Salicylate induces epithelial actin reorganization via activation of the AMP-activated protein kinase and promotes wound healing and contraction in mice.

Wounds that occur in adults form scars due to fibrosis, whereas those in embryos regenerate. If wound healing in embryos is mimicked in adults, scarring can be reduced. We found that mouse fetuses could regenerate tissues up to embryonic day (E) 13, but visible scars remained thereafter. This regeneration pattern requires actin cable formation at the epithelial wound margin via activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK). Here, we investigated whether the AMPK-activating effect of salicylate, an anti-inflammatory drug, promotes regenerative wound healing. Salicylate administration resulted in actin cable formation and complete wound regeneration in E14 fetuses, in which scarring should have normally occurred, and promoted contraction of the panniculus carnosus muscle, resulting in complete wound regeneration. In vitro, salicylate further induced actin remodeling in mouse epidermal keratinocytes in a manner dependent on cell and substrate target-specific AMPK activation and subsequent regulation of Rac1 signaling. Furthermore, salicylate promoted epithelialization, enhanced panniculus carnosus muscle contraction, and inhibited scar formation in adult mice. Administration of salicylates to wounds immediately after injury may be a novel method for preventing scarring by promoting a wound healing pattern similar to that of embryonic wounds.

Fisetin, a potential skin rejuvenation drug that eliminates senescent cells in the dermis.

Accumulation of senescent fibroblasts, chronic inflammation, and collagen remodeling due to aging-related secretory phenotypes have been hypothesized to cause age-related skin aging, which results in wrinkles and loss of skin elasticity, thus compromising appearance attractiveness. However, the rejuvenating effects of removing senescent cells from the human skin and the efficacy of related therapeutic agents remain unclear. Here, we investigated the effects of fisetin, a potential anti-aging component found in various edible fruits and vegetables, on senescent human dermal fibroblasts (HDFs) and aging human skin. Senescence was induced in primary HDFs using long-term passaging and treatment with ionizing radiation, and cell viability was assessed after treatment with fisetin and a control component. A mouse/human chimeric model was established by subcutaneously transplanting whole skin grafts from aged individuals into nude mice, which were treated intraperitoneally with fisetin or control a component for 30 d. Skin samples were obtained and subjected to senescence-associated-beta-galactosidase staining; the extent of aging was evaluated using western blotting, reverse transcription-quantitative PCR, and histological analysis. Fisetin selectively eliminated senescent dermal fibroblasts in both senescence-induced cellular models; this effect is attributable to cell death induction by caspases 3, 8, and 9-mediated endogenous and exogenous apoptosis. Fisetin-treated senescent human skin grafts showed increased collagen density and decreased senescence-associated secretory phenotypes (SASP), including matrix metalloproteinases and interleukins. No apparent adverse events were observed. Thus, fisetin could improve skin aging through selective removal of senescent dermal fibroblasts and SASP inhibition, indicating its potential as an effective novel therapeutic agent for combating skin aging.

Cistanche deserticola Polysaccharide Reduces Inflammation and Aging Phenotypes in the Dermal Fibroblasts through the Activation of the NRF2/HO-1 Pathway.

Dermal fibroblasts maintain the skin homeostasis by interacting with the epidermis and extracellular matrix. Their senescence contributes to functional defects in the skin related to aging. Therefore, there is an urgent need for novel therapeutic agents that could inhibit fibroblast senescence. In this study, we investigated the effects of Cistanche deserticola polysaccharide (CDP), a natural anti-inflammatory component, on the progression of senescence in human dermal fibroblasts. Normal human dermal fibroblasts (NHDFs) were cultured in passages, and highly senescent cells were selected as senescent cells. CDP treatment increased the cell proliferation in senescent NHDFs and decreased the proportion of senescence-associated-ß-galactosidase-positive cells. The treatment suppressed the senescence-related secretory phenotype, and reactive oxygen species (ROS) production was reduced, alleviating H2O2-induced oxidative stress. CDP mitigated ROS formation via the nuclear factor erythroid 2-related factor/heme oxygenase-1 pathway in senescent cells and was involved in the suppression of upstream p-extracellular signal-regulated kinase. These results indicate that CDP is an antioxidant that can alleviate age-related inflammation and may be a useful compound for skin anti-aging.

