Crosslinked hyaluronic acid dermal fillers: a comparison
`of rheological properties
`
`Samuel J. Falcone, Richard A. Berg
`FzioMed Inc., 231 Bonetti Drive, San Luis Obispo, California 93401
`
`Received 30 November 2006; revised 20 April 2007; accepted 31 July 2007
`Published online 15 January 2008 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.a.31675
`
`Abstract: Temporary dermal fillers composed of cross-
`linked hyaluronic acid (XLHA) are space filling gels that
`are readily available in the United States and Europe. Sev-
`eral families of dermal fillers based on XLHA are now
`available and here we compare the physical and rheologi-
`cal properties of these fillers to the clinical effectiveness.
`The XLHA fillers are prepared with different crosslinkers,
`using HA isolated from different sources, have different
`particle sizes, and differ substantially in rheological prop-
`erties. For these fillers, the magnitude of the complex vis-
`cosity, |h*|, varies by a factor of 20, the magnitude of the
`complex rigidity modulus, |G*|, and the magnitude of the
`complex compliance, |J*| vary by a factor of 10, the per-
`cent elasticity varies from 58% to 89.9%, and the tan d
`varies from 0.11 to 0.70. The available clinical data cannot
`be correlated with either the oscillatory dynamic or steady
`
`flow rotational rheological properties of the various fillers.
`However, the clinical data appear to correlate strongly
`with the total concentration of XLHA in the products and
`to a lesser extent with percent elasticity. Hence, our data
`suggest the following correlation: dermal filler persistence
`5 [polymer] 3 [% elasticity] and the clinical persistence of
`a dermal filler composed of XLHA is dominated by the
`mass and elasticity of the material implanted. This work
`predicts that the development of future XLHA dermal fil-
`ler formulations should focus on increasing the polymer
`concentration and elasticity to improve the clinical persist-
`ence. Ó 2008 Wiley Periodicals, Inc. J Biomed Mater Res
`87A: 264–271, 2008
`
`Key words: dermal fillers; hyaluronic acid; soft tissue aug-
`mentation; oscillatory dynamic properties
`
`INTRODUCTION
`
`Dermal fillers for cosmesis of the face were intro-
`duced many years ago and received a large follow-
`ing with the introduction of injectable bovine colla-
`gen.1 As the procedures for wrinkle correction with
`dermal fillers became more popular,
`the search
`began for safer, longer lasting fillers. The next gener-
`ation filler after bovine collagen was crosslinked hy-
`aluronic acid (XLHA), which was an improvement
`over bovine collagen in that it did not require a skin
`test for hypersensitivity and appeared to be longer
`lasting.2 The search for safer and more effective der-
`mal fillers has continued because patients desire
`temporary fillers that are safe and predictably last
`longer than 6 months.3,4 In spite of a plethora of
`new temporary fillers composed of XLHA that are
`available in Europe and are undergoing clinical test-
`ing in the United States, it is not understood what
`parameters control the performance of these fillers.
`Although several studies have demonstrated that
`
`Correspondence to: R. A. Berg; e-mail: raberg@fziomed.com
`
`Ó 2008 Wiley Periodicals, Inc.
`
`XLHA fillers are more persistent than bovine colla-
`gen,5 the differences among the various XLHA fillers
`have not demonstrated obvious improvements in
`performance. One recent study compared two XLHA
`fillers, Perlane and Hylaform,6 and additional stud-
`ies are expected as more fillers become available.7
`Comparison of dermal fillers’ effectiveness is com-
`pounded by the injection techniques,8 and the bio-
`logical response of tissue to an implant in terms of
`degree of inflammation.3
`Here we compare the rheological properties of
`several commercial XLHA dermal fillers to under-
`stand the differences between them in terms of their
`physical properties and to attempt to correlate physi-
`cal properties with performance. One feature of hy-
`aluronic acid (HA) that has been useful
`in some
`medical device applications is its ability to form a
`cohesive gel.9 Cohesiveness is a function of con-
`centration and molecular weight. The property of
`cohesiveness, although an advantage for certain
`applications,
`is generally not an advantage as a
`dermal filler9 because highly cohesive (HA) materi-
`als are generally dilute solutions of noncrosslinked
`HA with low elasticity and short persistence in use.
