Porous orbital implant structure

A porous structure for implantation into the orbital cavity of a mammal who has had an ocular enucleation, evisceration or who needs to have an orbital implant replaced, the structure comprising pores having a mean size of less than 200 micrometers. Also disclosed is a surgical method for placing an implant into a mammal who has had an ocular enucleation, evisceration or who needs implant replacement, whereby the implant obtains rapid ingrowth of connective and vascular tissues. The method comprises: selecting a porous ocular implant comprising pores with a mean size of less than 200 micrometers; and, placing the implant into an orbital cavity of a mammal.

FIELD OF THE INVENTION 
This invention concerns biocompatible compositions of matter. More 
particularly, it concerns an ocular implant capable of rapid and effective 
fibrovascular integration of vascular and/or connective tissues following 
implantation into the orbital cavity of an anophthalmic mammal. 
BACKGROUND ART 
Since at least 1884, surgeons have sought a means to improve the cosmetic 
rehabilitation of the anophthalmic patient. Accordingly, improvements have 
been sought by increasing the support of the artificial eye and by 
attempts to transfer all latent muscle movement directly to the artificial 
eye via some form of direct coupling between the eye and the implant. 
(Ruedemann A. D., "Plastic Eye Implant" Amer J Ophthalmol (1946) 
29:947-952). 
Ocular implants are used to replace the volume lost after enucleation or 
evisceration to improve the cosmetic psychological and rehabilitation of 
the anophthalmic patient. Many materials have been used for this purpose, 
starting with Mules's hollow glass spheres in 1884. (Mules, P. H., 
"Evisceration of the Globe, with Artificial Vitreous" Trans Ophthalmol Soc 
UK (1885) 5:200-206) Mules employed a hollow glass sphere; this sphere 
offered some support for the upper eyelid but was unable to relieve the 
chronic downward pressure on the lower lid. (Id.) It is necessary to avoid 
chronic downward pressure on the lower lid to alleviate the lid sag 
characteristic of long-term anophthalmic patients. (Ruedemann A. D., 
"Plastic Eye Implant" Amer J Ophthalmol (1946) 29:947-952, Durham D. G., 
"The New Ocular Implants" Am J Ophthalmol (1949) 32:79-89). 
Numerous implant innovations followed Mules' implant, including implants 
composed of: gold, cartilage, fat, fascia lata, bone, xenogeneic animal 
eyes, silver, Vitallium, platinum, aluminum, sponge, wool, rubber, silk, 
catgut, peat, agar, asbestos, cork, ivory, paraffin, Vaseline, celluloid, 
and silicone. For example, silicone implants have included spheres of 
various designs, including those which are solid, hollow, and inflatable. 
Glass beads have also been used to fill irregular cavities in the orbit 
(Gougelmann, H. P., "The Evolution of the Ocular Motility Implant" Int 
Ophthalmol Clin (1976) 10:689-711). Most of the implants composed of these 
materials were fully buried in the orbit, which was the usual procedure 
prior to 1941. (Gougelmann, H. P., "The Evolution of the Ocular Motility 
Implant" Int Ophthalmol Clin (1976) 10:689-711). 
In 1941, a combined implant and acrylic prosthesis was introduced by 
Ruedemann (Ruedemann, A. D., "Plastic Eye Implant" Amer J Ophthalmol 
(1946) 29:947-952). The extraocular muscles were attached to the posterior 
portion of this implant, which was covered with gauze. This Ruedemann 
implant was eventually abandoned, since it had to be manufactured before 
each operation, and further because secondary strabismus procedures were 
often required to correct late position problems. This implant was 
partially exposed and partially buried. 
There have been many other design variations of orbital implants since the 
Ruedemann eye, including several implants that when placed were partially 
exposed and partially buried, these implants allowed "interaction" with an 
externally placed, contact lens-type artificial eye through some exposed 
means, such as pegs, pins, or screws (Gougelmann, H. P., "The Evolution of 
the Ocular Motility Implant" Int Ophthalmol Clin (1976) 10:689-711). 
These partially exposed implants imparted good motility to the artificial 
eye, but were prone to infection and extrusion. Buried implants were then 
developed to provide motility through special contours on the anterior 
aspect of the implant which matched corresponding contours on the 
posterior aspect of the eye. Other buried implants employed magnets to 
achieve a form of coupling between the implant and the eye. 
Cutler employed a prosthesis comprising an implant with a peg to completely 
support the weight of the artificial eye and transfer all latent movement 
to the eye; however, these Cutler prostheses resulted in high rates of 
infection due to the inability of the material from which the implant was 
formed to support robust wound closure at the peg-implant interface. 
(Cutler M. L., "A Positive Contact Ball and Ring Implant for Use after 
Enucleation" Arch Ophthal (1947) 37:73-81). 
In recent years, porous ocular implants composed of hydroxyapatite (HA) and 
porous polyethylene (PP) have become accepted alternatives to traditional, 
nonporous spheres composed of silicone or acrylic. 
