The adverse consequences resulting from diabetes are often presented as severe complications. Diabetic wounds are one of the most commonly occurring complications in diabetes, and the control and treatment of this is costly. Due to a series of pathophysiological mechanisms, diabetic wounds remain in the inflammatory phase for a prolonged period of time, and face difficulty in entering the proliferative phase, thus leading to chronic non-healing wounds. The current consensus on the treatment of diabetic wounds is through multidisciplinary comprehensive management, however, standard wound treatment methods are still limited and therefore, more effective methods are required. In recent years, defensins have been found to play diverse roles in a variety of diseases; however, the molecular mechanisms underlying these activities are still largely unknown. Defensins can be constitutively or inductively produced in the skin, therefore, their local distribution is affected by the microenvironment of these diabetic wounds. Current evidence suggests that defensins are involved in the diabetic wound pathogenesis, and can potentially promote the early completion of each stage, thus making research on defensins a promising area for developing novel treatments for diabetic wounds. In this review, we describe the complex function of human defensins in the development of diabetic wounds, and suggest potential thera-peutic benefits.

Diabetes was estimated to affect over 536.6 million people worldwide in 2021, and an increase in prevalence occurs at a faster rate among middle-income countries[1]. Diabetes mellitus gradually causes a series of complications, such as neuropathy, retinopathy, nephropathy, cardiovascular diseases, and diabetic wounds, as it develops. Due to lesions in the small blood vessels and peripheral nerves that are particularly prominent in the feet, diabetic wounds are usually presented as diabetic foot ulcers (DFUs), and are characterized by a delayed tissue growth and increased susceptibility to infection. Patients with DFUs have an increased risk of lower limb amputation, which has poor short-term prognosis associated with high mortality[2]. Statistically, the five-year mortality and direct costs of care for patients with DFUs have been comparable to that of cancer[3]. The large patient population and medical expenditure urgently requires an effective treatment method.

The mechanisms for diabetic wound development involve multifactorial etiologies, including a hyperglycemic microenvironment, abnormal host immune resistance, and neuropathy (Figure 1)[4]. These mechanisms influence each other, instead of occurring independently, causing irreversible diabetic complications. The local damage to vessels and nerves, reduced growth factors expression, and lower collagen accumulation contribute to repeated outbreaks and the protracted course of diabetic wounds, leading to further infections. Although current treatments, including glycemic control, anti-infective treatment, and advanced dressing application, promote wound healing by regulating the local microenvironment, they also have disadvantages, such as protracted treatment periods, high costs, and occasionally inefficient results[5]. Hence, studying methods that promote diabetic wound healing is ongoing.

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Figure 1 Mechanism of refractory diabetic wounds. The mainstream views include: Hyperglycemic microenvironment, abnormal immune system and neuropathy. Hyperglycemic microenvironment results in the complex formation of advanced glycation end products (AGEs) and cytokines, as well as circulating progenitor cell dysfunction. AGEs can significantly inhibit the proliferation of endothelial cells and alter the structure of collagen and elastin in the vascular wall, causing microvascular injury in the wound. The hallmarks of abnormal immune system are polymorphonuclear cell dysfunction, late neutrophil infiltration and suppressed macrophage polarization. As a result, diabetic wound healing is delayed and susceptible to bacterial infections and even biofilm formation. Neuropathy is occasion of subcutaneous hemorrhage, ultimately leads to skin breakdown.

With concern about antibiotic resistance growing more prominent, antimicrobial peptides (AMPs) have garnered attention as a new method of antibacterial therapy, including development of different formulation strategies for effective delivery to wounds, including AMPs loaded in nanoparticles, hydrogels, creams, gels, etc. As a representative AMP, the defensins properties are gradually being researched (Table 1)[6-23]. Human defensins are divided into α-defensins and β-defensins[23]. Human α-defensins mainly occur in neutrophils (human neutrophil peptide1-4, HNP1-4) or small intestinal Paneth cells (human defensin5-6, HD5-6). More extensively, 31 human β-defensins (HBDs) have been described, and HBD1-4 is most widely-studied[24]. Reportedly, the direct primary role of defensins is controlling microbial infections by killing bacteria and modulating the immune system. Moreover, defensins play different roles in different environments within the body, such as infected wounds, malignancy, atherosclerosis, pulmonary fibrosis, etc. In this review, we focused on investigating their mechanism of action in wound healing, especially chronic diabetic wounds.

