Long-term results of tissue-engineered small-caliber vascular grafts in a rat carotid arterial replacement model.

The concept of tissue engineered small-caliber vascular grafts (TE-SCVGs) is theoretically ideal. In this study, we evaluated the long-term (more than 1 year) course of TE-SCVGs using a rat carotid arterial replacement model. We fabricated a TE-SCVG scaffold (0.7 mm in diameter) with electrospun nano-scale fibers. Poly-ε-caprolactone was used as a biodegradable polymer. These artificial vessels were then used in carotid arterial replacement performed on Sprague-Dawley rats. The implanted grafts were removed at an early phase (1, 2, 6 weeks), middle phase (12, 24 weeks), and late phase (48, 72 weeks) after implantation. Twenty-nine patent grafts from among the 40 implanted grafts (patency 72.5 %) could be evaluated. No aneurysm formation was observed during the follow-up period. Endothelial cells positive for immunostaining with von Willebrand factor were found to be already attached to the inner surface of the TE-SCVGs in the early phase. The percentage of smooth muscle cell specific marker (α-smooth muscle actin and calponin with fluorescent immunostaining) positive cells, which seemed to be mesenchymal cells in the graft wall, increased with time, while, in contrast, the scaffold material decreased. Even after 72 weeks, however, although the scaffold material had degraded, it had not disappeared completely. These results show that the novel TE-SCVGs we developed were still functioning in the rat carotid arterial circulation after more than 1 year. However, further investigations will be required with regard to regeneration of the SMC layer and the complete degradation of graft materials.

Regeneration of rotator cuff tear using electrospun poly(d,l-Lactide-Co-Glycolide) scaffolds in a rabbit model.

PURPOSE: The purpose of this study was to evaluate an application of poly(d,l-lactide-co-glycolide) (PLG) scaffold created by electrospinning in a rabbit rotator cuff defect model. METHODS: Forty-two Japanese white rabbits were used in this study. Defects of the infraspinatus tendon were created, and the PLG scaffolds were implanted. Contralateral infraspinatus tendons were reattached without creating defects. Histologic analyses were performed 4, 8, and 16 weeks after the operation, and mechanical evaluations were performed 0, 4, 8, and 16 weeks after the operation. RESULTS: Scaffold fibers remained without dissolution and spindle-shaped cells were observed inside of the scaffold at 4 weeks postoperatively. At 8 weeks, the PLG scaffold had dissolved and bone formation was observed at the scaffold-bone interface. At 16 weeks, the scaffold-bone interface matured and expression of type II collagen was observed. A statistical difference in ultimate failure load was not seen between the scaffold group and reattachment group or normal tendon after 8 weeks postoperatively. The stiffness in the scaffold group was not significantly different from that in the reattachment group at each time point. However, it was significantly weaker than normal tendon at each time point. CONCLUSIONS: Transplantation of cell-free PLG scaffold showed cell migration and type II collagen and proteoglycan expression at the scaffold-bone junction by 16 weeks postoperatively with a sufficient ultimate failure load in a rabbit rotator cuff defect model. CLINICAL RELEVANCE: The PLG scaffold could be applied to bridge rotator cuff defects. The results showed that bridging with scaffold can be equivalent to reattachment.

Electrospun synthetic polymer scaffold for cartilage repair without cultured cells in an animal model.

