Preparation and in vitro evaluation of vancomycin hydrochloride@polylactic acid-glycolic acid copolymer-chitosan-hyaluronic acid composite sustained-release microspheres北大核心

BACKGROUND: Local antibiotic slow-release system can solve the problems of total toxicity caused by systemic antibiotics and short half-life of short-term local antibiotics. OBJECTIVE: To prepare polylactic acid-glycolic acid copolymer-chitosan-hyaluronic acid composite sustained-release microspheres loaded with vancomycin and evaluate its performance. METHODS: Vancomycin-loaded polylactic acid-glycolic acid copolymer-chitosan-hyaluronic acid composite sustained-release microspheres and unloaded polylactic acid-glycolic acid copolymer-chitosan-hyaluronic acid composite microspheres were prepared by emulsion method. Mass concentrations of vancomycin in the drug-loaded microspheres were 25, 50, and 100 g/L. The drug-loading amount, encapsulation efficiency, and in vitro sustained release properties of the drug-loaded microspheres were detected. The three kinds of drug-loaded microspheres were co-cultured with Staphylococcus aureus bacteria separately, and the antibacterial rate was detected within the corresponding time points. The four kinds of microsphere extracts were co-cultured with MC3T3-E1 cells and MG-63 cells, and the cytotoxicity was detected by CCK-8 method after 1, 3, and 7 days of culture. RESULTS AND CONCLUSION: (1) The encapsulation efficiencies of 25, 50, and 100 g/L drug-loaded microspheres were (79.70±5.11)%, (86.41±3.91)%, and (63.18±1.96)%, and the drug loading was (3.98±0.26)%, (8.64±0.39)%, and (12.63±0.39)%. The encapsulation efficiency of 50 g/L drug-loaded microspheres was higher than that of 100 g/L drug-loaded microspheres (P < 0.05). The drug loading of 100 g/L drug-loaded microspheres was higher than that of the other two groups (P < 0.05). (2) Three kinds of drug-loaded microspheres had no obvious burst release within 24 hours, of which the drug release rate of 50 g/L drug-loaded microspheres at different time points was faster than that of the other two groups. The drug release amount of 100 g/L drug-loaded microspheres at different time points was higher than that of the other two groups, and the drug mass concentration of the three groups was higher than minimum antibacterial concentration of vancomycin at 56 days. (3) All three kinds of drug-loaded microspheres could effectively kill Staphylococcus aureus within a certain period of time. From 14 to 28 days, the relative colony rate of the three kinds of microspheres was lower than 3%, indicating that three kinds of drug-loaded microspheres can continuously and effectively kill Staphylococcus aureus. (4) The 25 and 50 g/L drug-loaded microspheres have no obvious cytotoxicity to MC3T3-E1 cells and MG-63 cells, and 100 g/L drug-loaded microspheres have certain cytotoxicity. (5) The results show that the VA@PLGA-CS-HA microspheres have good sustained-release performance, antibacterial ability and biological tissue compatibility. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

N-乙酰半胱氨酸丝素微球复合磷酸钙骨水泥的制备及表征

BACKGROUND: Calcium phosphate bone cement has been applied to clinical surgery because of its good biocompatibility and osteoconduction. However poor mechanical properties and lack of osteoinductivity limit its wide application. OBJECTIVE: To develop calcium phosphate cement incorporated with N-acetylcysteine (NAC) loaded silk fibroin microspheres (SFM), which is a kind of new injectable bone graft material with slow-release function, and evaluate its physical and chemical properties and cell compatibility. METHODS: Empty SFMs were prepared with emulsion solvent evaporation to absorb NAC solution of different concentrations by NAC-SFM and the concentration of NAC at the maximum drug loading ratio was determined. Then, NAC-SFM was loaded into calcium phosphate bone cement to test the drug release properties in vitro. MC3T3-E1 osteoblasts were cultured on the surface of NAC-SFM calcium phosphate bone cement and cell attachment and growth were observed by scanning electron microscope. Additionally, MC3T3-E1 cells were cultured with three kinds of bone cement extracts (calcium phosphate cement, SFM-calcium phosphate cement, NAC-SFM-calcium phosphate cement, as well as cultured in the α-minimum essential medium containing a volume fraction of 10% fetal bovine serum and 1% penicillin-streptomycin double antibody as the control. MTS assay was used to evaluate cell proliferation. RESULTS AND CONCLUSION: Microspheres in the composite bone cement presented with smooth surface, same size, diffused distribution and no obvious destroy. Thus, the SFM could remain stable in the reaction process of the composite bone cement. The double slow release system which contained silk fibroin microspheres and calcium phosphate bone cement showed a significant decrease in the cumulative release percentage of NAC within the first 24 hours compared with the control group (P < 0.05). In the next 28 days, the release speed of NAC was significantly lower in the NAC-SFM-calcium phosphate cement group than the calcium phosphate cement group (P < 0.05). In addition, different extracts had no significant cytotoxicity to the growth of MC3TC-E1 cells. Thus, the NAC-SFM-calcium phosphate cement has good cytocompatibility, which provide a new insight into the development of bone repair biomaterials. 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

