Objective To investigate the biocompatibility of p(3HB-co-3HH) and marrow mesenchymal stell cells (MSCs).Methods MSCs were inoculated to p(3HB-co-3HH), and then cultured for 2-4 weeks in vitro and embedded for 2 weeks in vivo. The growth, proliferation, morphology and phenotype properties of MSCs were observed by use of phase contrast microscope, electron microscope, HE staining and staining of type Ⅰ collagen. Results p(3HB-co-3HH) hadgood compatibility. The inoculated MSCs could be well-distributed, attached well and obtain the phenotype of MSCs in p(3HB-co-3HH). After osteogenic inducer were added, MSCs differentiated to osteoblasts and secreted matrix. Type Ⅰ collagen was stained positively by immunohistochemical techenique. Conclusion The above results demonstrate that there is satisfactory biocompatibility betweenp(3HB-co-3HH) and MSCs.
Objective To study the biocompatibility of tendon mixedextraction of bovine collagen(tMEBC) and to explore the feasibility of using the threedimensional framework as periodontal tissue engineering scaffold. Methods After being prepared, the tMEBC were cultured with the P4P6 of human periodontal ligament fibroblasts (HPDLFs) in vitro. Threedimensional framework was prepared from bovine tendon. The P4-P6 of HPDLFs (with an initial density of 5×106 cells／ml) were cultured in vitro. Cell attachment andproliferation were measured by cell counting 1 day, 3,5, and 10 days after cell seeding. Histological examination was performed with light microscope and scanning electron microscope 5 and 10 days after cell seeding. Results Porous structure, which supported the proliferation and attachment of HPDLFs, was found in tMEBC. The density of cell increased from 0.556×104 cells／ml 24 hours after cell seeding to 3.944×104 cells／ml 10 days after seeding. Light and scanning electron microscope examinationindicated that HPDLFs were attached and extended on the three-dimensional scaffolds and were well embedded in the newly formed tissue matrix. ConclusiontMEBC has good biocompatibility with the HPDLFs, and can be used as scaffold for cell transplantation in periodontal tissue engineering.
ObjectiveTo evaluate the biocompatibility of poly lactic acid/bone matrix gelatin (PLA/BMG) composite biomaterial so as to lay a foundation for bone defect repair.
MethodsRats'MC3T3-E1 cells were cultured with leaching solution of PLA/BMG and PLA material respectively for 7 days. The cell proliferation rate was tested by MTT and cell toxicity grading was carried out everyday. The PLA/BMG and MC3T3-E1 cells were co-cultured, the cell shape and proliferation were observed by inverted phase contrast microscope at 1, 3, and 5 days and cell adhesion by scanning electron microscope at 5 days. The PLA and PLA/BMG were implanted subcutaneously in 15 Wistar rats. The histological observation was done, and the thickness of fibrous membrane, the number of inflammatory cells, and the vascularization area were measured at postoperative 2nd, 4th, and 8th week.
ResultsThe tests for cytotoxicity in vitro showed that the cell proliferation rates were over 100% and the cell cytotoxic grades were grade 0 at 1-7 days in PLA/BMG group. While in PLA group, the cell proliferation rates were less than 100% and the cell cytotoxic grades were grade 1 at 2, 4, and 7 days. After co-culture of PLA/BMG and MC3T3-E1 cells, cells grew on the surface and in the pores of PLA/BMG, and the cellular morphology was triangle or polygon with abundant microvillus on the surface. After subcutaneous implantation, the rats survived to the end of experiment, and incision healed well. PLA was wrapped by connective tissue where there were a lot of lymphocytes and neutrophilic granulocytes. The cells and tissue grew slowly in PLA. The PLA/BMG materials were wrapped by little connective tissue where there were a few inflammatory cells. The connective tissue ingrowth was observed in the center of PLA/BMG. There was no significant difference in the thickness of fibrous membrane between 2 groups at each time point (P>0.05). The number of inflammatory cells of PLA/BMG group were significantly less than those in PLA group at 2, 4, and 8 weeks (P<0.05); the vascularization area was significantly larger than that in PLA group (P<0.05).
ConclusionPLA/BMG composite biomaterials prepared by super critical-CO2 technique are good in cell and tissue biocompatibilty.
