Method for producing implantable ligament and tendon prostheses and prostheses produced thereby

A method of making implantable ligament and tendon prostheses from natural collagen-containing tissues is described. The tissues used are those wherein the collagen fibers are aligned in one direction such as in ligaments or tendons. The method comprises disrupting the interfibrillar matrix physically by mechanical means such as a roller or rollers in order to generate an expanded network of fibers that are more easily further treated chemically and which also generate a more favorable substrate for tissue ingrowth after implantation. Prostheses made from separated collagen fiber bundles which retain their natural configuration and length (as opposed to reconstituted collagen), exhibit improved softness and flexibility when compared with prostheses made from conventional chemically fixed (e.g., glutaraldehyde cross-linked) tendons which have not been separated as described herein. Another implementation of the developed new technology allows for the formation of composites between the dissociated collagen fibers involving two or more tissues or between collagen and synthetic fibers. These composites can also include absorbable materials.

BACKGROUND OF THE INVENTION 
This invention relates to a method of preparing collagen-containing tissues 
for use as surgically implantable prostheses such as ligament or tendon 
replacements and to the protheses made by such a method. 
Collagen in the form of fibers represents the most abundant animal protein 
in mammals; it accounts for over 30% of all proteins. After being 
manufactured by the cells, the collagen molecules assemble into very 
characteristic fibrils. These fibrils vary in diameter from tissue to 
tissue and with the species and age of the animal. In general they range 
in diameters between 50 and 400 nanometers and are packed into larger 
bundles which we call fibers. Between these fibrils and fibers lies an 
interfibrillar material described in the earlier literature as the "ground 
substance" which we now believe to be composed of a highly organized 
network of negatively charged polysaccharides and associated polypeptides 
referred to as proteoglycans. In some structures, such as in tendons, the 
bundles of collagen fibers are surrounded by a thin connective tissue 
network which has not been well characterized biochemically. This 
composite of collagen and proteoglycans is responsible for the structural 
integrity of the supportive structures of the body such as skin, bone, 
tendons, blood vessels, etc. Cross-links between the collagen molecules 
within the fibrils are a prerequisite for these to be able to withstand 
the physical stresses to which they are subjected. In humans and animals 
these cross-links are generated by a rather complicated series of 
intracellular and extracellular events which generate reactive groups on 
the surface of the collagen molecules. These proceed to form 
intramolecular and intermolecular cross-links after the molecules assemble 
in the extracellular space. 
Not all the collagen molecules in a given organism are chemically 
identical. There is great similarity but nevertheless there are various 
well-defined types of collagen which in some cases are unique to 
particular tissues. For instance, the collagen in cartilage differs from 
that of the cornea, tendon, bone matrix, dermis and most other tissues. 
There are now nine well-defined types of collagen in vertebrates and many 
new ones are being discovered. Whereas in certain tissues collagen fibrils 
are considered to be almost permanent, in others such as bone, which 
undergoes constant remodeling, collagen is constantly being replaced. 
One of the earliest chemical modifications of collagen is associated with 
the process of leather tanning, a technology that has evolved over the 
ages. During the last 15 years increased interest has developed in the use 
of collagen and collagen-containing tissues in the manufacturing of 
medical devices. In some instances, chemical cross-linking with the use of 
bifunctional reagents such as glutaraldehyde, generates materials which 
are readily usable (i.e. pericardial patches). In other cases such as when 
heart valves are manufactured, porcine aortic valves, following chemical 
treatment, are mounted on frames or stents in order to provide them with a 
suitable framework. The stents are often covered with a porous material 
such as woven DACRON.RTM. polyester (DACRON is a trademark of E. I. duPont 
de Nemours and Company, Wilmington, Del.) to facilitate suturing and 
tissue ingrowth leading to attachment. 
In situations such as tendons and ligaments this type of covering is not 
very practical because of the magnitude of the forces imposed upon such 
prostheses. Attaining growth into a DACRON.RTM. polyester sleeve. for 
instance, which is covering a cross-linked tendon-ligament prosthesis as 
it traverses the bone will not assure the attachment of the ligament per 
se. If the prosthesis is partially bio-degradable as is the case of some 
incompletely cross-linked collagen materials, some degradation of the 
prosthesis and growth of tissue will occur, but the continued resorption 
and degradation of the intraarticular portion of the prosthesis will lead 
to eventual failure. 
