Pathology of bioprosthetic cardiac valves

Pathology of Bioprosthetic Cardiac Valves. VICTOR J. FERRANS, MD, PHD,* YOSHIFUMI TOMITA, MD, PHD,t STEPHEN L. HILBERT, PHD,+ MICHAEL JONES, MD,t AND WILLIAM C. ROBERTS, MD* .A description of the normal and abnormal anatomic features of bioprosthetic cardiac valves (BP) is ~rese~ted in this article, .with emphasis on cuspal calclficat~on. and perforatlo~, which are their major c~mphcatIons. The following types of bioprosthetic (tissue) heart valves are considered: porcine aortic valvular b~oprostheses (~erein referred to as porcine BPs); bovme and porcme parietal pericardial BPs; human dura mater valves; fascia lata valves and human aortic homografts. '

Abnormal Angulation

Angulation of a BP can result from insertion and suturing of the valve away from the usual axis. This c~n ~ccur ~vhen the anulus is distorted· by calcificatwn, mfectlon, dense scarring. Angulation also can result from partIal detachment of sutures (i.e., because of infection associated with a ring abscess).

01:

Disproportion and Hemodynamic Obstruction

ABNORMAlITIES EVIDENT ON IN SITU EXAMINATION OF THE BIOPROSTHESIS

. A~normali.ties.that are best (or only) diagnosed by m SItu exa~lnatlon of the BP include: perivalvular leak; obstructIon to the left ventricular outflow tract o~ to a co~onary ostium; entanglement of sutures: dIsproportIon; abnormal angulation of the BP; and compression of an anomalous left main or left circumflex coronary artery by the fixation rings of one or more BPs. Perivalvular leak (Para-anular Prosthetic Ring Discontinuity)

A peribasilar leak may result from the use of too few sutures to secure the BP and from inadequate placement or from pulling loose of sutures between t~e patie~t's .tissues and the bioprosthetic sewing rmg. 1 Penbasllar leaks tend to occur in instances in which the sutures are placed in tissues that are abnormal because of infection, extensive calcific deposits, or preexisting connective tissue disorders such as Marfan's syndrome or valvular prolaps~ (floppy valves). However, they also can develop in patients who have papillary muscle dysfunction and normal perivalvular tissues. l Residual glutaraldehyde may remain in tissue valves after prolonged storage, even after careful preimplantation washing of the BP.2 It has been suggested that it exerts toxic effects including induction of necrosis of the host tissues of the valve ring, which are directly in contact with the .sewing ring; such a necrosis could evolve into a perivalvular leak. . . Received from lhe *Palhology and tSurgery Branches, NalIonal Hearl, Lung, and Blood Inslilule, Nalional Instilules of Heahh, Belhesda, and lhe tCenter for Devices and Radiological Heahh, Food and Drug Adminislralion, Rockville, Maryland. . . A.ddress correspondence and reprint requesls lO Dr. Ferrans: Bulldmg 10, Room 7N-236, Nalional Instilules of Heahh, Belhesda, 1\10 20892. 0046-8177/87 SO.OO + .25

Nearly all bioprosthetic cardiac valves present some degree of obstruction to forward flow, and some degree (usually minimal) or regurgitant flow. The central.flow pattern of bioprosthetic valves generally provldes .good hemodynamic function, but ~alves of small sIzes may cause significant reduction m flow because the sewing ring and struts may occupy a larger fraction of the total orifice area of the BP than is the case in larger-sized BPs. Disproportion occurs when ~he substitute .cardiac valve is too large for the ventncle or ascending aorta into which it is inserted. Obstruction to a coronary ostium can result from mall?ositio~~ng of the stent in a BP implanted in the aortic posItion. I The left ventricular outflow tract may be obstructed by a strut of a BP that has not been placed properly in the mitral position;I,3 lowprofile BPs are less likely to cause this complication. Ventricular Rupture