`Dermal fillers are injected into the connective tissue
`
`Exhibit 1039
`Prollenium v. Allergan
`
`

`

`CROSSLINKED HYALURONIC ACID DERMAL FILLERS
`
`265
`
`of the dermis and therefore must be elastic in a low-
`shear environment. It is hypothesized that elasticity
`of a dermal filler leads to increased persistence but
`comparisons have not been performed. For noncros-
`slinked HA, dynamic rheological analysis demon-
`strated that as the frequency decreases the elastic
`properties decrease and hence, at low frequencies,
`the material becomes less elastic, and more viscous.
`Therefore, HA used in dermal fillers is always cross-
`linked to form gel particles that have high elasticity
`at lower frequencies.10 Dermal fillers prepared from
`XLHA are predominately elastic at low-shear envi-
`ronments and must have low viscosity under high
`shear to be able to be delivered through a small-bore
`needle. These unusual requirements have prompted
`us to compare commercially available dermal fillers
`prepared from XLHA in terms of their rheological
`properties.
`Clinical data comparing different XLHA dermal
`fillers are only starting to become available in the lit-
`erature. This study was undertaken to compare the
`physical properties of currently marketed XLHA fill-
`ers to determine which physical properties of XLHA
`dermal fillers are responsible for effectiveness when
`injected intradermally for soft tissue augmentation.
`Since the clinical data directly comparing commer-
`cial fillers to each other in the same study are not
`available and most studies have compared a given
`filler to bovine collagen in nasolabial folds in a split-
`face design, we have confined our clinical data set to
`using the wrinkle severity rating scale (WSRS) scor-
`ing system for fillers used in clinical studies reported
`to the FDA.11
`
`MATERIALS AND METHODS
`
`Materials
`
`The dermal fillers were obtained from commercial sour-
`ces. Restylane, Restylane SubQ, Restylane Perlane, Resty-
`lane Touch, and Restylane LIPP were obtained from Q-
`Med AB, (Uppsala, Sweden), or Medicis, (Scottsdale, AZ).
`Hylaform, Hylaform Plus, Juvederm 24, Juvederm 24HV,
`Juvederm 30, and Juvederm 30HV were obtained from
`Allergan (Inamed)
`(Irvine, CA) or LEA Derm (Paris,
`France). Puragen was obtained from Mentor (Edinburgh,
`UK), and Esthelis Basic was obtained from Anteis S.A
`(Geneva, Switzerland).
`
`Rheological measurements
`
`Small deformation oscillation dynamic rheological meas-
`urements were carried out with a Thermo Haake RS300
`Rheometer, Newington, NH, fitted in the cone and plate
`geometry. All measurements were performed with a
`35-mm/18 titanium cone sensor at 258C. Oscillation mea-
`surements were made over a frequency range of 0.628–198
`
`(rad/s). Percent elasticity is calculated as: Percent elasticity
`5 (100 3 G’)/(G’ þ G@).10,12
`
`RESULTS
`
`The HA dermal filler formulations evaluated in
`this study are all crosslinked. The exact nature of the
`crosslinking reaction conditions, crosslinker
`type,
`crosslink density, resultant particle size, and particle
`shape, all affect the physical properties of the XLHA
`formulation. While noncrosslinked HA forms a vis-
`cous solution when dissolved in aqueous solvents,
`chemically XLHA produces a material that swells in
`aqueous solution but does not dissolve. Hence, the
`XLHA dermal fillers are not solutions of XLHA but
`suspensions of swollen XLHA particles in aqueous
`solution. The properties of the XLHA products are
`influenced by the XLHA particle size and amount of
`polymer per unit volume. These materials do not
`have a smooth appearance and depending on the
`injection technique used can be lumpy. Also, suspen-
`sions of crosslinked polymers require larger gauge
`needles for injection into the dermis compared with
`the solutions of noncrosslinked polymers.
`Table I lists some properties of several commer-
`cially available dermal fillers containing XLHA. The
`source of HA is from bacterial fermentation except
`for the Hylaform products where the source of HA
`is animal
`(Avian). The crosslinkers used include
`vinyl sulfone, for the Hylaform products, 1,4-butane-
`diol diglycidyl ether (BDDE), for the Restylane and
`Juvederm series as well as Esthelis Basic. Puragen is
`crosslinked with 1,2,7,8-diepoxyoctane (DEO). Much
`has been written concerning the nature of the cross-
`linking reactions of XLHA dermal fillers. The differ-
`ences in the products from different manufacturers
`are generally ascribed to the physical state of the
`swollen gel after crosslinking HA. Products are
`described as being single or double crosslinked; par-
`ticulate or nonparticulate; monophasic, or biphasic.
