Glutaraldehyde cross-linked hydroxyapatite/collagen self-organized nanocomposites

To control the mechanical properties and biodegradability of self-organized hydroxyapatite/collagen (HAp/Col) nanocomposites, cross-linkage was introduced into the composites with glutaraldehyde (GA). The HAp/Col composite suspensions, prepared by a simultaneous titration method and aged for 3h, were cross-linked with the reagents for 10min under vigorous stirring. The precipitates obtained were filtrated and compacted by dehydration under a uniaxial pressure. The particle size distribution, 3-point bending strength, contained water amount and swelling ratio of the composites were examined as a function of cross-linkage amount; the biodegradability was estimated by animal tests using rabbits. As regards the cross-linked composites, no long-rage alignment of HAp crystals along collagen molecules was found with a transmission electron microscope, suggesting that the cross-linking reagents suppressed their long-range self-organization mechanism. The 3-point bending strength increased with the GA content and took a maximal value at 1.35mmol/g(col). The animal tests indicated no toxicity and osteoclastic resorption with good osteoconductivity. The resorption rate was decreased with increasing GA concentration. These results suggest that GA cross-linkage controls mechanical properties and resorption rate without reducing high biocompatibility of the composite.     

Stabilized collagen scaffolds for heart valve tissue engineering

Scaffolds for heart valve tissue engineering must function immediately after implantation but also need to tolerate cell infiltration and gradual remodeling. We hypothesized that moderately cross-linked collagen scaffolds would fulfill these requirements. To test our hypothesis, scaffolds prepared from decellularized porcine pericardium were treated with penta-galloyl glucose (PGG), a collagen-binding polyphenol, and tested for biodegradation, biaxial mechanical properties, and in vivo biocompatibility. For controls, we used un-cross-linked scaffolds and glutaraldehyde-treated scaffolds. Results confirmed complete pericardium decellularization and the ability of scaffolds to encourage fibroblast chemotaxis and to aid in creation of anatomically correct valve-shaped constructs. Glutaraldehyde cross-linking fully stabilized collagen but did not allow for tissue remodeling and calcified when implanted subdermally in rats. PGG-treated collagen was initially resistant to collagenase and then degraded gradually, indicating partial stabilization. Moreover, PGG-treated pericardium exhibited excellent biaxial mechanical properties, did not calcify in vivo, and supported infiltration by host fibroblasts and subsequent matrix remodeling. In conclusion, PGG-treated acellular pericardium is a promising scaffold for heart valve tissue engineering.     

Nature-derived epigallocatechin gallate/duck’s feet collagen/hydroxyapatite composite sponges for enhanced bone tissue regeneration

Abstract

Scaffolds mimicking structural and chemical characteristics of the native bone tissues are critical for bone tissue engineering. Herein, we have developed and characterized epigallocatechin gallate/duck’s feet collagen/hydroxyapatite (EGCG/DC/HAp) composite sponges that enhanced the bone tissue regeneration. The three-dimensional composite sponges were synthesized by loading various amounts (i.e. 1, 5 and 10 μM) of EGCG to duck feet derived collagen followed by freeze-drying and then coating with hydroxyapatite. Several measuremental techniques were employed to examine the properties of the as-fabricated composite sponges including morphology and structure, porosity, compressive strength, etc. and as well compared with pristine duck feet derived collagen. SEM observations of EGCG/DC/HAp sponges showed the formation of a highly porous collagen matrix with EGCG embodiment. The porosity and pore size of sponges were found to increase by high EGCG content. The compressive strength was calculated as 3.54 ± 0.04, 3.63 ± 0.03, 3.89 ± 0.05, 4.047 ± 0.05 MPa for 1, 5 and 10 μM EGCG/DC/HAp sponges, respectively. Osteoblast-like cell (BMSCs isolated from rabbit) culture and in vivo experiments with EGCG/DC/HAp sponges implanted in nude mouse followed by histological staining showed enhanced cell internalization and attachment, cell proliferation, alkaline phosphatase expressions, indicating that EGCG/DC/HAp sponges have ahigh biocompatibility. Moreover, highEGCG content in the EGCG/DC/HAp sponges have led to increased cellular behavior. Collectively, the 5 μM of EGCG/DC/HAp sponges were suggested as the potential candidates for bone tissue regeneration.     