There is some variation in the art concerning the term "integration". The 
term is used to denote any connection between tissues of the recipient and 
the implant (e.g. suturing an extraocular muscle to a wire loop). However, 
as used in the context of the present invention, integrated implants are 
porous implants capable of sustaining fibrovascular in growth. Porous 
implants have the advantage of becoming infiltrated by fibrovascular 
tissue, thereby providing resistance to infection, migration, and 
extrusion. (Merritt, K., et al., "Implant Site Infection Rates with Porous 
and Dense Materials" J Biomed Mater Res (1979) 13:101-8; Rosen, H. M., 
"The Response of Porous Hydroxyapatite to Contiguous Tissue Infection" 
Plast Recontr Surg (1991) 88:1076-80; Dutton, J. J., "Coralline 
Hydroxyapatite as an Ocular Implant" Ophthalmology (1991) 98:370-7; 
Shields, C. L, et al., "Lack of Complications of the Hydroxyapatite 
Orbital Implant in 250 Consecutive Cases" Trans Am Ophthalmol Soc (1993) 
91:177-189; discussion 189-95). 
An integrated implant also offers the possibility of good motility for an 
artificial eye by use of a motility/support peg. Furthermore, an 
integrated implant that incorporates a motility/support peg may (by 
supporting the artificial eye) also help prevent the development of a deep 
superior sulcus and entropion or ectropion of the lower lid, and may 
reduce other long-term structural defects due to chronic weight and 
pressure from the artificial eye. (Gougelmann, H. P., "The Evolution of 
the Ocular Motility Implant" Int Ophthalmol Clin (1976) 10:689-711). 
Not all porous implants can transfer implant movement directly to the 
artificial eye via a motility/support peg. Porous HA implants have the 
ability to accept a motility/support peg because, when fully vascularized, 
they can support complete epithelialization of the internal surfaces of 
the peg hole, thereby sealing the implant from the external environment 
and preventing infection. Porous polyethylene has only recently been 
coupled to the eye in this manner. Generally, porous polyethylene implants 
impart some motility through movement of the fornices, when the 
extraocular muscles are surgically connected to the fornices or to the 
implant. 
Vascularization can be a lengthy process in porous implants, requiring 6 to 
10 months or longer in some cases (Dutton, J. J., "Coralline 
Hydroxyapatite as an Ocular Implant" Ophthalmology (1991) 98:370-7). Peg 
placement is usually delayed until the implant shows a high degree of 
fibrovascular ingrowth, as established by some objective means, such as a 
bone scan or MRI. (Baumgarten, D., et al., "Evaluation of Biomatrix 
Hydroxyapatite Ocular Implants with Technetium-99m-mdp" J Nucl Med (1993) 
34:467-468; DePotter, P., et al., "Role of Magnetic Resonance Imaging in 
the Evaluation of the Hydroxyapatite Orbital Implant" Ophthalmology (1992) 
99:824-830) Since drilling of the implant for placement of the 
motility/support peg is usually delayed until the implant is fully 
vascularized, the complete rehabilitation of the patient can be limited by 
the speed and degree of fibrovascular ingrowth. Therefore, rapid, complete 
vascularization of these implants is desirable. Previous efforts to speed 
the process have included drilling holes in HA implants (Ferrone, P. J., 
and Dutton, J. J., "Rate of Vascularization of Coralline Hydroxyapatite 
Ocular Implants" Ophthalmology. (1992) 99:376-379) and cutting windows in 
any coating materials, such as donor sclera, used to wrap the implant, in 
order to increase direct contact between the HA material and the highly 
vascular tissues of the orbit. 
In particular, porous HA implants have the known ability to accept a 
motility/support peg, making possible the direct transfer of implant 
movement to the artificial eye. Preferred support pegs include those such 
as disclosed in copending application Ser. No. 08/241,960, filed May 12, 
1994; Ser. No. 08/853,647 filed May 9, 1997; and, Ser. No. 08/886,600 
filed Jul. 1, 1997, each in the name of Arthur C. Perry, and each of which 
are fully incorporated by reference herein. 
Since the cosmetic and psychological rehabilitation of the anophthalmic 
patient may depend on life-like movement and position of an artificial 
eye, compositions and methods are needed to increase the speed of 
fibrovascular ingrowth into an implant, since such ingrowth is a 
precondition of drilling the implant to accept the motility/support peg. 
Popular coralline HA implants currently available have a reported pore size 
of 500 .mu.m (HA500) (Interpore 500, Interpore International, Irvine, 
Calif.); these implants provide excellent fibrovascular ingrowth, but have 
a rough outer surface that may lead to exposure of the implant following 
surgery, due to abrasion of the overlying conjunctiva and Tenon's capsule. 
To avoid implant exposure, current practice calls for coating the implant 
in some material, such as donor sclera or fascia lata. (Perry, A. C., 
"Integrated Orbital Implants" Adv Ophthalmic Plast Reconstr Surg (1990) 
8:75-81). However, concerns about HIV infection and the additional 
surgeries needed to harvest a donor coating material have led to a search 
for suitable alternative coatings. (Dutton, J. J., "Coralline 
Hydroxyapatite as an Ocular Implant" Ophthalmology (1991) 98:370-7). 
Accordingly, there is a need for an ocular implant material having a 
smoother implant surface. A smoother implant surface could reduce abrasion 
on the orbital tissues during and after implantation, facilitates deeper 
placement of the implant in the orbit, and can reduce intraoperative time 
because the implant may not need to be surrounded by an additional 
coating. 
