Novel chitosan hydrogel formed by ethylene glycol chitosan, 1,6-diisocyanatohexan and polyethylene glycol-400 for tissue engineering scaffold: in vitro and in vivo evaluation.

Traditional chitosan hydrogels were prepared by chemical or physical crosslinker, and both of the two kinds of hydrogels have their merits and demerits. In this study, researchers attempted to prepare one kind of chitosan hydrogel by slightly crosslinker, which could combine the advantages of the two kinds of hydrogels. In this experiment, the crosslinker was formed by a reaction between the isocyanate group of 1,6-diisocyanatohexan and the hydroxyl group of polyethylene glycol-400 (PEG-400), then the crosslinker reacted with the amidine and the hydroxyl group of ethylene glycol chitosan to form the network structure. Physical properties of the hydrogel were tested by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and biodegradation. Biocompatibility was assessed by cell implantation in vitro and the scaffold was used as a cartilage tissue engineering scaffold to repair a defect in rabbit knee joints in vivo. FTIR results show the formation of a covalent bond during thickening of the ethylene glycol chitosan. SEM and degradation experiments showed that the ethylene glycol chitosan hydrogel is a 3-D, porous, and degradable scaffold. The hydrogel contained 2% ethylene glycol chitosan and 10 µl crosslinker was selected for the biocompatibility experiment in vitro and in vivo. After chondrocytes were cultured in the ethylene glycol chitosan hydrogel scaffold for 1 week cells exhibited clustered growth and had generated extracellular matrix on the scaffold in vitro. The results in vivo showed that hydrogel-chondrocytes promoted the repair of defect in rabbits. Based on these results, it could be concluded that ethylene glycol chitosan hydrogel is a scaffold with excellent physicochemical properties and it is a promising tissue engineering scaffold.

Cascade-Catalyzed Nanogel for Amplifying Starvation Therapy by Nitric Oxide-Mediated Hypoxia Alleviation.

Glucose oxidase (Gox)-mediated starvation therapy offers a prospective advantage for malignancy treatment by interrupting the glucose supply to neoplastic cells. However, the negative charge of the Gox surface hinders its enrichment in tumor tissues. Furthermore, Gox-mediated starvation therapy infiltrates large amounts of hydrogen peroxide (H2O2) to surround normal tissues and exacerbate intracellular hypoxia. In this study, a cascade-catalyzed nanogel (A-NE) was developed to boost the antitumor effects of starvation therapy by glucose consumption and cascade reactive release of nitric oxide (NO) to relieve hypoxia. First, the surface cross-linking structure of A-NE can serve as a bioimmobilization for Gox, ensuring Gox stability while improving the encapsulation efficiency. Then, Gox-mediated starvation therapy efficiently inhibited the proliferation of tumor cells while generating large amounts of H2O2. In addition, covalent l-arginine (l-Arg) in A-NE consumed H2O2 derived from glucose decomposition to generate NO, which augmented starvation therapy on metastatic tumors by alleviating tumor hypoxia. Eventually, both in vivo and in vitro studies indicated that nanogels remarkably inhibited in situ tumor growth and hindered metastatic tumor recurrence, offering an alternative possibility for clinical intervention.

Magnetic resonance imaging using a nonuniform Bo (NuBo) field-cycling magnet.

Magnetic resonance imaging (MRI) is a powerful noninvasive diagnostic tool with superior soft tissue contrast. However, access to MRI is limited since current systems depend on homogeneous, high field strength main magnets (B0-fields), with strong switchable gradients which are expensive to install and maintain. In this work we propose a new approach to MRI where imaging is performed in an inhomogeneous field using radiofrequency spatial encoding, thereby eliminating the need for uniform B0-fields and conventional cylindrical gradient coils. The proposed technology uses an innovative data acquisition and reconstruction approach by integrating developments in field cycling, parallel imaging and non-Fourier based algebraic reconstruction. The scanner uses field cycling to image in an inhomogeneous B0-field; in this way magnetization is maximized during the high field polarization phase, and B0 inhomogeneity effects are minimized by using a low field during image acquisition. In addition to presenting the concept, this work provides experimental verification of a long-lived spin echo signal, spatially varying resolution, as well as both simulated and experimental 2D images. Our initial design creates an open MR system that can be installed in a patient examination table for body imaging (e.g., breast or liver) or built into a wall for weighted-spine imaging. The proposed system introduces a new class of inexpensive, open, silent MRIs that could be housed in doctor's offices much like ultrasound is today, making MRI more widely accessible.

