Ann Thorac Surg 2005;79:672-676
© 2005 The Society of Thoracic Surgeons
New technology
Experimental Study of Replacing Circumferential Tracheal Defects With New Prosthesis
Hongcan Shi, MD, PhDa,*,
Zhifei Xu, MDb,
Xiong Qin, MDb,
Xuewei Zhao, MDb,
Dan Lu, MDa
a Department of Thoracic and Cardiovascular Surgery, Medical College of Yangzhou University, Yangzhou, Jiangsu Province, People's Republic of China
b Department of Thoracic and Cardiovascular Surgery, Second Military Medical University, Shanghai, People's Republic of China
Accepted for publication January 9, 2004.
* Address reprint requests to Dr Shi, Department of Thoracic and Cardiovascular Surgery, Medical College of Yangzhou University, 6 Huaihai Rd, Yangzhou, Jiangsu Province, PR China, P R China
shcan111{at}yahoo.com.cn
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Abstract
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PURPOSE: To investigate the feasibility of using a new tracheal prosthesis made of biomaterials to replace extensive circumferential tracheal defects.
DESCRIPTION: Three types of tracheal prostheses were developed and studied. Type I prosthesis was used in 8 mongrel dogs (group A), type II in 4 dogs (group B), and type III in 4 dogs (group C).
EVALUATION: In group A, one died from prosthetic dehiscence, another from anastomotic leakage, and the others had uneventful postoperative courses. The implanted prosthesis was completely incorporated with the recipient trachea, where different lengths of reepithelialization occurred on the luminal surface of the reconstructed trachea. Macroscopic examination showed scattered and different sizes of neo-ossification surrounding the implanted prosthesis. The prosthesis was radioopaque when exposed to routine x rays. In contrast, a relatively high number of complications occurred postoperatively in groups B and C.
CONCLUSIONS: Type I tracheal prosthesis, with further improvements, may be the optimal tracheal graft to replace circumferential tracheal defects, and appears very promising for clinical application.
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Introduction
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Since the first successful tracheal resection and reanastomosis was performed in the late 19th century [1], a large variety of surgical methods and techniques for tracheal reconstruction have been developed, depending on the degree of severity and the cause of tracheal diseases. An end-to-end anastomosis is preferred in the reconstruction of tracheal defects less than 50 to 60 mm in humans [2]. However, when a primary anastomosis is not feasible for the trachea, grafting is usually required. So far, various substitutes for tracheal replacement have been made, including the use of autologous tissue, autografts, allografts, prosthetic materials, tissue-engineered trachea, or a combination of these approaches [3–7]. Unfortunately, these approaches only have limited success experimentally and clinically due to anastomotic stenosis, immunologic rejection, local infections, prosthetic dislocation, and material failure [1, 3, 4].
In recent years, the perfect integration of polymeric sciences and life sciences has provided necessary substitutes for restoring and improving the functions of the impaired tissues and organs [8]. Polymeric biomaterials mainly consist of synthetic material and natural material. The former has the advantage of being able to better control the properties of materials such as biomechanical strength, which provides appropriate scaffolds for structural support and cell attachment. The latter, as an extracellular matrix component, may play a crucial role in facilitating cell growth and guiding the regeneration of the new tissue. Most recently, we integrated the two kinds of biomaterials and developed a new type of tracheal prosthesis. In our experiment, it was used on dogs with circumferential replacement of the cervical trachea and its efficacy was evaluated periodically after operation so as to obtain valuable scientific data and evaluate the technical feasibility for further clinical studies.
