Abstract
Background
Diabetic wound infections, exacerbated by multidrug-resistant pathogens
like MRSA, remain a critical challenge due to biofilm persistence and
dysregulated oxidative-inflammatory-metabolic crosstalk.
Results
In this work, we engineered COG-Z@P200 hydrogel, a chitosan-based
hydrogel integrating polydopamine-coated ZIF-8 nanoparticles, to
synergize mild photothermal therapy (40–45 °C) with metabolic-immune
reprogramming. Upon NIR irradiation, COG-Z@P200 disrupted MRSA through
Zn^2⁺-mediated membrane destabilization and localized hyperthermia,
achieving >99.5% eradication via combined physical puncture and
metabolic interference. Multi-omics analyses revealed suppression of
glycolysis (eno, gap downregulation), TCA cycle arrested (sucC, sdhA,
icd inhibition), and disruption of arginine biosynthesis (arcA, arcC,
arcD downregulation), impairing biofilm formation and pathogenicity.
Concurrent silencing of quorum sensing and virulence genes (agr, sec,
lac, opp, sdrD) further destabilized MRSA, while upregulation of
stress-response genes (yidD, nfsA, kdpA) indicated bacterial metabolic
paralysis. In diabetic murine models, the hydrogel attenuated oxidative
stress (DHE-confirmed ROS reduction), polarized macrophages to
pro-healing M2 phenotypes (Arg-1⁺/TNF-α↓), and enhanced angiogenesis
(VEGF/CD31↑) alongside aligned collagen deposition. This
multifunctional action accelerated wound closure by 48% versus
controls, outperforming clinical standards. By converging
nanomaterial-enabled bactericidal strategies with host microenvironment
recalibration, COG-Z@P200 hydrogel redefined diabetic wound management,
offering an antibiotic-free solution against multidrug-resistant
infections.
Conclusion
Our work established a biomaterial paradigm that concurrently targets
pathogen vulnerabilities and restores tissue homeostasis, addressing
the multidimensional complexity of chronic wounds.
Graphical abstract
[44]graphic file with name 12951_2025_3451_Figa_HTML.jpg
Supplementary Information
The online version contains supplementary material available at
10.1186/s12951-025-03451-6.
Keywords: Multifunctional hydrogel dressing, ZIF-8 nanoparticles,
Photothermal therapy, Metabolic interference, Diabetic wound
microenvironment
Introduction
Diabetic wound infections (such as diabetic foot ulcers) have become a
major burden on global public health due to their high risk of
complications and prolonged non-healing characteristics [[45]1, [46]2].
Hyperglycemia, excessive accumulation of reactive oxygen species (ROS)
and infection with multidrug-resistant bacteria (such as MRSA) in the
chronic wound microenvironment jointly lead to inflammatory disorders
[[47]3–[48]6], blocked angiogenesis and failure of extracellular matrix
(ECM) remodeling, ultimately forming a vicious cycle [[49]5–[50]10].
Traditional treatment strategies (such as systemic administration of
antibiotics and local debridement) are often limited in efficacy due to
the inability to simultaneously regulate the microenvironment,
especially the synergistic deterioration of drug-resistant bacterial
infection and high-sugar microenvironment [[51]3, [52]11–[53]13].
Therefore, the development of new multifunctional therapeutic platforms
to simultaneously address infection control, metabolic regulation and
tissue regeneration has become a core direction of current research
[[54]7, [55]14–[56]16].
Photothermal agents (PTAs) such as polydopamine (PDA), a natural analog
of melanin, have shown great potential as effective PTAs due to their
ease of synthesis, excellent photostability, biodegradability, and
biocompatibility [[57]17–[58]19]. PDA can convert light energy into
heat, leading to bacterial cell membrane damage, DNA damage, protein
denaturation, and subsequent bacterial death. However, conventional PTT
often requires temperatures above 60 °C, which can damage healthy
tissues and limit its therapeutic potential in vivo [[59]20]. Recent
studies suggest that maintaining temperatures below 50 °C can minimize
damage to normal tissues while still achieving effective bacterial
eradication [[60]21, [61]22].
In addition to PTT, Zeolitic Imidazolate Framework-8 (ZIF-8) has
emerged as a promising material for wound healing applications. ZIF-8
is a metal–organic framework (MOF) known for its high specific surface
area, porosity, and excellent biocompatibility, making it an ideal
candidate for drug delivery and antibacterial applications [[62]23].
ZIF-8 exerts its antibacterial effect by disrupting bacterial cell
membranes and generating reactive oxygen species (ROS), which inhibit
bacterial growth and reduce the risk of infection [[63]24, [64]25].
Moreover, ZIF-8 can be utilized as a carrier for the controlled release
of growth factors, anti-inflammatory agents, and antimicrobial drugs,
thereby promoting wound healing and tissue regeneration [[65]26].
Despite the potential of ZIF-8, its application in wound healing has
been limited by several challenges, including its stability and the
need for more efficient antibacterial strategies. To address these
limitations, multicomponent synergistic therapies have emerged as a
promising solution. By combining various therapeutic components, such
as metal nanoparticles, PDA, and hydrogels, it is possible to enhance
the antibacterial effect, reduce the risk of bacterial resistance, and
accelerate wound healing [[66]27–[67]29]. For example, metal
nanoparticles, such as those incorporated in hydrogels, can enhance the
PTT effect by preventing nanoparticle aggregation, improving the
porosity structure [[68]28, [69]29], and providing mild antibacterial
properties at low temperatures.
This study presents a multifunctional hydrogel dressing (COG-Z@P200
+ NIR5) engineered to accelerate diabetic wound healing through
synergistic metabolic intervention and dynamic microenvironment
regulation. The hydrogel integrates dopamine-modified ZIF-8
nanoparticles with a double-crosslinked framework composed of
carboxymethyl chitosan (CMCS), gelatin, and oxidized sodium alginate
(OSA). CMCS disrupts bacterial membranes via Zn^2⁺ release and
synergizes with near-infrared (NIR)-triggered mild photothermal therapy
(40–45 °C), achieving dual bactericidal action through physical
puncture and metabolic interference. Gelatin enhances biocompatibility
and mechanical adhesion, while OSA reinforces structural stability. The
hydrogel’s therapeutic efficacy arises from three interconnected
mechanisms: NIR-activated hyperthermia disrupts MRSA energy metabolism
by suppressing glycolysis (eno, gap) and quorum sensing (agr),
immunomodulatory polarization of macrophages toward M2 phenotypes
(Arg-1⁺/TNF-α↓) promotes angiogenesis (VEGF/CD31↑) and collagen
alignment, and pH-responsive ROS scavenging with ECM remodeling
dynamically adapts to microenvironmental fluctuations. Validated in
diabetic murine models, COG-Z@P200 + NIR5 reduced healing time by 48%
and eradicated >99.5% MRSA, outperforming clinical benchmarks. This
work establishes an antibiotic-free paradigm for chronic wound
management, bridging nanomaterial-enabled bactericidal strategies with
host-directed metabolic-immune crosstalk, and offers a scalable
solution for multidrug-resistant infections in hyperglycemic niches.
Materials and methods
Materials and instruments
Zinc nitrate hexahydrate (Zn(NO[3])[2]·6H[2]O, 99.9%),
2-methylimidazole (2-MIm), carboxymethyl chitosan (CMCS, Shanghai
yuanye Bio-TechnologyCo., Ltd. carboxylation degree ≥80%, Mw = 650
kDa), Sodium alginate (SA, Aladdin™, assay ≥98%, viscosity 350–550
mPas, Mw = 100–150 kDa, M:G ratio = 1:1), pig skin-derived gelatin
(Gel, type A), dopamine (DA, purity ≥99%), streptozotocin (STZ, Beijing
Solebao), sodium hydroxide (NaOH), ethanol (C[2]H[5]OH),
β-galactosidase kit (β-galactosidase, Beyotime, Haimen, China),
2′,7′-dichlorodihydrofluorescein diacetate detection kit (DCFH-DA,
Beyotime, Haimen, China), enhanced ATP detection kit (Beyotime, Haimen,
China). All chemicals and reagents were employed as received unless
stated otherwise. X-ray diffraction (XRD, D8 Advance, Bruker, Germany).