Two-in-One Polymeric NO-Donor Prodrugs Mediate Precision and Synergistic Prostate Cancer Treatment.

Chemotherapy predominates in clinical treatment of prostate cancer (PCa), while irreversible resistance to chemotherapeutics and severe side effects hinder the therapeutic efficacy, especially in castration-resistant PCa (CRPC). Herein, a bombesin (BBN)-decorated two-in-one prodrug (T-NO/E2-PMs) incorporating a polymeric nitric oxide (NO) donor and acetal-linked 17ß-estradiol (E2) in one backbone is developed, aiming to inhibit androgen receptor (AR) expression, reprogram the tumor microenvironment of CRPC, and enhance estradiol-mediated hypoxic CRPC therapy. Following efficient internalization mediated by BBN, T-NO/E2-PMs releases estradiol and NO in response to the unique intracellular environments. Both in vitro and in vivo studies demonstrate that the T-NO/E2-PMs nano-prodrug along with NO release potently downregulates AR levels to reverse CRPC and further enhances the chemo-sensitization of estradiol to PCa PC-3 cell apoptosis and the inhibition of metastasis. Collectively, this two-in-one nano-prodrug strategy offers a promising platform for construction of advanced nanomedicine to boost the therapeutic efficacy.

Application of hyperspectral remote sensing for supplementary investigation of polymetallic deposits in Huaniushan ore region, northwestern China.

A gold-silver-lead-zinc polymetallic ore was selected in Huaniushan, Gansu Province as the study area. Hyperspectral aerial images as the primary information source, ground spectrum tests, and sampling analysis were used as auxiliary techniques. They were combined with large-scale mineral and geological maps and other high-resolution satellite remote sensing images. Hyperspectral remote sensing classification identification and quantitative analysis methods were used to study the main mineral resources and rock mass occurrence. Finally, deposit distribution information was extracted and validated. The results showed that the effective classification methods by hyperspectral images were spectral angle mapping, minimum noise fraction transform, and mixed tuned matched filtering. Based on the ground survey, combined with sampling analysis, the accuracy of classification was 80%. The recognition rate of the main ore body-the iron-manganese cap lead-zinc oxide ore-was as high as 81%. This research showed that hyperspectral remote sensing in this mining area has excellent demonstration effects and is worth completing and supplementing original mineral and geological maps. The targets are important areas for detailed follow-up on mineral resource exploration.

Adenovirus-mediated expression of growth and differentiation factor-5 promotes chondrogenesis of adipose stem cells.

The repair of articular cartilage injuries is impeded by the avascular and non-innervated nature of cartilage. Transplantation of autologous chondrocytes has a limited ability to augment the repair process due to the highly differentiated state of chondrocytes and the risks of donor-site morbidity. Mesenchymal stem cells can undergo chondrogenesis in the presence of growth factors for cartilage defect repair. Growth and differentiation factor-5 (GDF5) plays an important role in chondrogenesis. In this study, we examined the effects of GDF5 on chondrogenesis of adipose-derived stem cells (ADSCs) and evaluate the chondrogenic potentials of GDF5 genetically engineered ADSCs using an in vitro pellet culture model. Rat ADSCs were grown as pellet cultures and treated with chondrogenic media (CM). Induction of GDF5 by an adenovirus (Ad-GDF5) was compared with exogenous supplementation of GDF5 (100 ng/ml) and transforming growth factor-beta (TGF-beta1; 10 ng/ml). The ADSCs underwent chondrogenic differentiation in response to GDF5 exposure as demonstrated by production of proteoglycan, and up-regulation of collagen II and aggrecan at the protein and mRNA level. The chondrogenic potential of a one-time infection with Ad-GDF5 was weaker than exogenous GDF5, but equal to that of TGF-beta1. Stimulation with growth factors or CM alone induced transient expression of the mRNA for collagen X, indicating a need for optimization of the CM. Our findings indicate that GDF5 is a potent inducer of chondrogenesis in ADSCs, and that ADSCs genetically engineered to express prochondrogenic growth factors, such as GDF5, may be a promising therapeutic cell source for cartilage tissue engineering.

