Research on Hyaluronic Acid/Polyethylene oxide Electrospun Fiber Membranes for Skin Defect Repair

Research on Hyaluronic Acid/Polyethylene oxide Electrospun Fiber Membranes for Skin Defect Repair | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Research on Hyaluronic Acid/Polyethylene oxide Electrospun Fiber Membranes for Skin Defect Repair Weiwei Zhao, Zhenfeng Song, Huanhuan Li, Suping Shi, Yonghui Zhang, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5315474/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Skin tissue defects are a common clinical issue. Electrospun nanofibrous membranes, mimicking the extracellular matrix (ECM) structure, show promising potential for skin defect repair. This study utilizes electrospinning technology to prepare a soft, lightweight, and breathable fibrous membrane wound dressing using hyaluronic acid and polyethylene oxide as substrates for skin defect repair. Scanning electron microscopy (SEM) measurements reveal an average diameter of 280.7 ± 10.24 nm for the fibers. The study evaluates the fibrous membrane's absorbance, hydrophilicity, porosity, water vapor permeability, and antibacterial property.and assesses wound healing and histological outcomes through in vivo experiments. The hyaluronic acid/polyethylene oxide (HA/PEO) combination demonstrated an excellent 92.53±2.32% wound healing rate and the thickest connective tissue thickness of (1597.04± 272.71μm) µm on day 14, along with the highest collagen area deposition （33.20 ± 5.58% ）. These results indicate that the hyaluronic acid/polyethylene oxide fibrous membrane exhibits excellent mechanical properties, hydrophilicity, biocompatibility, and significant potential for promoting wound healing. Physical sciences/Nanoscience and technology Physical sciences/Nanoscience and technology/Nanomedicine Skin defect Tissue engineering Electrospinning Hyaluronic acid Polyethylene oxide Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Highlights 1.HA is a natural polymer that, as a major component of the skin ECM,It has good hydrophilic character, biocompatibility, biodegradability and chemical modification ability. 2.PEO is a synthetic polymer with good biocompatibility and low toxicity, which can increase the spinnability of the blend fiber membrane and improve the mechanical properties. 3.This study combines natural polymer HA and synthetic polymer PEO, possessing more complete biological characteristics. 1. Introduction The skin, the largest organ in the human body, covers approximately 20,000 square centimeters and constitutes about 15% of an adult's total body weight. It is composed of three layers: the epidermis, dermis, and subcutaneous layer, and serves primarily as a protective barrier against microbial contamination, harmful substances, and excessive fluid loss. It also plays a role in regulating body temperature [1, 2]. Wound healing is traditionally divided into four distinct stages: (1) Hemostasis: occurring within hours post-injury, (2) Inflammation: where exposed collagen at the wound site activates the coagulation cascade (intrinsic and extrinsic pathways), initiating the inflammatory phase, (3) Proliferation: involving key processes such as epithelialization, angiogenesis, granulation tissue formation, and collagen deposition, and (4) Remodeling: characterized by the deposition of collagen networks [3]. Ideal wound healing results in the restoration of the normal anatomical structure, function, and appearance of the skin [4]. Current wound treatment strategies include: (1) autologous skin grafts [5], (2) allogenic skin grafts [6], (3) xenografts [7], (4) amniotic membrane [8], and (5) tissue engineering [9, 10]. The design of skin tissue engineering constructs involves a combination of seed cells, growth factors, and scaffold materials. When designing scaffold materials, factors such as biocompatibility, biodegradability, mechanical properties, structure, and manufacturing methods must be considered. Choosing appropriate biomaterials is a crucial factor in designing functional wound dressings [11]. Electrospun fibers, due to their nanoscale characteristics mimicking the natural extracellular matrix, have been studied as promising tissue engineering scaffolds. They serve as bacterial barriers, promoting hemostasis, absorbing excess exudate, and allowing adequate gas exchange. As shown in Figure 1，The electrospinning process typically consists of four components: a high-voltage power supply, a syringe pump, a spinneret, and a collector [12]. This process can be divided into the following four steps: (1) Formation of a Taylor cone from the polymer solution under an electric field; (2) Extension of the charged jet along a straight line under the electric field; (3) Thinning of the jet and increased electrostatic bending instability (also known as whipping instability); (4) Solidification of the jet into fibers, which are collected on a grounded collector [13]. The simplicity and versatility of this setup make it an ideal choice for tissue engineering, as it is compatible with a wide range of natural and synthetic polymers as well as various polymer combinations. The type of polymer used significantly affects the final properties of nanofiber wound dressings [14]. Hyaluronic acid (HA), also known as hyaluronan, is a high-molecular-weight polysaccharide composed of repeating disaccharide units of N-acetyl-D-glucosamine and D-glucuronic acid, with a molecular weight range of 1 kDa to 3 MDa. Commercially available hyaluronic acid is typically in the form of its sodium salt, known as sodium hyaluronate. Each α-1,4-glucuronic acid and β-1,3-N-acetyl-D-glucosamine disaccharide unit carries a negative charge, making it one of the most abundant glycosaminoglycans in the extracellular matrix, with the highest concentration in soft connective tissues. It is characterized by high swelling, water absorption, biodegradability, biocompatibility, and antibacterial properties. Additionally, hyaluronic acid can mimic biological properties, particularly those of the skin, which gives it significant potential in wound dressing applications [15]. Polyethylene oxide (PEO) is widely used due to its water solubility, hydrophilicity, high viscosity, and biocompatibility with other bioactive substances. It also possesses low toxicity, semi-crystallinity, and antibacterial properties [16, 17]. In this study, a mixed spinning solution was prepared using hyaluronic acid and polyethylene oxide as substrates, with chloroform as the solvent, and electrospinning technology was employed to produce nanofibrous membranes. The study examined the microscopic morphology, hydrophilicity, and the potential of these membranes to promote the healing of full-thickness skin defects in rats. This research provides valuable insights for the development of new wound dressings. 2. Materials And Methods 2.1 Main Experimental Materials Low Molecular Weight Sodium Hyaluronate (HA-TLM): Molecular weight 10 kDa-1.0 MDa, Huaxi Biotech Co., Ltd. Polyethylene Oxide (PEO): Molecular weight 4 kDa, average Mv 4,000,000, containing less than 1000 ppm of BHT, Sigma-Aldrich (Shanghai) Trading Co., Ltd. Cocamide DEA: 6501 Coconut Oil Diethanolamide, active agent with an amine value (mg KOH/g) ≤ 45, Karunbilu Skincare Raw Materials Store. 2.2 Preparation of HA/PEO and PEO Fiber Membranes Solution Preparation**:Weigh sodium hyaluronate and polyethylene oxide powder in a 1:1 ratio. Dissolve in chloroform with cocamide DEA as an activator, and mix in a 20 ml vial to obtain a 2% (w/v) solution of HA/PEO. Stir magnetically for 12 hours to remove bubbles, resulting in the HA/PEO spinning solution. We found that a concentration lower than 2% resulted in insufficient viscosity and poor fiber formation due to low surface tension, while concentrations above 2% resulted in excessively high viscosity, causing difficulties in spinning and needle clogging. Since pure hyaluronate cannot be electrospun alone, we compared it with pure PEO fibers. Prepare a 2% (w/v) PEO spinning solution using chloroform with cocamide DEA as an activator, and stir magnetically for 12 hours to remove bubbles. Electrospinning**: Pour the HA/PEO and PEO spinning solutions into separate 10 ml syringes. Set the electrospinning parameters: spinneret-to-collector distance at 15 cm, pump feed rate at 1 ml/h, voltage at 20 kV, and rotation speed at 300 rpm. Collect the fibers on a foil-covered drum. Spin at room temperature for 10 hours, then dry the spun membranes in a 37°C oven. 2.3 Characterization of HA/PEO and PEO Fiber Membranes 2.3.1 Surface Structure Analysis: Use a scanning electron microscope (SEM) to analyze the surface structure. Fix the prepared fiber membranes on a sample stage with double-sided conductive tape, gold-coat for 20 seconds, and observe under a SU8 100 SEM. Set the voltage to 3 kV and observe cross-sections at different magnifications. Randomly measure 50 fibers using Image software to calculate the average diameter and variance. 