Decellularized tissue engineering scaffolds appear to have the properties of similar structure and mechanical characteristics to native tissues，good biocompatibility，suitability for cell adhesion，growth and angiogenesis induction，and non-immunogenicity. Genipin has anti-inflammatory，antithrombotic and antioxidative features which can considerably suppress vascular and endothelial inflammatory activation，increase mechanical strength of biological scaffolds，inhibit inflammatory response and decrease degradation rate of biological scaffolds. By cross-linking with decellularized matrices，Genipin can further improve corresponding performance of tissue engineering matrices，which is very helpful to promote the application of tissue engineering into clinical practice of cardiothoracic surgery. This review focuses on recent research process and possible prospects of Genipin cross-linking in tissue engineering in the field of cardiothoracic surgery.
Objective To fabricate in situ crosslinking hyaluronic acid hydrogel and evaluate its biocompatibility in vitro. Methods The acrylic acid chloride and polyethylene glycol were added to prepare crosslinking agent polyethylene glycol acrylate (PEGDA), and the molecular structure of PEGDA was analyzed by Flourier transformation infrared spectroscopy and 1H nuclear magnetic resonance spectroscopy. Hyaluronic acid hydrogel was chemically modified to prepare hyaluronic acid thiolation (HA-SH). And the degree of HA-SH was analyzed qualitatively and quantitatively by Ellman method. HA-SH solution in concentrations (W/V) of 0.5%, 1.0%, and 1.5% and PEGDA solution in concentrations (W/V) of 2%, 4%, and 6% were prepared with PBS. The two solutions were mixed in different ratios, and in situ crosslinking hyaluronic acid hydrogel was obtained; the crosslinking time was recorded. The cellular toxicity of in situ crosslinking hyaluronic acid hydrogel (1.5% HA-SH and 4% PEGDA mixed) was tested by L929 cells. Meanwhile, the biocompatibility of hydrogel was tested by co-cultured with human bone mesenchymal stem cells (hBMSCs). Results Flourier transformation infrared spectroscopy showed that most hydroxyl groups were replaced by acrylate groups; 1H nuclear magnetic resonance spectroscopy showed 3 characteristic peaks of hydrogen representing acrylate and olefinic bond at 5-7 ppm. The thiolation yield of HA-SH was 65.4%. In situ crosslinking time of hyaluronic acid hydrogel was 2 to 70 minutes in the PEGDA concentrations of 2%-6% and HA-SH concentrations of 0.5%-1.5%. The hyaluronic acid hydrogel appeared to be transparent. The toxicity grade of leaching solution of hydrogel was grade 1. hBMSCs grew well and distributed evenly in hydrogel with a very high viability. Conclusion In situ crosslinking hyaluronic acid hydrogel has low cytotoxicity, good biocompatibility, and controllable crosslinking time, so it could be used as a potential tissue engineered scaffold or repairing material for tissue regeneration.
Objective To evaluate the biocompatibil ity of a new nano TCP/ gelatin / velvet antler polypeptide material. Methods The nano TCP/ gelatin / velvet antler polypeptide material was prepared, and the morphous was observed by scanning electron microscope. L929 and NIH/3T3 cell l ines were cultured conventionally. Acute toxicity test, hemolysis test, cell prol iferation and cytotoxicity test were used to evaluate the biocompatibil ity of the material. Results The compositemicrosphere material was about 10 μm in diamerter and had good spherical geometry, high monodispersity with nanometer size holes on the surface. Toxic symptoms such as hyperspasmia, palsy and death did not appear during the observing stage in acute toxicity test. Maximum hemolysis rate of the material was less than 5% which met the requirement of hemolysis test standard as a medical material. Different concentrations of the materials leaching l iquor could enhance the prol iferation of NIH/3T3 cells, which showed the good biologic activity. Toxicity grade was 0, and the material was no cytotoxic. Conclusion Nano TCP/ gelatin / velvet antler polypeptide material has good biocompatibil ity.
Objective To choose the best procedure on preparation of acellularbovine pericardium (ABP) guided bone regeneration (GBR) material. Methods The BP was decellularized with 0.25% Trypsin+0.5% Triton X-100. The acellular bovine pericardiums (ABPs) were treated with phosphatebuffered saline(PBS) (group A), 95% glycerol (group B), EDAC (group C), and EDAC and 95% glycerol (group D) respectively. The treated ABPs were implanted subcutaneously in the back of SD rats respectively at random and no material was implanted as control. Seven rats were sacrificed at 2 weeks, twelve at 4 weeks, twelve at 8 weeks, seven at 16 weeks. Local reaction was studied grossly. The amount of antigen presenting cell (APC) and the percentage of ABP degeneration were reckoned by images analysis system. Results The ABPs were replaced by fibroblasts completely in group A at 8 weeks， in group C at 16 weeks, but only less than 50% till 16 weeks in groups B and D. In all groups, the depth of surrounding fibres attenuated timedependingly. The APC amount of the groups B and D was higher than that of the control group, and the ABP of the groups B and D degraded partly at 16 weeks. Conclusion The ABP treated with EDAC can be replaced by the surrounding tissues and has good biocompatibility.
