Publication InformationAbstract Title: Helium Plasma Skin Regeneration: Evaluation of Skin Tissue Effects in

a Porcine Model and Comparison to Nitrogen Plasma Skin RegenerationJournal: Lasers in Surgery and Medicine, Published 06 October 2019

Author: J. David Holcomb, MD, Adrienne Schucker, DVM, DACVP

APYX MEDICAL DISCLOSURESFINANCIAL DISCLOSURESJ. David Holcomb is a consultant, grant recipient, and clinical research investigatorwith Apyx Medical and receives compensation in the form of hourly compensationand grants.CLEARED INDICATIONS & MANUFACTURING DISCLOSURES The use of the terms “skin rejuvenation” in relation to the use of the Renuvion/J-

Plasma technology has not been approved or cleared by the FDA. The dermalresurfacing indication for use discussed has not been approved or cleared by theFDA.

Apyx Medical wants to present you with current scientific discourse. Specific usageoutside of the general cleared indication may not be safe or effective. Renuvion/J-Plasma has received a general clearance and has not been determined to be safeor effective for use in any specific indication or anatomical location including skintightening indication referenced in this article.

Apyx Medical manufactures and owns the Renuvion/J-Plasma technologydiscussed in this article.

(Continued on next page)

The following document may only be disseminated with this disclaimer cover page attached

Publication InformationAbstract Title: Helium Plasma Skin Regeneration – Evaluation of Skin Tissue Effects in

a Porcine Model and Comparison to Nitrogen Plasma Skin RegenerationConference: American Society for Laser Medicine & Surgery 2019 Annual Conference

Chapter Title: Helium Plasma-Driven Radiofrequency in Body ContouringAuthor: J. David Holcomb, MD

APYX MEDICAL DISCLOSURES, CONTINUEDRISKS: Risk associated with the use of Renuvion/J-Plasma for skin/dermal rejuvenation mayinclude: unintended burns, temporary nerve injury, pain, tenderness, itching, bleeding,bruising/hematoma/seroma, allergic reaction, hypersensitivity to the treatment (resulting inerythema, swelling, induration and/or urticaria), temporary or permanent post-inflammatory hyperpigmentation, telangiectasias, skin overheating/burn, hypertrophicscarring, hypopigmentation, calcification, discoloration/temporary or permanenthypopigmentation, loss of correction, vessel laceration or occlusion, abscess at treatmentsite which may result in induration and/or scar formation, and prolonged wound healing.There may be additional risks associated with the use of other devices along with Renuvionand there may be an increased risk for patients who have undergone prior surgical oraesthetic procedures in the treatment area. As with any procedure, individual results mayvary. As with all energy devices there are inherent risks associated with its use, refer to theIFU for further information.INTENDED USE DISCLOSURE: The Renuvion®/J-Plasma® Precise Open Handpiece is intendedto be used with compatible J-Plasma electrosurgical generators for the delivery ofradiofrequency energy and/or helium plasma for cutting, coagulation and ablation of softtissue during open surgical procedures. The Renuvion®/J-Plasma® Precise & Precise Open®Handpieces are compatible with electrosurgical Generators BVX-200H, and BVX-200P. Referto the Instructions for Use for the currently approved or cleared indications.

The following document may only be disseminated with this disclaimer cover page attached

Lasers in Surgery and Medicine

Helium Plasma Skin Regeneration: Evaluation of SkinTissue Effects in a Porcine Model and Comparison toNitrogen Plasma Skin RegenerationJ. David Holcomb, MD

1* and Adrienne Schucker, DVM, DACVP2

1Division of Facial Plastic Surgery, Holcomb—Kreithen Plastic Surgery and MedSpa, Sarasota, Florida2Department of Pathology, American PreClinical Services, Minneapolis, Minnesota