Transmission electron microscope (TEM, JEOL JEM-F200, Japan). Scanning
Electron Microscope (SEM, ZEISS Sigma 300, Germany). X-ray
photoelectron spectroscopy (XPS, ESCALAB 250Xi, Thermo Scientific,
USA).
Synthesis of ZIF-8@PDA nanoparticles
Based on previous studies, we synthesized ZIF-8 nanoparticles with a
particle size of around 533 nm through an enhanced solvothermal
technique [[70]30]. Subsequently, 20 mg of ZIF-8 particles were
suspended in 100 ml of 10 mM Tris–HCl solution at pH 8.5. Subsequently,
0.2 g of dopamine hydrochloride was included and agitated for 10 min
until fully dissolved. The pH was subsequently adjusted to 8.5, and the
mixture was agitated for 6 h at ambient temperature in darkness.
Subsequently, the mixture underwent centrifugation and was rinsed three
times with deionized water to eliminate any unpolymerized dopamine. The
black ZIF-8@PDA particles were produced ultimately. The comprehensive
preparation protocols are outlined in the Supporting Information.
Synthesis and characterization of COG-Z@P hydrogel
Various concentrations of ZIF-8@PDA nanoparticles (50, 100, 150 and 200
µg//mL) were dissolved in CMCS and thoroughly mixed. Subsequently, 10%
gel (w/v) and the pre-prepared OSA solution (15%, w/v) were included.
The solution was stirred vigorously under vortex conditions until
gelation occurred, OSA is chemically double-cross-linked with Gel and
CMCS via a dynamic Schiff base reaction to generate COG-Z@P hydrogel
(COG hydrogel does not contain ZIF-8@PDA NPs).
Rheological assessments were performed with a rheometer (MCR 302) to
evaluate the mechanical characteristics of hydrogels. The hydrogel was
made in circles with a diameter of 1.5 cm and a thickness of 1–2 mm,
and it was examined at 37 °C using a time-oscillating scan at 1 Hz and
1% strain. The variations in storage modulus (G′) and loss modulus (G″)
were quantified, respectively. The self-repairing capabilities of
hydrogels were evaluated by the oscillatory strain scanning technique.
The test cycle comprised a modest strain (γ = 1%) at a constant
frequency of 1 rad/s for 100 s, thereafter transitioning to a large
strain (γ = 400%) for an additional 100 s. This cycle was reiterated
2.5 times. The macroscopic self-healing characteristics of hydrogels
were assessed utilizing two hydrogel discs: a brownish-black disc
(Z@P-COG200) and a light yellow disc (COG). The hydrogels were
bisected, and the semicircular segments of varying colors were aligned
along the incision. The hydrogels were incubated at 37 °C for 30 min,
following which they were removed. The technique was documented through
video and photography to capture the self-healing characteristics of
the hydrogel.
In vitro photothermal and antibacterial properties
1 mL of ZIF-8@PDA nanoparticles at varying concentrations (50, 100, 150
and 200 µg/mL) was distributed in either ultrapure water or COG
hydrogel. The experiment was performed in a 24-well plate. Thereafter,
the samples were exposed to a near-infrared laser (808 nm, 1 W/cm^2)
for 10 min, utilizing ultrapure water or COG hydrogel as controls.
Temperature variations were documented via an infrared thermal imaging
camera. The photothermal stability of the nanoparticles was assessed by
comparing the temperature profiles prior to and following near-infrared
irradiation, in addition to after four heating and cooling cycles.
Gram-negative E. coli and drug-resistant MRSA bacteria were tested in
vitro to assess the material’s antibacterial activity. The MARS
infected wound model was employed to evaluate the in vivo antibacterial
efficacy of the hydrogel. The comprehensive processes are presented in
the Supporting Information.
Transcriptomic analysis
Total RNA isolation, cDNA library construction, and sequencing
MRSA cultures were exposed to Z@P-COG200 + NIR for 5 min, with
PBS-treated bacterial suspensions serving as negative controls.
Bacterial pellets were harvested via centrifugation at 6,000×g (4 °C,
5 min) and subjected to three sequential PBS washes. Three biological
replicates per group (Control: CTRL1, CTRL2, CTRL3; Experimental:
Sample1, Sample2, Sample3) were processed under standardized
conditions. Total RNA was extracted using TRIzol® reagent, with RNA
integrity confirmed by Bioanalyzer (RIN > 8.0). Strand-specific cDNA
libraries were constructed following Illumina® TruSeq™ Stranded Total
RNA Library Prep Kit protocols (Majorbio Bio-pharm Technology,
Shanghai, China), including ribosomal RNA depletion via Ribo-Zero™ Gold
rRNA Removal Kit. Libraries were quantified by Qubit® 4.0 Fluorometer
and size-validated by Agilent 2100 Bioanalyzer. Paired-end sequencing
(2 × 150 bp) was performed on an Illumina NovaSeq 6000 platform
(Majorbio Bio-pharm Technology) with a minimum depth of 20 million
reads per sample.
Differentially expressed genes (DEGs) screening and bioinformatics analysis
Following normalization and variance stabilization of raw count data
using DESeq2 (v1.24.0), differentially expressed genes (DEGs) were
identified through a rigorous statistical framework incorporating
Benjamini–Hochberg multiple testing correction. Genes meeting the
threshold of absolute log2 fold change (|log2 FC|) ≥1 and adjusted
p-value (padj) <0.05 were classified as significant DEGs. Functional
annotation was performed through hierarchical orthologous group
classification via the Clusters of Orthologous Genes (COG) database
(v5.0) and pathway enrichment analysis using the Kyoto Encyclopedia of
Genes and Genomes (KEGG) database (Release 107.1). Enrichment
significance was assessed via hypergeometric testing with false
discovery rate (FDR) correction (q-value <0.05), as implemented in the
clusterProfiler R package (v4.6.2).
Metabolomic analysis
Metabolite extraction and LC‒MS-based untargeted metabolomics analysis
Metabolomic sample preparation followed the protocol in Transcriptomic
analysis, with six biological replicates per group (Control: CTRL1,
CTRL2, CTRL3, CTRL4, CTRL5, CTRL6; Experimental: Sample1, Sample2,
Sample3, Sample4, Sample5, Sample6). Metabolites were extracted using
400 μL of 80% methanol containing 0.02 mg/mL L-2-chlorophenylalanine
(internal standard). Tissues were homogenized in a cryogenic grinder
(−10 °C, 50 Hz, 6 min), sonicated (40 kHz, 30 min) at 5 °C, incubated
at −20 °C for 30 min, and centrifuged (13,000×g, 15 min, 4 °C). The
supernatant was analyzed via LC–MS (Thermo Scientific Vanquish Horizon
UHPLC-Q-Exactive system) by Majorbio Bio-pharm Technology (Shanghai,
China).
Differentially expressed metabolites (DEMs) screening and bioinformatics
analysis
Raw metabolomics data were analyzed using Progenesis QI (Waters
Corporation, USA). Metabolite identification was conducted via HMDB and
MetLin databases, followed by pathway annotation using KEGG
([71]http://www.genome.jp/kegg/). Differential metabolites (DEMs) were
screened with the ropls R package (v1.6.2), applying thresholds of p <
0.05 and variable importance in projection (VIP) >1.
Biocompatibility of hydrogels
Cytocompatibility study
The substance underwent sterilization, and the extract was procured.
Cells were inoculated at a density of 2.0 × 10^4 cells/cm^2 and
incubated under regulated conditions. To evaluate the proliferation of
L929 cells co-cultured with different hydrogels after 1, 2 and 3 days,
10% MTT in serum-free medium was introduced to each well, followed by a
4-h incubation. Subsequently, DMSO was added, and the plate was
agitated for 10 min. The absorbance at 490 nm was then quantified using
a multifunctional microplate reader.All trials were conducted in
quintuplicate, and the mean value was utilized as the final result.
