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The use of the tissue adhesive, methyl-2-cyanoacrylate monomer, in invertebrate surgery
JOURNAL
OF INVERTEBRATE
PATHOLOGY
12,412-414
(1968)
The Use of the Tissue Adhesive, Methyl-2-cyanoacrylate in Invertebrate Surgery1 RONALD L. TA~OR AND WILLIAM C.
Monomer,
FRECKLETON,
JR.
Center for Pathobiology, University of California, Irvine, Cdifornia 92664 Received March 11, 1968 The tissue adhesive, methyl-2-cyanoacrylate monomer, which heretofore has been used exclusively in experimental and clinical studies on higher animals and man, was demonstrated to be of value in sealing surface incisions of cockroaches, In contrast to a beeswax sealant, or no sealant at all, the tissue adhesive provided more favorable conditions for normal repair and wound-healing processes. It would seem to have unexplored possibilities in other research involving invertebrates.
INTRODUCTION
Methyl-2-cyanoacrylate monomer,2 first used as an industrial adhesive, came to the attention of medical researchers in 1960 when its potentials as a tissue adhesive became apparent. Since then it has been used almost exclusively by physicians in both experimental and clinical studies on higher animals and man. Its uses have included repairing neural anurisms, plugging bladder or urinary tube fissures, mending skin incisions, anastomosing small blood vessels, and controlling hemorrhages. It has been used, in fact, wherever nonsuture repair is desirable or necessary following surgical procedures. Some of the properties that make use of methyl-2-cyanoacrylate monomer attractive as a tissue adhesive are its biodegradability, stability, flexibility, rapid polymerization in the presence of body fluids, low toxicity, ease of application, and strength of bond relative to the inherent 1 This investigation was supported by Public Health Service Research Grant No. 5358 (to E. A. Steinhaus) from National Institute of Allergy and Infectious Diseases. 2 Methyl-2-cyanoacrylate monomer was supplied sterile in 1 ml tubes through the courtesy of Ethicon, Inc., Somerville, New Jersey.
tensile strength of adjacent tissues. (For a comprehensive review of its properties and uses see Page, 1966). Of the more than three hundred publications concerning this sealant and its homologues, none consider its use in surgery on invertebrate animals. We report here on the use of this adhesive for sealing incisions in the body wall on the Madeira cockroach, Leucophaea maderae, and suggest other possible uses for it in research involving invertebrates. MATERIALS
AND
METHODS
Cockroaches were subjected to hypothermia, which functioned primarily to immobilize them and secondarily to minimize bleeding through thickening the blood. Incisions approximately 7 mm long were made on the dorsum of the fourth and fifth abdominal segments lateral and parallel to the heart. Blood and tissue fluids were swabbed away leaving the surface clean and relatively dry. For application of the adhesive, a 2 X 12 mm strip of cellophane was picked up at one end with forceps, dipped into a drop of the adhesive (kept on a microscope slide in a petri dish), and dragged the length of the incision (Fig. 1). A quick puff of air on the incision accel-
method of application of tissue FIG. 1. Photograph, approximately actual size, demonstrating adhesive. The incision (lined here with thread for purposes of illustration) is seen through the 2 x 12 mm strip of cellophane which is used to apply the adhesive. This strip is held at one end by forceps. FIG. 2. Cross section through incision 2 days after application of the tissue adhesive. Observe mass of blood cells beneath incision. 94 x. FIG. 3. Cross section through incision 30 days after application of adhesive. Note regenerated epithelium and cuticle (arrow) beneath wound tissue. 160 x , FIG. 4. Cross section through incision 2 days after application of beeswax sealant. Note separation of both sides of incision and mass of necrotic blood cells protruding through incision. 79 X. FIG. 5. Cross section through incision 30 days after application of beeswax sealant. Note regenerated integument (arrows) beneath wound tissue. Also note separation of both sides of incision. 51 x . FIG. 6. Cross section through untreated incision 30 days after incision was made. Observe large mass of necrotic blood cells involved in the lesion and the separation of the integument. 44~. 413
114
TAYLOR
ASI)
erated polymerization of the adhesive. Minimal amounts of adhesive had to be used because it was instantly absorbed by the tissues, killing and glueing together all those it penetrated. The method described here provided just enough adhesive to seal the incision and not penterate the deeper tissues. When properly applied there was virtually no indication that a sealant had been used. For comparison, control incisions were either left unsealed or sealed with IO% beeswax in Paraplast (mp 5657°C ) using a heated glass rod. Aseptic procedures were used throughout. RESULTS
AND
DISCUSSION
Subsequent observations showed that incisions sealed with the tissue adhesive did not bleed; however, cockroaches with unsealed incisions suffered considerable blood loss, especially when the wings were laid back over the wound. Apparently juxtaposition of the wings over the incision tended to interfere with coagulation and/or to draw blood out by capillarity. Bleeding also frequently occurred from incisions treated with beeswax sealant which lacked flexibility and was easily loosened and lost from the incisions as a result of the animal’s normal body movements. Sometimes fatbody tissue was lost through the open wounds. Although not determined, use of the tissue adhesive would probably afford the cockroach better protection against infection than would no sealant or even use of the beeswax mixture. Two days and 30 days after the incisions were made, a rectangular piece of integument, including the incisions, was fixed in Zenker’s fluid, embedded in Paraplast, sectioned at 6 CL,and stained with hemotoxylin and eosin. At both time periods, the incisions sealed with tissue adhesive showed a relatively normal coaptation of the integument on both sides of the wound (Figs. 2, 3), a situation which contrasts to that of using the beeswax sealant (Figs, 4, 5) or
FRECKLETOS,
JR.