Background and Objectives: Helium plasma skin re-generation (PSR) is a novel skin rejuvenation technologywith significant differences compared with nitrogen PSRtechnology but that may exert similar skin tissue effects.Study objectives included a comparison of acute andchronic skin tissue changes among the two plasmas in aporcine animal model.Study Design/Materials and Methods: In this study,both helium and nitrogen gas plasmas were used to treatthe dorsal skin of Yorkshire cross mini pigs with 20%(8.6 J/cm2) and 40% (17.8 J/cm2) power helium plasmasingle pass treatment (4 liter gas flow, continuous energydelivery, and linear non‐overlapping passes) comparedwith high energy nitrogen plasma double pass treatment(PSR3 @ 14.1 J/cm2: 4.0 J, 2.5 Hz pulse rate, overlappinghorizontal, and vertical passes). Acute and chronicskin contraction, maximum acute depth of injury andchronic reparative healing depth were assessed alongwith representative histopathology in each treatmentparadigm.Results: High‐energy nitrogen plasma treatment exhibitedgreatest mean depth of acute tissue injury 4 hours post‐treatment whereas helium plasma treatment exhibitedgreater acute skin tissue contraction. Then, 20% and 40%power helium plasma treatment results were each verysimilar among animals as a percentage of nitrogen plasmatreatment results for both depths of acute tissue injury andacute skin tissue contraction. Mean depths of reparativetissue healing were similar among treatment paradigms 30days after treatment with significant intra‐ and inter‐an-imal variability observed within each treatment paradigm.Thirty‐day mean skin tissue contraction was greater forhelium plasma treatment; however, the data varied sig-nificantly between animals in all paradigms. Histopatho-logic tissue evaluation after 30 days showed similar findingsamong the treatment paradigms with epidermal hyper-plasia, flattening of rete ridges and with regenerativegranulation tissue expanding the superficial and papillarydermis.Conclusions: This study demonstrates modestly reduceddepth of the thermal effect, greater skin tissue contractionand similarity of acute and chronic histopathological findingsfor helium plasma when compared with nitrogen plasma in a

porcine animal model. © 2019 The Authors. Lasers inSurgery and Medicine published by Wiley Periodicals, Inc.

Key words: nitrogen plasma; helium plasma; radio-frequency; skin regeneration; coagulation; impedance;energy density

INTRODUCTION

Nitrogen plasma skin regeneration (PSR) remainsa viable option for skin rejuvenation. The unique heatsignature and healing profile of nitrogen plasma results ina non‐ablative (at time of treatment), non‐chromophore‐dependent thermal wound with initial preservation ofthe upper layers of desiccated skin tissue that serve as anatural biological dressing during the skin’s regenerativehealing process [1,2]. The lack of an open wound, rela-tively rapid epidermal recovery (typically within 7 daysafter treatment), non‐fractionated (effectively full field)energy delivery and suitability for diverse skin types areamong the more significant clinical merits of this skinrejuvenation treatment [3,4].

Nitrogen PSR histological studies (porcine, human)showed significant neo‐collagenesis and a correspondingreduction in elastosis in the upper dermis [4–6]. Clinicalevaluation of nitrogen PSR treatment effectiveness dem-onstrated benefits in the appearance of treated skin withimprovements in dyschromia, photodamage and skintexture including reduction of acne scarring and rhyti-dosis [1,2,4,7–9]. Complications resulting from nitrogenPSR treatment include focally delayed healing, post‐in-flammatory hyperpigmentation, and rare hypertrophic

© 2019 The Authors. Lasers in Surgery and Medicine published by Wiley Periodicals, Inc.

This is an open access article under the terms of the CreativeCommons Attribution‐NonCommercial‐NoDerivatives License,which permits use and distribution in any medium, provided theoriginal work is properly cited, the use is non‐commercial and nomodifications or adaptations are made.

Accepted 20 September 2019Published online in Wiley Online Library(wileyonlinelibrary.com).DOI 10.1002/lsm.23167

*Correspondence to: J. David Holcomb, MD, Division of FacialPlastic Surgery, Holcomb—Kreithen Plastic Surgery and MedSpa,Sarasota 34237‐6045, FL.E‐mail: drholcomb@sarasota-med.com

scarring events [1,4,10]. Despite treatment protocols thatinclude darker skin types permanent hypopigmentationhas not been reported.Benefits of nitrogen PSR include relatively quick

healing, low incidence of complications, suitability for

intermediate skin types (e.g., Fitzpatrick III, IV),full‐field treatment of dyschromia and photodamage,diversity of protocols for various skin conditions, highpatient acceptance and the potential for improvement ofrhytidosis. One significant disadvantage, however, is