Cell cytotoxicity was evaluated using a Calcein-AM/PI dual staining
kit. A Leica MD IL inverted fluorescent microscope was used to observe
live cells at 495 nm/515 nm (excitation/emission) and dead cells at 495
nm/635 nm.
Blood compatibility study
A hemolytic activity assay was conducted to evaluate the blood
compatibility of the hydrogel. C57/BL6 diabetic mice were anesthetized,
and blood was obtained from the abdomen vein using a blood collection
tube. A blood dilution was created by combining 4 mL of mouse blood
with 5 mL of PBS. Subsequently, 500 μL of various hydrogels and 500 μL
of the blood dilution were combined in a 1.5 mL centrifuge tube and
incubated at 100 rpm in a stirrer at 37 °C for 1 h. Following
incubation, the tube was centrifuged at 1,000 rpm for 10 min, and 100
μL of the supernatant was aliquoted into a 96-well plate. The
absorbance of the solution at 540 nm was assessed utilizing a
multifunctional enzyme marker. A positive control comprised 500 μL of
water, whereas a negative control comprised 500 μL of PBS buffer. The
hemolysis rate was ultimately determined using Eq. ([72]1):
[MATH: Hemolysisratio=(Ae-Ap/Aw-Ap)×100% :MATH]
1
Ae—represents absorbance of the experimental group; Ap—denotes
absorbance of the negative control group; Aw—signifies absorbance of
the positive control group.
Construction of whole skin wound model in diabetic mice infected with MRSA
All animal experiments were conducted in strict accordance with the
National Regulation of China for the Care and Use of Laboratory
Animals, as well as the National Research Council Guidelines for the
Care and Use of Laboratory Animals. The study was approved by the
Ethics Committee for Medical and Experimental Animals at Northwestern
Polytechnical University (Approval No. 202202091). C57/BL6 male mice
(SPF, male, 6–8 weeks old) were acquired from the Animal Experiment
Center at Xi’an Jiaotong University. The mice were given D12492
high-sugar, high-fat standard food and plenty water for one week. The
fasting blood glucose levels of mice were measured after a 12-h fast.
Thereafter, the mice were weighed and administered an intraperitoneal
injection of a prepared streptozotocin (STZ, Solarbio) solution at a
dosage of 55 mg/kg for five consecutive days. One week later, the mice
underwent fasting blood glucose testing via blood collection from the
tail vein. Blood glucose levels surpassing 16.7 mmol/L, along with
noted weight loss and polyuria, confirmed the effective establishment
of the diabetic mouse model.
A 0.3% sodium pentobarbital solution (0.1–0.2 mL/10 g) was administered
to anesthetize the mice. A 6 mm diameter punch was employed to generate
a circular wound on the backs of mice, then followed by the application
of 20 μL of Staphylococcus aureus (1 × 10⁸ CFU/mL) drops onto the wound
surface to construct a full-thickness skin injury infection model. Mice
were randomly allocated into five groups: Control group, COG group, COG
+ NIR5 group, COG-Z@P200 group, and COG-Z@P200 + NIR5 group.
To evaluate wound tissue regeneration (epidermal thickness, new
granulation tissue, and collagen deposition), mice were euthanized on
days 3, 7 and 14, and skin tissues were harvested. The collected
tissues were fully submerged in 4% paraformaldehyde (G1101, Servicebio)
for fixation, then embedded in paraffin, sectioned, and stained using
hematoxylin–eosin (H&E, GP1031, Servicebio) and Masson’s trichrome
staining kit (G1006, Servicebio). Images were obtained using a tissue
biopsy scanner (Science, WINMEDIC) and analyzed with ImageJ software.
Statistical analysis
In this experiment, each group contained a minimum of three independent
replicates. The data results were presented as mean ± standard
deviation (SD). The experimental data were subjected to statistical
analysis by One-way ANOVA to evaluate significant differences among
groups. A *p < 0.05 denotes statistical significance, **p < 0.01
signifies a high level of statistical significance, ***p < 0.001
represents a very high level of statistical significance, and
****p < 0.0001 shows an extremely high level of statistical
significance.
Results and discussion
Synthesis and characterization of ZIF-8 NPs and ZIF-8@PDA NPs
ZIF-8 nanoparticles were produced according to established protocols
reported in the literature [[73]31, [74]32]. Subsequently, DA was
incorporated, resulting in the formation of ZIF-8@PDA nanoparticles
(Fig. [75]1A). A SEM image demonstrated that ZIF-8 NPs exhibited a
normal dodecahedron shape (Fig. [76]1A-a1), but ZIF-8@PDA NPs, coated
with polydopamine, presented an irregular spherical form
(Fig. [77]1A-a2). Based on TEM morphology, ZIF-8 (Fig. [78]1B-b1)
exhibited a uniform coloration, but ZIF-8@PDA NPs (Fig. [79]1B-b2)
displayed a lighter hue on the surface compared to the interior. The
transformation of nanoparticles from white to black, before and after
polydopamine coating, was observable to the naked eye. The EDS spectra
of ZIF-8@PDA NPs (Fig. [80]1C) indicated an elevation in carbon and
nitrogen components, potentially attributable to the PDA coating.
1 mg/mL ZIF-8@PDA nanoparticles in ddH[2]O were analyzed for Zeta
potential and hydrodynamic particle size using a Nano-ZS dynamic light
scattering instrument (Fig. [81]1D, E). The zeta potential of ZIF-8@PDA
NPs (−45 mV) was significantly lower than that of ZIF-8 NPs (24.7 mV),
indicating enhanced colloidal stability and improved dispersion of
ZIF-8@PDA NPs. The average particle size of ZIF-8 NPs and ZIF-8@PDA NPs
was found to be reasonably consistent in an aqueous environment, with
ZIF-8 exhibiting a positive charge and ZIF-8@PDA NPs displaying a
negative charge attributed to the hydroxyl groups of DA (Fig. [82]1F).
Fig. 1.
[83]Fig. 1
[84]Open in a new tab
Characterization of ZIF-8 NPs and ZIF-8@PDA NPs. A–B SEM and TEM. Scale
= 200 nm. C Mapping of element. Scale = 50 nm. D–E Particle size
distribution. F Zeta potentials. G FT-IR spectrum. H–K XRD and XPS
analysis
In comparison to ZIF-8 NPs, the FTIR spectra of ZIF-8@PDA NPs exhibited
a peak at 1500 cm^−1, indicative of the stretching vibration of the
carbon–carbon double bond (C=C group), primarily derived from the
benzene ring structure of dopamine (Fig. [85]1G). The characteristic
XRD peaks of ZIF-8 are seen in the XRD patterns of NPs (Fig. [86]1H) at
2θ angles of 7.20°, 12.60°, and 17.90°. These angles correspond to the
(110), (211), and (222) crystallographic planes [[87]33]. The crystal
morphology of ZIF-8 was confirmed, and the peak position exhibited
minimal alteration before and after dopamine coating. The chemical
composition and electrical structure of ZIF-8 nanoparticles exhibited
significant alterations for dopamine encapsulation, as demonstrated by
XPS (Fig. [88]1I). ZIF-8@PDA NPs comprised zinc (Zn), oxygen (O),
carbon (C), and nitrogen (N). The zinc ions exhibited two peaks
(Fig. [89]1J) at 1022 and 1045 eV, corresponding to the 2p[3/2] and
2p[1/2] orbitals, respectively. Simultaneously, the C1 s peak
bifurcated into two components at 284.8 and 291.2 eV, associated with
the C–C and C–N bonds (Fig. [90]1K). The characteristic N1 s and Zn 2p
peaks remained following dopamine encapsulation, indicating that
ZIF-8@PDA nanoparticles have a stable framework structure. The
characteristic N1 s and Zn 2p peaks remained following dopamine
encapsulation, indicating that ZIF-8@PDA nanoparticles have a stable
framework structure. Previous findings indicated that the N1 s XPS
spectrum exhibited two peaks at 400.98 and 399.38 eV (Figure S1 A),
corresponding to C=N and C–N in 2-methylimidazole. Additionally, a
novel signal associated with C=O at 531.73 eV (Figure S1B) suggests the
potential presence of polydopamine on the surface of ZIF-8.
