Absorptive supramolecular elastomer wound dressing based on polydimethylsiloxane–(polyethylene glycol)–polydimethylsiloxane copolymer: preparation and characterization

Abstract

Polydimethylsiloxanes (PDMS) are soft materials with high elasticity and excellent biocompatibility, giving them high potential for widespread applications as biomaterials. However, the application of PDMS to wound dressing is limited due to its shortcomings, particularly its non-absorptive and poor adhesive properties. This work addresses these limitations in two ways: first, a hydrophilic and biocompatible polyethylene glycol (PEG) block was introduced to the PDMS main chain to obtain carboxyl acid terminated PDMS-b-PEG-b-PDMS copolymers (EPMDS–COOH₂); second, EPMDS–COOH₂ polymers were reacted with diethylenetriamine and urea via a two-stage route to obtain a novel supramolecular elastomer (ESESi) based on multiple hydrogen bond association. Compared with the supramolecular elastomer based on simple PDMS (SESi), the hydrophilicity, water-absorption rate, adhesive ability and rate of water vapor permeation were significantly improved, enhancing the properties of ESESi films for application to wound dressing. After fully evaluating the bio-compatibilities of ESESi films, a full-thickness dermal wound model was chosen to estimate the healing performance, and traditional vaseline gauze, SESi film and commercialized Tegaderm™ film were chosen as controls. The results confirmed that the ESESi polymer could promote the healing of a skin wound to some extent, mainly due to its improved absorption and adhesive properties.

---

1. Introduction

Wound healing is a dynamic and complex process that includes inflammatory, proliferation, and remodeling phases.¹ Therefore, wound dressings play a critical role during healing, with the aim of providing an ideal healing environment. Studies have shown that in addition to preventing infection, presenting biocompatibility and allowing gaseous exchange, an ideal wound dressing would also manage the wound exudate to provide a suitable environment for wound healing.^2–4 Wound exudate is often perceived as undesirable because exudate can require frequent dressing changes and can lead to maceration of the wound bed, delayed healing, imbalances between fluids and electrolytes, odor and infection.^5–9 Therefore, it is important that wound dressings have a good absorptive capacity. The importance of absorbent wound dressings did not receive significant attention for more than a century until George Winter put forward the theory of wet healing and demonstrated that the wound healing rate was accelerated almost two-fold if wounds were kept moist.¹⁰ Since then, an increasing number of studies about absorbent wound dressings have been found in wound dressing research.^11,12 These papers have investigated modern first-line absorbent wound dressings, including alginate, hydrofiber, foam, hydrocolloid and polysaccharide bead wound dressings.^13–17 These novel absorbent wound dressings may exhibit some favorable effects in managing exudates and wound healing. However, these absorbent wound dressings still do not meet all clinical requirements, with shortcomings such as poor strength, lack of adhesiveness, opacity, discomfort, inconvenience and even a propensity to cause inflammation.^18–20 Therefore, better wound dressings remain a challenge. Polydimethylsiloxanes (PDMS) are commonly used polymers with many excellent properties, including softness, transparency, outstanding physical properties, chemical durability, and high oxygen permeability.^21,22 More importantly, PDMS exhibit remarkable biocompatibility and have been approved by the US Food and Drug Administration (FDA). As a result, PDMS have been widely applied in medical products. Although PDMS may be an ideal wound dressing material,^23–26 polydimethylsiloxane materials have thus far mostly been used as an accessory for wound dressings, for example, as a scaffold.²⁷ This is due to some obvious disadvantages of PDMS, for instance, their lack of adhesiveness and poor absorption due to their hydrophobicity. To address these limitations and develop an “absorbent polydimethylsiloxane wound dressing”, in previous studies,^28–30 we developed a novel way to prepare wound dressing materials based on a supramolecular elastomer that generated multiple hydrogen bonds among PDMS chains (SESi). To augment the inherent properties of PDMS, SESi was endowed with cementitiousness and some absorptivity. However, wound healing experiments demonstrated that the SESi was not sufficiently absorptive, resulting in unsatisfactory promotion of wound healing. Therefore, the SESi films do not meet the requirements of “absorptive wound dressings”.