TABLE 1. Comparison of Thermal and Physical Properties of Helium and Nitrogen Gas Plasmas Used in thisStudy

Helium plasma Nitrogen plasma

Molar heat volume (J/mol·K) 12.4 19.9Plasma generation Direct discharge Local dischargeTop‐down thermal conduction Yes YesIn‐depth joule tissue heating Yes NoPlasma beam diameter at target 3mm 6mmEnergy delivery Continuous (or pulsed) Pulsed (1.0–2.5 Hz)Study treatment energy (Watts or J/s) 20% power= 3.2 4 Joules @ 2.5 Hz= 10

40% power= 6.6Energy density (J/cm2) 20% power= 8.6 14.1

40% power= 17.8

The study treatment energies for the helium plasma soft tissue coagulation device were determined with calorimetry performed withcontinuous energy delivery. Because energy delivery with the helium plasma device is dynamic (treatment tip continuously moving over thetissue) an average tip velocity of 1 cm/s was used to determine tissue area treated in 1 second (3mm spot width× 10mm length) withresulting oval surface area of 0.37 cm2 used in the helium plasma energy density calculations.

Fig. 1. Acute tissue injury. (A) Control, hematoxylin and eosin (H&E) stain, ×10magnification. (B) Helium plasma 20% power (single pass), Masson’s trichrome stain,×10 magnification. (C) Helium plasma 40% power (single pass), Masson’s trichrome stain, ×10magnification. (D) Nitrogen plasma PSR3 (4.0 J), Masson’s trichrome stain, ×10magnification. Scale bars = 0.1 mm. Vertical black bars designate depth of dermal injury,extending into the papillary and reticular dermis. Scarlet stained tissue in 1B, C, D indicatesdenatured tissue characterized by coagulative necrosis.

2 HOLCOMB AND SCHUCKER

that reduction of moderate to severe rhytidosis remainsinferior to conventional full‐field, multi‐pass, ablativeCO2 or erbium YAG laser skin resurfacing. A novel,previously Food and Drug Administration (FDA) clearedhelium gas plasma soft tissue coagulation device(initially developed for surgical use) with potential forskin resurfacing applications was evaluated for similartreatment outcomes versus nitrogen PSR (the positivecontrol) prior to initiating clinical studies—includinginitial characterization of helium gas plasma skin tissueeffects and comparison to nitrogen PSR in a live porcineskin tissue treatment model.

METHODS

The animal studies were conducted at AmericanPreclinical Services (Minneapolis, MN) using four ju-venile porcine Yorkshire cross animals. The animalstudies were conducted at Americal Preclinical Servicesusing four juvenile porcine Yorkshire cross animals,with two animals survived four hours following treat-ment and two animals survived thirty days followingtreatment. Thirty‐six test locations were identified andmarked along the dorsum of each animal using tattooink. The dorsum was chosen for this initial study due tothe need to survive the animal for 30 days while mini-mizing the potential for additional non‐study‐relatedinjury to the treatment sites.The helium gas plasma soft tissue coagulation device

(Bovie J‐Plasma; Bovie Medical Corporation, Clear-water, FL) was evaluated in a commercially availableFDA‐approved format with energies selected based onthe depth of thermal effects observed for non‐dermato-logic tissues at much higher energy levels [11]. Heliumplasma energy delivery consisted of single, linear,non‐overlapping passes with continuous energy de-livery and 4 L/min helium gas flow at 20% and 40%power levels correlating to energy densities below(8.6 J/cm2) and above (17.8 J/cm2) that used for nitrogenPSR treatment (14.1 J/cm2).The nitrogen PSR device (Energist Medical Group

formerly Rhytec, Swansea, United Kingdom) was eval-uated with the highest energy (4.0 J, 2.5 Hz pulse rateand energy density 14.1 J/cm2—see Table 1) double pass(PSR3) treatment protocol available to maximize depthof skin tissue effects and to enable this device to serveas the positive control for these studies. Of note, thetechnical specifications for the more recently FDA‐ap-proved device (NeoGen PSR System, 2013) are identicalto those of the original FDA‐approved device (PortraitPSR System). While highest energy, double pass ni-trogen PSR treatment is certainly not appropriate forall skin types or conditions in the clinical setting, theseparameters are appropriate for patients with lighterskin types and deep rhytids in the cheek and peri‐oralareas.All measurements, necropsies, tissue preparation,

and histological staining (Hemoatoxylin and Eosin,Masson’s trichrome) were performed by American