Preparation and structural characterization of COG-Z@P hydrogels
An environmentally sustainable one-pot method was employed to
incorporate varying concentrations of Z@P nanoparticles into COG,
resulting in hydrogels designated as COG-Z@P50, COG-Z@P100, COG-Z@P150
and COG-Z@P200. The COG hydrogel was synthesized by doubly
cross-linking the aldehyde group of oxidized sodium alginate (OSA) with
the amino groups of CMCS and gelatin via a Schiff base reaction
(Fig. [91]2A). OSA was synthesized through the oxidation of sodium
alginate (SA), as described in previous studies [[92]34]. The FTIR
spectrum of OSA exhibited a unique absorption band at 1720 cm⁻^1,
corresponding to the stretching vibration of the (−CHO) C=O bond in the
SA aldehyde group (Figure S2 A). The oxidation degree of OSA was found
to be 27.28% (Figure S2B), determined via the hydroxylamine
hydrochloride reaction, which facilitated the subsequent Schiff base
reaction.
Fig. 2.
[93]Fig. 2
[94]Open in a new tab
Preparation and characterization of COG-Z@P200 hydrogel. A–B Schematic
diagram of hydrogel synthesis; C SEM image; D Frequency scan; E Dynamic
strain scan test; F Cyclic scan test under shear strains of 1 and 400%
(ω = 1 rad/s); G Self-healing performance; H Demonstration of
injectability of hydrogel; I Adhesion and tensile properties; J–K
Thermal infrared image of COG-Z@P hydrogel under NIR radiation. L
Temperature change curves of different concentrations of ZIF-8@PDA NPs
and COG-Z@P hydrogels with laser irradiation time. M Temperature
changes of COG-Z@P200 hydrogel during four cycles of on/off 1.0 W
cm^−2, 808 nm laser irradiation
ZIF-8@PDA nanoparticles were incorporated into CMC to produce a fluid
black suspension, as shown in Fig. [95]2B. This suspension was then
combined with the OSA solution and rapidly mixed. The COG-Z@P200
hydrogel was synthesized in less than 10 s. For comparison, the COG
hydrogel was synthesized without the addition of ZIF-8@PDA
nanoparticles. Both hydrogels exhibited a homogeneous, porous, and
interconnected network structure, with ZIF-8@PDA nanoparticles
uniformly distributed (Fig. [96]2C, red arrow). The self-healing
properties of the hydrogels were evaluated by bisecting a black
COG-Z@P200 hydrogel disc and a transparent COG hydrogel. The two
resulting semicircular hydrogel pieces were placed in contact along the
cutting line to assess the reformation of the gel. As shown in
Fig. [97]2D, within 30 min, the two halves autonomously reassembled
into a complete hydrogel with distinct colors. The reformed hydrogel
demonstrated excellent self-healing capabilities, maintaining
structural integrity when stretched with forceps perpendicular to the
incision line.
Mechanical properties and self-healing properties
Three distinct hydrogel samples were subjected to dynamic frequency
scanning experiments, resulting in the storage modulus (G′) and loss
modulus (G″), along with the rheological test results, as presented in
Fig. [98]2E. All hydrogels exhibited G′ values exceeding G″ throughout
the entire testing range, demonstrating robust gel strength and a
stable gel state. The G′ and G″ values of the COG hydrogel increased
with frequency, indicating physical entanglement within the network of
reversible dynamic chemical cross-linking. However, the G′ and G″
values of the hydrogel samples COG-Z@P100 and COG-Z@P200 remained
largely unchanged. This resulted mainly from the stable network
structure and elevated cross-linking density generated by the higher
concentration of ZIF-8@PDA NPs. The COG hydrogel exhibited a storage
modulus greater than its loss modulus throughout the entire testing
range, suggesting that the hydrogel possesses significant gel strength
and maintains a stable gel state. As illustrated in Figures S3 A-B and
[99]2F, the storage modulus (elastic modulus) of COG, COG-Z@P100 and
COG-Z@P200 hydrogels was 808.21, 1040.3 and 1748.3 Pa, respectively, at
a frequency of 1 rad/s. This indicates that an increase in the content
of ZIF-8@PDA nanoparticles correlates with a rise in cross-linking
density and the elastic modulus of the hydrogel. The G″ values of the
three hydrogels converge at a strain of 385%, indicating a transitional
state between solid-like and liquid-like behavior. Complete hydrogel
degradation occurred at a critical strain exceeding 385%. To evaluate
the hydrogel’s self-healing capability, sequential step-strain testing
was performed (Figures S3 C-D and [100]2G). Upon an initial strain of
400%, G′ decreased significantly from 2102.6 to 339.24 Pa, indicating
the disruption of the hydrogel network. However, when the strain was
reduced to 1%, G′ recovered to 2081.02 Pa, suggesting that the
cross-linked network had largely reformed. After five cycles of
alternating high and low strain, the G′ and G″ values of the repaired
hydrogel closely resembled those in the initial cycle, confirming its
robust self-healing properties.
The excellent injectability of the COG-Z@P200 hydrogel makes it
particularly suitable for complex wound healing applications, including
those involving irregularly shaped wounds. Using a 27-gauge needle
(diameter: 0.41 mm), the hydrogel was smoothly injected to form the
inscription ‘NPU’ without clogging (Fig. [101]2H). Upon injection into
water, the hydrogel maintained its structural integrity, forming
continuous spiral thin lines, thereby demonstrating its outstanding
injectability and shape retention. This high injectability is
attributed to its optimized viscosity and shear-thinning behavior,
which are further enhanced by the presence of dynamic non-covalent
interactions within the hydrogel network. Furthermore, the hydrogel
exhibited remarkable self-healing capabilities due to the reversibility
of these interactions. As shown in Fig. [102]2I, the hydrogel adhered
firmly to human finger joints, maintaining stable attachment during
flexion and extension without cracking or peeling. This strong adhesion
and mechanical resilience under dynamic conditions underscore its
potential for clinical applications in wound healing.
Near-infrared triggered photothermal properties and antioxidant capacity
Photothermal therapy has shown considerable potential in biomedicine,
with polydopamine (PDA) capable of absorbing visible and near-infrared
(NIR) light and converting it into thermal energy [[103]35–[104]37]. To
evaluate the photothermal properties of ZIF-8@PDA nanoparticles (NPs)
and COG-Z@P hydrogels, we tested their temperature responses under 808
nm NIR irradiation. As shown in Fig. [105]2J–K, the temperature of the
system increased with rising concentrations of ZIF-8@PDA NPs after 10
min of irradiation. Notably, the COG-Z@P200 hydrogel exhibited a higher
temperature rise than the ZIF-8@PDA NPs alone, with ΔT values of 38.4
and 36.1 °C, respectively (Figures S4 A-B). These findings were further
confirmed by infrared thermal imaging (Fig. [106]2L), which showed a
consistent temperature increase.
The addition of ZIF-8@PDA NPs into the COG hydrogel effectively
prevented nanoparticle aggregation, which can lead to the loss of
photothermal efficiency. The photothermal conversion efficiency (η) was
calculated for both COG-Z@P200 and ZIF-8@PDA NPs at a concentration of
200 μg/mL under 808 nm laser irradiation, yielding values of 16.88 and
25.27%, respectively. These results indicate that the photothermal
performance of the COG-Z@P200 hydrogel and ZIF-8@PDA NPs are
comparable, confirming the efficacy of ZIF-8@PDA NPs as a photothermal
agent in hydrogel formulations. The following Eqs. ([107]2)–([108]4)
were used to determine the photothermal conversion efficiency:
[[109]38, [110]39]
[MATH: Q=cmΔT :MATH]
2
where
[MATH: Q :MATH]
,
[MATH: c :MATH]
,
[MATH: m :MATH]
,
[MATH: ΔT :MATH]
are the total heat, specific heat (H[2]O: 4200 J kg^−1·K^−1) and
gelatin: 1830 J kg^−1·K^−1), mass, and the change of temperature,
respectively. The total energy of the light could be calculated using
following equation:
[MATH: E=PSt
:MATH]
3
where
[MATH: P :MATH]
is the power density of the light,
[MATH: S :MATH]
and
[MATH: t :MATH]
are the irradiation area (
[MATH: S :MATH]
=2 cm^2) and irradiation time. The photothermal conversion efficiency
of COG-Z@P200 hydrogel was calculated as follows:
[MATH: η=Q/E=cmΔT/PSt :MATH]
4
The maximum temperature was similarly attained after near-infrared
irradiation, consistent with the first cycle, during repeated heating
and cooling experiments following four “ON/OFF” cycles, indicating
effective photothermal reversibility (Fig. [111]2M). After five minutes
of exposure to 808 nm radiation, the temperature of COG-Z@P200 attained
49.5 °C. The temperature was suitable for photothermal antibacterial
activity and may facilitate the rapid realization of photothermal
antibacterial capabilities.