It has been found that both hydroxyapatite and porous polyethylene implants 
are capable of complete vascularization. The hydroxyapatite implants 
vascularize more rapidly than the commercially available porous 
polyethylene (MedPor, Porex Surgical, College Park, Ga.). When the 
interstitial pore size of the PP was increased to a reported pore size of 
approximately 400 microns, which corresponded to the reported pore size of 
commercially available HA (e.g., Interpore, Irvine, Calif.), the rate and 
extent of vascularization of PP and HA were more similar, a is finding 
that indicated that increasing the interstitial pore size favorably 
influenced vascularization of porous implants. (Rubin, P. A. D., et al. 
"Comparison of Fibrovascular Ingrowth Into Hydroxyapatite and Porous 
Polyethylene Orbital Implants" Ophthalmic Plastic and Reconstructive 
Surgery 10(2):96-103 (1994)). 
Thus, it was found that PP with pore sizes in the 400 micron range, 
resulted in more optimal orbital tissue ingrowth than a denser PP implant 
having an interstitial pore size of approximately 150 microns. It was 
noted by the authors of the previous study that it was not clear to what 
extent a further increases in pore size would enhance vascularization, 
providing guidance in the art that even larger pore sizes were desirable. 
Again, it was noted that there was a need for maximizing the rate and 
extent of fibrovascular ingrowth, while minimizing the inflammation and 
the potential for infection with the relatively large orbital implant. 
Maximized soft tissue ingrowth into the depths of an implant decreases the 
inflammatory cell response and potential for infection. (Rubin, P. A. D., 
et al. "Comparison of Fibrovascular Ingrowth Into Hydroxyapatite and 
Porous Polyethylene Orbital Implants" Ophthalmic Plastic and 
Reconstructive Surgery 10(2):96-103 (1994)). 
Plaster of Paris is a biocompatible material which is composed of the 
hemihydrate form of calcium sulfate produced by heating gypsum calcium 
sulfate dihydrate to drive off water. (Alexander, H., et al., 
"Calcium-based Ceramics and Composites in Bone Reconstruction" CRC 
Critical Reviews in Biocompatibility (1987) 4:43-77) It is highly 
biocompatible and has been successfully used to fill defects in bone 
(Peltier, L. F., "The Use of Plaster of Paris to Fill Defects in Bone" 
Clin Orthop (1961) 21:1-31), in dental surgery, and for orbital 
augmentation (Geist, C. E., et al., "Orbital Augmentation by 
Hydroxylapatite-based Composites. A Rabbit Study and Comparative Analysis" 
Ophthalmic Plast Reconstr Surg (1991) 7:8-22). When mixed with HA 
particles for orbital augmentation, calcium sulfate has been shown to 
resorb within several weeks of implantation. Moreover, connective tissue 
ingrowth has been noted in mixtures of HA and calcium sulfate, with 
minimal inflammation (Geist, C. E., et al., "Orbital Augmentation by 
Hydroxylapatite-based Composites. A Rabbit Study and Comparative Analysis" 
Ophthalmic Plast Reconstr Surg (1991) 7:8-22). 
Thus, there is need for an orbital implant with as many of the following 
characteristics as possible: It should be biocompatible, readily 
vascularized, and have little or no tendency toward extrusion, migration, 
or infection (see, e.g., Dutton, J. J., "Coralline Hydroxyapatite as an 
Ocular Implant" Ophthalmology (1991) 98:370-7); it should also serve to 
impart motility to an artificial eye while supporting the weight of the 
eye to preserve the delicate, and cosmetically important, structures of 
the lid; and preferably is capable of being attached to an artificial eye. 
DISCLOSURE OF THE INVENTION 
A porous structure for implantation into the orbital cavity of a mammal 
comprising pores having a mean size of less than 200 micrometers. In 
preferred embodiments, the pores have a mean size of from 50 to 150 
micrometers; a mean size of from 60 to 90 micrometers; a mean size of from 
75 to 85 micrometers; or, a mean size of about 77 micrometers. The 
structure can comprise a growth factor, such as recombinant human basic 
fibroblast growth factor beta. The structure can comprise a biocompatible 
coating, such as calcium sulfate, polylactic acid, polyglycolic acid, or 
animal tissue. 
Also disclosed is a surgical method for placing a implant into a mammal who 
has had an ocular enucleation, evisceration or who needs implant 
replacement (i.e. "secondary implantation"), whereby the implant obtains 
rapid ingrowth of connective and vascular tissues, said method comprising: 
selecting a porous ocular implant comprising pores with a mean size of 
less than 200 micrometers; and, placing the implant into an orbital cavity 
of a mammal. The method can further comprise a step of burying the implant 
beneath conjunctival tissues of the mammal. The method can further 
comprise a step of covering the implant before the placing step, with a 
material such as scleral tissue or calcium sulfate. The method can further 
comprise applying a growth factor to the implant, such as recombinant 
human basic fibroblast growth factor beta.

MODES FOR CARRYING OUT INVENTION 
As disclosed herein, fibrovascular ingrowth into various porous ocular 
implants was investigated in an animal model, as a function of implant 
material composition, porosity, addition of growth factors, and use of 
coatings. Eighty-one new Zealand white rabbits underwent unilateral 
enucleation and implantation with ocular implants composed of the 
following materials: 
coralline hydroxyapatite (HA) ("HA200") (Interpore 200, Interpore 
International, Irvine, Calif.); 
coralline hydroxyapatite ("HA500") (Interpore 500, Interpore International, 
Irvine, Calif.); 
synthetic HA ("synHA"); and, 
high-density porous polyethylene ("PP"). 