PURPOSE: The purpose of our study was to explore the possibility that an electrospun bioabsorbable scaffold could be used in the treatment of a full-thickness articular defect without the addition of exogenous cells in a rabbit model. METHODS: Two types of poly(D,L-lactide-co-glycolide) (PLG) scaffolds, a solid cylindrical type and a cannulated tubular type, were made with the electrospinning method. Osteochondral defects, 5 mm in diameter and 5 mm in depth, made on the femoral condyles of rabbits were filled with these scaffolds, and the repair process was investigated histologically. RESULTS: In the groups in which the defect was filled with the scaffold, fibrous tissue at the articular surface of the scaffold was observed at postoperative week 2. Thereafter cartilage at the articular surface and bone at the subchondral zone were regenerated, and the repaired cartilage was maintained through postoperative week 24. By contrast, the untreated defect was filled with hematoma at postoperative week 2; thereafter regenerated cartilage and bone were observed. However, the surface of the articular cartilage was not regular, and regenerated cartilage was not well organized. The histologic scores of the groups in which the defect was filled with cannulated tubular electrospun PLG scaffolds were significantly higher than those of the untreated defect group at postoperative weeks 12 and 24 (P < .01). CONCLUSIONS: The electrospun PLG scaffold could repair a 5-mm osteochondral defect created in the rabbit model without exogenous cultured cells. CLINICAL RELEVANCE: The electrospun PLG scaffold could repair full-thickness osteochondral defects. The cannulated type of PLG scaffold has the possibility to lead not only to good regeneration of cartilage but also to easy transplantation by use of a guidewire through the cannulas in the scaffold.

Local delivery of mesenchymal stem cells with poly-lactic-co-glycolic acid nano-fiber scaffold suppress arthritis in rats.

Mesenchymal stem cells (MSC) have been used recently for the treatment of autoimmune diseases in murine animal models due to the immunoregulatory capacity. Current utilization of MSC requires cells in certain quantity with multiple courses of administration, leading to limitation in clinical usage. Here we efficiently treated collagen-induced arthritis rats with a single local implantation with reduced number of MSC (2∼20% of previous studies) with nano-fiber poly-lactic-co-glycolic acid (nano-fiber) scaffold. MSC seeded on nano-fiber scaffold suppressed arthritis and bone destruction due to inhibition of systemic inflammatory reaction and immune response by suppressing T cell proliferation and reducing anti- type II collagen antibody production. In vivo tracing of MSC demonstrated that these cells remained within the scaffold without migrating to other organs. Meanwhile, in vitro culture of MSC with nano-fiber scaffold significantly increased TGF-ß1 production. These results indicate an efficient utilization of MSC with the scaffold for destructive joints in rheumatoid arthritis by a single and local inoculation. Thus, our data may serve as a new strategy for MSC-based therapy in inflammatory diseases and an alternative delivery method for bone destruction treatment.

Prevention of arterial graft spasm in rats using a vasodilator-eluting biodegradable nano-scaled fibre.

OBJECTIVES: Arterial graft spasm occasionally causes circulatory collapse immediately following coronary artery bypass graft. The aim of this study is to evaluate the efficacy of our developed materials, which were composed of milrinone (phosphodiesterase III inhibitor) or diltiazem (calcium-channel blocker), with nano-scaled fibre made of biodegradable polymer to prevent arterial spasm. METHODS: Milrinone- or diltiazem-releasing biodegradable nano-scaled fibres were fabricated by an electrospinning procedure. In vivo milrinone- or diltiazem-releasing tests were performed to confirm the sustained release of the drugs. An in vivo arterial spasm model was established by subcutaneous injection of noradrenalin around the rat femoral artery. Rats were randomly divided into four groups as follows: those that received 5 mg of milrinone-releasing biodegradable nano-scaled fibre (group M, n = 14); 5 mg of diltiazem-releasing biodegradable nano-scaled fibre (group D, n = 12); or those that received fibre without drugs (as a control; group C, n = 14) implanted into the rat femoral artery. In the fourth group, sham operation was performed (group S, n = 10). One day after the implantation, noradrenalin was injected in all groups. The femoral arterial blood flow was measured continuously before and after noradrenalin injection. The maximum blood flow before noradrenalin injection and minimum blood flow after noradrenalin injection were measured. RESULTS: In vivo drug-releasing test revealed that milrinone-releasing biodegradable nano-scaled fibre released 78% of milrinone and diltiazem-releasing biodegradable nano-scaled fibre released 50% diltiazem on the first day. The ratios of rat femoral artery blood flow after/before noradrenalin injection in groups M (0.74 ± 0.16) and D (0.72 ± 0.05) were significantly higher than those of groups C (0.54 ± 0.09) and S (0.55 ± 0.16) (P < 0.05). CONCLUSION: Noradrenalin-induced rat femoral artery spasm was inhibited by the implantation of milrinone-releasing biodegradable nano-scaled fibre or diltiazem-releasing biodegradable nano-scaled fibre. These results suggested that our materials might be effective for the prevention of arterial graft spasm after coronary artery bypass graft.