The capacity of hydrogels to scavenge free radicals (DPPH) was utilized
to evaluate their antioxidant properties. The catechol group in
unoxidized SA resulted in a 32.55% reduction of DPPH by COG hydrogel
after 60 min of incubation, as shown in Figure S5. The free radical
scavenging rates of the COG-Z@P hydrogel increased to 51.88, 51.77,
57.19, and 56.60%, respectively, upon the incorporation of ZIF-8@PDA.
The catechol groups in COG-Z@P hydrogel enhance its antioxidant
capacity, with an increase in ZIF-8@PDA correlating to a greater
antioxidant capacity.
Antibacterial activity
This study explored the antibacterial effects of COG-Z@P200 hydrogel
under 1.0 W·cm^−2 NIR radiation at various conditions, including the
effect of using NIR light alone. Figure [112]3A shows the colony
distribution of E. coli and MRSA after 2 h of treatment under different
conditions, including the control group, NIR irradiation for 5 min, and
COG-Z@P200 hydrogel with or without NIR exposure. Statistical analysis
revealed that NIR irradiation alone showed very weak antibacterial
activity, but when combined with COG-Z@P200 hydrogel, the antibacterial
activity was significantly enhanced (Fig. [113]3B–C). Moreover, the
antibacterial effect continued to improve with prolonged NIR exposure.
After 5 min of NIR irradiation, the hydrogel demonstrated an
antibacterial efficacy exceeding 99%, with a measured temperature of
approximately 49.5 °C, which remained within the safe threshold for
PTT, thus preventing further harm to the body [[114]21]. Additionally,
the incorporation of ZIF-8@PDA nanoparticles into the COG hydrogel
significantly enhanced its antibacterial effect against MRSA and E.
coli, highlighting the crucial role of ZIF-8@PDA nanoparticles in
antibacterial performance. Further, bacterial morphology observed by
SEM showed that E. coli and MRSA exposed to NIR light alone had
relatively intact bacterial membranes with no obvious damage
(Fig. [115]3D). However, when COG-Z@P200 hydrogel was combined with
NIR, the bacterial membranes exhibited wrinkling and severe damage,
indicating that the photothermal effect of NIR plays a crucial role in
the antibacterial activity of the hydrogel. The extent of bacterial
damage increased with prolonged NIR exposure times, as evidenced by the
treatments COG-Z@P200 + NIR5 and COG-Z@P200 + NIR10. And with the
addition of ZIF-8@PDA and the synergistic effect of NIR, we can clearly
see the nanoparticles attached to the surface of bacteria.
Fig. 3.
[116]Fig. 3
[117]Open in a new tab
Antibacterial properties of COG-Z@P200 hydrogel against E. coil and
MARS for 2 h. A Colony growth of bacteria; B–C Survival rates. D SEM
images of bacteria; E TEM images of bacteria. F β-galactosidase levels;
G Protein leakage; H DNA and RNA leakage; I ATP levels. J Schematic
diagram of the antibacterial mechanism and *P < 0.05, ***P < 0.001,
****P < 0.0001
In vitro results led to the selection of MRSA for further exploration
of its antibacterial mechanisms.The Zn^2⁺ ion release curve in Figure
S6 was analyzed by kinetic model, revealing that the release process of
Zn^2⁺ was mainly controlled by diffusion (Higuchi model, R^2 > 0.93).
The study found that the cumulative release of Zn^2⁺ gradually
increased over time at different pH values. Under pH 5.4 and 6.8
conditions, NIR radiation significantly accelerated the release rate of
Zn^2⁺, indicating that the synergistic effect of pH environment and NIR
radiation has an important influence on the Zn^2⁺ release process. As
reported in the literature, ZnO nanoparticles are non-toxic to the
human body and have strong antibacterial activity. Photothermally
enhanced ZnO nanoparticles release Zn^2⁺, promote bacterial cell
membrane rupture and ROS generation to synergistically kill bacteria.
This may be attributed to the protonation competition between Zn^2⁺ and
carboxylic acid groups under low pH conditions, resulting in the
breaking of coordination bonds, thereby accelerating the release of
Zn^2⁺. In addition, under the action of NIR radiation, the local
temperature of the hydrogel increases, further reducing the activation
energy of Zn^2⁺ diffusion and accelerating the diffusion process of
Zn^2⁺. NIR radiation may also promote the release of Zn^2⁺ through the
plasma enhancement effect. Metal nanostructures can excite local plasma
resonance under NIR radiation, generate strong electric fields, enhance
the reactivity of nearby molecules, further promote the release of
Zn^2⁺, and significantly enhance the antibacterial properties of
hydrogels. However, repeated near-infrared irradiation may affect the
photothermal stability of COG-Z@P200 hydrogel. Thermal interactions may
lead to the aggregation of nanoparticles due to the aggregation of
nanoparticles or the oxidation effect of PDA, thereby reducing the
photothermal efficiency, while the oxidation of PDA may gradually
affect the structural integrity and photothermal performance of the
hydrogel. The PDA coating can effectively stabilize ZIF-8 nanoparticles
and mitigate the negative effects of nanoparticle aggregation and
oxidation through its self-healing properties [[118]28]. Nevertheless,
these effects are mild and usually occur during longer periods of use,
and do not significantly affect the short-term stability of the
hydrogel. The release of Zn^2⁺ is highly sensitive to pH, which helps
to effectively exert antibacterial effects without compromising
short-term structural stability. Similarly, the oxidation of PDA may
slightly affect the mechanical strength during long-term use, but its
self-healing ability can effectively offset this effect. Overall, the
COG-Z@P200 hydrogel can maintain sufficient integrity during typical
wound treatment processes to ensure its therapeutic effect and
biocompatibility.
The TEM results, shown in Fig. [119]3E, revealed that the COG-Z@P200
+ NIR5 treatment caused significant structural damage to the bacteria,
including wrinkling of the cell membrane and leakage of cellular
contents. These findings were consistent with those of other studies.
The permeability of the bacterial cell membrane was further assessed
using β-galactosidase, which exhibits fluorescence when the cell
membrane is compromised [[120]23]. Both the COG-Z@P200 + NIR5 and
COG-Z@P200 + NIR10 treatments showed high fluorescence intensity at
OD420, as demonstrated in Fig. [121]3F, indicating increased membrane
permeability. Additionally, Fig. [122]3G shows a significant increase
in protein leakage in MRSA treated with COG-Z@P200, while Fig. [123]3H
shows a marked rise in ATP levels with prolonged NIR exposure.
Moreover, the COG-Z@P200 + NIR5 treatment group exhibited the highest
levels of leakage of cellular components, including DNA and RNA
(Fig. [124]3I), suggesting a strong antibacterial effect. These results
indicated that COG-Z@P200 combined with NIR treatment led to membrane
disruption, elevated intracellular ATP levels, and ultimately,
bacterial death. The effect of NIR treatment on bacterial energy
consumption and intracellular ATP levels was also examined, given the
strong correlation between bacterial metabolism and intracellular
temperature [[125]40, [126]41]. Figure [127]3J summarizes the proposed
antibacterial mechanism of COG-Z@P hydrogel against bacteria.