The HA200, HA500, and PP implants were implanted untreated or after 
treatment with recombinant human basic fibroblastic growth factor 
(Rh-.beta.FGF). Nine HA500 implants were implanted after coating with 
calcium sulfate (Plaster of Paris) to provide a smooth outer surface. 
Implants were harvested at 1-, 2-, 4-, or 8-week intervals and were 
examined histologically. 
As discussed below, no significant differences in the degree of 
fibrovascular ingrowth were found as a function of implant composition. 
Surprisingly, significant increases in ingrowth were found in HA200 
compared with HA500 implants. Increases in ingrowth were also found in 
Rh-.beta.FGF-treated implants compared with untreated controls. The 
calcium sulfate coated implants showed less vascularization compared with 
the uncoated implants, although the difference was not significant. 
Composition 
The two most popular materials used to manufacture porous implants, i.e., 
implants of PP and implants of coralline HA, were tested in the studies 
disclosed herein. Also, tested was synthetic hydroxyapatite. No 
significant differences in ingrowth were noted between implants composed 
of these materials (FIGS. 3 and 4), although they represent markedly 
different technologies. 
Porosity 
Porous implant materials, such as HA and PP, possess many of the 
characteristics presently deemed preferred for an ocular implant, and they 
offer clear advantages over solid, nonporous spheres. These preferred 
porous compositions allow fibrovascular tissue to grow into the implants; 
the presence of fibrovascular ingrowth is believed to prevent migration 
within the orbit and may help to minimize the chance of infection and 
exposure through breakdown of the thin overlying tissues of the 
conjunctiva and Tenon's capsule (Buettner, H. and Bartley, G., "Tissue 
Breakdown and Exposure Associated with Orbital Hydroxyapatite Implants" Am 
J Ophthalmol (1992) 113:669-673). A further advantage of porous implants 
is their ability to be directly integrated with the extraocular muscles, 
thereby maximizing the transfer of all latent muscle movement to the 
implant. 
The importance of pore size in determining the nature of fibrovascular 
tissue was investigated in the present study. It was a surprising finding 
that the HA200 showed significantly better ingrowth than HA500. This 
result is believed to be due to the fact that smaller pores, in this case 
pores reported as 200-.mu.m, rather than pores reported as 500-.mu.m, may 
encourage fibrovascular ingrowth. This is a clinically important finding, 
for an additional reason: HA200 implants have a smoother surface and may 
thus be less prone to abrade the overlying tissues and are less likely to 
lead to exposures. It may also be easier to place these implants more 
deeply within the orbit. 
Growth Factors 
Growth factors have been successful in associated medical fields, and 
relatively recently have been used in ophthalmology. Thus, the present 
studies were performed to determine the applicability of growth factors in 
ocular implant surgery. Traditional methods of extracting growth factors 
from human placenta or bovine brain were laborious and ineffective (Rieck, 
P., et al., "Human Recombinant .beta.FGF Stimulates Endothelial Wound 
Healing in Rabbits." Current Eye Research (1992) 11:1161-1172); but recent 
progress in recombinant DNA technology has made it possible to produce 
growth factors on a scale large enough to make their therapeutic use a 
practical consideration. 
Basic fibroblast growth factor (FGF) is stored within basement membranes 
and may exhibit angiogenic activity. Recombinant human basic fibroblast 
growth factor is derived from E. coli through recombinant DNA techniques. 
Rh-.beta.FGF is a 146 amino acid polypeptide from a family of growth 
factors that show a high affinity for heparin and have been extracted from 
a number of tissues such as eye, retina, brain, pituitary, and human 
placenta (Folkman, J., and Klagsbrun, M., "Angiogenic Factors" Science 
(1987) 235:442-447; Rieck, P., et al., "Recombinant Human Basic Fibroblast 
Growth Factor (Rh-.beta.FGF) in Three Different Wound Models in Rabbits: 
Corneal Wound Healing Effect and Pharmacology" Exp Eye Res (1992) 
54:987-998). It is one of several angiogenic factors which in recent years 
have been fully purified, their amino acid sequences determined, and their 
genes cloned. 
Recombinant human fibroblast growth factor beta (Rh-.beta.FGF) has been 
used in rabbit corneal studies to promote epithelial and endothelial wound 
healing (Rieck, P., et al., "Human Recombinant .beta.FGF Stimulates 
Endothelial Wound Healing in Rabbits." Current Eye Research (1992) 
11:1161-1172). 
Rh-.beta.FGF was used in the present study because of the demonstrated 
ability of this and related growth factors to induce new capillary blood 
vessel ingrowth in vitro and in vivo (Montesano, R., et al., "Basic 
Fibroblast Growth Factor Induces Angiogenesis In Vitro" Proc Natl Acad Sci 
USA (1986) 83:7297-7301; Baird, A., and Bohlen, P., "Fibroblast Growth 
Factors", In: Sporin, M. B., and Roberts, A. B., eds. Peptide Growth 
Factors and Their Receptors 1. (New York, Springer-Verlag, 1991). 