SUMMARY OF THE INVENTION 
We have therefore designed an approach to facilitate ingrowth and 
attachment of native and cross-linked collagenous materials to host 
tissues. This approach comprises expanding physically the collagenous 
network of the prosthesis prior to implantation to assure the penetration 
of host tissue through its interstices in order to ensure attachment and 
which also provides improved chemical treatment of the tissues. 
In accordance with one object of this invention, a means for dissociating 
the collagen fibrils and fibers present in natural and chemically 
cross-linked tissues is provided that improves the handling properties of 
such tissue (flexibility and elasticity) and enhances the potential for 
tissue ingrowth leading to attachment to the host. 
In the process of the present invention, natural collagen-containing 
tissues such as tendons and ligaments are obtained from animal sources. 
During the processing of such tissues, the protein structure of the 
tissues is covalently cross-linked in one step or step-wise to the degree 
desired to protect the tissue from excessive swelling and other losses of 
physical integrity after implantation in the body of an animal. The 
cross-linking comprises treating the tissue with a cross-linking agent, 
and a permanently implantable prosthesis is constructed from the collagen 
fiber bundles present in the collagen-containing tissues. At some point 
prior to implantation, disruption of the interfibrillar soft connective 
tissues is carried out by a method, such as via the action of a mechanical 
action of a roller under pressure for providing separation of the collagen 
fiber bundles that comprise the collagen-containing tissues while 
retaining substantially all of the natural configuration and length of the 
individual collagen fibers making up the fiber bundles. 
The collagen fibers in native tissues present themselves as a composite of 
closely packed bundles of fibrils embedded in a proteoglycan matrix. This 
composite, particularly after the prior art treatments to render it 
non-biodegradable and antigenic, tends to be resistant to tissue ingrowth 
and host cell proliferation. As a result of the disruption of this 
interfibrillar matrix generated by the procedure of the present invention, 
the collagen fibers become separated in such a way as to generate large 
interfibrillar spaces suitable for tissue ingrowth. In some tissues, such 
as the tendon. the collagen fibers are encapsulated by tight fitting 
connective tissue membranes, as indicated above which in effect form a 
constraining jacket around the fibers. It has been found that 
concomitantly with the separation of the collagen fibers according to the 
present invention, one sees a marked increase in flexibility of the 
prosthesis. Apparently the release of the fibers from their constraining 
jacket provides enhanced flexibility of the cross-linked collagen network. 
The material so treated is now particularly suitable for use in the 
production of composites. For instance, several of these natural tissues 
can be interwoven by weaving or braiding to form larger aggregates of 
increased cross-section. This provides composites of still more enhanced 
elasticity. Another aspect of this fiber expansion and dissociation is 
that it allows the formation of composites between this naturally derived 
material and synthetic fibers. Such composites should enhance the tensile 
strength and allow for new modalities for attachment of the prosthesis to 
bone and soft connective tissues. This modality of treatment also allows 
for the introduction of other materials into the weave. Degradable fibers 
can be introduced and intermingled with the natural or cross-linked 
collagen in such a way that following its resorption, the spaces occupied 
by such a material will now be replaced by host connective tissue or bone 
ingrowth. 
In addition, the network of collagen fibrils, when separated in accordance 
with this invention, becomes more readily accessible to any subsequent 
chemical treatment. Chemicals, in particular cross-linking reagents, 
penetrate slowly through dense connective tissues such as tendons as 
becomes evident under many circumstances where fixation is observed to be 
inadequate or incomplete. This expansion of the fibrillar network prior to 
completion of chemical treatment is particularly useful when the chemicals 
to be used are very reactive, of large molecular weight or are unstable in 
nature. By properly choosing combinations of greatly expanded structure 
with a less expanded structure and combining other fibers with the 
naturally derived material, various heretofore unavailable configurations 
of tendon and ligament prostheses can be obtained. For example, there is 
described hereinafter a prosthesis specifically designed for replacement 
of the cruciate ligament of the knee wherein an expanded portion is 
provided at each end for bone fixation and a relatively smooth center 
portion is provided for passing through the central knee joint space.