Transcriptome sequence and metabolomics analysis
Transcriptome and metabolomic analysis were carried out to investigate
the antibacterial mechanism of COG-Z@P200 + NIR5 on MRSA isolates in
more detail. The sample group was designated as COG-Z@P200 + NIR5 for
photothermal therapy, whereas the CTRL group was designated as
untreated MRSA. Each sample’s uniform overall gene expression levels
were displayed in a boxplot (Fig. [128]4A), and Table S1 reported that
all samples had good biological reproducibility, with all Pearson
correlation coefficients better than 0.98. The findings of the
transcriptome comparison show which genes are differentially expressed
(DEGs) in the various groups. A total of 278 down-regulated and 192
up-regulated genes were identified within the 470 differentially
expressed gene sets (FC > 2, FDR < 0.05) (Fig. [129]5B–C).
Fig. 4.
[130]Fig. 4
[131]Open in a new tab
Transcriptome of MRSA after different treatments with hydrogel. A Box
plot of the expression number of DEGs; B Volcano plot showing the
identified up- and down-regulated genes. C Statistical histogram of
DEGs. D GO enrichment analysis of DEGs. E KEGG pathway enrichment
analysis of DEGs. F Heatmap and cluster analysis of glycolysis and
TCA-related genes. G Heatmap and cluster analysis of arginine-related
genes. H Heatmap and cluster analysis of quorum sensing and virulence
factor-related genes
Fig. 5.
[132]Fig. 5
[133]Open in a new tab
Changes in MRSA metabolomics after different treatments. A PCA analysis
of samples; B Column plot of enriched pathways. C Metabolite heat map
cluster analysis. D KEGG pathway enrichment analysis. E Antimicrobial
mechanism diagram of COG-Z@P200 hydrogel
Gene Ontology (GO) analysis and pathway enrichment from the Kyoto
Encyclopedia of Genes and Genomes (KEGG) were used to clarify the
connection between differentially expressed genes (DEGs) and
antibacterial action. The DEGs were functionally annotated using GO
enrichment analysis, which categorized them into groups such molecular
functions (MF), cellular components (CC), and biological processes (BP)
(Fig. [134]4D). The majority of the enriched genes were linked to
biological processes that included glycolysis, arginine catabolism, ATP
biosynthesis, translation, ion transmembrane transport, macromolecule
biosynthesis, and catabolic processes. Enriched genes were
predominantly associated with membrane protein complexes, ribosomes,
and proton-transporting ATP synthase complexes under the cellular
component category. In terms of molecular function, the DEGs were
mostly engaged in the transmembrane transporter activity of
glucose-6-phosphate, the transmembrane transporter activity of
ATPase-coupled ions, the activity of ATP-dependent peptidase, and a
variety of other transmembrane transport activities, including those
involving carbohydrates. Important pathways such ABC transporters,
glycolysis/gluconeogenesis, the tricarboxylic acid (TCA) cycle,
two-component systems, and arginine biosynthesis were identified by
KEGG enrichment analysis (Fig. [135]4E). The metabolism of galactose,
the phosphotransferase system (PTS), the production of valine, leucine,
and isoleucine, Staphylococcus aureus infection, quorum sensing, and
the metabolism of starch and sucrose were among the other noteworthy
processes. These results imply that Zn^2+ release caused by COG-Z@P200
+ NIR5 therapy, which probably interferes with protein production and
jeopardizes membrane integrity in MRSA, may be responsible for the
antibacterial effects seen [[136]42]. These pathways shed light on the
underlying processes of the COG-Z@P200 hydrogel’s photothermal
antibacterial activity.
The three main categories of antibacterial processes associated with
DEGs were glycolysis, arginine biosynthesis, and quorum
sensing/virulence factors. As demonstrated in Fig. [137]4F, the
reduction in ATP generation caused by the downregulation of
glycolysis-related genes (eno, gap) has an impact on MRSA’s energy
supply [[138]43, [139]44]. The TCA cycle-related genes sucC, sdhA and
icd were inhibited, which reduced bacterial metabolism and oxidative
stress tolerance [[140]45, [141]46]. Downregulation of arcA, arcC and
arcD interfered with arginine biosynthesis, which is necessary for the
development of biofilms and reduced MRSA’s pathogenic potential
(Fig. [142]4G) [[143]47, [144]48]. The downregulation of sec, lac, opp
and sdrD genes resulted in lower virulence and biofilm stability, as
demonstrated in Fig. [145]4H, indicating that quorum sensing, which
controls virulence factors, was also impacted [[146]49]. The response
of MRSA to environmental stress was shown by the upregulation of
stress-related genes (yidD, nfsA and kdpA) [[147]50]. According to
these results, treatment with COG-Z@P200 + NIR5 suppresses MRSA
development by interfering with energy metabolism, biofilm formation,
and expression of virulence.
PCA analysis (Fig. [148]5A) showed clear differences between the CTRL
and Sample groups, with strong sample correlation (Figure S7),
according to metabolic analysis. Glycolysis/gluconeogenesis, pyruvate
metabolism, TCA cycle, arginine and proline metabolism, ABC transport,
galactose metabolism, the two-component system, PTS, and quorum sensing
were identified as important metabolic pathways by KEGG pathway
classification analysis (Fig. [149]5B, D). The heatmap clustering
results revealing that COG-Z@P200 hydrogel exhibits antibacterial
action against MRSA are compatible with the downregulation of
metabolites such as N[2]-acetyl-L-ornithine, norleucine, aspartate,
methionine, and succinat (Fig. [150]5C). These results suggest a
possible antimicrobial mechanism of COG-Z@P200 hydrogel against MRSA
isolates (Fig. [151]5E).
Biocompatibility and antioxidant capacity
In order to systematically evaluate the cell compatibility of the
hydrogel, we cultured L929 fibroblasts in the presence of different
concentrations of ZIF-8 and ZIF-8@PDA nanoparticles (Fig. [152]6A–B).
The results showed that ZIF-8@PDA NPs had no obvious toxicity to cells
in the concentration range of 200 μg/mL and below, and the cell
activity was maintained at a high level during the three-day culture
process, which was significantly better than the unmodified ZIF-8 NPs,
which had already shown certain cytotoxicity at the same concentration.
This shows that dopamine modification can effectively improve the
biocompatibility of ZIF-8 nanoparticles.
Fig. 6.
[153]Fig. 6
[154]Open in a new tab
Biocompatibility of ZIF-8 NPs, ZIF-8@PDA NPs and COG-Z@P composite
hydrogels. A–C MTT of ZIF-8, ZIF-8@PDA and hydrogels co-cultured with
L929 cells for 1, 2 and 3 days; D Pictures of live/dead staining of
hydrogel co-cultured with L929 cells after 3 days; E Pictures of
hemolytic activity test of hydrogel; F Hemolysis rate of hydrogel; G
Pictures of hydrogel hemostatic effect; H Bleeding quantitative
statistics. n = 3, **p < 0.01, ****P < 0.0001
In addition, the MTT assay (Fig. [155]6C) and live/dead staining
results (Fig. [156]6D) of the hydrogel system as a whole further
verified its excellent cell compatibility. The cell viability of all
groups was close to that of the 10% FBS control group, and there was
almost no obvious cell death (green is live cells, red is dead cells).
Further observing the effect of different concentrations on therapeutic
performance, we combined the thermal response experimental data in
Fig. [157]2K–M and found that a ZIF-8@PDA concentration of 200 μg/mL
has an ideal photothermal conversion efficiency under near-infrared
laser irradiation, can quickly raise the temperature to a therapeutic
window of 40–45 °C, and maintain thermal stability in multiple rounds
of illumination cycles without aggregation or degradation. Therefore,
this concentration not only ensures cell safety, but also meets the
functional requirements of photothermal therapy. Based on the above
results, 200 μg/mL was finally selected as the loading concentration of
ZIF-8@PDA NPs to ensure that COG-Z@P200 hydrogel has both good
therapeutic performance and biosafety, and has the potential for
clinical transformation as a wound dressing.
