**3. Wounds**

Wounds are divided into two categories, acute and chronic. Acute wounds can be healed predictably with normal wound healing approaches, and chronic wounds develop following one or more failures in the normal wound healing process. The wound healing process has four phases and has been described in detail above.

Acute wounds, sudden injuries to the skin, vary from superficial scratches to deep skin damages that can happen anywhere on the body. The causes of acute wounds vary but primarily include abrasion, puncture, laceration, and incision. Classified by causality, the two dominant types of acute wounds are surgical and traumatic. Surgical wounds are intentionally created for medical reasons, and traumatic wounds are randomly caused by external force. Severe pain is associated with wounds, which are especially prevalent among patients with fragile skin.

Chronic wounds remain in an inflamed state that delays healing for long periods lasting several months. Chronic wounds may take years to recover or may never heal, leading to physical and psychological su ffering as well as considerable pressure on the social healthcare system. Diabetic patients with chronic wounds are at especially high risk for bacterial infections, because the slightly alkaline pH of chronic wounds promotes bacterial colonization. Figure 3a shows the pH values of acute wounds and chronic wounds in Figure 3b (adapted from [10]). During healing, the pH of acute wounds gradually declines to a level approximating that of the acid mantle. Chronic wounds, however, remain slightly alkaline. Monitoring and manipulating pH can be a critical tool for preventing and treating chronic wounds. Studies have reported on pH value monitoring combined with pH-activated drug control release systems and smart wound dressings and bandages [11–15].

**Figure 3.** (**a**) Course of the pH milieu in acute wounds. (**b**) Course of the pH milieu in chronic wounds (adapted from [10]).

Severe chronic wounds with bacterial infections can evolve into bacteremia, sepsis, or septicemia and severe deterioration in wound status. Severely chronic wounds can result in amputation, a frequent occurrence with diabetic foot ulcers, and a major issue in modern healthcare [76,77]. Diabetic patient wound care managemen<sup>t</sup> must be undertaken with care and rigor. In 2014, chronic wounds impacted 15% of all Medicare beneficiaries in the United States, with an estimated cost of \$28–\$32 billion annually [78]. Among the 15% of Medicare beneficiaries (8.2 million in population), who had at least one type of wound or infection, surgical infections were most prevalent (4.0%), followed by diabetic infections (3.4%) [79]. The huge cost of nonhealing chronic wounds has become an urgen<sup>t</sup> issue that should be taken seriously.

#### *3.1. The Role of pH Value in Wound Healing*

pH value is one of the critical factors involved in the wound healing process for both acute wounds and chronic wounds; it affects matrix metalloproteinase activity, fibroblast activity, keratinocyte proliferation, microbial proliferation, biofilm formation, and immunological responses [80]. Different stages of the wound healing process may require environments of different pH to recover from skin damage and infection. Imbalances in pH can result in serious chronic wounds.

Matrix metalloproteinases (MMPs) are a family of more than 20 proteases. They are able to degrade extracellular components and facilitate the removal of damaged tissues after new tissues have formed. However, excessive protease amounts can interrupt the wound healing process, since they cause endothelial cell damage and degrade the epidermal–dermal junction, which eventually destroys the newly forming tissues [9,80]. To limit expression of MMPs, metalloproteinases (TIMPs) act as inhibitors; they are often at low levels in chronic wounds. Balancing the levels of MMPs and TIMPs is necessary for successful wound healing. Figure 4 (adapted from [9]) depicts the pH-dependent activity of four proteases important in wound healing. Because chronic wounds remain in an alkaline milieu, this could be one cause for high expression of MMPs that lead to unrecovered chronic wounds. However, most proteases can be inactivated in an acid milieu, so decreasing the pH value in wound sites may be an effective method for treating chronic wounds.

**Figure 4.** Assessment of pH-dependent activity profiles of four proteases important in wound healing (adapted from [9]).

Lowering the pH of the wound environment may enhance wound contraction. Pipelzadeh et al. [8] stimulated the responses of strips of rat superficial fascia in vitro using physiological solutions at different pH values (5.5, 6.6, 7.3, 8.1) and containing adenosine, calcium and potassium ions, and mepyramine. Their results sugges<sup>t</sup> that there was an at least sixfold increase in the responsiveness to adenosine under acidic conditions (pH 5.5) compared with controls, while in alkaline conditions, responses were not significantly different from control responses.

Lönnqvist et al. [81] found that human keratinocytes cultured in medium with a pH value of 6.0 showed a decreased ability to re-populate the scratched area compared to controls, and no wounds presented re-epithelialization when cultured at pH 5.0. These results sugges<sup>t</sup> that any efforts to alter local wound pH should remain over pH 5.0, which could ensure better re-epithelialization.