DETAILED DESCRIPTION OF THE INVENTION AND PREFERRED EMBODIMENTS 
Exemplary starting materials useful in practicing the invention include 
animal tissues of diverse origin with relatively long lengths of collagen 
fibers in the natural state such as are present in ligaments and tendons, 
which are the preferred starting materials. The tissue used in this 
invention is required to be in such a state that, to the greatest degree 
possible, the natural state and length of the collagen fibers is retained. 
According to this invention, the tissues are preferably cleaned from 
adherent fat and loose connective tissue as soon as possible after 
harvesting, although cleaning may occur after storage or partial 
processing if more convenient. If other treatments according to the 
invention are not to be started immediately the tissues are placed for 
storage in a balanced electrolyte solution that is calcium free and 
buffered at a neutral pH with a buffer, such as sodium phosphate. This 
solution is kept cool (e.g., 4.degree.-8.degree. C. in a preferred 
embodiment). 
Several possibilities now exist for further treatment in accordance with 
the invention. In one embodiment of the invention the tissue is at least 
partially stabilized by chemical treatment in accordance with prior art 
processing techniques such as those described, for example, in Nimni et 
al. U.S. Pat. No. 4,378,224 granted Mar. 29, 1983. The processes described 
in that patent are incorporated herein by reference, but in a preferred 
embodiment of this invention comprise treatment of a bovine tendon in a 
solution containing a conventional cross-linking agent such as a 
dialdehyde. For example, a 0.2 volume percent aqueous glutaraldehyde 
solution buffered with phosphate at pH 7.4 may be used. Penetration of 
glutaraldehyde is enhanced by shaking the solution containing the 
prosthesis. Since this is a slow process, it is allowed to proceed for at 
least 7 days with daily changes of the solution. This causes a partial 
cross-linking of the collagen and the protein-like compounds naturally 
associated with it. 
This tissue is then rinsed with buffered saline (pH 7.4) and extended on a 
hard surface. A roller device such as a wooden dowel or a metal rolling 
pin which exerts a downward pressure is moved over the tendon 
perpendicular to the long axis of the tendon. The amount of pressure used 
is that amount which is sufficient to mechanically disrupt and thereby 
cause the fiber bundles of the tissue to separate without destroying the 
natural configuration and length of the individual collagen fibers. 
Avoiding such destruction is important because the natural state of the 
fiber bundles provides strength to the finished prosthesis. The mechanical 
disruption process is continued until the tendon exhibits a large number 
of longitudinal striations visible to the naked eye which can be distended 
laterally by pulling sidewise. For example, a tendon which was originally 
6 mm in diameter can be flattened to about 20-30 mm in width. Further 
details on the mechanical disruption process will become apparent 
hereinafter. 
The disrupted tissue is placed again in the above-described 0.2% 
glutaraldehyde solution for continued fixation and further treatment in 
accordance with prior art techniques such as those described in the 
aforementioned Nimni. et al. patent. This Nimni, et al. procedure 
incorporates additional covalent cross- links involving the free carboxyl 
groups of collagen using the carbodiimide reaction and an aliphatic 
diamine, preferably hexanediamine. 
Referring to the Drawings, wherein like reference characters designate 
corresponding parts throughout the Figures thereof, FIG. 1 shows the 
manner by which hard roller 18 is used to mechanically disrupt a partially 
cross-linked bovine extensor longus tendon 10 by placing it against hard 
flat surface 19 (e.g., a hard table top) and repeatedly drawing roller 18 
back and forth in the direction shown by arrows 17 over the tendon 10 and 
produce portion 12 composed of separated collagen fiber bundles, one of 
the many present being indicated by reference numeral 16. FIG. 1 further 
shows the result of distending portion 12 laterally and exerting gentle 
tension to expose the criss-crossed network of collagen fiber bundles 14 
shown which reflects the intricate wavy zig-zag pattern of the native 
fibrillar structure. After mechanical disrupting, the tendon is further 
processed chemically as described above. The resulting prosthesis material 
obtained is characterized by intrafibrillar spaces and by a softer, more 
flexible nature than the stiff prostheses which are obtained by chemically 
treating tendons in the same manner, but without including the step of 
mechanically disrupting the tissue as described herein. 
