Electrical fields in wound healing—An overriding signal that directs cell migration

Injury that disrupts an epithelial layer instantaneously generates endogenous electric fields (EFs), which were detected at human skin wounds over 150 years ago. Recent researches combining molecular, genetic and imaging techniques have provided significant insights into cellular and molecular responses to this “unconventional” signal. One unexpected finding is that the EFs play an overriding guidance role in directing cell migration in epithelial wound healing. In experimental models where other directional cues (e.g., contact inhibition release, population pressure etc.) are present, electric fields of physiological strength override them and direct cell migration. The electrotaxis or galvanotaxis is mediated by polarized activation of multiple signaling pathways that include PI3 kinases/Pten, membrane growth factor receptors and integrins. Genetic manipulation of PI3 kinase/Pten (Phosphoinositide 3-kinases/phosphatase and tensin homolog) and integrin β4 demonstrated the importance of those molecules. The electric fields are therefore a fundamental signal that directs cell migration in wound healing. One of the most challenging question is: How do cells sense the very weak electric signals? Clinically, it is highly desirable to develop practical and reliable technologies for wound healing management exploiting the electric signaling.     

Airway epithelial wounds in rhesus monkey generate ionic currents that guide cell migration to promote healing

Damage to the respiratory epithelium is one of the most critical steps to many life-threatening diseases, such as acute respiratory distress syndrome and chronic obstructive pulmonary disease. The mechanisms underlying repair of the damaged epithelium have not yet been fully elucidated. Here we provide experimental evidence suggesting a novel mechanism for wound repair: endogenous electric currents. It is known that the airway epithelium maintains a voltage difference referred to as the transepithelial potential. Using a noninvasive vibrating probe, we demonstrate that wounds in the epithelium of trachea from rhesus monkeys generate significant outward electric currents. A small slit wound produced an outward current (1.59 μA/cm(2)), which could be enhanced (nearly doubled) by the ion transport stimulator aminophylline. In addition, inhibiting cystic fibrosis transmembrane conductance regulator (CFTR) with CFTR(Inh)-172 significantly reduced wound currents (0.17 μA/cm(2)), implicating an important role of ion transporters in wound induced electric potentials. Time-lapse video microscopy showed that applied electric fields (EFs) induced robust directional migration of primary tracheobronchial epithelial cells from rhesus monkeys, towards the cathode, with a threshold of <23 mV/mm. Reversal of the field polarity induced cell migration towards the new cathode. We further demonstrate that application of an EF promoted wound healing in a monolayer wound healing assay. Our results suggest that endogenous electric currents at sites of tracheal epithelial injury may direct cell migration, which could benefit restitution of damaged airway mucosa. Manipulation of ion transport may lead to novel therapeutic approaches to repair damaged respiratory epithelium.     

The role of electrical signals in murine corneal wound re-epithelialization

Ion flow from intact tissue into epithelial wound sites results in lateral electric currents that may represent a major driver of wound healing cell migration. Use of applied electric fields (EF) to promote wound healing is the basis of Medicare-approved electric stimulation therapy. This study investigated the roles for EFs in wound re-epithelialization, using the Pax6(+/-) mouse model of the human ocular surface abnormality aniridic keratopathy (in which wound healing and corneal epithelial cell migration are disrupted). Both wild-type (WT) and Pax6(+/-) corneal epithelial cells showed increased migration speeds in response to applied EFs in vitro. However, only Pax6(+/+) cells demonstrated consistent directional galvanotaxis towards the cathode, with activation of pSrc signaling, polarized to the leading edges of cells. In vivo, the epithelial wound site normally represents a cathode, but 43% of Pax6(+/-) corneas exhibited reversed endogenous wound-induced currents (the wound was an anode). These corneas healed at the same rate as WT. Surprisingly, epithelial migration did not correlate with direction or magnitude of endogenous currents for WT or mutant corneas. Furthermore, during healing in vivo, no polarization of pSrc was observed. We found little evidence that Src-dependent mechanisms of cell migration, observed in response to applied EFs in vitro, normally exist in vivo. It is concluded that endogenous EFs do not drive long-term directionality of sustained healing migration in this mouse corneal epithelial model. Ion flow from wounds may nevertheless represent an important component of wound signaling initiation.     