Thus, the primary goal of this work is to design a biocompatible and absorbent polydimethylsiloxane material for use as a wound dressing. In this work, a polyethylene glycol (PEG) block, which is both hydrophilic and biocompatible, was introduced into the PDMS main-chain to obtain a carboxyl-terminated PDMS–PEG–PDMS tri-block copolymer (EPDMS–COOH₂). After a “two step” reaction with diethylenetriamine (DETA) and urea, another novel supramolecular elastomer (ESESi) was obtained, consisting of a PDMS–PEG–PDMS tri-block copolymer cross-linked by multiple hydrogen bonds. This study describes the synthesis and characterization of ESESi, followed by the evaluation of its biocompatibility, water vapor and bacterial permeability, and its effect on wound healing in comparison with the unmodified supramolecular elastomer film based on polydimethylsiloxane (SESi) and with the commercially available Tegaderm™ film. The results showed that the inclusion of PEG blocks endowed the ESESi film with excellent water swelling ability. Furthermore, the water vapor permeability, adhesiveness and mechanical properties of the ESESi film were also improved by the introduction of PEG blocks. Surprisingly, the supramolecular material synthesized here is also cohesive with itself. Combining these new properties with the existing characteristics of polydimethylsiloxane materials, ESESi film wound dressings showed better performance than all control wound dressings, including traditional vaseline gauze, SESi films, and commercialized Tegaderm™ films, demonstrating that ESESi films may be effective absorptive wound dressings.

2. Materials and methods

2.1 Materials and animals

Octamethylcyclotetrasiloxane (D₄, >99.5%, Dow Corning, USA), 1,1,3,3-tetramethyldisiloxane (HMM, >98%, Kaihua Taicheng Silicone Co., Ltd., Quzhou, China), tert-butyl methacrylate (tBMA, 98%, Chemlin Chemical Industry Co., Ltd., Nanjing, China), platinum(0)-1,3-divinyl-1,1,3,3-tetramethyl-disiloxane complex solution (Karstedt's catalyst solution, Pt ∼ 2%, Aladdin Reagent Co. Ltd., China), and polyethylene glycol (PEG, M_n = 600, Sigma-Aldrich, 99%) were used as received. Tegaderm™ films (transparent film dressing, type 1624W) were supplied by 3M Health Care (St. Paul, MN, USA). SESi film wound dressings were prepared as in our previous work.³⁰ The culture medium RPMI 1640 was obtained from Invitrogen (New York, USA). Fetal bovine serum was provided by Hangzhou Sijiqing Biological Engineering Materials Co., Ltd. (Hangzhou, China). MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide) was obtained from Alfa-Aesar. Tetrahydrofuran (THF) was purchased from Aladdin and dried with sodium. Diethylenetriamine (DETA, 98%), allyl chloride (AC, 99%), urea (99%), phosphoric acid (85%), sodium hydride (60%) and solvents, were used as received without any further purification.

Male Wistar rats (weight range, 200–250 g) were used to evaluate the wound healing characteristics of the materials under study. All experiments were completed with the approval of the Animal Research Ethics Committee of the South China Agricultural University. During the study, all rats had access to chow and water ad libitum.

2.2 Preparation of ESESi

2.2.1. Preparation of linear carboxyl-terminated PDMS–PEG–PDMS tri-block copolymer (EPDMS–COOH₂). EPDMS–COOH₂ was synthesized through the five-step process described in Scheme 1. Diallyl polyethylene glycol (APEG) was prepared by Williamson ether synthesis. Then, two successive hydrosilylation reactions were performed with PHMS and tert-butyl methacrylate (tBMA). Finally, EPDMS–COOH₂ was obtained through an acidic hydrolysis reaction of the t-butyl ester terminated polydimethylsiloxane (EPDMS–tBMA₂). Three types of EPDMS–COOH₂ were prepared with different PDMS/PEG ratios, i.e., EPDMS_1/9–COOH₂, EPDMS_1/11–COOH₂, and EPDMS_1/15–COOH₂, in which the [–CH₂–CH₂–O–]/[–Si(CH₃)–O–] molar ratios were 1/9, 1/11, and 1/15, respectively.

Scheme 1 Synthetic route of EPDMS_x–COOH₂.

2.2.2. Synthesis of ESESi. The ESESi elastomer was synthesized via a two-step process shown in Scheme 2, which is similar to the reaction in our previous work^28–30 and Leibler's report.^31,32

Scheme 2 The preparation and structure of the ESESi elastomer.

In the first step, EPDMS_x–COOH₂ (x = 1/9, 1/11, or 1/15) and DETA were added at room temperature in a thermostated reactor fitted with a Dean–Stark trap, a stirring system, and a nitrogen inlet, with a [–NH₂]/[–COOH] molar ratio of 2.2/1. Then, the mixture was heated to 120 °C at a heating rate of 40 °C h⁻¹ and held at 120 °C for 1 h. Then, the mixture was heated to 135 °C at a rate of 10 °C h⁻¹. After reacting for 3.5 h, the mixture was cooled to room temperature, solubilized in chloroform and washed with a water/methanol mixture (weight ratio 5/2) at least five times. The oligoamide was collected as a solution in chloroform.