京尼平交联3D打印胶原/壳聚糖支架的表征北大核心

背景:胶原/壳聚糖支架需交联才能达到相应力学性能,有研究表示调节交联剂浓度可以在一定范围内调控支架的理化性能。目的:探究京尼平浓度对胶原/壳聚糖支架理化性能的影响,制备理化性能可调节的组织工程支架。方法:将胶原和壳聚糖粉末分别溶于弱酸后混合均匀,作为打印墨水,利用生物3D打印机低温打印胶原支架与胶原/壳聚糖支架,经冻干、中和处理后分别以1,3,5 mmol/L的京尼平进行交联。检测各组支架的表观结构稳定性、抗拉能力、溶胀性能、降解性能与生物相容性。结果与结论:①将支架在PBS中浸泡3 d后,对比未交联的冻干支架,交联后胶原支架表面维持规则的孔结构,但是支架出现明显变形;交联后胶原/壳聚糖支架表面结构规则,仅1 mmol/L京尼平交联的胶原/壳聚糖支架存在轻微变形。②随着京尼平浓度的增加,各组支架的力学性能增加,并且对应交联浓度下的胶原/壳聚糖支架力学性能好于胶原支架。③随着京尼平浓度的增加,胶原支架的溶胀率下降,胶原/壳聚糖支架的溶胀率无明显变化。④浸泡于胶原酶溶液中后,不同浓度京尼平交联的胶原支架在1 h内被完全降解,胶原/壳聚糖支架的降解速率随京尼平浓度的增加而降低,均呈现先快速后平缓的趋势。⑤将骨髓间充质干细胞接种于各组交联支架3 d后,1,3 mmol/L京尼平交联的胶原/壳聚糖支架(或胶原支架)上的细胞数量明显多于5 mmol/L京尼平交联的胶原/壳聚糖支架(P<0.05)。⑥结果表明,京尼平可在一定范围调节胶原/壳聚糖支架理化性能,其中3 mmol/L京尼平交联的胶原/壳聚糖支架具有较好的力学性能、抗酶解能力与生物相容性。     

Comparison of the properties of collagen–chitosan scaffolds after γ-ray irradiation and carbodiimide cross-linking

The property of collagen–chitosan porous scaffold varies according to cross-linking density and scaffold composition. This study was designed to compare the properties of collagen–chitosan porous scaffolds cross-linked with γ-irradiation and carbodiimide (CAR) for the first time. Eleven sets of collagen–chitosan scaffolds containing different concentrations of chitosan at a 5% increasing gradient were fabricated. Fourier transform infrared spectroscopy was performed to confirm the success of cross-linking in the scaffolds. The scaffold morphology was evaluated under scanning electron microscope (SEM). SEM revealed that chitosan was an indispensable material for the fabrication of γ-ray irradiation scaffold. The microstructure of γ-ray irradiation scaffold was less stable than those of alternative scaffolds. Based upon swelling ratio, porosity factor, and collagenase degradation, γ-ray irradiation scaffold was less stable than CAR and 25% proportion of chitosan scaffolds. Mechanical property determines the orientation in γ-irradiation and CAR scaffold. In vitro degradation test indicated that γ-irradiation and CAR cross-linking can elevate the scaffold biocompatibility. Compared with γ-ray irradiation, CAR cross-linked scaffold containing 25% chitosan can more significantly enhance the bio-stability and biocompatibility of collagen–chitosan scaffolds. CAR cross-linked scaffold may be the best choice for future tissue engineering.     