鞘内注射97-9-G4 PLGA缓释微球对神经病理陛疼痛大鼠痛行为的影响

Objective To study the pain killing effects of intratheeal injection of 97-9-G4 PLGA sustained-release microspheres in the SD rat model of CCI of sciatic nerve. Methods SD rats with intrathecal catheter deployment were randomly divided into 4 groups(n=8): normal saline group(NS group), 97-9-G4 group (G group), 97-9-G4 PLGA sustained-release microspheres group(SRG group) and blank PLGA group(P group). On the 11th day after CCI, the animals in NS group and G group were administered the relevant drugs three times (7 am, 1 pm and 7 pm; i.t.). Those in SRG group and P group were administered only once (7 am; i.t.). Paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) were measured 5 times at the fixed time-point within the period of study. Results The drug entrapment efficiency was 80.4%, drug loading efficiency was 27.78%, and the mean diameter of microspheres was 77 μm. Two peaks of 97-9-G4 were observed in vitro release measurement. About 25.28% was released in the first 30 rain and about 71.33% was accumulated released in the period from 30 rain to 24 hours. Till 10 days, about 98.3% of 97-9-G4 was accumulated released from the microspheres. In vivo, compared with basic pain threshold, all groups, except sham group exhibited similar values of PWMT and PWTL at the beginning of drug administration. After drug administration, compared with NS group, the values of PWMT and PWTL were significantly increased at 8 am, 2 pm and 8 pro(P＜0.05). At 12 am and 6 pm, there was no significant difference (P＞0.05). Compared with NS group, the values of PWMT and PWTL in group SRG were significantly increased at all the time points except at 8 am. No significant difference was demonstrated in the values of group P as compared with group NS (P＞0.05). Conclusion 97-9-G4 PLGA sustained-release microspheres demonstrate good appearance and significant in vitro sustained-release characteristics. In rat CCI models, the microspheres demonstrate good sustained-release effect and efficiently raised the PWMT and PWTL. The sustained-release effect could last more than 12 hours in vivo.     

Atorvastatin effects on proliferation and apoptosis of rat bone marrow-derived endothelial progenitor cells in vitro北大核心

BACKGROUND: Atorvastatin has a cardiovascular protective effect that significantly improves endothelial function and promotes the mobilization, migration, and differentiation of endothelial progenitor cells. However, the screening of atorvastatin concentration for in vitro cell culture is not well documented. OBJECTIVE: To investigate the effects of different concentrations of atorvastatin on rat bone marrow-derived EPCs growth characteristics. METHODS: Bone marrow mononuclear cells from Sprague-Dawley rats were induced in selective culture fluid to culture EPCs. Immunofluorescence staining was used to identify cell surface markers. Harvested EPCs were divided into control group and atorvastatin groups with four different concentrations (0.01, 0.1, 1, and 10 µmol/L) for culture. The growth and proliferation of EPCs were observed under light microscope and MTT assay. Flow cytometry was used to detect apoptosis in EPCs. Nitric oxide and endothelial nitric oxide synthase levels in the culture fluid were measured by nitrate reductase method. RESULTS AND CONCLUSION: The number of cells tended to increase in the control and atorvastatin groups, and it was highest in the 1 µmol/L atorvastatin group. The cell number in the 10 µmol/L atorvastatin group began to decrease at 7 days of culture. Among the five groups, the apoptotic rate of cells was lowest in the 1 µmol/L atorvastatin group and highest in the 10 µmol/L atorvastatin group. The levels of nitric oxide and endothelial nitric oxide synthase were significantly higher in the 0.01, 0.1 and 1.0 µmol/L atorvastatin groups compared with the control group (P < 0.01), but lower in the 10 µmol/L atorvastatin group compare with the other groups (P < 0.01). Overall, atorvastatin can promote the proliferation of endothelial progenitor cells and reduce apoptosis by increasing the production of endothelial nitric oxide synthase and nitric oxide, and 1 µmol/L atorvastatin is most suitable for the EPCs culture. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Preparation and characteristics of ciprofloxacin hydrochloride/Carboxymethyl chitosan implanted microsphere

