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GRHL3 is upregulated in wound-front keratinocytes of acute wounds, and its expression is altered in human chronic wounds. (A) Schematic representation of the full-thickness mouse wound model for expression studies, showing mice reporting Grhl3 expression with βGal (LacZ) expression (left panel). A wound showing location of the proliferating epidermis (PE), the migrating wound-front epidermis (ME), and a scab overlying the wound (right panel). (B) IHC localization of GRHL3 expression (anti-βGal antibody) in mouse skin sections at 0, 1, 3, and 7 days after wounding. Right panels show higher magnifications of the indicated areas in the black boxes. Scale bar: 60 μm. Black arrows point to the nuclear expression of GRHL3 in keratinocytes. (C) Schematic representation of a biopsy from a healing human wound (left panel). H&E staining of a healing human wound (right panel). (D) IHC staining for GRHL3 (anti-GRHL3 antibody) in a healing human wound. Right panel shows higher magnification of the indicated area in the black box. Black arrows point to the nuclear expression of GRHL3 in suprabasal cells in reepithelized human epidermis. Scale bar: 36 μm. (E) Schematic representation of a biopsy collected from a human chronic nonhealing wound (left panel). H&E staining of a human chronic nonhealing wound (right panel). (F) IHC staining for GRHL3 (anti-GRHL3 antibody) in a human chronic nonhealing wound. Right panel shows higher magnification of the indicated area in the black box. Black arrows point to the increased expression of GRHL3 in the deeper part of the chronic wound epidermis. Scale bar: 36 μm.
Source publication
The migrating keratinocyte wound front is required for skin wound closure. Despite significant advances in wound healing research, we do not fully understand the molecular mechanisms that orchestrate collective keratinocyte migration. Here, we show that, in the wound front, the epidermal transcription factor Grainyhead like-3 (GRHL3) mediates decre...
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... created small wounds (4 mm) on the back skin of Grhl3-Cre;LacZ reporter mice that have previously been shown to faithfully mark the endogenous expression of Grhl3 in skin and other tissues (14). We preformed IHC analysis for β-Galactosidase (βGal) at several time points after wounding ( Figure 1A; n = 3). In unwounded skin, βGal expression is sparse and within the suprabasal layer of the epidermis ( Figure 1B). ...
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... preformed IHC analysis for β-Galactosidase (βGal) at several time points after wounding ( Figure 1A; n = 3). In unwounded skin, βGal expression is sparse and within the suprabasal layer of the epidermis ( Figure 1B). Twenty-four hours after wounding, when a migrating epithelium is forming, βGal is upregulated and more uniformly expressed in keratinocytes located in the wound front -in particular, suprabasally ( Figure 1B). ...
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... unwounded skin, βGal expression is sparse and within the suprabasal layer of the epidermis ( Figure 1B). Twenty-four hours after wounding, when a migrating epithelium is forming, βGal is upregulated and more uniformly expressed in keratinocytes located in the wound front -in particular, suprabasally ( Figure 1B). At day 3 after wounding, when keratinocyte migration is prominent, βGal is highly upregulated in migrating suprabasal keratinocytes in the wound front ( Figure 1B). ...
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... hours after wounding, when a migrating epithelium is forming, βGal is upregulated and more uniformly expressed in keratinocytes located in the wound front -in particular, suprabasally ( Figure 1B). At day 3 after wounding, when keratinocyte migration is prominent, βGal is highly upregulated in migrating suprabasal keratinocytes in the wound front ( Figure 1B). At day 7 after wounding, when epidermal cells have completed migration across the wound, increased expression of βGal persists in suprabasal keratinocytes ( Figure 1B), consistent with the known role of GRHL3 in epidermal barrier restoration after injury (14). ...
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... day 3 after wounding, when keratinocyte migration is prominent, βGal is highly upregulated in migrating suprabasal keratinocytes in the wound front ( Figure 1B). At day 7 after wounding, when epidermal cells have completed migration across the wound, increased expression of βGal persists in suprabasal keratinocytes ( Figure 1B), consistent with the known role of GRHL3 in epidermal barrier restoration after injury (14). These data, showing upregulation of Grhl3 in wound-front keratinocytes, are consistent with an active role of GRHL3 in keratinocyte migration during healing of acute wounds. ...
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... analysis of GRHL3 in punch biopsies collected from healthy individuals with healing acute wounds also shows upregulation of GRHL3 in the wounded epidermis (Figure 1, C and D, and Supplemental Figure 1A; supplemental material available online with this article; https://doi. org/10.1172/jci.insight.142577DS1; ...
