Hydrogel inspired by "adobe" with antibacterial and antioxidant properties for diabetic wound healing

Abstract

With the aging population, the incidence of diabetes is increasing. Diabetes often leads to restricted neovascularization, antibiotic-resistant bacterial infections, reduced wound perfusion, and elevated reactive oxygen species, resulting in impaired microenvironments and prolonged wound healing. Hydrogels are important tissue engineering materials for wound healing, known for their high water content and good biocompatibility. However, most hydrogels suffer from poor mechanical properties and difficulty in achieving sustained drug release, hindering their clinical application. Inspired by the incorporation of fibers to enhance the mechanical properties of "adobe," core-shell fibers were introduced into the hydrogel. This not only improves the mechanical strength of the hydrogel but also enables the possibility of sustained drug release. In this study, we first prepared core-shell fibers with PLGA (poly(lactic-co-glycolic acid)) and PCL (polycaprolactone). PLGA was loaded with P2 (Parathyroid hormone-related peptides-2), developed by our group, which promotes angiogenesis and cell proliferation. We then designed a QTG (QCS/TA/Gel, quaternary ammonium chitosan/tannic acid/gelatin) hydrogel, incorporating the core-shell fibers and the anti-inflammatory drug celecoxib into the QTG hydrogel. This hydrogel exhibits excellent antibacterial properties and biocompatibility, along with good mechanical performance. This hydrogel demonstrates excellent water absorption and swelling capabilities. In the early stages of wound healing, the hydrogel can absorb the wound exudate, maintaining the stability of the wound microenvironment. This hydrogel promotes neovascularization and collagen deposition, accelerating the healing of diabetic wounds, with a healing rate exceeding 95 % by day 14. Overall, this study provides a promising strategy for developing tissue engineering scaffolds for diabetic wound healing.

Keywords: Antibacterial; Antioxidant; Core-shell fibers; Diabetic wound healing; Hydrogel.

Graphical abstract

Scheme. 1

Schematic illustration of the molecular design, fabrication process, and treatment of diabetic wounds with QTG@C@PPP hydrogel. (a) Chemical structures of QCS, TA and Gel. (b) The flow chart of the hydrogel preparation. (c) The treatment process and mechanism of action of the hydrogel on diabetic wounds in rats.

Fig. 1

Formation and mechanism of QTG@C@PPP. (a) and (b) FT-IR spectra of the QTG hydrogel and PPP electrospun fibers. (c) Zeta potential of QTG@C@PPP. (d) TEM image of the PPP electrospun fibers. (e) Pore size distribution analysis for (f). (f)–(h) SEM images of the QTG@C@PPP at magnifications of 100x, 200x, and 5000x. (i)–(k) EDS elemental analysis of (g).

Fig. 2

Characterization of QTG@C@PPP. (a) Degradation behavior of hydrogels in vitro. (b) Degradation behavior of electrospun fibers in vitro. (c) Swelling behavior. (d) Water absorption performance. (e) P2 in vitro release. (f) Cel in vitro release. (g) Rheological properties. (h) Compressive stress-strain curve. (i) Tensile stress-strain curve.

Fig. 3

Characterization of QTG@C@PPP. (a) and (b) Macroscopic views of the hydrogel. (c) Photographs of the hydrogel under tensile stretching at different angles. (d) Photographs demonstrating the adhesion of the hydrogel to various materials(The white dashed lines indicate QTG@C@PPP). (e) Images demonstrating the injectability of the hydrogel. (f) and (g) Images demonstrating the transparency of the hydrogel.

Fig. 4

Antibacterial properties of QTG@C@PPP. (a), (b) and (c) Images from the inhibition zone measurement showing the effects of the hydrogel on S. aureus, E. coli, and MRSA. (d), (e) and (f) Quantitative statistics from the inhibition zone measurement. (g) and (h) Images from the plate coating method for S. aureus, E. coli, and MRSA.(i), (j) and (k) Quantitative statistics from the plate coating method for S. aureus, E. coli, and MRSA.

Fig. 5

Antioxidant properties of QTG@C@PPP. (a) DCFH-DA fluorescence images of cells in the presence of 1 mM H₂O₂ for each group. (b) and (c) Statistical analysis of DCFH-DA fluorescence intensity. (d) Detection of DPPH radical scavenging ability.

Fig. 6

Biocompatibility and cell proliferation-promoting properties of QTG@C@PPP. (a) and (b) CCK-8 results of cells on days 1, 3 and 5. (c) Live/Dead staining images of cells. (d) and (e) Statistical analysis of cell viability. (f) The images of the EdU proliferation assay for cells. (g) and (h) Statistical analysis of EdU-positive cell rates.

Fig. 7

The properties in promoting cell migration and protecting mitochondria of QTG@C@PPP. (a) Transwell assay images of cells. (b) and (c) Statistical analysis of migration efficiency.

Fig. 8

The properties in promoting tube formation of QTG@C@PPP. (a) Bright-field and fluorescence images of the tube formation assay. (b) Statistical analysis of the number of tubes. (c)–(e) Statistical analysis of the total tube length, number of segments, and number of junctions in the tube formation.

Fig. 9

The expression of COL-1, VEGF, FGF, and CD31 in HUVEC. (a)–(d) Statistical analysis of relevant genes in HUVEC using qRT-PCR. (e) The immunofluorescence staining images of relevant genes in HUVEC. (f)-(i)Statistical analysis of immunofluorescence intensity.

Fig. 10

In vivo assessment of QTG@C@PPP for wound healing. (a) The images of wounds at days 0, 3, 7 and 14 after different treatments. (b) Healing traces of different treatment groups at day 14. (c) -(e)Statistical analysis of wound healing rates in different treatment groups at days 3, 7 and 14.

Fig. 11

Histological evaluation of wound regeneration. (a) H&E stained images of wound tissues in each group at days 3, 7 and 14. (b) Masson trichrome stained images of wound tissues in each group (at days 3, 7, and 14). (c) Statistical analysis of epithelial layer thickness at day 14 in H&E stained images (epithelial thickness indicated between two yellow dashed lines). (d)–(f) Collagen density at different time points in each group. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 12

Immunohistochemical analysis of regenerated wound tissue. (a) Immunohistochemical staining images of wound tissues at day 14. (b)–(e) Statistical data of the relative expression levels of inflammatory factors and vascular formation-related factors at day 14.