The applications of biodegradable polymeric material as drug carrier in medicineare triggering more interest,especially the microspheres prepared by wrapping drugin polymeric material. For its target to specific organ and tissue and its ability tocontrolled- release of drug,the microspheres can be used to reduce the toxic side ef-fect of drug and improve the biological utilization percentage of drug.In this study,with water- soluble carboxymethyl chitosan( CMC) as carrier and ciprofloxacin hy…     

Influences of Organic Solvents on Particle Size and Drug-loading Efficiency for 5-Fluorouracil Poly（lactic acid） Nanoparticles

The objective of this study was to investigate the influences of organic solvents on particle size, drug content, loading efficiency and yield for 5-Fluorouracil Poly (lactic acid) nanoparticles . The 5-Fluorouracil was entrapped into poly(lactic acid)(PLA) nanoparticles using a water-in-oil-in-water solvent evaporation technique. During the preparation process, ethyl acetate and acetone were used as organic solvents since they are less toxic than the more commonly used dichloromethane. The effect of the three solvents on particle size, drug content, loading efficiency and yield of nanopartcles was compared. When the solvent of the oil phase was acetone, the highest drug content, smallest particle size and lowest yield were obtained for the PLA nanoparticles.     

The precursor frequency of alloreactive CTLs is proportional to the number of T cell epitope specificities

The aim of this study is to find the experimental evidence that the precursor frequency of alloreactive CTLs is proportional to the number of the T-cell epitope specificities. The number of T-cell epitope specificities was manipulated by pulsing different number of HLA-A2 restricted peptide(s) onto the T2 cells, which acted as stimulating cells to elicit allo-reaction by co-culturing with peripheral blood lymphocytes (PBLs) of HLA-A2 negative individual. Ten HLA-A2 restricted peptides (all were normal cell components) were synthesized, and cell peptide extract was prepared by frozen and thawed.T2 cells loaded with different number of peptide(s) were co-cultured with PBLs of an HLA-A2 negative individual; the latter were stained with PKH67 in advance. Then the proliferation was monitored with flow cytometry, and the precursor frequency of the effector cells was analyzed by the ModFit Software. After 6 d of culture, no proliferation was observed in the bulk culture of PBL alone, and obvious proliferation took place when PBLs of the HLA-A2 negative were co-cultured with T2 cells loaded with or without loading peptide(s). The precursor frequency of the alloreactive CTLs was 0.052 819 for co-culture with T2 cells loaded without peptide; however it was 0.030 429 for T2 cells with EBV/ LMP2A and 0. 030 528 for T2 cells loaded with a single autogeneic peptide, and increased up to 0.144 942 for T2 cells loaded with 10 autogeneic peptides; the precursor frequency was 0.203 649 when co-cultured with T2 cells loaded with miscellaneous peptides extracted from the cytoplasm of T2 cells. This study reveals that the precursor frequency of alloreactive CTLs is proportional to the number of T-cell epitope specificities, and independent of the density of the allogeneic HLA ClassⅠmolecule. Our findings support the hypothesis that the alloreactive T cell populations comprise miscellaneous T cell clones; each is specific to corresponding pMHC. The novel constellation of peptides presented by allogeneic MHC molecules makes thousands of different epitopes, which account for the exceptional high precursor frequency of alloreactive T cells.     

