Cell Death In Keratinocytes Research Articles

Single-cell analysis of Kaposi's sarcoma-associated herpesvirus infection in three-dimensional air-liquid interface culture model.

The oral cavity is the major site for transmission of Kaposi’s sarcoma-associated herpesvirus (KSHV), but how KSHV establishes infection and replication in the oral epithelia remains unclear. Here, we report a KSHV spontaneous lytic replication model using fully differentiated, three-dimensional (3D) oral epithelial organoids at an air-liquid interface (ALI). This model revealed that KSHV infected the oral epithelia when the basal epithelial cells were exposed by damage. Unlike two-dimensional (2D) cell culture, 3D oral epithelial organoid ALI culture allowed high levels of spontaneous KSHV lytic replication, where lytically replicating cells were enriched at the superficial layer of epithelial organoid. Single cell RNA sequencing (scRNAseq) showed that KSHV infection induced drastic changes of host gene expression in infected as well as uninfected cells at the different epithelial layers, resulting in altered keratinocyte differentiation and cell death. Moreover, we identified a unique population of infected cells containing lytic gene expression at the KSHV K2-K5 gene locus and distinct host gene expression compared to latent or lytic infected cells. This study demonstrates an in vitro 3D epithelial organoid ALI culture model that recapitulates KSHV infection in the oral cavity, where KSHV undergoes the epithelial differentiation-dependent spontaneous lytic replication with a unique cell population carrying distinct viral gene expression.

The role of tumor necrosis factor-alpha (TNF-α) inhibitor in steven johnson syndrome/toxic epidermal necrolysis (SJS/TEN) management: a systematic review

Background. Steven Johnson Syndrome and Toxic Epidermal Necrolysis are dermatologic emergencies with high mortality rates characterized by extensive skin involvement. TNF-α has a role in inducing the production of granulysin that enhances keratinocyte cell death. Thus, the use of TNF-α inhibitors such as etanercept and infliximab is promising to halt the disease progression. This systematic review aims to evaluate the efficacy of TNF-α inhibitor in SJS/TEN management. Method. A systematic review using an online database was conducted based on PRISMA guidelines. Inclusion criteria were studied about the role of TNF-α inhibitor in SJS/TEN management. The exclusion criteria were a letter to the editor, commentary report, review, meta-analysis, study not used humans as a study subject, not available in full text and not in English or Bahasa Indonesia. Result. Fourteen studies are included in this systematic review consisting of ten case reports, three cohort retrospective studies, and one randomized controlled trial. TNF-α inhibitors such as etanercept and infliximab are beneficial for SJS/TEN management because they accelerate skin healing time and re-epithelization, reduce in-hospital duration and mortality rate, and minimal complications and side effects. TNF-α inhibitors should be given quickly after the disease onset, and combination with other immunosuppressive agents should be avoided to prevent the excessive immunosuppressive effect. Conclusion. TNF-α inhibitors such as etanercept and infliximab promise therapy to halt SJS/TEN progression.

416 Pyroptosis of keratinocyte in Stevens-Johnson syndrome / toxic epidermal necrolysis

Stevens Johnson syndrome (SJS)/ toxic epidermal necrolysis (TEN) are characterized by the remarkable cell death of keratinocytes. Our previous studies have revealed that keratinocyte death pathway involve apoptosis and necroptosis. However, in recent years, it has been found that there is a crosstalk between apoptosis, necroptosis and pyroptosis. Pyroptosis is an inflammatory programmed cell death characterized by caspase 1-mediated cleavage of GSDMD and the release of IL-1b and IL-18. The aim of our study is to reveal whether pyroptosis is associated with the pathogenesis of SJS/TEN and develop specific therapeutic agents that can suppress these cell death programs in SJS/TEN. Expression of cleaved GSDMD and cleaved caspase1, which were key players of pyroptosis, were observed in the keratinocytes of SJS/TEN patients by immunofluorescent staining. To prove that pyroptosis can occur in keratinocytes, we stimulated HaCaT cells with pyroptosis inducers, LPS and nigertin, and evaluated pyroptosis reaction by live/dead staining assay, observation of morphological changes and cleaved GSDMD expression. HaCaT cells after LPS and nigertin stimulation formed large bubbles and a punctate positive reaction of cleaved GSDMD on the cell membrane, occurring as a result of membrane pore-forming activity of the gasdermin-N domain. Pyroptosis occurred even after stimulating HaCaT cells with SJS serum. Our study showed that pyroptosis occurred in keratinocytes of SJS/TEN patients and substances that induce pyroptosis to keratinocyte were present in the serum of the patients with SJS/TEN. We have investigated SJS/TEN-specifically elevated proteins by selected reaction monitoring. We are currently exploring the substances that induce pyroptosis. It is expected that understanding the mechanisms of keratinocyte death in SJS/TEN lead us to develop a specific therapeutic drug that can suppress cell death in SJS/TEN.

