222 AMERICAN JOURNAL OF OPHTHALMOLOGY AUGUST, 1967

chamber, along with ejection of an aqueous stream. Thickened cornea consisted of both "fused" and nonfused lamellae. Some of the lesions that did not penetrate into the eye were accompanied by a depression of the anterior lens surface, apparently a result of heat transmission. Deeper intraocular changes did not occur in the nonpenetrated eye within the limited time interval between irradiation and these preliminary observa­tions.

A form of corneal thickening here termed "fusion" of corneal lamellae that occurred at the periphery of the corneal lesion lost affinity for alcian blue, Masson, and Van Gieson stains. Electron microscopy of this region revealed scattered amorphous foci along the collagen fibrils in the stromal lamellae.

A clear plastic face shield 0.060 inches thick was found to be an effective protection to the eye under the limited conditions of these experiments. This shield may also serve as an indicator of accidental expo­sure.

The recent introduction of reconstituted collagen as suture material offered the first new commercially available absorbable suture material since catgut sutures were introduced by Joseph Lister in 1867.1 Re­ports of the use of collagen in general surgi­cal procedures and in some ophthalmic pro­cedures have already appeared.2-5 To date, no mention has been made of the use of re­constituted collagen sutures in corneal sur­gery. Because the cornea is composed most-

From the Division of Ophthalmology, Univer­sity of North Carolina School of Medicine, Chapel Hill, and McPherson Hospital, Dur­ham. This study was supported in part by a grant from the North Carolina Association for the Blind.

Armed Forces Institute of Pathology (20305)

REFERENCES 1. Yanoff, M. and Fine, B. S. : Glutaraldehyde

fixation of routine surgical eye tissue. Am. J. Ophth. 63:137, 1967.

2. Fine, B. S. and Zimmerman, L. E. : Miiller's cells and the "middle limiting membrane" of the human retina. Invest. Ophth. 1:304, 1962.

3. Fine, S., Klein, E., Peacock, G. R., Fine, B. S., Hansen, W. P. and Litwin, M. S. : Control of laser hazards and management of accidents. Sem­inar on laser safety, Martin Company, Orlando, Florida, May 19-20, 1966.

4. Meyer, K. and Anderson, B. : The chemical specificity of the mucopolysaccharides of the cor­nea. Exper. Eye Res. 4:346, 1965.

5. Offret, G., Payrau, P., Pouliquen, Y., Faure, J. P. and Hameau, J. P. : The mucopolysacchar­ides of the cornea; distribution, role, and modifications in normal and pathologic corneas. (Abstract) Surv. Ophth. 8:518, 1963.

6. Nordschow, C. D. : Aspects of aging in human collagen: an exploratory thermoelastic study. Exper. Molec. Path. 5:350, 1966.

7. Fine, B. S. and Fine, S. : Unpublished obser­vations.

8. Langley, R. K., Mortimer, C. B. and McCul-loch, C. : The experimental production of cata­racts by exposure to heat and light. Arch. Ophth. 63 :473, 1960.

9. Fine, S. : Unpublished data.

ly of collagen, sutures of the same material should be ideal for the repair of surgical wounds. This study was undertaken to evaluate the use of collagen sutures in clos­ing surgical wounds of the cornea.

MATERIAL

The suture material was derived from frozen flexor tendon of beef cattle.8 Bulk tendon was placed in a proteolytic enzyme to remove impurities and the remaining ma­terial placed in an organic acid-methanol-water medium in which controlled homoge-nization of collagen fibers took place. The resulting mixture was extruded into an ace­tone-ammonia bath producing strands in a

RECONSTITUTED COLLAGEN SUTURES IN CORNEAL SURGERY A N EXPERIMENTAL AND CLINICAL EVALUATION

GEORGE F. BRUMBACK, M.D., AND S A M U E L D. M C P H E R S O N , JR . , M.D. Durham, North Carolina

VOL. 64, NO. 2 COLLAGEN SUTURES 223

manner similar to the production of synthet­ic textile fibers. Strands were dried and ap­propriately tanned, resulting in the final su­ture.