聚偏氟乙烯中空纤维血液透析器的研究

Objective Currently applied membrane materials for hemodialysis are mainly polysulfone and polyethers. However, defects still remain in the removal of β2-MG and biocompatibility. Polyvinylidene fluoride (PVDF) has not been put in use in hemodialysis. This study focused on the evaluation of mechanics performance and the dialysis performance of PVDF membrane material. Methods PVDF hollow fiber membranes were spun by changing the membrane forming conditions. The membranes were sealed to dialyzers by centrifugal moulding method. The simulation solution, instead of patient blood, was used to evaluate the dialysis performance of PVDF hollow fiber membrane dialyzers. Results The tensile failure strength of PVDF membrane was about 20 cN. The tensile elongation of PVDF membrane was about 200%. The clearance rates of the dialyzers prepared with the fiber whose wall thickness was 30μ m to urea and lysozyme were 90% and 75%, separately,while the rejection to BAS reached 90% above. With the mass content of (polyethylene glycol, PEG) was 19%,the rejection rate of bovine serum was 91%, which was higher than that with the PEG mass content of 22%. When the flow velocity of dialysis fluid was increased, the clearance rate of urea and lysozyme was improved, but the rejection of BAS was not much affected. Conclusion The results show that the mechanics performance can satisfy the demand of dialysis. The clearance rate of urea and lysozyme has greatly improved. The retention of bovine serum albumin by using PVDF dialyzers is ideal. The PVDF can be exploited as a new membrane material.     

Preparation of Curcumin Chitosan Nanoparticles and Its Effect on Free Radical Injury in Mice with Mycoplasma Pneumoniae Pneumonia

Objective: Curcumin(Cur) and Chitosan(CS) were utilized as primary components for the production of curcumin chitosan nanoparticles. The impact of these nanoparticles on oxidative stress in mycoplasma pneumoniae-infected mice was assessed.Methods: The drug loading and entrapment efficiency of Cur-CS nanoparticles were determined for various feeding ratios, and the release profiles of Cur-CS nanoparticles in different release media were investigated using the dynamic membrane dialysis method.The ...     

THE EXPERIMENTAL STUDY OF NEW COMPOSITED BIOMATERIAL-TITANIUM/BIOCERAMIC FOR REPLACEMENT OF JAW DEFECT

A new composited biomaterial (TGH)was prepared with titanium powder,glasspowder and hydroxylapatite powder,which were mixed well at certain preparationand were sintered in high vacuum and temperature condition.The TGH materialswere used for experimental repairing jaw bone defect,and inserted in mandibles andfemur of native dogs.In addition,they were composited with bovine bone morpho-genetic protein (bBMP)implanted in the same bone.The investigation were per-formed by means of LM.FM.SEM.EDXA.XRD and mechanical test machine,andthe dogs were sacrificed at 1,2,4,8,12,16,20 weeks postimplantation.The results     

紫杉醇聚合物纳米囊泡的制备和体外释放研究

Objective To develop paclitaxol (PTX)-loaded polymersomes based on poly (ε-caprolactone)-block-poly (ethylene glycol)-block-poly (ε-caprolactone)(PCL-b-PEG-b-PCL, PCEP) amphiphilic triblock copolymers. Methods A series of PCEP copolymers was synthesized by ring-opening polymerization of ε-caprolactone initiated by PEG. The block copolymers were characterized by FT-IR、1H NMR and GPC. PTX-loaded polymersomes were prepared by thin-film and ultrasonic dispersion method and characterized in terms of morphology,particle size and size distribution, encapsulation efficiency and in vitro release. The effect of different hydrophilic and hydrophobic chain length on the drug-loading content, entrapment efficiency, size and size distribution and in vitro release were also investigated. Results The PTX-loaded polymersomes showed nanometer size and spherical morphology with core and shell. The sizes of PTX-loaded polymersomes increased with the increasing of the molecular weight of PCEP. The PTX-loaded polymersomes showed a continuous and steady release of PTX without initial burst release. The release rate increased with the increasing of hydrophilic PEG chain content and decreased with the increasing of hydrophobic PCL chain content. Conclusion For the first time, a novel PTX-loaded polymersomes was developed based on PCL-b-PEG-b-PCL amphiphilic triblock copolymers. The PTX-loaded polymersomes showed nanometer size, narrow size distribution and high drug encapsulation efficiency.These results indicate that PTX-loaded polymersomes could be a promising novel controlled release dosage form to increase therapeutic effect with decreased toxic and side effect of PTX.     