409 Corneoptosis, functional keratinocyte cell death, is tightly associated with spaciotemporal dynamics of epidermal tight junctions

Epidermis has two physical barrier components, stratum corneum (SC) and tight junction (TJ). Formation of SC or cornification is initiated by the cell death of the top layer (SG1) of stratum granulosum, termed ‘corneoptosis’ after prolonged intracellular Ca2+([Ca2+]i) elevation (∼60min), followed by rapid intracellular acidification. However, the exact timing of the onset of corneoptorsis remains to be clarified. In this study, we aimed to determine the spatiotemporal relationship between the turnover of TJs and corneoptosis. We performed intravital confocal microscopic imaging of mouse ear co-expressing ZO-1-Venus (TJ strand marker) and GCaMP3 (intracellular Ca2+ marker). Hoechst 33342 was intradermally injected to monitor the change of nuclear pH. Consistent with our previous observation, ZO-1-Venus signals showed two TJ strand-pattens in upper SG. According to the TJ turnover, three phases are observed; TJ-phase 1 which shows single old TJ strand formed in SG2 cells; TJ-phase 2 which shows double TJ strands; TJ-phase 3 which shows new single TJ strand. Time lapse imaging analysis revealed that the elevation of GCaMP3 signal was observed only in TJ-phase 2 which shows double TJs, but not in other TJ-phases. The elevation of [Ca2+]i lasted for approximately 60min. Subsequently, intracellular acidification was induced in these SG1 cells as detected by both the decrease of GCaMP3 signal and the increase of Hoechst 33342 signals. Simultaneously, ‘upper larger TJ strand’ of double-TJ strand in SG1 cells disappeared, resulting in change from TJ-phase 2 to TJ-phase 3. These findings indicated that the onset of corneoptosis is tightly associated with the spaciotemporal dynamics of TJs.

Impaired LEF1 Activation Accelerates iPSC-Derived Keratinocytes Differentiation in Hutchinson-Gilford Progeria Syndrome.

Hutchinson–Gilford progeria syndrome (HGPS) is a detrimental premature aging disease caused by a point mutation in the human LMNA gene. This mutation results in the abnormal accumulation of a truncated pre-lamin A protein called progerin. Among the drastically accelerated signs of aging in HGPS patients, severe skin phenotypes such as alopecia and sclerotic skins always develop with the disease progression. Here, we studied the HGPS molecular mechanisms focusing on early skin development by differentiating patient-derived induced pluripotent stem cells (iPSCs) to a keratinocyte lineage. Interestingly, HGPS iPSCs showed an accelerated commitment to the keratinocyte lineage than the normal control. To study potential signaling pathways that accelerated skin development in HGPS, we investigated the WNT pathway components during HGPS iPSCs-keratinocytes induction. Surprisingly, despite the unaffected β-catenin activity, the expression of a critical WNT transcription factor LEF1 was diminished from an early stage in HGPS iPSCs-keratinocytes differentiation. A chromatin immunoprecipitation (ChIP) experiment further revealed strong bindings of LEF1 to the early-stage epithelial developmental markers K8 and K18 and that the LEF1 silencing by siRNA down-regulates the K8/K18 transcription. During the iPSCs-keratinocytes differentiation, correction of HGPS mutation by Adenine base editing (ABE), while in a partial level, rescued the phenotypes for accelerated keratinocyte lineage-commitment. ABE also reduced the cell death in HGPS iPSCs-derived keratinocytes. These findings brought new insight into the molecular basis and therapeutic application for the skin abnormalities in HGPS.

Stratum corneum as polymer sheet: concept and cornification processes.