Ventricular rupture, which can be located either at the l.e~'el of the atrioventricular groove, at the site o~ exclswn of one or both papillary muscles, or midway. between the ~itral ring and the stumps of the papIllary muscles, IS a major complication of mitral valvular replacement with any kind of prosthetic valve. 4 STRUCTURE OF UNIMPLANTED BIOPROSTHESES Bef~re. describing the .p~thologic changes that develop m Implanted BPs, It IS convenient to review briefly the morphologic features of normal, unimplanted BPs. Porcine BPs, pericardial BPs, fascia lata valves, an? dura mater valves have all been config~red .as tnl.eaflet valves, regardless of the valve posit~on m wh~ch they are implanted. The only exceptwns to thIS generalization are the pericardial unicusp or unileaflet valve 5 and the bicuspid pericardial valve 6 ; neither has been used extensively. Most BPs u.ndergo preimplantation chemical treatment: porcine BPs and pericardial BPs with 0.25 to 0.6 per cent

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glutaraldehyde, and dura mater valves with glycerol. Homografts are sterilized, usually with antibiotics (several other methods were found to be less satisfactory), but are not treated with al~ehyde fixatives, ~nd fascia lata valves were used wahout any chemical treatment. Some types of glutaraldehyde-treated BPs are stored in dilute formalin after processing, to improve sterility, because the antifu.ngal and antibac. terial (especially against spores) actIOn of low concentrations of glutaraldehyde is po~r.7,8 In gene.ral, the durability of unfixed and formalu~-fix~d po.rcme and pericardial BPs is poor. 9 - IO AnticalclficatlOn treatments have been devised to mitigate the problem of calcific deposits developing in BPs. These include the surfactant processes (T6 and PV2). These processes do not appear to alter the morphology of unimplanted BPs significantly. Clinical trials of valves pr~­ pared according to these processes are currently m progress. In the sheep model, b~th I;>rocess~s are e~­ fective in mitigating the calcificatIon m porcme aortic . valvular BPs, but not in pericardial BPs. The stents on which the bioprosthetic tissue is mounted vary in structural design, flexibility, and composition (plastic and metal) from one manufacturer to another, and among different models o~ the same BP, in which the stents may have been modified (e.g., to make them lower in profile); different ,stents may be used for the sa~e type o~ valve ~ccordmg to whether it is to be used m the atnoventncular or the aortic valve position. Similarly, the techniques by which the tissues are attached to the stents vary, particularly among pericardial BPs from different manufacturers (in efforts to minimize chances of cuspal disruption near the stent posts). Porcine Aortic Valvular Bioprostheses Glutaraldehyde-treated porcine BPs are the most widely used type of BP a!1d consist of a I;>0rcine aortic valve and adjacent regIOn of the aortic root. The porcine valve is asymmetric. The ri.ght coronary cusp is larger than the other two cusps; Its most basal region contains a layer of ventricular muscle c~lls that represent an extension of septal muscle and Impar~ a darker color a':ld a firmer co~;istency to th,i,s regIOn of the cusp. ThiS muscle layer ( mu.scle shelf ) sometimes is also found in the basal regIOns of the other two cusps. It can cause delayed and less complete opening of the right corona.ry cusp (comI;>ared with the other two cusps) of porcme BPs, and a can become the site of calcific deposits involving the myocytes. For these rea.sons,. a "mo~ified orifice valve" has been designed m which the nght coronary cusp is removed and replaced ~y ~ cusp from another valve; this BP has had very hmaed use. Other attempts have been made to eliminate part of this muscle shelf, either by curetting from the external surface before the valve is mounted on its stent or by tucking it into the sewing ring duri':lg mou~ting. Th.e structure of the unimplanted porcme aortic valve IS illustrated in figures lA to C.