紫杉醇聚合物纳米囊泡的制备和体外释放研究

Objective To develop paclitaxol (PTX)-loaded polymersomes based on poly (ε-caprolactone)-block-poly (ethylene glycol)-block-poly (ε-caprolactone)(PCL-b-PEG-b-PCL, PCEP) amphiphilic triblock copolymers. Methods A series of PCEP copolymers was synthesized by ring-opening polymerization of ε-caprolactone initiated by PEG. The block copolymers were characterized by FT-IR、1H NMR and GPC. PTX-loaded polymersomes were prepared by thin-film and ultrasonic dispersion method and characterized in terms of morphology,particle size and size distribution, encapsulation efficiency and in vitro release. The effect of different hydrophilic and hydrophobic chain length on the drug-loading content, entrapment efficiency, size and size distribution and in vitro release were also investigated. Results The PTX-loaded polymersomes showed nanometer size and spherical morphology with core and shell. The sizes of PTX-loaded polymersomes increased with the increasing of the molecular weight of PCEP. The PTX-loaded polymersomes showed a continuous and steady release of PTX without initial burst release. The release rate increased with the increasing of hydrophilic PEG chain content and decreased with the increasing of hydrophobic PCL chain content. Conclusion For the first time, a novel PTX-loaded polymersomes was developed based on PCL-b-PEG-b-PCL amphiphilic triblock copolymers. The PTX-loaded polymersomes showed nanometer size, narrow size distribution and high drug encapsulation efficiency.These results indicate that PTX-loaded polymersomes could be a promising novel controlled release dosage form to increase therapeutic effect with decreased toxic and side effect of PTX.     

3D生物打印构建聚乳酸羟基乙酸/纳米羟基磷灰石支架骨形态发生蛋白2缓释复合体的实验研究

BACKGROUND: Tissue-engineered bone scaffold fabricated by 3D-bioprinting technique has good controllability in morphology and structure. However, construction of tissue-engineered bone/cell growth factor complex and time-dose effect of sustained-release factors are needed to be further researched.  OBJECTIVE: To fabricate a sustained-release composite of polylactic-co-glycolic acid (PLGA)/nano-hydroxyapatite (n-HA) scaffold carrying bone morphogenetic protein-2 (BMP-2) using 3D-bioprinting technique, and test the biological properties of the PLGA/n-HA scaffold carrying BMP-2 and the sustained-release properties, thereby to discuss its feasibility as the tissue-engineered bone scaffold composite.  METHODS: Temperature-sensitive chitosan hydrogel was prepared using chitosan and β-glycerophosphate to construct a sustained-release composite, chitosan nanoparticles carrying BMP-2 . 3D-bioprinting technique was utilized to fabricate the PLGA/n-HA scaffold carrying BMP-2. Biological features of the scaffold composite were tested, and time-dose effect of BMP-2 sustained-release was observed.  RESULTS AND CONCLUSION: The average pore size of the scaffold-cytokine composite was (431.31±18.40) μm, and the porosity was (73.64±1.82)%. The cumulative release rate of BMP-2 from the scaffold-cytokine composite that effectively controlled the burst release during 48 hours and 30 days were suitable for the physiological needs. In conclusion, the porosity, pore size, release property, degradation rate, and mechanical strength of the scaffold-cytokine composite all meet the biological requirements of tissue-engineered bone construction. 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

Study on Preparation and Release of Dexamethasone Hyaluronan Microspheres

Hyaluronan （HA） , the consistent glycosaminoglycans in extracellular matrix, is a kind of biomaterials with wonderful biocompatibility. To develop drug release system （DDS） with HA as drug carrier is a new hotspot in the field of pharmaceutics. In this paper, we applied technique of ultrosound and reversed phase （Water/Oil） emulsification to develop dexamethasone （DEX）-HA-STMP cross-linking microspheres （DEX-HA MS） with STMP as cross-linker. DEX-HA MS has a wonderful shape and property of dispersion. There is a negative correlation between diameter of DEX-HA MS and the content of cross-linker, or the content of emulsifier, and a positive correlation between the diameter and CHA. When CHA ≤ 1% ,DEX/HA≤ 1/10 （g/g） ,there is a positive correlation between the factors mentioned below and drug loading （ DL% ）/loading efficiency （ LE% ）, the content of STMP, the content of emulsifier, CHA and the content of DEX. DEX-HA-MS can realize function of slow release. In vitro drug release experiment shows that cumulative release （CR%） of DEX-HA MS fits in with pervasion-corrosion equation, and there is a negative correlation between the content of STMP, CHA and CR%, a positive correlation between emulsifier and CR%. When DEX/HA ≤ 1/5 （g/g） there is a negative correla- tion between the content of DEX and CR%.     