Objective To investigate the influence of different dose levels of hydroxyapatite/tricalcium phosphate (HA/TCP) on the proliferation and alkalinephosphatase (ALP) activity of rabbit osteoblasts. Methods Three different doselevels of HA/TCP (10%, 40%, 70%) were co-cultivated with rabbit osteoblasts respectively. The proliferation and ALP expression capacity of osteoblasts were examined with MTT method and enzyme histochemistry once every 24 hours until 5 days. Three control groups of other materials were treated and examined in the sameway: rabbit osteoblasts as normal control; polyvinylchloride as positive control; titanium alloy as negative control. Results There was remarkable timeeffect relationship in the proliferation of osteoblasts. Ten percent HA/TCP did not affect osteoblasts growth while 40% HA/TCP could slow the cell growth rate down though time-effect relationship still existed. The proliferation of osteoblasts stagnated when co-cultivated with 70% HA/TCP. On the other hand, 10% HA/TCP could cause reversible damage on ALP activity of osteoblasts, whereas when the dose was40%, and the cultivation lasted 6 days the damage was irreversible. Three different dose levels of titanium alloy (10%, 40%, 70%) had no effect on the proliferation or ALP activity of osteoblasts. Conclusion Dosage is an important factor affecting the biocompatibility evaluation of biomaterial. It suggests that dose choosing should be more specified upon each individual biomaterial. It also indicates that ALP may be a good supplementary index of the cell compatibility of material.
Objective To evaluate the biocompatibility and in vivo degradation of novel chest wall prosthesis materials and provide some data for their clinical application. MethodsAccording to the standard for the biological evaluation of the medical devices, several tests were performed to evaluate the tissue toxic effects induced by polydioxanone (Group A), chitosan (Group B), and hydroxyapitite/collagen (Group C),which were tested as component materials of the chest wall prosthesis. In the hemolysis test, 0.2 ml of the anticoagulant rabbit blood was added to the component materials and the normal saline (negative control) and to the distilled water(positive control). Five samples were made in each group. Absorbency was measured and the hemolysis rate was determined. In the acute systemic toxicity test, 20 mice were randomly divided into 4 groups (Groups A, B and C, and the normal saline group, n=5). The leaching liquid (50 ml/kg) was injected through the caudal vein, which was observed at 24, 48 and 72 hours. In the pyrogen test, 12 rabbits were randomly divided into 4 groups (Groups A, B, C and the normal saline group, n=3) the leaching liquid(10 ml/kg) was injected through the ear vein,and the body temperature was recorded within 3 hours. In the in vivo degradable test, the component materials (10 mm×10 mm) were implanted in 12 rabbits at 2, 4, 8, 12, 16 and 24 weeks, respectively, after operation. Two rabbitswere sacrificed for the macroscopic and the microscopic examinations. Results The chest wall component materials had no hemolytic reaction, no acute systemic toxicity, and no pyrogen reaction. The results demonstrated that the implanted materials had only a mild inflammatory reaction during the early days of the grafting, which subsided gradually. There was no tissue denaturation, necrosis or pathological hyperplasia when the prosthesis materials were degraded. Conclusion The degradable materials of the chest wall prosthesis have a good biocompatibility and agreat biological safety though their surgical application still requires a further clinical research.
Objective To evaluate the biocompatibility and safety of a novel orthopedics materials-graded zirconia(ZrO2)hydroxyapatite(HA) composite biomaterials. Methods First, ultrafine powers of ZrO2 and HA powder were prepared by chemical precipitation method, then graded ZrO2-HA composite was synthesized by dry-laying and sintering method. After the physiological saline and culture medium extracts of the composite were prepared, four experiments were conducted as follows:① The mouse acute toxic test consists of 2 groups(n=10). The extracts were intravenously injected to mice in the first group, and physiological saline to mice in the second group. The dose was 50 g/kg. Their toxicity manifestation, morality and the change of weight were recorded.② The standard curve of proliferation and metabolism of L929 cells was established. ③ The cytotoxinic test consists of 3 groups: materials group (extracts of the materials), positive control group (culture fluid with 0.64% phenol), and negative control group (RPMI-1640 culture fluid). Each of three was cultured with cell suspension, and then the morphology of the cells was observed, the relative proliferation rate (RGR) was calculated, and the toxicity was classified. ④ In vitrohemolytic test was divided into 3 groups: extracts, sterile distilled water (positive control) and 0.9% physiological saline. In each of three, 0.2 ml anticoagulant diluted fresh rabbit blood was added. The percentage of hemolysis was tested. ⑤ The muscle and implantation test were divided into 4 groups(n=3). The composite biomaterials were implanted into pygal muscleson either side and lateral condyles of femurs. After surgery, the rats of four groups were sacrificed at 12 and 24 weeks respectively.Tissue slice and scanning electronic microscopy were performed. Results General acute toxic test: no mouse died within 3 weeks; no toxicity symptom or adverse effects were shown within 3 days. The weight of materials group increased by 3.57±0.49 g, and the control group by 3.62±0.61 g, showing no statistically significant difference（Ρgt;0.05）.The standard curve of L929 cell perliferation and metabolism showed that their existed a positive correlation between the number of L929 cells and the perliferation. ③ Cytotoxinic test: cytosomes in the positive control group diminished and appeared round, there were pyknotic nucleus, the attached cells agglomerated; the toxicity was level Ⅳ. The morphology of cells in materials groupand negative control group was normal, and the number of them increased; the toxicity was level Ⅰand level 0, respectively. The MTT color experiments showed that positive control group was significantly lower than materials group and negative control group, showing statistically significant difference (Plt;0.01); there was no statistically significant difference between materials group and negative group.④ Hemolytic test: in vitrohemolytic rate of negative control group was0, of positive control group was 100%, and of materials group was 1.66%, which accords with the standard that hemolytic rate should be lower than 5% specified in ISO. ⑤ Implant test:No apparent rejection reaction took place after the composite was implanted; the composite bonded with the bones of the receptors firmly, which had good bonedinduced effect. Conclusion Graded ZrO2-HA composite bioceramic has good biocompatibility and is suitable for orthopedic biomaterials.