Biphasic scaffold for annulus fibrosus tissue regeneration.

Intervertebral disc (IVD) degeneration is the major cause of lower back pain, while the currently available treatments are symptomatic rather than curative. Tissue engineering is a powerful therapeutic strategy that can restore the normal biomechanical motion of the human spine. The ability of a biphasic elastic scaffold to structurally and elastically simulate the annulus fibrosus (AF) tissue of the IVD was explored. The outer phase of the scaffold was a ring-shaped demineralized bone matrix gelatin (BMG) extracted from cortical bone, which mimicks the type I collagen structure and ligamentous properties of outer AF. The inner phase of the scaffold was a bio-biomaterial poly(polycaprolactone triol malate) (PPCLM) orientated in concentric sheets and seeded with chondrocytes to recapitulate the inner layer of the AF, which is rich in type II collagen and proteoglycan. The mechanical properties and degradation of PPCLM could be adjusted by controlling the post-polymerization time of the pre-polymer. PPCLM also demonstrated good biocompatibility in a foreign body response in vivo assay. Incorporation of BMG into the scaffold enhanced the compressive strength compared with PPCLM alone. In addition, the tensile stress of the BMG/PPCLM scaffold was 50-fold greater than that of PPCLM alone, and close to that of normal rabbit AF. Finally, the biphasic scaffold supported the growth of rabbit chondrocytes, as confirmed by Safranin-O and type II collagen immunostaining. The excellent mechanical properties and biocompatibility of the BMG/PPCLM scaffold make it a promising candidate for AF repair.

Mouse growth and differentiation factor-5 protein and DNA therapy potentiates intervertebral disc cell aggregation and chondrogenic gene expression.

BACKGROUND CONTEXT: Growth and differentiation factor-5 (GDF-5)-deficient mice showed abnormalities in intervertebral disc (IVD) structure and extracellular matrix. Adenovirus-mediated GDF-5 delivery can promote the growth of rabbit disc cells. PURPOSE: The aim of the present study was to investigate the effect of recombinant GDF-5 protein and GDF-5 complementary DNA (cDNA) on the metabolism of IVD cells. STUDY DESIGN: The effects of recombinant GDF-5 protein and GDF-5 cDNA on mouse IVD cells will be evaluated in vitro. METHODS: Mouse disc cells in vitro were treated with recombinant GDF-5 protein. Mouse GDF-5 cDNA was cloned into an expression vector and was used to transfect mouse disc cells in vitro. Therapy with GDF-5 protein and cDNA was assessed by measuring cell proliferation, proteoglycan production, and extracellular matrix gene expression. RESULTS: Biochemical assays revealed an elevated sulfated glycosaminoglycan (GAG)/DNA ratio in mouse IVD cells that were cultured in the presence of various concentrations of mouse GDF-5(mGDF-5) protein. Real-time reverse transcription-polymerase chain reaction (RT-PCR) demonstrated that treating the cells with GDF-5 protein increased the expression of the collagen Type II and aggrecan genes in a dose-dependent manner but decreased matrix metalloproteinase (MMP)-3 gene expression. Immunohistochemistry showed an increase in the aggregation of mouse IVD cells that were treated with mGDF-5 in culture compared with the control group. The mouse GDF-5 gene was successfully cloned into an expression plasmid vector, and GDF-5 protein production was confirmed by Western blot analysis. Type II collagen and aggrecan gene expression by the cells increased significantly in the cells that were transfected by nucleofection with the GDF-5 plasmid compared with cells that were transfected with a control plasmid. CONCLUSIONS: This is the first report of the cloning of the mouse GDF-5 gene and use of the nucleofection method to transfer DNA into IVD cells. The data suggest that both recombinant protein and the cDNA forms of GDF-5 can increase the expression of genes for extracellular matrix proteins in mouse IVD cells. Future attempts at gene therapy to treat degenerative disc disease with a novel ex vivo gene transfer technique are needed to develop a therapy that would alleviate the condition of patients with clinically relevant axial spine pain.