`The Restylane, Juvederm, and Esthelis Basic prod-
`ucts all use BDDE as the crosslinking agent for HA.
`In the Restylane series, the crosslinking reaction pro-
`duces particles of crosslinked HA that are swollen in
`the aqueous phase. The number of particles/mL
`and the size of the particles differentiate the prod-
`ucts. As the size of the particles increases the num-
`ber of particles/mL decreases, and the products are
`advertised for use in the correction of deeper facial
`defects. The number of particles/mL for some of the
`Restylane family of products is listed in Table I.
`The crosslinking reaction for the Juvederm family
`of products is a patented process that produces a
`single phase, nonparticulate, crosslinked HA gel
`according to the manufacturer. This crosslinking
`technology is reported by the manufacturer to give
`
`Journal of Biomedical Materials Research Part A
`
`

`

`266
`
`FALCONE AND BERG
`
`Juvederm a softer feel when injected into the dermis
`and good persistence without the stiffness of other
`XLHA dermal fillers. Esthelis Basic products use a
`proprietary technology, cohesive polydensified ma-
`trix (CPM) to produce a single-phase nonparticulate
`XLHA dermal filler according to the manufacturer.
`Puragen uses DEO in a double crosslinking reaction
`that results in both ether and ester bonds crosslink-
`ing HA chains. According to the manufacturer, this
`highly crosslinked material is expected to improve
`the persistence of the Puragen products. The concen-
`trations of XLHA in these formulations varies from
` 5 mg/mL to 24 mg/mL. Increasing the concentra-
`tion of XLHA above  25 mg/mL becomes problem-
`atic because the products become too difficult to be
`injected through a small-bore needle.
`Table II lists some rheological properties, at 0.628
`(rad/s), of several commercially available dermal
`fillers containing XLHA. The magnitude of the com-
`plex viscosity (|h*|), at 0.628 (rad/s) of the dermal
`filler formulations, Figure 1, varies widely from 58
`to 1199 Pa s, almost 20 fold. Restylane LIPP has the
`highest magnitude of complex viscosity, 1199 Pa s
`and Esthelis Basic, using the CPM technology, the
`lowest. The XLHA products have a wide range of
`complex viscosities at low frequency.
`The magnitude of the complex viscosity, |h*|, for
`the Restylane family of products varies from 330 Pa
`s for Restylane SubQ to 1199 Pa s for Restylane
`LIPP. For the Restylane family of products, the rheo-
`logical properties could be affected by the number of
`XLHA particles contained per milliliter in the prod-
`uct. The rheological properties of Restylane SubQ,
`Restylane Perlane, Restylane, and Restylane Touch
`were measured and the
`results
`indicate
`that
`although the number of particles/mL changes and
`the products all have different particle sizes, and
`indications for use of these products are different,
`Table I, the magnitude of the complex viscosities for
`all of these products are similar. Restylane Touch
`has 500,000 particles and a magnitude of complex
`viscosity of 422.5 Pa s, Restylane has 100,000 par-
`ticles/mL and a magnitude of complex viscosity of
`532.4 Pa s, and Restylane Perlane has 10,000 parti-
`cle/mL with a magnitude of complex viscosity of
`486.4 Pa s. The |h*|, at 0.628 (rad/s), for these
`Restylane family of products does not relate to the
`number of XLHA particles contained per milliliter.
`Figure 2 demonstrates that for the Restylane family
`products, Perlane, Restylane and Restylane Touch,
`all have similar percent elasticity that is not a func-
`tion of the number of particles/mL. A change in
`number of particles/mL from 106 to 104 per mL is
`not associated with a significant change in the per-
`cent elasticity indicating that the percent elasticity
`for the product is independent of number of par-
`ticles/mL and hence particle size.
`
`BDDE:1,4butanedioldiglycidylether,CPM:cohesivepolydensifiedmatrixtechnologyresultsinmonophasicdoublecrosslinkedHA,DOE:1,2,7,8-diepoxyoctane—
`aQ-medproductliterature.
`
`doublecrosslinked—bothetherandesterlinksformedduringcrosslinkingreaction,ND:notdetermined.