The skin barrier protects our body from external insults and water loss through a specialized layer, the stratum corneum. The stratum corneum, an accumulation of dead keratinocytes (corneocytes), comprises lipids and supporting cell bodies. We propose a framework of lipid-filled polymer sheet of corneocytes, a unique structure that achieves flexibility and robustness, updating the rigid image of the historical bricks-and-mortar model. The polymerization of polymer sheet (cornification) by cell death of keratinocytes (corneoptosis) is delicately and dynamically controlled by cytoplasmic calcium ion and pH. Understanding the structure and formation of the stratum corneum can lead to better treatments for skin diseases and a better understanding of the evolution of the stratum corneum.

Increase in the Intracellular Bulk Water Content in the Early Phase of Cell Death of Keratinocytes, Corneoptosis, as Revealed by 65 GHz Near-Field CMOS Dielectric Sensor.

While bulk water and hydration water coexist in cells to support the expression of biological macromolecules, how the dynamics of water molecules, which have long been only a minor role in molecular biology research, relate to changes in cellular states such as cell death has hardly been explored so far due to the lack of evaluation techniques. In this study, we developed a high-precision measurement system that can discriminate bulk water content changes of ±0.02% (0.2 mg/cm3) with single-cell-level spatial resolution based on a near-field CMOS dielectric sensor operating at 65 GHz. We applied this system to evaluate the temporal changes in the bulk water content during the cell death process of keratinocytes, called corneoptosis, using isolated SG1 (first layer of stratum granulosum) cells in vitro. A significant irreversible increase in the bulk water content was observed approximately 1 h before membrane disruption during corneoptosis, which starts with cytoplasmic high Ca2+ signal. These findings suggest that the calcium flux may have a role in triggering the increase in the bulk water content in SG1 cells. Thus, our near-field CMOS dielectric sensor provides a valuable tool to dissect the involvement of water molecules in the various events that occur in the cell.

Trehalose Protects Keratinocytes against Ultraviolet B Radiation by Activating Autophagy via Regulating TIMP3 and ATG9A.

Trehalose, a natural disaccharide, is synthesized by many organisms when cells are exposed to stressful stimuli. On the basis of its ability to modulate autophagy, trehalose has been considered an innovative drug for ameliorating many diseases, but its molecular mechanism is not well described. Previous findings demonstrated that trehalose plays a photoprotective role against ultraviolet (UV) B-induced damage through autophagy induction in keratinocytes. In this study, coimmunoprecipitation, label-free quantitative proteomic and parallel reaction monitoring, and western blot analysis demonstrated that trehalose promotes the interaction between tissue inhibitor of metalloproteinase (TIMP) 3 and Beclin1. Western blot and immunofluorescence staining analysis suggested that trehalose increases ATG9A localization in lysosomes and decreases its localization in the endoplasmic reticulum. Furthermore, in the presence or absence of UVB radiation, we evaluated the influence of TIMP3 and ATG9A small interfering RNA (siRNA) on the effect of trehalose on autophagy, cell death, migration, or interleukin-8 expression in keratinocytes, including HaCaT, A431, and human epidermal keratinocytes. The results revealed that in HaCaT cells, TIMP3 and ATG9A siRNA resulted in attenuation of trehalose-induced autophagy and inhibited cell death. In A431 cells, TIMP3 and ATG9A siRNA led to attenuation of trehalose-induced autophagy and cell death and inhibited migration. In human epidermal keratinocytes, trehalose-induced autophagy and inhibition of the interleukin-8 expression were blocked by ATG9A but not TIMP3 siRNA. In addition, the results of quantitative real-time PCR and immunohistochemistry analysis demonstrated the abnormal expression of TIMP3 and ATG9A in actinic keratosis and cutaneous squamous cell carcinoma skin tissues. These findings suggest the protective effects of trehalose in normal keratinocytes and its inhibitory effects on cancerous keratinocytes, possibly mediated by activation of autophagy and regulation of TIMP3 and ATG9A, providing the mechanistic basis for the potential use of trehalose in the prevention or treatment of UVB-induced skin diseases.

Immune mechanisms in oral lichen planus.