The suture* used in this study was 6-0 plain collagen with a three-eighths curve re­verse cutting needle. The suture was pliable, handled easily and had excellent tying and knot-holding characteristics.

EXPERIMENTAL METHODS

The following study was undertaken to determine the gross and histologie reactions of rabbit corneas to collagen and the disap­pearance time of collagen sutures in experi­mental incisions.

Adult New Zealand white rabbits were anesthetized with intravenous sodium pento-barbital with a dose of 33 mg/kg. Lashes were clipped, the eyes irrigated with sterile saline and several drops of 0.5% propara-caine hydrochloride applied. The field was draped with sterile gauze squares and an eye sheet. A four-mm incision was made in the central cornea and closed with two tri­ply tied sutures. Tissue bites of approxi­mately half thickness were taken one mm from the wound edge (fig. 1). An antibiotic ophthalmic ointment was instilled and the rabbits returned to their cages for observa­tion. Ointment was applied daily for one week and observations recorded.

Thirty rabbits were used in this study and were killed at one, two, three, four, eight and 12 weeks postoperatively. Oper­ated eyes were enucleated, fixed in 10% for­malin and embedded in paraffin. Serial sec­tions stained with hematoxylin and eosin were examined with the microscope.

GROSS OBSERVATIONS

Conjunctival hyperemia, iris congestion and corneal edema developed the day after operation and subsided in 10 days. Eyes oc­casionally became inflamed for 24 to 48 hours at the time of suture disappearance.

* Furnished by Ethicon, Inc., Sommerville, New Jersey.

Fig. 1 (Brumback and McPherson). Four-mm corneal wounds were closed with two 6-0 collagen sutures.

This was thought to be due to the suture being pulled from the cornea by the com­bined actions of the lids and nictitating membrane and was most pronounced in three eyes in which suture sloughed at 25, 31 and 34 days, respectively.

The rate of disappearance of the extra-corneal portion of the suture was carefully observed. Suture knots began to disappear four days after operation and all were gone in 34 days; 95% of suture knots disap­peared between the fourth and 21st day (fig. 2) . The average time of knot disappearance was 13.3 days. Intralamellar sutures varied markedly in their time of disappearance.

MICROSCOPIC OBSERVATIONS

Sections of eyes removed one week after operation showed moderate edema, most marked at the incision and adjacent to the su­ture. Epithelization of the suture tracks was complete in seven of the 10 eyes examined. There was moderate infiltration of the wound and suture tracks with mononuclear and polymorphonuclear cells. Some macro­phage and fibroblastic cellular infiltration was seen in the suture track and wound. The suture did not appear fragmented or invaded by cells.

224 AMERICAN JOURNAL OF OPHTHALMOLOGY AUGUST, 1967

PLAIN COLLAGEN SUTURES (73) "NON-SENSITIZED" RABBITS

Fig. 2 (Brumback and McPher­son). Ninety-five percent of extra-corneal sutures disappeared be­tween the fourth and 21st day.

8 9 10 II 12 13 14 IS 16 17 18 192021 Time (days)

Two weeks after operation there was less corneal edema and fewer inflammatory cells. Fibroblastic proliferation at the wound in­terface was marked. Sections through su­tures showed early fragmentation and cellu­lar invasion of the suture. Epithelial cells in suture-free tracks were crenated. The over­all decrease in stromal infiltrate was closely paralleled by the decrease in cells at the lim-bus.

Three weeks after operation, most eyes had a normal cell population except for some large corneal corpuscles. One eye con­tained suture which was noted to be frag­mented and heavily invaded by macrophages (fig. 3) . In other eyes sutures were intact. Fibroblasts, macrophages and degenerating epithelial cells were seen in other suture tracks.