`
`27
`27
`27
`30
`27
`27
`27
`
`Lg.bore
`
`27
`30
`30
`27
`30
`
`Size(ga)
`Needle
`
`Middermisandlips
`Midordeepdermisandlips
`Mediumtodeepwrinklesandlips
`Middermisandlips
`Middermisandlips
`Mediumtodeepwrinkles
`Lips
`SubQ,addvolume
`Deepdermisandlips
`Moderatetoseverewrinkles
`Superficiallines
`Moderatetoseverewrinkles
`Moderatetoseverewrinkles
`
`ND
`ND
`ND
`ND
`ND
`ND
`1000
`10,000
`100,000
`500,000
`
`crosslinked
`DEO,double
`BDDE,CPM
`BDDE
`BDDE
`BDDE
`BDDE
`BDDE
`BDDE
`BDDE
`BDDE
`BDDE
`Vinylsulfone
`Vinylsulfone
`
`(EU)CEMark
`
`Indication
`
`Particles/mL
`
`Crosslinker
`
`No.of
`
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Bacteria
`Avian
`Avian
`
`Source
`
`HA
`
`25
`24
`24
`24
`24
`20
`20
`20
`20
`20
`
`4.5–6.0
`4.5–6.0
`
`(mg/mL)
`
`Concentration
`
`Mentor
`Anteis
`LEADerm
`LEADerm
`LEADerm
`LEADerm
`Q-MedAB
`Q-MedAB
`Q-MedAB
`Q-MedAB
`Q-MedAB
`Inamed
`Inamed
`
`Puragen
`EsthelisBasic
`Juvederm30HV
`Juvederm30
`Juvederm24HV
`Juvederm24
`RestylaneLIPPa
`RestylaneSubQa
`RestylanePerlanea
`Restylanea
`RestylaneToucha
`HylaformPlus
`Hylaform
`
`Manufacturer
`
`Product
`
`ThePhysicalPropertiesofXLHADermalFillers
`
`TABLEI
`
`Journal of Biomedical Materials Research Part A
`
`

`

`CROSSLINKED HYALURONIC ACID DERMAL FILLERS
`
`267
`
`TABLE II
`The Rheological Properties, at 0.628 (rad/s) of HA-Based Dermal Fillers
`
`Product
`
`|h*| (Pa s)
`
`|G*| (Pa)
`
`|J*| (1/Pa)
`
`tan (d)
`
`% Elasticity
`
`Puragen
`Hylaform
`Hylaform Plus
`Restylane LIPP
`Restylane
`Restylane Perlane
`Restylane Touch
`Restylane SubQ
`Juvederm 30HV
`Juvederm 24HV
`Juvederm 30
`Juvederm 24
`Esthelis Basic
`
`941.8
`136.4
`108.2
`1199.0
`532.4
`486.4
`422.5
`330.4
`81.89
`151.5
`93.9
`58.4
`61.6
`
`591.7
`85.7
`68.0
`753.5
`334.5
`305.6
`265.5
`207.6
`51.46
`95.2
`59.0
`36.7
`38.7
`
`For the Juvederm family of products, the |h*|, at
`0.628 (rad/s), varies from 152 to 58 Pa s, Figure 1.
`The magnitude of the low frequency complex viscos-
`ity increases for Juvederm 24, to Juvederm 30HV,
`next is Juvederm 30, with Juvederm 24HV having
`the highest magnitude of |h*|, and the products are
`intended for different indications. However, Juve-
`derm 24HV and Juvederm 30HV are stated to have
`higher viscosities and perceived to be the most
`persistent of the Juvederm products even though
`the magnitude of the complex viscosity, |h*|, of
`Juvederm 30HV is no higher than Juvederm 30.
`Although the magnitude of the complex viscosity of
`Juvederm 24 HV is higher than Juvederm 24, the
`magnitude of the complex viscosity,|h*| at 0.628
`(rad/s), are all below 200 Pa s for this family of
`products. Again, the magnitude of the low frequency
`complex viscosity does not seem to correlate to per-
`sistence or indicated use for the Juvederm family of
`products.