Novel biodegradable poly(1,8-octanediol malate) for annulus fibrosus regeneration.

A novel elastic scaffold that simulates the deformability of annulus fibrosus (AF) and has good biocompatibility was developed. The scaffold was formed of a malic acid-based polyester poly(1,8-octanediol malate) (POM), which was synthesized by direct polycondensation. The tensile strength of POM gradually increased with the extension of the polymerization time, while the degradation rate decreased. Rat AF cells proliferated on the POM films and maintained their phenotype. The 3D scaffold also supported the growth of the AF cells, as confirmed by Safranin-O and type II collagen staining. POM also demonstrated a good biocompatibility in an in vivo foreign body response assay, an important prerequisite for tissue engineering applications. This study suggests that elastic POM scaffold may be an ideal candidate for AF tissue engineering.

Porcine-derived xenogeneic bone/poly(glycolide-co-lactide-co-caprolactone) composite and its affinity with rat OCT-1 osteoblast-like cells.

Porcine-derived xenogeneic bone (PDXB) was derived from cancellous bone of adult porcine. Its morphology and structure were characterized by SEM, FTIR and XRD. A series of composite films consisting of PDXB and poly(glycolide-co-lactide-co-caprolactone) (PGLC) polymer were prepared. Because of the introduction of PGLC polymer, the PDXB/PGLC composites especially PDXB/PGLC(30/70) and PDXB/PGLC(50/50) showed good processability and mechanical properties. In addition, the hydrophilicity of the composites was enhanced as well since the PDXB component was hydrophilic. Osteoblast-like cells (OCT-1) were used as an in-vitro model to assess the affinity of the PDXB/PGLC composites. It was found that compared with the pure PGLC film, PDXB/PGLC(30/70) and PDXB/PGLC(50/50) composite films promoted cell attachment, proliferation and ALP (alkaline phosphatase) activity obviously. In addition, the cells preferred growing on the areas of exposed PDXB. It was considered that the hydrophilicity, osteoconductivity and appropriate surface roughness (Sa=3.30, 4.00 microm) induced by PDXB facilitate cell growth. However, the introduction of too much PDXB, such as PDXB/PGLC(70/30) film, would obtain an adverse effect on the cell growth since the value of Sa was up to 7.33 microm. It indicated that only the composites with appropriate surface topography could favor cell growth. Surface topography probably has a more important effect on cell growth process than surface chemistry.

Influences of ammonia plasma treatment on modifying depth and degradation of poly(L-lactide) scaffolds.

Hydrophobicity of poly(L-lactide) scaffolds is a main drawback in obtaining a sufficient mass of seeded cells for satisfying the requirements of tissue engineering. Plasma treatment is a useful technique to enhance the hydrophilicity of the scaffolds. However, the effect of this technique on the modifying depth and degradation of the scaffolds should be considered. In this paper, the influence of NH3 plasma treatment on the modifying depth and degradation of scaffolds were investigated. The results showed that the modifying depth of the scaffolds increased with treating time and the plasma power ranging from 20 to 80 W influenced the depth slightly. However, the degradation of the scaffolds increased with increasing treatment time and plasma power. The results also showed that the plasma intruded the scaffolds gradually from top to bottom. For a 4 mm thick scaffold, the optimized treatment condition was 20 W of power in a 30 Pa ammonia atmosphere for 30 min of treating time. Under this condition, the integrity of scaffold could be relatively well kept. NH3 plasma treatment enabled the penetration of cells into scaffolds and facilitated the proliferation of cells in them.

