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Experimental Studies |
1 From the Departments of Radiology (A.R.F., N.L.D.) and Pathology (C.G.A.T.), University of Michigan Medical Center, Ann Arbor, Mich. Received July 18, 2003; revision requested September 25; revision received August 3, 2005; accepted September 6; final version accepted November 18. Supported by a grant from the Cardiovascular and Interventional Radiology Research and Education Foundation (now known as the Society of Interventional Radiology Foundation). Address correspondence to A.R.F., Diagnostic Radiology Department, Dartmouth-Hitchcock Medical Center, One Medical Center Dr, Lebanon, NH 03756 (e-mail: andrew.r.forauer{at}Hitchcock.org).
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ABSTRACT |
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Materials and Methods: The proposal was approved by the University Committee on the Use and Care of Animals. Tunneled silicone 7-F catheters were placed via a jugular vein in eight swine. The animals were separated into four groups of two pigs each according to catheter indwelling times of 7, 14, 30, and 45 days. After the animals were sacrificed, the catheter, access vein, and cranial vena cava were dissected, removed en bloc, and fixed in formalin. Histologic evaluation was performed by using standard light microscopy on hematoxylin-eosin stained specimens; immunohistochemistry was also performed to confirm specific cell populations.
Results: Catheter-related sheaths that covered 33%–100% of the intravascular catheter length were identified in all eight catheter specimens. After 7 days, catheters had a partial or circumferential mixed cellular and noncellular covering consisting of smooth muscle cells, thrombus, and areas with endothelial cell populations. Sheaths from catheters excised at 14 days were characterized by prominent endothelial cell and smooth muscle cell proliferation. Catheters excised at 30 and 45 days showed less prominent cellularity and more prominent collagen content, in a well-developed sheath, than did those excised at 7 and 14 days. With longer catheter indwelling times, an endothelial layer, indistinguishable from the adjacent vein wall, covered the catheter surface.
Conclusion: The sheath that develops around central venous catheters in the swine model consists of cellular and noncellular components. A substantial proportion of the sheath is made up of a smooth muscle cell and collagen layer with overlying endothelial cells.
© RSNA, 2006
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INTRODUCTION |
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Despite catheter malfunction due to noninfectious complications in up to one-third of patients (4,7,8), there have been few complete examinations of the composition and biologic evolution of the catheter-related sheath (9,10). O'Farrell and colleagues (9) studied the reaction to silicone catheter materials in rats. The catheters were placed via a jugular approach, and the animals were sacrificed at 3, 7, and 60 days. They observed catheter-related thrombus with points of attachment to the vein wall in the earliest group. The thrombus underwent the expected changes of organization at the 7- and 60-day observation points.
In 1998 and 2001, Xiang et al (10,11) at the Catholic University of Leuven, Belgium, performed detailed studies of catheter-related sheath formation in a small-animal (rat and rabbit) model. This work showed that the catheter sleeve possessed a cellular component that included smooth muscle and endothelial cells (10–12).
The purpose of the present study was to evaluate the development and histologic features of jugular vein catheter–related (fibrin) sheaths in a swine model.
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MATERIALS AND METHODS |
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Catheter Insertion
Each swine's right neck was shaved, prepared, and draped in a sterile fashion. By using ultrasonographic guidance, either the right external jugular vein (n = 7) or internal jugular vein (n = 1) was identified and punctured with a 19-gauge needle. (The first 6.6-F catheter was inserted via the internal jugular vein. The remaining catheters were placed via the external jugular vein, simply because of the anatomy of the pig and the ease of creating the short subcutaneous tunnel necessary to conceal and fix the end of the catheter.) A 0.032-inch 3-mm J guidewire (Bard Access, Salt Lake City, Utah) was advanced with fluoroscopic guidance into the cranial vena cava. (The cranial vena cava in swine is analogous to the superior vena cava in humans.)
The venotomy site was dilated to 7 or 8 F, and a short peel-away sheath (Bard Access; or Cook, Bloomington, Ind) was placed. A track of between 3 and 5 cm in length was selected cephalic and lateral to the venotomy for the subcutaneous tunnel. A small (<1-cm) incision was made with a number 11 scalpel. A blunt-ended tunneling device was then used to create the tunnel. The catheter was pulled through the tunnel. A subcutaneous port (M.R.I. Low Profile; Bard Access) with a 6.6-F single-lumen silicone catheter was implanted in one animal. In the remaining seven pigs, 7-F dual-lumen silicone catheters (Bard Access) were used for insertions. The catheter was sized, cut, and inserted via the peel-away sheath. The catheter tip was positioned in the distal part of the cranial vena cava.