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Pericardia I Bioprostheses Practically all pericardial BPs are I~ade of bovine parietal pericardium; the only exception appears to be the Polystan valves, which are made of porcine parietal pericardium and are ~sed a~ I?arts of va~ved conduits that also have an mner hnmg of panetal pericardium. II The thic.kness ~f the cusps ~s greate.r in pericardial BPs than m porcme BPs. Panetal 'per~­ cardium l2 ,13 is composed of the serosal layer, .whlCh IS formed by mesothelial cells; the fibrosa, which contains collagen, elastic fibers,. ne~ves, ?lood vess<:ls, and lymphatics; and the eplpencardlal connective tissue which has more loosely arranged collagen and elasti~ fibers. The collagen fibrils in pa~ietal perica~­ dium form relatively short bundles, which ~re mult~­ directional and overlapping rather than lughly onented and welllayered. 14 The stn~ct~r~ of the uni~­ planted pericardial bioprosthesls IS Illustrated m figures ID and E. Dura Mater Valves Normal human dura mater is composed of two distinct layers: an outer (or endosteal) layer and an inner (or meningeal) layer. IS The outer layer co~­ tains large bundles of collagenous fibers and constitutes about two thirds of the total thickness of the dura. About 50 per cent of the bundles in this layer appear oriented in the same direction; the other 5.0 per cent are multidirectional. The inner layer IS thinner and has smaller bundles of collagen, many of which are arranged irregularly, often obliquely or at right angles to the main direction of orientation of the bundles in the outer layer. In some areas the two layers of the dura are closely apposed; in others they are separated by spaces of variable width. Blood vessels are present in some of these spaces. In both layers the collagen is wavy, elastic fibers are few and small, and proteoglycan content (judged by ~taining with alcian blue) is low. Cellular elements m dura mater are scarce and consist of elongated fibroblasts with spindle-shaped nuclei and scanty cytoplasm. These valves have been used extensively in Latin America 16. in a clinical trial in England they were frequently'found to develop cuspal tears.I' Their use has decreased sharply in recent years. Fascia Lata Valves Fascia lata valves constructed of three cusps of autologous fascia lata mounted on a s.tent ,~ere used 20 years ago, but their use has been discontInued because of poor results due to overgrowth of fibrous sheath of host origin and disintegration of cuspal connective tissue. I8 - 20 Homografts Human aortic valve homografts consist of a sleeve of tissue containing the aortic valve and a portion of ascending aorta; this sleeve is trimmed just below the cuspal attachments and is scalloped along

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PATHOLOGY OF BIOPROSTHETIC CARDIAC VALVES (Ferrans et al.)

FIGURE 1. Structure of unimplanted porcine (A through C) and pericardial (D and E) bioprosthetic cardiac valves (BPs). A, Histologic section of cusp of porcine BP. showing inflow surface at bottom and outflow surface at top. Note the ventricularis (V), the spongiosa (S). and the fibrosa (F). (Hematoxylin-eosin stain. x150.) B, Basal portion of right coronary cusp, showing the darkly stained muscle shelf (arrowheads). (Toluidine blue stain. x 25.) C Collagenous cords in cusp are sectioned transversely and appear as focal thickenings of the fibrosa (compare with A). (Hematoxylin-eosin stain. x 100.) D. Section of cusp of bovine pericardial BP. showing the inflow surface (the epipericardial layer) at bottom and the outflow surface (serosal layer) at top. The layers of connective tissue are poorly defined. (Hematoxylin-eosin stain. x 100.) E. Polarized light micrograph of section taken parallel to the surface of cusp of pericardial BP, showing multidirectional pattern of birefringence of collagen fibrils. (Hematoxylin-eosin stain. x 100.)