FT-IR study for hydroxyapatite/collagen nanocomposite cross-linked by glutaraldehyde

FT-IR analysis was performed for the hydroxyapatite (HAp)/collagen (COL) nanocomposite cross-linked by glutaraldehyde (GA). The amide bands I, II and III from COL matrix, and phosphate and carbonate bands from HAp were identified. The amide B band arising from C–H stretching mode showed a sensitive conformation by the degree of cross-linking. The amide I band showed a complicate conformational change by the degree of cross-linking. The characteristic amide I band at 1685 cm⁻¹, which is known as an aging parameter in the biological bone, did not show a monotonous tendency by the degree of cross-linking. The relative contents of the organics in the cross-linked HAp/COL nanocomposite were evaluated as an integration ratio between the amide I band at 1600–1700 cm⁻¹ and PO₄³⁻ band at 900–1200 cm⁻¹. The increase of the organics content by the cross-linking is enabled by the further organization of Ca²⁺ ions of HAp crystals in HAp/COL nanocomposite. The complicate conformational behavior in the amide I, II and III bands seems to be affected by the cross-linking induced directional arrangement of HAp/COL nanocomposite fibrils.     

聚乙烯醇-壳聚糖-胶原组织工程支架的研究

目的 以聚乙烯醇（PVA）、壳聚糖（Cs）和胶原（Col）为主要原料制备PVA-Cs—Col复合支架,并研究其作为组织工程支架材料的可行性。方法 把聚乙烯醇、壳聚糖和胶原按一定配比复合,测定复合材料的含水率、膨胀率和力学性能,扫描电镜观察材料横截面的组织形态。结果 不同分子量PVA与不同质量的cs和Col复合,得到的复合支架材料湿态抗张强度为5．70MPa,含水率在60．15％-72．50％,膨胀率在185．33％～317．57％。不同配比的复合支架具有不同的内部组织形态结构。结论 PVA—Cs—Col复合支架材料具有较高的含水率和适宜的膨胀率,内部孔洞丰富,Cs：PVA：Col的质量比为30：15：0．20时,复合支架综合性能较佳,适合用于组织工程支架材料。     

Tissue response of defined collagen-elastin scaffolds in young and adult rats with special attention to calcification

Collagen-elastin scaffolds may be valuable biomaterials for tissue engineering because they combine tensile strength with elasticity. In this study, the tissue response to and the calcification of these scaffolds were evaluated. In particular, the hypothesis was tested that calcification, a common phenomenon in biomaterials, may be due to microfibrils within the elastic fibre, and that these microfibrils might generate a tissue response. Four scaffolds were subcutaneously implanted, viz. collagen, collagen + pure elastin, collagen+microfibril-containing, and collagen + pulverised elastic ligament (the source for elastin). Explants were evaluated at day 3, 7 and 21. In young Sprague Dawley rats, collagen + ligament calcified substantially, whereas collagen + elastin (with and without microfibrils) calcified less, and collagen did not. Calcification started at elastic fibres. In both Sprague Dawley and Wistar adult rats, however, none of the scaffolds calcified. Mononuclear cell infiltration was prominent in young and adult Sprague Dawley rats. In adult Wistar rats, this infiltration was associated with the presence of microfibrils. Degradation of scaffolds and new matrix formation were related with cellular influx and degree of vascularisation. In conclusion, absence of microfibrils from the elastic fibre does not prevent calcification in young Sprague Dawley rats, but does reduce the tissue response in adult Wistar rats. Cellular response and calcification differs with age and strain and therefore the choice of animal model is of key importance in biomaterial evaluation.     

Comparative study of bovine, porcine and avian collagens for the production of a tissue engineered dermis

Combining bovine collagen with chitosan followed by freeze-drying has been shown to produce porous scaffolds suitable for skin and connective tissue engineering applications. In this study collagen extracted from porcine and avian skin was compared with bovine collagen for the production of tissue engineered scaffolds. A similar purity of the collagen extracts was shown by electrophoresis, confirming the reliability of the extraction process. Collagen was solubilized, cross-linked by adding chitosan to the solution and freeze-dried to generate a porous structure suitable for tissue engineering applications. Scaffold porosity and pore morphology were shown to be source dependant, with bovine collagen and avian collagen resulting into the smallest and largest pores, respectively. Scaffolds were seeded with dermal fibroblasts and cultured for 35 days to evaluate the suitability of the different collagen–chitosan scaffolds for long-term tissue engineered dermal substitute maturation in vitro. Cell proliferation and scaffold biocompatibility were found to be similar for all the collagen–chitosan scaffolds, demonstrating their capability to support long-term cell adhesion and growth. The scaffolds contents was assessed by immunohistochemistry and showed increased deposition of extracellular matrix by the cells as a function of time. These results correlate with measurements of the mechanical properties of the scaffolds, since both the ultimate tensile strength and tensile modulus of the cell seeded scaffolds had increased by the end of the culture period. This experiment demonstrates that porcine and avian collagen could be used as an alternative to bovine collagen in the production of collagen–chitosan scaffolding materials.     