2.3.2 Water Absorption Test: Calculate the water absorption rate using the formula [18]. Cut the fully dried HA/PEO fiber membranes into approximately 2 cm × 2 cm squares. Weigh the initial dry mass, then immerse the samples in PBS and incubate at 37°C for 24 hours. After removing excess water with filter paper, weigh the samples again. Due to the high solubility of pure PEO fibers, we used commercially available hemostatic gauze as a control. Measure each sample six times and average the results. The water absorption rate is calculated as: Water Absorption Rate (%) = (W - W1) / W1 × 100%, where W is the wet weight after PBS soaking, and W1 is the initial dry weight. 2.3.3 Porosity Test: Measure the porosity of the electrospun fiber membranes using the liquid displacement technique [19]. Cut fully dried HA/PEO fiber membranes into approximately 2 cm × 2 cm squares. Record the mass (W1) and volume (V1) of each sample before immersion. Soak in 95% ethanol in a six-well cell culture plate, incubate at 37°C for 24 hours, then record the saturated mass (W2). Use pure PEO fibers and commercial hemostatic gauze as controls. Measure each sample six times and average the results. Porosity (P) is calculated as: P = (W2 - W1) / (p × V1), where P is the porosity, W1 and W2 are the weights before and after soaking, V1 is the volume before soaking, and p is the ethanol density. 2.3.4 Water Vapor Permeability Test: Determine the water vapor permeability by placing 5 ml of saline in a glass bottle with a screw cap, covering the bottle opening with the HA/PEO fiber membrane, and weighing (m1). Measure the membrane surface area (s), then incubate at 37°C for 24 hours (t). After incubation, weigh the membrane again (m2) [20]. Use pure PEO fibers and commercial hemostatic gauze as controls. Measure each sample six times and average the results. Water vapor transmission rate (WVTR) is calculated as: WVTR (g/m²/day) = (m1 - m2) / (s × t). 2.3.5 Antibacterial experiment The antibacterial test method in this study conducts quantitative antibacterial tests according to GB/T 20944.3 - 2008. The test bacteria are Gram-negative Escherichia coli (ATCC 25922TM) and Gram-positive Staphylococcus aureus (ATCC 25923TM). The test includes preparing bacterial suspensions and samples as well as determining colony counts. At the initial stage of the test, the required culture medium is prepared, and the culture medium, test tubes, and pipette tips are sterilized. Add about 3 - 5 ml of culture medium to each test tube and tilt it at a certain angle to form a slant for refrigeration. Use a bamboo stick to select a thallus for subculture and then place it in an incubator at 37 °C for cultivation. Prepare a liquid culture medium and culture the colonies to prepare a bacterial suspension. Use PBS without samples as a blank control group. Each sample in the test has 5 parallel control samples. Incubate the samples in a constant temperature oscillator at 37 ± 1 °C and 150 rpm/min for 10 hours. Use a spectrophotometer to measure the optical density (OD) at 600 nm. Record the OD value generated at 600 nm. 2.4 In Vivo Evaluation of HA/PEO and PEO Fiber Membranes for Skin Defect Repair 2.4.1 Animal Model Creation: All animal models used in these studies were conducted according to standard guidelines approved by the Ethics Committee of Xinxiang Medical University (# K-2021-033-01 ). This experiment was conducted in accordance with the guidelines for the Management and Use of Experimental Animals and the ARRIVE guidelines published by the national Science and Technology in animal husbandry management, experimental surgery, and euthanasia. Create a full-thickness skin defect model on the backs of male SD rats to evaluate the effects of different materials on wound healing. Select 18 male SD rats weighing 200-250 g, divide them into 3 groups of 6 rats each, housed separately with free access to food and water. After acclimatization, anesthetize the rats with pet-grade isoflurane, adjusting the oxygen flow and anesthetic dose appropriately. After anesthesia, place the rats on the operating table, shave and disinfect the skin on both sides of the back. Use a punch to create a 1 cm diameter full-thickness skin defect. Sterilize the PEO and HA/PEO fiber membranes under UV light for 1 hour and apply them to the wounds. Set up a gauze group as a control. Photograph the wounds at different time points for documentation. After the rats recover, they resume normal drinking and eating, indicating successful model establishment. 2.4.2 Sample Collection: At days 3, 7, and 14 post-wounding, anesthetize the rats with isoflurane, excise the full-thickness skin samples down to the superficial fascia, fix them in 4% paraformaldehyde for 24 hours, embed in paraffin, and section. After obtaining samples from rats, they were euthanized by spinal dislocation method according to the AVMA Guidelines for the Euthanasia of Animals. 2.4.3 Wound Area and Healing Rate Calculation: Measure the wound area using image software to determine the wound healing rate [21]. Calculate the wound healing rate as: Healing Rate = (1 - Area on Day N / Area on Day 0) × 100%, where N = 3, 7, 14, 21 days. 2.4.4 Histological Analysis: Fix tissue samples in 4% paraformaldehyde at 4°C for 48 hours, then embed in paraffin and perform staining for microscopic observation. Use Hematoxylin and Eosin (HE) staining to assess wound closure and Masson staining to evaluate collagen deposition. and capture images using an upright optical microscope. 2.5 Statistical Analysis All data are expressed as mean ± standard deviation (n ≥ 6). Analyze group differences using ANOVA. Statistical analysis is performed using GraphPad Prism 8.00, with a p-value of less than 0.05 considered statistically significant. 3. Results 3.1 Characterization of HA/PEO and PEO Fiber Membranes 3.1.1 Surface Structure Analysis by Scanning Electron Microscopy (SEM) Figure 2A shows the surface morphology of the poly (ethylene oxide) (PEO) and hyaluronic acid/poly (ethylene oxide) (HA/PEO) fiber membranes. SEM images reveal that the PEO fiber membrane is smooth and loosely structured with a beaded appearance, as shown in figure 2B .with an average diameter of 473.1 ± 25.94 nm, consistent with the diameter of pure PEO nanofibers [22]. In contrast, as shown in figure 2Bthe HA/PEO fiber membrane features finer fibers with a dense network structure and no beaded appearance, with an average diameter of 280.7 ± 10.24 nm. The rigidity of hyaluronic acid makes its electrospinning challenging; however, as an important component of the extracellular matrix (ECM), HA provides excellent hydrophilicity. PEO is known for its favorable electrospinning properties, indicating that the appropriate ratio of HA to PEO forms a mixed interpenetrating network structure. The smaller fiber diameter positively impacts cell proliferation [23]. 3.1.2 Fourier Transform Infrared Spectroscopy (FTIR) Analysis The chemical composition of the fiber membrane was also confirmed by the Fourier transform infrared spectroscopy instrument DTGS KBr. figure 3 shows the infrared characteristic absorption peaks in the range of 400 cm⁻¹ to 4000 cm⁻¹. For a pure PEO membrane, its typical absorption band is located at 1145.41 cm⁻¹, which is due to the stretching vibration of CH₂ [25]. We found the stretching vibration absorption peak of primary amide C-H at 2883.09 cm⁻¹, that is, -CH₂ and -CH₃, which is consistent with the PEO absorption band. The HA/PEO fiber membrane has very strong peaks at 1722.19 cm⁻¹ and 1167.41 cm⁻¹. The characteristic band of HA was also found near 1362.93 cm⁻¹, which can be attributed to the amide II vibration generated by the coupling of N-H bending and C-N stretching vibration of secondary amide [24]. 1722.19 cm⁻¹ is the stretching vibration absorption peak of carbonyl primary amide (C=O); 1294.5 cm⁻¹ and 1362.93 cm⁻¹ are the deformation vibration absorption peaks of secondary amide C-H [26]; 1167.41 cm⁻¹ is the deformation vibration absorption peak of C-N; 1047.65 cm⁻¹ is the stretching vibration absorption peak of C-O-C [27]; 959.67 cm⁻¹ is the out-of-plane rocking peak of {-CH₂-}n, which may be due to the coordination between hyaluronic acid and polyethylene oxide. The appearance of the above peaks indicates that hyaluronic acid has been successfully doped with polyethylene oxide. 