Objective To observe the biocompatibil ity of self-assembled FGL peptide nano-fibers scaffold with neural stem cells (NSCs). Methods FGL peptide-amphiphile (FGL-PA) was synthesized by sol id-phase peptide synthesistechnique and thereafter It was analyzed and determined by high-performance l iquid chromatography (HPLC) and massspectrometry (MS). The diluted hydrochloric acid was added into FGL-PA solution to reduce the pH value and accordinglyinduce self-assembly. The morphological features of the assembled material were studied by transmission electron microscope (TEM). NSCs were cultured and different concentrations of FGL-PA assembled material were added with the terminal concentrations of 0, 50, 100, 200, 400 mg/L, respectively. CCK-8 kit was used to test the effect of FGL assembled material on prol iferation of NSCs. NSCs were added into differentiation mediums (control group: DMEM/F12 medium containing 2% B27 supplement and 10% FBS; experimental group: DMEM/F12 medium containing 2% B27 supplement, 10% FBS and 100 mg/L FGL-PA, respectively). Immunofluorescence was appl ied to test the effect of FGL-PA assembled material on differentiation of NSCs. Results FGL-PA could be self-assembled to form a gel. TEM showed the self-assembled gel was nano-fibers with diameter of 10-20 nm and length of hundreds nanometers. After NSCs were incubated for 48 hours with different concentrations of FGL-PA assembled material, the result of CCK-8 assay showed that FGL-PA with concentrations of 50, 100 or 200 mg/L could promote the prol iferation of NSCs and absorbance of them was increased (P lt; 0.05). Immunofluorescence analysis notified that the differentiation ratio of neurons from NSCs in control group and experimental group were 46.35% ± 1.27% and 72.85% ± 1.35%, respectively, when NSCs were induced to differentiation for 14 days, showing significant difference between 2 groups (P lt; 0.05). Conclusion FGL-PA can self-assemble to nano-fiber gel, which has good biocompatibil ity and neural bioactivity.
OBJECTIVE To investigate the feasibility of freeze-dried demineralized bone matrix (FDBM) as scaffold material in bone tissue engineering. METHODS Osteoblasts which were isolated from cranial periosteum of New Zealand rabbits were cultured as the seeding cells, then the cells were cocultured with heterogenous FDBM in vitro. The cell-material complex was observed under phase microscope, light microscope and electronic scanning microscope in order to evaluate the interaction between cells and FDBM. RESULTS Eight hours after coculture, the osteoblasts adhered to FDBM scaffolds. Seven days later, the osteoblasts differentiated and proliferated in FDBM network. Extracellular matrix was secreted and calcium nodes were formed among osteoblasts. CONCLUSION FDBM is a good scaffold material for the bone tissue engineering.
Objective To review the current researches of scaffold materials for skeletal muscle tissue engineering, to predict the development trend of scaffold materials in skeletal muscle tissue engineering in future. Methods The related l iterature on skeletal muscle tissue engineering, involving categories and properties of scaffold materials, preparative techniqueand biocompatibil ity, was summarized and analyzed. Results Various scaffold materials were used in skeletal muscle tissue engineering, including inorganic biomaterials, biodegradable polymers, natural biomaterial, and biomedical composites. According to different needs of the research, various scaffolds were prepared due to different biomaterials, preparative techniques, and surface modifications. Conclusion The development trend and perspective of skeletal muscle tissue engineering are the use of composite materials, and the preparation of composite scaffolds and surface modification according to the specific functions of scaffolds.