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... analysis of GRHL3 in punch biopsies collected from healthy individuals with healing acute wounds also shows upregulation of GRHL3 in the wounded epidermis (Figure 1, C and D, and Supplemental Figure 1A; supplemental material available online with this article; https://doi. org/10.1172/jci.insight.142577DS1; ...
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... = 3). In contrast to mouse skin, in humans, we detected some GRHL3 expression in the basal layer, but as in the mouse, its highest expression is in the suprabasal layers of the epidermis, the regenerating spinous and granular layers ( Figure 1D). Contrasting to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). ...
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... contrast to mouse skin, in humans, we detected some GRHL3 expression in the basal layer, but as in the mouse, its highest expression is in the suprabasal layers of the epidermis, the regenerating spinous and granular layers ( Figure 1D). Contrasting to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). ...
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... contrast to mouse skin, in humans, we detected some GRHL3 expression in the basal layer, but as in the mouse, its highest expression is in the suprabasal layers of the epidermis, the regenerating spinous and granular layers ( Figure 1D). Contrasting to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). ...
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... contrast to mouse skin, in humans, we detected some GRHL3 expression in the basal layer, but as in the mouse, its highest expression is in the suprabasal layers of the epidermis, the regenerating spinous and granular layers ( Figure 1D). Contrasting to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). ...
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... contrast to mouse skin, in humans, we detected some GRHL3 expression in the basal layer, but as in the mouse, its highest expression is in the suprabasal layers of the epidermis, the regenerating spinous and granular layers ( Figure 1D). Contrasting to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). ...
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... to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). Collectively, these data show that GRHL3 is upregulated in the wound front of healing wounds, consistent with a role in skin wound healing, and that GRHL3's expression is abnormally increased in the basal layer in chronic human wounds. ...
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... to acute healing wounds, chronic wounds ( Figure 1, E and F, and Supplemental Figure 1A) have a higher proportion of basally located cells expressing GRHL3 (Supplemental Figure 1B; n = 3), although the total proportion of keratinocytes expressing GRHL3 is similar between healing and chronic wounds (Supplemental Figure 1C). Notably, basal keratinocytes in chronic wounds with high GRHL3 expression exhibit increased intercellular spaces and elongated, migratory-like cell morphology -features that are not observed in basal keratinocytes in acutely healing wounds ( Figure 1F and Supplemental Figure 1A). Collectively, these data show that GRHL3 is upregulated in the wound front of healing wounds, consistent with a role in skin wound healing, and that GRHL3's expression is abnormally increased in the basal layer in chronic human wounds. ...
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... GRHL3 is highly expressed in human chronic wounds (Figure 1), we next investigated whether components of the GRHL3/FSCN1/E-cadherin pathway are altered in mouse diabetic wounds, a model of delayed wound healing where mice are made diabetic through high-fat feeding (27) (Figure 6D; n = 3); diabetes was confirmed with fasting glucose measurements (fasting blood glucose > 200 mg/dL). Although FSCN1 was detected in some basal keratinocytes in diabetic wounds, it was downregulated in suprabasal keratinocytes in the wound front compared with controls ( Figure 6E and Supplemental Figure 6A). ...
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... the GRHL3/FSCN1/E-cadherin pathway needs to be further investigated in chronic wounds, our work already suggests that it may be disrupted in conditions associated with impaired wound healing. We show an altered GRHL3 expression pattern in human chronic wounds (Figure 1). We also show that the FSCN1/E-cadherin pathway is altered in diabetic wounds, although this appears to be independent of GRHL3 ( Figure 6). ...
Citations
... In this study, we revealed the novel role of GRHL3/Lnc-DC/ZNF750 axis in regulating keratinocytes differentiation. GRHL3 and ZNF750 have been reported to be important regulators in skin diseases [43][44][45], so we speculate that lnc-DC may play important roles in diseases such as refractory wounds, skin squamous cell carcinoma and psoriasis, but more detailed and systematic experiments are needed to verify it. ...
... During epithelialization, keratinocytes, which constitute the outermost layer of the skin, migrate to the top of the newly formed granulation tissue, and the GRHL3 (grainyhead like 3)/FSCN1 (fascin actin-bundling protein 1)/E-cadherin pathway is activated in the cells of the injured skin (Figure 4), and these keratinocytes meet with keratinocytes from the contralateral edge, these keratinocytes meet migrating keratinocytes from the opposite edge, and the wound closes. 105 Growth factor-induced re-epithelialization occurs when epidermal growth factor (EGF) is generated by platelets, keratinocytes, and adipocytes, respectively, meanwhile Anti-inflammatory macrophages are activated as well as pro-repairing macrophages. 107 As a "find me" signal, S1P inhibits keratinocyte proliferation, induces keratinocyte differentiation and migration, and plays a role in wound healing. ...