Poly(ethylene glycol) hydrogels cross-linked by hydrolyzable polyrotaxane containing hydroxyapatite particles as scaffolds for bone regeneration

Poly(ethylene glycol) (PEG) hydrogels cross-linked by a hydrolyzable polyrotaxane containing hydroxyapatite particles (PRX-HAp) were developed as scaffolds for bone regeneration. Five scaffolds with various composition of the polyrotaxane, PEG and HAp particles were prepared to examine cell adhesion in vitro using rat primary cultured osteoblast. Cells were observed to attach well on a PRX-HAp that have the same weight ratio of the polyrotaxane and HAp particles at 7 days after seeding. These results indicate that HAp particles are necessary for cell adhesion and survival, but a higher ratio of the particles is not suitable for cell adhesion. The composites of rat osteoblast and the PRX-HAp were implanted subcutaneously in syngeneic rats and harvested at 5 weeks after implantation. In histological analysis, osteoblast-like cells became arrayed along the surface of the PRX-HAp, and osteoid-like tissues were observed in the region between a queue of osteoblast-like cells and PRX-HAp. These images are similar to intramembranous ossification, and it is expected that bone regeneration occurs on the surface of the PRX-HAp. This study strongly suggests the great potential of the PRX-HAp as scaffolds for bone regeneration.     

Preparation and evaluation of molecularly-defined collagen-elastin-glycosaminoglycan scaffolds for tissue engineering

Extracellular matrix components are valuable building blocks for the preparation of biomaterials involved in tissue engineering, especially if their biological, chemical and physical characteristics can be controlled. In this study, isolated type I collagen fibrils, elastin fibres and chondroitin sulphate (CS) were used for the preparation of molecularly-defined collagen-elastin-glycosaminoglycan scaffolds. A total of 12 different scaffolds were prepared with four different ratios of collagen and elastin (1:9, 1:1, 9:1 and 1:0), with and without chemical crosslinking, and with and without CS. Collagen was essential to fabricate coherent, porous scaffolds. Electron microscopy showed that collagen and elastin physically interacted with each other and that elastin fibres were enveloped by collagen. By carbodiimide-crosslinking, amine groups were coupled to carboxylic groups and CS could be incorporated. More CS could be bound to collagen scaffolds (10%) than to collagen-elastin scaffolds (2.4-8.5% depending on the ratio). The attachment of CS increased the water-binding capacity to up to 65%. Scaffolds with a higher collagen content had a higher tensile strength whereas addition of elastin increased elasticity. Scaffolds were cytocompatible as was established using human myoblast and fibroblast culture systems. It is concluded that molecularly-defined composite scaffolds can be composed from individual, purified, extracellular matrix components. Data are important in the design and application of tailor-made biomaterials for tissue engineering.     

Poly(ethylene glycol) hydrogels cross-linked by hydrolyzable polyrotaxane containing hydroxyapatite particles as scaffolds for bone regeneration