EGF and TGF-alpha in wound healing and repair

Wound healing is a localized process which involves inflammation, wound cell migration and mitosis, neovascularization, and regeneration of the extracellular matrix. Recent data suggest the actions of wound cells may be regulated by local production of peptide growth factors which influence wound cells through autocrine and paracrine mechanisms. Two peptide growth factors which may play important roles in normal wound healing in tissues such as skin, cornea, and gastrointestinal tract are the structurally related peptides epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha). EGF/TGF-alpha receptors are expressed by many types of cells including skin keratinocytes, fibroblasts, vascular endothelial cells, and epithelial cells of the GI tract. In addition, EGF or TGF-alpha are synthesized by several cells involved in wound healing including platelets, keratinocytes, and activated macrophages. Healing of a variety of wounds in animals and patients was enhanced by treatment with EGF or TGF-alpha. Epidermal regeneration of partial thickness burns on pigs or dermatome wounds on patients was accelerated with topical application of EGF or TGF-alpha, and EGF treatment accelerated healing of gastroduodenal ulcers. EGF also increased tensile strength of skin incisions in rats and corneal incisions in rabbits, cats, and primates. Additional research is needed to better define the roles of EGF, TGF-alpha and their receptor in normal wound healing, to determine if alterations have occurred in the EGF/TGF-alpha system in chronic wounds, and optimize vehicles for effective delivery of peptide growth factors to wounds.     

Mesenchymal stem cell-derived exosomes: A novel and potential remedy for cutaneous wound healing and regeneration

Poor healing of cutaneous wounds is a common medical problem in the field of traumatology. Due to the intricate pathophysiological processes of wound healing, the use of conventional treatment methods, such as chemical molecule drugs and traditional dressings, have been unable to achieve satisfactory outcomes. Within recent years, explicit evidence suggests that mesenchymal stem cells (MSCs) have great therapeutic potentials on skin wound healing and regeneration. However, the direct application of MSCs still faces many challenges and difficulties. Intriguingly, exosomes as cell-secreted granular vesicles with a lipid bilayer membrane structure and containing specific components from the source cells may emerge to be excellent substitutes for MSCs. Exosomes derived from MSCs (MSC-exosomes) have been demonstrated to be beneficial for cutaneous wound healing and accelerate the process through a variety of mechanisms. These mechanisms include alleviating inflammation, promoting vascularization, and promoting proliferation and migration of epithelial cells and fibroblasts. Therefore, the application of MSC-exosomes may be a promising alternative to cell therapy in the treatment of cutaneous wounds and could promote wound healing through multiple mechanisms simultaneously. This review will provide an overview of the role and the mechanisms of MSC-derived exosomes in cutaneous wound healing, and elaborate the potentials and future perspectives of MSC-exosomes application in clinical practice.     

Circulating fibrocyte mobilization in negative pressure wound therapy

Non‐healing diabetic wounds are difficult to treat. They also create heavy financial burdens for both patients and society. Negative pressure wound therapy (NPWT) has been adopted to treat intractable wounds and has proved to be effective. However, the mechanisms that underlie the effects of this treatment are not entirely understood. Circulating fibrocytes are unique haematopoietic‐derived stem cells that have been reported to play a pivotal role in wound healing. Here, we have investigated the effect of NPWT on fibrocyte mobilization and the role of fibrocyte mobilization in the healing of diabetic wounds during NPWT. We show that the NPWT group exhibited 2.6‐fold to 12.1‐fold greater numbers of tail vein‐injected PKH‐26‐labelled fibrocytes in the diabetic wound sites compared with the control group. We also demonstrate that the full‐thickness skin wounds treated with NPWT exhibit significantly reduced mRNA and protein expression, blood vessel density and proliferating cells when exogenous fibrocyte mobilization is inhibited. We speculate that systemic mobilization of fibrocytes during NPWT may be a mechanism for healing intractable wounds in a diabetic rat model experiment and that enhancement of cell mobilization may represent a potential treatment idea for intractable wound healing across all fields of surgery.     

Effects of Senescence in Wound Healing

With the advent of aging society, the effects of aging on all aspects of the body are attracting more and more attention. Among them, the increasing incidence of chronic wounds in the elderly not only affects the quality of the elderly’s life significantly, but also brings a heavy medical burden on society. Delayed and poor wound healing increases the possibility of severe infections. To find out a solution for infection and chronic wounds, it is necessary to clarify the specific mechanisms of wound healing and possible intervention targets. Wound healing is a complex physiological process in the human body, which involves the coordinated activation of multiple cell types and signaling pathways. The role of senescent cells in wound healing is causing growing attention in recent years. It was thought that wound healing needs to take a longer time in elder people. In recent years, it has been found that senescent cells promote wound healing. So far, the effects of senescent cells on the efficiency and quality of wound healing and its specific mechanism have not been fully clarified. What is certain is that different types of senescent cells and even different subtypes of the same senescent cells play different roles in fast and chronic wound healing. It is not only the heterogeneity of the senescent cell itself, but also the difference in the surrounding microenvironment that determines the effect of senescent cells on wound healing. The study of its mechanism is helpful to find a way to promote the healing of wounds. It is worth noting that senescent cells themselves may also induce poor wound prognosis, such as chronic wounds, inflammation and decreased anti-infection ability. Therefore, the ideal treatment strategy to apply senescent cells will be a comprehensive plan that maximizes the efficiency and quality of wound healing, while minimizing the risk of senescent cells itself becoming an inducement for chronic wounds.     