... During remodeling, F I G U R E 4 (A-G) GRHL3 upregulates FSCN1 in migrating pre-wound keratinocytes, and FSCN1 increases keratinocyte migration, which is important for wound re-epithelialization. Reproduced under terms of the CC-BY license. 105 Copyright 2021, The Authors, published by the American Society for Clinical Investigation. (H) In addition, S1P signaling molecules can inhibit keratinocyte proliferation and induce keratinocyte differentiation and migration to promote wound healing. ...
Effectively eliminating apoptotic cells is precisely controlled by a variety of signaling molecules and a phagocytic effect known as efferocytosis. Abnormalities in efferocytosis may bring about the development of chronic conditions, including angiocardiopathy, chronic inflammatory diseases and autoimmune diseases. During wound healing, failure of efferocytosis leads to the collection of apoptosis, the release of necrotic material and chronic wounds that are difficult to heal. In addition to the traditional phagocytes‐macrophages, other important cell species including dendritic cells, neutrophils, vascular endothelial cells, fibroblasts and keratinocytes contribute to wounding healing. This review summarizes how efferocytosis‐mediated immunomodulation plays a repair‐promoting role in wound healing, providing new insights for patients suffering various cutaneous wounds. image
... [1][2][3][4][5][6] Traditionally, studies on cell migration have primarily focused on understanding the regulatory mechanisms driven by chemical signals. [7][8][9][10][11][12][13] However, recent research has shed light on the cooperative role of physical cues present in the cellular environment in conjunction with chemical signals. [14][15][16][17][18] For instance, cells can perceive and migrate along gradients of substrate stiffness, [14] exhibiting collective movement through physical interactions, [15,16] and undergoing polarization within microconfinement. ...
Efficient cell migration is crucial for the functioning of biological processes, e.g., morphogenesis, wound healing, and cancer metastasis. In this study, we monitored the migratory behavior of 3D fibroblast clusters using live cell microscopy, and found that crowded environment affects cell migration, i.e., crowding leads to directional migration at the cluster's periphery. The number of cell layers being stacked during seeding determines the directional-to-random transition. Intriguingly, the migratory behavior of cell clusters resembles the dispersion dynamics of clouds of passive particles, indicating that the biological process is driven by physical effects (e.g., entropy) rather than cell communication. Our findings highlight the role of intrinsic physical characteristics, such as crowding, in regulating biological behavior, and suggest new therapeutic approaches targeting at cancer metastasis.
... Re-epithelialization is an important step in the wound healing process. During re-epithelialization, KCs, the major cellular constituent of the epidermis, migrate to the top of the granulation tissue, and meet KCs from the other edge of the wound, thereby closing the wound (30). In diabetic wounds, excessive apoptosis of KCs is a key cellular event in wound closure. ...
Diabetic wounds are characterized by delayed and incomplete healing. As one of the most common complications of diabetes, diabetic wounds can be fatal in some cases. Programmed cell death (PCD) is an active and ordered cell death mode determined by genes, including apoptosis, autophagy, pyroptosis, necroptosis, ferroptosis, and cuproptosis. It is currently believed that PCD plays a crucial role in diabetic wound healing. Diabetic hyperglycemic environments can lead to abnormal PCD in various cells during healing processes, thereby affecting the activity and function of cells and interfering with diabetic wound healing. Therefore, this review focuses on the new roles and mechanisms of PCD in diabetic wound healing. Moreover, the challenges and perspectives related to PCD in diabetic wound healing are presented, which will bring new insights to improve diabetic wound healing.
... Rights reserved. [191][192][193][194][195][196] [191] [ 155,197] [ [198][199][200] [201] Comparison of the major phenotypic characteristics of several mouse gene knockout or knockin models displaying similar phenotypes as RIPK4-deficient mice. The similarity in phenotypes may indicate several of these genes act in common signaling pathways. ...
The receptor interacting protein kinases (RIPK) are a family of serine/threonine kinases that are involved in the integration of various stress signals. In response to several extracellular and/or intracellular stimuli, RIP kinases engage signaling cascades leading to the activation of NF-κB and mitogen-activated protein kinases, cell death, inflammation, differentiation and Wnt signaling and can have kinase-dependent and kinase-independent functions. Although it was previously suggested that seven RIPKs are part of the RIPK family, phylogenetic analysis indicates that there are only five genuine RIPKs. RIPK1 and RIPK3 are mainly involved in controlling and executing necroptosis in keratinocytes, while RIPK4 controls proliferation and differentiation of keratinocytes and thereby can act as a tumor suppressor in skin. Therefore, in this review we summarize and discuss the functions of RIPKs in skin homeostasis as well as the signaling pathways involved.