`
`0.0017
`0.0117
`0.0147
`0.0013
`0.0030
`0.0033
`0.0038
`0.0048
`0.01943
`0.0105
`0.0170
`0.0272
`0.0258
`
`0.24
`0.14
`0.11
`0.18
`0.28
`0.30
`0.32
`0.39
`0.27
`0.31
`0.35
`0.53
`0.70
`
`80.4
`88.0
`89.9
`84.9
`78.2
`77.2
`75.6
`71.8
`78.7
`76.2
`74.1
`65.2
`58.8
`
`The magnitude of the complex rigidity modulus,
`|G*|, at low frequency, 0.628 (rad/s), for all prod-
`ucts is listed in Table II. The magnitude of |G*| at
`low frequency relates to the overall stiffness of the
`dermal filler at low deformation rate. The magnitude
`of the complex rigidity modulus, |G*|, versus fre-
`quency, for the dermal fillers is shown in Figure 3,
`and the higher the magnitude of the complex modu-
`lus, the stiffer the material. There is a large range,
` 10-fold, in the magnitude of the complex modulus
`versus frequency response of the XLHA dermal fill-
`ers studied here. Puragen has the highest magnitude
`of stiffness and Juvederm 24 or Esthelis Basic has
`the lowest magnitude of stiffness. The Restylane
`family of products has higher magnitudes of com-
`plex modulus than the Juvederm family of products.
`For the Restylane and Juvederm product families,
`
`Figure 1. A plot of the magnitude of the complex viscos-
`ity, |h*| (Pa s), at 0.628 (rad/s) for the XLHA dermal fill-
`ers listed in Table II. The magnitude of |h*| varies widely
`from 1199 to 58 Pa s for these products. Puragen and the
`Restylane series have the highest magnitudes of complex
`viscosities while Hylaform, Juvederm, and Esthelis Basic
`have much lower magnitudes of complex viscosity.
`
`Figure 2. A plot of the percent elasticity at 0.628 (rad/s),
`(100 3 G’/(G’ þ G@), versus the number of particles con-
`tained per milliliter for Restylane Touch, Restylane, Resty-
`lane Perlane, and Restylane SubQ. The data indicate that
`there is no correlation between the percent elasticity and
`the number of particles/mL and hence, the particle size in
`this Restylane product series. For this Restylane family of
`products, the number of particles/mL do not correlate to
`any of the low frequency (0.628 rad/s) rheological parame-
`ters listed in Table II.
`
`Journal of Biomedical Materials Research Part A
`
`

`

`268
`
`FALCONE AND BERG
`
`Figure 3. A plot of the magnitude of complex modulus,
`|G*| (Pa), versus frequency for the dermal filler listed in
`Table II. In this figure, Hylaform Plus and Restylane LIPP
`have been omitted. In this figure, the legend order is in de-
`scending magnitude of |G*| at 0.628 (rad/s). Puragen has
`the highest magnitude of |G*|at 0.628 (rad/s) and Juve-
`derm 24 has the lowest magnitude of |G*|at 0.628 rad/s.
`For this set of products, the magnitude of |G*| varies
`over 10 fold from the stiffest material, Puragen, to the least
`stiff, Juvederm 24. The magnitude of the modulus or over-
`all stiffness for these products is probably due to the cross-
`link density. It is also of interest to note that although they
`have different magnitudes, the slope of the |G*| versus
`frequency curves is very similar for all of the XLHA prod-
`ucts described here. This is not surprising since the struc-
`ture of the crosslinked polymer swollen in the matrix is
`very similar for all XLHAs.
`
`the complex rigidity modulus
`the magnitude of
`is more similar within each family than between
`families.
`Several studies concerning XLHA products have
`referred to a rheological property called the percent
`elasticity.10,12,13 In these studies percent elasticity is
`calculated as (100 3 G’)/(G’ þ G@) and is reported
`as the proportion of elasticity in an XLHA formula-
`tion. The percent elasticity versus frequency, for the
`XLHA dermal filler materials, is shown in Figure 4.
`The XLHAs have percent elasticity that range from
`60 to 90% with the Restylane series more closely
`grouped than the others. There is a considerable
`range in the percent elasticity of the XLHA products
`with Juvederm 24 and Esthelis being the least elastic
`and Hylaform the most elastic.
`The magnitude of the complex compliance, |J*|,
`versus frequency, for these materials, is shown in
`Figure 5. The compliance is the inverse of the modu-
`lus, and is a measure of how easy it is to deform a
`material. Again, the data indicate that the XLHA
`dermal fillers have a wide range of magnitudes of
`complex compliance of over 10 fold. Puragen has the
`lowest magnitude of complex compliance of all the
`dermal fillers studied here.
