of silver sulfadiazine (SD-Ag) group. Keratinocytes were cultured on"nano-gold / chitosan film"bionic-function scaffold. 6h, 12h, 24h after inoculation, the cell attached rations were calculated respectively. The cultured cells were characterized and identified by immunohistochemistry and transmissive electron microscope (TEM).Results: We found that this nano-silver wound dressing was sterility, no pyretogen. It had no irritation and systemic acute toxicity. 10 and 13 post scald day, the wound healing rates were 88.50±4.03% and 98.98±6.09% respectively in nano-silver dressing group. While the healing rates were 67.78±3.86% and 81.67±6.30% in silver sulfadiazine (SD-Ag) group (control group) and were 73.70±3.25%and 88.60±5.21% in chitosan film group (control group). In addition, there were few infected SD rats in nano-silver dressing group. The blood silver level in nano-silver dressing group was lower than that in SD-Ag group (p<0.01). The concentration of blood silver in nano-silver dressing group was 6 times higher than normal level, while in SD-Ag group was 25 times higher than normal level. On day 13 post-operation, the silver level in blood was 0.11±0.06μg/g, which returned normal level nearly (p>0.05, normal silver level was 0.07±0.03μg/g), in nano-silver dressing group and the silver level in SD-Ag group was 4 times higher than normal level. The concentration of tissue silver was higher than normal level in nano-silver dressing and SD-Ag group. But the silver level in tissues was higher than that in nano-silver dressing group (p<0.01). The concentration of liver silver was 11.48±0.32μg/g and 1.64±0.12μg/g in SD-Ag group and nano-silver dressing group, respectively.In comparison to control groups,"nano-gold/chitosan film"bionic-function scaffold could significantly (P<0.01) increase the attached ration of keratinocytes and promote their growth. 24h after inoculation, the attached rations were 58.72±3.40% and 28.61±2.23% on the scaffold and cell culture plastic, respectively. After inoculation 12h, 24h, the attached rations on the scaffold were the highest. However, the attached ration (25.52±3.08%) after inoculation 6h on the scaffold was lower than that of on chitosan film (33.41±1.42%) (P<0.05). In addition, the attached rations (25.52±3.08%, 40.34±1.12%,58.72±3.40%) after inoculation 6h, 12h, 24h of keratinocytes cultured on the scaffold changed obviously. There were nearly no change on chitosan film (33.41±1.42%, 35.52±2.69%, 39.45±1.32%) and cell culture plastic (18.02±6.22%, 26.11±2.56%, 28.61±2.23%).Conclusion: Nano-silver bionic wound dressing was of sterility, no- pyretogen, no irritation and systemic acute toxicity and can accelerate wound healing, and it can decrease the risk of silver poisoning. It could be used on deep partial-thickness wound with a prospect of wide-rage clinical application."Nano-gold/chitosan film"bionic-function scaffold could significantly (P<0.01) increase the attached ration of keratinocytes and promote their growth. The rapidly proliferating keratinocytes, which were cultured on this scaffold, maintained their bioactivity. It was a good candidate for wound dressing in skin tissue engineering.
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