ObjectiveTo explore the influence of three central venous catheter biomedical materials (polyurethane, silicone, and polyvinyl chloride) on the proliferation, apoptosis, and cell cycle of Xuanwei Lung Cancer-05 (XWLC-05) cells so as to provide the basis for clinical choice of central venous catheter. MethodsXWLC-05 cells were cultured and subcultured, and the cells at passage 3 were cultured with polyurethane, silicone, and polyvinyl chloride (1.0 cm × 1.0 cm in size), and only cells served as a control. At 24, 48, and 72 hours after cultured, MTT assay was used to detect the cellular proliferation and flow cytometry to detect the cell cycle and apoptosis. At 72 hours after cultured, inverted microscope was used to observe the cell growth. ResultsInverted microscope showed the cells grew well in control group, polyurethane group, and silicone group. In polyvinyl chloride group, the cells decreased, necrosed, and dissolved; residual adherent cells had morphologic deformity and decreased transmittance. At 24 and 48 hours, no significant difference in proliferation, apoptosis, and cell cycle was found among 4 groups (P gt; 0.05). At 72 hours, the proliferations of XWLC-05 cells in three material groups were significantly inhibited when compared with control group (P lt; 0.05), and the cells in polyvinyl chloride group had more significant proliferation inhibition than polyurethane group and silicone group (P lt; 0.05), but there was no signifcant difference in proliferation inhibition between polyurethane group and silicone group (P gt; 0.05). Compared with the control group, three material groups had significant impact on the rate of apoptosis and cell cycle: polyvinyl chloride group was the most remarkable, followed by silicone group, polyurethane group was minimum (P lt; 0.05). ConclusionPolyvinyl chloride can significantly impact the proliferation, apoptosis, and cell cycle of XWLC-05 cells; polyurethane has better biocompatibility than polyvinyl chloride and silicone
Objective To investigate the feasibility of a new kind of porous β tricalcium phosphate (β-TCP) as a scaffold for the bone tissue engineering Methods The inverted phase contrast microscope was used to observe the growth of the marrow mesenchymal stem cells (MSCs) in the experimentalgroup and the control group at 10 days.In the experimental group, the MSCs were cultured with β-TCP(3 mm×3 mm×3 mm) in the 24-hole cultivation board, and in the control to control group, only MSCs were cultivated. The scanning electron microscope was used to observe growth of MSCs at 6 days. Cultivated with β-TCP at 3, 6, 9, 12 days, the MTT assay was used to judge the biocompatibility. The cytotoxicity was analyzed with the method that used the different density(100%， 50%， 10%， 1%，0%) leaching liquor gained from β-TCP to raise MSCs. MSCs were induced into the osteoblasts and were mixed with β-TCP, and the composite was used to repair a large radius bone defect in the rabbit. The specimens were made at 2,6,12 weeks. The histology imageology, and the radionuclide bone scan were used to analyze the bone formation. Results Some MSCs had a good adherence 4 hours after MSCs were inoculated and had a complete adherence at 12 hours. The cells were shaped like polyangle, spindle or converge monolayer after 8-10 days. The cells in the two groups had no difference. The cell adhesion was good, when observed by the inverted phase contrast microscope and the scanning electron microscope at 6 days. MTT showed that the absorbance (A)was not statistically different between the experimental group and the control group (P＞0.05); the different density leaching liquor had no cytotoxicity at the different time points. Histology, X-ray, and CT tomograph showed that itcould repair the large radius bone defect in the rabbit and its in vivo degradationrate was the same as the bone formation rate. Conclusion The new porous β-TCP has a unique three dimensional (3D) stereochemical structure and superordinary physicochemical property, and so it is a good scaffold for the bone tissue engineering.
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 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 obtain highly purified and large amount of Schwann cells (SCs) by improved primary culture method, to investigate the biocompatibility of small intestinal submucosa (SIS) and SCs, and to make SIS load nerve growth factor (NGF) through co-culture with SCs.
Methods
Sciatic nerves were isolated from 2-3 days old Sprague Dawley rats and digested with collagenase II and trypsin. SCs were purified by differential adhesion method for 20 minutes and treated with G418 for 48 hours. Then the fibroblasts were further removed by reducing fetal bovine serum to 2.5% in H-DMEM. MTT assay was used to test the proliferation of SCs and the growth curve of SCs was drawn. The purity of SCs was calculated by immunofluorescence staining for S-100. SIS and SCs at passage 3 were co-cultured in vitro. And then the adhesion, proliferation, and differentiation of SCs were investigated by optical microscope and scanning electron microscope (SEM). The NGF content by SCs was also evaluated at 1, 2, 3, 4, 5, and 7 days by ELISA. SCs were removed from SIS by repeated freeze thawing after 3, 5, 7, 10, 13, and 15 days of co-culture. The NGF content in modified SIS was tested by ELISA.