Poly(ethylene glycol) (PEG) hydrogels cross-linked by a hydrolyzable polyrotaxane containing hydroxyapatite particles (PRX-HAp) were developed as scaffolds for bone regeneration. Five scaffolds with various composition of the polyrotaxane, PEG and HAp particles were prepared to examine cell adhesion in vitro using rat primary cultured osteoblast. Cells were observed to attach well on a PRX-HAp that have the same weight ratio of the polyrotaxane and HAp particles at 7 days after seeding. These results indicate that HAp particles are necessary for cell adhesion and survival, but a higher ratio of the particles is not suitable for cell adhesion. The composites of rat osteoblast and the PRX-HAp were implanted subcutaneously in syngeneic rats and harvested at 5 weeks after implantation. In histological analysis, osteoblast-like cells became arrayed along the surface of the PRX-HAp, and osteoid-like tissues were observed in the region between a queue of osteoblast-like cells and PRX-HAp. These images are similar to intramembranous ossification, and it is expected that bone regeneration occurs on the surface of the PRX-HAp. This study strongly suggests the great potential of the PRX-HAp as scaffolds for bone regeneration.     

Effect of collagen on the mechanical properties of hydroxyapatite coatings

In this study, the mechanical properties of bioactive coatings on Ti6Al4V substrates were investigated using instrumented nanoindentation. The aim was to observe the differences in the mechanical properties before and after immersion in collagen solution. The hydroxyapatite coatings were prepared through two processes: self-assembly in simulated body fluid and a hydrothermal method. Sintered hydroxyapatite disks were used as controls. The test samples were then incubated in a dilute collagen solution for 24 hours to produce composite coatings. The materials were investigated using XRD, SEM and nanoindentation. The results showed that the grain sizes of the hydroxyapatite coatings formed using two processes were 1?μm and 10?μm, respectively. The Young's modulus of the pure hydroxyapatite, the disk and the coatings, was 3.6 GPa. After collagen incubation treatment, the composites had a Young's modulus of 7.5 GPa. The results also showed that the strengthening phenomena of collagen were more obvious for homogeneous and small-grain hydroxyapatite coatings. These results suggest that there are similarities between these HAp/collagen composited and natural composite materials, such as teeth and bones.     

Novel Scaffolds Based on Poly(2-hydroxyethyl methacrylate) Superporous Hydrogels for Bone Tissue Engineering

In this study, poly(2-hydroxyethyl methacrylate) (pHEMA)-based superporous hydrogels were synthesized by radical polymerization of 2-hydroxyethyl methacrylate (HEMA) in the presence of a gas blowing agent, sodium bicarbonate. These hydrogels are: pHEMA, pHEMA–gelatin, glycerol phosphate (GP) cross-linked pHEMA–gelatin, glutaraldehyde (GA) cross-linked pHEMA–gelatin superporous hydrogels (SPHs) and pHEMA–hydroxyapatite (HA) superporous hydrogel composites (SPHCs). The hydrogels have a structure of interconnected pores with pore sizes of approx. 500 μm. Although the extent of swelling decreased when gelatin and HA were incorporated to the pHEMA structure, the time to reach the equilibrium swelling (approx. 20 s) was not affected so much. In the presence of gelatin and cross-linkers, mechanical properties significantly improved when compared with pHEMA SPH. Among all the synthesized hydrogels, pHEMA–HA SPHC showed great improvement in mechanical strength and its elastic modulus value was 0.027±0.002 N/mm². Osteogenic activities of pHEMA-based scaffolds were examined by preosteoblastic MC3T3-E1 cell-culture studies. The mitochondrial activity test (MTT) showed that gelatin-containing scaffolds stimulated cell proliferation compared with other scaffolds, while alkaline phosphatase levels (ALP) and mineralization were found highest for the GP cross-linked pHEMA–gelatin SPH. However, pHEMA SPH and pHEMA–HA SPHC did not support cell proliferation and also differentiation. In conclusion, pHEMA–gelatin SPH and GP cross-linked pHEMA–gelatin SPH can be considered as potential scaffolds for bone tissue-engineering applications.     