`
`Journal of Biomedical Materials Research Part A
`
`Figure 4. A plot of the percent elasticity, (100 3 G’/(G’ þ
`G@), versus frequency of the dermal fillers listed in Table
`II. In this figure, Hylaform Plus and Restylane LIPP have
`been omitted and the legend order is in descending per-
`cent elasticity at 0.628 (rad/s). Hylaform has the highest
`percent elasticity at 0.628 (rad/s) and Esthelis Basic has
`the lowest percent elasticity at 0.628 (rad/s). The percent
`elasticity for these XLHA products varies widely from
` 60 to 90%. Since Hylaform has the highest percent elas-
`ticity but the lowest 6-month improvement WSRS scores,
`(see Fig. 6), percent elasticity of the XLHA dermal filler
`itself does not correlate to product persistence and concen-
`tration must be taken into effect.
`
`Figure 5. A plot of the magnitude of the complex compli-
`ance, |J*| (1/Pa), versus frequency of the dermal fillers
`listed in Table II. In this figure, Hylaform Plus and Resty-
`lane LIPP have been omitted. The magnitude of |J*| is a
`measure of the overall ease of deformation of a material
`and hence, a material with a lower magnitude of complex
`compliance is harder to deform than a material with a
`higher magnitude of complex compliance. In this figure,
`the legend order is in descending complex compliance at
`0.628 (rad/s). Puragen has the lowest compliance and is
`the most difficult to deform, at 0.628 (rad/s). Juvederm 24
`has the highest magnitude of |J*| and is easiest to deform,
`at 0.628 (rad/s). The magnitude of |J*| for these XLHA
`products varies widely ( 10 fold). Puragen is the least
`compliant dermal filler followed by the Restylane products
`all of which have a magnitude of |J*| below 0.01. The
`remaining products all have a magnitude of |J*| above
`0.01 at 0.628 (rad/s).
`
`

`

`CROSSLINKED HYALURONIC ACID DERMAL FILLERS
`
`269
`
`TABLE III
`Effectiveness Data for Dermal Fillers at
`6-Month Post-Treatment
`
`Product
`
`Zyplast
`Restylane
`Zyplast
`Juvederm 30
`Zyplast
`Juvederm 24HV
`Zyplast
`Juvederm 30HV
`
`6-Month
`Improvement
`Score WSRS
`
`0.36
`0.93
`0.5
`1.4
`0.3
`1.3
`0.4
`1.4
`
`Reference
`
`Restylane control
`Ref. 14
`Juvederm 30 control
`Ref. 7
`Juvederm 24HV control
`Ref. 7
`Juvederm 30HV control
`Ref. 7
`
`In these studies all of the XLHA products were com-
`pared to Zyplast as a control.
`
`for any relationship
`In attempting to account
`between rheological properties and clinical benefit or
`persistence, the available data are quite limited. This
`is because few studies compare one filler to another.
`Some fillers like Esthelis and Puragen have no
`published data on performance. The available data
`on fillers where one product was compared with
`another is summarized in Tables III–V. The data set
`is only for the clinical studies involving filling the
`nasolabial folds in studies submitted to the FDA in
`support of approval of a Premarket Application.
`Only the 6-month data point was evaluated in each
`of the studies. If one compares the persistence of the
`fillers for which there are data with the measured
`values given in Table II, the persistence, as measured
`by the 6-month WSRS improvement score, correlates
`
`TABLE IV
`Effectiveness Data for Dermal Fillers
`at 3-Months Post-Treatment
`
`Product
`
`Zyplasta
`Hylaforma
`Zyplast
`Juvederm 30
`Zyplast
`Juvederm 24HV
`Zyplast
`Juvederm 30HV
`Hylaforma
`Hylaform Plusa
`
`3-Month
`Improvement
`Score WSRS
`
`1.2
`1.1
`1.0
`1.6
`1.0
`1.7
`0.9
`1.6
`0.8
`0.9
`
`Reference
`
`Hylaform control
`Ref. 15
`Juvederm 30 control
`Ref. 7
`Juvederm 24HV control
`Ref. 7
`Juvederm 30HV control
`Ref. 7
`Control for Hylaform Plus
`Ref. 15
`
`aTo compare Hylaform and Hylaform Plus to the
`products in Table III, studies comparing Hylaform and
`Hylaform Plus to Zyplast are compared. In these studies,
`a 6-point scale with a live evaluator was used to score the
`3-month time point in the 3-month study demonstrating
`that Zyplast, Hylaform, Hylaform Plus are all similar.
`Since Zyplast was used as a control in the Restylane and
`Juvederm studies in Table III, Juvederm, Restylane, and
`Hylaform can all be used to estimate effectiveness.