Biosorptive-flotation of copper(II) from environmental water samples using sugar beet pulp as sorbent and oleic acid as surfactant

Abstract

Wounds are likely to have existed ever since mankind has existed. Wound healing is an enormously complicated process and the actual scientific mechanisms and events that take place during healing are far more complex and dynamic than might be imagined. Essential elements, especially trace elements are believed to be pivotal to the wound healing process. Their involvement in tissue regeneration and repair appears to be wide ranging and their deficiencies have been reported to impair the healing process. However, further research is required to establish the involvement of trace elements and their specific species in the wound healing process.

Thus assessments of trace element levels in wound tissues using new, reliable, verified and validated technologies could be beneficial for trace element based wound healing. Here, is an attempt made to assess the link between trace element concentrations and healing processes of chronic and acute wounds. Wound tissue samples from 58 chronic leg ulcers and 50 acute wounds were analysed for concentrations of Fe, Zn, Cu, Mn, Ca, Sn, Cr, Cd and Pb using atomic absorption spectrophotometer. Blood samples were also collected from the same patients and analysed in the same manner. All the data were tested for normality by Ryan–Joiner normality test (α = 0.05) and one-way ANOVA was done for the normally distributed data.

The results showed that the concentration of Ca, Zn, Cu and Mn were similar in both acute and chronic wounds. Apart from Ca, which showed elevated concentrations, other metal concentrations are either similar or lower than the analysed concentrations in blood. It appears that there is a significant difference in the concentrations of iron accumulated in the tissues of chronic and acute wounds.

The concentrations of Sn, Cr, Cd and Pb, which are considered as toxic metals, were not present in detectable levels with the graphite furnace atomic absorption spectrophotometry in both types of wound tissues.     

Placenta mesenchymal stem cell accelerates wound healing by enhancing angiogenesis in diabetic Goto-Kakizaki (GK) rats

Multipotent mesenchymal stem cells have recently emerged as an attractive cell type for the treatment of diabetes-associated wounds. The purpose of this study was to examine the potential biological function of human placenta-derived mesenchymal stem cells (PMSCs) in wound healing in diabetic Goto-Kakizaki (GK) rats. PMSCs were isolated from human placenta tissue and characterized by flow cytometry. A full-thickness circular excisional wound was created on the dorsum of each rat. Red fluorescent CM-DiI-labeled PMSCs were injected intradermally around the wound in the treatment group. After complete wound healing, full-thickness skin samples were taken from the wound sites for histological evaluation of the volume and density of vessels. Our data showed that the extent of wound closure was significantly enhanced in the PMSCs group compared with the no-graft controls. Microvessel density in wound bed biopsy sites was significantly higher in the PMSCs group compared with the no-graft controls. Most surprisingly, immunohistochemical studies confirmed that transplanted PMSCs localized to the wound tissue and were incorporated into recipient vasculature with improved angiogenesis. Notably, PMSCs secreted comparable amounts of proangiogenic molecules, such as VEGF, HGF, bFGF, TGF-β and IGF-1 at bioactive levels. This study demonstrated that PMSCs improved the wound healing rate in diabetic rats. It is speculated that this effect can be attributed to the PMSCs engraftment resulting in vascular regeneration via direct de novo differentiation and paracrine mechanisms. Thus, placenta-derived mesenchymal stem cells are implicated as a potential angiogenesis cell therapy for repair-resistant chronic wounds in diabetic patients.     