Preparation and Evaluation in Vitro of BCNU-Loaded PLA Nanoparticles

In this study, using a spontaneous emulsification/solvent extraction method, BCNU-Ioaded PLA nanoparticles (NPs) with small particle size and narrow size distribution have been acquired. The particle size of the NPs ranged from 40-60 nm and 100-200 nm according to different requirements. SEM and TEM showed that the particle size considerably decreases with increasing emulsification concentration and decreasing PLA concentration and ratio of oil to water. The highest drug loading ratio and drug encapsulation efficiency of NPs were 5. 63% and 33.45%. The results demonstrated that decrease of initial BCNU content resuited in a noticeably increased encapsulation yield. A thorough study in vitro showed that the drug could be steadily released from NPs for one week. In addition, drug-loaded NPs had higher antitumor activity, compared with free BCNU,and sustained drug release characteristics as well.     

Study on Tat Mediated Having Composite Magnetic Nanoparticles Targeting Function

This paper describes a new formulation of magnetic nanoparticles coated by a novel polymer matrix-O-Carboxylmethylated Chitosan （O-CMC） as a drug/gene carrier. The O-CMC magnetic nanoparticles were derivatized with a peptide sequence from the HIV-tat protein and transferrin to improve the translocational property and cellar uptake of the nanoparticles. To evaluate the O-MNPs-Tat-Tf as a drug carrier, Methotrexate （MTX） was incorporated as a model drug and MTX-loaded O-MNPs-Tat-Tf with an average diameter of 75 nm were prepared and characterized by TEM, AFM and VSM. The cytotoxicity of MTX-loaded O- MNPs-Tat-Tf was investigated with C6 cells. The results showed that the MTX-loaded O-MNPs-Tat-Tf retained significant antitumor toxicity.     

Entrapment and Sustained Release of Hydrophobic Drugs with Different Molecular Weights from PHBHHx-PEG Nanoparticles

Biodegradable polymeric nanoparticles are more and more frequently used in drug delivery systems, which represent one of the most rapidly developing areas. In our previous study, a novel natural hybrid polyester, polyethylene glycol 200 （PEG200） end-capped poly （3-hydroxybutyrate-co-3-hydroxyhcxanoate） （PHBHHx-PEG） was directly produced by Aeromonas hydrophila fermentation. In this study, the performance of the novel biodegradable PHBHHx-PEG copolyester as a sustained release carrier for hydrophobic drugs with different molecular weights and the in vitro sustained release profile were investigated. 5-Fluorouracil （5-Fu, Mw=130.1）, TGX221 （Mw=364.4）, and Rapamycin （RAP, Mw=914.2） were used as the model drugs. PHBHHx-PEG nanoparticles entrapped with 5-Fu, TGX221 and RAP were fabricated by a modified emulsification/solvent evaporation method, respectively. The average diameter of 5-Fu, TGX221, and RAP loaded PHBHHx-PEG nanoparticles was between 198.2-217.4 nm, and the entrapment efficiency of the three drugs was 62.5%, 93.4% and 91.9%, respectively. The in vitro release profiles of 5-Fu, TGX221 and RAP from PHBHHx-PEG nanoparticles were different. 5-Fu showed faster release rate and an obvious initial burst release phase. TGX221 and RAP were demonstrated to be released more slowly and steadily. The release percentages of 5-Fu, TGX221 and RAP were 97.7%, 85.1% and 74.7% after releasing for 72 h. PHBHHx-PEG is a kind of promising material as a carrier for the entrapment and delivery of hydrophobic drugs especially for those drugs with high molecular weight.     