`
`TABLE V
`Correlations from Tables III and IV
`
`WSRS
`Improvement
`Score
`(6 months)
`
`Concentration
`(% Solids)
`
`%
`Elasticity
`
`Complex
`Viscosity
`
`Complex
`Modulus
`
`0.4
`0.93
`1.4
`
`0.5
`2.0
`2.4
`
`88–89
`71–78a
`65–76
`
`136
`330–530a
`58–151
`
`68–85
`207–334a
`36–95
`
`aExcluding Restylane LIPP.
`
`inversely to the percent elasticity as shown in Figure
`6. Hence, as the percent elasticity increases, the per-
`sistence decreases. From the previous discussion,
`this result is counterintuitive and indicates that high
`elasticity alone cannot account for the persistence of
`XLHA dermal fillers; however, the concentration of
`polymer was not taken into effect. The effectiveness
`of the dermal fillers listed in Table V does correlate
`to the concentration of polymer in the formulation
`as shown in Figure 7. Here, the persistence of der-
`mal filler is directly related to the mass of polymer
`in the formulation. Intuitively, it would be reasona-
`ble that both polymer concentration and elasticity
`would relate to the effectiveness of dermal filler.
`This relationship is shown in Figure 8, where effec-
`tiveness for a series of dermal fillers is plotted versus
`
`Figure 6. Described in this plot is the relationship of the
`6-month improvement score WSRS to the XLHA polymer
`percent elasticity at 0.628 (rad/s), for Hylaform, Restylane,
`Juvederm 24HV, Juvederm 30, and Juvederm 30HV. The
`improvement scores are from Tables III and IV. The WSRS
`score is a measure of the relative effectiveness of these
`XLHA dermal fillers after 6 months. For these XLHA der-
`mal fillers, the WSRS improvement score increases as the
`percent elasticity decreases. The WSRS improvement score
`correlates in a linear manner to the percent elasticity at
`0.628 (rad/s). Since the prevailing theory suggests that
`higher percent elasticity results in dermal fillers with
`higher persistence, this result is somewhat counterintui-
`tive. These data suggest that high elasticity at low fre-
`quency is not
`the only factor affecting persistence for
`XLHA dermal filler.
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`

`270
`
`FALCONE AND BERG
`
`HA, is that one can assume that the biological or tis-
`sue response is similar for each filler in the series.
`Many different filler materials have been tested and
`the differences in tissue responses and injection tech-
`niques make it very difficult to correlate the clinical
`persistence of these different materials to physical
`properties (for reviews, see Refs. 1, 17–20). Attempts
`to compare the available XLHA dermal fillers have
`been limited. Two products Hylaform and Restylane
`have been compared in terms of some chemical
`properties16 but since these products differ in con-
`centration and particle size, the data set is too lim-
`ited to draw conclusions. Rheologically, all of the
`XLHA products generally have high elastic to loss
`modulus characteristics exemplified by low tan (d) val-
`ues throughout the entire frequency range. Attempts
`have been made in the past to estimate the elastic
`nature of XLHAs by calculating the percent elasticity
`from the viscoelastic modulii. This approach is lim-
`ited by the effect of concentration of the polymer.
`Hylaform is highly elastic but is dilute compared
`with other products and also is less persistent than
`Restylane and Juvederm (Table V). Another possibil-
`ity is that the magnitude of the complex viscosity or
`the magnitude of the complex rigidity modulus is
`the controlling factor. Juvederm has magnitudes of
`complex viscosity and complex modulus that are
`considerably lower than Restylane (Table II), and it
`appears to be more persistent (Table V).
`Other suggestions have been that the persistence
`is proportional to particle size. The only product
`family where particle size is estimated is the Resty-
`lane family of products, and for this family there
`
`Figure 8. This figure describes the correlation of the 6-
`month WSRS improvement score to the product of the
`concentration (mg/mL) and percent elasticity for XLHA
`dermal fillers Hylaform, Restylane, Juvederm 24HV, Juve-
`derm 30, and Juvederm 30HV. There is a linear correlation
`that indicates that both properties (concentration and elas-
`ticity) are factors influencing the persistence of XLHA der-
`mal fillers.
`
`the 6-month improvement score
`Figure 7. A plot of
`WSRS versus XLHA polymer concentration (mg/mL) for
`Hylaform, Restylane, Juvederm 24HV, Juvederm 30, and
`Juvederm 30HV. The improvement scores are from Tables
`III and IV. The WSRS score is a measure of the relative
`effectiveness of these XLHA dermal fillers after 6 months.