Calcium Ion Flow Permeates Cells through SOCs to Promote Cathode-Directed Galvanotaxis

Sensing and responding to endogenous electrical fields are important abilities for cells engaged in processes such as embryogenesis, regeneration and wound healing. Many types of cultured cells have been induced to migrate directionally within electrical fields in vitro using a process known as galvanotaxis. The underlying mechanism by which cells sense electrical fields is unknown. In this study, we assembled a polydimethylsiloxane (PDMS) galvanotaxis system and found that mouse fibroblasts and human prostate cancer PC3 cells migrated to the cathode. By comparing the effects of a pulsed direct current, a constant direct current and an anion-exchange membrane on the directed migration of mouse fibroblasts, we found that these cells responded to the ionic flow in the electrical fields. Taken together, the observed effects of the calcium content of the medium, the function of the store-operated calcium channels (SOCs) and the intracellular calcium content on galvanotaxis indicated that calcium ionic flow from the anode to the cathode within the culture medium permeated the cells through SOCs at the drift velocity, promoting migration toward the cathode. The RTK-PI3K pathway was involved in this process, but the ROCK and MAPK pathways were not. PC3 cells and mouse fibroblasts utilized the same mechanism of galvanotaxis. Together, these results indicated that the signaling pathway responsible for cathode-directed cellular galvanotaxis involved calcium ionic flow from the anode to the cathode within the culture medium, which permeated the cells through SOCs, causing cytoskeletal reorganization via PI3K signaling.     

Effects of synthetic defensin fragments on aggregation and adhesion of epitheliolike cells

Normal course of processes of regeneration and epithelization of damaged tissues has been shown to be based on the capability of cells participating in these processes for selective adhesion. In the case of the complete or partial absence of this capability in the cells-participants of the wound healing process, the so-called non-healing wounds appear. In this connection, it remains actual to search for natural agents promoting healing of chronic non-healing wounds. In the present work, we studied effects of synthetic fragments of leukocytic antimicrobial peptides defensines—GER, FGER, and GERA—on aggregation and adhesion of epitheliolike cells of the CHO-K1 line. These peptides have been established to have aggregate-stimulating properties; besides, they enhance adhesion of the cells to the untreated plastic and inhibit fibronectinmediated cell adhesion. Possible pathways of regulation by peptides of processes of intercellular and cell-matrix interaction are discussed as well as ways of release of these compounds in an organism and their functional role in an organism.     

Effects of synthetic fragments of defensins on aggregation and adhesion of epitheliolike cells

Normal course of processes of regeneration and epitheliazation of damage tissues has been shown to be based on the capability of cells participating in these processes for selective adhesion. In the case of the complete or partial absence of this capability in the cells-participants of the wound healing process, the so-called non-healing wounds appear. In this connection, it remains actual to search for natural agents promoting healing of chronic non-healing wounds. In the present work, we studied effects of synthetic fragments of leukocytic antimicrobial peptides defensines--GER, FGER, and GERA--on aggregation and adhesion of epitheliolike cells of the CHO-K1 line. These peptides have been established to have aggregate-stimulating properties; besides, they enhance adhesion of the cells to the untreated plastic and inhibit fibronectinmediated cell adhesion. Possible pathways of regulation by peptides of processes of intercellular and cell-matrix interaction are discussed as well as ways of release of these compounds in an organism and their functional role in an organism.     

The role of oxidised regenerated cellulose/collagen in chronic wound repair and its potential mechanism of action

Normal wound healing is a carefully controlled balance of destructive processes necessary to remove damaged tissue and repair processes which lead to new tissue formation. Proteases and growth factors play a pivotal role in regulating this balance, and if disrupted in favour of degradation then delayed healing ensues; a trait of chronic wounds. Whilst there are many types of chronic wounds, biochemically they are thought to be similar in that they are characterised by a prolonged inflammatory phase, which results in elevated levels of proteases and diminished growth factor activity. This increase in proteolytic activity and subsequent degradation of growth factors is thought to contribute to the net tissue loss associated with these chronic wounds.

In this study, we describe a new wound treatment, comprising oxidised regenerated cellulose and collagen (ORC/collagen), which can redress this imbalance and modify the chronic wound environment. We demonstrate that ORC/collagen can inactivate potentially harmful factors such as proteases, oxygen free radicals and excess metal ions present in chronic wound fluid, whilst simultaneously protecting positive factors such as growth factors and delivering them back to the wound.

These characteristics suggest a beneficial role for this material in helping to re-balance the chronic wound environment and therefore promote healing.     