Capillary blood vessel formation is a complex process which includes 
endothelial cell proliferation, the sprouting of new capillaries, the 
migration of endothelial cells, and the breakdown of extracellular matrix 
surrounding existing capillaries. 
Exogenously applied Rh-.beta.FGF may stimulate all of the biological 
activities required to elicit neovascularization (Rieck, P., et al., 
"Human Recombinant .beta.FGF Stimulates Endothelial Wound Healing in 
Rabbits." Current Eye Research (1992) 11:1161-1172). 
The present studies investigated whether Rh-.beta.FGF could be affixed to 
PP and HA implants and, if so, whether they would enhance the rate of 
vascularization. 
In the present studies, all implants treated with Rh-.beta.FGF showed 
significant increases in fibrovascular ingrowth compared with identical, 
untreated implants (FIG. 5). This finding is clinically significant, in 
view of the objective to decrease the latency of peg-fitting following 
enucleation, evisceration, or secondary implantation. It is also 
noteworthy that none of the implants treated with Rh-.beta.FGF in the 
present studies were observed to be exposed at any time, despite marked 
postoperative inflammation in orbits containing these implants. 
Although previous corneal studies showed that Rh-.beta.FGF is tolerated in 
rabbits, a greater-than-normal inflammatory response was observed during 
the first 72 hours, characterized by increased erythema and 
orbital/peri-orbital edema. However, there were no episodes of extrusion 
or infection, and by 72 hours the rabbits showed no unusual symptoms. As 
appreciated by one of ordinary skill in the art, this finding suggests 
that the Rh-.beta.FGF did bind to some degree to the implants and that a 
lower dose might be used in any future situations. 
As appreciated by those of ordinary skill in the art, concerns regarding 
the safety of growth factors, such as systemic absorption, must be taken 
into account when used as a means to enhance fibrovascular ingrowth into 
these implants. At least one previous study showed no evidence of systemic 
absorption when Rh-.beta.FGF was applied to the rabbit cornea to 
investigate its role in healing corneal epithelium (Rieck, P., et al., 
"Recombinant Human Basic Fibroblast Growth Factor (Rh-.beta.FGF) in Three 
Different Wound Models in Rabbits: Corneal Wound Healing Effect and 
Pharmacology" Exp Eye Res (1992) 54:987-998); although its behavior when 
placed within orbital tissues may be different. 
Calcium sulfate coating 
Donor sclera and other coverings are used by surgeons to provide for 
several therapeutic advantages, such as to: facilitate attachment of 
extra-ocular muscles to HA implants, to allow placement of the implant 
deeply within the orbit, and to prevent tissue breakdown over the rough 
anterior surface of HA implants. Biocompatible coatings that are used with 
orbital implants include polylactic acid, polyglycolic acid, and animal 
tissues such as sclera and fascia. Presently preferred coatings are 
disclosed in copending U.S. application Ser. No. 08/241,960 filed May 12, 
1994 in the name of Arthur C. Perry. As disclosed herein, calcium sulfate 
was used to coat the HA spheres so as to provide some of the 
above-described therapeutic advantages of scleral wrappings. 
An HA implant manufactured with a smooth covering that achieves these goals 
while not impeding vascularization would offer several benefits. For 
example, the cost of obtaining banked sclera would be saved and surgical 
time would be reduced, particularly relative to use of autologous sclera. 
Additionally, concerns of infectivity from donor tissue could be reduced, 
since some patients have refused to accept an HA implant coated with 
allogeneic sclera due to even a theoretical risk of virus transmission 
such as HIV (Dutton, J. J., "Coralline Hydroxyapatite as an Ocular 
Implant" Ophthalmology (1991) 98:370-7). Although a pre-coated HA implant 
may not replace the function of sclera or other tissue wrappings as a 
facile means of attaching the extraocular muscles, some surgeons already 
routinely attach the muscles directly to the HA, without the use of a 
scleral covering. 
The implants disclosed herein were coated with a relatively thick layer of 
plaster of Paris, i.e., calcium sulfate of about 1 to 1.5 mm thickness. 
The calcium sulfate coating was still visible at the time of explantation 
in all cases (FIG. 6). While the material appeared to be well tolerated by 
the rabbit orbit, two of the calcium sulfate-coated HA implants were lost 
in the present studies and were presumed to have been exposed and extruded 
prior to harvesting. The orbital tissues in these rabbits were otherwise 
healthy in appearance. 
Although the calcium sulfate-coated implants did not show significantly 
lower degrees of fibrovascular ingrowth, the loss of 2 specimens may have 
affected the statistical power of these results, as visual inspection 
showed some inhibition of ingrowth. Calcium sulfate coating may still be 
useful, although thinner layers of calcium sulfate may better achieve 
desired effects. Additionally, a more-controlled method of application of 
the coating may be desirable to minimize infections and to ensure the 
desired thickness can be achieved. 
The present data regarding fibrovascular ingrowth is relevant due to its 
status as a prerequisite of complete integration of porous ocular 
implants, and because it may be necessary to achieve the complete cosmetic 
and psychological rehabilitation of an anophthalmic patient, since only a 
fully vascularized implant can accommodate a motility/support peg. 
EXAMPLES 
The present examples evaluated the rate and degree of fibrovascular 
ingrowth in common porous ocular implants as a function of material 
composition, porosity, treatment with growth factors, and the application 
of a calcium sulfate (Plaster of Paris) coating. 