Oral lichen planus (OLP) is a T cell-mediated inflammatory disease of the oral mucosa that has been extensively researched over many years but as yet the mechanisms of pathogenesis are still not fully understood. Whilst the specific aetiological factors driving OLP remain ambiguous, evidence points to the development of a chronic, dysregulated immune response to OLP-mediating antigens presented by innate immune cells and oral keratinocytes leading to increased cytokine, chemokine and adhesion molecule expression. These molecules recruit T cells and mast cells to the diseased site and orchestrate a complex interplay between cells that culminates in keratinocyte cell death, mucosal basement membrane destruction and long-term chronicity of the disease. The main lymphocytes involved are thought to be CD8+ cytotoxic and CD4+ Th1 polarised T cells although recent evidence indicates the involvement of other Th subsets such as Th9, Th17 and Tregs, suggesting that a more complex immune cell relationship exists during the disease process. This review provides an overview of the immune mechanisms at play in OLP pathogenesis with particular emphasis on the role of the different Th subsets and how these recent discoveries may guide research towards identifying potential therapeutic targets.

Single-cell multi-omic approaches define common molecular and cellular signals of dominant antigen-driven cells at the site of drug-induced Stevens Johnson Syndrome and Toxic Epidermal Necrolysis (SJS/TEN) tissue damage

Human leukocyte antigen (HLA)-restricted CD8+ T-cells expressing dominant T-cell receptor (TCR) clonotypes are recently implicated drivers of keratinocyte cell death, cutaneous blistering, and mortality in drug-induced Stevens Johnson Syndrome/Toxic epidermal necrolysis (SJS/TEN). Signatures of these effector population(s) remain undefined in affected tissue but hold utility for early diagnosis and targeted therapy.

Hydrogen-Rich Water Prevents Dehydration-Induced Cellular Oxidative Stress and Cell Death in Human Skin Keratinocytes

Hypohydration is linked to increased risk of a variety of diseases and can be life-threatening, especially in elderly populations. Dehydration induces cellular damage partially through the production of reactive oxygen species (ROS) in cells, tissues and organs. Hydrogen molecules are known to convert ROS to harmless water. Therefore, theoretically hydrogen-rich water (HW) might eliminate dehydration-induced ROS and reverse its harmful effects in cells. In this in vitro study, we demonstrated that air-drying for 5 min could induce ROS generation in both nucleus and cytoplasm of human keratinocytes HaCaT as quantified by CellROX® Green/Orange reagents (Thermo Fisher Scientific, Waltham, Massachusetts, U.S.), respectively. Conversely, when the air-drying time was increased to 10 and 20 min, HaCaT cells lost the ability to produce ROS. Scanning electron microscopic (SEM) images showed that 10 min air-drying could induce severe membrane damage in HaCaT cells. PrestoBlue assay showed that, when HaCaT cells were air-dried for 20 min, cell viability was decreased to 27.6% of the control cells 48 h later. However, once HaCaT cells were pretreated with HW-prepared media, dehydration-induced intracellular ROS, cell membrane damage and cell death were significantly reduced as compared with double distilled water (DDW) under the same conditions. In conclusion, our data suggested that HW can decrease dehydration-induced harmful effects in human cells partially through its antioxidant capacity.

Hyperoside and Quercitrin in Houttuynia cordata Extract Attenuate UVB-Induced Human Keratinocyte Cell Damage and Oxidative Stress via Modulation of MAPKs and Akt Signaling Pathway.

Ultraviolet radiation is a major environmental harmful factor on human skin. In this paper, we investigate the potential mechanism of Houttuynia cordata extract on UVB-induced HaCaT keratinocyte cell death and inflammation. We found that Houttuynia cordata ethyl acetate extract fraction (HC-EA) protected against UVB-induced cell damage. The HPLC results indicate that quercitrin and hyperoside are the major polyphenolics in HC-EA and are responsible for providing protection against UVB-induced cell death. These responses were associated with the regulation of caspase-9 and caspase-3 activation, which rescued HaCaT cells from UVB-induced apoptosis. In addition, HC-EA, quercitrin, and hyperoside attenuated UVB-induced inflammatory mediators, including IL-6, IL-8, COX-2, and iNOS. Furthermore, the treatment of cells with HC-EA and its active compounds abolished intracellular ROS and increased levels of heme oxygenase-1 and superoxide dismutase. UVB-induced ROS production mediated Akt and mitogen activated protein kinases (MAPKs) pathways, including p38, ERK, and JNK. Our results show HC-EA, quercitrin, and hyperoside decreased UVB-induced p38 and JNK phosphorylation, while increasing ERK and Akt phosphorylation. MAPKs and Akt mediated cell survival and death were confirmed by specific inhibitors to Akt and MAPKs. Thus, HC-EA, which contains quercitrin and hyperoside, protected keratinocyte from UVB-induced oxidative damage and inflammation through the modulation of MAPKs and Akt signaling.