Preclinical Services Pathology Department. The studypathologist (co‐author AS) was blinded to the treatmentidentification information for each site during histo-pathologic slide review. This was a GLP (Good Labo-ratory Practice) study performed in compliance with thestudy protocol and amendments and Pre‐Clinical Path-ology Consulting Service standard operating proce-dures.

Dermal tissue injury was evaluated for each treat-ment site with a depth of injury measured from thedermal‐epidermal junction (basement membrane) to thedeepest focus of dermal injury. This focus was measuredin the approximate center of the treatment site wherepossible with a depth of the skin tissue injury meas-urements captured using a validated ocular reticle inthe Olympus BX45 microscope with each site measuredin millimeters. The treatment site squares had beenpreviously outlined with blue tattoo ink. The linear di-mensions of the upper and lower horizontal sides andthe right and left vertical sides were measured in mil-limeters with the horizontal and vertical measurementsaveraged for each square and then multiplied to obtainaverage surface area dimensions for each square. Rawdepth data and treatment site dimensions obtained ineach animal consisted of eight treatment sites for the

Fig. 2. Graph: Relative depth of acute tissue injury (percent) forhelium plasma versus nitrogen plasma control (y‐axis) withtreatment paradigm on x‐axis and mean depth of tissue injury inmicrometers below basement membrane also shown. Note thegreater depth of effect for the nitrogen plasma treatmentparadigm and non‐overlap of helium and nitrogen plasmastandard deviation error bars. While the mean depth of tissueinjury was greater in one animal (18P0444), the relative depth ofinjury as a percentage of nitrogen plasma injury depth wasremarkably similar among the helium plasma treatmentparadigms in the two animals.

HELIUM PLASMA SKIN REGENERATION 3

helium plasma groups and the high‐energy nitrogenplasma group with the exception of only four treatmentsites for high energy nitrogen plasma in animal18P0439. These data were analyzed and converted intoa graphical format using Kaleidagraph 4.0 (SynergySoftware, Reading, Pennsylvania).

RESULTS

Acute Histopathology and Injury Depth Data

In the acute animals, nitrogen PSR treatment (doublepass or PSR3, 4.0 J) and helium gas plasma treatmentat both 20% and 40% power all resulted in acutethermal injury histopathologically evidenced by ne-crotic epidermis adherent to basement membrane anddeeper dermal soft tissue coagulative (necrotic) injuryextending into the superficial reticular dermis (Fig. 1,note retention of scarlet stain in tissue heated above itsdenaturation temperature as well as the relativelysharp demarcation between the scarlet‐stained tissueabove and the airline blue‐stained tissue below with thelatter representative of non‐denatured tissue deeper inthe dermis [12]). Depth of acute tissue injury variedamong animals where high energy nitrogen plasmatreatments exhibited greatest depth and where 20% and

40% power helium plasma treatments were each verysimilar among animals as a percentage of nitrogenplasma treatment depth (Fig. 2). Acute tissue injurydepth for 20% and 40% helium plasma treatment ineach of two animals was 54% and 57% and 66%and 71%, respectively, that of nitrogen plasma.Measured acute injury depths below the basementmembrane (all sites, both animals) averaged 404 μm fornitrogen plasma treatment and 222 and 281 μm for 20%and 40% helium plasma, respectively.