FTIR (KBr, cm⁻¹): 3410 (ν_NH), 3024 (δ_NH), 2962 (ν_{as CH3}), 1652 (ν_{C=O amide}), 1606 (ν_{CN imidazoline}), 1549 (δ_NH), 1441 (δ_{as CH3}), 1263 (δ_{s Si–CH3}), 1020–1100 (ν_Si–O), 867 (r_Si–CH3), 792 (ν_{as Si–CH3}), 699(ν_{s Si–CH3}) (Fig. S1†).

¹H-NMR δ (CDCl₃, ppm): 2.57 (t, NHCH₂CH₂NH₂), 2.73 (t, C(O)NHCH₂CH₂NH), 2.75 (t, NHCH₂CH₂NH₂), 2.78 (t, NH₂CH₂CH₂N–imidazoline), 3.17 (t, NH₂CH₂CH₂N–imidazoline), 3.32 (m, C(O)NHCH₂CH₂NH), 3.34 (m, C(O)NHCH₂CH₂N–imidazoline), 3.65 (m, –CH₂–CH₂–O–), 3.81 (m, N(C)CH₂CH₂N–imidazoline) (Fig. S2†).

In the second step, urea was added to the organic phase collected above, and the mixture was stirred under a 200 mL min⁻¹ nitrogen flow at 70 °C overnight, with a [urea]/[–COOH] ratio of 2.2/1. The mixture was heated to 135 °C at a heating rate of 40 °C h⁻¹. After 1 h, the mixture was heated to 160 °C in 5 °C increments every 30 min. After further incubation, the mixture rose up along the stirring stem. Once the mixture had risen completely for 1–2 h, the reaction was stopped. After the product cooled to room temperature, it was cut into small fragments and washed in water at 50 °C for 2–3 days. Finally, the washed product was dried at 110 °C under vacuum for 24 h, and a faint yellow and semi-transparent elastomer (ESESi_x, x = 1/9, 1/11, or 1/15) was obtained.

FTIR (KBr, cm⁻¹): 3116 (ν_NH), 2965 (ν_{as CH3}), 1660 (ν_{C=O amide}), 1600 (ν_{CN imidazoline}), 1540 (δ_NH), 1452 (δ_{as CH3}), 1260 (δ_{s Si–CH3}), 1020–1100 (ν_Si–O), 869 (r_Si–CH3), 797 (ν_{as Si–CH3}), 695(ν_{s Si–CH3}) (Fig. S3†).

¹H-NMR δ (CDCl₃, ppm): 3.33–3.47 (m, C(O)NHCH₂CH₂N–imidazolidone), 3.65 (m, –CH₂–CH₂–O–), 3.82 (t, C(O)NHCH₂CH₂N–imidazolidone) (Fig. S4†).

2.2.3. Preparation of the ESESi film and SESi film. The ESESi elastomer was heated and pressed at 145 °C for 15 min in a mold and then cool pressed at room temperature for 15 min to obtain a 0.75 mm thick ESESi film. The SESi films were obtained using the same method.

2.3 Characterization of ESESi

The hydrogen bonding interactions in the ESESi matrix and their related thermal properties were determined with temperature-dependent infrared spectroscopy. Infrared spectra were obtained using a Bruker Vertex 70 Fourier transform-infrared spectrometer equipped with a Harrick ATC-024 temperature controller.

Differential scanning calorimetry (DSC) analysis was used to evaluate the thermal transitions and crystallization temperature of these ESESi materials. Thermal analyses were performed at a heating and cooling rate of 10 °C min⁻¹ in a nitrogen atmosphere with a Netzsch DSC 204C apparatus. The ESESi specimens were first heated and held at 100 °C for 5 min and then cooled and held at −150 °C for 5 min. The thermal transitions of these materials were evaluated from thermograms collected during the second heating process.

XRD analyses were used to observe the crystallization of ESESi materials at room temperature. This was performed with a PANalytical X'Pert-Pro X-ray diffractometer with filtered monochromatic Cu Kα radiation in the 2θ range of 5° to 90°.

An RPA 2000 rubber process analyzer was used for rheological temperature sweeps (30 to 160 °C) and frequency sweeps (0.05 to 200 rad s⁻¹). The dynamic temperature sweep was performed at strain amplitude of 3% and frequency of 1 Hz. The dynamic frequency sweep was performed at strain amplitude of 3% over the temperature range from 40 to 100 °C.

The mechanical properties of the ESESi film were determined by a tensile testing machine (Zwick GmbH & Co. KG, Ulm, Germany) at a stretching rate of 500 mm min⁻¹. Tensile tests were performed on rectangular specimens (50 mm × 10 mm × 1 mm).

