Cold Plasma Welding System for Surgical Skin Closure: In Vivo Porcine Feasibility Assessment

Background. Cold plasma skin welding is a novel technology that bonds skin edges through soldering without the use of synthetic materials or conventional wound approximation methods such as sutures, staples, or skin adhesives. The cold plasma welding system uses a biological solder applied to the edges of a skin incision, followed by  the application of cold plasma energy. The objectives of this study were to assess the feasibility of a cold plasma welding system in approximating and fixating skin incisions compared with conventional methods and to evaluate and define optimal plasma welding parameters and histopathological tissue response in a porcine model. Methods and Materials. The cold plasma welding system (BioWeld1 System, IonMed Ltd, Yokneam, Israel) was used on porcine skin incisions using variable energy parameters. Wound healing was compared macroscopically and histologically to incisions approximated with sutures. Results. When compared to sutured skin closure, cold plasma welding in specific system parameters demonstrated comparable and favorable wound healing results histopathologically as well as macroscopically. No evidence of epidermal damage, thermal or otherwise, was encountered in the specified parameters. Notably, bleeding, infection, and wound dehiscence were not detected at incision sites. Skin incisions welded at extreme energy parameters presented second-degree burns. Conclusion. Implementation of cold plasma welding has been shown to be feasible for skin closure. Initial in vivo results suggest cold plasma welding might provide equal, if not better, healing results than traditional methods of closure. 

Skin wounds and incisions are involved in almost all traumatic and surgical interventions. Wound closure techniques vary widely and include the use of many different biological and synthetic materials including sutures, staplers, and skin adhesives.^1-3 Recently, several products have been introduced to simplify skin closure and reduce infection, scarring, and other complications. These advances include dressing materials, antibiotic-coated sutures,⁴ automated suture tools, and improved topical skin adhesives.^2,5,6

Tissue welding has shown promising results as an alternative method of skin wound closure and may offer advantages over conventional methods of wound closure including a watertight tissue seal, reduction in surgical procedure time, the elimination of a foreign-body reaction to sutures or clips, and reduction of trauma.^7,8 The concept of skin soldering is relatively new. Laser soldering devices have been described for skin approximation,^9-11 although initial experience suggests additional heat formed by the process may cause thermal damage to the incision and adjacent tissue.^7,12 Despite its efficacy in wound approximation to produce both physiologically and mechanically seamless closure with tensile strength, thermal-mediated tissue fusion, unlike cold plasma technology, is not cost effective and has gained only limited acceptance.¹³ 

Advancements in laser welding have been reported within the literature. Concentrated protein solutions used as a solder have demonstrated an increase in tensile strength.¹⁴ A study by Fried and Walsh¹⁵ that investigated laser skin welding and tensile strength of wounds found that weld strengths were significantly higher than sutured apposition strengths and, over time, continued to strengthen. Solders widely used are concentrated serum albumin solutions. As albumin is the prominent component of plasma, this may be beneficial in the process of welding tissue.⁸ In fact, albumin solder has been reported to improve the strength of the weld, decrease the incidence of thermal damage, and generally produce a more steadfast weld.¹⁶ However, studies^17,18 have demonstrated albumin to be an immunological risk.Nevertheless, Bleustein et al¹⁶ report albumin has been used as a tissue expander for years. 

Cold plasma tissue welding is a novel technology that bonds soft tissues without the use of synthetic materials. Recent technological advancements have enabled the use of a low-power cold plasma jet welding device. This device activates albumin-based adhesives and achieves approximation and fixation of skin incisions and wounds without causing any additional thermal damage to the incision or adjacent tissues. Therefore, the purpose of this study was to assess the feasibility of a cold plasma-based welding system in approximating and fixating skin incisions compared with conventional suture-enabled skin incision closure and to evaluate and define optimal plasma welding parameters and histopathological tissue response in a porcine model. 