Adhesion and proliferation of OCT-1 osteoblast-like cells on micro- and nano-scale topography structured poly(L-lactide).

The impact of the surface topography of polylactone-type polymer on cell adhesion was to be concerned because the micro-scale texture of a surface can provide a significant effect on the adhesion behavior of cells on the surface. Especially for the application of tissue engineering scaffold, the pore size could have an influence on cell in-growth and subsequent proliferation. Micro-fabrication technology was used to generate specific topography to investigate the relationship between the cells and surface. In this study the pits-patterned surfaces of polystyrene (PS) film with diameters 2.2 and 0.45 microm were prepared by phase-separation, and the corresponding scale islands-patterned PLLA surface was prepared by a molding technique using the pits-patterned PS as a template. The adhesion and proliferation behavior of OCT-1 osteoblast-like cells morphology on the pits- and islands-patterned surface were characterized by SEM observation, cell attachment efficiency measurement and MTT assay. The results showed that the cell adhesion could be enhanced on PLLA and PS surface with nano-scale and micro-scale roughness compared to the smooth surfaces of the PLLA and PS. The OCT-1 osteoblast-like cells could grow along the surface with two different size islands of PLLA and grow inside the micro-scale pits of the PS. However, the proliferation of cells on the micro- and nano-scale patterned surface has not been enhanced compared with the controlled smooth surface.

Repair of articular cartilage defects in rabbits through tissue-engineered cartilage constructed with chitosan hydrogel and chondrocytes.

OBJECTIVE: In our previous work, we prepared a type of chitosan hydrogel with excellent biocompatibility. In this study, tissue-engineered cartilage constructed with this chitosan hydrogel and costal chondrocytes was used to repair the articular cartilage defects. METHODS: Chitosan hydrogels were prepared with a crosslinker formed by combining 1,6-diisocyanatohexane and polyethylene glycol. Chitosan hydrogel scaffold was seeded with rabbit chondrocytes that had been cultured for one week in vitro to form the preliminary tissue-engineered cartilage. This preliminary tissue-engineered cartilage was then transplanted into the defective rabbit articular cartilage. There were three treatment groups: the experimental group received preliminary tissue-engineered cartilage; the blank group received pure chitosan hydrogels; and, the control group had received no implantation. The knee joints were harvested at predetermined time. The repaired cartilage was analyzed through gross morphology, histologically and immunohistochemically. The repairs were scored according to the international cartilage repair society (ICRS) standard. RESULTS: The gross morphology results suggested that the defects were repaired completely in the experimental group after twelve weeks. The regenerated tissue connected closely with subchondral bone and the boundary with normal tissue was fuzzy. The cartilage lacuna in the regenerated tissue was similar to normal cartilage lacuna. The results of ICRS gross and histological grading showed that there were significant differences among the three groups (P<0.05). CONCLUSIONS: Chondrocytes implanted in the scaffold can adhere, proliferate, and secrete extracellular matrix. The novel tissue-engineered cartilage constructed in our research can completely repair the structure of damaged articular cartilage.

Synthesis and cell affinity of functionalized poly(L-lactide-co-beta-malic acid) with high molecular weight.

A novel functionalized biodegradable poly(L-lactide-co-beta-benzyl malolactonate) (p-PLMA) with high molecular weight was synthesized through ring-opening copolymerization. Three p-PLMA copolymers with different beta-benzyl malolactonate content were synthesized. The molecular weight (M(w)) and tensile strength of the copolymer with 4 mol% beta-benzyl malolactonate content were 179,800 and 19.0MPa respectively, the molecular weight (M(w)) and tensile strength of p-PLMA decreased with beta-benzyl malolactonate content increasing. The hydrophilicity of the de-protected product: poly(L-lactide-co-beta-malic acid) (d-PLMA) increased with malic acid content increasing. The results of 3T3 mice fibroblasts cultivated on d-PLMA films showed that the cell adhesion on d-PLMA was better than that of PLLA and the cell attached efficiency of d-PLMA with 8 mol% malic acid content was the highest. The cells grew well both on the surface and inside of d-PLMA scaffolds. The cell affinity of d-PLMA was better than that of PLLA.