The catheter was flushed with a mixture of heparin and saline (10 U of heparin per milliliter of saline), and two 2-0 nylon sutures (Prolene; Ethicon, Somerville, NJ) were tied around the catheter to prevent back bleeding. No suture was placed around the access vein, and the vessel was allowed to remain patent. The overlying skin was approximated by using a single-layer closure technique with absorbable suture material (3-0 Vicryl; Ethicon). Finally, a small amount of tissue glue (Nexaband; Veterinary Products Laboratories, Phoenix, Ariz) was applied at the surface of the venotomy and remote tunneling site. After the catheter was placed, a digital chest radiograph was obtained to document final catheter position. A single investigator with 5 years of experience in interventional radiology (A.R.F.) performed all catheter insertions.
The animals were returned to their cages, placed under standard warming lamps, and allowed to recover from the anesthesia. They were extubated once swallowing reflexes returned. They were observed throughout the duration of the study period for signs of local infection, systemic infection, and any other manifestations of illness. The animals did not receive anticoagulants, antiplatelet agents, or antibiotics during the postoperative course.
Catheter Removal
Catheter removal was performed on two animals in each group after catheter indwelling times of 7, 14, 30, and 45 days. Each animal was sedated by using intramuscular telazol (6 mg/kg) and xylazine (2.2 mg/kg). The animal was weighed so that we could calculate the dose of heparin to administer. A preremoval digital radiograph of the chest was obtained to document the final position of the catheter. Before we euthanized the animal, a weight-based heparin bolus (300 U/kg) was administered. The heparin was allowed to circulate for 5 minutes. Euthanasia was performed by using an intravenous injection (390 mg/mL [0.22 mL/kg]) of sodium pentobarbital (Fatal-Plus solution; Vortech Pharmaceuticals, Dearborn, Mich).
After death was documented, the animal was placed in the supine position. The sternum was cut by using a sternal saw (Stryker, Kalamazoo, Mich). The right anterior and lateral ribs were cut away, enabling better exposure of the mediastinal structures. A skin incision was made at the base of the neck, and, by using sharp and blunt dissection, the subcutaneous portion of the catheter was located. The tissues surrounding the catheter were dissected free. The venotomy was identified, and the dissection was carried along the vessel (external or internal jugular vein) to its confluence with the vena cava. The distal part of the vena cava and the cuff of the right atrium were also dissected out. The catheter, jugular vein, vena cava, and right atrium were removed as an intact specimen, pinned on a styrofoam block, and immersed in 10% formalin. Again, a single investigator (A.R.F.) performed all the removal procedures.
Specimen Processing
The tissues were fixed in formalin for a minimum of 7 days. The veins, with their indwelling catheters still in place, were then sliced beginning at the site of the catheter's entry into the vein and continuing distally at 1-cm intervals. Transverse slices of the vein, including the catheter, of approximately 0.4–0.6 cm thickness were placed in cassettes and embedded in paraffin. Standard hematoxylin-eosin slides were produced from each cassette.
The hematoxylin-eosin slides were examined with low-power (magnification, x25) light microscopy, and the extent of sleeve formation was mapped onto a diagram that depicted the intravascular portion of the catheter. The slides were also examined with high-power (magnification, up to x400) light microscopy to further delineate histologic characteristics. Specific cell populations (eg, smooth muscle cells, endothelial cells) and their arrangements were noted. The appearance of the extracellular space was also observed. The slides were interpreted by two of the authors (A.R.F. and C.G.A.T., who were junior faculty members in the Departments of Radiology and Pathology, respectively). The slides were also reviewed by a senior pathologist who had more than 30 years of experience in vascular pathology, as well as by a member of the veterinary faculty who had more than 15 years of experience in veterinary research.
Unstained slides (blanks) from specimen cassettes that were obtained from within 1 cm of the venotomy site and less than or equal to 1 cm from the distal end of the catheter-related sheath were then selected. Immunohistochemistry staining (Dako; Carpinteria, Calif) was performed on these slide specimens. The following stains were used: anti–smooth muscle actin (mouse-antihuman; dilution, 1:50), anti–von Willebrand factor (rabbit-antihuman; dilution, 1:500), anti-myeloperoxidase (rabbit-antihuman; dilution, 1:16 000), and anti-CD68 (mouse-antihuman; dilution, 1:1600). These were used to identify smooth muscle cells, endothelial cells, neutrophils, and monocytes and macrophages, respectively.
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RESULTS |
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The cranial vena cava remained patent in all animals during the postinsertion time course. At postmortem examination, no specimen was thrombosed. Catheter patency was not evaluated and was not a part of the study.
Catheter-related sheaths could be identified grossly and microscopically (Fig 1) in all eight catheter specimens, particularly when the specimens were placed in a transverse orientation within the cassette before being embedded in paraffin. No vena caval specimen was opened longitudinally before fixation because of the possibility of disrupting catheter-to–vein wall bridges that may have formed.
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Hematoxylin-eosin staining revealed well-formed catheter-related sheaths in all specimens. The sheath began at the site where the catheter entered the access vessel and was continuous along the catheter distally.