The present results indicated that smaller pore sizes, minimal physical 
barriers to ingrowth, and active growth induction by means of growth 
factors are means of achieving the speed and level of ingrowth needed to 
achieve the best surgical result and patient satisfaction with porous 
ocular implants. Particularly surprising was that smaller pore sizes led 
to enhanced fibrovascular ingrowth. 
Example 1 
Surgical Enucleation 
The animals involved in the studies regarding the present invention were 
procured, maintained, and used in accordance with the Animal Welfare Act 
of 1966, as amended, and the Guide for the Care and Use of Laboratory 
Animals prepared by the Institute of Laboratory Animal Resources, National 
Academy of Sciences--National Research Council, as required by SECNAVINST 
3900.38B. 
Enucleation of the right eye was performed on 81 New Zealand white rabbits 
averaging 3 kg in weight. Intramuscular anesthesia was administered with 
50 mg/kg of Ketamine and 5 mg/kg of Xylazine. The rabbits were also given 
an IM dose of 0.25 cc of penicillin G. The peri-orbital fur was shaved and 
the right eye and peri-orbital area were prepped with Betadine solution. A 
retrobulbar injection of Xylocaine 1% with epinephrine 1:100,000 was given 
and the surgical site was draped in a sterile fashion. 
Following a complete peritomy, the extraocular muscles were tagged with 
double-armed 5-0 Vicryl suture and were then released from the globe. 
Muscles were isolated in the infer-nasal quadrant and the supero-lateral 
quadrant. Upon close inspection, the muscle complex in the supero-lateral 
quadrant appeared to be composed of two muscles, i.e., a rectus muscle 
with an attached oblique muscle. The globe was enucleated and pressure was 
applied to the posterior orbit for hemostasis. 
Example 2 
Calcium Sulfate Coating 
To control the level of calcium sulfate infiltration into the pores, the 
coralline HA implants were filled with water and frozen prior to 
application of the calcium sulfate. All implants, except those treated 
with Rh-.beta.FGF, were sterilized by autoclaving prior to implantation. 
The Rh-.beta.FGF-treated implants were autoclaved prior to the application 
of the growth factor in a sterile solution. 
Example 3 
Surgical Implantation of the Various Implants 
Twelve-millimeter ocular implants were prepared for implantation in all 
cases. A previous study found that 14-mm implants resulted in high rates 
of exposure (Rubin, P. A., et al., "Comparison of Fibrovascular Ingrowth 
into Hydroxyapatite and Porous Polyethylene Orbital Implants" Ophthalmic 
Plast Reconstr Surg (1994) 10:96-103), probably due to the small size of 
the rabbit orbit (J. K. Popham, personal communication, 1995). 
All implants were immersed in a 20-mg/ml solution of gentamicin prior to 
implantation, except for those treated with fibroblast growth factor 
(Rh-.beta.FGF), which was in a sterile solution. Rh-.beta.FGF was obtained 
from the Department of Cell Biology, Scripps Research Institute, La Jolla, 
Calif. It was stored at -80.degree. C. and was passed over an endotoxin 
column to remove bacterial endotoxin before dilution to a concentration of 
10 .mu.g/ml. The concentration was verified by a protein assay and 
spectrophotometry. 
The implants were separated by type and placed in sterile 30-ml syringes, 
which were then filled with enough solution to completely immerse the 
implant. Each syringe was capped, the plunger was withdrawn to create a 
mild vacuum, and the barrel was agitated to release residual air from the 
implant. The implants were incubated overnight at 4.degree. C. Prior to 
implantation, the implants were gently rinsed twice with sterile PBS. 
All implants were supplied by their respective manufacturers. The calcium 
sulfate and Rh-.beta.FGF treatments were applied as described herein. 
Each 12-mm implant was placed in the center of the orbit between the 
muscles, and the muscles were sutured to each other over the anterior 
aspect of the implant. Since all implants were placed unwrapped (i.e., 
were not within a scleral shell), sterile plastic sleeves were created 
around the implants, fashioned from surgical drapes or gloves, to 
facilitate insertion of the implants deeply into the orbit. After the 
implant insertion, the sleeve material was removed, the conjunctiva was 
closed meticulously with 5-0 Vicryl suture, standard ophthalmic antibiotic 
ointment was applied to the orbit and, the lids were sutured together with 
at least one 5-0 Vicryl suture to allow instillation of ointment 
postoperatively. The rabbits were individually caged following recovery 
from anesthesia and each was observed closely during the first 
postoperative week. Antibiotic ophthalmic ointment was applied to the 
orbit daily during the first postoperative week. 
Example 4 
Surgical Explantation 
For explantation, the rabbits were sacrificed at the predetermined 
intervals with intramuscular Ketamine and Xylazine, followed by 2.5 ml of 
intra-cardiac beuthanasia D. The implants and surrounding tissues were 
removed and fixed in Formalin. Each specimen was then soaked in water, 
dehydrated in alcohol, vacuum infiltrated, and then imbedded in methyl 
methacrylate. 
Example 5 
Specimen Analysis 
The specimens were cut through the center into 1- to 2-mm sections using a 
wet diamond band saw. The slices were mounted on slides, recut using the 
sandwich method, and ground down to a thickness of approximately 200 
.mu.m. All specimens were stained using the fibrin stain of Ladewig. 