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Material and Methods
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Tracheal Prosthesis
Three types of tracheal prostheses were developed, whose basic skeleton of tubular mesh (length: 50 mm; thickness: 1 mm; internal diameter: 20 mm; mesh pore size: 300 µm) was knitted with polypropylene (PP) monofilament and poly (lactic-co-glycolic acid) (PLGA) fiber by means of a special microcaliber knitting machine. The inner side of type I tubular mesh was first coated with polyurethane solution (PU, with a concentration of 3.5%) and then with collagen solution (C, with a concentration of 2%). The exterior of type I was then immobilized with collagen-hydroxyapatite (C/HA) composites according to the following procedures: shape-casting, freeze-drying, and thermal cross-linking, which made the composites a microporous structure with pore sizes ranging from 100 to 150 µm. All of these processes kept the inner wall of type I airtight and the external wall microporous, and further increased its biocompatibilities and bioactivities (Fig 1). In contrast, the internal and external walls of type II were coated with polyurethane solution, which produced a prosthesis similar to a nonporous one, while type III was coated only with collagen solution so as to make it porous after the rapid postoperative biodegradation of the collagen.
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Fig 1. Schema of the structure of the type I tracheal prosthesis. (C/HA = collagen-hydroxyapatite; PLGA = poly (lactic-coglycolic acid); PP = polypropylene; PU/C = polyurethane solution with collagen solution.)
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Animal Experiments
Sixteen adult mongrel dogs, weighing 16.3 to 24.6 kg, were used to establish the experimental models. The dogs were divided into three subgroups: group A, with 8 dogs, used type I prosthesis; group B, having 4 dogs, received type II; and group C, again 4 dogs, used type III. General anesthesia was performed with an intramuscular injection of ketamine hydrochloride (15 mg/kg), and maintained intravenously with ketamine hydrochloride and Diazepam under spontaneous respiration. The establishment of animal models in groups A, B, and C followed the same operative procedures. The cervical trachea was exposed through an anterior midline incision in a supine position. A 45-mm-long segment of circumferential defect including 7 to 10 tracheal cartilage rings was created with a scalpel. Traction points were conducted to both proximal and distal cut ends of the native trachea to prevent retraction of the edges. Both ends of the graft were invaginated into the inside of the anastomotic sites of the native trachea to avoid excessive tension in suture line. A sterilized flexible endotracheal tube was intubated into the distal trachea through the operative field to maintain ventilation. After both anastomoses were completed using interrupted suturing with 3 to 0 Vicryl (Ethicon Inc, Cincinnati, OH), the cervical incision was then closed layer by layer.
Postoperatively, each dog was allowed to recover spontaneously. Three to six months after reconstruction, all long-term surviving dogs were killed with an overdose injection of sodium pentobarbital, followed by radiography, bronchoscopy, and microscopy at autopsy. All of the surgical and anesthetic procedures were done in accordance with the "Guide for the Care and Use of Laboratory Animals" published by the National Institutes of Health (National Institutes of Health Publication No. 85 to 23, revised 1985).
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Results
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In group A only one died from prosthetic dehiscence 96 days after implantation, another from anastomotic leakage caused by severe cervical incision infection 45 days after operation, and the rest had uneventful postoperative courses and did not show any respiratory symptoms after a follow-up of 3 to 6 months. There were no severe complications such as erosion of the surrounding organs, abscess formation, anastomotic dehiscence, or prosthetic dislocation in any of the dogs when they were killed. In all dogs living longer than 2 months after operation, tracheal prosthesis was completely incorporated with the recipient trachea and the fibrous connective tissue, together with new capillary vessels, had entirely infiltrated into the tubular mesh pores. In most cases, scattered and different sizes of neo-ossification substances, in which many irregular tunnels connect with bone marrow cavity containing affluent neonate vessels, bone marrow, and bone trabecula, were discovered in regions surrounding the implanted prosthesis (Figs 2 and 3). Endoscopic and histologic examinations revealed that the epithelium, whose color and luster are similar to the native trachea, had grown over the prosthesis, but the degree of reepithelialization varied in each dog. In these specimens, the neoepithelium was composed of pseudostratified ciliated columnar epithelium near the anastomotic ends (Figs 4 and 5). Scanning electron microscopy showed that the epithelial cells near the anastomoses displayed long and uniform cilia similar to those of normal tracheal epithelium (Fig 6). The implanted prosthesis was roentgenopaque when exposed to routine x rays because of the introduction of hydroxyapatite (Fig 7). Computed tomography revealed the implanted prosthesis had no structural changes and showed a perfect patent airway (Fig 8). At autopsy, the prostheses were found to retain original shape, and have some elasticity and rigidity to resist extrinsic compression.