The adhesiveness of the ESESi films was measured using an inclined type ball tack device (HT-6032 Tape Initial Adhesion Tester, Dongguan, China) at room temperature according to GB/T 4852-2002, with a 30° angle of inclination. Initial adhesion was defined as the maximum ball number (corresponding to the diameter of the ball) that remained adhered to the film, with larger ball numbers indicating higher tack properties of the films.

The water vapor transition rate (WVTR) was used to evaluate the permeability of the ESESi, SESi and Tegaderm™ films to moisture. WVTRs were measured according to ASTM E96. The film samples were used to cover the mouth of a conical flask (25 mm in diameter) filled with 40 mL of distilled water. The flask was then maintained in an incubator at 37 °C and 40 ± 2% RH. The WVTR was calculated using the following eqn (1):

where WVTR is expressed in g m⁻² d⁻¹; G is the weight change in g; A is the area of the cup mouth (m²); and t is time (d).

The static water contact angles (WCAs) of the SESi and ESESi film surfaces were measured with a contact angle meter (DropMeter A-100, MAIST Vision Inspection & Measurement Co. Ltd., China) at ambient temperature. One drop of water (10 μL) was placed onto the surface and observed with an optical microscope. The WCA values were calculated from three measurements at different positions.

The water absorption ratio (A_w) was determined by soaking the samples in normal saline at 37 °C and 40 ± 2% RH until reaching the swelling equilibrium state. The films were then removed and wiped gently with filter paper. The A_w values were calculated by eqn (2):

where W_e and W_d are the film weights in the equilibrium and dry state, respectively.

2.4 Evaluation of cytotoxicity

In this study, cytotoxicity was evaluated via the MTT cytotoxicity and direct contact tests, according to GB/T 16886.5-2003 (equivalent to ISO 10993-5: 1999).

2.4.1. MTT cytotoxicity. The test patches of the test samples (1 cm × 1 cm × 0.1 cm) were soaked in deionized water for 48 h and dried. Then, they were sterilized for 3 h with ultraviolet light and then immersed in 10 mL of RPMI 1640 culture medium for 24 h at a film/solution ratio of 1/10 (cm² mL⁻¹). After 24 h, the medium incubated with the film was extracted and diluted with fresh RPMI 1640 culture medium for testing at different concentrations (75%, 50% and 25%).

L929 cells (murine aneuploid fibrosarcoma cells) were cultured in RPMI 1640 culture medium supplemented with 10% FBS in 96 well plates (100 μL medium per well) at a density of 1.0 × 10⁵ cells per mL. Then, the cells were cultured overnight at 37 °C in a humidified 5% CO₂ incubator. After culturing the cells for 24 and 48 h in different concentrations (25%, 50%, 75% and 100%) of diluted extract of ESESi film, respectively, a solution of 5.0 mg mL⁻¹ MTT (in PBS) was added to these wells. After incubation for another 4 h at 37 °C, the growth medium was removed, and 150 μL of DMSO was added, followed by vigorous shaking to dissolve the purple formazan crystals that formed. Absorbance was measured with a MK-III Microplate Reader (Thermo, USA) at a wavelength of 492 nm.

2.4.2. Direct contact test. A suspension of L929 cells (2 mL) was injected into the vessels and incubated at 37 °C in a humidified 5% CO₂ incubator. Afterwards, fresh RPMI 1640 culture medium was added. Sterilized sample patches (1 cm × 1 cm × 0.1 cm) were gently placed on the layer of cells in the center of each of the replicate vessels. One tenth of the cell layer surface was covered by the test sample. The cells were incubated in the same environment for 24 h. An LDPE film (negative control) and an organotin-stabilized PVC film (positive control) were selected as comparisons.

2.5 Detection of bacterial permeation

The membranes were sterilized by irradiation with 75% ethanol and then ultraviolet light for 30 min. E. coli and S. cremoris were cultured aerobically at 37 °C and shaken at 150 rpm for 12–16 h. Sterilized samples of the ESESi film, the SESi film, the Tegaderm™ film and the vaseline gauze were cut into circular discs (diameter 10 mm) and placed on the center of an agar dish. Next, the bacterial suspensions were diluted to 1 × 10⁹ colony-forming units (CFU) mL⁻¹, 20 mL of these suspensions were added to the center of each sample, and the inoculum was uniformly plated on the surface of each sample. After incubation at 37 °C for 24 h, the agar under each sample was cut with a knife and placed into a tube containing 2 mL of PBS. The bacteria on the agar were detached for 4 min using an ultrasonic cleaner, and the numbers of bacteria were determined by routine CFU analysis on an agar dish with different dilutions.