'" its upper border into anchoring projections above the cusp commissures. 21 - 23

Attachment of Blood Cells to the Surfaces of Bioprostheses

POSTIMPLANTATION CHANGES IN BIOPROSTHESES

Many pathologic changes that occur in BPs implanted as substitute cardiac valves are common to all types of these devices, reflecting three basic facts: (1) collagen is the most important structural element in these valves; (2) in BPs there is no synthetic or renewal mechanism (such as that provided by the fibroblasts present in native valves) to replace the collagen that is gradually broken down, either because of proteolysis (although glutaraldehyde-treated collagen is very resistant to the effects of proteolytic enzymes) or because of the mechanical stresses to which the cusps are subjected-thus, the cumulative effect of this gradual breakdown can lead to the formation of cuspal tears and perforations; and (3) collagen in implanted BPs tends to undergo a time-dependent process of focal calcification, which may result in stenosis and/or regurgitation and loss of mobility of the cuSpS.9,24-28

Within a few days after implantation, BPs become covered with a layer of fibrin, platelets, and a few erythrocytes; inflammatory cells usually are relatively few and consist mainly of macrophages and multinucleated giant cells; lymphocytes and plasma cells are scarce, as are neutrophils (fig. 2).9 The presence of neutrophils in implanted BPs is highly suggestive of infection. Plasma cells were relatively numerous in the BPs of children who were considered to have an immunologic reaction to the BPs29; such cases, however, are rare and there is little evidence to suggest that rejection or an immunologic reaction is of importance in the pathogenesis of calcification or other postimplantation changes. 3o Most leukocytes and macrophages remain confined to the surfaces, with little tendency to infiltrate the bioprosthetic tissue. 9 Clusters of lymphocytes, macrophages, and a few plasma cells are sometimes found in connective tissue of host origin at the junction between valve tissue and the sewing ring, particularly in the right coronary cusp of porcine BPs. l\lultinucleated giant cells also may be numerous in these areas, where they may be associated with suture materials.

FIGURE 2. A, Macrophages and multinucleated giant cells line the surface of a bioprosthetic cardiac valve (BP) implanted in a sheep for 20 weeks. (Hematoxylin-eosin stain. x 250.) B. Scanning electron micrograph showing numerous platelets on the surface of a BP removed because of dysfunction after being implanted for 76 months in the mitral position. (x 2.800.)

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The overall incidence of infection is generally similar in patients with mechanical vah'es and in patients with BPs. Infection of implanted BPs is covered in the article on infective endocarditis. Cuspal Tears and Perforations

FIGURE 3. Classification of cuspal lesions in bioprosthetic valves. Type I lesions are tears that involve the free edge of the cusp. Type II are linear perforations along the base of the cusp. Type III lesions are larger perforations in the center of the cusp. Type N are small. often multiple perforations that appear as pinholes and usually are associated with calcific deposits.

In addition to the changes just described, plasma proteins penetrate into the substance of the BPs because of the absence of an endothelial cell barrier, and the connective tissue structure becomes gradually endothelialized, particularly in the basal regions of the cusps, and also may become covered with a fibrous sheath of host origin. This process may take from several months to several years. 31 Aggregates of platelets also may develop on nonendothelialized regions of the surfaces, where they are in direct contact with collagen fibrils, which form part of the surfacesY·32 This finding is not surprising in view of the tendency of collagen to induce platelet aggregation. This tendency is minimized by preimplantation treatment of BPs with glutaraldehyde. 32 Activation, spreading, and hyperactive responses of platelets as well as circulating aggregates of platelets have been reported in some patients with BPs-particularly in those with degenerated BPs32; however, the clinical significance of these findings remains uncertain. Thrombi in Bioprostheses and Embolic Phenomena

Although the lack of need for the use of anticoagulants is often cited as one of the advantageous features of BPs, compared with mechanical cardiac valves, thrombi and thromboembolic episodes do occur in patients with implanted BPs, even in those receiving anticoagulants, but less frequently than in patients with mechanical valves.16.24.25.28 Thrombi in BPs may occur as early or late postimplantation phenomena. They tend to be localized on the outflow surfaces of the cusps and may be associated with calcific deposits. Bioprosthetic Infection

Infection of an implanted BP continues to be a major complication of cardiac valvular replacement.