Results
The purity of SCs was more than 98%. MTT assay showed that the SCs entered the logarithmic growth phase on the 3rd day, and reached the plateau phase on the 7th day. SCs well adhered to the surface of SIS by HE staining and SEM; SCs were fusiform in shape with obvious prominence and the protein granules secreted on cellular surface were also observed. Furthermore, ELISA measurement revealed that, co-culture with SIS, SCs secreted NGF prosperously without significant difference when compared with the control group (P gt; 0.05). The NGF content increased with increasing time. The concentration of NGF released from SIS which were cultured with SCs for 10 days was (414.29 ± 20.87) pg/cm2, while in simple SIS was (4.92 ± 2.06) pg/cm2, showing significant difference (P lt; 0.05).
Conclusion
A large number of highly purified SCs can be obtained by digestion with collagenase II and trypsin in combination with 20-minute differential adhesion and selection by G418. SIS possesses good biocompatibility with SCs, providing the basis for further study in vivo to fabricate the artificial nerve conduit.
To explore the histological and the hematological change of rabbits after implanting novel injectable artificial nucleus prostheses, and to evaluate the biological safety. Methods In accordance with Biological Evaluation of Medical Devices, materials of polyurethane, sil icone rubber and macromolecular polyethylene for medical use were made into short column 1 cm in length and 0.3 cm in diameter. Forty-eight SPF New Zealand white rabbits weighing 2.5-3.0 kg were used, and cavity 1 cm in depth was made in the area 2 cm away from the spinal midl ine by separating muscle.Then according to different material being implanted, the rabbits were divided into 3 groups (n=16): Group A, polyurethane; group B, sil icone rubber; group C, macromolecular polyethylene for medical use as negative control. General condition of the rabbits was observed after operation. Gross and histology observation were conducted 1, 4, 12 and 26 weeks after operation. Blood routine, biochemical function and electrolyte assays were performed 26 weeks after operation to observe pathological changes of organs. Meanwhile, physicochemical properties of the materials were detected, and the material in the same batch was used as negative control. Results All rabbits survived until the end of experiment, and all wounds healed by first intention. In each group, red swollen muscles were observed 1 week after operation and disappeared 4 weeks after operation, connective tissue around the implanted materials occurred 12 and 26 weeks after operation. At 26 weeks after operation, there were no significant differences among three groups in blood routine, biochemical function and electrolyte assays (P gt; 0.05). Organs had smooth surface without ulceration, ecchymosis, obvious swell ing, hyperemia or bleeding, and nodules. There were no significant differences among three groups in percentage weight of each organ (P gt; 0.05). Histology observation: granulation tissue prol iferation and inflammatory cell infiltration were observed in each group 1 week after operation, fibrous capsule formation around the materials and the disappearance of inflammatory cell infiltration were evident 4 weeks after operation, cyst wall grewover time and achieved stabil ity 12 weeks after operation. The inflammatory response and the fiber cyst cavity of groups A and B met the standard of GB/T 16175 and were in l ine with group C. No specific pathological changes were discovered in the organs 26 weeks after operation. For group A, no significant difference was evident between before and after material implantation in terms of weight average molecular weight, number average molecular weight, tensile strength at break and elongation at break (P gt; 0.05). For group B, no significant difference was evident between before and after material implantation in shore hardness (P gt; 0.05). Conclusion Novel injectable nucleus pulposus prostheses do not damage local tissue and function of organs, but provide good biocompatibil ity and biological safety.
The poor mechanical property and vulnerability to bacterial infections are the main problems in clinic for dental restoration resins. Based on this problem, the purpose of this study is to synthesize silver-titanium dioxide (Ag-TiO2) nanoparticles with good photocatalytic properties, and add them to the composite resin to improve the mechanical properties and photocatalytic antibacterial capability of the resin. The microstructure and chemical composition of Ag-TiO2 nanoparticles and composite resins were characterized. The results indicated that Ag existed in both metallic and silver oxide state in the Ag-TiO2, and Ag-TiO2 nanoparticles were uniformly dispersed in the resins. The results of mechanical experiments suggested that the mechanical properties of the composite resin were significantly improved due to the incorporation of Ag-TiO2 nanoparticles. The antibacterial results indicated that the Ag-TiO2 nanoparticle-filled composite resins exhibited excellent antibacterial activities under 660 nm light irradiation for 10 min due to the photocatalysis, and the Ag-TiO2 nanoparticle-filled composite resins could also exhibit excellent antibacterial activities after contact with bacteria for 24 h without light irradiation because of the release of Ag ions. In summary, this study provides a new antibacterial idea for the field of dental composite resins.
