Skin closure by Nd: YAG laser welding

Skin closure by Nd:YAG laser welding R. Patrick Abergel, M.D.,* Richard F. Lyons, M.D., Rodney A. White, M.D., Gary Lask, M.D., Lois Y. Matsuoka, M.D.,** Richard M. Dwyer, M.D., and Jouni Ditto, M.D., Ph.D. Torrance, CA, and Springfield, IL Skin incisions 6 mm in length were made on the backs of hairless mice. Control wounds were closed with interrupted 5-0 Prolene sutures, and experimental wounds were approximated and closed by laser welding using a Nd: YAG laser. The wounds were examined daily, and specimens were excised at weekly intervals for histopathologic study, transmission electron microscopy, tensile strength determination, and type I collagen-specific messenger ribonucleic acid measurements. The laser-welded wounds demonstrated rapid healing. Histologic study showed a functional scar tissue at day 7 and a minimal residue of the original wound at day 14. Tensile strength of the control and experimental wounds was similar at all time points. The levels of collagen-specific messenger ribonucleic acid were significantly higher in the sutured group in comparison with the laser group or with normal control skin. This study suggests that laser welding might have advantages over suturing, since the former is sterile and nontactile, does not require introduction of foreign material into the wound, and provides subjectively improved cosmetic results. (J AM ACAD DERMATOL 14:810-814, 1986.)

Conventional technic for closure of cutaneous wounds entails approximation of the wound edges by sutures or staples. In most instances, healing and scar formation are satisfactory. However, suturing technics may lead to various complications, From the Departments of Medicine and Surgery, University of California at Los Angeles School of Medicine, the Divisions of Dermatology and Vascular Surgery, Harbor-UCLA Medical Center, Torrance, and the Department of Medicine, Division of Dermatology, Southern Illinois University School of Medicine, Springfield.** Supported in part by grants AM-284S0, AM-35297, and OM-28833 from the U.S. Public Health Service, National Institutes of Health, and by Laser Endoscopy Medical Group, Inc., Los Angeles, CA. Presented at the Annual Scientific Meeting, Southern California Chapter, American College of Surgeons, Coronado, CA, Jan. 1820, 1985, and at the Fifth Annual Meeting of the American Society for Laser Medicine and Surgery, Orlando, FL, May 27-29, 1985. (Published as an abstract in Lasers Surg Med 5:180, 1985.) Reprint requests to: Dr. R. Patrick Abergel, Division of Dermatology, Harbor-UCLA Medical Center, 1000 West Carson SI., Torrance, CA 90509/213-533-2217. *Recipient of the 1984 Ken Burdick Memorial Fellowship from Syntex, awarded by the Dermatology Foundation.

810

such as foreign body reaction, development of keloid or hypertrophic scar, and infections. Recently, lasers have been used in surgery for removal of tissue by cutting and vaporization. 1 Since laser surgery is anoncontact, sterile, bloodless technic, it may have clear advantages over conventional surgery. Furthermore, laser energy can be precisely delivered through fiberoptics or a microscope. 2 In our study, we have further developed the concept of wound closure by laser welding. Specifically, we have examined the applicability of an Nd: YAG laser for wound closure. Laser-induced wound healing was studied clinically and by histopathologic and electron microscopic technics. Mechanical properties of the scar were determined by tensile strength and extensibility measurements. Furthermore, biochemical analyses were performed through assay of type I collagen-specHic messenger ribonucleic acid in the wound as an index of collagen synthesis.

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Skin closure by Nd:YAG laser 811

Fig. 1. Skin welding by Nd: YAG laser. Full-thickness wounds, 6 mm in length, were produced on the backs of hairless mice (A). The wounds w~re closed either by an Nd: YAG laser (B, right wound) or by a Prolene suture (B, left wound). C, Appearance of the wounds at day 21.

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Table I. Extensibility of skin wounds closed by Nd: YAG laser or suture Extensibility* in wounds

No. of days after closure

Closed by laser

Closed by suture

7 21

52.0 56.8

81.3 76.9

*Calculated as the difference between the initial length of the skin strip and the length at breaking point; the values are expressed as the percent of the unwounded control skin.

Fig. 2. Transmission electron microscopy of the laser-

welded wound at 21 days. Note the presence of tightly packed collagen fibers in the proximity of a fibroblastic cell with active endoplasmic reticulum and multiple mitochondria.