no sealant at all (Fig. 6). Apparently contraction of the musculature surrounding a fresh incision results in separation of the two sides, but the bond formed by the tissue adhesive is strong enough to counteract this pull. After 2 days, blood cells have accumulated directly beneath the wound in all cases (Figs. 2, 4), and have protruded slightly through the incision in those cases where no sealant or beeswax sealant (Fig. 4) was used. After 30 days, normal repair processes are essentially complete in all lesions (Figs. 3, 5). The epidermal cells have proliferated around the lesion, migrated under the wound tissue forming a continuous sheet of epidermal cells, and secreted new cuticle (see Wigglesworth, 1937). By this time the incision and wound tissue are completely isolated from the body cavity. In conclusion, the tissue adhesive, methyl-2-cyanoacrylate monomer, provided favorable conditions for normal repair and wound healing processes of surface incisions in the cockroach. We have used it effectively for 2 years to seal incisions in the body wall of cockroaches, and to seal off necks antiseptically in decapitation experiments. Although it undoubtedly will not prove to be the ideal material for all problems where an adhesive or sealant is required, it would seem to have unexplored possibilities in invertebrate research involving implantations, parabiotic fusions, and other experiments where it is necessary or desirable to “glue” one tissue to another. ACKNOWLEDGMENT The authors wish to express their sincere appreciation ito Miss Paula Butler for her expert technical
assistance. REFERENCES
PAGE,
R. C. 1966. Methyl-2-cyanoacrylate monomer, a biodegradable plastic tissue adhesive. Cohesive New, 6 (4), 2. WIGGLESWORTH, V. B. 1937. Wound healing in an insect (Rhodnius prolixus Hemiptera). J. Exptl. Bid., 14, 364-381.
OF INVERTEBRATE
PATHOLOGY
12,412-414
(1968)
The Use of the Tissue Adhesive, Methyl-2-cyanoacrylate in Invertebrate Surgery1 RONALD L. TA~OR AND WILLIAM C.
Monomer,
FRECKLETON,
JR.
Center for Pathobiology, University of California, Irvine, Cdifornia 92664 Received March 11, 1968 The tissue adhesive, methyl-2-cyanoacrylate monomer, which heretofore has been used exclusively in experimental and clinical studies on higher animals and man, was demonstrated to be of value in sealing surface incisions of cockroaches, In contrast to a beeswax sealant, or no sealant at all, the tissue adhesive provided more favorable conditions for normal repair and wound-healing processes. It would seem to have unexplored possibilities in other research involving invertebrates.
INTRODUCTION
Methyl-2-cyanoacrylate monomer,2 first used as an industrial adhesive, came to the attention of medical researchers in 1960 when its potentials as a tissue adhesive became apparent. Since then it has been used almost exclusively by physicians in both experimental and clinical studies on higher animals and man. Its uses have included repairing neural anurisms, plugging bladder or urinary tube fissures, mending skin incisions, anastomosing small blood vessels, and controlling hemorrhages. It has been used, in fact, wherever nonsuture repair is desirable or necessary following surgical procedures. Some of the properties that make use of methyl-2-cyanoacrylate monomer attractive as a tissue adhesive are its biodegradability, stability, flexibility, rapid polymerization in the presence of body fluids, low toxicity, ease of application, and strength of bond relative to the inherent 1 This investigation was supported by Public Health Service Research Grant No. 5358 (to E. A. Steinhaus) from National Institute of Allergy and Infectious Diseases. 2 Methyl-2-cyanoacrylate monomer was supplied sterile in 1 ml tubes through the courtesy of Ethicon, Inc., Somerville, New Jersey.
