Biocompatibility Study Of The Nano-copper Complex/Polymer Composite As A Novel Intra-vas Device Material

Part I Cytotoxicity evaluation of the nano-copper complex/polymer composite in vitroObjective To evaluate the cytotoxicity of the nano-copper complex/polymer composite in vitro so as to provide experimental evidence for its clinical application.Methods The cytotoxicity of the nano-copper complex/polymer composite was evaluated with MTT assay and direct contact assay using the cell line L929 in vitro. L929 cells were cultured with different concentrations of the extracts of the nano-copper complex/polymer composite (25%, 50% and 100%, V/V) in vitro, and MTT assay was performed to evaluate the cell proliferation at 2, 4 and 7 days. The absorption value was recorded at 492 nm wave lenth on enzyme analyzer. The relative growth rate was calculated, and the toxicity of the nano-copper complex/polymer composite was evaluated. L929 cells were cultured directly with the nano-copper complex/polymer composite. The morphologic changes of the cells were observed by inverted microscope at intervals of 24 h, 48 h and 72 h.Results At each point of culture time, the cytotoxicity of all groups were grade 0 or 1 with more than 90% cell proliferation rate except the positive control group was grade 4 with 0～20% cell proliferation rate; except the positive control group, the OD values of experimental groups and negative control group were increased with culture days,and the OD value of the positive control group on day 2, 4 and 7 was much lower than other groups (P<0.01); there were no significant differences between negative control group and experimental group (P>0.05).In the direct contact assay, no side effects were found in regard to the cell growth, proliferation and attachment to the nano-copper complex/polymer composite as compared with the negative control group. The cytotoxicity grade of the nano-copper complex/polyer composite was 0, suggesting that the material was almost free of cytotoxicity.Conclusion The nano-copper complex/polymer composite possesses ideal cellular compatibility, and therefore has great potential for clinical use as a intra-vas device material.PartⅡAcute systemic toxicity evaluation of the nano-copper complex/polymer composite in vivoObjective To investigate the acute systemic toxic reaction induced by the extracts of the nano-copper complex/polymer composite, which were injected in the abdominal cavity of mice.Methods Twenty healthy mice about 20 g body weight were equally divided between male and female and divided into two groups randomly, their initial body weight was recorded. Mice of the experimental group were injected the extracts of the material through intraperitoneal injection, the dosage was 50 g/kg body weight. And the control group were injected with saline. After 24 h, 48 h and 72 h, the general behavior, toxicity manifestation and death number was observed. The change of body weight was recorded. And their venous blood was extracted to detect indicatrix of hematology after experiment. Results No death of the mice was found in all the mice within the 3 days, and no toxicosis symptom or ill effect was found within the 3 days. There was no significant difference between the experimental group and control group (P>0.05). Data of ALT, AST, TP, ALB, GLU, BUN and CRE had no significant difference between the experimental group and control group (P>0.05).Conclusion The nano-copper complex/polymer composite has no acute toxicity in mice.PartⅢMuscular implantation test of the nano-copper complex/polymer composite in vivoObjective To evaluate the tissue reaction and tissue biocompatibility of the nano-copper complex/polymer composite, which was implanted into the back of rabbits. The specimens were harvested at different point respectively and examined by histology after implanting in vivo.Methods The nano-copper complex/polymer composite was implanted into the spinal muscle of fifteen New Zealand white rabbits at four points in one side. and the contralateral was sham-operated, every point is between 2 cm at interval, the depth of the material is inserted into 1～2 cm muscle, then sutured muscle and skin. The rabbits were executed after 1, 4 and 12 weeks respectively, took out both sides of the spinal muscles, macroscopic observation of subcutaneous implantation site of abnormal lesions and muscle tissue were performed. At the same time, cut wrapped sample around 0.5～1cm of muscle tissue, hematoxylin-eosin staining and the optical microscopic observation were done to compare the tissue reaction around the implanting site, as well as the thickness of fibrous cysts, inflammatory cells and other cells with that of the sham-operated side. Results In 1 week of implantation, acute inflammatory reaction was the main characteristic of the organism action. Inflammatory response of grade 4 occurred in the muscular tissues around the implants without formation of complete capsule. The acute inflammatory response disappeared gradually in 4 weeks with the traits that heterophil granulocyte disappeared and lymph-cell reduced, and there was compact fibrous capsule formed on the material surface of the tissue. Inflammatory response lowered to grade 2 and the capsular reaction to grade 3. The fibrous capsule reaction became more stable at 12 weeks and both of the inflammatory response and capsular reaction were at grade 1.Conclusion The nano-copper complex/polymer composite causes only mild inflammation after implantation in rabbits, it suggests good biocompatibility of the material in the tissure of the animal body.