Large-Scale Nanofiber Manufacturing| Electrospun robust, biodegradable, bioactive, and nanostructuredsutures to accelerate the chronic wound healing

Professor Wu Shaohua (Qingdao University) & Assistant Researcher Zhang Li (Huashan Hospital): Absorbable, High-Strength, Bioactive Nanofiber Sutures Significantly Promote Chronic Diabetic Wound Healing

Sutures are essential medical devices for postoperative tissue closure and wound healing. However, diabetic patients have far weaker self-healing capabilities than normal patients, making sutures critically important for diabetic wound recovery. Currently, most surgical sutures lack sufficient biofunctionality, and there are no specialized sutures for diabetic chronic wounds. Additionally, traditional sutures' microfibers differ significantly from natural extracellular matrix (ECM) nanofibers, hindering cell activity and tissue regeneration.

Recently, Professor Wu Shaohua’s team (Qingdao University) and Assistant Researcher Zhang Li (Huashan Hospital) published research in Biofabrication titled "Electrospun Robust, Biodegradable, Bioactive, and Nanostructured Sutures to Accelerate Chronic Wound Healing" (first author: Li Yiran, Qingdao University). The team developed poly(L-lactic acid) (PLLA) nanofiber sutures loaded with a Salvia-Gegen herbal compound (SRHC) via electrospinning and thermal stretching. These sutures exhibit excellent mechanical properties, biocompatibility, and drug release capabilities that suppress inflammation while promoting angiogenesis, collagen deposition, and hair follicle regeneration, significantly accelerating healing in diabetic mice.

Fig. 1: Suture morphology/structure.

During the research process, three types of sutures were prepared based on the SRHC loading amount, named as drug-free PLLA, 1%SRHC/PLLA with 1% drug loading, and 5%SRHC/PLLA with 5% drug loading. SEM images showed that all three types of nanofiber sutures (PLLA, 1%SRHC/PLLA, and 5%SRHC/PLLA) had similar morphology and structure, consisting of continuous fibers without beading and uniform morphology, which could significantly improve operability and patient comfort. The fibers in all three sutures exhibited high orientation along the suture axis, with fiber diameters of 270.3±80.2 nm, 265.3±42.5 nm, and 264.3±51.5 nm, respectively, similar to the diameter of natural ECM collagen fibrils (50-500 nm).

Fig. 2: Physicochemical properties

FTIR spectra showed slight peak enhancement at ~1644 cm-1 in the two drug-loaded sutures, corresponding to C=O and C=C in puerarin and danshen. XRD results indicated that all three sutures exhibited diffraction peaks at ~16.5°, corresponding to the (110)/(200) planes of the α'-type crystal. PLLA material has certain hydrophobicity, and the addition of SRHC did not significantly affect the hydrophilicity of the sutures. Furthermore, due to the slow degradation rate of PLLA under mild conditions, the mass of all three sutures did not show significant changes after 21 days of in vitro degradation. However, the mass loss was greater in the two drug-loaded sutures due to drug release effects. DPPH antioxidant activity tests showed that as the drug loading concentration increased, the DPPH radical scavenging rate of the sutures significantly improved, and higher sample leachate concentrations also resulted in higher DPPH radical scavenging rates.

Fig. 3: Mechanical performance.

All three PLLA nanofiber sutures exhibited similar tensile mechanical behavior: after the linear elastic region, they yielded and entered the strengthening region before finally fracturing. Compared to the mechanical parameters of PLLA sutures (breaking load: 2.6±0.21 N, tensile strength: 34.5±2.9 MPa, initial modulus: 1260.9±150.8 MPa, elongation at break: 32.3±11.4%), the addition of SRHC did not significantly affect the mechanical properties. The 1% and 5% drug-loaded PLLA nanofiber sutures maintained breaking loads of 2.6±0.3 N and 2.6±0.2 N, tensile strengths of 39.0±4.0 MPa and 38.4±3.0 MPa, initial moduli of 1433.8±344.2 MPa and 1547.3±365.5 MPa, and elongation at break values of 27.7±4.9% and 27.6±2.7%, respectively. Knot strength tests showed that the knotted breaking strength was reduced by 26.9%~30.8% compared to regular tensile breaking strength, and the tensile strength was reduced by 27.2%~30.3%. The initial modulus after knotting did not decrease significantly, while the maximum elongation at break decreased to 17.6±3.3%, 18.2±1.5%, and 17.2±2.6%.

Fig. 4: Cell migration.

