Chronic wounds, which fail to heal within a normal timeframe, affect a significant portion of the population, posing a substantial healthcare burden. Current treatments lack effective targeted therapies, highlighting the need for innovative approaches. This study focuses on leveraging the combined benefits of bioactive peptides, hydrogels, and nanomaterials to address this challenge. Bioactive peptides, such as RL-QN15, have shown promise in wound healing. Hydrogels, particularly sodium alginate (SA) hydrogels crosslinked with Zn²⁺, offer advantages due to their biocompatibility and ability to mimic the extracellular matrix (ECM). Nanomaterials, like hollow polydopamine (HPDA) nanoparticles, provide enhanced drug delivery capabilities and antioxidant properties. Previous research demonstrated the therapeutic potential of RL-QN15, but optimization was needed to enhance its efficacy and reduce costs. This study aimed to optimize RL-QN15, creating CYRL-QN15, a cyclic heptapeptide, and integrate it within a HPDA/CYRL-QN15/ZA hydrogel for enhanced chronic wound healing.

The literature review highlights the limitations of current chronic wound treatments and the growing interest in bioactive peptides, hydrogels, and nanomaterials as therapeutic agents. Several bioactive peptides derived from amphibian skin, including AO-GL12, cathelicidin-OA1, cathelicidin-NV, and RL-QN15, have demonstrated efficacy in wound healing. The use of hydrogels as wound dressings is supported by their ability to maintain a moist environment and promote cell proliferation. Sodium alginate, with its biocompatibility and ability to form gels with divalent cations like Zn²⁺, is a suitable hydrogel candidate. Nanomaterials, especially HPDA nanoparticles, offer advantages in drug delivery and antioxidant activity. The researchers cite previous work demonstrating the potential of RL-QN15 and the benefits of loading it into HPDA nanoparticles, justifying the need for structural optimization and hydrogel formulation.

The study involved several key steps: 1. **Peptide Synthesis and Optimization:** RL-QN15 and its derivatives (RE-RL-QN15, LIRL-QN15, and CYRL-QN15) were synthesized. The structure and stability of CYRL-QN15 were characterized. 2. **Cell Culture:** Human keratinocytes (HaCaT), human skin fibroblasts (HSFs), human umbilical vein endothelial cells (HUVECs), and mouse macrophages (RAW 264.7) were cultured. 3. **In vitro Assays:** Scratch healing assays were used to assess the effects of peptides on keratinocyte migration. HUVEC migration and tube formation assays evaluated angiogenesis. Cytokine levels (TGF-β1, TNF-α) were measured using ELISA. Antioxidant activity was assessed using free radical scavenging assays. 4. **Hydrogel Preparation and Characterization:** A Zn²⁺-crosslinked SA hydrogel containing HPDA nanoparticles loaded with CYRL-QN15 (HPDA/CYRL-QN15/ZA hydrogel) was prepared. Loading and release efficiency, biocompatibility, and degradation were determined. 5. **In vivo Studies (Diabetic Mice):** Full-thickness skin wounds were created in diabetic mice. The HPDA/CYRL-QN15/ZA hydrogel was applied, and wound healing was assessed over time. Cytokine levels and histological analysis (H&E, Masson trichrome, PAS, immunohistochemistry, immunofluorescence) were performed to assess tissue regeneration and macrophage polarization. 6. **Ex vivo Human Studies:** A human ex vivo diabetic skin wound model was used to assess the hydrogel's efficacy in human tissue.

CYRL-QN15, a cyclic heptapeptide derived from RL-QN15, exhibited comparable or superior wound healing properties compared to RL-QN15 in both in vitro and in vivo models. In vitro, the HPDA/CYRL-QN15/ZA hydrogel promoted keratinocyte proliferation, migration, and angiogenesis, while regulating macrophage cytokine secretion and exhibiting antioxidant activity. In vivo studies in diabetic mice demonstrated that the hydrogel significantly accelerated wound healing, promoting M2 macrophage polarization, re-epithelialization, granulation tissue formation, collagen deposition, and angiogenesis. The improved wound healing was also observed in an ex vivo human diabetic skin wound model, confirming the translational potential of the hydrogel.

The study successfully demonstrated the therapeutic potential of the HPDA/CYRL-QN15/ZA hydrogel in promoting chronic wound healing. The optimization of RL-QN15 to CYRL-QN15 resulted in a shorter, more stable, and potentially more cost-effective peptide with enhanced activity. The incorporation of CYRL-QN15 within a Zn²⁺-crosslinked SA hydrogel, further enhanced by the inclusion of HPDA nanoparticles, provided a synergistic effect, leading to significant improvements in various aspects of the wound healing process. The findings support the combined use of peptides, hydrogels, and nanomaterials as a promising strategy for treating chronic wounds, particularly in diabetic patients. The ex vivo human studies provide strong evidence for the clinical translation of this hydrogel.

This research successfully developed a novel HPDA/CYRL-QN15/ZA hydrogel that significantly accelerated chronic skin wound healing in both diabetic mice and human ex vivo models. The optimization of RL-QN15 into the cyclic heptapeptide CYRL-QN15, combined with the delivery system of the hydrogel, resulted in a potent therapeutic agent. Future research could focus on clinical trials to evaluate the hydrogel's efficacy and safety in larger patient populations, exploring different formulations and potential modifications to further enhance its therapeutic potential.

While the study provides compelling evidence for the efficacy of the HPDA/CYRL-QN15/ZA hydrogel, some limitations exist. The sample sizes in both the animal and human studies were relatively small. Long-term effects of the hydrogel and its potential for scar formation require further investigation. The study primarily focused on diabetic wounds; further research is needed to determine its effectiveness in other types of chronic wounds.