FIG. 2 depicts an alternative embodiment of the present invention showing 
the effect of mechanically disrupting tendon 20 as described for FIG. 1 
above using hard roller 28 in the direction of arrow 27 on hard surface 
29. In this embodiment, however, the tendon 20 is used after cleaning but 
before any further treatment such as fixation by cross-linking is 
employed. The result shown at 22 is a flattening of the tendon into a 
compact belt-shaped configuration 22 without as much separation between 
fibers as can be seen in FIG. 1. The tissue is then fixed in this 
configuration by chemical processing as has been described above. The 
mechanical disruption process can also be applied to fully treated 
collagenous materials while they are in the wet state to obtain softer, 
more flexible prostheses although this is a less preferred process since 
the earlier use of the mechanical disruption process exposes more fiber 
bundles to the chemicals employed to treat the collagenous materials. 
Applying the mechanical disruption process prior to completion of the 
chemical treatment tends to eliminate the possibility of untreated areas 
in the collagenous materials thereby reducing any effects that such 
untreated portions might have on the performance of the prosthesis after 
implantation. As a further alternative, and providing still greater 
assurance of full chemical treatment a first disruption step can be 
performed, followed by partial chemical treatment and then a further 
disruption step can be performed before final chemical treatment. 
As an alternative to using a single roller against a flat surface, an 
apparatus composed to two rollers arranged similarly to those of a 
conventional washing machine roller ringer or a two roll mill where the 
spacing between the two rollers is adjustable can be used to cause 
mechanical disruption of the collagen fiber bundles in the tendons. One 
advantage of using such an apparatus is the fact that the some control can 
be exerted over the degree of pressure exerted against the tendons by 
adjusting the roller spacing. The rollers in either case can be made, for 
example, of hard rubber or stainless steel. 
Short, rocking, rolling strokes appear to work better in a mechanical 
disruption process utilizing a roller than do long continuous strokes. As 
the diameter of the roller or rollers used is increased, more force 
against the tendons is needed to achieve the same degree of disruption 
obtained with smaller diameter rollers. 
After the mechanical flattening procedure, the collagen fiber bundles 
within the tendon may be pulled apart to further increase compliance or to 
allow separation of long individual fiber bundles. After the mechanical 
disruption procedure and the fiber separation procedure, the tendons can 
be reshaped into their natural configuration by gently tugging at the 
ends. 
FIG. 3 depicts the result of braiding masses 32, 34 and 36 of collagen 
fiber bundles treated in accordance with the process of the present 
invention to produce a prosthesis 30 which can then be used as, for 
example, a tendon replacement prosthesis. More than just two masses of 
treated collagen fiber bundles can be braided. It is also possible to 
intertwine synthetic polymeric fibers such as DACRON polyester and other 
biocompatible polymers such as polytetrafluoroethylene, reconstituted 
collagen fibers (treated or untreated to reduce reabsorption), polylactic 
acid polymer fibers and the like in with the treated collagen fiber 
bundles to create composite braided products which have the properties of 
both types of fibers. Braiding is employed to enhance the physical 
properties of the prosthesis and to control the diameter of the prosthesis 
itself. 
FIG. 4 depicts a prosthesis which is composed of two mechanically disrupted 
and chemically treated (in accordance with the present invention) bovine 
tendons 50, 60 which have been partially expanded into two braided 
portions 40 and 40' where portion 40 is made by braiding masses 51, 52, 
61, and 62 of treated collagen fiber bundles and portion 40' is similarly 
made by braiding together masses 53, 54, 63 and 64 of treated collagen 
fiber bundles where the central portions of tendons 50 and 60 are left 
unbraided. 
The processed collagenous materials are then employed in manners known to 
those skilled in the art of making prostheses from treated collagenous 
materials to manufacture tendon and ligament prostheses. The embodiment 
illustrated in FIG. 4 could be used to replace the human cruciate ligament 
where the opposite ends of the prostheses are stapled or otherwise fixed 
to the bone and the central portion 50, 60 passes through the central knee 
joint space. 
Although presently preferred embodiments of the invention have been 
described above, modifications thereof will become apparent to those 
skilled in the art. Therefore it is to be understood that within the scope 
of the appended claims the invention may be practiced otherwise than as 
specifically described.