Synthesis and characterization of biodegradable polylactide-grafted dextran and its application as compatilizer.

Brush-like biodegradable polylactide-grafted dextran copolymer (PLA-g-dextran) was by a bulk polymerization reaction using a trimethylsilyl-protected (TMS) dextran as macroinitiator and stannous octoate as catalyst. After the polymerization, the TMS groups could be easily removed by immersing the copolymer in methanol for 48 h. The PLA-g-dextran copolymers were characterized by (1)H NMR, GPC and intrinsic viscosity measurements. Besides, mouse 3T3 fibroblasts were cultured on these copolymeric substrates together with pure polylactide (PLA). Although the copolymers exhibited better hydrophilicity and cell affinity compared to pure PLA because of the incorporation of glucose units and the brush-like architecture, it was found that the cells still could not migrate into the center part of scaffold made of PLA-g-dextran copolymer. In result, PLA-g-dextran copolymers themselves were not an appropriate choice for the cell scaffold material, however, it could be used as compatilizer to ameliorate the compatibility between hydrophilic dextran and hydrophobic PLA due to its amphiphilic structure, which could improve the mechanical properties of PLA/dextran blends by reducing the phase separation between PLA and dextran. Therefore, the PLA/dextran blends, which had good cell affinity and moderate mechanical strength, might be prospect cell scaffold materials.

Biodegradable poly(L-lactide)-poly(ethylene glycol) multiblock copolymer: synthesis and evaluation of cell affinity.

A series of poly(L-lactide)-poly(ethylene glycol) multiblock copolymers (Multi-PLE) with high molecular weight were synthesized and successfully used to fabricate three-dimensional scaffolds. Using mouse NIH 3T3 fibroblasts as model cells, the cell affinity of various Multi-PLE copolymers was evaluated and compared with that of poly(L-lactide) (PLLA) by means of cell attachment efficiency measurement, scanning electron microscopy observation and MTT assay. On one hand, the results showed that the cell attachment efficiency on Multi-PLE 4/1(4/1 refers to the molar ratio of lactidyl units to ethylene oxide units) films was close to that on PLLA film, however, the other Multi-PLE films exhibited much lower cell attachment efficiency than PLLA film, such as Multi-PLE 2/1 and Multi-PLE 1/1, which had higher PEG content. On the other hand, it was interesting to find that cell proliferation on Multi-PLE4/1 and Multi-PLE2/1 scaffolds was better than that on PLLA scaffold, which was closely related to the improved hydrophilicity of Multi-PLE copolymers due to the incorporation of PEG in comparison with pure PLLA. The Multi-PLE copolymer scaffolds with appropriate hydrophilicity were in favor of mass transportation, and then of cell proliferation and cell affinity. It meant that the cell proliferation would be much improved by increasing the hydrophilicity of the three-dimensional scaffolds, which even outweighed the disadvantages of the cell attachment efficiency reduction with the incorporation of PEG.

Cell adhesion on gaseous plasma modified poly-(L-lactide) surface under shear stress field.

A series of gases were used for plasma treatment of poly-(L-lactide) (PLLA) under various conditions such as atmosphere, electric power, pressure and time. The NH(3) was preferably selected for modifying the surface of PLLA because it can obtain appropriate hydrophilicity and surface energy with high polar component compared to other gases. Subsequently, cells were seeded onto NH(3) modified surface and exposed to 29.5N/m(2) of shear stress field by means of a parallel plate flow chamber in order to get good insight into the influence of N-containing incorporation on cell retention, cell morphology, and cell shape factor. The results showed that cell retention on the modified PLLA was much higher than that on the unmodified one. The NH(3) plasma modified PLLA with high cell affinity and resistance to shear stress was gained. Surface hydrophilicity, surface energy with high polar component and N-containing groups may play an important role in enhancing cell resistance to shear stress. It revealed that the parallel plate flow chamber is an effective device for evaluating the effects of surface modification on the cell affinity of a material.