Pannexin 2 is expressed in murine skin and promotes UVB-induced apoptosis of keratinocytes.

Pannexins (PANX) are a family of three channel-forming membrane glycoproteins expressed in the skin. Previous studies have focused on the role of PANX1 and PANX3 in the regulation of cellular functions in skin cells while PANX2, the largest member of this protein family, has not been investigated. In the current study, we explored the temporal PANX2 expression in murine skin and found that one Panx2 splice variant (Panx2-202) tends to be more abundant at the protein level and is continuously expressed in developed skin. PANX2 was detected in the suprabasal layers of the mouse epidermis and up-regulated in an in vitro model of rat epidermal keratinocyte differentiation. Furthermore, we show that in apoptotic rat keratinocytes, upon UV light B (UVB)-induced caspase-3/7 activation, ectopically overexpressed PANX2 is cleaved in its C-terminal domain at the D416 residue without increasing the apoptotic rate measured by caspase-3/7 activation. Notably, CRISPR-Cas9 mediated genetic deletion of rat Panx2 delays but does not impair caspase-3/7 activation and cytotoxicity in UVB-irradiated keratinocytes. We propose that endogenous PANX2 expression in keratinocytes promotes cell death after UVB insult and may contribute to skin homeostasis.

Cosmeceutical Potentials of Grammatophyllum speciosum Extracts: Anti-Inflammations and Anti-Collagenase Activities with Phytochemical Profile Analysis Using an Untargeted Metabolomics Approach

Grammatophyllum speciosum is the largest orchid species and a well-known traditional medicinal plant. Due to skin aging, natural products that inhibit this process can attract the attention of consumers and scientists because radical-scavenging activity, collagenase inhibition, and inflammatory suppression are valuable in dermatological applications. This study investigated the phytochemicals in G. speciosum leaves extracts that have cosmeceutical potentials, including radical-scavenging, anticollagenase, and anti-inflammatory abilities. G. speciosum leaves were extracted using water-based extraction methods. High-resolution mass spectrometry was used to identify the phytochemicals in the extracts. Fibroblast and keratinocyte cell cytotoxicity was determined. Antioxidant abilities were measured using DPPH and ABTS assays. The effect of the extracts on nitric oxide (NO) in macrophage cells was investigated. ELISA of the collagenase enzyme was determined. A total of 721 annotated metabolites were identified in the extracts. Vitexin and orientin were the most abundant metabolites. Cell viability was >80% in both cell lines when the extract concentration was <1 mg/mL. The IC50 values for DPPH and ABTS were 56 and 117 μg/mL, respectively. Furthermore, the extracts revealed that NO and collagenase activity were suppressed by 42% and 23%, respectively. The extracts can suppress ROS, inflammatory, and collagenase activities without causing fibroblast and keratinocyte cell death. Thus, this study provides information on metabolites in G. speciosum leaves, which is promising as cosmeceuticals or pharmaceuticals with anti-inflammatory and anti-collagenase activities.

NAMPT-derived NAD+ fuels PARP1 to promote skin inflammation through parthanatos cell death.

Several studies have revealed a correlation between chronic inflammation and nicotinamide adenine dinucleotide (NAD+) metabolism, but the precise mechanism involved is unknown. Here, we report that the genetic and pharmacological inhibition of nicotinamide phosphoribosyltransferase (Nampt), the rate-limiting enzyme in the salvage pathway of NAD+ biosynthesis, reduced oxidative stress, inflammation, and keratinocyte DNA damage, hyperproliferation, and cell death in zebrafish models of chronic skin inflammation, while all these effects were reversed by NAD+ supplementation. Similarly, genetic and pharmacological inhibition of poly(ADP-ribose) (PAR) polymerase 1 (Parp1), overexpression of PAR glycohydrolase, inhibition of apoptosis-inducing factor 1, inhibition of NADPH oxidases, and reactive oxygen species (ROS) scavenging all phenocopied the effects of Nampt inhibition. Pharmacological inhibition of NADPH oxidases/NAMPT/PARP/AIFM1 axis decreased the expression of pathology-associated genes in human organotypic 3D skin models of psoriasis. Consistently, an aberrant induction of NAMPT and PARP activity, together with AIFM1 nuclear translocation, was observed in lesional skin from psoriasis patients. In conclusion, hyperactivation of PARP1 in response to ROS-induced DNA damage, fueled by NAMPT-derived NAD+, mediates skin inflammation through parthanatos cell death.