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The connective tissue components of all BPs are subjected to chemical and degradative processes that place certain limits on their durability. The most important of these processes seems to be the mechanical disruption of collagen fibrils, which occurs as a consequence of the flexing and bending that occur with each cycle of valvular opening and closing. The contribution of proteolytic enzymes in blood cells and plasma to this process of "collagen degeneration" is unclear. As mentioned previously, such phenomena are accentuated at certain sites in which mechanical stresses are high, thus causing focal breakdown of the connective tissue structure and giving rise to tears and perforations. Ishihara et al. 33 have presented a system (figs. 3 and 4) for the classification of cuspal lesions in bioprosthetic valves. The collagen affected by these changes has a pale, hyaline appearance in histologic sections stained either with hematoxylineosin or with toluidine blue, and on transmission electron microscopic study the collagen fibrils have frayed tips, are decreased in number, and are associated with large amounts of finely granular material. 9 Examination by polarized light microscopy shows a marked decrease in the birefringence of collagen in such areas. For reasons that are unclear, collagen degradation is rapid in fascia lata valves l8 - 20 but slow in aortic homografts. 21 - 23 Type I lesions are most frequent in the cusps of pericardiap1.35 and dura mater 17 valves. They have been recently reported to be a relatively common complication in patients undergoing mitral valvular replacement with Ionescu-Shiley pericardial BPs,34.35 but they also have been encountered with other pericardial BPs of more recent design. In both pericardial and dura mater valves, these lesions (flail cusp) tend to occur near the stent post rather than along the central region of the free edge. They have a similar preferential location in porcine BPs. Type II lesions are thought to be related to the phenomenon called "hinging," in which the cusps do not open with an even motion, but tend to open by bending excessively at one localized site near the cuspal base. The.cumulative effects of such a repeated hinging are believed to lead to separation of bundles of collagen and to the formation of a linear defect along the base of the cusp.33 Type III lesions usually result Jrom infection but also can be produced by the confluence of several type IV lesions. Type III lesions caused by infection have shown extensive destruction and necrosis of cuspal tissue at their edges. Type IV lesions usually occur in the vicinity of calcific deposits, and are thought to result from mechanical forces acting on the collagen at the edges of such deposits. 33 .36

PATHOLOGY OF BIOPROSTHETlC CAADIAC VALVES (Ferrans et alJ

FIGURE 4. Cuspal tears and perforations. A. Type I lesion is a tear located near the commissure of a porcine bioprosthetic cardiac valve (BP) Implanted In a sheep. lesion has developed near the tree edge (which appears very thin but not yet torn) in the region of a

a

commissure of a porcine BP implanted in a sheep. C. Type II lesion has developed in the shape of an arc along the basal portion of a cusp of a porcine BP implanted in a sheep. D, Scanning electron micrograph of type N lesion in a dura mater valve. (x 50,) E. Delamination of cusp of porcine BP is characterized by separation of the connective tissue layers. Compare with his1ology of unlmplanted BP in figure 1. (Hematoxylil'l-eosin stain. x 100.) F, Electron micrograph of collagen fibrils undergoing disruption into finely granular material in BP removed because of tears 76 months after implantation in the mitral position. (lead citrate and uranyl acetate stain. x 22.000.)

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fIGURE 5. Histologic section of basal portion of cusp of porcine bioprosthetic cardiac valve showing fibrous sheath that has grown extensively (bottom)on the inflow surface and to a much lesser extent (upperright)on the outflow surface. (Hematoxylin-eosin stain. x 100.)