MATERIALS AND METHODS Full-thickness skin incisions, 6 mm in length, were made on the backs of 60-day-old hairless (hr/hr) mice* that were under general anesthesia (methoxyflurane). Control wounds were closed with a 5-0 Prolene suture, and experimental wounds were closed by laser welding. The wounds were left uncovered, and the animals were allowed to move freely in their individual cages. An Nd: YAG laser, t a continuous-wave laser with a wavelength of 1,060 nm, was used to perform experimental skin closure. The power was 1.0 W, the distance from the fiberoptic handpiece to the skin was kept constant at 1.5 em, and the spot surface was 0.02 cm2 , determining an irradiance of 50 W Icm2 • For the welding procedure the wound edges were approximated with forceps, and the wounds were closed by a continuous pass of the laser beam along the wound. The pass was repeated, if necessary, until complete closure occurred. Usually, two or three passes were sufficient for closure. All welding and suturing procedures were performed by one investigator (R. P. A). The wounds were examined daily, and biopsy spec*Jackson Laboratories, Bar Harbor, ME. tEndolase, Inc., Washington, DC.

imenswereobtainedat7, 14, and21 days and processed routinely for histopathologic study (hematoxylin-eosin stain). For transmission electron microscopy, specimens were fixed in 2.5% glutaraldehyde. For tensile strength determinations, identical skin strips, 3 em long and 0.6 em wide, were excised from anesthetized animals at days 7, 14, and 21. Each skin strip contained the original wound situated in the middle transversely to the long axis of the strip. Sutures were removed prior to excision of the strips from sutured wounds and before the mechanical testing was done. At each time point, control skin, as well as laser-welded wounds, were obtained from the same animal, and three animals were tested at each time point. The tensile strength and extensibility of the wounds were determined with a tensile strength apparatus 3 that measured total load (measured in kilograms) and extension of skin (measured in millimeters) at the breaking point. For collagen analyses, type I collagen messenger ribonucleic acid levels were determined in the wounded area of skin as an index of collagen biosynthesis. For this purpose, total ribonucleic acid was isolated,4 and the type 1 collagen messenger ribonucleic acid levels were determined by dot-blot hybridizations 5 using pro-IX I (I) collagen-specific complementary deoxyribonucleic acid (eDNA) probe. 6 RESULTS

Closure of skin wounds was successfully performed with Nd: YAG laser (Fig. 1). A total of thirty animals were subjected to skin closure by laser welding. The resulting cosmetic appearance of the laser-welded wounds was better than that of the sutured wounds by subjective assessment (Fig. 1, C). No epidermal necrosis of the wound edges or infection was noted in either the laserwelded or the sutured group. Histologic study of the wounds demonstrated normal wound healing with accumulation of con-

Volume 14 Number 5, Part 1

Skin closure by Nd:YAG laser

May, 1986

A

2.0

...'" t-

:J:

813

c

B

1.5 1.0

'"

W ~ 0.5

o

5 EXTENSION (mm)

Fig. 3. Tensile strength determinations of the wounds closed by laser welding or suture at days 7 (A), 14 (B), and 21 (C). The graph depicts the load (measured in kilograms) versus the extension (measured in millimeters) of the skin strip. The top of the curve on the y axis corresponds to the load at breaking point. Dashed line, Wounds closed with laser welding. Dotted line, Wounds closed with the suture. Solid line, Unwounded control skin.

Table II. Type I collagen-specific mRNA levels in the wounds closed by Nd; YAG laser welding or suture* Proal(I) collagen mRNA levels in wounds No. of days after surgery Control§

4

10

16

Closed by laser U/JLg RNAt

I

556 ± 43 982 966 911

Closed by suture

0/4

U/JLg RNAt

177 174 164

328 ± 13 2,425 2,421 2,652

I

%:1:

739 738 809

eDNA: Complementary deoxyribonucleic acid; mRNA: messenger ribonucleic acid. *Wounds closed by laser welding or suture had biopsy specimens taken at days indicated and processed for assay of procd(I) collagen mRNA level determinations, as described in the text. tThe values are expressed as arbitrary absorbance units (U) obtained by scanning the autoradiograms of mRNA-[32PlcDNA hybrids at 700 nml fLg total RNA. *Values are percent of the corresponding control. §The controls represent unwounded skin in the corresponding mice; the values are mean ± SO of three parallel determinations.

nective tissue. Transmission electron microscopy of the laser-welded wounds revealed the accumulation of newly synthesized collagen in the proximity of fibroblastic cells with pronounced rough endoplastic reticulum and an abundance of mitrochondria, features suggesting enhanced synthetic activity (Fig. 2). Similar electron microscopic observations were made in sutured wounds (not shown). Determination of the mechanical properties of the wounds revealed minimal tensile strength at day 1, when the wounds were closed by either laser welding or suturing. Tensile strength increased progressively with time, being similar in both groups at days 7, 14, and 21. However, these tensile strength values at day 21 were only about 30% of that in unwounded skin of the same animals

(Fig. 3). At days 7 and 21, wounds approximated by suture were more extensible than wounds welded by laser (Table I). Type I procollagen messenger ribonucleic acid levels in the wounded area were also determined as an indication of collagen synthetic activity. The results demonstrated that the messenger ribonucleic acid levels in the sutured wounds were markedly elevated, up to eightfold, in comparison with levels in unwounded control skin (Table II). A lesserincrease, approximately 1. 7-fold, was noted in laser-welded wounds (Table II). DISCUSSION