An inflammatory response was present at the site where the catheter entered each vein. This observation was true for all indwelling time groups. The more acute inflammatory cell populations (neutrophils) predominated in the animals with shorter indwelling times (7 and 14 days), while more chronic monocytic cells were present in the animals with longer indwelling times (30 and 45 days). Cells were concentrated within the vein wall, with the early cellular response surrounding the catheter. The observation was based on hematoxylin-eosin staining, however; this was not confirmed by the immunohistochemistry markers for neutrophils (anti-myeloperoxidase) or monocytes (anti-CD68).
The two 7-day catheters had sheaths that were partially or circumferentially composed of a mixed cellular and noncellular covering of smooth muscle cells in a background of collagen (Fig 2a). Included in this were red blood cells and areas of associated thrombus. Adjacent vein walls along the course of the catheter from the jugular vein to the vena cava exhibited focal areas of intimal thickening. Fourteen-day catheters (n = 2) had sheaths with endothelial and smooth muscle cell proliferation that was more prominent than those in the 7-day specimens. The phenomenon of cellular proliferation was attenuated in the animals with longer indwelling times of 30 (n = 2) and 45 (n = 2) days. Specifically, there was a lower observed proportion of smooth muscle cells in the animals with 30- and 45-day indwelling times than in those with 7- and 14-day times. Conversely, collagen content in the 30- and 45-day specimens was higher than that in the 7- and 14-day specimens (Fig 2b). The presence of smooth muscle cells was confirmed with the immunologic stains. Cells staining positive for smooth muscle actin were present in all specimens. These cells could be seen both within the substance of the sleeve material and in the neovessels within the sheath (Fig 2c). These cells assumed a typical orientation to the vessel lumen, with the long axis oriented with the circumference of the vessel.
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DISCUSSION |
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There are a number of articles in the literature that are concerned with the topic of catheter-related (fibrin) sheaths; a minority of these focus on the sheaths' histopathologic features and microscopic development. An often-referenced article (15) describes the findings at autopsy in 55 patients with subclavian vein catheters. In that study, sheaths were identified in all specimens, even as soon as 24 hours after catheter insertion. Microscopic evaluation was reported to reveal a predominately fibrin makeup, "with no evidence of endothelialization or organization." The sleeve was observed, in many cases, to be adherent to the adjacent vein wall.
Well into the 1990s, descriptions of a catheter-related sheath consisting of "fibrin and thrombocytes" (20) or a "layer of investing fibrin and proteinaceous material" (24), could still be found in the literature.
Later, two reports offered a more detailed microscopic and histologic description. In 1996, a study (9) was performed in which small-caliber silicone catheters were inserted in 15 rats. The catheters were placed via a jugular approach, and the animals were sacrificed at 3, 7, and 60 days. Catheter-related thrombus, with points of attachment to the vein wall, was observed in the earliest group. The thrombus underwent changes typical of organization at the 7- and 60-day observation points and evolved into what was described as a "dense fibrous connective tissue containing numerous spindle-shaped fibroblasts" (9).
In a second, larger study (10), again performed by using a rat model, catheters placed via the jugular vein in 123 animals were evaluated. Histologic changes were studied at catheter indwelling times that ranged from 1 day to 6 months. A true pericatheter thrombus was identified in all animals within the first 3 days after catheter insertion. A transformation occurred from pericatheter thrombus to a more cellular structure composed of collagen with smooth muscle and endothelial cells; this latter structure appeared 1–4 weeks after catheter placement.
Three kinds of catheter-associated thrombus have been described (11). The first variety is a meshlike thrombus that bridges the vein wall and catheter. This is thought to evolve into the mixed cellular and collagen catheter-related sleeve described by these authors in an earlier report (10). A second, nonorganized form of thrombus has been termed "sleeve-related thrombus" and is found on the distal aspects of the indwelling catheter itself. This variety has no attachment to the vein wall and is histologically and physically separate. Last, mural thrombus is found on the vein wall adjacent to the distal intravascular aspect of the catheter. This thrombus undergoes organization and is thought to become incorporated into the vein wall. No sleeve-related thrombus was found in our specimens. The other two forms—thrombus that evolves into the vein wall-catheter bridges and pericatheter thrombus that becomes incorporated into the vein wall itself—were observed in our specimens. It is uncertain why a thrombus at a particular location develops into a cellular bridge instead of incorporating into a vein wall, although catheter motion may influence this process (25).
The role of catheter-tip trauma and associated thrombus formation has been examined in a swine model (25). Silicone catheters with or without a 0.018-inch wire stabilizing loop at the distal indwelling tip were inserted, and their tips were positioned in the distal aspect of the superior vena cava. In the group in which catheter tips were stabilized by the wire loop, there was only a mild increase in vein wall thickness without vein wall thrombus. In the control group (without the stabilizing loop), mural thrombus formed at the site of local vein wall trauma caused by catheter tip motion. This thrombus subsequently underwent organization and resulted in vein wall thickening and intimal hyperplasia.