Suture-Induced Perforations

In addition to the four types of perforations described previously, suture-induced abrasions and perforations have been reported 37 in BPs implanted both in the mitral and in the aortic vah'e position, in instances in which the sutures anchoring and sewing ring had excessively long ends. These long ends may make contact with the bioprosthetic cusps and eventually erode them, causing cuspal perforations. These lesions have been produced by monofilament and by braided types of sutures, and they have been induced experimentally in sheep with BPs.37

as "neointima") also is observed in valved right ventricle-pulmonary artery conduits, in which it can involve the bioprosthetic cusps, which become adherent to the inner wall of the conduit, causing both stenosis and regurgitation. 39 Calcification

Fibrous Sheathing

Implanted BPs gradually become partially or covered by a fibrous sheath of host origin. 1 This fibrous sheath appears to 'grow in from the region of the stent and spreads over both of the surfaces of cusps (fig. 5). The collagen in fibrous sheaths usually is easily distinguished from that in the bioprosthetic cusps because it is less eosinophilic; the nuclei of the host connective tissue cells are much more basophilic than those in the cusp itself (fig. 5). The thickness of the fibrous sheath can be equal to, or greater than, that of the cusp itself. In some instances metaplastic cartilage and even bone develop at the border between fibrous sheath and bioprosthetic CUSp.38 Fibrous sheaths appear to have some protective effect in reinforcing the cuspal structure; however, in some instances, they can grow excessively and cause cuspal stiffening and even commissural fusion. 24 ,25,31 Thus, they can produce significant cuspal stenosis. Similar fibrous sheathing occurs in dura mater valves,16,17 homografts,21-23 and fascia lata valves. 18 - 20 Fibrous sheathing (sometimes referred to com~letely

592

Calcific deposits develop in most fa0rcine BPs implanted for three years or longer,9,24- 8 and can be the cause of stenosis because they can stiffen the cusps considerably and reduce their mobility. They also can be the cause of regurgitation, because they often are associated with cuspal tears and perforations. In all types of BPs, the most significant calcific deposits develop in association with collagen fibrils (intrinsic calcification) and with organizing surface microthrombi (extrinsic calcification). The gross and microscopic changes associated with calcification are illustrated in figure 6. Factors known to predispose to the calcification of BPs are: young age of the patient, chronic renal disease, other causes of hyperparathyroidism, and bioprosthetic infection. 4o Both porcine and pericardial BPs implanted in children may undergo rapid, clinically severe calcification within three years after implantation (see reference 40 for review). There is considerable, unexplained variation in the extent to . which calcific deposits develop in BPs implanted in adults, in whom such deposits usually do not become clinically significant until four years or longer after implantation. Calcific deposits are the single most important cause of dysfunction of implanted BPs in adults. The pathogenesis of calcific deposits is poorly understood. 4o Subcutaneous models of valve implantations have provided useful insights into this process. 41 .42

PAlHOLQGY OF BIOPROSlHETlC CARDIAC VALVES (Ferrans et 01.)

FIGURE 6. Calcific deposits in bioprosthetic cardiac valves (BPs). A, Radiograph of valve demonstrates mUltiple calcific nodules. 8. Dissecting microscopic view of multiple calcific deposits near a commissure of a porcine BP. (x 30.) C, Calcific deposit in porcine BP implanted for 20 weeks in a sheep. (Hematoxylin-eosin stain. x 150.) D, Calcific deposits in pericardiaI BP implanted for 20 weeks in a sheep. This region of the cusp is thickened and delaminated. (Hematoxylin-eosin stain. x 150.) E and F, Ultrastructure of calcific deposits associated with collagen. Deposits in Einvolve the entire thickness of the collagen fibrils. which have been sectioned transversely; deposits in Fare confined to the interfibrillary spaces so that the collagen fibrils appear lucent. (Uranyl acetate and lead citrate stain. x 54.000 and x 56.000.)