Four weeks after operation, two eyes still

Fig. 3 (Brumback and McPherson). Suture is being phagocytosed by wandering cells 21 days after operation. (Hematoxylin-eosin, χ330.)

VOL. 64, NO. 2 COLLAGEN SUTURES 225

contained suture fragments which were heavily invaded by macrophages. The re­maining eyes showed no sutures. No inflammatory cells were seen in the corneal stroma and wounds were bridged by lamel­lae of essentially normal thickness. Occa­sional epithelial cells were seen in suture tracks.

Six weeks after operation, sections re­vealed firm healing of wounds. Suture tracks were difficult to find and contained only occasional macrophages. No suture was found even with polarized light. Twelve weeks after operation sections showed es­sentially complete healing and appeared little different from those of eyes removed at six weeks.

Comment. Comparison with observations in a previous study by one of us demon­strated an over-all decrease of inflammatory cell response to collagen sutures as com­pared to sutures of plain catgut, chromic catgut and silk.7 In general, the microscopic observations were similar to those found by Taylor and associates8 in a study of rat-tail tendon as corneal suture material. The de­gree of tissue inertness to collagen sutures was the most striking single finding. This, combined with the predictability of suture loss, indicated that collagen sutures might be suitable for use in keratoplasty. A most de­sirable trait of collagen suture was the flat­tening of the strand incorporated in the knot. This appeared to give greater holding power to the knot.

SUTURE ANTIGENICITY

The following investigation was under­taken to determine the possible antigenicity of collagen suture and its influence on tis­sue response. Antigen was prepared from 3-0 plain collagen suture by trituration in a tissue homogenizer. The final suspension of suture material contained 1.0 mg/cc in ei­ther saline or a complete Freund's adjuvant mixture.9

Twelve rabbits were injected subcuta-

neously for an eight-week period with each animal receiving 11.3 mg of suture antigen. Six of the rabbits were given Freund's adjuvant mixture. All were skin tested with 0.01 mg of suture suspension intradermally, one week after completion of the injection schedule. All rabbits gave a negative reac­tion to skin test.

Corneal incisions were sutured as in the first study and the rabbits were treated and observed in the same manner. Rabbits were killed one, two and three weeks postopera-tively. Eyes were prepared for study as in the first group.

Observations. Gross reactions, knot dis­appearance and microscopic findings were not significantly different from those in the first study.

Comment. Others have investigated the antigenicity of collagen with more prolonged immunization schedules than used in this study. Only low titers of circulating anti­bodies were found.10'11 All rabbits in the present study, whether given plain or Freund's adjuvant antigen, gave negative results to the skin test. The rate of knot disap­pearance and the histologie findings were generally comparable to those in the nonin-jected rabbits.

CLINICAL

We have used 6-0 plain collagen sutures in 16 partial penetrating keratoplasties. The results were comparable to those previously obtained using silk or other nonabsorbable sutures as well as catgut. In most instances, 12 to 14 sutures were used to fix the grafts in place. Complications which might be at­tributed to suture material occurred in two patients. Iris prolapse occurred in one pa­tient on the first postoperative day. This was successfully repaired with similar su­ture material. Anterior synechiae formed, ne­cessitating synechiotomy on the 10th postop­erative day, in a second patient.

In 15 of 16 grafts, reliable statistics were available for 184 sutures. Three sutures

226 AMERICAN JOURNAL OF OPHTHALMOLOGY AUGUST, 1967

PLAIN COLLAGEN SUTURES IN PENETRATING KERATOPLASTY IN HUMAN EYES (15 EYES, 184 SUTURES)

10 T 12

r· 14

T· 16 20 22 30

Fig.

0 2 4 6 8 K) 12 14 16 18 20 22 24 26 28 Time (days)

4 (Brumback and McPherson). Most suture knots disappeared between 11 and 21 days after operation.

were removed for various reasons : two 21 days and one 14 days postoperatively. Dis­appearance of the extracorneal portion of the suture began at 11 days and all knots were gone in 30 days. The average time of knot disappearance was 16.3 days; 50% of the knots disappeared in 15 days and 75% in 18. Suture knot disappearance began and ended in a total span of 19 days (fig. 4) .

