Application of in-body tissue architecture-induced Biotube vascular grafts for vascular access: Proof of concept in a beagle dog model.

INTRODUCTION: The first choice of vascular access for hemodialysis is an autogenous arteriovenous fistula, because prosthetic arteriovenous grafts have a high probability of failure. In this study, Biotubes, in-body tissue architecture-induced autologous collagenous tubes, were evaluated for their potential use as vascular access grafts. Three animal implantation models were developed using beagle dogs, and the in vivo performance of Biotubes was observed after implantation in the acute phase as a pilot study. METHODS: Biotubes (internal diameter ca. 4.0 mm, length ca. 5.0 cm, and wall thickness ca. 0.7 mm) were prepared through subcutaneous embedding of specially designed molds in beagle dogs for 8 weeks. The Biotubes were then implanted between the common carotid artery and the jugular vein of beagles via three methods, including side-to-side (in) -end-to-end (out) as type 1 (n = 4), side-to-side (both) as type 2 (n = 4), and side-to-end (in) -end-to-side (out) as type 3 (n = 1 using a composite Biotube). RESULTS: Although two cases in type 1 and 2 resulted in Biotube deformation, all cases were patent for 4 weeks and maintained a continuous turbulent flow. At 4 weeks after implantation, percutaneous puncture could be performed repeatedly without aneurysm formation or hemorrhage. CONCLUSION: Within a short implantation period, with limited animal numbers, this proof-of-concept study showed that Biotubes may have a high potential for use in vascular access.

Shape memory of in-body tissue-engineered Biotube® vascular grafts and the preliminary evaluation in animal implantation experiments.

BACKGROUND: Tissues formed by in-body tissue architecture (iBTA) are soft and flexible. However, strongly bending iBTA-induced vascular grafts, called biotubes, may cause lumen collapse by kinking, subsequently leading to occlusion after implantation. In this study, we developed a method for biotube shape memory and verified its performance in preliminary animal implantation experiments. METHODS: Straight biotubes were prepared by subcutaneous embedding of straight molds into beagle dogs for two months. Upon overnight immersion of the obtained straight biotubes in a 70% ethanol solution under U-shape framing, the biotubes maintained their U shape even after washing with saline solution. Additionally, spiral-shaped biotubes formed from goats using spiral molds could be stretched straight via the same alcohol treatment. RESULTS: Within limited acute-phase animal implantation experiments, U-shaped biotubes functioned as AV shunt grafts in the femoral region of the beagle without deformation of vascular shape (N.=11). In addition, the long straight biotubes derived from spiral molds could be interposed between goat carotid arteries while maintaining their straight shape (N.=2). All implants maintained perfect patency at the 1-month follow-up period without any evidence of vascular deformation. CONCLUSIONS: By retaining iBTA-induced tissues in an alcohol solution in the target shape, the shape of the tissues was imprinted and maintained even after implantation within a limited acute period. Therefore, in order to obtain tissues of various shapes, it is unnecessary to use a mold design to maintain the individual shape.

Development of long in vivo tissue-engineered "Biotube" vascular grafts.

In-body tissue architecture (iBTA), a cell-free, in vivo tissue engineering technology that can produce autologous implantable tissues of the desired shape by subcutaneously embedding specially designed molds, was used to develop long tubular collagenous tissues called Biotubes. Spiral molds for long Biotubes were assembled with an outer pipe-shaped spiral shell and an inner spiral mandrel, and embedded into subcutaneous pouches of beagle dogs or goats for 1 or 2 months. Tubular collagenous tissues were formed at the space between the shell and the mandrel of the mold. Depending on the spiral turn number in the mold, Biotubes of 25 cm or 50 cm (internal diameter 4 mm or 5 mm) were prepared with nearly homogeneous mechanical and histological properties over their entire length. Biotubes stored in 70% ethanol were allogenically implanted into beagle dogs or goats to evaluate their in vivo performance. The 25-cm Biotubes functioned as arterial grafts with no need for luminal modification or mechanical support, and demonstrated vascular reconstruction within 3 months after implantation into dogs. The 50-cm Biotubes functioned as arteriovenous shunt grafts in the neck region of goats without thrombus formation and vascular deformation for 1 month. Thus, the world's longest tissue-engineered vascular grafts with small diameter could be developed using iBTA.