The sections were examined at a magnification of 50.times. using an 
overlying 1-mm grid. Each 1-mm increment of depth (level) from the edge of 
the implant was measured at 4 different quadrants by 2 investigators, for 
a total of 8 measurements per level, with the exception of the innermost 
level (the intersection of the axes) which was measured twice. 
Two investigators independently characterized each section at 1-mm 
intervals (grids) along two perpendicular axes through the center of the 
implant (FIG. 1), and each investigator estimated the fibrovascular 
ingrowth in each grid according to the following scale: 0%, .ltoreq.25%, 
.ltoreq.50%, .ltoreq.75%, and .ltoreq.100% of available pore space. For 
example, a 1.times.1-mm grid containing some available pore area with no 
evidence of tissue ingrowth was graded 0%, while an area containing even 
one cell or tissue fiber was graded .ltoreq.25% (FIG. 2). 
The inter-investigator assessments were consistent (correlation coefficient 
0.935) throughout the investigation (p&lt;0.0001). The assessments for the 
eight data points in each level were averaged to derive the 
percent-ingrowth per level. The percent-ingrowth for all levels within an 
implant were averaged to derive the percent-ingrowth per implant type (see 
Table 3). Statistical differences in the degree of fibrovascular ingrowth 
among the implant types was determined using a 3-way analysis of variance. 
Example 6 
Results Following Surgical Explanation 
Table 1 shows the breakdown of implant types, treatments and number of 
implants evaluated. 
TABLE 1 
______________________________________ 
Implant type and treatment 
NUMBER NUMBER 
IMPLANT TYPE 
NUMBER EXTRUDED EVALUATED 
______________________________________ 
HA200 12 0 12 
HA200 GF 9 0 9 
HA500 12 1 11 
HA500 GF 9 0 9 
HA500 CS 9 2 7 
PP 12 0 12 
PP GF 9 0 9 
SynHA 9 0 9 
______________________________________ 
Legend: 
HA200, reported 200 .mu.mporosity coralline hydroxyapatite; HA500, 
reported 500 .mu.mporosity coralline hydroxyapatite; PP, porous 
polyethylene; SynHA; synthetic porous hydroxyapatite; GF, growth 
factortreated; CS, calcium sulfatecoated. 
Three of each of the implants treated with growth factor (HA200 GF, HA500 
GF, PP GF) were explanted at intervals of 1, 2, and 4 weeks to assess 
fibrovascular ingrowth. The same implant types without growth factor 
(controls) were harvested at 1, 2, 4, and 8 weeks. The synHA implants and 
those coated with calcium sulfate were harvested at 2, 4, and 8 weeks (see 
Table 2). Three of the HA500 implants (2 HA500CS, 1 HA500) were not 
present at the time of harvesting and were presumed to have been extruded. 
TABLE 2 
______________________________________ 
Schedule of Explanation and Numbers Explanted, by 
Implant Type and Treatment 
Implant Type 
Week 1 Week 2 Week 4 
Week 8 
______________________________________ 
HA200 3 3 3 3 
HA200 GF 3 3 3 0 
HA500 3 2 3 3 
HA500 GF 3 3 3 0 
HA500 CS 0 3 2 2 
PP 3 3 3 3 
PP GF 3 3 3 0 
SynHA 0 3 3 3 
______________________________________ 
Exposure and Extrusion--At the time of explanation, two calcium 
sulfate-coated HA500 implants and one uncoated HA500 implant were absent 
and were presumed to have completely extruded from the orbit after the 
first postoperative week. 
Of the implants present during explantation, 8 of 79 (10.1%) showed 
evidence of exposure. Exposure was only noted in the HA implants, as 
follows: 3 HA500, 3 calcium sulfate-coated HA500, and 2 HA200. 
Inflammation and Infection--Most of the 81 rabbits demonstrated some thick, 
white conjunctival discharge from the orbit in the early postoperative 
period, which cleared spontaneously. In one rabbit, the discharge 
continued beyond this period but was responsive to additional applications 
of ointment. None of the rabbits showed signs of chronic orbital 
infection. 
Most (56%) of the orbits containing implants treated with growth factor 
showed elevated levels of edema and erythema of the lids and peri-orbital 
tissues during the first 72 hours postoperatively. The level of 
inflammation was characterized as follows: 9 moderate (PP, 5; HA500, 4) 
and 6 severe (PP, 2; HA500, 4). None of the HA200 growth factor-treated 
implants were associated with elevated levels of inflammation. By 
postoperative day 3, all of the growth factor rabbits showed normal levels 
of inflammation and none showed signs of infection. 
Notably, a significant difference (p=0.027) was found between the level of 
ingrowth in HA200 and HA500 implants, with the HA200 showing more complete 
ingrowth. This finding was surprising in view of disclosures regarding the 
impact of porosity on ingrowth. Previously, it had been reported that 
implants with larger pores would achieve better ingrowth relative to 
implants with smaller pores. 
Implants treated with growth factor showed significantly greater (p=0.014) 
fibrovascular ingrowth than untreated implants. 
Implants coated with calcium sulfate showed less ingrowth than uncoated 
implants, although the difference did not reach statistical significance 
(p=0.055). 