Device description
The BioWeld1 System (IonMed Ltd, Yokneam, Israel) is a cold plasma welding system that generates an atmospheric plasma jet that activates an albumin-based adhesive to allow skin incision closure. Helium (He), a nontoxic, inert, monatomic gas belonging to the noble gases, is ionized by radio frequency radiation, transforming it to plasma. The He plasma flows out of the probe and interacts with an albumin-based adhesive applied to a skin incision. Due to the plasma’s unique chemical and electrical characteristics, the albumin denaturizes¹⁹ while fixating skin edges in an approximated union of the wound. 

Albumin, a major protein found in the blood plasma, was used in the form of bovine serum albumin (BSA), which is derived from cows and commonly used in in vitro biological studies. The BSA is biodegradable and disappears from the incision site shortly after completion of the procedure. The study used BSA in a powder form  (Biological Industries, Kibbutz Beit-Haemek, Israel) infused with distilled water. Two different concentrations of albumin solutions were used: 40% and 45% BSA w/w. The albumin solution was prepared at least 72 hours prior to the experiment in a 10-mL syringe, covered with aluminum foil, and stored in the refrigerator at 4°C. Thirty minutes before application, the albumin solution was removed from the refrigerator and allowed to reach room temperature.

Study design
The experiment was performed on a domestic farm pig as a porcine dermal tissue model. All procedures were conducted in accordance with protocols approved by the experimental animal facility and ethical committee of the Technion – Israel Institute of Technology, Haifa, Israel.  Anesthesia was administered to the animal intravenously using ketamine (20 mg/kg) followed by inhaled isoflurane 1% to 2%.  Twenty-two depth incisions were performed under sterile conditions by a single operator. Twenty were cold plasma soldered, and the remaining 2 incisions, serving as a control group, were sutured. The incisions were made on the pig’s dorsal skin. Eighteen incisions were 20-mm long, 1 incision was 40 mm, and 3 incisions were 80 mm.  All incisions spanned the full thickness of the dermis. Each closure method (in terms of soldering parameters) was performed at least twice by the same operator (Table 1). The 2 control-group incisions were sutured in layers using 3 subcutaneous Vicryl 3-0 sutures (Ethicon, Somerville, NJ) and intradermal nylon 3-0 (Ethicon) continuous suture. The incisions were immediately covered with Steri-Strips (3M, St. Paul, MN). The 20 remaining incisions were closed with the BioWeld1 cold plasma welding system (Figure 1) as described below. No device was used to approximate incision edges except minimal manual stretching from the distal edges of the skin incision along the weld. The 45% BSA solution was topically applied (only externally) to the edges of the skin in a uniform layer along the incision. Cold plasma welding was performed in a vertical zigzag pattern; the incisions were immediately covered with adhesive skin closures.

Upon closure of the wound, 1 incision was closed with low power (4.32 watts) and medium gas-flow parameters. These were the baseline parameters for the welding procedure. Two incisions were welded using high gas flow, while another 2 incisions welded with low gas flow.  While most of the incisions were welded using 45% w/w (0.45 g of albumin powder to 0.55 g of water) concentration of BSA, 2 incisions were welded using the 40% concentration of BSA to estimate the qualities of welding with this concentration and the immediate acute tensile strength of welding. To evaluate the feasibility of longer incisions, 4 incisions were made up to 80 mm in length, 4 times longer than the previous incisions. Two standard 20-mm incisions were welded with a different technique that also used BSA in the subcutaneous layer. A 1-mm gap between wound edges (Figure 2) was advertently left; albumin was applied to the dermal side of the skin edge and subsequently welded, followed by welding the skin edges while another layer of albumin was applied. Two additional incisions were welded at a faster rate using higher energy parameters. The welding tips were in a dielectric breakdown discharge (DBD) configuration. Two kinds of DBD tips were used: round tip and wide tip. According to the manufacturer, the wide tip provides different and possibly favorable plasma dispersion because it allows a swift, efficient application over larger incisional surfaces. Two incisions were welded using the DBD wide tip. 