3.1.3 Water Absorption Rate Analysi s The water absorption and moisture retention properties of wound dressings are crucial for wound healing. An ideal dressing should absorb metabolic products and wound exudate continuously, keeping the wound moist and promoting healing [28,29]. figure 4A and figure 4D compares the water absorption rates of HA/PEO fiber membranes and commercial gauze. The HA/PEO membranes show a significant increase in water absorption (1089.5% ± 128.5%) compared to commercial gauze (108.67% ± 10.50%), attributed to water molecule infiltration within the nanofiber network. This indicates the superior water absorption capability of the fabricated fiber membranes. 3.1.4 Porosity Analysis Porosity is a fundamental evaluation criterion for ideal wound dressings. High porosity facilitates gas exchange and cell respiration, with an ideal porosity for tissue engineering applications being at least 90% [30]. The high porosity of the fiber membranes ensures close resemblance to the natural ECM structure, enhancing cell compatibility [31]. The results in figure 4B and figure 4E show that the gauze group has a porosity of 34.16% ± 6.73%, the PEO group 97.16% ± 27.25%, and the HA/PEO group 169.03% ± 36.75%, significantly higher than both the gauze and PEO groups. The increased porosity with smaller fiber diameters suggests that HA/PEO membranes offer better cell adhesion and facilitate gas, nutrient, and fluid exchange, thereby accelerating wound healing. 3.1.5 Water Vapor Transmission Rate (WVTR) Analysis A low WVTR impedes wound healing by hindering liquid exchange between tissue cells, leading to exudate accumulation and bacterial contamination, thus slowing fibroblast proliferation and granulation tissue growth [32]. figure 4C and figure 4F show the results，The WVTR for commercial gauze is 2627.40 ± 309.92 g/m²/day, while pure PEO membranes have a WVTR of 2871.57 ± 302.41 g/m²/day. The HA/PEO membranes exhibit a significantly higher WVTR of 3619.39 ± 292.85 g/m²/day compared to both commercial gauze and pure PEO membranes. The high WVTR of HA/PEO membranes reduces moisture and temperature buildup at the wound site, lowering the risk of bacterial infection. Thus, HA/PEO membranes facilitate greater evaporation of wound exudate, preventing accumulation. 3.1.6 Antibacterial activity in vitro The antibacterial properties of PEO and HA/PEO nanofiber membranes at different concentrations against two typical pathogens, Escherichia coli and Staphylococcus aureus, were determined according to the OD600 value. As can be seen from the figure 5, the obtained nanofiber membranes have certain antibacterial properties against Escherichia coli and Staphylococcus aureus. Compared with Escherichia coli, PEO and HA/PEO nanofiber membranes at different concentrations have a more excellent inhibitory effect on Staphylococcus aureus. Among them, the HA/PEO nanofiber membrane with a concentration of 2% shows the best antibacterial effect. It can protect the wound surface from microbial infection, thereby promoting wound healing. 3.2 In Vivo Experiments 3.2.1 Rat Wound Healing Analysis Wound healing rate is a critical indicator of the effectiveness of medical dressings. Table 1 results show that that on day 3, the healing rates of the PEO and HA/PEO groups were（29.46±9.34%）and（49.95±9.45%）, respectively, compared to（22.56 ± 6.63%） for the gauze control group, indicating that both PEO and HA/PEO membranes influence full-thickness wound healing in SD rats. On day 7, the HA/PEO group exhibited a wound healing rate of （62.36±4.75%）, significantly higher than PEO (43.89±7.92%) and gauze control (36.45±11.17%). By day 14, the healing rates were 63.82% for the gauze group, (84.85±2.91%) for PEO, and (92.53±2.32%) for HA/PEO, The results are shown in figure 6A and figure 6C that with the HA/PEO group showing the most pronounced wound contraction and visible new granulation tissue. On day 21, the HA/PEO group's wounds were nearly closed, with new skin and hair nearing normalcy. The larger gaps between traditional gauze fibers may result in secondary damage during removal. These results suggest that the HA/PEO group outperforms both the gauze control and PEO groups in promoting wound healing. 3.2.2 Histological Analysis HE staining analysis As shown in Figure 7a，The results indicate that on day 7, all groups exhibited inflammatory cell infiltration in the wound tissue. The gauze control group showed a large number of inflammatory cells and significant defects, while the PEO and HA/PEO groups displayed increased fibroblasts and more collagen regeneration and gland formation. The HA/PEO group demonstrated superior epithelialization. As shown in Figure 7c,On day 14, the HA/PEO group had a connective tissue thickness of （1597.04±272.71um）, compared to （1250.03±127.53um) for the PEO group and （1029.39μm±259.11um） for the gauze control group. Histologically, the HA/PEO group showed well-organized epidermal cells, sparse inflammatory cells, and abundant new fibrous tissue. The gauze control group still had numerous inflammatory cells, incomplete epithelialization, and poorly defined skin layers with incomplete accessory structures. Masson Staining Results As shown in Figure 7B,Collagen content, a crucial indicator of wound healing, was analyzed using Masson’s trichrome staining, which colors collagen blue and muscle fibers red.As shown in Table 2 ,Image software analysis revealed that on day 7, the collagen area was（4.19%±4.05）for the gauze control group, （11.06%±6.94）for the PEO group, and （20.42%±9.27） for the HA/PEO group. On day 14, the collagen area in the HA/PEO group was （33.20%±5.58）, significantly higher than the gauze control group （9.27%±6.72）and PEO（17.11%±6.94) groups. The HA/PEO group showed more pronounced collagen deposition and wavy collagen fibers, closely resembling normal skin. Western blot protein blot analysis: To confirm that the nanofiber membrane reduces the inflammatory response, we conducted Western blot protein blot analysis. Figure 8 shows the expression of IL-1β and TNF-α in wound tissues treated with the gauze control group, PEO group, and HA/PEO group. It can be seen that compared with the PEO group and the gauze control group, the inflammatory response in the HA/PEO group is significantly reduced. The grayscale value of the protein band is quantified using ImageJ software. Conclusion and Future Outlook In summary, nanofiber membranes are a promising choice for wound dressings. Effective dressings must possess excellent biocompatibility, biodegradability, hydrophilicity, and tensile strength. Hyaluronic acid (HA), a natural polymer and major component of skin ECM, enhances wound healing by producing pro-inflammatory cytokines and promoting fibroblast proliferation and differentiation. HA's good hydrophilicity, biocompatibility, biodegradability, and chemical modification capabilities complement PEO, a synthetic polymer known for its biocompatibility and low toxicity. PEO improves the electrospinnability and mechanical properties of the blend. Given that existing natural and synthetic polymers do not fully exhibit these properties, this study utilizes electrospinning to prepare PEO and HA/PEO fiber membranes. In vitro characterization shows that these materials exhibit desirable fiber diameters, water absorption, porosity, and WVTR. In vivo experiments demonstrate their ability to accelerate wound healing, increase collagen deposition, and significantly reduce inflammation. Therefore, HA/PEO fiber membranes are a promising wound dressing for clinical application, with plans to extend research to large animal models in the near future. Declarations Ethics approval and consent to participate All protocols for experimental in animals were approved by the Animal Care and Experimental Committee of Xinxiang Medical University ( # K-2021-033-01 ) CRediT authorship contribution statement Weiwei Zhao: Writing – original draft, Visualization, Software, Resources, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. Zhenfeng Song: Visualization, Methodology. Huanhuan Li: Conceptualization, Investigation. Suping Shi : Data curation. Hongying Chen: Data curation, Xianwei Wang: Funding acquisition. Zhikun Guo: Funding acquisition. Yongkun Sun: Writing – review & editing, Funding acquisition, Conceptualization. Conflict of interest The authors declare no competing interests. Data Availability Statement The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request. Funds : Supported by the National Key R&D Program Grant (2018YFC1105800), Open Project of the Third Affiliated Hospital of Xinxiang Medical University (KFKTZD202106), Open Program of Henan Joint International Research Laboratory of Stem Cell Medicine (KFKT202105), Henan Key Laboratory of Medical and Protective Products (KFKT2023). References Pastushenko, I., et al., Skin Stem Cells: At the Frontier Between the Laboratory and Clinical Practice. Part 1: Epidermal Stem Cells. Actas Dermosifiliogr, 2015. 