Construction of 3 animal experimental models in the development of honeycomb microporous covered stents for the treatment of large wide-necked cerebral aneurysms.

The treatment of large or wide-necked cerebral aneurysms is extremely difficult, and carries a high risk of rupture, even when surgical or endovascular methods are available. We are developing novel honeycomb microporous covered stents for treating such aneurysms. In this study, 3 experimental animal models were designed and evaluated quantitatively before preclinical study. The stents were prepared using specially designed balloon-expandable stents (diameter 3.5-5.0 mm, length 16-28 mm) by dip-coating to completely cover their struts with polyurethane film (thickness 20 µm) and microprocessing to form the honeycomb pattern after expansion. (1) In an internal carotid artery canine model (n = 4), all stents mounted on the delivery catheter passed smoothly through the tortuous vessel with minimal arterial damage. (2) In an the large, wide-necked, outer-sidewall aneurysm canine model, almost all parts of the aneurysms had embolized immediately after stenting (n = 4), and histological examination at 2 months revealed neointimal formation with complete endothelialization at all stented segments and entirely organized aneurysms. (3) In a perforating artery rabbit model, all lumbar arteries remained patent (n = 3), with minimal change in the vascular flow pattern for over 1 year, even after placement of a second, overlapping stent (n = 3). At 2 months after stenting, the luminal surface was covered with complete thin neointimal formation. Excellent embolization performance of the honeycomb microporous covered stents without disturbing branching flow was confirmed at the aneurysms in this proof-of-concept study.

Development of an in vivo tissue-engineered vascular graft with designed wall thickness (biotube type C) based on a novel caged mold.

Small-diameter biotube vascular grafts developed by in-body tissue architecture had high patency at implantation into rabbit carotid arteries or rat abdominal aortas. However, the thin walls (34 ± 14 µm) of the original biotubes made their implantation difficult into areas with low blood flow volumes or low blood pressure due to insufficient mechanical strength to maintain luminal shape. In this study, caged molds with several windows were designed to prepare more robust biotubes. The molds were assembled with silicone tubes (external diameter 2 mm) and cylindrical covers (outer diameter 7 mm) with 12 linear windows (1 × 9 mm). After the molds were embedded into beagle dorsal subcutaneous pouches for 4 weeks, type C (cage) biotubes were obtained by completely extracting the surrounding connective tissues from the molds and removing the molds. The biotube walls (778 ± 31 µm) were formed at the aperture (width 1 mm) between the silicone rods and the covers by connective cell migration through the windows of the covers. Excellent mechanical properties (external pressure resistance, approximately 4 times higher than beagle native femoral arteries; burst strength, approximately 2 times higher than original biotubes) were obtained. In the acute phase of implantation of the biotubes into beagle femoral arteries, perfect patency was obtained with little stenosis and no aneurysmal dilation. The type C biotubes may be useful for implantation into peripheral arteries or veins in addition to aortas.

Development of an in vivo tissue-engineered valved conduit (type S biovalve) using a slitted mold.

In autologous valved conduits (biovalves) using in-body tissue architecture, the limited area available for leaflet formation is a concern. In this study, we designed a novel biovalve mold with slits to enhance in vivo cell migration, regardless of size. As a control, the original mold without slits was used. When both types of molds were embedded into subcutaneous pouches in beagle dogs for 8 weeks, the outer surfaces of all molds were completely covered with connective tissue to form conduit tissue. In the molds without slits, the leaflet size was limited to half of the design. In contrast, in the mold with slits, the complete leaflet area was formed. Upon trimming excess peripheral tissues, removing the mold, and cutting the connective tissue formed at the slits, completely autologous connective tissue biovalves with the designed leaflet area were obtained as type S (diameter, 6-28 mm) biovalves. The slit structure customized to the mold was effective for allowing cells to enter, thereby facilitating cell migration and contributing to the successful preparation of reliable biovalves of various physiological sizes suitable for all clinical uses.