Development of novel drug-eluting biodegradable nano-fiber for prevention of postoperative pulmonary venous obstruction.

Pulmonary venous obstruction (PVO) after correction of total anomalous pulmonary venous connection (TAPVC) frequently occurs due to intimal-hyperplasia and the required re-operation. We have developed a novel sustained-release drug delivery system, using Tacrolimus-eluting biodegradable nano-fiber (TEBN). It consists of nano-scale fiber composed of biodegradable polymer and Tacrolimus. This study evaluated the effects of TEBN for prevention of venous anastomotic stricture in a rat model to apply to PVO operation. Tacrolimus was incorporated into poly (L-lactide-co-glycolide). The venous stricture model was made by rat inferior vena cava anastomosis. The IVC anastomosis was covered with TEBN with 1.0 wt% Tacrolimus (n=12) or without TEBN as a control (n=12), and evaluated histologically at 1, 2, and 4 weeks after operation. The ratio of intimal area was significantly reduced in the TEBN group compared with the control group (ratio; 1 week: 0.43+/-0.26 vs. 0.07+/-0.04, P=0.04, 2 weeks: 0.39+/-0.19 vs. 0.05+/-0.02, P=0.01, 4 weeks: 0.31+/-0.15 vs. 0.09+/-0.04, P=0.03, control vs. TEBN, respectively). Histological findings showed endothelialization along the inner surface of the vein even in TEBN. The TEBN reduced intimal hyperplasia and preserved endothelialization even in a venous stricture. These results suggested that this strategy might be useful for prevention of recurrent PVO after TAPVC correction.

A new strategy for prevention of anastomotic stricture using tacrolimus-eluting biodegradable nanofiber.

OBJECTIVE: We developed a novel sustained drug-eluting device using tacrolimus-eluting biodegradable nanofiber to prevent anastomotic stricture and evaluated the effects in a rat abdominal aortic anastomosis model. METHODS: In vitro and in vivo tacrolimus release tests for tacrolimus-eluting biodegradable nanofiber were performed to confirm its sustained release. To verify the prevention of anastomotic stricture, tacrolimus-eluting biodegradable nanofiber was placed around the end-to-end anastomosis of abdominal aorta in rats. Five rats were allocated to the following 5 groups: (1) control without tacrolimus-eluting biodegradable nanofiber, (2) 5 mg of nanofiber only (0 wt% of tacrolimus), (3) 5 mg of tacrolimus-eluting biodegradable nanofiber containing 0.04 wt% of tacrolimus, (4) 5 mg of tacrolimus-eluting biodegradable nanofiber containing 0.1 wt% of tacrolimus, and (5) 5 mg of tacrolimus-eluting biodegradable nanofiber containing 1.0 wt% of tacrolimus. Morphometric and histologic analyses including immunohistochemistry were performed in each of the groups 2 weeks after the operation. RESULTS: The tacrolimus-eluting biodegradable nanofiber gradually released tacrolimus for at least 1 month in vitro and in vivo. The ratio of intimal area was significantly reduced in the 1.0 wt% tacrolimus-eluting biodegradable nanofiber group compared with the other groups (0.26, 0.24, 0.25, 0.21, and 0.08 in control, 0 wt%, 0.04 wt%, 0.1 wt%, and 1.0 wt%, respectively, P < .05). The cells, which constitute intimal hyperplasia, were positive for smooth muscle actin and SMemb, and factor VIII revealed that endothelial cells covered the surface of the aortic lumen even in the 1.0 wt% tacrolimus-eluting biodegradable nanofiber group in immunohistochemistry. CONCLUSION: Tacrolimus-eluting biodegradable nanofiber reduced neointimal hyperplasia and preserved endothelialization. This device may be useful in the prevention of anastomotic stricture.