2.6 The survey of wound contraction

The wound dressings developed in this paper were applied to full-thickness skin defects in rats.

The experiments were repeated multiple times to obtain a sample size of five rats per treatment per time point.

After surgery, the treated rats were fed in individual cages, and wound conditions were observed and recorded at different healing times. On days 4, 6, 8, 10, and 14 post-surgery, the treated rats were anesthetized and sacrificed by cervical dislocation. The new wound area was determined by measuring the diameters of the two wound sites. The wound sites were excised and processed for histological evaluation. The percent wound contraction was calculated with the following formula (3):

where A₀ and A_t are the original wound area and the area of the wound at the time of biopsy, respectively. The wound area was determined by examining images of the wounds. The wound tissue and adjacent normal skin were carefully removed for histological analysis. Then, the tissues were fixed with 10% phosphate-buffered formalin and stained with hematoxylin and eosin (H&E) reagent for histological observations. Digital photos were acquired by a light microscope (Nikon DS-Ri1, Japan) at 40× and 200× magnification.

Statistical analyses were performed with at least five samples of each ESESi film, SESi film, Tegaderm™ film and vaseline gauze. All data were presented as the mean ± standard deviation (SD), and the significance level was set at p < 0.05. Intergroup differences were determined using Microsoft Excel's statistical function to compare test groups with a control group using Student's t-test.

3. Result and discussion

3.1 Hydrogen bonding interactions of ESESi

The association of the hydrogen bonds were characterized by collecting FT-IR spectra of ESESi at various temperatures (30 °C, 50 °C, 70 °C, 90 °C, 110 °C, 130 °C and 150 °C), as shown in Fig. 1. According to the results, the C=O stretching signals of amide groups (1656 cm⁻¹) shift to higher wave numbers (1683 cm⁻¹), whereas the N–H bending signals (1537 cm⁻¹) shift toward lower wave numbers (1514 cm⁻¹) with increasing temperature. Meanwhile, the FT-IR spectra collected following cooling were almost identical to those collected during heating. Both these shifts are due to the multiple hydrogen bonds between N–H and C=O species.

Fig. 1 FT-IR spectra of ESESi at various temperatures: (A) heating and (B) subsequent cooling.

3.2 Physical properties of ESESi

3.2.1. Thermal transition and crystallization of ESESi. The behavior of the thermal transition and crystallization of ESESi were analyzed by DSC and XRD, respectively.

As shown in Fig. 2A, different ESESi materials exhibit similar trends in their thermal transitions. Generally, two clear low transition temperatures (T_g) of siloxane chains (T_g1) and PEG chains (T_g2) could be observed. Moreover, melting peaks (T_m) were found less than 10 °C. However, not all ESESi materials contained two melting and two low transition peaks, as shown in Fig. S5.† This may be due to the formation of hydrogen bonds between PEG chains and other chains, forming an analogous homogeneous chain. Moreover, no crystalline peak was observed in the X-ray diffraction pattern of ESESi at room temperature. The XRD and DSC results confirm that ESESi was not able to crystallize at room temperature (Fig. 2B). Due to its low T_g and T_m values, the ESESi material could exhibit good elasticity at room temperature or even at a lower temperature.

Fig. 2 Typical curves (A) DSC and (B) XRD curves of ESESi_1/9.

3.2.2. Viscoelastic properties of ESESi. The viscoelastic properties were evaluated by measuring the storage modulus (G′) and loss modulus (G′′) during dynamic temperature sweeps (Fig. 3). These results indicate a clear glass transition at low temperature, and a highly elastic rheological response (G′ > G′′) was observed below 150 °C. In addition, a wide temperature-independent rubbery plateau in G′ was also detected in the temperature range from 80 °C to 110 °C. Nevertheless, G′ decreases sharply when the temperature exceeds 110 °C. In accordance with the results of the FT-IR analysis, a significant decrease in the degree of hydrogen bond associations is observed at higher temperatures.

Fig. 3 The temperature dependence of the dynamic storage modulus (G′) and loss modulus (G′′) for ESESi material.

3.2.3. Tensile properties of ESESi film. Tensile testing was used to determine the mechanical properties of the ESESi film in comparison with the SESi film. As depicted in Fig. 4, the tensile strength of the ESESi film was greater than that of the SESi film. This result indicates that the hydrogen bond interactions between the PEG blocks and other chains promote the tensile strength of the PDMS material. In addition, higher PEG block ratios are associated with greater tensile strength.