On the fourth postoperative day, the pig was examined for evidence of dermal thermal damage and macroscopic evidence of other complications such as wound dehiscence. The classic signs and symptoms of infection (ie, erythema, edema, heat, and purulence²⁰) were assessed. In addition, digital pictures of the incisions were taken with a PowerShot A2400 (Canon, Tokyo).

On the seventh postoperative day, the incisions were examined again, digital pictures were taken, samples of skin incisions and adjacent untreated skin tissue were harvested, and 26 skin samples were sent for histological examination. Incisions treated with extreme welding parameters with macroscopic evidence of thermal skin damage were excluded. Up to 3 histological slices were taken from each incision that presented macroscopic changes. A total of 30 slices were prepared (Table 2). Subsequently, the pig was euthanized as mandated by the ethics committee due to the number of cuts, their area and length, and the improbable closure of the wounds after the excisions. 

Tissue samples were fixed in 10% formaldehyde solution, routinely processed, and stained with hematoxylin and eosin for histological examination. Histological examination and evaluation was performed by a single pathologist in a blinded, randomized manner. Histological examination included evaluation of the following parameters: epidermal changes and integrity, the presence of thermal damage (encrustation or skin necrosis), the presence of surface albumin, and abnormal reepithelization in the form of hyperplasia. Dermis and subcutaneous tissue were examined for the presence or absence of necrosis, edema, and granulation tissue along the incision line. A semi-quantitative grading scheme incorporating 5 grades (0 = no change, 1 = minimal change, 2 = mild change, 3 = moderate change, 4 = marked change) was used to evaluate the extent of the lesions in the sections. 

All incisions to the porcine model healed uneventfully with no signs of bleeding, infection, or surgical wound dehiscence. Incisions welded with high energy and gas-flow parameters presented macroscopic evidence of thermal skin damage in the form of second-degree burns.

Using the cold plasma welding system, the time required for wound closure ranged between 49 seconds for fast welding and up to 125 seconds for 1-mm gap incision welding. The average time was 79 seconds. Intradermal suture-aided skin closure required between 48 and 102 seconds, with an average time of 75 seconds. On the seventh postoperative day, scars of plasma welded (except for extreme flow parameters) looked similar to sutured incisions in terms of skin alignment and approximation, wound redness, and crust. On macroscopic appearance, the epidermal closure was satisfactory. Incisions treated using extreme flow parameters had up to second-degree burns, which manifested as a red-purple halo around the treated area (Figure 3). 

Microscopic findings
Histological assessment (Table 3) revealed no evidence of epidermal integrity damage in all examined incisions. There was no evidence of residual BSA in incisions, including above the epidermis and within the incision itself. A layer of encrustation was found in most of the epidermal layers of welded or sutured incision; however, this ranged from minimal to mild encrustation. Only 2 incisions had an encrustation layer, and these incisions were cold plasma welded and not sutured. Furthermore, no epidermal necrosis was found in the soldered incisions. 

The dermis layer showed no signs of thermal damage in all welded incisions. In a single incision welded with the wide tip and low-energy parameters, minimal edema was detected in the superficial dermis while the subcutaneous layer was unaffected in all histological skin samples. In addition, the incision line was filled by maturing granulation tissue. No evidence of delayed healing process was found in either the welded or sutured incisions. The lowest amount of granulation tissue (grade 1) was present in incisions welded with the wide DBD tip. The subcutaneous layer was only marginally affected by the plasma welding. However, evaluation of the subcutaneous layer in the sutured incisions showed some distortion (Figure 4A-D, 4E-H). 

This study sought to assess the feasibility of cold plasma-based welding system in approximating and fixating skin incisions compared with conventional methods and to evaluate and define optimal plasma welding parameters and histopathological tissue response in a porcine model. These results demonstrate that the cold plasma welding method of wound closure has the potential to produce comparable, if not better, results than conventional suturing methods. This includes both macroscopic and microscopic healing and scarring. 