106(9): p. 725-32. Wong, R., et al., The dynamic anatomy and patterning of skin. Exp Dermatol, 2016. 25(2): p. 92-8. Broughton, G.N., J.E. Janis and C.E. 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Liu, X., et al., Polyvinylpyrrolidone/chitosan-loaded dihydromyricetin-based nanofiber membrane promotes diabetic wound healing by anti-inflammatory and regulating autophagy-associated protein expression. Int J Biol Macromol, 2024. 259(Pt 1): p. 129160. Additional Declarations No competing interests reported. Supplementary Files Tab1.png Table 1 :In vivo experiment: Wound healing area (mm) measured in male SD rats on days 0, 3, 7, and 14. Tab2.png Table 2 :In vivo experiment: Collagen staining area (%) by Masson staining. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5315474","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":377954537,"identity":"4794576a-4e23-40a9-b34e-da6a65abb376","order_by":0,"name":"Weiwei Zhao","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAzUlEQVRIiWNgGAWjYBACNmbmww8+VNjYMbY3HyBOCx87W5rhjDNpycw9xxKI0yLHz2Mgzdt2mLF9Ro4BsQ5jMDCcwZbGzDsj5+ONNwx2croNhLUkPPjAY8Mn2fN2s+UchmRjswOEtRwwnCGRxmzYnrtNmofhQOI2wloYG6R5DA4z7j+Q84xYLcwM0jwJhxkbO3LYiNXCxmY440BaMmPPMWPLOQZE+EW+//znBx//gaPy4Y03FXZyBLWgAAkeIqMGWQupOkbBKBgFo2BEAADZAkCVmps1wAAAAABJRU5ErkJggg==","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":true,"prefix":"","firstName":"Weiwei","middleName":"","lastName":"Zhao","suffix":""},{"id":377954538,"identity":"bff3c673-1e18-49ca-877e-89b20bb8ec37","order_by":1,"name":"Zhenfeng Song","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Zhenfeng","middleName":"","lastName":"Song","suffix":""},{"id":377954539,"identity":"16b57eda-9289-47a6-8ad3-29c29c641c87","order_by":2,"name":"Huanhuan Li","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Huanhuan","middleName":"","lastName":"Li","suffix":""},{"id":377954540,"identity":"2286ef4e-91fa-43a1-b53f-d0b1b4da8334","order_by":3,"name":"Suping Shi","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Suping","middleName":"","lastName":"Shi","suffix":""},{"id":377954541,"identity":"fa7dda5e-5548-4e46-b30c-c8269eff7c0e","order_by":4,"name":"Yonghui Zhang","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Yonghui","middleName":"","lastName":"Zhang","suffix":""},{"id":377954542,"identity":"7a9221f7-0398-4bb5-bad8-fa7d357fff47","order_by":5,"name":"Xianwei Wang","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Xianwei","middleName":"","lastName":"Wang","suffix":""},{"id":377954543,"identity":"d274a77b-949e-4dec-8b39-5d701dc7cd89","order_by":6,"name":"Zhikun Guo","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Zhikun","middleName":"","lastName":"Guo","suffix":""},{"id":377954544,"identity":"d0b2910d-04b1-4211-979a-f92579f2fd90","order_by":7,"name":"Yongkun Sun","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Yongkun","middleName":"","lastName":"Sun","suffix":""},{"id":377954545,"identity":"b04eaa60-aa11-4aae-adaf-735138c85dea","order_by":8,"name":"Hongying Chen","email":"","orcid":"","institution":"Xinxiang Medical University","correspondingAuthor":false,"prefix":"","firstName":"Hongying","middleName":"","lastName":"Chen","suffix":""}],"badges":[],"createdAt":"2024-10-23 04:08:14","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5315474/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5315474/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":69995741,"identity":"e4854fae-4f57-47dc-a0e8-30d0bd1bd739","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":116772,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic diagram of electrospinning process.\u003c/p\u003e","description":"","filename":"Slide1.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/09587faf39fb667e209de6f0.png"},{"id":69996695,"identity":"9243dc7f-2d11-44ed-b337-84ae46b77c4c","added_by":"auto","created_at":"2024-11-27 10:29:20","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":237505,"visible":true,"origin":"","legend":"\u003cp\u003eMaterial characterization: A is the scanning electron microscope image of PEO and HA/PEO. B is the frequency distribution of PEO fiber diameter. C is the frequency distribution of HA/PEO fiber diameter. D is the statistical analysis of the diameter size of HA/PEO and PEO. ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Slide2.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/3f2fbabab0bd5e25d5a3a580.png"},{"id":69995743,"identity":"a3f56f1c-4b3e-44f7-8d28-bd8bb15f5162","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":69580,"visible":true,"origin":"","legend":"\u003cp\u003eMaterial characterization: The upper one is the infrared characteristic absorption peak of HA/PEO nanofiber membrane. The lower one is the infrared characteristic absorption peak of PEO nanofiber membrane.\u003c/p\u003e","description":"","filename":"Slide3.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/734c2810c17d64187d6b3783.png"},{"id":69995738,"identity":"c5af3be3-e7c6-4038-b7c0-78686793c89a","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":123963,"visible":true,"origin":"","legend":"\u003cp\u003eA shows the dry weight and wet weight of HA/PEO fiber membrane and gauze before and after soaking in PBS for 24 hours, and D is the water absorption analysis; B shows the dry weight and wet weight of HA/PEO fiber membrane, PEO fiber membrane and gauze before and after soaking in 95% absolute ethanol, and E is the statistical analysis of the porosity of HA/PEO fiber membrane, PEO fiber membrane and gauze; C shows the dry weight and wet weight of HA/PEO fiber membrane, PEO fiber membrane and gauze before and after soaking in normal saline, and F is the statistical analysis of the water vapor transmission rate of HA/PEO fiber membrane, PEO fiber membrane and gauze. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Slide4.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/dfaa7e7ff8c470d3ddd8c7c0.png"},{"id":69995982,"identity":"4c036de1-5cba-4f4c-860f-df6a8563cd53","added_by":"auto","created_at":"2024-11-27 10:21:20","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":133412,"visible":true,"origin":"","legend":"\u003cp\u003eA shows the OD values for inhibiting Escherichia coli at different concentrations, and B shows the OD values for inhibiting Staphylococcus aureus at different concentrations.\u003c/p\u003e","description":"","filename":"Slide5.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/a8ae5a7c0a39cdb1ad9278dd.png"},{"id":69995748,"identity":"5cf3cb6c-711d-402b-916c-8958b2cfbb9d","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":334171,"visible":true,"origin":"","legend":"\u003cp\u003ePanel A shows the wound area of three groups of SD male rats on days 0, 3, 7, 14, and 21 of the experiment. The scale bar = 1 cm. Panel B shows the statistical analysis of the wound healing rate. Panel C is the contour map of the wound healing process. Panel D is the curve of the wound healing rate. *p \u0026lt; 0.05, ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Slide6.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/d12fe9171ae8b533c6e873ba.png"},{"id":69995745,"identity":"02353831-2f93-4f7f-99d7-b6e78bc80c81","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":761421,"visible":true,"origin":"","legend":"\u003cp\u003ePanel A shows HE staining results and Masson staining results. HE staining results: The green rectangle represents the junction area between the normal incision and normal skin. The scale bars are 500 μm and 100 μm respectively. (Blue arrow: epithelial layer; green arrow: sweat gland; purple arrow: new capillaries; red arrow: hair follicle; black arrow: fibroblasts.); Masson staining results: scale bar = 500 μm; Panel B shows the statistical results of the area of collagen after Masson staining. The area of collagen in normal skin is set as 100%. Panel C shows the statistical results of the thickness of connective tissue on day 14. *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Slide8.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/b649d175a37f762bdbe701b7.png"},{"id":69995746,"identity":"b8f0e7a4-8819-40e6-8d7b-a6675c7648d9","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":133785,"visible":true,"origin":"","legend":"\u003cp\u003eAnalysis of the Western Blot imprinting of IL-1 β and TNF- α in the three groups of wound tissues *p \u0026lt; 0.05, **p \u0026lt; 0.01, ***p \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Slide9.