Insights into the activation of oral keratinocyte cell death by Candida albicans and Staphylococcus aureus biofilms

Polymicrobial biofilms comprising Candida albicans and Staphylococcus aureus can increase the frequency and severity of oral diseases. This study assessed oral keratinocyte cell death, apoptosis and/or necrosis, promoted by soluble factors from single and dual biofilms of S. aureus and C. albicans. The soluble factors were obtained from the 16-h biofilm growth media. Cell viability was assessed by MTT and cell membrane damage by LDH. SEM was used for morphology changes. Assessment of apoptosis and necrosis was performed using annexin V and propidium iodide and caspases -2, -3, -6, -8 and -9. Statistical analysis was conducted with ANOVA and Tukey tests (α = 5%). Dual biofilms promoted the greatest harmful effect on oral cells, with a viability rate of 31.76%, damage to cell membranes and LDH released. Dual biofilms also induced higher percentages of necrotic cells (24.95%). Apoptosis was associated with caspases -2, -3, -6 and -8 activation.

OTULIN maintains skin homeostasis by controlling keratinocyte death and stem cell identity

OTULIN is a deubiquitinase that specifically cleaves linear ubiquitin chains. Here we demonstrate that the ablation of Otulin selectively in keratinocytes causes inflammatory skin lesions that develop into verrucous carcinomas. Genetic deletion of Tnfr1, knockin expression of kinase-inactive Ripk1 or keratinocyte-specific deletion of Fadd and Mlkl completely rescues mice with OTULIN deficiency from dermatitis and tumorigenesis, thereby identifying keratinocyte cell death as the driving force for inflammation. Single-cell RNA-sequencing comparing non-lesional and lesional skin reveals changes in epidermal stem cell identity in OTULIN-deficient keratinocytes prior to substantial immune cell infiltration. Keratinocytes lacking OTULIN display a type-1 interferon and IL-1β response signature, and genetic or pharmacologic inhibition of these cytokines partially inhibits skin inflammation. Finally, expression of a hypomorphic mutant Otulin allele, previously shown to cause OTULIN-related autoinflammatory syndrome in humans, induces a similar inflammatory phenotype, thus supporting the importance of OTULIN for restraining skin inflammation and maintaining immune homeostasis.

206 OTULIN maintains skin homeostasis by controlling keratinocyte death and stem cell identity

OTULIN is a deubiquitinase that specifically cleaves linear ubiquitin chains. We demonstrate that ablation of OTULIN selectively in keratinocytes, results in the appearance of inflammatory skin lesions that develop into verrucous carcinomas. Genetic ablation of TNFR1, knockin expression of kinase-inactive RIPK1 or keratinocyte-specific deletion of FADD and MLKL completely rescues mice with OTULIN deficiency from dermatitis and tumorigenesis, identifying keratinocyte cell death as the driving force for inflammation.

S2k guideline: Diagnosis and management of cutaneous lupus erythematosus - Part1: Classification, diagnosis, prevention, activity scores.

S2k guideline: Diagnosis and management of cutaneous lupus erythematosus - Part1: Classification, diagnosis, prevention, activity scores.

Burn Injury: Mechanisms of Keratinocyte Cell Death

Cutaneous burn injury is associated with epidermal loss in the zone of coagulation zone and delayed tissue loss in the zone of stasis. Thus, thermal stress can trigger both necrosis and regulated cell death (RCD) or apoptosis. Experimental in vitro and in vivo work has clearly demonstrated apoptotic events of thermally injured keratinocytes that are accompanied by morphological and biochemical markers of regulated cell death. However, in vivo data for the different pathways of regulated cell death are sparse. In vitro experiments with heat-stressed human keratinocytes have demonstrated death receptor involvement (extrinsic apoptosis), calcium influx, and disruption of mitochondrial membrane potential (intrinsic apoptosis) in regulated cell death. In addition, caspase-independent pathways have been suggested in regulated cell death. Keratinocyte heat stress leads to reduced proliferation, possibly as a result of reduced keratinocyte adhesion (anoikis) or oncogene involvement. Understanding the underlying mechanisms of RCD and the skin’s responses to thermal stress may lead to improved strategies for treating cutaneous burn trauma.