The organization of intravascular thrombus involves an infiltration by smooth muscle cells and the development of a vascularized connective tissue that includes collagen, smooth muscle cells, and endothelial cells (26,27). Inflammatory cells are also known to be involved in venous thrombosis (28). In the present study, cells resembling inflammatory cells were seen at hematoxylin-eosin staining, but the antibody-specific stains did not confirm inflammatory cell lines. This raised the question of the antibody specificity and reactivity of these particular stains (anti-CD68 of mouse origin and anti-myeloperoxidase of rabbit origin) in the swine model. The use of formalin-fixed specimens may also have influenced antibody labeling.
The development of the catheter-related sheath is postulated to begin with thrombus that develops after the trauma associated with the catheter insertion procedure (12,16). Local trauma occurs at the venotomy site. Factors contributing to thrombus formation include disturbance of normal flow through the venous segment and stasis that occurs between the catheter and the vein wall. Other locations of trauma occur at foci of friction of the catheter against the vein wall or catheter tip impact against the vein wall and in segments where catheters lie in acute angles within the course of the vein (12,25). In addition, acute or chronic (organized) thrombus has been confirmed in catheter stripping specimens (29).
A mixed cellular and noncellular covering was observed in all catheter specimens in our study. The development of a sheath containing collagen and proliferating smooth muscle cells and the progressive covering by endothelial cells was well documented in this swine model. Although similar findings have been reported in a rodent model (9,10), the pig is a widely accepted large-animal model for translational research in cardiovascular processes and pathology. In our swine model, the endothelial cells appeared in two locations within the thrombus: First, as a component of the developing neovascularity within the catheter-related sheath; and second, as a covering layer, located either over the mural thrombus on the vein wall or over the pericatheter thrombus that evolved into the catheter-related sheath.
The process of catheter-related sheath formation is a dynamic and ongoing response of the components of the vein wall to the catheter and associated thrombus. The sequence of the steps of sheath formation is similar among animals and humans, but we would not begin to equate the timing seen in a rodent or swine model with that seen in patients. Inflammatory, endothelial, and smooth muscle cells are involved in this response, and these are all biologically active cell types. Our findings support the hypothesis that a pathologic process occurs when thrombus organizes adjacent to a synthetic scaffold—a catheter. This process differs from intravascular thrombus formation because the presence of the catheter within the vessel lumen allows the process to continue with only limited focal vein wall contact.
Our study had some limitations. Two criticisms could be raised with regard to our results. First, the number of animals in each group was small. The other principal detailed histologic study (10) of the catheter-related sheath was performed in a rodent model, within which research can be more easily conducted with a larger number of animals. Our study was designed to confirm previous observations in small-animal models by using a well-accepted large-animal model of cardiovascular processes. The second limitation of our study was that it is typically difficult to apply results of animal studies to humans. However, the pig is a widely used and accepted model for vascular research. We do not mean to imply that the time course of sheath development would be similar in swine and in humans; however, the sequence of events should be similar.
In conclusion, the sheath that forms around central venous catheters in the swine model consists of cellular and noncellular components; its development is a dynamic process that represents more than simply the accumulation of noncellular material and thrombus.
Practical application: Much of the current interventional vascular research is focused on drug-eluting coatings for stents (30). These coatings consist of cytostatic or cytotoxic agents that target cell populations involved in stent-related restenosis. The characterization of the cellular basis of catheter-related sheath formation may initiate further developments in the area of catheter technologies (31) that could include the development of materials with or without coatings that prevent, retard, or eliminate the sheath.
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ADVANCES IN KNOWLEDGE |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONADVANCES IN KNOWLEDGEReferences |
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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Author contributions: Guarantor of integrity of entire study, A.R.F.; study concepts, all authors; study design, A.R.F.; literature research, A.R.F.; experimental studies, A.R.F., N.L.D.; data acquisition, A.R.F., C.G.A.T.; data analysis/interpretation, A.R.F., N.L.D.; manuscript preparation, A.R.F.; manuscript definition of intellectual content, all authors; manuscript editing, A.R.F., C.G.A.T.; manuscript revision/review and final version approval, all authors
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References |
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  M. J. Oliver, D. C. Mendelssohn, R. R. Quinn, E. P. Richardson, D. K. Rajan, R. A. Pugash, J. A. Hiller, A. J. Kiss, and C. E. Lok Catheter Patency and Function after Catheter Sheath Disruption: A Pilot Study Clin. J. Am. Soc. Nephrol., November 1, 2007; 2(6): 1201 - 1206. [Abstract] [Full Text] [PDF]
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