Thirty‐Day Histopathology and Reparative HealingDepth Data

In the 30‐day‐survived animals, nitrogen PSR treat-ment (double pass or PSR3, 4.0 J) and helium gasplasma treatment at both 20% and 40% power all ex-hibited epidermal hyperplasia, loss of rete ridges andregenerative granulation tissue expanding the super-ficial and papillary dermis (Fig. 3). Significant intra‐and inter‐animal variability was observed within eachtreatment paradigm and mean depths of reparativetissue healing were similar among treatment para-digms at 30 days after treatment (Fig. 4). Measuredchronic reparative healing depths below the basementmembrane (all sites, both animals) averaged 360 μm for

Fig. 3. Thirty‐day reparative healing depth. (A) Control, hematoxylin and eosin (H&E)stain, ×10 magnification. (B) Helium plasma 20% power (single pass), Masson’s trichromestain, ×10 magnification. (C) Helium plasma 40% power (single pass), Masson’s trichrome stain,×10 magnification. (D) Nitrogen plasma PSR3 (4.0 J), Masson’s trichrome stain, ×10magnification with smaller inset lower left at ×4 magnification for orientation. Scale bars(except inset in D)= 0.1mm. Vertical black bars designate the depth of reparative tissue healingextending into the dermis.

4 HOLCOMB AND SCHUCKER

nitrogen plasma treatment and 283 and 272 μm for 20%and 40% helium plasma, respectively.

Acute and 30‐Day Skin Contraction Data

Representative gross treatment site photographs foracute pre‐ and post‐treatment and chronic pre‐treatmentand pre‐necropsy at 30 days are shown in Figures 5 and 6.Acute skin tissue contraction was observed in all treat-ment paradigms (Fig. 7) and averaged −5.4% among theanimals for nitrogen plasma treatment. Acute skin tissuecontraction was much greater for helium plasma treat-ments at both 20% and 40% power with low inter‐animalvariability (4–16%) as a percentage of nitrogen plasmatreatment (193–231% greater contraction at 20% powerand 258–269% greater contraction at 40% power, Fig. 7).The treatment site area reductions were also observed inall treatment paradigms at thirty days following treat-ment. Significant inter‐animal variability was observedfor all treatment paradigms at 30 days following treat-ment with greatest skin tissue contraction observed forhelium plasma treatment (Fig. 8). A potentially importantfactor impacting these results is that the survived ani-mals gained 10% body mass (and a corresponding increasein girth that resulted in an increase in treatment site

areas for the untreated control sites) during the 30‐daypost‐treatment interval—the increase in area of thecontrols was accounted for in the calculations as afurther decrease in area that had been overcome by thetreatment.

DISCUSSION

The purpose of this study was to evaluate the suit-ability of helium gas plasma for skin resurfacing pro-cedures in a pre‐clinical model and to compare thetechnology to the predicate nitrogen PSR technologythat is currently FDA cleared for that indication.Thermal injury depth measurements provided data re-lated to the safety of the helium plasma device for theapplication. Soft tissue contraction measurements pro-vided further insight as to the potential efficacy of thehelium plasma device for wrinkle reduction through thecontraction of soft tissue. Representative histology fromthe two plasmas provided a detailed comparison of theskin tissue effects at both time points.

To better understand and analyze the results of thestudy, it is important to outline differences in the mech-anism of plasma beam generation and in the exerted skintissue effects between the helium and nitrogen plasmadevices. Nitrogen PSR treatment delivers heat to thetissue through the expulsion of energy generated by ion-izing nitrogen gas through the use of microwave energy.Helium PSR treatment delivers heat to the tissue in asimilar fashion but also via electrical current travelingfrom one electrode to another through an ionized heliumgas bridge interface.

While both nitrogen and helium gases are stable,inert, ideal gases helium gas is monatomic with threedegrees of freedom for subatomic thermal energytransitions whereas nitrogen gas is diatomic with fivedegrees of freedom for such energy transitions [note twoadditional degrees of freedom (total of seven) related tovibrational energy are also found at very high temper-atures far exceeding those encountered in clinical ap-plications] [13]. The additional two degrees of freedomgive nitrogen gas greater ability to store heat energyand comparison on an equi‐molar basis (where the samenumber of atoms of helium or molecules of nitrogen arearriving at the surface per unit time and transferringthermal energy) results in a higher molar volume heatcapacity for nitrogen than for helium (Table 1: 19.9 vs.12.4 J/mol·K, respectively).