The results obtained in this study with an Nd: YAG laser at 1,060 nm suggest that laser welding may provide a new and efficient method for closing

Journal of the American Academy of Dermatology

814 Abergel et al

cutaneous wounds. Previously, other tissues, such as blood vessels, have been successfully welded with various types of lasers. 7- w Morphologic examination of the laser-welded wounds demonstrated evidence of active healing similar to that noted in sutured wounds. In addition, the tensile strength of the healing wounds was identical, whether a wound was closed by laser welding or by suture. Thus, laser welding appears to lead to similar wound healing processes with corresponding tensile properties of the wound. It should be noted, however, that in this study, no-tension wounds, consisting of a single incision, were studied; the applicability of laser welding to the closure of wounds with tension remains to be tested. Previous studies have demonstrated that in most situations the control of collagen production in skin fibroblasts resides on the transcriptional leveP 1,12; thus assays of procollagen messenger ribonucleic acid levels are expected to reflect collagen biosynthesis. In this study, type I procollagen messenger ribonucleic acid. levels were mar~edly increased in wounds closed by suture, whereas the messenger ribonucleic acid levels in laser-welded wounds were increased to a lesser degree. This observation could be interpreted to reflect the possibility that less active collagen synthesis takes place in the laser-welded wounds; the active synthesis in suture-closed wounds may resuIt from foreign body reaction and/or more extensive injury caused by the suture material. As an end result, laser-welded wounds may have less tendency to form hypertrophic scars and keloids. Although laser welding of the skin appears to be promising, several potential problems can be identified. For example, incomplete closure of the wound could result in immediate dehiscence, or excessive irradiation might cause burn. Thus accurate energy measurement is of critical importance for the success of the laser welding, and further development of automated and computerassisted welding devices may allow more precise delivery of the energy. An important observation in this study was that the cosmetic appearance of laser-welded wounds was significantly better than in sutured wounds assessed subjectively. Specifically, suture tracks present in the suture-closed wounds were absent from the laser-welded wounds. This feature ofla-

ser welding is of particular interest in cosmetic surgery. The mechanisms of laser welding are not apparent at this point. One could speculate activation of the tissue reactions necessary for normal wound healing, such as fibrin clot formation. Irrespective of the mechanism, skin closure by laser welding appears to have several advantages over conventional suturing technics: the laser welding is sterile and requires less tissue manipulation without introduction of foreign material into the wound, thus limiting the risk of infection and foreign body reaction. Nevertheless, longer follow-up and human trials are needed to attest to the ultimate benefit of wound closure by laser welding. REFERENCES 1. Hall RB: Incision of tissue by CO, laser. Nature 232: 131132, 1971. 2. Goldman L: The biomedical laser: Technology and clinical applications. New York, 1981, Springer-Verlag New York, Inc. 3. Coulson WF, Carnes WH: Cardiovascular studies on copper-deficient swine. II. Mechanical properties of the aorta. Lab Invest 11:1316-1321, 1962. 4. Chirgwin JM, Przybyla AE, McDonald RJ, Rutter WJ: Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18:52945299, 1983. 5. Abergel RP, Pizzurro D, Meeker CA, et al: Biochemical composition of the connective tissue in keloids and analyses of collagen metabolism in keloid fibroblast cultures. J Invest Dermatol 84:384-390, 1985. 6. Chu M-L, Myers JC, Bernard MP, et al: Cloning and characterization of five overlapping cDNAs specific for human procl.! (I) collagen chain. Nucleic Acids Res 10:5925-5934, 1982. 7. White RA, Abergel RP, Klein S, ct al: Biological effects of laser welding on vascular healing. Lasers Surg Med 5:146, 1985. (Abst.) 8. White RA, Abergel RP, Klein SR, et al: Laser welding of venotomies. Arch Surg. (In press.) 9. Dew DK, Serbent R, Hart WS, ct al: Laser-assisted microsurgical vessel anastomosis techniques: The use of argon and CO, lasers. Lasers Surg Med 3: 135-137, 1983. 10. Jain KV: Sutureless microvascular extra-intracranial anastomoses with Nd: YAG laser. Lasers Surg Med 4:311-316, 1984. 11. Focht RJ, Adams SL: Tissue specificity of type I collagen gene expression is determined at both transcriptional and posttranscriptional levels. Mol Cell BioI 4: 1843-1852, 1984. 12. Uitto J, Perejda AJ, Abergel RP, et al: Altered steady state ratio of type IIIII procollagen mRNAs correlates with selectively increased type I procolJagen biosynthesis in cultured keloid fibrosis. Proc Natl Acad Sci USA. 82:5935-5939, 1985.