First, MTT staining was used to observe the migration trajectory of human dermal fibroblasts (HDFs) on the sutures. Drug-free PLLA nanofiber sutures were tightly arranged to form a high-density strip fabric, and HDFs were seeded within a 5 mm restricted area at one end of the fabric. After 14 days of normal culture, the cell migration process was recorded. During the two-week culture period, the purple trajectory along the PLLA nanofiber sutures extended to approximately 20 mm. Additionally, scratch assays demonstrated SRHC's excellent guidance function for cell migration. The cell scratches cultured in 5%SRHC/PLLA suture leachate almost disappeared after 12 hours, while other groups still showed significant gaps.

Fig. 5: Cytotoxicity/anti-inflammatory effects

HDFs were seeded on the surfaces of all three PLLA nanofiber sutures, and after 3 days of co-culture, the cytoskeleton (green) and nuclei (blue) were stained to observe cell morphology. All three suture surfaces were uniformly adhered to by numerous spindle-shaped cells with highly extended filamentous cytoskeletons, and the cell extension direction closely aligned with the nanofiber orientation, indicating that the microstructure of PLLA nanofiber sutures could effectively regulate cell elongation and alignment. MTT assays performed on days 1 and 3 showed that HDFs could survive on all three PLLA nanofiber suture surfaces. By day 3, the cell viability in all three groups was significantly higher than on day 1, demonstrating that HDFs could adhere well to the suture surfaces and perform normal cellular activities. Moreover, the addition of SRHC further provided favorable conditions for cell growth and proliferation.

Mouse macrophages activated by LPS were cultured in leachates from the three sutures, and the supernatants were collected for ELISA testing. The control group and drug-free group showed similar inflammation levels, while the two drug-loaded groups exhibited significantly reduced concentrations of pro-inflammatory factors TNF-α and IL-6, along with markedly increased expression levels of anti-inflammatory factors IL-10 and IL-13. 

Fig. 6: Diabetic wound healing progression.

Continuously recorded wound images showed that during the first 3 postoperative days, wounds treated with all four sutures were similar, with visible incision marks and suture materials. By day 6, most of the three PLLA nanofiber sutures had degraded and fallen off, and the wounds had closed, with slight linear scars visible to the naked eye. By day 12, the scar in the PLLA group showed no significant change compared to the previous time point, the 1%SRHC/PLLA group had a smoother wound surface, while the 5%SRHC/PLLA group only retained scars at the suture sites, with clearly visible white new hair growth. In contrast, the polyester suture control group showed basically healed wounds by day 12 but with obvious scars and non-degradable suture materials remaining.

Fig. 7: Histological analysis.

Skin tissue from diabetic mouse incisions on day 12 was collected for histological analysis using H&E and MT staining. All four suture-treated wounds showed complete epidermal layers, but only the 1%SRHC/PLLA and 5%SRHC/PLLA groups exhibited relatively continuous dermal tissue and hair follicle distribution, with the 5%SRHC/PLLA group showing the highest structural integrity and maturity of new skin tissue. The control group contained numerous inflammatory cells, while the 5% drug-loaded group had significantly fewer inflammatory cells. In MT-stained images, the control, PLLA, and 1%SRHC/PLLA groups contained abundant blue collagen fibers, with disordered arrangement in the control group and more aligned fibers in the nanofiber suture groups. The 5%SRHC/PLLA group showed the most organized and mature collagen fiber arrangement, with new tissue structure approaching normal.

Fig. 8: Immunofluorescence (Ki67/CD31).

Ki67 is an important indicator of cell division activity. Red Ki67 staining showed that the fluorescence area and intensity in the 5%SRHC/PLLA group were significantly higher than in the other three groups. Semi-quantitative analysis revealed that compared to the control group's fluorescence intensity, the relative fluorescence intensities of the two drug-loaded groups reached ~1.9-fold and ~5.0-fold, respectively, proving that drug-loaded PLLA nanofiber sutures could effectively promote cell proliferation, especially when the drug loading reached 5%, where cell proliferation speed increased substantially.

CD31 staining of vascular endothelial cells was used to detect angiogenesis in wound tissues. Almost no fluorescence was observed in the control group, while sparse CD31 signals were seen in the PLLA and 1%SRHC/PLLA groups. The 5%SRHC/PLLA group showed more red fluorescence-labeled sites. Statistical results indicated that the relative fluorescence intensity of the 5%SRHC/PLLA experimental group was ~2.1-fold that of the control group, demonstrating that 5%SRHC/PLLA nanofiber sutures could effectively promote vascular regeneration at wound sites, thereby shortening healing time.