Interestingly, incisions lines of welded wounds were evenly approximated. This may be due to the absence of tension points as seen with sutured wounds. Histologically, the assessment revealed no evidence of damage to the integrity of the epidermis in any of the welded incisions. Furthermore, no histological evidence of BSA residue was present either above the epidermis or inside the welded incisions. 

As reported in the results, the sutured incisions in the subcutaneous layer showed a distortion. In the long term, this distortion may yield a wider scar and has the potential to develop abnormal scar formation. Distortions in the restored layers, such as those formed by a reaction to a foreign material left in the tissue, and physical pressure on the tissue, can have a negative effect on wound healing.²¹ The authors found in the current and preliminary experiments that distortions existed in the sutured incisions. Because cold plasma welding requires minimal contact with the outer skin, significantly less interference is experienced, which promotes closure of the incision and an uninterrupted healing process. 

By using albumin as a welding medium, adjusting flow and energy parameters, and using a wide tip for the welding system, thermal damage was avoided. When albumin was used, thermal damage was mostly a byproduct of 2 factors: time and power (both of which were significantly reduced). Denaturated albumin adheres to the epidermal skin layer and forms a bond that contributes to the stability of the incision and to the immediate and long-term healing process. A natural biocompatible substance, albumin attenuates the amount of thermal damage⁷ by forming a coagulum buffer that subsequently disappears during the healing process via biodegradation.¹⁹ In addition to the welding functionality, plasma is considered an efficient disinfectant^22,23 during welding; however, this was not assessed in this trial. The use of the wide DBD tip improves plasma behavior on wider incision areas, which facilitates better temperature control and a swift closure avoiding the risk of tissue thermal damage. 

Unlike other tissue approximation methods, cold plasma welding joins incised tissue in an approximated union while avoiding burns, necrosis, and the use of synthetic or potentially irritating materials. It uses a low-heat delivery method aimed at the preservation of the adjacent tissue structure and functionality. Cold plasma welding enables the normal wound healing process of primary closure and complete adaptation of skin tissue layers without the use of synthetic materials within the tissue. 

Of note, surgical smoke inhalation remains an occupational hazard in operating theaters and poses significant chemical and biological dangers.²⁴ The smokeless capability of the tissue welding technology offers an important benefit to patients and health professionals by enabling wound closure without inhalation of toxins and viruses.^25-27 While thermal tissue destruction exposes surgic- al personnel to smoke plumes, the cold welding system offers a smoke-free environment, reducing potential occupational hazards. In addition, there is an added benefit of reducing the potential and incidence of needle stick injuries from suture needles. 

The average time to close an incision using the cold plasma system was 79 seconds (49–125 seconds), which was comparable to the average 75 seconds (48–102 seconds) for traditional methods of suturing. It is therefore possible to reduce incision closure time by 35% without compromising the wound union, which is significant in cases of multiple, longer incisions. This is related to skill acquisition as plasma application was reported to be mastered swiftly compared with sutured skin closure, which requires more training and surgical skill.  

Due to the nature of this experiment, a blinded assessment of the macroscopic immediate results could not be performed since the results could easily differentiate cold plasma welding and skin sutures. However, the microscopic results were blinded and evaluated by a single pathologist who was able to impartially evaluate the results after they were blinded. Therefore, further long-term blinded macroscopic results evaluations should be completed. Possible limitations of this study include the fact that study parameters were based on small, linear incisions, a clean environment, avoiding tension on skin edges, and a short follow-up period for study feasibility. A longer follow-up period is necessary for the assessment and evaluation of possible long-term complications, scar formation, inflammatory response, quality, and tensile strength. Longer and deeper incisions should be evaluated with and without tension on incision edges to further elucidate the recommended treatment parameters. Despite these limitations, the encouraging positive results of this feasibility study mandates further investigation that evaluates alternatives to traditional skin closure techniques.

Cold plasma welding could prove beneficial in providing optimal conditions for tissue healing, expediting wound closure, and attaining positive outcomes in primary wound closure that is physiologically and mechanically seamless and provides sufficient tensile strength.