Mechanical properties of cross-linked collagen meshes after human adipose derived stromal cells seeding

The main goal of this study was to evaluate the potential of collagen meshes derived from porcine dermis as scaffolds for repairing pelvic organ prolapses. Mechanical properties of collagen meshes with different cross-linking percentages before and after Adipose Derived Stromal Cells (ADSC) seeding were studied as well as the cell-scaffold interaction. Uniaxial tensile tests of the collagen meshes with three different cross-linking percentages (full-, partial-, and noncross-linked) were carried out along orthogonal directions. Their mechanical properties were studied with the same tests before and after seeding with human derived adipose stem cells (ADSC) after 1 and 7 days. Histological analyses were performed to determine adhesion and proliferation of ADSC. Significant differences in mechanical properties of the unseeded meshes were observed between each orthogonal direction independently of the cross-linking percentage. A better cell adhesion rate was observed in the cross-linked meshes. An increase in the mechanical properties after cell seeding was observed with a direct relation with the degree of cross-linking. All meshes analyzed showed a marked anisotropy that should be taken into account during the surgical procedure. The cross-linking treatment increased cell adhesion and the mechanical properties of the collagen meshes after seeding. These results suggest that the mechanical properties of this type of collagen mesh could be useful as scaffolds for repair of pelvic organ prolapse.     

Photochemical cross-linking for collagen-based scaffolds: a study on optical properties, mechanical properties, stability, and hematocompatibility

Collagen presents an attractive biomaterial for tissue engineering because of its excellent biocompatibility and negligible immunogenicity. However, some intrinsic features related to the mechanical stability and thrombogenicity limit its applications in orthopedic and vascular tissue engineering. Photochemical cross-linking is an emerging technique able to stabilize tissue grafts and improve the physicochemical properties of collagen-based structures. However, other important properties of collagen-based structures and the effect of processing parameters on these properties have not been explored. In this study, we aim to investigate the dose dependence of tensile and swelling properties on two parameters, namely, laser energy fluence and rose Bengal photosensitizer concentration. We also study the compression properties using cyclic compression test, long-term stability using subcutaneous implantation, and hematocompatibility using platelets adhesion test, of cross-linked collagen structures. Moreover, because limited optical penetration in turbid media is the major obstacle for light-based techniques, we also characterize the optical properties, which partially determine the effective optical penetration depth in collagen gel samples, during photochemical cross-linking. Laser energy fluence and rose Bengal concentration are important parameters affecting the cross-linking efficiency, which was characterized as the mechanical and the swelling properties, in a dose-dependent manner. Under the experimental conditions in this study, the peak fluence was 12.5 J/cm2 and the minimal rose Bengal concentration for effective cross-linking was >0.00008% (0.786 micromol). Photochemical cross-linking also enhanced the compression strength and long-term stability of collagen structures without compromising the tissue compatibility. Furthermore, photochemical cross-linking reduced platelet adhesion and abolished fibrin mesh formation, thereby improving the hematocompatibility of collagen structures. These results suggest the feasibility of using the photochemically cross-linked collagen structures for orthopedic and vascular tissue engineering. Finally, the effective optical penetration depth in collagen gel samples is wavelength and rose Bengal concentration dependent, and was approximately 12 mm at 514 nm at 0.001% (9.825 micromol), the rose Bengal concentration mostly used in this study.     

Pectin-chitosan-PVA nanofibrous scaffold made by electrospinning and its potential use as a skin tissue scaffold

Scaffolds made of chitosan nanofibers are often too mechanically weak for their application and often their manufacturing processes involve the use of harmful and flammable organic solvents. In the attempt to improve the mechanical properties of nanofibrous scaffolds made of chitosan without the use of harmful chemicals, pectin, an anionic polymer was blended with chitosan, a cationic polymer, to form a polyelectrolyte complex and electrospun into nanofibers for the first time. The electrospun chitosan-pectin scaffolds, when compared to electrospun chitosan scaffolds, had a 58% larger diameter, a 21% higher Young’s modulus, a 162% larger strain at break, and a 104% higher ultimate tensile strength. Compared to the chitosan scaffolds, the chitosan-pectin scaffolds’ swelling ratios decreased by 55% after 60?min in a saline solution and more quickly released the preloaded tetracycline HCl. The L929 fibroblast cells proliferated slightly slower on the chitosan-pectin scaffolds than on the chitosan scaffolds. Nonetheless, cells on both materials deposited similar levels of extracellular type I collagen on a per DNA basis. In conclusion, a novel chitosan-pectin nanofibrous scaffold with superior mechanical properties than a chitosan nanofibrous scaffold was successfully made without the use of harmful solvents.     