There was a significant difference (p=0.001) in ingrowth between the 4 time 
periods (1 wk, 2 wks, 4 wks, 8 wks) in which the implants were explanted. 
In all cases, except in the case of PP during week 2, and in 3 instances 
of extruded implants which restricted the number of data points in a 
particular cell to 2 implants (see Table 3), more ingrowth was noted with 
each successive week. 
TABLE 3 
______________________________________ 
Percent-ingrowth per implant as a function of 
material composition, porosity, growth factors, and coatings. 
Implant Type 
Week 1 Week 2 Week 4 Week 8 
______________________________________ 
HA200 60.6 64.9 78.5 91.3 
HA200 GF 44.5 69.3 100.0 NA 
HA500 54.7 35.2 [n = 2] 
66.0 82.8 
HA500 GF 61.6 81.8 79.3 NA 
HA500 CS NA 35.8 62.8 [n = 2] 
41.9 [n = 2] 
PP 73.8 58.3 82.9 90.5 
PP GF 76.0 81.1 85.4 NA 
SynHA NA 53.8 64.3 86.6 
______________________________________ 
Example 7 
Pore Size Calculations 
Previously, it has been reported that the HA 200 implants had pores sized 
at 200 micrometers, and that the HA 500 implants had pores sized at 500 
micrometers. To confirm these pore sizes, and to determine the pore sizes 
of the synthetic HA (SynHA) and the porous polyethylene (PP), the 
following protocol was undertaken. Samples of materials were photographed 
in a scanning electron microscope (SEM) (Leica STEREOSCAN 400.RTM., Leica, 
Inc., Deerfield, Ill.). Coralline hydroxyapatite (e.g., HA200 and HA500) 
has an anisotropic structure. Because of this anisotropy, these samples 
were prepared for analysis by grinding both parallel and transverse to the 
long axis of the pore structure. Synthetic HA has a generally isotropic 
structure. Porous polyethylene is known to have a radial pore structure 
gradient (see, e.g., Klawitter, J. J. An Evaluation Bone Growth into 
Porous High Density Polyethylene J. Biomed. Mater Res. 10:311-323 (1976)) 
Accordingly, only the exterior surfaces of the porous polyethylene samples 
were evaluated. 
Photographs of all samples were enlarged a defined amount, and 
cross-sections of pores were measured manually. The calibration scale 
printed on the photograph by the SEM was used to convert the measurements 
to pore dimensions. The longest dimension and the shortest dimension of 
each pore in the plane of the surface were determined. The two 
measurements were averaged to obtain a measurement of each pore. The mean 
of all measured pore sizes was calculated, and a histogram of pore size 
distribution was prepared for each implant-type measured. 
FIG. 7 depicts a histogram of pore size measurements for transverse 
sections of the HA 200 samples. Forty samples were measured in this plane. 
The mean pore size was 64 micrometers. 
FIG. 8 depicts a histogram of the pore size measurements from longitudinal 
sections of the HA 200 implants. Forty-three specimens were examined in 
this plane. The mean pore size was 89 micrometers. 
FIG. 9 depicts a histogram for the pore size measurements of HA 500 
implants taken along transverse sections. Fifty-eight specimens were 
examined in this plane. The mean pore size for these measurements was 262 
micrometers. 
FIG. 10 depicts a histogram of pore size measurements taken from 
longitudinal sections of HA 500 implants. Fifty-four specimens were 
examined in this plane. The mean pore size from these specimens was 220 
micrometers. 
FIG. 11 depicts a histogram of pore size measurements for the implants of 
porous polyethylene. These pore size measurements were taken at the 
surface of the implants. Twenty-one specimens were examined. The mean pore 
size was 563 micrometers. 
FIG. 12 depicts a histogram for the pore size measurements taken from 
synthetic hydroxyapatite implants. Fifty-two specimens were examined. The 
mean pore size for these samples was 220 micrometers. 
It was a surprising finding that the pore size data for the coralline 
hydroxyapatite implant differed so significantly from what was reported to 
be the approximate pore size for these materials. Moreover, the newly 
determined pore size information taken together with the data for 
fibrovascular ingrowth for the various samples, manifested the 
particularly surprising finding that enhanced fibrovascular ingrowth took 
place for hydroxyapatite implants (HA 200) having a mean pore size of 64 
micrometers in a transverse section and 89 micrometers in longitudinal 
section. Taking an average of the transverse and longitudinal pore size 
determinations for this material, it was found that enhanced fibrovascular 
ingrowth took place in a material having an average pore size of 
approximately 77 microns. Previously reported studies directed and 
encouraged selection of implant materials having substantially larger pore 
sizes than such samples. 
Closing 
It must be noted that as used herein and in the appended claims, the 
singular forms "a," "and," and "the" include plural referents unless the 
context clearly dictates otherwise. Thus, for example, reference to "a 
formulation" includes mixtures of different formulations and reference to 
"the method of treatment" includes reference to equivalent steps and 
methods known to those skilled in the art, and so forth. 
Unless defined otherwise, all technical and scientific terms used herein 
have the same meaning as commonly understood by one of ordinary skill in 
the art to which this invention belongs. Although any methods and 
materials similar to equivalent to those described herein can be used in 
the practice or testing of the invention, the preferred methods and 
materials are now described. All publications or applications mentioned 
herein are fully incorporated by reference herein.