Artificial cornea preparation using collagen/chondroitin sulfate/fibroblast growth factor composite film北大核心

BACKGROUND: The traditional corneal scaffolds exhibit poor strength and biological compatibility. Little is reported on the artificial cornea prepared by collagen and chondroitin sulfate (CS), which consist of the natural corneal tissue. OBJECTIVE: To prepare the collagen/CS/fibroblast growth factor (FGF) composite artificial cornea with slow-release growth factor, high strength and light transmittance, as well as good biocompatibility. METHODS: Regenerated collagen films were prepared by 1%, 5%, 10% collagen solutions using flow casting method, and the regenerated collagen film with the best bioactivity that was prepared by 5% collagen solution was screened through a biomechanical test. Then, the CS/collagen composite film was achieved by cross-linking the CS (2, 20, 80 g/L) with collagen by using N-(3-Dimethylaminopropyl)- N’S-ethylcarbodimide hydrochloride-N-Hydroxysuccinimide. The composite film made of 20 g/L CS was confirmed to have the best transparency, which was used to be mixed with 5, 25, 50 mg/L FGF in PBS for 24 hours to prepare the collagen/CS/FGF composite films. ELISA method was used to detect the FGF level in the supernatant. Afterwards, corneal epithelial cells were co-cultured with regenerated collagen film, collagen/CS composite film and collagen/CS/FGF composite film, respectively. After 48 hours of co-culture, cell proliferation was detected by MTT method, based on which we could screen the optimal collagen/CS/FGF composite film. After co-culture with the collagen/CS/FGF composite film for 48 and 72 hours, cell morphology was observed by confocal microscope and scanning electron microscope, respectively. RESULTS AND CONCLUSION:The release amount of FGF from the composite films was dependent on the initial loading amount of FGF. Meanwhile, FGF released slowly from the three kinds of composite films, and the release amount was 11%, 23%, 30% at 72 hours after culture, in accordance with the pharmacokinetic process. MTT findings indicated that the optimal loading concentration of FGF was 25 mg/L. Under the microscope, the collagen/CS/FGF composite film promoted the adhesion, growth and proliferation of corneal epithelial cells. To conclude, the collagen/CS/FGF composite film is expected to be an ideal scaffold material for artificial cornea preparation. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Adsorption and Step Elution of Urokinase Using, Affinity Chromatography -Comparison of Data with Rate Model Simulation

A non-equilibrium chromatographic rate model was employed to simulate the affinity chromatography of urokinase. The chromatography process was developed to a yield of high purity product of urokinase from crude materials. The affinity gel used in the process was prepared by an epichlorohydrin-activation method using epichlorohydrin activated Sepharose 4B as a matrix and p-aminobenzamidine as a ligand. The chromatographic process were numerically simulated and analyzed with the aid of VERSE-LC computer simulator. Considering the basic principles, rate model with the back mixing in column inlet was utilized in simulating and studying the effect of the column inlet pattern on other parameters. Comparison of the simulation results with the experimental data showed that the rate model can be used to describe the affinity chromatography of urokinase in a fixed bed column with satisfactory accuracy.     

Proliferation and differentiation of periosteum cells induced by icariin北大核心

BACKGROUND: Icariin has a broad prospect for promoting cell proliferation. Differentiation direction of periosteal cells is uncertain, but the cells are easy to be induced by ultrasound, oxygen or bone morphogenetic protein 7 (BMP7). Periosteal cells have been applied in bone tissue engineering; however, icariin effects on the proliferation and differentiation of periosteal cells is little reported. OBJECTIVE: To investigate the effect of icariin on the proliferation and differentiation of human periosteal cells, thus providing theoretical basis for icariin applied in bone tissue engineering. METHODS: The human periosteum was obtained and the primary cells were isolated in vitro. After culture and expansion, periosteal cells were cultured in 24-well plates, and induced by 0.001, 0.01 and 0.1 mg/L icariin and 50 µg/L BMP7, respectively. The corresponding avsorbance values of different groups were detected. The levels of alkaline phosphatase and calcium nodules in periosteal cells were measured at 1, 3, 5 and 7 days, and the mRNA levels of osteocalcin, osteopontin and Runx2 were detected at 3, 5 and 7 days. RESULTS AND CONCLUSION: The periosteal cells proliferated well after induction with icariin, and could proliferate well in different concentrations of icariin and the positive control group (P < 0.05). Compared with the control group, the periosteal cells induced by icariin were able to produce more alkaline phosphatase and calcium nodules (P < 0.05). The mRNA expression of osteocalcin, osteopontin and Runx2 in periosteal cells could be up-regulated by icariin (P < 0.05). These findings imply that icariin can promote proliferation and differentiate of periosteal cells into osteoblasts, and it can be used as an inducer for the preparation of seed cells in bone tissue engineering. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.