Characterization of surface property of poly(lactide-co-glycolide) after oxygen plasma treatment.

In this study, poly (lactide-co-glycolide) (PLGA) films were treated by oxygen plasma. The surface structure, topography and surface chemistry of treated PLGA films were characterized by contact angle measurement, scanning electron microscope observation, atomic force microscopy, and X-ray photoelectron spectrum analysis. The cell affinity of the oxygen plasma treated films was evaluated under dynamic conditions by Parallel Plate Flow Chamber (PPFC). The results showed that the hydrophilicity increased greatly after oxygen plasma treatment. High quantities of -C-O groups, such as hydroxyl and peroxyl groups could be incorporated into the surface of PLGA (70/30) by controlling appropriate plasma treatment conditions. Moreover, the oxygen plasma treatment resulted in formation of peaks and valleys on the sample surfaces, and the roughness increased with treatment time. Cells stretched very well and the ability to endure the shear stress was improved greatly after the PLGA (70/30) was modified by appropriate plasma treatment, i.e. under 50W for 2 or 10 min. However, when the treatment time was increased to 20 min, the percentage of adherent cells on the roughest surface decreased because the content of polar groups incorporated onto the surface decreased. The results showed that improved cell adhesion was attributed to the combination of surface chemistry and surface morphology of PLGA during plasma etching.

Plasma-treated, collagen-anchored polylactone: Its cell affinity evaluation under shear or shear-free conditions.

Poly(L-lactic acid)(PLLA) and poly(L-lactic-co-glycolic acid) (PLGA) (85/15) were modified by plasma treatment. Then they were collagen anchored (PT/CA), and the cell affinity was evaluated by cell culture under shear or shear-free conditions. A convenient and "intuitionistic" dyeing method has been proposed for measuring the modified depth when plasma treatment is applied for the treatment of porous scaffolds. A parallel plate flow chamber was developed in order to study the cell affinity of a material under shear stress. Our results show that a porous scaffold can be modified by plasma treatment and that a depth of about 4.0 mm for this modification can be reached with NH(3) plasma treatment (50 w, 20 Pa, 5 min). PT/CA modification is an effective surface modification method for facilitating cell transplantation and improving the cell affinity of three-dimensional porous cell scaffolds in tissue engineering. It can solve the problem of non-uniform cell distribution in most synthetic porous cell scaffolds. Using the flow chamber system, a series of quantitative data, including cell adherent fraction, cell area, and mean shape, were compared to evaluate the cell affinity of PLLA before and after PT/CA modification. The results indicate that the quality of cell attachment on PT/CA-modified PLLA apparently is better than that on unmodified PLLA. The flow chamber system potentially may be a tool for evaluating surface modification methods.

Acceleration effect of basic fibroblast growth factor on the regeneration of peripheral nerve through a 15-mm gap.

In this study, nerve guides composed of poly(D,L-lactide) (PDLLA) were fabricated and used in the repair of transected sciatic nerves (15-mm gaps) of rats. Nerve guides with a two-ply structure (inner layer dense, outer layer microporous) were prepared by controlling the solvent evaporation rate. Then basic fibroblast growth factor (bFGF) was embedded in the inner layer of the nerve guides. Thus the inner dense layer not only could prevent the ingrowth of fibroblast and avoid the outgrowing nerve cable, but it also could retain the released bFGF in the guide lumen. The outer porous layer allowed vascular ingrowth and the diffusion of essential nutrients into the guide lumen. The data show that by using this nerve guide, the transected 15-mm sciatic nerve was regenerated successfully within 4 months. The recovery of function of the regenerated nerves was significantly accelerated by bFGF, as indicated by an electrostimulation test and histologic assays. In addition, the bFGF retained its bioactivity during embedding and continuously was released from the matrix, as confirmed by the results of both the dorsal root ganglia (DRG) and the Schwann cell culture in the presence of PDLLA matrix containing bFGF. The released bFGF enhanced the ability of the nerve fibers to sprout from dorsal root ganglia, and it accelerated the proliferation of Schwann cells.