`For the data shown here, the Hylaform 6-month WSRS
`score was assumed to be equivalent to the 6-month WSRS
`Zyplast score (Table III), because when Hylaform and
`Zyplast were compared in a 3-month study, they had very
`similar WSRS scores. Zyplast was also used as the control
`in the 6-month study comparing Restylane to Juvederm.
`For these data, the 6-month WSRS improvement score cor-
`relates very well to the concentration of XLHA contained
`in the product.
`
`the product of the polymer concentration in solution
`and the percent elasticity, at 0.628 rad/s, of the for-
`mulation. All of the XLHA fillers for which there are
`clinical data correlate linearly with the polymer con-
`centration and elasticity.
`
`DISCUSSION
`
`The clinical persistence and characteristics of HA-
`based dermal fillers may be influenced by their
`physical properties.16 When HA is formulated as a
`biomaterial for a specific indication, it is frequently
`crosslinked.10 HA is subject to degradation by hyalu-
`ronidase in the body and crosslinking is expected to
`reduce susceptibility to enzymatic degradation. The
`dynamic rheological properties of HA need to be
`improved for the polymer to perform its intended
`use as dermal filler. Crosslinking the polymer
`increases the elastic modulus of the polymer, at low
`frequency, making the XLHA more elastic than non-
`crosslinked HA. Hence, for dermal filler formula-
`tions, HA is chemically crosslinked to increase the
`elastoviscous properties and increase the residence
`time at the site of injection.
`The advantage of comparing a series of dermal
`fillers manufactured from the same material, that is
`
`Journal of Biomedical Materials Research Part A
`
`

`

`CROSSLINKED HYALURONIC ACID DERMAL FILLERS
`
`271
`
`appears to be no correlation between particle size
`and percent elasticity (Fig. 1), the magnitude of the
`complex viscosity (Fig. 4), the magnitude of the com-
`plex modulus (Fig. 5), or percent elasticity (Fig. 6).
`The major difference between members of the Resty-
`lane family of products with different particle sizes
`is the injectability of the products. Restylane and
`Restylane Touch are administered through 30 gauge
`needles whereas Perlane requires a 27-gauge needle
`and SubQ requires a larger cannula. Unfortunately,
`comparisons of persistence among the Restylane
`products are not available.
`In conclusion, the oscillatory dynamic rheological
`properties of XLHA dermal fillers have been deter-
`mined and it was found that they are quite variable
`with respect to the magnitude of the complex rigid-
`ity modulus, overall stiffness (|G*|), the magnitude
`of the complex viscosity (|h*|), and percent elastic-
`ity. Although there are limited controlled clinical
`data, the most straightforward correlation is between
`persistence of the product and polymer concentra-
`tion, when the products are compared in a typical
`clinical study of paired bilaterally placed fillers in
`the nasolabial folds. Within the same concentrations,
`persistence appears to be related to elasticity. A lin-
`ear correlation exists between persistence and poly-
`mer concentration and also between persistence and
`the polymer concentration and elasticity. The data
`presented here indicate that the persistence of XLHA
`dermal fillers appears to be a function of the concen-
`tration of polymer in solution and to a lesser extent
`the elasticity of the material. The maximum concen-
`tration of XLHA polymers in the current XLHA der-
`mal filler formulations appear to be limited to  25
`mg/mL. Finding new methods
`to increase the
`XLHA concentration in dermal filler formulations
`may be the important hurdle to overcome for new
`filler candidates prepared from XLHA.
`
`References
`
`1. Klein AW, Elson ML. The history of substances for soft tissue
`augmentation. Dermatol Surg 2000;26:1096–1105.
`2. Fernandez EM, Mackley CL. Soft
`tissue augmentation: A
`review. J Drugs Dermatol 2006;5:630–641.
`
`3. Lemperle G, Morhenn V, Charrier U. Human histology and
`persistence of various injectable filler substances for soft
`tissue augmentation. Aesthetic Plast Surg 2003;27:354–366;
`discussion 367.
`4. Bosniak S, Cantisano-Zilkha. Restylane and Perlane: A six
`year clinical experience. Operative Tech Oculoplastic Orbital
`Reconstr Surg 2001;4:89–93.
`5. Narins RS, Brandt F, Leyden J, Lorenc ZP, Rubin M, Smith S.
`A randomized, double-blind, multicenter comparison of the
`efficacy and tolerability of Restylane versus Zyplast for the
`correction of nasolabial folds. Dermatol