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/4798e311d895c191887a94d6.png"},{"id":77669935,"identity":"fa122835-f9fe-4679-84ac-92934d0d1b03","added_by":"auto","created_at":"2025-03-04 06:47:14","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2944809,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/83915840-653c-496d-bf16-e80188a85856.pdf"},{"id":69995736,"identity":"fcfc01a8-5215-4fbf-aff8-80b2e8f26228","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":25417,"visible":true,"origin":"","legend":"\u003cp\u003eTable 1 :In vivo experiment: Wound healing area (mm) measured in male SD rats on days 0, 3, 7, and 14.\u003c/p\u003e","description":"","filename":"Tab1.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/9037cbf5c1ffde7ac4f58cd8.png"},{"id":69995737,"identity":"1d699539-ea53-4b40-be6a-958b4e06602b","added_by":"auto","created_at":"2024-11-27 10:13:20","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":17550,"visible":true,"origin":"","legend":"\u003cp\u003eTable 2 :In vivo experiment: Collagen staining area (%) by Masson staining.\u003c/p\u003e","description":"","filename":"Tab2.png","url":"https://assets-eu.researchsquare.com/files/rs-5315474/v1/e6bfe7072b97d594e965690a.png"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eResearch on Hyaluronic Acid/Polyethylene oxide Electrospun Fiber Membranes for Skin Defect Repair\u003c/p\u003e","fulltext":[{"header":"Highlights","content":"\u003cp\u003e1.HA is a natural polymer that, as a major component of the skin ECM,It has good hydrophilic character, biocompatibility, biodegradability and chemical modification ability.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.PEO is a synthetic polymer with good biocompatibility and low toxicity, which can increase the spinnability of the blend fiber membrane and improve the mechanical properties.\u003c/p\u003e\n\u003cp\u003e3.This study combines natural polymer HA and synthetic polymer PEO, possessing more complete biological characteristics.\u003c/p\u003e"},{"header":"1. Introduction","content":"\u003cp\u003eThe skin, the largest organ in the human body, covers approximately 20,000 square centimeters and constitutes about 15% of an adult\u0026apos;s total body weight. It is composed of three layers: the epidermis, dermis, and subcutaneous layer, and serves primarily as a protective barrier against microbial contamination, harmful substances, and excessive fluid loss. It also plays a role in regulating body temperature [1, 2]. Wound healing is traditionally divided into four distinct stages: (1) Hemostasis: occurring within hours post-injury, (2) Inflammation: where exposed collagen at the wound site activates the coagulation cascade (intrinsic and extrinsic pathways), initiating the inflammatory phase, (3) Proliferation: involving key processes such as epithelialization, angiogenesis, granulation tissue formation, and collagen deposition, and (4) Remodeling: characterized by the deposition of collagen networks [3]. Ideal wound healing results in the restoration of the normal anatomical structure, function, and appearance of the skin [4]. Current wound treatment strategies include: (1) autologous skin grafts [5], (2) allogenic skin grafts [6], (3) xenografts [7], (4) amniotic membrane [8], and (5) tissue engineering [9, 10]. The design of skin tissue engineering constructs involves a combination of seed cells, growth factors, and scaffold materials. When designing scaffold materials, factors such as biocompatibility, biodegradability, mechanical properties, structure, and manufacturing methods must be considered. Choosing appropriate biomaterials is a crucial factor in designing functional wound dressings [11].\u003c/p\u003e\n\u003cp\u003eElectrospun fibers, due to their nanoscale characteristics mimicking the natural extracellular matrix, have been studied as promising tissue engineering scaffolds. They serve as bacterial barriers, promoting hemostasis, absorbing excess exudate, and allowing adequate gas exchange. As shown in Figure 1，The electrospinning process typically consists of four components: a high-voltage power supply, a syringe pump, a spinneret, and a collector [12]. This process can be divided into the following four steps: (1) Formation of a Taylor cone from the polymer solution under an electric field; (2) Extension of the charged jet along a straight line under the electric field; (3) Thinning of the jet and increased electrostatic bending instability (also known as whipping instability); (4) Solidification of the jet into fibers, which are collected on a grounded collector [13]. The simplicity and versatility of this setup make it an ideal choice for tissue engineering, as it is compatible with a wide range of natural and synthetic polymers as well as various polymer combinations. The type of polymer used significantly affects the final properties of nanofiber wound dressings [14].\u003c/p\u003e\n\u003cp\u003eHyaluronic acid (HA), also known as hyaluronan, is a high-molecular-weight polysaccharide composed of repeating disaccharide units of N-acetyl-D-glucosamine and D-glucuronic acid, with a molecular weight range of 1 kDa to 3 MDa. Commercially available hyaluronic acid is typically in the form of its sodium salt, known as sodium hyaluronate. Each \u0026alpha;-1,4-glucuronic acid and \u0026beta;-1,3-N-acetyl-D-glucosamine disaccharide unit carries a negative charge, making it one of the most abundant glycosaminoglycans in the extracellular matrix, with the highest concentration in soft connective tissues. It is characterized by high swelling, water absorption, biodegradability, biocompatibility, and antibacterial properties. Additionally, hyaluronic acid can mimic biological properties, particularly those of the skin, which gives it significant potential in wound dressing applications [15].\u003c/p\u003e\n\u003cp\u003ePolyethylene oxide (PEO) is widely used due to its water solubility, hydrophilicity, high viscosity, and biocompatibility with other bioactive substances. It also possesses low toxicity, semi-crystallinity, and antibacterial properties [16, 17]. In this study, a mixed spinning solution was prepared using hyaluronic acid and polyethylene oxide as substrates, with chloroform as the solvent, and electrospinning technology was employed to produce nanofibrous membranes. The study examined the microscopic morphology, hydrophilicity, and the potential of these membranes to promote the healing of full-thickness skin defects in rats. This research provides valuable insights for the development of new wound dressings.\u003c/p\u003e"},{"header":"2. Materials And Methods","content":"\u003cp\u003e\u003cstrong\u003e2.1 Main Experimental Materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eLow Molecular Weight Sodium Hyaluronate (HA-TLM): Molecular weight 10 kDa-1.0 MDa, Huaxi Biotech Co., Ltd.\u003c/p\u003e\n\u003cp\u003ePolyethylene Oxide (PEO): Molecular weight 4 kDa, average Mv 4,000,000, containing less than 1000 ppm of BHT, Sigma-Aldrich (Shanghai) Trading Co., Ltd.\u003c/p\u003e\n\u003cp\u003eCocamide DEA: 6501 Coconut Oil Diethanolamide, active agent with an amine value (mg KOH/g) \u0026le; 45, Karunbilu Skincare Raw Materials Store.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2 Preparation of HA/PEO and PEO Fiber Membranes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSolution Preparation**:Weigh sodium hyaluronate and polyethylene oxide powder in a 1:1 ratio. Dissolve in chloroform with cocamide DEA as an activator, and mix in a 20 ml vial to obtain a 2% (w/v) solution of HA/PEO. Stir magnetically for 12 hours to remove bubbles, resulting in the HA/PEO spinning solution. We found that a concentration lower than 2% resulted in insufficient viscosity and poor fiber formation due to low surface tension, while concentrations above 2% resulted in excessively high viscosity, causing difficulties in spinning and needle clogging. Since pure hyaluronate cannot be electrospun alone, we compared it with pure PEO fibers. Prepare a 2% (w/v) PEO spinning solution using chloroform with cocamide DEA as an activator, and stir magnetically for 12 hours to remove bubbles.\u003c/p\u003e\n\u003cp\u003eElectrospinning**: Pour the HA/PEO and PEO spinning solutions into separate 10 ml syringes. Set the electrospinning parameters: spinneret-to-collector distance at 15 cm, pump feed rate at 1 ml/h, voltage at 20 kV, and rotation speed at 300 rpm. Collect the fibers on a foil-covered drum. Spin at room temperature for 10 hours, then dry the spun membranes in a 37\u0026deg;C oven.