Genipin-Cross-linked Electrospun Collagen Fibers

The fabrication of a fibrous collagen scaffold using electrospinning is desirable for tissue-engineering applications. Previously, electrospun collagen fibers were shown to be unstable in aqueous environments and, therefore, cross-linking is essential to stabilize these fibers. In this study genipin, a significantly less cytotoxic cross-linking agent compared to glutaraldehyde, was used to cross-link electrospun collagen fibers. The significance of this research lies in the use of four alcohol/water solvent systems to carry out the crosslinking reaction to maintain fibrous morphology during cross-linking. The four cross-linking conditions established were: (1) ethanol, 5% water and 3 days, (2) ethanol, 3% water and 5 days, (3) ethanol, 5% water and 5 days, and (4) isopropanol, 5% water and 5 days at a genipin concentration of 0.03 M. Results illustrated that genipin-cross-linking was effective in maintaining collagen fiber integrity in aqueous and cell culture media environments for up to 7 days. In addition, it was shown that fiber swelling could be controlled by using different cross-linking conditions. Swelling of cross-linked fibers immersed in Dulbecco's modified eagle medium for 7 days ranged from 0 to 59 ± 4%. The cross-linked fibers were analyzed using scanning electron microscopy, Fourier transform infrared spectroscopy and ninhydrin assay. Finally, studies using primary human fibroblasts indicated good cell adhesion to these scaffolds. Overall, our data suggest that these stabilized fibrous collagen scaffolds provide a promising environment for tissue-regeneration applications.     

Study on the physical properties of tissue-engineered blood vessels made by chemical cross-linking and polymer-tissue cross-linking

In this study, we attempted to chemically cross-link decellularized blood vessel tissue and to perform cross-linking with a polymer in order to control its stability and functionalization. For this purpose, we cross-linked tissue by intrahelical, interhelical, and intermolecular cross-linking between the polymer and the collagen helix, which is a component of the native tissue. The intrahelically cross-linked tissue showed weaker stability against heat and degradation caused by collagenase compared to the interhelically cross-linked tissue. The tissue intermolecularly cross-linked with polymer showed the highest stability against heat and degradation caused by collagenase. The mechanical strength test showed that the Young’s moduli were different for the intra/interhelically and intermolecularly cross-linked tissues, with the latter being stiffer. This is thought to be because the cross-linked polymer functions in the same way as elastin, whereas simple collagen cross-linking provides a supportive matrix that holds the collagen and elastin together.     

应用不同物理加强条件的Ⅰ型胶原支架的大白鼠动物模型与组织学分析

BACKGROUND: In previous studies, the dehydrothermal cross-linking method was modified by the authors to improve the degradation property of collagen scaffolds. The cross-linking time was increased from 24 to 48 hours, and the cross-linking temperature increased from 105 to 115 ℃. OBJECTIVE: To verify the anti-degradation ability of collagen scaffolds prepared using the modified dehydrothermal cross-linking method and to obtain the optimal efficacy of the scaffolds on damaged tissue repair and regeneration. METHODS: Highly-purified type I collagen scaffolds with native triple helix structure were prepared and subjected to three different dehydrothermal cross-linking conditions: 105 ℃ for 24 hours, 105 ℃ for 48 hours and 115 ℃ for 24 hours. Material samples, 1 cm×1 cm, were implanted subcutaneously into the rat dorsum. The specimens were harvested at 3 days, 14 days and 42 postoperative days followed by fixation and histological analysis using hematoxylin-eosin staining. RESULTS AND CONCLUSION: No untoward foreign body and immunological reactions were observed in any groups. In the group of 105 ℃ for 48 hours, the scaffold retention and degree of pore openness were better than the other two groups at 14 days after scaffold implantation (P < 0.05). These findings indirectly suggest that the anti-degradation ability of collagen scaffolds can be strengthened under certain dehydrothermal cross-linking conditions: the cross-linking time is increased from 24 to 48 hours.