Fig. 4 Stress–strain curves of ESESi and SESi at a 500 mm min⁻¹ stretch rate.

3.2.4. Initial adhesion of ESESi. Adhesion is an indispensable characteristic of an ideal wound dressing. Excellent adhesion makes a wound dressing convenient to use and aids in the creation of a suitable healing environment. In this paper, the adhesion of ESESi was determined by testing the initial adhesion. In the dry state, ESESi films show outstanding initial adhesion, as reported in Fig. 5, higher than those of the SESi and Tegaderm™ films. Surprisingly, the ESESi films become nearly non-adhesive when they reach saturation. This unique characteristic would significantly reduce the difficulty and pain involved in removing saturated wound dressings.

Fig. 5 Comparison of the initial adhesion of test samples in the dry and saturated states.

3.3 Water vapor transition rate (WVTR)

A good wound dressing must have an adequate WVTR. A suitable WVTR is beneficial for enabling vapor and gas exchange, which aid in the healing process. This paper tested films with a thickness of approximately 0.7 mm. According to the results, the introduction of PEG blocks resulted in a higher WVTR for the ESESi film than for the SESi film, as shown in Fig. 6. As with tensile strength, the WVTR also improved with an increasing PEG block ratio.

Fig. 6 Water vapor transition rate of ESESi films and SESi films.

3.4 Evaluation of hydrophilicity and water absorption of ESESi films

The water contact angles on the film surface were tested to determine the hydrophilicity of the ESESi and SESi films. Before modification by PEG, the SESi film is hydrophobic (113.1 ± 2.1°). However, as revealed in Fig. 7, the hydrophilicity of the ESESi films was enhanced as the PEG block ratio was increased. In other words, PEG blocks endow ESESi films with hydrophilicity.

Fig. 7 Static water contact angle of the films.

In addition to hydrophilicity, the water absorption of these films was also measured. The water absorption of the films was determined by soaking the films in normal saline at 37 °C. Similar to the results for hydrophilicity, the water absorption of the films was improved almost 50 fold at a PEG/PDMS ratio of 1/9 (Fig. 8).

Fig. 8 The normal saline absorption of ESESi films and SESi films.

3.5 Cytotoxicity test

Cytotoxicity is an extremely important property to consider for both in vitro and in vivo applications. In this paper, cell viability was used to determine the in vitro cytotoxicity of ESESi materials. ESESi_1/9 films were chosen for testing in all of the in vitro cytotoxicity and in vivo wound healing experiments due to its overall excellent performance.

In the MTT cytotoxicity test, the ESESi_1/9 film was soaked in RPMI 1640 culture medium for 24 h or 48 h. Then, the ESESi_1/9 film was removed from the media, and different dilutions of the conditioned media were used to evaluate cytotoxicity. Fig. 9 shows that cell viability was near 100% for all dilutions tested. The direct contact assay was also used to evaluate in vitro cytotoxicity. Fig. 10 shows the morphology of L929 cells after incubation with the ESESi film, an HDPE film (negative control), and an organotin-stabilized PVC film (positive control) for 24 h. These results indicate that cells cultured in the presence of the ESESi film were similar to those cultured with the negative control, with almost all of the cells in the form of blooms. Both cytotoxicity results showed that the ESESi film is non-toxic to L929 cells.

Fig. 9 Comparison of the cytotoxicity of different concentrations of diluted extract of ESESi film (25%, 50%, 75% and 100%), based on the MTT assay.

Fig. 10 Comparison on the cytotoxicity of (A) negative control (high-density polyethylene film, HDPE), (B) ESESi film, and (C) positive control (organotin-stabilized polyvinyl chloride film, PVC) against L929 cells, based on the direct contact test.

3.6 Bacterial permeation test

As wound dressings must provide protection against bacterial invasion, bacterial permeation tests were conducted. Vaseline gauze, Tegaderm™ film and SESi film were chosen as comparisons. As shown in Table 1, vaseline gauze cannot protect against E. coli or S. cremoris permeation. In contrast, ESESi films completely prevent bacterial invasion, as do the controls (SESi film and Tegaderm™ film). More details are presented in Fig. S6.†

Table 1 Bacterial permeation of vaseline gauze, Tegaderm™ film, SESi and ESESi films

	E. coli	S. cremoris
Vaseline gauze	12.8 × 10^7 CFU mL^−1	3.33 × 10^7 CFU mL^−1
Tegaderm™ film	0	0
SESi film	0	0
ESESi film	0	0

---

3.7 In vivo wound healing model in rats

3.7.1. Macroscopic examination of wound healing. The in vivo wound healing experiment demonstrated that the ESESi film, Tegaderm™ film, and SESi film exhibited better promotion of healing than the controls of vaseline gauze. The wounds covered with vaseline gauze were dehydrated, and a scab formed very early in the healing process. In contrast, the wounds covered by the ESESi film, Tegaderm™ film, and SESi film were more neat and moist, without apparent inflammatory exudates. The macroscopic appearances of the wounds at different healing times are presented in Fig. 11, and the statistic results were shown in Fig. 12. Although none significant difference (p < 0.05) could be found among the group of ESESi film, Tegaderm™ film, and SESi film, the significant level (p < 0.01) between ESESi film and vaseline gauze are higher than the other two groups (p < 0.05), especially on the 6^th, 8^th and 14^th day.