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3 Characterization of HA/PEO and PEO Fiber Membranes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3.1 Surface Structure Analysis:\u003c/strong\u003e Use a scanning electron microscope (SEM) to analyze the surface structure. Fix the prepared fiber membranes on a sample stage with double-sided conductive tape, gold-coat for 20 seconds, and observe under a SU8 100 SEM. Set the voltage to 3 kV and observe cross-sections at different magnifications. Randomly measure 50 fibers using Image software to calculate the average diameter and variance.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3.2 Water Absorption Test:\u003c/strong\u003e Calculate the water absorption rate using the formula [18]. Cut the fully dried HA/PEO fiber membranes into approximately 2 cm \u0026times; 2 cm squares. Weigh the initial dry mass, then immerse the samples in PBS and incubate at 37\u0026deg;C for 24 hours. After removing excess water with filter paper, weigh the samples again. Due to the high solubility of pure PEO fibers, we used commercially available hemostatic gauze as a control. Measure each sample six times and average the results. The water absorption rate is calculated as: Water Absorption Rate (%) = (W - W1) / W1 \u0026times; 100%, where W is the wet weight after PBS soaking, and W1 is the initial dry weight.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3.3 Porosity Test:\u003c/strong\u003e Measure the porosity of the electrospun fiber membranes using the liquid displacement technique [19]. Cut fully dried HA/PEO fiber membranes into approximately 2 cm \u0026times; 2 cm squares. Record the mass (W1) and volume (V1) of each sample before immersion. Soak in 95% ethanol in a six-well cell culture plate, incubate at 37\u0026deg;C for 24 hours, then record the saturated mass (W2). Use pure PEO fibers and commercial hemostatic gauze as controls. Measure each sample six times and average the results. Porosity (P) is calculated as: P = (W2 - W1) / (p \u0026times; V1), where P is the porosity, W1 and W2 are the weights before and after soaking, V1 is the volume before soaking, and p is the ethanol density.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3.4 Water Vapor Permeability Test:\u0026nbsp;\u003c/strong\u003eDetermine the water vapor permeability by placing 5 ml of saline in a glass bottle with a screw cap, covering the bottle opening with the HA/PEO fiber membrane, and weighing (m1). Measure the membrane surface area (s), then incubate at 37\u0026deg;C for 24 hours (t). After incubation, weigh the membrane again (m2) [20]. Use pure PEO fibers and commercial hemostatic gauze as controls. Measure each sample six times and average the results. Water vapor transmission rate (WVTR) is calculated as: WVTR (g/m\u0026sup2;/day) = (m1 - m2) / (s \u0026times; t).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.3.5 Antibacterial experiment\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe antibacterial test method in this study conducts quantitative antibacterial tests according to GB/T 20944.3 - 2008. The test bacteria are Gram-negative Escherichia coli (ATCC 25922TM) and Gram-positive Staphylococcus aureus (ATCC 25923TM). The test includes preparing bacterial suspensions and samples as well as determining colony counts. At the initial stage of the test, the required culture medium is prepared, and the culture medium, test tubes, and pipette tips are sterilized. Add about 3 - 5 ml of culture medium to each test tube and tilt it at a certain angle to form a slant for refrigeration. Use a bamboo stick to select a thallus for subculture and then place it in an incubator at 37 \u0026deg;C for cultivation. Prepare a liquid culture medium and culture the colonies to prepare a bacterial suspension. Use PBS without samples as a blank control group. Each sample in the test has 5 parallel control samples. Incubate the samples in a constant temperature oscillator at 37 \u0026plusmn; 1 \u0026deg;C and 150 rpm/min for 10 hours. Use a spectrophotometer to measure the optical density (OD) at 600 nm. Record the OD value generated at 600 nm.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.4 In Vivo Evaluation of HA/PEO and PEO Fiber Membranes for Skin Defect Repair\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.4.1 Animal Model Creation:\u0026nbsp;\u003c/strong\u003eAll animal models used in these studies were conducted according to standard guidelines approved by the Ethics Committee of\u0026nbsp;Xinxiang Medical University\u0026nbsp;(#\u003cstrong\u003eK-2021-033-01\u003c/strong\u003e).\u0026nbsp;This experiment was conducted in accordance with the guidelines for the Management and Use of Experimental Animals and the ARRIVE guidelines published by the national Science and Technology in animal husbandry management, experimental surgery, and euthanasia.\u0026nbsp;Create a full-thickness skin defect model on the backs of male SD rats to evaluate the effects of different materials on wound healing. Select 18 male SD rats weighing 200-250 g, divide them into 3 groups of 6 rats each, housed separately with free access to food and water. After acclimatization, anesthetize the rats with pet-grade isoflurane, adjusting the oxygen flow and anesthetic dose appropriately. After anesthesia, place the rats on the operating table, shave and disinfect the skin on both sides of the back. Use a punch to create a 1 cm diameter full-thickness skin defect. Sterilize the PEO and HA/PEO fiber membranes under UV light for 1 hour and apply them to the wounds. Set up a gauze group as a control. Photograph the wounds at different time points for documentation. After the rats recover, they resume normal drinking and eating, indicating successful model establishment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.4.2 Sample Collection:\u0026nbsp;\u003c/strong\u003eAt days 3, 7, and 14 post-wounding, anesthetize the rats with isoflurane, excise the full-thickness skin samples down to the superficial fascia, fix them in 4% paraformaldehyde for 24 hours, embed in paraffin, and section.\u0026nbsp;After obtaining samples from rats, they were euthanized by spinal dislocation method according to the AVMA Guidelines for the Euthanasia of Animals.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.4.3 Wound Area and Healing Rate Calculation:\u003c/strong\u003e Measure the wound area using image software to determine the wound healing rate [21]. Calculate the wound healing rate as: Healing Rate = (1 - Area on Day N / Area on Day 0) \u0026times; 100%, where N = 3, 7, 14, 21 days.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.4.4 Histological Analysis:\u003c/strong\u003e Fix tissue samples in 4% paraformaldehyde at 4\u0026deg;C for 48 hours, then embed in paraffin and perform staining for microscopic observation. Use Hematoxylin and Eosin (HE) staining to assess wound closure and Masson staining to evaluate collagen deposition. and capture images using an upright optical microscope.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.5 Statistical Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll data are expressed as mean \u0026plusmn; standard deviation (n \u0026ge; 6). Analyze group differences using ANOVA. Statistical analysis is performed using GraphPad Prism 8.00, with a p-value of less than 0.05 considered statistically significant.\u0026nbsp;\u003c/p\u003e"},{"header":"3. Results","content":"\u003cp\u003e\u003cstrong\u003e3.1 Characterization of HA/PEO and PEO Fiber Membranes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.1 Surface Structure Analysis by Scanning Electron Microscopy (SEM)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFigure 2A shows the surface morphology of the poly (ethylene oxide) (PEO) and hyaluronic acid/poly (ethylene oxide) (HA/PEO) fiber membranes. SEM images reveal that the PEO fiber membrane is smooth and loosely structured with a beaded appearance, as shown in figure 2B .with an average diameter of 473.1 \u0026plusmn; 25.94 nm, consistent with the diameter of pure PEO nanofibers [22]. In contrast, as shown in figure 2Bthe HA/PEO fiber membrane features finer fibers with a dense network structure and no beaded appearance, with an average diameter of 280.7 \u0026plusmn; 10.24 nm. The rigidity of hyaluronic acid makes its electrospinning challenging; however, as an important component of the extracellular matrix (ECM), HA provides excellent hydrophilicity. PEO is known for its favorable electrospinning properties, indicating that the appropriate ratio of HA to PEO forms a mixed interpenetrating network structure. The smaller fiber diameter positively impacts cell proliferation [23].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.2 Fourier Transform Infrared Spectroscopy (FTIR) Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe chemical composition of the fiber membrane was also confirmed by the Fourier transform infrared spectroscopy instrument DTGS KBr.