Blood compatibility is a crucial property for biomedical hydrogels. In
the hemolysis assay, PBS and 0.1% Triton X-100 were used as the
negative and positive controls, respectively. Figure [158]6E shows that
neither the COG-Z@P200 hydrogel group nor the negative control group
exhibited significant hemolysis after one hour of incubation with
diluted whole blood (the supernatant remained clear). In contrast, the
positive control group, containing 0.1% Triton, exhibited a distinct
red supernatant, indicating red blood cell rupture and hemoglobin
release. The hemolysis rates of all hydrogel groups were well below the
safe threshold of 5.0%, as determined by absorbance measurement at 540
nm, confirming the excellent blood compatibility of the hydrogels
(Fig. [159]6F).
In the diabetic mouse bleeding model, bleeding was induced by
puncturing the liver with a needle, and hemostasis was achieved by
applying a prefabricated COG-Z@P200 hydrogel patch. Due to the
hydrogel’s ability to seal the wound and stop the bleeding, it
significantly reduced liver hemorrhage (Fig. [160]6G). As shown in
Fig. [161]6H, blood loss in the COG-Z@P200 hydrogel group was 111.02
mg, while the untreated control group lost 246.47 mg of blood,
indicating the hydrogel’s effective hemostatic properties.
The ROS-scavenging capability of the hydrogel was assessed by
co-culturing it with L929 fibroblasts treated with the ROS inducer
Rosup. In addition to stimulating excessive ROS production, Rosup
enables the quantification of intracellular ROS levels using the
DCFH-DA fluorescent probe. As shown in Figure S8, the positive control
group (treated with Rosup alone) exhibited a significant increase in
fluorescence intensity, indicating elevated ROS levels. In contrast,
treatment with 5 mg/mL COG-Z@P200 hydrogel markedly reduced
fluorescence intensity, suggesting effective ROS scavenging. These
findings demonstrate that COG-Z@P200 hydrogel can efficiently mitigate
oxidative stress, highlighting its potential as a wound dressing for
managing ROS-related tissue damage.
Full thickness MRSA infected skin wound healing
To establish a subcutaneous infection model, full-thickness skin
defects were created on the dorsal regions of mice, and wound healing
was monitored following hydrogel treatment at days 0, 3, 7 and 14
(Fig. [162]7A). No adverse effects were observed in any animal group
throughout the procedure. Representative wound images at different time
points (0, 3, 7 and 14 days) are shown in Fig. [163]7B–C, with the
COG-Z@P200 + NIR5 group exhibiting the smallest wound area.
Fig. 7.
[164]Fig. 7
[165]Open in a new tab
COG-Z@P200 hydrogel promotes wound healing. A Establishment of
MASA-infected diabetic wound model; B Pictures of wound healing; C
Wound healing simulation diagram; D H&E staining images; E Wound
healing rate; F Dermal gap; G Masson staining images. *P < 0.05,
***P < 0.001, ****P < 0.0001
Histological analysis via H&E staining (Fig. [166]7D) revealed
inflammatory cell infiltration in all groups by day 3, marking the
inflammatory phase. By day 7, new epidermis formation was evident in
the COG-Z@P200 and COG-Z@P200 + NIR5 groups, with the latter exhibiting
more pronounced healing. In the COG, COG + NIR5, and COG-Z@P200 groups,
scab formation was observed, accompanied by epithelial and inflammatory
cell migration toward the wound center. However, in the control group,
neither epidermal regeneration nor significant healing was observed.
By day 14, the COG-Z@P200 group had entered the remodeling phase,
exhibiting a thicker epidermal layer and a narrower dermal gap compared
to the control and NIR groups. In the COG-Z@P200 + NIR5 group, tissue
remodeling was nearly complete, with the dermal structure closely
resembling that of normal skin and the formation of numerous hair
follicles, indicating advanced healing. High-magnification images of
tissue sections (Fig. [167]7D) revealed the emergence of new hair
follicles in the dermis of the COG-Z@P200 group, whereas the control
and COG groups displayed a thicker epithelial layer with delayed dermal
remodeling. Notably, in the COG-Z@P200 + NIR5 group, the dermis
underwent more rapid maturation, characterized by the presence of
abundant newly formed hair follicles (red arrows) and a thinner, more
structurally normal epithelial layer. These findings demonstrate that
both the COG-Z@P200 and COG-Z@P200 + NIR5 groups significantly
outperformed the control and NIR groups in promoting wound healing. The
superior wound healing observed in the COG-Z@P200 + NIR group is likely
attributed to its photothermal antibacterial effects, which facilitated
epithelialization and accelerated dermal tissue maturation.
The wound healing rate (Fig. [168]7E) indicated that the COG-Z@P200
+ NIR5 group exhibited significantly accelerated wound closure compared
to other groups at days 3, 7 and 14. By day 14, the epidermal thickness
in the control, COG, COG + NIR5, and COG-Z@P200 groups remained
markedly different from that of normal skin tissue, whereas the
epidermal thickness in the COG-Z@P200 + NIR5 group closely resembled
that of normal skin (Figure S9). Moreover, the COG-Z@P200 + NIR5 group
displayed the smallest dermal gap (135.2 μm) among all groups,
approximating the normal skin tissue gap (Fig. [169]7F), suggesting
that COG-Z@P200 + NIR5 significantly promoted dermal tissue repair. The
Masson staining results of the wound sites in each group are shown in
Fig. [170]7G. On day 3, only a small amount of collagen deposition was
observed in localized regions of the wound areas in all groups.
Notably, the collagen content in the COG-Z@P200 + NIR5 group was
significantly higher than in the other groups, indicating that this
group exhibited stronger collagen synthesis ability during the early
stages of wound healing. By day 7, the number of fibroblasts had
generally increased in all groups, but significant differences were
observed in the distribution pattern of collagen fibers. In the Control
and COG groups, collagen fibers were mainly concentrated on the sides
of the wound, while the collagen deposition in the area beneath the
wound was sparse. In contrast, the COG-Z@P200 and COG-Z@P200 + NIR5
groups exhibited more prominent collagen deposition in the granulation
tissue, with a more uniform distribution. Among these, the COG-Z@P200
+ NIR5 group showed the most intense collagen staining, suggesting its
superior collagen synthesis capacity.
Histological analysis of internal organs using H&E staining (Figure
S10) revealed severe organ damage in the control group, including
cardiac rupture (blue arrow), hepatic sinusoidal dilation (black
arrow), indistinct red and white pulp junctions in the spleen (white
circles), inflammatory infiltration in the lungs (yellow circles),
renal tubular dilation (blue arrows), and vacuolar degeneration. In
contrast, mice treated with COG-Z@P200 + NIR5 exhibited no signs of
inflammatory infiltration, and their internal organs appeared
histologically normal. These findings indicate that the COG-Z@P200
+ NIR5 hydrogel possesses excellent biocompatibility and does not
induce systemic toxicity, underscoring its potential for safe in vivo
applications.
Antibacterial and microenvironmental regulation in vivo
In vivo investigation was conducted to assess the antibacterial
efficacy and tissue integration of COG-Z@P200 under near-infrared (NIR)
radiation. On the third day, wound tissue was excised to examine
bacterial growth at the wound site through Gram staining. The results,
presented in Fig. [171]8A, indicated a noticeable purple coloration in
the control and COG groups, suggesting significant bacterial presence
and potential ongoing infection. In contrast, the COG + NIR5 and
COG-Z@P200 groups exhibited much less purple staining, indicating a
reduction in bacterial infection. The COG-Z@P200 + NIR5 group showed
almost no purple staining, reflecting the substantial bacterial
clearance, which aligns with previous in vitro antibacterial results.
Further, bacterial cultures from the skin surface (Fig. [172]8B–C)
revealed that the COG-Z@P200 + NIR5 group had almost no bacterial
colonies, corresponding to an antibacterial rate of up to 98%. Analysis
of live and dead bacteria in the collected bacterial solution
(Fig. [173]8D) demonstrated a significantly higher proportion of dead
bacteria in the COG-Z@P200 + NIR5 group. These results confirm that the
incorporation of ZIF-8@PDA nanoparticles and NIR irradiation enhanced
the antibacterial properties of the hydrogel.
Fig. 8.