Fig. 5 (Brumback and McPherson). Suture struts are present seven weeks after operation.

This was quite in contrast to our experience with plain and chromic catgut sutures where the intervals of time involved were 33 and 56 days.7

The intralamellar suture struts remained across the wound for varying periods of time. Struts seldom disappeared in less than three weeks and in many instances persisted for months (fig. 5). Even in these instances there was a noticeable lack of foreign-body reaction or stimulus to corneal vasculariza-tion and this marked degree of tissue inert­ness to the suture material was striking in each case (fig. 6). Such longevity of intra-stromal suture has been observed for as long as 12 months without apparent detriment to either the graft or recipient eye.

In our opinion reconstituted collagen su­tures are satisfactory for the closure of clear corneal wounds in penetrating kerato-plasty or corneal lacerations.

SUMMARY AND CONCLUSIONS

1. The reaction to plain reconstituted col­lagen sutures was studied in both normal rabbits and in rabbits which we attempted

VOL. 64, NO. 2 COLLAGEN SUTURES 227

Fig. 6 (Brumback and McPherson). Suture struts present six months after operation excite little corneal reaction.

to sensitize to the suture. Little difference was noted in the reaction to sutures or the disappearance of extracorneal portions of the sutures in these two groups of animals.

2. Collagen sutures were used in pene­trating keratoplasties on 16 human eyes with satisfactory results.

3. Minimal reaction was present to colla­gen sutures in both human and rabbit cor­neas.

4. Collagen sutures were found to be sat­isfactory for keratoplasty ; demonstrated ex­cellent handling characteristics, excited little

tissue reaction and possessed low antigenici-ty.

1110 West Main Street

ACKNOWLEDGMENT

Technical assistance was provided by Miss San­dra Strickland and Mr. William Brown.

REFERENCES

1. Guthrie, Douglas, Lord Lister. Edinburg, E. and S. Livingstone, Ltd., 1949.

2. Miller, J. M., Zoll, D. R., Brown, E. O. and Howard, F. : Clinical observations on the use of an extruded collagen suture. Arch. Surg. 88:167, 1964.

3. Miller, J. M., Bulls, A. A. Jr. and Howard, F. : Use of a plain extruded collagen suture : Clinical observations. Arch. Surg. 90:385, 1965.

4. McPherson, S. D., Jr. and Young, J. A. : Extruded collagen sutures in cataract surgery. Arch. Ophth. 73 :463, 196S.

5. Regan, E. F. and Dunnington, J. H. : Colla­gen sutures in cataract surgery: Clinical and ex­perimental observations Tr. Am. Ophth. Soc. 64:39, 1966.

6. Van Winkle, W., Jr. : Physical, chemical an biologic properties of absorbable collagen sutures. Sommerville, New Jersey, Ethicon, Inc.

7. McPherson, S. D., Jr.: The use of absorb­able sutures in surgery of the cornea. Tr. Am. Ophth. Soc. 57:700, 1959.

8. Taylor, S. R., Jr., McPherson, S. D., Jr. and Peacock, E., Jr. : The experimental use of rat tail tendon as corneal suture material. Am. J. Ophth. 56:549, 1963.

9. Freund, J. : The effect of paraffin oil and mycobacteria on antibody formation and sensitiza-tion. Am. J. Clin. Path. 21:645, 1951.

10. Steffen, C, Timpl, R. and Wolff, T. : Immu­nology and specificity of collagen: 11. Investiga­tions about the specificity of collagen and its de­rivatives by hemagglutination and hemagglutinin inhibition of anticollagen and antiparent gelatin immune sera. J. Immunol. 93:656, 1964.

11. Watson, R. F., Rothbard, S. and Vanamee, P. : Antigenicity of rat collagen. J. Exp. Med. 99:535,1954.