\u0026nbsp;figure 3 shows the infrared characteristic absorption peaks in the range of 400 cm⁻\u0026sup1; to 4000 cm⁻\u0026sup1;. For a pure PEO membrane, its typical absorption band is located at 1145.41 cm⁻\u0026sup1;, which is due to the stretching vibration of CH₂ [25]. We found the stretching vibration absorption peak of primary amide C-H at 2883.09 cm⁻\u0026sup1;, that is, -CH₂ and -CH₃, which is consistent with the PEO absorption band. The HA/PEO fiber membrane has very strong peaks at 1722.19 cm⁻\u0026sup1; and 1167.41 cm⁻\u0026sup1;. The characteristic band of HA was also found near 1362.93 cm⁻\u0026sup1;, which can be attributed to the amide II vibration generated by the coupling of N-H bending and C-N stretching vibration of secondary amide [24]. 1722.19 cm⁻\u0026sup1; is the stretching vibration absorption peak of carbonyl primary amide (C=O); 1294.5 cm⁻\u0026sup1; and 1362.93 cm⁻\u0026sup1; are the deformation vibration absorption peaks of secondary amide C-H [26]; 1167.41 cm⁻\u0026sup1; is the deformation vibration absorption peak of C-N; 1047.65 cm⁻\u0026sup1; is the stretching vibration absorption peak of C-O-C [27]; 959.67 cm⁻\u0026sup1; is the out-of-plane rocking peak of {-CH₂-}n, which may be due to the coordination between hyaluronic acid and polyethylene oxide. The appearance of the above peaks indicates that hyaluronic acid has been successfully doped with polyethylene oxide.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.3 Water Absorption Rate Analysi\u003c/strong\u003es\u003c/p\u003e\n\u003cp\u003eThe water absorption and moisture retention properties of wound dressings are crucial for wound healing. An ideal dressing should absorb metabolic products and wound exudate continuously, keeping the wound moist and promoting healing [28,29]. figure 4A and figure 4D compares the water absorption rates of HA/PEO fiber membranes and commercial gauze. The HA/PEO membranes show a significant increase in water absorption (1089.5% \u0026plusmn; 128.5%) compared to commercial gauze (108.67% \u0026plusmn; 10.50%), attributed to water molecule infiltration within the nanofiber network. This indicates the superior water absorption capability of the fabricated fiber membranes.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.4 Porosity Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePorosity is a fundamental evaluation criterion for ideal wound dressings. High porosity facilitates gas exchange and cell respiration, with an ideal porosity for tissue engineering applications being at least 90% [30]. The high porosity of the fiber membranes ensures close resemblance to the natural ECM structure, enhancing cell compatibility [31]. The results in figure 4B and figure 4E show that the gauze group has a porosity of 34.16% \u0026plusmn; 6.73%, the PEO group 97.16% \u0026plusmn; 27.25%, and the HA/PEO group 169.03% \u0026plusmn; 36.75%, significantly higher than both the gauze and PEO groups. The increased porosity with smaller fiber diameters suggests that HA/PEO membranes offer better cell adhesion and facilitate gas, nutrient, and fluid exchange, thereby accelerating wound healing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.5 Water Vapor Transmission Rate (WVTR) Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA low WVTR impedes wound healing by hindering liquid exchange between tissue cells, leading to exudate accumulation and bacterial contamination, thus slowing fibroblast proliferation and granulation tissue growth [32]. figure 4C and figure 4F show the results，The WVTR for commercial gauze is 2627.40 \u0026plusmn; 309.92 g/m\u0026sup2;/day, while pure PEO membranes have a WVTR of 2871.57 \u0026plusmn; 302.41 g/m\u0026sup2;/day. The HA/PEO membranes exhibit a significantly higher WVTR of 3619.39 \u0026plusmn; 292.85 g/m\u0026sup2;/day compared to both commercial gauze and pure PEO membranes. The high WVTR of HA/PEO membranes reduces moisture and temperature buildup at the wound site, lowering the risk of bacterial infection. Thus, HA/PEO membranes facilitate greater evaporation of wound exudate, preventing accumulation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.1.6 Antibacterial activity in vitro\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe antibacterial properties of PEO and HA/PEO nanofiber membranes at different concentrations against two typical pathogens, Escherichia coli and Staphylococcus aureus, were determined according to the OD600 value. As can be seen from the figure\u0026nbsp;5, the obtained nanofiber membranes have certain antibacterial properties against Escherichia coli and Staphylococcus aureus. Compared with Escherichia coli, PEO and HA/PEO nanofiber membranes at different concentrations have a more excellent inhibitory effect on Staphylococcus aureus. Among them, the HA/PEO nanofiber membrane with a concentration of 2% shows the best antibacterial effect. It can protect the wound surface from microbial infection, thereby promoting wound healing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.2 \u003cem\u003eIn Vivo\u003c/em\u003e Experiments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.2.1 Rat Wound Healing Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWound healing rate is a critical indicator of the effectiveness of medical dressings. Table 1 results show that that on day 3, the healing rates of the PEO and HA/PEO groups were（29.46\u0026plusmn;9.34%）and（49.95\u0026plusmn;9.45%）, respectively, compared to（22.56\u0026nbsp;\u0026plusmn;\u0026nbsp;6.63%）\u0026nbsp;for the gauze control group, indicating that both PEO and HA/PEO membranes influence full-thickness wound healing in SD rats. On day 7, the HA/PEO group exhibited a wound healing rate of\u0026nbsp;（62.36\u0026plusmn;4.75%）, significantly higher than PEO (43.89\u0026plusmn;7.92%) and gauze control (36.45\u0026plusmn;11.17%). By day 14, the healing rates were 63.82% for the gauze group, (84.85\u0026plusmn;2.91%) for PEO, and (92.53\u0026plusmn;2.32%) for HA/PEO, The results are shown in figure 6A and figure 6C that with the HA/PEO group showing the most pronounced wound contraction and visible new granulation tissue. On day 21, the HA/PEO group\u0026apos;s wounds were nearly closed, with new skin and hair nearing normalcy. The larger gaps between traditional gauze fibers may result in secondary damage during removal. These results suggest that the HA/PEO group outperforms both the gauze control and PEO groups in promoting wound healing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.2.2 Histological Analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eHE staining analysis\u003c/p\u003e\n\u003cp\u003eAs shown in Figure 7a，The results indicate that on day 7, all groups exhibited inflammatory cell infiltration in the wound tissue. The gauze control group showed a large number of inflammatory cells and significant defects, while the PEO and HA/PEO groups displayed increased fibroblasts and more collagen regeneration and gland formation. The HA/PEO group demonstrated superior epithelialization. As shown in Figure 7c,On day 14, the HA/PEO group had a connective tissue thickness of\u0026nbsp;（1597.04\u0026plusmn;272.71um）, compared to\u0026nbsp;（1250.03\u0026plusmn;127.53um) for the PEO group and\u0026nbsp;（1029.39\u0026mu;m\u0026plusmn;259.11um）\u0026nbsp;for the \u0026nbsp;gauze control group. Histologically, the HA/PEO group showed well-organized epidermal cells, sparse inflammatory cells, and abundant new fibrous tissue. The gauze control group still had numerous inflammatory cells, incomplete epithelialization, and poorly defined skin layers with incomplete accessory structures.\u003c/p\u003e\n\u003cp\u003eMasson Staining Results\u003c/p\u003e\n\u003cp\u003eAs shown in Figure 7B,Collagen content, a crucial indicator of wound healing, was analyzed using Masson\u0026rsquo;s trichrome staining, which colors collagen blue and muscle fibers red.As shown in Table 2 ,Image software analysis revealed that on day 7, the collagen area was（4.19%\u0026plusmn;4.05）for the gauze control group,\u0026nbsp;（11.06%\u0026plusmn;6.94）for the PEO group, and\u0026nbsp;（20.42%\u0026plusmn;9.27）\u0026nbsp;for the HA/PEO group. On day 14, the collagen area in the HA/PEO group was\u0026nbsp;（33.20%\u0026plusmn;5.58）, significantly higher than the gauze control group\u0026nbsp;（9.27%\u0026plusmn;6.72）and PEO（17.11%\u0026plusmn;6.94) groups. The HA/PEO group showed more pronounced collagen deposition and wavy collagen fibers, closely resembling normal skin.