[174]Fig. 8
[175]Open in a new tab
Microenvironmental status of the wound site. A Gram staining of
bacteria (G+ : purple; G−: red); B Photos of bacterial colonies; C
Antibacterial rate; D Staining of live and dead bacteria. E–F
Immunofluorescence staining of TNF-α (red) and Arg-1 (red) at the wound
site on 7 th day; G–H Immunofluorescence staining of CD31 (red), VEGF
(green) and Col1 (red) on 14 th day. (Blue fluorescence indicates cell
nuclei) ***P < 0.001, ****P < 0.0001
In order to systematically evaluate the comprehensive regulatory
effects and in vivo safety of COG-Z@P200 hydrogel in diabetic wound
repair, we focused on analyzing its effects on immunomodulation,
angiogenesis, and collagen deposition. As shown in Fig. [176]8E, F, the
immunofluorescence staining results showed that COG-Z@P200 hydrogel
could effectively inhibit the expression of proinflammatory factor
TNF-α, while upregulating Arg-1, promoting the polarization of
macrophages from M1 to M2, alleviating inflammatory response, and
accelerating the transformation of wounds from the inflammatory phase
to the repair phase. The expression of angiogenesis markers CD31 and
VEGF was enhanced, and green ring structures appeared, indicating that
the new blood vessels gradually matured and the vascularization level
was significantly improved (Fig. [177]8G). In addition, as shown in
Fig. [178]8H, the immunofluorescence staining results of type I
collagen showed that COG-Z@P200 and its combined NIR irradiation group
significantly enhanced collagen deposition on the 14 th day, which is
related to tissue repair, further verifying the role of hydrogel in
promoting connective tissue regeneration. Overall, COG-Z@P200 hydrogel
synergistically regulates the wound microenvironment through multiple
mechanisms, significantly promoting the healing of diabetic infected
wounds.
To investigate the biodegradability of the hydrogels, we conducted a
systematic exploration of their degradation kinetics by simulating
various wound microenvironment conditions (including pH, enzyme
presence, and ROS levels) through in vitro experiments. The
experimental results showed that the degradation rate of hydrogels was
closely related to the intensity of microenvironmental stimulation. In
the pH-dependent degradation experiment (Figure S11 A), the acidic
environment significantly accelerated the degradation. Under pH 5.4
conditions, the residual mass of the hydrogel dropped to 30% within
7 days, while the degradation rate was significantly slowed down under
neutral (pH 6.8) and weakly alkaline (pH 7.4) environments. In the
enzymatic degradation experiment (Figure S11B), the higher the lysosome
concentration, the stronger the degradation effect. The 1.0 mg/mL
lysosome treatment group had only 20% of the residual mass on the 7 th
day, which was more thoroughly degraded than the 0.5 mg/mL group. In
the oxidative stress experiment (Figure S11 C), when the hydrogen
peroxide concentration increased from 1.0 to 10.0 mM, the residual mass
decrease rate of the hydrogel increased by nearly 3 times, and the high
concentration ROS environment significantly destroyed the stability of
the material. Similarly, the hydrogels were implanted subcutaneously to
observe their decomposition over time. By day 14, the hydrogels had
completely degraded (Figure S12 A). Histological examination using H&E
and Masson staining (Figure S12B–C) showed that the COG-Z@P200-treated
tissues did not show obvious inflammatory responses compared with the
control group. Obvious collagen deposition was seen in the treated
tissues, and after 14 days, there was no significant difference between
the treated tissues and the surrounding normal tissues. To evaluate the
antioxidant properties of the hydrogels in vivo, dihydroethidium (DHE)
staining was performed on the third day to assess the intracellular ROS
levels at the wound site (Figure S13). Compared with the control group,
the red fluorescence of ROS staining was significantly reduced in the
COG-Z@P200 and COG-Z@P200 + NIR5 hydrogel groups, indicating that the
COG-based hydrogels have excellent antioxidant capacity.
Discussion
A multicomponent synergistic photothermal hydrogel dressing
(COG-Z@P200) was engineered for the treatment of methicillin-resistant
MRSA infected wounds, combining robust antibacterial activity with
exceptional tissue compatibility. The hydrogel achieves its
bactericidal efficacy through a dual-action mechanism involving
Zn^2⁺-mediated membrane destabilization and near-infrared
(NIR)-activated mild photothermal therapy (PTT). Specifically, Zn^2⁺
ions released from COG-Z@P200 hydrogel electrostatically interact with
the negatively charged bacterial membrane, inducing structural
permeabilization. Concurrently, under NIR irradiation, localized
hyperthermia (40–45 °C) synergistically enhances membrane disruption,
facilitating rapid leakage of intracellular proteins, nucleic acids
(DNA/RNA), and cytoplasmic components, ultimately leading to bacterial
death. Crucially, this low-temperature PTT strategy can maintain the
integrity of healthy tissue while maximizing antibacterial efficiency,
effectively avoiding the thermal damage that may occur in traditional
photothermal therapy [[179]51, [180]52]. NIR light is an ideal trigger
due to its good penetration and non-invasiveness [[181]53].
Mechanistically, transcriptomic and metabolomic profiling revealed that
COG-Z@P200 hydrogel disrupts MRSA pathogenicity via multimodal
metabolic interference. Specifically, it suppresses glycolysis, blocks
the tricarboxylic acid (TCA) cycle, and impairs arginine biosynthesis,
thereby inducing DNA fragmentation and metabolic collapse.
Concurrently, the hydrogel inhibits biofilm formation by downregulating
quorum sensing-related genes (e.g., agr, operon) and suppresses cell
membrane synthesis through fatty acid metabolism inhibition.
In vivo evaluations in diabetic murine models demonstrated the
COG-Z@P200 hydrogel’s therapeutic superiority through coordinated
antioxidant, anti-inflammatory, and pro-regenerative mechanisms. DHE
staining confirmed the hydrogel’s potent ROS scavenging capacity,
significantly attenuating oxidative stress in wound tissues. Concurrent
immunofluorescence analysis revealed upregulated Arg-1 expression and
suppressed TNF-α levels, indicating M2 macrophage polarization and
resolution of chronic inflammation. These effects synergistically
preserved fibroblast functionality, enhanced angiogenesis (evidenced by
elevated VEGF/CD31 expression), and promoted structured collagen
deposition (as demonstrated via Masson’s trichrome staining),
collectively accelerating wound closure compared to conventional
treatments. By concurrently remodeling oxidative, inflammatory, and
metabolic microenvironments, the COG-Z@P200 hydrogel establishes a
multifunctional paradigm for combating drug-resistant infections in
diabetic wounds.
Compared with other wound dressings based on similar hydrogels (such as
chitosan-based, collagen-based, or silver nanoparticle-loaded
hydrogels), COG-Z@P200 + NIR5 hydrogel exhibits clear advantages in
terms of its dual antibacterial mechanisms (Zn^2⁺ release and
photothermal effect), strong mechanical properties, biodegradability,
and exceptional wound healing potential (Table S2). Additionally,
compared to the commonly used clinical antibiotic vancomycin,
COG-Z@P200 + NIR5 hydrogel demonstrates broader-spectrum antibacterial
activity, faster wound healing, and a dual-mode mechanism that prevents
bacterial resistance. Notably, vancomycin is associated with various
adverse effects, including nephrotoxicity, infusion-related “red man
syndrome,” and ototoxicity from prolonged or high-dose use. In
contrast, COG-Z@P200 + NIR5 hydrogel shows excellent biocompatibility,
very low cytotoxicity, and does not require systemic administration,
thus avoiding the risk of systemic side effects. These advantages
highlight the potential of this hydrogel as a safe and effective
non-antibiotic alternative for treating infected diabetic wounds. In
summary, COG-Z@P200 hydrogel has good scalability, mature production
process, and is suitable for large-scale production. Sterilization can
be done by pasteurization, and freeze-dried storage at −80 °C is
recommended to maintain its stability. Regarding regulation, it is
expected to be approved as a Class II medical device in the United
States and may be approved as a Class III medical device in the
European Union, which requires clinical trial verification.
Supplementary Information
[182]Additional file 1.^ (29.5MB, docx)
Acknowledgements