... In vitro studies of migrating epithelial cell sheets have shown that adherens junctions are essential for cells to migrate collectively (18), and that E-cadherin is tightly regulated in collective cell migration during embryonal development, tumor metastasis, and wound healing (19,20). In an acute wound model using human xenografts, a downregulation of E-cadherin is observed in the wound front (21). AQPs are pivotal players in collective cell migration and it is thought that the facilitated water permeability aids in cell shape changes required for cell migration in restricted 3D environments (22). ...
... Like adherens junctions, tight junctions are also essential in collective cell migration in skin wound healing (30). Loss of tight junction proteins claudin-1 and occludin has been observed in wound edges of human chronic wounds, whereas they were present found to be almost absent in the wound front in an acute wound model using human skin xenograft (21). These observations are made in the wound front with intact epidermis, without any observations from the specific wound edge (transition from intact epidermis to loss of epidermal cells). ...
Chronic wounds are defined as wounds that fail to proceed through the normal phases of wound healing; a complex process involving different dynamic events including migration of keratinocytes in the epidermis. Chronic wounds are estimated to affect 1-2% of the human population worldwide and are a major socioeconomic burden. The prevalence of chronic wounds is expected to increase with the rising number of elderly and patients with diabetes and obesity, who are at high risk of developing chronic wounds. Since E-cadherin and the water channel aquaporin-3 are important for both skin function and cell migration, and aquaporin-3 is furthermore involved in wound healing of the skin demonstrated by impaired wound healing in aquaporin-3-null mice, we hypothesized that E-cadherin and aquaporin-3 expression may be dysregulated in chronic wounds. Therefore, we investigated the expression of E-cadherin and aquaporin-3 in biopsies from the edges of chronic wounds from human patients. This was accomplished by immunohistochemical stainings of E-cadherin and aquaporin-3 on serial sections followed by qualitative evaluation of staining patterns, which revealed low expression of both E-cadherin and aquaporin-3 at the wound edge. Future studies are needed to reveal if this downregulation is associated with the pathophysiology of chronic wounds.
The clinical treatment of chronic diabetic wounds is a long-standing thorny issue. Strategies targeting the diabetic micro-environment have been developed to promote wound healing. However, it remains challenging to reverse...
Tissue development is mediated by a combination of mechanical and biological signals. Currently, there are many reports on biological signals regulating repair. However, insufficient attention is paid to the process of mechanical regulation, especially the active mechanical regulation in vivo, which has not been realized. Herein, a novel dynamically regulated repair system for both in vitro and in vivo applications is developed, which utilizes magnetic nanoparticles as non‐contact actuators to activate hydrogels. The magnetic hydrogel can be periodically activated and deformed to different amplitudes by a dynamic magnetic system. An in vitro skin model is used to explore the impact of different dynamic stimuli on cellular mechano‐transduction signal activation and cell differentiation. Specifically, the effect of mechanical stimulation on the phenotypic transition of fibroblasts to myofibroblasts is investigated. Furthermore, in vivo results verify that dynamic massage can simulate and enhance the traction effect in skin defects, thereby accelerating the wound healing process by promoting re‐epithelialization and mediating dermal contraction.
Objective:
Throughout our existence, the skin senses and analyses the mechanical forces imposed by the environment. In response to these environmental forces, skin can deform itself and achieve a biological response. The subsequent cutaneous plasticity emerges from mechanical properties arising from the collective action of the skin cells, particularly keratinocytes, that govern the tensile strength via cell-to-cell adhesions and via cell-matrix adhesion structures. In addition to serving as force-bearing entities, keratinocytes respond to forces by activating signaling pathways to control their own fate and function. To detect and adapt to mechanical signals, keratinocytes possess a panel of sensory receptors and junctional intercellular structures. Mechanically activated ion channel Piezo1 has been described as a force sensor and as being involved in pleasant touch perception. In this study, relationships between Piezo1 modulation and oxytocin synthesis were investigated.
Methods:
The expression of Piezo1 in the skin was studied and was compared to the expression of TRPV1. Dooku1 antagonist and Jedi1 agonist were used to modulate Piezo1. The level of E-cadherin and oxytocin was monitored in ex vivo skin biopsies by immunodetection.
Results:
Taken together, our results illustrate the major role of mechanosensitive ion channel Piezo1 in skin barrier integrity, and in peripheral oxytocin synthesis in the skin.
Conclusion:
In conclusion, this study highlights the relationships between pleasant touch, soft touch, and local oxytocin synthesis.