\u003c/p\u003e\n\u003cp\u003eWestern blot protein blot analysis: To confirm that the nanofiber membrane reduces the inflammatory response, we conducted Western blot protein blot analysis. Figure 8 shows the expression of IL-1\u0026beta; and TNF-\u0026alpha; in wound tissues treated with the gauze control group, PEO group, and HA/PEO group. It can be seen that compared with the PEO group and the gauze control group, the inflammatory response in the HA/PEO group is significantly reduced. The grayscale value of the protein band is quantified using ImageJ software.\u003c/p\u003e"},{"header":"Conclusion and Future Outlook","content":"\u003cp\u003eIn summary, nanofiber membranes are a promising choice for wound dressings. Effective dressings must possess excellent biocompatibility, biodegradability, hydrophilicity, and tensile strength. Hyaluronic acid (HA), a natural polymer and major component of skin ECM, enhances wound healing by producing pro-inflammatory cytokines and promoting fibroblast proliferation and differentiation. HA\u0026apos;s good hydrophilicity, biocompatibility, biodegradability, and chemical modification capabilities complement PEO, a synthetic polymer known for its biocompatibility and low toxicity. PEO improves the electrospinnability and mechanical properties of the blend. Given that existing natural and synthetic polymers do not fully exhibit these properties, this study utilizes electrospinning to prepare PEO and HA/PEO fiber membranes. In vitro characterization shows that these materials exhibit desirable fiber diameters, water absorption, porosity, and WVTR. In vivo experiments demonstrate their ability to accelerate wound healing, increase collagen deposition, and significantly reduce inflammation. Therefore, HA/PEO fiber membranes are a promising wound dressing for clinical application, with plans to extend research to large animal models in the near future.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll protocols for experimental in animals were approved by the Animal Care and Experimental Committee of Xinxiang Medical University (\u003cstrong\u003e#\u003c/strong\u003e\u003cstrong\u003eK-2021-033-01\u003c/strong\u003e)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCRediT authorship contribution statement\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eWeiwei Zhao:\u0026nbsp;\u003c/strong\u003eWriting \u0026ndash; original draft, Visualization, Software, Resources, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. \u003cstrong\u003eZhenfeng Song:\u0026nbsp;\u003c/strong\u003eVisualization, Methodology. \u003cstrong\u003eHuanhuan\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;Li:\u0026nbsp;\u003c/strong\u003eConceptualization, Investigation.\u003cstrong\u003eSuping Shi\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eData curation. \u003cstrong\u003eHongying Chen:\u003c/strong\u003eData curation,\u003cstrong\u003eXianwei Wang:\u0026nbsp;\u003c/strong\u003eFunding acquisition. \u003cstrong\u003eZhikun Guo:\u0026nbsp;\u003c/strong\u003eFunding acquisition. \u003cstrong\u003eYongkun Sun:\u0026nbsp;\u003c/strong\u003eWriting \u0026ndash; review \u0026amp; editing, Funding acquisition, Conceptualization.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunds\u003c/strong\u003e\u003cstrong\u003e:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSupported by the National Key R\u0026amp;D Program Grant (2018YFC1105800), Open Project of the Third Affiliated Hospital of Xinxiang Medical University (KFKTZD202106), Open Program of Henan Joint International Research Laboratory of Stem Cell Medicine (KFKT202105), Henan Key Laboratory of Medical and Protective Products (KFKT2023).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003ePastushenko, I., et al., Skin Stem Cells: At the Frontier Between the Laboratory and Clinical Practice. 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Biomater Adv, 2022. 134: p. 112554\u003c/li\u003e\n \u003cli\u003eGuo, J., et al., Preparation and evaluation of dual drug-loaded nanofiber membranes based on coaxial electrostatic spinning technology. Int J Pharm, 2022. 629: p. 122410.\u003c/li\u003e\n \u003cli\u003eFouad, H., et al., Development of New Bio-Composite of PEO/Silk Fibroin Blends Loaded with Piezoelectric Material. Polymers (Basel), 2022. 14(19).\u003c/li\u003e\n \u003cli\u003eChen, H., et al., Molecular Interaction, Chain Conformation, and Rheological Modification during Electrospinning of Hyaluronic Acid Aqueous Solution. Membranes (Basel), 2020. 10(9).\u003c/li\u003e\n \u003cli\u003eSHIN Y C,SHIN D M,LEE E J,\u003cem\u003eet al\u003c/em\u003e.Hyaluronic Acid/PLGA Core/Shell Fiber Matrices Loaded with EGCG Beneficial to Diabetic Wound Healing [J] .\u003cem\u003eAdv Healthc Mater\u003c/em\u003e, 2016, 5(23):3035-3045.\u003c/li\u003e\n \u003cli\u003eSagawa, N. and T. Shikata, Hydration behavior of poly(ethylene oxide)s in aqueous solution as studied by near-infrared spectroscopic techniques. J Phys Chem B, 2013. 117(37): p. 10883-8.\u003c/li\u003e\n \u003cli\u003eDong, Q., et al., Understanding hyaluronic acid induced variation of water structure by near-infrared spectroscopy. Sci Rep, 2020. 10(1): p. 1387.\u003c/li\u003e\n \u003cli\u003eGuo, J., et al., Preparation and evaluation of dual drug-loaded nanofiber membranes based on coaxial electrostatic spinning technology. Int J Pharm, 2022. 629: p. 122410.\u003c/li\u003e\n \u003cli\u003eBazmandeh, A.Z., et al., Dual spinneret electrospun nanofibrous/gel structure of chitosan-gelatin/chitosan-hyaluronic acid as a wound dressing: In-vitro and in-vivo studies. Int J Biol Macromol, 2020. 162: p. 359-373.\u003c/li\u003e\n \u003cli\u003eDu P, et al., In vivo and in vitro studies of a propolis-enriched silk fibroin-gelatin composite nanofiber wound dressing. Heliyon, 2023. 9(3): p. e13506.\u003c/li\u003e\n \u003cli\u003e左凌楠等, 基于壳聚糖的静电纺丝纳米纤维膜用于伤口敷料的研究进展. 中国药学杂志, 2019. 54(14): 第1126-1131页.\u003c/li\u003e\n \u003cli\u003eSafonova, L., et al., Silk Fibroin/Spidroin Electrospun Scaffolds for Full-Thickness Skin Wound Healing in Rats. Pharmaceutics, 2021. 13(10).\u003c/li\u003e\n \u003cli\u003eLiu, X., et al., Polyvinylpyrrolidone/chitosan-loaded dihydromyricetin-based nanofiber membrane promotes diabetic wound healing by anti-inflammatory and regulating autophagy-associated protein expression. Int J Biol Macromol, 2024. 259(Pt 1): p. 129160.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Skin defect, Tissue engineering, Electrospinning, Hyaluronic acid, Polyethylene oxide","lastPublishedDoi":"10.21203/rs.3.rs-5315474/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5315474/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Skin tissue defects are a common clinical issue. Electrospun nanofibrous membranes, mimicking the extracellular matrix (ECM) structure, show promising potential for skin defect repair. This study utilizes electrospinning technology to prepare a soft, lightweight, and breathable fibrous membrane wound dressing using hyaluronic acid and polyethylene oxide as substrates for skin defect repair. Scanning electron microscopy (SEM) measurements reveal an average diameter of 280.7 ± 10.24 nm for the fibers. The study evaluates the fibrous membrane's absorbance, hydrophilicity, porosity, water vapor permeability, and antibacterial property.and assesses wound healing and histological outcomes through in vivo experiments. The hyaluronic acid/polyethylene oxide (HA/PEO) combination demonstrated an excellent 92.53±2.32% wound healing rate and the thickest connective tissue thickness of (1597.04± 272.71μm) µm on day 14, along with the highest collagen area deposition （33.20 ± 5.58% ）. These results indicate that the hyaluronic acid/polyethylene oxide fibrous membrane exhibits excellent mechanical properties, hydrophilicity, biocompatibility, and significant potential for promoting wound healing.","manuscriptTitle":"Research on Hyaluronic Acid/Polyethylene oxide Electrospun Fiber Membranes for Skin Defect Repair","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-11-27 10:13:15","doi":"10.21203/rs.3.rs-5315474/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"b772b88e-d60e-4162-b445-77c118267d5c","owner":[],"postedDate":"November 27th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":40825544,"name":"Physical sciences/Nanoscience and technology"},{"id":40825545,"name":"Physical sciences/Nanoscience and technology/Nanomedicine"}],"tags":[],"updatedAt":"2025-03-04T06:39:04+00:00","versionOfRecord":[],"versionCreatedAt":"2024-11-27 10:13:15","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5315474","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5315474","identity":"rs-5315474","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}