Soft tissue fibroblasts from well healing and chronic human wounds show different rates of myofibroblasts in vitro

Due to an increasing life expectancy in western countries, chronic wound treatment will be an emerging challenge in the next decades. Because therapies are improving slowly appropriate diagnostic tools enabling the early prediction of the healing success remain to be developed. We used a well-established in vitro assay in combination with the analysis of 27 cytokines to discriminate between fibroblasts from chronic (n = 6) and well healing (n = 8) human wounds. Proliferation and migration of the cells as well as their response to hypoxia and their behaviour in co-culture with microvascular endothelial cells were analyzed. Myofibroblast differentiation, a time-limited essential process of regular wound healing, was also quantified. Besides weaker proliferation and migration significantly higher rates of myofibroblasts were detected in chronic wounds. With respect to the cytokine release, there was a clear trend within the group of chronic wound fibroblasts, which were releasing interferon-γ, monocyte chemotactic protein-1, granulocyte–macrophage colony stimulating factor and basic fibroblast growth factor in higher amounts than fibroblasts from healing wounds. Although the overall response of both groups of fibroblasts to hypoxia and to the contact with endothelial cells was similar, especially chronic wound fibroblasts seemed to benefit from the endothelial interaction during hypoxia and displayed better migration characteristics. The study shows (1) that the assay can identify specific features of fibroblasts derived from different human wounds and (2) that wound fibroblasts are varying in their response to the chosen parameters. Thus, current therapeutic approaches and individual healing prediction might benefit from this assay.     

Cellular mechanisms of skin repair in humans and other mammals

The increased incidence of non-healing skin wounds in developed societies has prompted tremendous research efforts on the complex process known as “wound healing”. Unfortunately, the weak relevance of modern wound healing research to human health continues to be a matter of concern. This review summarizes the current knowledge of the cellular mechanisms that mediate wound closure in the skin of humans and laboratory animals. The author highlights the anatomical singularities of human skin vs. the skin of other mammals commonly used for wound healing research (i.e. as mice, rats, rabbits, and pigs), and discusses the roles of stem cells, myofibroblasts, and the matrix environment in the repair process. The majority of this review focuses on reepithelialization and wound closure. Other aspects of wound healing (e.g. inflammation, fibrous healing) are referred to when relevant to the main topic. This review aims at providing the reader with a clear understanding of the similarities and differences that have been reported over the past 100 years between the healing of human wounds and that of other mammals.     

Bioelectric potential gradients may initiate cell cycling: ELF and zeta potential gradients may mimic this effect

When a number of experimental studies in bioelectromagnetics were reviewed, those in which weak, exogenous extremely low frequency (ELF) fields were applied in fixed juxtaposition to their target tissues, were found to initiate mitogenesis or mitogenesis-related signals more successfully than when the target tissue moved freely during the irradiation. It is suggested that ELF fields in fixed juxtaposition to their target tissue and implanted foreign bodies or endogenous tissues with a significant zeta potential, mimic bioelectric fields generated at wounds. When the potential is high enough, they assist healing by moving cells into the wound and stimulating quiescent cells at the wound margin to cycle. Electrophoresis may help the initial migration of cells into the wound to form a clot, and migration of fibroblasts and epithelial cells from the wound margin. When exposed for a long time in a fixed juxtaposition to a potential gradient too weak to show in situ microelectrophoresis along the cell membrane surface, surface particles may coalesce to form microclusters, where like-charged surface particles are in close proximity, and growth factor receptor oligomerization and other cycle-initiating reactions are facilitated. Bioelectromagnetics 18:341–348, 1997. © 1997 Wiley-Liss, Inc.     

Mathematical Modeling in Wound Healing,Bone Regeneration and Tissue Engineering

The processes of wound healing and bone regeneration and problems in tissue engineering have been an active area for mathematical modeling in the last decade. Here we review a selection of recent models which aim at deriving strategies for improved healing. In wound healing, the models have particularly focused on the inflammatory response in order to improve the healing of chronic wound. For bone regeneration, the mathematical models have been applied to design optimal and new treatment strategies for normal and specific cases of impaired fracture healing. For the field of tissue engineering, we focus on mathematical models that analyze the interplay between cells and their biochemical cues within the scaffold to ensure optimal nutrient transport and maximal tissue production. Finally, we briefly comment on numerical issues arising from simulations of these mathematical models.     

Growth Inhibition in Candida albicans due to Low Intensity Constant Direct Current

Electrical stimulation with weak constant direct currents is the treatment of choice for enhancement of wound healing. The use of a negative electrode placed in the wound has been reported to be successful in treating infected ulcers by inhibiting bacterial growth. The aim of our study was to examine the in vitro effects of a low intensity constant direct current, of 0.2 to ImA, applied for 2 to 18 hours on Candida albicans yeast. Electric current was applied using two systems: over Pt-Ir electrodes directly immersed in the culture medium and over agar bridges, which prevented the electrochemical reactions at the metal electrodes and their byproducts from influencing the microbe growth. It was found that even such weak direct currents inhibit the Candida albicans' growth. The inhibitory action was proportional to the magnitude and application time of the electric current.