tensile strength of adjacent tissues. (For a comprehensive review of its properties and uses see Page, 1966). Of the more than three hundred publications concerning this sealant and its homologues, none consider its use in surgery on invertebrate animals. We report here on the use of this adhesive for sealing incisions in the body wall on the Madeira cockroach, Leucophaea maderae, and suggest other possible uses for it in research involving invertebrates. MATERIALS
AND
METHODS
Cockroaches were subjected to hypothermia, which functioned primarily to immobilize them and secondarily to minimize bleeding through thickening the blood. Incisions approximately 7 mm long were made on the dorsum of the fourth and fifth abdominal segments lateral and parallel to the heart. Blood and tissue fluids were swabbed away leaving the surface clean and relatively dry. For application of the adhesive, a 2 X 12 mm strip of cellophane was picked up at one end with forceps, dipped into a drop of the adhesive (kept on a microscope slide in a petri dish), and dragged the length of the incision (Fig. 1). A quick puff of air on the incision accel-
method of application of tissue FIG. 1. Photograph, approximately actual size, demonstrating adhesive. The incision (lined here with thread for purposes of illustration) is seen through the 2 x 12 mm strip of cellophane which is used to apply the adhesive. This strip is held at one end by forceps. FIG. 2. Cross section through incision 2 days after application of the tissue adhesive. Observe mass of blood cells beneath incision. 94 x. FIG. 3. Cross section through incision 30 days after application of adhesive. Note regenerated epithelium and cuticle (arrow) beneath wound tissue. 160 x , FIG. 4. Cross section through incision 2 days after application of beeswax sealant. Note separation of both sides of incision and mass of necrotic blood cells protruding through incision. 79 X. FIG. 5. Cross section through incision 30 days after application of beeswax sealant. Note regenerated integument (arrows) beneath wound tissue. Also note separation of both sides of incision. 51 x . FIG. 6. Cross section through untreated incision 30 days after incision was made. Observe large mass of necrotic blood cells involved in the lesion and the separation of the integument. 44~. 413
114
TAYLOR
ASI)
erated polymerization of the adhesive. Minimal amounts of adhesive had to be used because it was instantly absorbed by the tissues, killing and glueing together all those it penetrated. The method described here provided just enough adhesive to seal the incision and not penterate the deeper tissues. When properly applied there was virtually no indication that a sealant had been used. For comparison, control incisions were either left unsealed or sealed with IO% beeswax in Paraplast (mp 5657°C ) using a heated glass rod. Aseptic procedures were used throughout. RESULTS
AND
DISCUSSION
Subsequent observations showed that incisions sealed with the tissue adhesive did not bleed; however, cockroaches with unsealed incisions suffered considerable blood loss, especially when the wings were laid back over the wound. Apparently juxtaposition of the wings over the incision tended to interfere with coagulation and/or to draw blood out by capillarity. Bleeding also frequently occurred from incisions treated with beeswax sealant which lacked flexibility and was easily loosened and lost from the incisions as a result of the animal’s normal body movements. Sometimes fatbody tissue was lost through the open wounds. Although not determined, use of the tissue adhesive would probably afford the cockroach better protection against infection than would no sealant or even use of the beeswax mixture. Two days and 30 days after the incisions were made, a rectangular piece of integument, including the incisions, was fixed in Zenker’s fluid, embedded in Paraplast, sectioned at 6 CL,and stained with hemotoxylin and eosin. At both time periods, the incisions sealed with tissue adhesive showed a relatively normal coaptation of the integument on both sides of the wound (Figs. 2, 3), a situation which contrasts to that of using the beeswax sealant (Figs, 4, 5) or
FRECKLETOS,
JR.
no sealant at all (Fig. 6). Apparently contraction of the musculature surrounding a fresh incision results in separation of the two sides, but the bond formed by the tissue adhesive is strong enough to counteract this pull. After 2 days, blood cells have accumulated directly beneath the wound in all cases (Figs. 2, 4), and have protruded slightly through the incision in those cases where no sealant or beeswax sealant (Fig. 4) was used. After 30 days, normal repair processes are essentially complete in all lesions (Figs. 3, 5). The epidermal cells have proliferated around the lesion, migrated under the wound tissue forming a continuous sheet of epidermal cells, and secreted new cuticle (see Wigglesworth, 1937). By this time the incision and wound tissue are completely isolated from the body cavity. In conclusion, the tissue adhesive, methyl-2-cyanoacrylate monomer, provided favorable conditions for normal repair and wound healing processes of surface incisions in the cockroach. We have used it effectively for 2 years to seal incisions in the body wall of cockroaches, and to seal off necks antiseptically in decapitation experiments. Although it undoubtedly will not prove to be the ideal material for all problems where an adhesive or sealant is required, it would seem to have unexplored possibilities in invertebrate research involving implantations, parabiotic fusions, and other experiments where it is necessary or desirable to “glue” one tissue to another. ACKNOWLEDGMENT The authors wish to express their sincere appreciation ito Miss Paula Butler for her expert technical
assistance. REFERENCES
PAGE,
R. C. 1966. Methyl-2-cyanoacrylate monomer, a biodegradable plastic tissue adhesive. Cohesive New, 6 (4), 2. WIGGLESWORTH, V. B. 1937. Wound healing in an insect (Rhodnius prolixus Hemiptera). J. Exptl. Bid., 14, 364-381.