Fig. 11 Comparison of the promotion of wound healing by vaseline gauze, Tegaderm™ film, SESi film and ESESi film.

Fig. 12 The wound contraction of the wound dressings. Results (mean ± SD) were obtained from the four experimental groups: Tegaderm™ film (n = 5), vaseline gauze film (n = 5), SESi film (n = 5) and ESESi film (n = 5) (*: significant at p < 0.05; **: significant at p < 0.01).

According to these results, the ESESi film could promote greater average wound contraction in comparison with vaseline gauze, even slightly higher than SESi film and the commercially available Tegaderm™ film (Fig. 12). Both of these results indicate that the ESESi film a little better promotes the acceleration of wound healing.

3.7.2. Histological analysis. H&E staining of wound tissue was performed to precisely evaluate the conditions of the wounds during wound healing. The overall conditions of the wounds are depicted in Fig. S7,† but several higher magnification (200×) images were also acquired to enable more detailed analysis, as shown in Fig. 13.

Fig. 13 H&E staining of wound tissues from all of the groups. (Notes: IE, inflammatory exudates; BS, blood scab; CH, congestion and hemorrhage; GT, granulation tissue; BV, blood vessel; CF, collagenous fibers; E, re-epithelialization; MOCF, more organized fibroblasts).

In general, wound healing proceeded in three phases: (i) inflammation; (ii) proliferation, including the formation of granulation tissue; and (iii) matrix formation and remodeling. This sequential process requires the interaction of cells in the dermis and epidermis, as well as the activity of chemical mediators released from inflammatory cells, fibroblasts and keratinocytes.

According to histological observations, the wounds were in the inflammatory phase on the 4^th day post-wounding. In all wounds, the infiltration of lymphocyte cells and fibroblasts, as well as vasodilatation, could be observed. Moreover, the wound covered with the Tegaderm™ film formed a blood scab, and the wound covered with the SESi film displayed congestion and hemorrhage.

On the 6^th day post-wounding, congestion and hemorrhage were observed on the wound covered by the SESi film. In contrast with the other wounds, a large amount of granulation tissue was observed, and capillaries began to grow perpendicularly to the wound treated by the ESESi film. In addition, fibroblasts began to migrate along the fibrin threads in the wounds treated by the Tegaderm™, SESi and ESESi films.

On the 8^th day, congestion and hemorrhage could still be observed in the wound covered by the vaseline gauze film. At this point, the wounds covered by the Tegaderm™ film, SESi film and ESESi film were forming capillaries and fibroblasts, but this process was more organized for the wound covered by the ESESi film.

After 10 days, the wounds were in the proliferative phase, and the scabs on the wound covered with the Tegaderm™ film were reduced in size. At the same time, the wound covered with vaseline gauze began to form capillaries and fibroblasts. On the 14^th day, all surfaces of the test wounds were nearly healed. During this stage, fibroblasts were still proliferating, and the collagenous fibers became more organized. Moreover, skin cuticles began to form and the re-epithelialization of the wound treated by the ESESi film was clearly superior.

In summary, similar with the Tegaderm™ film and SESi film, the ESESi film could also able to create a wet wound-healing environment and promote the wound healing, compared with the vaseline gauze. In which the ESESi film showed slightly higher efficiency than the other groups from the view point of the re-epithelialization.

4. Conclusions

A novel hydrogen bond associated supramolecular elastomer (ESESi) film based on PDMS–PEG–PDMS tri-block copolymers was prepared. This material is semi-transparent, soft, and exhibits mechanical properties similar to those of cross-linked rubber. In addition, the ESESi films possess nice water absorption ability, water vapor permeability, adhesion, and biocompatibility, mainly due to the introduction of the hydrophilic and biocompatible PEG blocks. We further demonstrated that the ESESi films created a suitable wet wound-healing environment. Thus, to some degree, it could promote the wound healing process. These results make the absorptive PDMS film ESESi a promising material for application to wound dressings.