This greater heat capacity for nitrogen gas is ideallysuited for delivery of heat to the tissue in single briefpulses. The nitrogen beam applicator uses a “localdischarge” process with the ionization process confinedto the handpiece where microwaves heat a tungstenwire (susceptor) that then heats a flow of nitrogen gasas well as partially ionizing the nitrogen gas. Once theionized nitrogen gas leaves the applicator nozzle theexcited electrons move to lower energy states and emitthe characteristic yellow optical emission (Lewis Ray-leigh afterglow) as photons of a specifically visible

Fig. 4. Graph: Relative 30‐day reparative healing depth(percent) for helium plasma versus nitrogen plasma control(y‐axis) with a treatment paradigm on x‐axis and meanreparative healing depth in micrometers below basementmembrane are also shown. Although the greater depth ofeffect was observed for the nitrogen plasma treatmentparadigm, overlap of nitrogen and helium plasma standarddeviation error bars was observed in both paradigms in eachanimal. Depth of reparative healing tissue was greater in oneanimal (18P0436) for both helium and nitrogen plasmatreatment paradigms.

HELIUM PLASMA SKIN REGENERATION 5

wavelength escape and the plasma converts back to thestable gas state. Heat distribution within the nitrogenplasma pulse is Gaussian in nature and tissue heatingoccurs with top‐down thermal transfer from the flowinghot nitrogen gas as much of the nitrogen gas may nolonger be ionized at the time of thermal energy transfer.In contrast, with its lower heat capacity and unique

electrical properties helium gas is more suited for avery different energy delivery paradigm wherein he-lium plasma is created through a “direct discharge”process where a continuous electrical discharge pathexists from the tip of the handpiece (positive electrodeor cathode) to the target tissue (negative ground oranode). While a small percentage of the helium gas isionized in the non‐heated state (cold atmosphericplasma) addition of radiofrequency (RF) energy onlyincrementally increases ionization of the helium gasflow (e.g., <0.1%) but does so continuously along thebeam path down to the target tissue. RF energy is de-livered to the handpiece by the generator and used toenergize the positive electrode. When helium gas ispassed over the energized electrode, helium plasma is

generated, which enables heat to be applied to tissue intwo distinct modes.

First, top‐down thermal transfer from the flowing hothelium gas occurs with heat generated by the actualproduction of the plasma beam itself through the ioniza-tion and rapid neutralization of the helium atoms. Theneutralization of the helium atoms gives rise to thecharacteristic violet optical emission (Lewis Raleigh af-terglow). Second, as plasmas are very good electricalconductors, a portion of the RF energy used to energizethe electrode and generate the plasma passes from theelectrode to the tissue. Electrical displacement current isformed in the tissue region immediately surrounding thehelium plasma beam contact point, both across the tissuesurface and in‐depth. In one‐half cycle, current flows intothis region, accumulating charge that is withdrawn in thenext half‐cycle. The flow of current through the resistanceof the tissue creates Joule (resistive) heating.

The direct discharge process and the supply gas uti-lized by the investigative device may account for thedifferences in skin tissue effects noted in this studywhen compared with the nitrogen PSR system. The

Fig. 5. Gross photographs (animal 18P0444): (A) Forty percent of helium plasma pre‐treatment.(B) Forty percent of helium plasma 4 hours post‐treatment. (C) PSR3 pre‐treatment. (D) PSR34 hours post‐treatment. Scale bars= 5mm. Note the similarity of superficial coagulation (necroticepidermis) in (B) and (D) and the greater erythematous reaction of surrounding tissue in (D)along with grossly visible tissue contraction in (B) versus (A) and (D) versus (C).

6 HOLCOMB AND SCHUCKER

conductive helium plasma beam produced by the directdischarge process can be thought of as a flexible wire orelectrode bridge that “connects” to the tissue that rep-resents the path of least resistance to the flow of the RFenergy. The tissue that represents the path of leastresistance is typically either the tissue that is in closestproximity to the tip of the device or the tissue that hasthe lowest impedance (is the easiest to pass energythrough). As tissue is treated, it coagulates and desic-cates, and the impedance of the tissue increases. Asthis occurs, the path of least resistance is constantlychanging, and subsequently, the permissive, seeminglychaotic energy (plasma beam) branches out laterallyfrom treated, higher impedance tissue to untreated,lower impedance tissue. This results in non‐Gaussianenergy deposition with impedance‐dependent lateralthermal spread and a decrease in the depth of thermaleffect for the helium plasma device. As heliumgas has a simple molecular structure consisting ofonly two electrons it is easily ionized by low current RFenergy. Since the current of the RF energy is so low, it is

dispersed before it is able to penetrate deep intothe tissue. This allows for effective soft tissue heatingand contraction with a relatively shallow depth ofthe thermal effect. This self‐limiting nature of heliumplasma’s thermal depth of tissue effect has beendemonstrated in several other tissue types and withmuch higher energy settings than used in thisstudy [11].