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Fig 2. Photomicrograph of cartilaginous ossification and calcification (arrow) containing affluent neonate vessels, bone marrow, and bone trabecula (from a dog in group A 45 days after reconstruction). (Hematoxylin & eosin; original magnification, x200.)
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Fig 3. Scanning electron photomicrograph of transitional area (arrow) between tracheal cartilage and newly formed bone (from a dog in group A, 45 days after reconstruction). (Original magnification, x100.)
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Fig 4. Bronchoscopic appearance revealed neither stenosis nor granulation in the anastomotic region with good airway patency, and reengineered epithelial lining was recognized (from a dog in group A, 180 days after reconstruction).
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Fig 5. Columnar ciliated neoepithelium line the prosthesis near the anastomosis (from a dog in group A, 180 days after reconstruction). (Hematoxylin & eosin; original magnification, x200.)
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Fig 6. Scanning electron microscopic views of the luminal surface. Dense cilia like those of native tracheal epithelium are present on the prosthesis near the anastomosis (from a dog in group A, 180 days after reconstruction). (Original magnification, x3,000.)
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Fig 7. The implanted prosthesis is radiopaque when exposed to routine x-rays (from a dog in group A, 30 days after reconstruction).
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Fig 8. Computed tomography revealed the implanted prosthesis had no structural changes and showed a perfect patent airway (from a dog in group A, 30 days after reconstruction).
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In contrast, a relatively high number of postoperative complications occurred in group B, in which all 4 dogs surviving less than 90 days died of prosthetic dehiscence and displacement, or of progressive granuloma formation at the anastomoses. No ingrowth of ciliated epithelium on the luminal surface of the prosthesis was discovered. In group C, all 4 dogs died within 28 days of graft application. The main causes of death were mesh exposure, inward collapse, local infection, and air leakage resulting from the prosthesis itself.
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Comment
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Tracheal reconstruction, for which primary end-to-end anastomosis cannot be performed, remains an unsolved clinical problem. In the past 50 years, a variety of artificial trachea have been designed and assessed, but so far none have been satisfactory for clinical use. The failures are mainly due to their high mortality and morbidity rates such as prosthetic dislodgement, granuloma formation, local infection, air leakage, anastomotic stenosis, or obstruction [1].
Neville and colleagues [2] stated that the ideal tracheal prosthesis should be airtight, with a nonwettable inner surface, and adequate consistency to prevent respiratory collapse. It must be well accepted by the host, cause minimal inflammatory reaction, and can be molded into various sizes and configurations, and it should permit ingrowth of respiratory epithelium along the lumen. Above all, the most demanding requirements for tracheal prosthesis consist in its lateral rigidity and longitudinal flexibility. Though a surface of ciliated respiratory epithelium has proved to be desirable, it is not essential [1]. Kojima and colleagues [9] further emphasized that the trachea must maintain flexibility in the longitudinal direction to allow for free movement of the head, while maintaining the necessary rigidity to prevent collapse of the trachea during breathing.
The trachea is not a sterile organ. Okumura and colleagues [10] further argued that one of the main reasons why prosthetic tracheal reconstruction is difficult is that the trachea is located at a site facing the "external environment." Early reports showed the replacement of the trachea had been done with tubular conduits composed of solid and porous materials [1]. Although the solid graft such as silicone elastomer [2] might maintain an open airway for some time, the major drawback of this approach is lack of healing with the host tissue. In group B, dehiscence and migration of the prosthesis often occurred after reconstruction, which inevitably resulted in severe postoperative complications associated with local infection at the interface between foreign material and host tracheal bed. Epithelium rarely occurred overlying the prosthesis. We speculated that the type II prosthesis, similar to nonporous sealed structure and lack of rapid tissue and cell invasion into the pores of the mesh, might be responsible for such discouraging results. In contrast, the porous prosthesis like Marlax [10] mesh might permit ingrowth of host connective tissue into the mesh, but eventually tended to fail mainly due to its lack of lateral rigidity. In group C, with the rapid biodegradation and resorption of collagen postoperatively before the native tissue has entirely infiltrated the mesh pores, the exposure of tubular mesh may become inevitable, and local infection, inward collapse, and air leakage from the trunk of the prosthesis eventually occurred.