It is possible that different stages of wound healing require environments of different pH. An acid milieu could improve fibroblast proliferation [82], whereas neutral and alkaline environments with a pH range of 7.21–8.34 are better for re-epithelialization [83,84]. To prevent the development of chronic

wounds, an acid milieu is useful for depressing excessive amounts of MMPs and helps to form the protective acid mantle and normal skin barrier.

#### *3.2. pH in Infected Wounds*

Breidenbach et al. [85] showed that tissue microbial levels equal to or higher than 10<sup>4</sup> cfu/g were responsible for delayed wound healing. However, bacterial biofilm can a ffect the condition of chronic wounds, alter host immune response, a ffect interactions with other microorganisms, and alter pH, temperature, and nutrient levels in wound regions [86].

Ammonia, a by-product of bacterial metabolism, raises tissue pH [87]. Human pathogenic bacteria grow well when the pH value is above 6, which could lead to more complicated situation for chronic wounds. Because the pH fluctuation in chronic wounds (Figure 3b) is significantly distinct from that of the normal acid mantle, the skin fails to prevent bacterial colonization. Furthermore, the activity of MMPs is heightened when the milieu changes from neutral to alkaline, and infected chronic wounds are subjected to repeated inflammatory states that prevent healing. Changes in pH change can also influence the toxicity of bacterial end products and thus a ffect their enzyme activity. For instance, lowering pH has been shown to induce structural changes in staphylococcal enterotoxin (SEC2), which has been discussed above [51]. Decreasing pH in wound regions may not only suppress excess MMP expression but also inhibit bacterial growth and toxicity, making it a synergistical method for treating chronic wounds.

Percival et al. [88] reviewed the e ffect of some antiseptics on pH. In other research, Percival and coworkers [89] found that the antimicrobial performance of silver dressing was significantly enhanced at a pH of 5.5 when compared with a pH of 7.0. It was presumed that the oxidized form of silver present in an acidic environment was active for inhibiting bacterial growth. Molecular iodine (I2) with its active forms (I2, H2OI+) in solution is calculated to be in the pH range of 3–9 and is thought to have grea<sup>t</sup> antimicrobial activity. The greatest antimicrobial e ffect is found within an acid milieu having a pH between 3 and 6, since H2OI+ is the critical biocidal agen<sup>t</sup> most released in this environment [90]. Polyhexamethylene biguanide (PHMB) is a cationic antimicrobial that can inactivate the e fflux pump of bacterial cell membranes. The maximal antibacterial e ffect of PHMB has been shown to occur at a pH range of 5–6 [91]. Chlorhexidine (CHX) is a biguanide cationic detergent similar to PHMB and is more stable at a pH range from 5 to 8 in solution, but its antimicrobial e fficacy is optimal in a range of pH 5.5–7.0, unlike PHMB, which is more e ffective in an acidic milieu [92]. There are a number of other examples not cited in this review, but overall, optimizing pH values can benefit wound healing and inhibit microbial growth.

## *3.3. Wound Dressings*

Wound dressings are barriers that prevent wounds from further mechanical damage or infection. An ideal wound dressing should not only serve as protection but should also be able to absorb wound exudates, maintain a wet environment around the wound, and allow normal transportation of nutrients and gases. There are various types of wound dressings already available in clinical use and on the market, such as plasters, strips, tapes, foams, beads, occlusive films, hydrocolloids, hydrogels, alginate, charcoal dressings, silicone gel, etc.

In recent years, some studies have focused on developing smart wound dressings or bandages di fferent from conventional wound dressings and featuring additional functions, such as radiation-activated drug control-released system, the detection of wound status, and wireless connections between smart wound dressings and mobile phones. Smart wound dressings equipped with pH and temperature sensors can utilize the pH change in wounds to activate drug release for monitoring and treating proposes. Mirani et al. [11] designed a multifunctional hydrogel-based wound dressing capable of colorimetric measurement of pH, indication of bacterial infection, and the release of antibiotic agents at the wound site. They also developed an image analysis application that can record and analyze digital images captured by a smartphone. This facilitates monitoring of treatment strategies