Variables that may potentially impact skin tissue ef-fects of both plasma devices include handpiece velocity,treatment tip configuration and distance from the tissuesurface as well as gas flow rates, energy pulsingschemes, energy “beam” configuration and number ofpasses. The treatment tip to target tissue distance maybe of more importance for the nitrogen plasma treat-ment as an increase or decrease from the recommendedoffset distance will have a similar direct effect on thedepth of tissue injury. For the helium plasma device in-creasing the distance from the target tissue beyond aminimum electrical coupling distance (e.g., 5 mm) willcause the helium plasma RF bridge to sever resulting in

Fig. 6. Gross photographs (animal 18P0436): (A) Forty percent of helium plasma pre‐treatment.(B) Forty percent of helium plasma 30 days post‐treatment. (C) PSR3 pre‐treatment. (D) PSR3 30days post‐treatment. Scale bars= 5mm. Note the similarity of healing in (B) and (D) andambiguity of visible tissue contraction in (B) versus (A) and (D) versus (C) (see text).

HELIUM PLASMA SKIN REGENERATION 7

no tissue effect. Moving the helium plasma electrodecloser to the target tissue will not significantly increasethe depth of effect because of the more stable nature ofthe energized gas flow, continuously changing skintissue impedance that effectively expands the beam pe-ripherally (away from areas with higher impedance andtowards areas with lower impedance) and device char-acteristics that may limit energy density‐independent ofhandpiece velocity. The electrocoupling characteristicsand distance may be somewhat advantageous as holdingthe treatment tip closer to the skin surface will notsignificantly increase energy density and holding thetreatment tip beyond the electrocoupling distance willsever the radiofrequency bridge resulting in no tissueeffect—versus inverse relationship between treatmenttip distance from skin surface and resulting tissue ef-fects with the predicate device (increased/reduced tissueeffect with decreasing/increasing distance betweentreatment tip and the skin). Additional specific variablesthat may affect helium plasma skin tissue effects in-clude extrinsic factors that may affect the skin tissueimpedance (e.g., injected local or tumescent anesthesia;e.g., topical anesthetic creams, gels, ointments) and de-vice electrical configuration (e.g., reduced current lim-iting Joule tissue heating).

Available energy densities for the helium andnitrogen plasma systems overlap with maximum energydensity for nitrogen PSR (14.1 J/cm2) corresponding to apower level of approximately 32% for helium plasma(Fig. 9). Although the molar heat volume capacity ofnitrogen gas is higher than for helium gas, the heliumplasma system is capable of much higher energydensities due to its RF generator design (availablepower range from 2W up to 40W), smaller plasma beamdiameter at the target (spot size), continuous pulsetrain energy delivery (vs. maximum single‐pulse rate of2.5 Hz for nitrogen PSR) and its additional Joule tissueheating (enabled by maintenance of the helium gasplasma bridge via continuous re‐ionization of heliumalong the beam path) beyond the common top‐down orsurface tissue heating of both systems (Table 1).

Despite these perceived advantages, the depth of RFJoule tissue heating during treatment with the heliumgas plasma soft tissue coagulation device is again subjectto skin tissue impedance and device characteristics(voltage, duration of current flow). Whereas unregulatedRF energy and increased skin tissue impedance mayparadoxically result in a greater extent of skin tissue in-jury [14] the helium plasma device’s low current andcontinuous (five thousand times per second) impedancemonitoring are designed to prevent unintended electricalcurrent flow deep into the target tissue. Even so, it isestablished that full field and even fractional energy‐based skin resurfacing treatments at high energy den-sities may result in delayed healing and scarring, espe-cially in more permissive treatment areas (e.g., neck)where the skin is thinner and/or less vascular [15].