Acknowledgements

The authors acknowledge financial support from the National Natural Science Foundation of China (grants 51473051 and 51003032), the Science and Technology Planning Project of Guangdong Province (grant 2016A010103007) and the Fundamental Research Funds for the Central Universities, SCUT (grants 2015ZZ062).

Notes and references

1. A. Gosain and L. DiPietro, World J. Surg., 2004, 28, 321.
2. S. Lin, K. Chen and L. Run-Chu, Biomaterials, 2001, 2, 2999.
3. S. Purna and M. Babu, Collagen based dressings – a review, Burns, 2000, 26, 54.
4. J. Boateng, K. Matthews, H. Stevens and G. Eccleston, J. Pharm. Sci., 2008, 97, 2892.
5. C. Ratliff, Advance for Nurse Practitioners, 2008, 16(7), 32–35.
6. Z. Moore and H. Strapp, Br. J. Nurs., 2015, 24, S12.
7. M. Fonder, G. Lazarus, D. Cowan, B. Aronson-Cook, A. Kohli and A. Mamelak, J. Am. Acad. Dermatol., 2008, 58, 185.
8. D. Brett, The Journal of Wound, Ostomy and Continence Nursing, 2006, 33(suppl. 6), S3.
9. I. Anderson, Prof. Nurse, 2002, 18, 145.
10. G. Winter, Nature, 1962, 193, 293.
11. S. Seaman, J. Am. Podiatr. Med. Assoc., 2002, 92, 24.
12. L. Horken, G. Stansfield and M. Miller, J. Wound Care, 2009, 18, 298.
13. A. Thomas, K. Harding and K. Moore, Biomaterials, 2000, 21, 1797.
14. Y. Barnea, J. Weiss and E. Gur, Ther. Clin. Risk Manage., 2010, 6, 21.
15. H. Vermeulen, D. Ubbink, A. Goossens, R. de Vos and D. Legemate, Br. J. Surg., 2005, 6, 665.
16. A. Mohamed and M. Xing, Int. J. Burns Trauma, 2012, 2, 29.
17. J. Bianchi, J. Wound Care, 2001, 10, 225.
18. N. Trengove, M. Stacey, S. MacAuley, N. Bennett, J. Gibson, F. Burslem, G. Murphy and G. Schultz, Wound Repair Regen., 1999, 7, 442.
19. S. Thomas, J. Wound Care, 1992, 1, 27.
20. S. Palfreyman, E. Nelson and J. Michaels, BMJ, 2007, 335, 244.
21. J. McDonald and G. Whitesides, Acc. Chem. Res., 2002, 35, 491.
22. T. Goda, T. Konno, M. Takai, T. Moro and K. Ishihara, Biomaterials, 2006, 27, 5151.
23. P. Losi, E. Briganti, A. Magera, D. Spiller, C. Ristori, B. Battolla, M. Balderi, S. Kull, A. Balbarini, R. Di Stefano and G. Soldani, Biomaterials, 2010, 31, 5336.
24. E. Van den Kerckhove, K. Stappaerts, W. Boeckx, H. Van den, S. Monstrey, A. Van der Kelen and J. De Cubber, Burns, 2001, 27, 205.
25. H. Bi and Y. Jin, Int. J. Burns Trauma, 2013, 1, 63.
26. R. Xu, G. Luo, H. Xia, W. He, J. Zhao and B. Liu, et al., Biomaterials, 2015, 40, 1.
27. M. Fourman, B. Phillips, J. Fritz, N. Conkling, S. McClain and M. Simon, et al., Ann. Plast. Surg., 2014, 73, 150.
28. L. Yang, Y. Lin, L. Wang and A. Zhang, Polym. Chem., 2014, 5, 153.
29. A. Zhang, L. Yang, Y. Lin, H. Lu, Y. Qiu and Y. Su, J. Biomater. Sci., Polym. Ed., 2013, 24, 1883.
30. A. Zhang, W. Deng, Y. Lin, J. Ye, Y. Dong and Y. Lei, et al., J. Biomater. Sci., Polym. Ed., 2014, 25, 1346.
31. P. Cordier, F. Tournilhac, C. Soulié-Ziakovic and L. Leibler, Nature, 2008, 451, 977.
32. D. Montarnal, P. Cordier, C. Soulié-Ziakovic, F. Tournilhac and L. Leibler, J. Polym. Sci., Part A: Polym. Chem., 2008, 46, 7925.

---

Footnote

† Electronic supplementary information (ESI) available: Detailed characterization, including FTIR, DSC and XRD curves of ESESis, bacterial permeation results and the photos of H&E stained wound tissue (40×). See DOI: 10.1039/c6ra07146e

---