that patients can manage at home. In 2018, Mostafalu and coworkers [13] reported a smart bandage for treating and monitoring chronic wounds. This hydrogel-based dressing is loaded with a temperature sensor, a pH sensor, a flexible heater, and a thermo-responsive drug release system. It may be well suited for the treatment of chronic wounds. Moreover, the sensor can be connected to a mobile phone via Bluetooth for visual recording of wound status that allows patients to self-manage wounds. Liu and coworkers [14] developed a smart hydrogel wound patch incorporating modified pH indicator dyes. It showed optimal mechanical properties under different calcium and water concentrations. When pH increased, the color of the hydrogel patches underwent a color transition from yellow to orange and red, which matched the clinically meaningful pH range of chronic or infected wounds. Kiaee and coworkers [15] reported on an electronic wound dressing with active topical drug delivery in response to electrically induced pH change. In basic environments, this dressing released drugs in response to the dehydration process. With this smart dressing, chronic wounds can be systematically and effectively treated.

#### *3.4. Wound Care*

Debridement, transplant, negative-pressure wound therapy (NPWT or VAC), and antibiotic ointments are commonly used for wound management. Debridement literally means the removal of dead or infected tissues to improve wound healing. There are several techniques to remove damaged tissues including surgical, mechanical, chemical, autolytic, and maggot-based debridement. Skin transplantation is successfully used to close open chronic wounds by placing autologous grafts or allogeneic grafts on the wound surface. This treatment option can support the rate of epithelialization of the wound areas, and rapid covering of central wound areas with keratinocytes [93]. To investigate the effect of NPWT, Armstrong and coworkers [94] analyzed 162 patients (77 assigned to NPWT and 85 control) in a randomized clinical trial and found that more patients healed in the NPWT group. The speed of wound closure and tissue formation was faster in the NPWT group than in control groups. NPWT seems to be safe and effective for treating complex diabetic foot wounds and could potentially reduce re-amputation cases compared to standard care. Antibiotic ointments are widely used for preventing wound infections. It is a fairly common wound managemen<sup>t</sup> practice in clinic and personal wound care. However, multidrug-resistant (MDR) pathogens have rapidly emerged due to the abuse of antibiotics and have created a growing a threat to human health.

Bowler and coworkers [95] reviewed infections in various wounds and suggested some associated approaches for wound management. Wound cleansing and surgical debridement are complementary with antimicrobial therapy because they could help reduce the opportunity for infection by reducing microbial load and provide better antibiotic penetration. Moreover, debridement can also avoid blood clots accumulating in tissue debris and consequently reduce the potential for microbial growth. To address the issue of MDR pathogens, nanoparticles have been applied to wound dressings to prevent infection. There are already several review articles discussing the effects of silver and silver nanoparticles in wound managemen<sup>t</sup> [96–99]. However, because silver is toxic towards mammalian cells, therapeutic window questions remain unanswered. Svitlana Chernousova and Matthias Epple [100] reviewed the toxicity of silver (silver ions and silver nanoparticles) in bacterial, cellular, and animal tests, and examined its impact on the environment. Silver toxicity is associated with particle size, size distribution, morphology, crystallinity, surface functionalization, charge, dose, and concentration. For these reasons, silver should be critically assessed and well-characterized before being used in consumer products, cosmetics, and medical products. Additionally, there is an emerging concern regarding silver-resistant bacteria [101].

Infants have fragile skin and poor skin barriers that result in easy damage. For these reasons, wound care for infants should be particularly careful and rigorous. Strodtbeck and coworkers [102] sugges<sup>t</sup> several strategies for infant wound care that are shown in Figure 5 (adapted from [102]). The scientific rationales for each care strategy are further explained in Strodtbeck's article. Stahl and coworkers [103] reported that cleansing and topical antibacterial use has helped prevent infant open wounds from getting infected from synthetic heterografts, septal patches, valves, conduits, or homograft outflow tracts used during the reconstruction of thoracic wounds. Those prostheses could increase wound complications and the risk of endovascular infection. Bookout and coworkers [104] reported on successful treatment of an infant, a toddler, and an adolescent NPWT. Notably, the infant's labial/buttock wound was completely closed without skin transplantation. They also suggested that adjustments of NPWT should be relative to age, size, and tolerance to therapy.

**Figure 5.** Strategies to promote wound healing in newborns/infants (adapted from [102]).

We sugges<sup>t</sup> monitoring pH change during wound managemen<sup>t</sup> as a means of evaluating wound healing. Measuring pH value in wound sites is relatively simple and fast. It could be used to provide a rough reference for physicians to diagnose and assess wound healing state, including whether wounds are receiving effective treatment or are becoming chronic, unhealing wounds. Additional examination and analysis, including ultrasound, wound depth measurement, and infection testing could supplement this initial, rough assessment to best determine treatment protocol.