Previous study of nitrogen PSR tissue effects ina Yucatan minipig model showed quick epidermal

Fig. 7. Graph: Relative net acute change in treatment site area(percent) for helium plasma versus nitrogen plasma control(y‐axis) with treatment paradigm on x‐axis and mean change intreatment site area in millimeters squared are also shown.Greatest percent reduction in treatment site area was observedfor the 40% helium plasma treatment paradigm with similarfindings among the two animals.

Fig. 8. Graph: Relative net 30‐day change in treatment site area(percent) for helium plasma versus nitrogen plasma control(y‐axis) with treatment paradigm on the x‐axis and mean changein treatment site area in millimeters squared are also shown.Greatest tissue contraction was observed for both helium plasmatreatment paradigms and the relative area decrease was muchhigher in one animal (18P0439).

8 HOLCOMB AND SCHUCKER

recovery with preservation of rete ridge formation atlower fluences and more extensive epidermal injurywith irreversible thermal damage extending into thesuperficial dermis at higher fluences [3]; the overalldepth of irreversible tissue injury was more superficialthan that observed in this study. We believe that thedifferences in histopathology observed in this studymay be related to the enhanced depths of tissue effectsand the still relatively early tissue sampling at a re-covery timepoint of just 30 days. A longer post‐treat-ment recovery period before tissue sampling may helpresolve this question. The use of a different pig breed(Yorkshire cross) in the current study may have af-fected the depth of tissue injury findings since priorevaluation of skin thickness in these two breeds dem-onstrated that the epidermis is approximately one‐third thinner in the Yorkshire breed [16]. Differences innitrogen plasma treatment tip position relative to theskin surface may have also contributed to the relativeincreased depth of tissue injury noted in this study.Inadvertently holding the nitrogen plasma treatmenttip closer to or farther away from the skin surface di-rectly impacts the effective treatment energy observedat the skin surface, increasing or lowering energydensity, respectively. It is possible that the nitrogenplasma treatment tip position was held relatively

closer to the skin in this study. Of note, similarmovement of the helium plasma treatment tip does notsignificantly impact the effective treatment energyobserved at the skin surface unless increased treat-ment tip to skin surface distance becomes sufficient todisrupt the RF bridge interface.

CONCLUSIONS

The differences between the helium and nitrogen plasmadevices evaluated in this study result in differences in depthof thermal effect and in skin tissue contraction. The lowerdepths of thermal effect and larger percentages of skin tissuecontraction for the helium plasma device compared to theFDA cleared nitrogen plasma device point to its potentialsuitability for use in skin resurfacing procedures. Despitesignificant differences in plasma generation and character-istics and in plasma—skin tissue interaction, high energydouble‐pass nitrogen plasma and helium gas plasma treat-ment (20% and 40% power) of porcine skin (continuous en-ergy delivery, single‐pass) exhibit similar acute and chronichistopathological changes. The greater skin tissue con-traction exhibited by the helium plasma system at the lowerenergy density evaluated (40% lower than nitrogen PSR)may be explained by its unique bimodal energy delivery andits more complete full‐field energy delivery to the tissue.These effects may be compounded at the higher energydensity evaluated (20% higher than nitrogen PSR).

The helium plasma device has been available commer-cially in an FDA‐approved format for general indicationsof ablation, coagulation, and cutting of soft tissue even asthese pre‐clinical studies have been conducted. Duringthis time the device has been in use (off label) by manypractitioners for facial skin rejuvenation, both in previousgenerator design and in the updated generator designused in this study. While anecdotal clinical evidenceshows much promise for this new technology for thetreatment of facial rhytidosis, detailed study of heliumplasma skin tissue effects and a formal clinical studyevaluating the efficacy and safety are ongoing.

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Fig. 9. Graph: Helium plasma power (percent, x‐axis) versushelium plasma energy density (J/cm2, y1‐axis) with an overlay ofnitrogen plasma energy density (J/cm2, y2‐axis). The energydensity for helium plasma at 20% and 40% power isapproximately 40% less and 20% higher, respectively, than thatof nitrogen plasma. Maximum energy density for the nitrogenplasma system as tested is approximately equivalent to that ofthe helium plasma system at 32% power.

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