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FIGURE 7. Intracuspol hematoma in a porcine bioprosthetic cardiac valve (BP). A, View of the explanted BP, showing dark discoloration of

basal regions of the cusps. B, Histologic examination shows penetration of blood (arrowheads) Into spongiosa of the cusp, which has become thickened. (J-Iematoxylin-eosin stain. x 15,)

REFERENCES

Intracuspal Hematomas and Bending of the Struts

Intracuspal hematomas and bending of the struts are infrequent causes of stenosis of porcine aortic valvular BP. Intracuspal hematomas43 are due to the penetration of red blood cells between the layers of the cusps, especially in the spongiosa, leading to an increase in cuspal thickness and stiffness (fig. 7). The presence of blood imparts a characteristic dark color to the affected cusps. It has been suggested that the blood penetrates between the bioprosthetic cusps and the sewing ring. 43 Progressive inward bending of the polypropylene struts ("stem creep") of porcine BP may cause bioprosthetic stenosis. 44 Infiltration of Lipid Material and Amyloid

Infiltration of lipid material, presumably derived from blood plasma, has been reported to occur in extraordinary amounts in some implanted BPs. In some,45,46 large crystals of cholesteryl esters have been demonstrated, and in others,9 lipid infiltration has been very marked in areas adjacent to type I cuspal tears. The true prevalence of such deposits is uncertain. Goffin et al.4' found microscopic deposits of amyloid in the sewing ring of one fascia lata valve and in cuspal tissue in 10 of 30 porcine BPs. All amyloid-containing porcine BPs had been in place for at least 33 months, and all except two showed dysfunction or severe degeneration of cuspal tissue. In the majority of the valves, the amyloid deposits were permanganate sensitive and tryptophan positive. The pathogenesis and significance of these deposits are unknown.

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I. Roberts WC, Ferrans VJ: Complications of replacement of either the mitral or aortic \'al\e or both by either mechanical or bioprosthetic vah'es. In Cohn LH, Gallucci V (eds): Cardiac Bioprostheses. r\ew York, Yorke, 1982, pp 331-345 2. Gendler E, Gendler S, r\imni ME: Toxic reactions evoked by glutaraldeh}'de-fixed pericardium and cardiac vah'e tissue bioprostheses. J Biomed Mater Res 18:727, 1984 3. Roberts WC: Complications of cardiac val\'e replacement: characteristic abnormalities of prostheses pertaining to any or specific site. Am HeartJ 103:113, 1982 4. Roberts WC, Isner JM, Virmani R: Left ventricular incision midway between the mitral anulus and the stumps of the papillary muscles during mitral \'ah'e excision with or without rupture or aneurysmal formation: analysis of 10 necropsy patients. Am Heart J 104:1278, 1983 5. Gabbay S, Frater RW: The unileaflet heart \'ah'e bioprosthesis: new concept. In Cohn LH, Gallucci V (eds): Cardiac Bioprostheses. r\ew York, Yorke, 1982, pp 411-424 6. Bodnar E, Bowden r\L, Drury PJ, et al: Bicuspid mitral bioprosthesis. Thorax 36:45, 1981 7. Center for Disease Control: Isolation of m)'cobacteria species from porcine heart \al\'e prostheses. MMWR 26:42, 1977 8. Laskowski LF, Marr JJ, Spernoga JF, et al: Fastidious m)'cobacteria grown from porcine prosthetic-heart-\,ah'e cultures. N Engl J Med 197:101, 1977 9. Ferrans VJ, Spray TL, Billingham ME, et al: Structural changes in glutaraldeh}'de-treated porcine heterografts used as substitute cardiac valves. Transmission and scanning electron microscopic observations in 12 patients. AmJ CardioI41:1159, 1978 10. Sade RM, Greene WB, Kurtz S~I: Structural changes in a porcine xenograft after implantation for 105 months. AmJ Cardio144:761, 1979 II. Pol)'stan Bioprostheses Information Bulletins BPA, BPB, BPC, BPD, BPE, PBF, and BPI. Copenhagen, Denmark, Polptan NS, 1980 12. Ishihara T, Ferrans VJ,Jones M, et al: Structure ofbo\ine parietal pericardium and of unimplanted lonescu-Shiley pericardial \ah'ular bioprostheses. J Thorac Cardio\asc Surg 81 :747, 1981 13. Allen DJ, DiDio LJ, Zacharias A, et al: Microscopic study of normal parietal pericardium and unimplanted Puig.Zerbini pericardial valvular heterografts.J Thorac Cardio\asc Surg 87:845,1984 14. Hilbert SL, Ferrans VJ, Swanson WM: Optical methods for the non-destructive C\'aluation of collagen morphology in bioprosthetic heart \alves.J Biomed Mat Res 20:1411,1986 15. Allen DJ, DiDio LJA: Scanning and transmission electron microscopy of the encephalic meninges in dogs.J Submicrosc C)'toI9:1, 1977 16. Puig LG, Verginelli G, Ir)'ia K, et al: Homologous dura mater cardiac \ah'es. Study of 533 surgical cases. J Thorac Cardio\asc Surg 69:722, 1975