Usefulness of Harmonic ACE+7 Scalpel in Breast Reconstruction with Extended Latissimus Dorsi Flap: An Open-label Single Institution Pilot Study.

The extended latissimus dorsi (ELD) flap is a safe and aesthetically acceptable method to reconstruct small to medium-sized breasts. However, the long time required for flap elevation and intraoperative bleeding contributes to various postoperative complications. We investigated the use of alternative devices, such as the Harmonic ACE+7, which has a long arm that can help simultaneously detach and seal tissues to prevent such complications. Methods: We compared 27 patients who underwent breast reconstruction with the ELD flap using the Harmonic ACE +7 scalpel, and 28 patients who underwent breast reconstruction using an electrocautery scalpel, between May 2019 and March 2022. Data on patient demographics, surgery, and postoperative complications were collected. Surgical outcomes were compared between electrocautery (EC) and Harmonic ACE+7 (HA) groups. Results: The median age of the patients was 50.2 years. The patient demographics between the groups did not show significant differences. Flap necrosis and hematomas did not occur, and seroma was the major postoperative complication (65.7% in the EC group and 70% in the HA group). The time required for flap elevation was significantly shorter in the HA group than in the EC group (286.0 minutes and 179.0 minutes, respectively). Blood loss reduced significantly in the HA and EC groups (138.5 mL and 78.2 mL, respectively). Moreover, decreased drainage was observed for the breast area. There were no significant differences in other end points. Conclusion: In breast reconstruction with ELD flaps, using the Harmonic ACE+7 can help reduce the rate of seroma, operative time, and intraoperative bleeding without further disadvantages.

Identification of resibufogenin, a component of toad venom, as a novel senolytic compound in vitro and for potential skin rejuvenation in male mice.

Senescent cells that accumulate with age have been shown to contribute to age-related diseases and organ dysfunction and have attracted attention as a target for anti-aging therapy. In particular, the use of senescent cell-depleting agents, or senolytics, has been shown to improve the aging phenotype in animal models. Since senescence has been implicated in the skin, particularly in fibroblasts, this study used aged human skin fibroblasts to investigate the effects of resibufogenin. A component of the traditional Chinese medicine toad venom, resibufogenin was investigated for senolytic and/or senomorphic activity. We found that the compound selectively caused senescent cell death without affecting proliferating cells, with a marked effect on the suppression of the senescence-associated secretory phenotype. We also found that resibufogenin causes senescent cell death by inducing a caspase-3-mediated apoptotic program. Administration of resibufogenin to aging mice resulted in an increase in dermal collagen density and subcutaneous fat, improving the phenotype of aging skin. In other words, resibufogenin ameliorates skin aging through selective induction of senescent cell apoptosis without affecting non-aged cells. This traditional compound may have potential therapeutic benefits in skin aging characterized by senescent cell accumulation.

Development of a Novel Senolysis Approach Targeting the Senescent Fibroblast Marker HTR2A via Antibody-Dependent Cellular Cytotoxicity.

Abnormal remodeling of collagen and extracellular matrix caused by the accumulation of senescent fibroblasts in the dermis is the most likely cause of skin aging. Therefore, the application of "senolysis," in which only senescent cells are cleared from the body, has a potential in the development of antiaging treatments for skin. However, markers that label senescent fibroblasts only reflect the state of senescence, and it is important to develop markers as therapeutic targets to aid senolysis application. We investigated the potential of serotonin 2A receptor (HTR2A), which is involved in melanin production in response to ultraviolet light, as a senescent cell marker. The results showed that HTR2A is upregulated in aging dermal fibroblasts but is expressed at low levels in proliferating young cells. Flow cytometry demonstrated the presence of many HTR2A-positive cells in the aging cell population and few in the young cells. Furthermore, antibody-dependent cytotoxicity assays revealed that HTR2A preferentially sensitizes senescent fibroblasts and specifically damages only senescent cells by natural killer cells that recognize it. In conclusion, selective labeling of the novel senescent cell marker, HTR2A, could preferentially eliminate senescent cells and may contribute to the future development of novel skin senolysis approaches.

Compound 13 Promotes Epidermal Healing in Mouse Fetuses via Activation of AMPK.