Although little is known about defensins involvement in diabetic wounds, existing studies indicate that they play potential roles in complex pathophysiological changes of diabetic wounds[25,26]. This review summarizes and analyzes known experimental data about the role of defensins in diabetic wound healing, particularly for inflammation, cell proliferation and migration, regeneration of blood vessels and nerves, and antibacterial activities. Research articles on the role of defensins in diabetic wounds, published between inception and September 10, 2022, were collected from various search engines, such as PubMed, Google Scholar, Web of Science, and Science Direct using the following keywords: AMPs, defensins, host defense peptides, diabetic, refractory, and chronic wounds, wound healing, etc. Identified studies and relevant citations within these studies were reviewed.

The response to tissue injury involves multiple cellular and extracellular events, including inflammation, re-epithelialization, and angiogenesis, followed by fibroplasia with collagen synthesis, and tissue remodeling. Defensins may be a multifactorial modulator in the management of this process, which interferes in diabetic wounds (Table 2, Figure 2).

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Figure 2 Role of defensins in diabetic wound healing. VEGF: Vascular endothelial growth factor; Ang: Angiogenin.

Although there is ambiguity regarding its exact role, refractory healing of diabetic wounds is speculated result from interactions between multiple pathophysiological changes in the microenvironment of hyperglycemic and persistent inflammation. This affects immune cell function and composition of defensins at the wound site. In human skin, HBD1 is constitutively expressed in epithelial cells, while inducible HNP1-4 by neutrophils and HBD2-3 by keratinocytes mainly[65]. It was obtained through a biopsy that HBD2-4 were overexpressed in the border area of DFUs[25]. Studies generally agree that inadequate HBD expression is associated with poor wound healing, and many methods that promote diabetic wound healing are seemingly carried out by promoting defensins expression[66,67]. In diabetic wounds, higher HNP1, HNP3, and HNP4 expressions are more common in the central part than in the marginal areas, thus causing a significant increase in IL-8 expression under the influence of advanced glycation end products (AGEs)[29].

Defensins affect the expression and secretion of cytokines, cell proliferation, migration, and apoptosis, and are also involved in all stages of wound healing. Contrary to its proven activity in fighting pathogens and promoting tissue reconstruction, the role of defensins in inflammation and vascularization remains unclear. This discrepancy could be due to pro-inflammatory and anti-inflammatory properties being attributed to HBDs at lower concentrations[28,29], compared to antibacterial effects at higher concentrations exhibited in different experiments[48]. Thus, effects of defensins may vary depending on concentration. Furthermore, HNP1 and HBD3 exhibit increased cytotoxic effects with the increased concentration, which can also be related to a greater hydrophobicity[43,68]. Therefore, changing the local distribution or structure of defensins can have beneficial effects and prevent toxic side effects.

Studying every type of defensins within a single experiment is difficult. Additionally, certain defensins can exhibit different or contradictory effects within the same environment due to differences in experimental complexity and aims of the experiment. These factors create a huge obstacle in horizontal comparison among similar experiments. Cytotoxicity caused by defensins is difficult to assess, which indicates that topical application may be more appropriate than the systemic administration. Considering the unstable biochemical properties of defensins, topical application alone may be insufficient. To overcome this limitation, researchers are studying biological dressings as an alternative; however, formulation of an ideal material has not yet been achieved. However, animal studies on defensins exhibit improved healing outcomes, and display stable effects through application of new materials or genetic engineering methods[17,20,69]. These findings present defensins as a promising therapeutic approach owing to modern techniques, such as development of new materials to efficiently load active factors and novel protein sequences to highlight their beneficial effects.

Defensins regulates chronic inflammation, tissue regeneration, angiogenesis, and nerve recovery, as well as antimicrobial properties; therefore, they are a promising treatment for diabetic wounds. There is an urgent need to find the appropriate dosing regimens and develop new biological dressing alternatives to incorporate active factors. Hence, further preclinical investigations are necessary to understand extensive molecular mechanisms of defensins in the treatment of diabetic wounds, and consequently determine suitable therapeutic strategies.