PATHOLOGY OF BIOPROSTHETIC CARDIAC VALVES (Ferrans et 01.) 17. Martelli V. Wain WHo Bodnar E. et al: Mitral valve replacement with dura mater bioprosthesis. ArtifOrgans 4:168.1980 18. Willen R. Willen H. Dubiel WT. et al: Composition of surface layers in unfixed autologous fascia lata heart valve grafts. A transmission electron microscopal study. UpsalaJ Med Sci 88:25. 1983 19. Silver MD. Trimble AS: Structure of autologous fascia lata heart valve prosthesis. Arch Pathol Lab Med 93:109.1972 20. Olsen EG, AI-Janabi N. Salamao CS. et al: Fascia lata valves: clinicopathological study. Thorax 30:528. 1975 21. Reichenbach DO. Mohri H, Merendino KA: Pathological changes in human aortic valve homografts. Circulation 39.40(suppl 1):1-47.1969 22. Aparicio SR. Donnelly RJ. Dexter F, et al: Light and electron microscopy studies on homograft and heterograft heart valves. J Pathol 115:147,1975 23. Barratt-Bo)'es BG. Roche HG. Brandt PWT, et al: Aortic homograft valve replacement: a long-term follow-up of an initial series of 101 patients. Circulation 40:763. 1969 24. Spray TL. Roberts WC: Structural changes in porcine xenografts used as substitute cardiac valves. Gross and histologic observations in 51 glutaraldehyde-preserved Hancock valves in 41 patients. Am J CardioI40:319. 1977 25. Fishbein MC, Gissen SA. Collins JJ Jr, et al: Pathologic findings after cardiac valve replacement with glutaraldehyde-fixed porcine valves. AmJ CardioI40:331, 1977 26. Cipriano PRo Billingham ME. Oyer PE. et al: Calcification of porcine prosthetic heart valves: a radiographic and light microscopic study. Circulation 66: 1100, 1982 27. Ferrans VJ. Boyce SW, Billingham ME, etal: Calcific deposits in porcine bioprostheses. Structure and pathogenesis. Am J Cardiol 46:721, 1980 28. Schoen FJ, Hobson CE: Anatomic analysis of removed prosthetic heart valves: causes of failure in 33 mechanical valves and 58 bioprostheses. 1980 to 1983. HUM PATHOL 16:549, 1985 29. Rocchini AP. Weesner KM. Heidelberger K. et al. Porcine xenograft valve failure in children: an immunologic response. Circulation 64(suppl 11): 11-162, 1981 30. Levy RJ. Schoen FJ. Howard SL: Mechanism of calcification of porcine bioprosthetic aortic valve cusps: role of T lymphocytes. Am J Cardiol 52:629, 1983 31. Ishihara T, Ferrans VJ. Jones M. et al: Occurrence and significance of endothelial cells in implanted porcine bioprosthetic valves. AmJ Cardiol 48:443. 1981 32. MagiIligan OJ Jr. Oyama C. Klein S. et al: Platelet adherence to bioprosthetic cardiac valves. AmJ CardioI53:945. 1984 33. Ishihara T, Ferrans VJ, Boyce SW, et al: Structure and classification of

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