To overcome the above shortcomings, we developed the type I prosthesis with multilayer structures, which might fulfill the previously mentioned requirements of ideal tracheal prosthesis. The tubular mesh, used in this experiment and knitted with polypropylene and poly (lactic-co-glycolic acid), is the main structural component responsible for good rigidity and elasticity. An optimal microporosity of the external surface of the tubular mesh provides proper scaffolds for cell attachment, fibroblastic proliferation, and neovascular invasion into the mesh, which contributes to the growth and organization of connective tissues at the interface between the prosthesis and adjacent tissues. Polypropylene monofilament, with fairly good biomechanical properties, is one of the nonabsorbable materials. The soft poly (lactic-co-glycolic acid) fiber is characterized by its controllable biodegradability. The two kinds of biomaterials have the properties of the ability to be knitted and molded into various sizes and configurations. The elasticity and compliance of the type I prosthesis, after coating procedure with polyurethane, were enhanced. An impervious inner liner completely prevented air leakage from the mesh and the overgrowth of obstructing granulation tissue after operation. It was found at autopsy that the type I prosthesis remained mechanically strong enough to retain its original shape with a patent airway against extrinsic compression. In addition, approximately six weeks after operation, the ciliated columnar epithelium resembling normal respiratory mucosa near the anastomotic ends was found, and the degrees of reepithelialization varied in each dog. However, confluent regenerated epithelium over the entire width of the reconstructed prosthesis was not discovered, which might be an enduring and challenging issue to surgeons.
In order to enhance the tissue affinity of the type I prosthesis itself, we initially introduced biomaterials synthesized with collagen and hydroxyapatite. The exterior of the prosthesis was immobilized with collagen-hydroxyapatite spongelike microporous composites with a pore size range of 100 to150 µm, which provides sufficient three-dimensional templates and nutritional microenvironment to facilitate cell attachment, migration, and invasion into the mesh pores [11]. The collagen-hydroxyapatite composites, in which the hydroxyapatite nanocrystals align along the collagen molecules, possess excellent biocompatibility, biodegradability, and osteoconductive activities. The hydroxyapatite was biologically reabsorbed by phagocytosis of osteoclastlike cells and conducted osteoblasts to form new bone in the surrounding areas [12]. Macroscopic observation indicated that many scattered and different sizes of neo-ossification substances were found in regions surrounding the implanted prosthesis in dogs surviving longer than 45 days after operation. The newly formed bone, containing abundant bone trabecula as well as bone marrow, not only further reinforced the rigidity of the implanted prosthesis to resist airway pressure during breathing, but also produced a markedly coordinating effect in maintaining the supportive strength of the implanted prosthesis to keep the airway fully open. In addition, hydroxyapatite introduced into the extracoating layer materials made the prosthesis radiopaque when exposed to the routine roentgenogram, which contributed to periodic dynamic observation after graft application.
The type I tracheal prosthesis, with an optimal structure of intraluminal impermeability and extraluminal microporosity, has sufficient lateral elasticity and longitudinal flexibility. The reconstruction of the trachea makes it desirable to use the type I prosthesis, which meets the requirements of reliability, stability, biocompatibility, and few complications. This prosthesis, as an appropriate alternative, may prove potential for the long-segment circumferential tracheal reconstruction, and appears very promising for clinical studies.
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Disclosures and Freedom of Investigation
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The tested technology involved in this study was not purchased or borrowed. The authors had full control of the design of the study, methods used, outcome parameters, analysis of data, and production of the written report.
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Footnotes
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The Society of Thoracic Surgeons, the Southern Thoracic Surgical Association, and The Annals of Thoracic Surgery neither endorse nor discourage use of the new technology described in this article.
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References
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Abstract
Introduction