Unlike adults, early developing fetuses can completely regenerate tissue, and replicating this could lead to the development of treatments to reduce scarring. Mice epidermal structures, including wound healing patterns, are regenerated until embryonic day (E) 13, leaving visible scars thereafter. These patterns require actin cable formation at the epithelial wound margin through AMP-activated protein kinase (AMPK) activation. We aimed to investigate whether the administration of compound 13 (C13), a recently discovered AMPK activator, to the wound could reproduce this actin remodeling and skin regeneration pattern through its AMPK activating effect. The C13 administration resulted in partial formations of actin cables, which would normally result in scarring, and scar reduction during the healing of full-layer skin defects that occurred in E14 and E15 fetuses. Furthermore, C13 was found to cause AMPK activation in these embryonic mouse epidermal cells. Along with AMPK activation, Rac1 signaling, which is involved in leaflet pseudopodia formation and cell migration, was suppressed in C13-treated wounds, indicating that C13 inhibits epidermal cell migration. This suggests that actin may be mobilized by C13 for cable formation. Administration of C13 to wounds may achieve wound healing similar to regenerative wound healing patterns and may be a potential candidate for new treatments to heal scars.

Corrigendum: Single-cell RNA-seq analysis reveals cellular functional heterogeneity in dermis between fibrotic and regenerative wound healing fates.

[This corrects the article DOI: 10.3389/fimmu.2022.875407.].

New Senolysis Approach via Antibody-Drug Conjugate Targeting of the Senescent Cell Marker Apolipoprotein D for Skin Rejuvenation.

Senescent cells accumulate in aging skin, causing age-related changes and a decline in functional efficiency. Therefore, senolysis, a treatment that specifically removes senescent cells and rejuvenates the skin, should be explored. We targeted apolipoprotein D (ApoD), a previously identified marker expressed on senescent dermal fibroblasts, and investigated a novel senolysis approach using a monoclonal antibody against this antigen and a secondary antibody conjugated with the cytotoxic drug pyrrolobenzodiazepine. Observations using fluorescently labeled antibodies revealed that ApoD functions as a surface marker of senescent cells and that the antibody is taken up and internalized only by such cells. The concurrent administration of the antibody with the PBD-conjugated secondary antibody specifically eliminated only senescent cells without harming young cells. The antibody-drug conjugate treatment of aging mice combined with the administration of antibodies reduced the number of senescent cells in the dermis of mice and improved the senescent skin phenotype. These results provide a proof-of-principle evaluation of a novel approach to specifically eliminate senescent cells using antibody-drug conjugates against senescent cell marker proteins. This approach is a potential candidate for clinical applications to treat pathological skin aging and related diseases via the removal of senescent cells.

Navitoclax (ABT-263) Rejuvenates Human Skin by Eliminating Senescent Dermal Fibroblasts in a Mouse/Human Chimeric Model.

Chronic senescence, such as aging, contributes to age-related tissue dysfunction and disease development. The accumulation of senescent fibroblasts and the senescence-associated secretory phenotype is particularly implicated in this process. Removal of senescent cells has been reported to prevent tissue dysfunction and to extend the life span during aging. ABT-263 (navitoclax), which inhibits antiapoptotic proteins, is a leading antiaging drug; however, its role in human skin aging is unclear. This study aimed to determine the rejuvenating effects of ABT-263 on aging skin using a human skin graft mouse model. We assessed the viability of ABT-263-treated skin fibroblasts after inducing senescence. Aged human skin was transplanted under the back skin of nude mice and injected intraperitoneally with the drug or control. Analysis of the skin specimens revealed that ABT-263 induced selective elimination of senescent dermal fibroblasts. Senescent human skin treated with ABT-263 exhibited a decrease in the number of senescent cells and in the expression of aging-related secretory phenotype molecules, such as matrix metalloproteinases and interleukins and an increase in collagen density. Our results indicate that selective removal of senescent skin cells with ABT-263 can improve the aging phenotype of human skin without side effects. ABT-263 is, thus, a novel potential therapeutic agent for skin aging.

Identification of Apolipoprotein D as a Dermal Fibroblast Marker of Human Aging for Development of Skin Rejuvenation Therapy.

The current understanding of skin aging is that senescent fibroblasts accumulate within the dermis and subcutaneous fat to cause abnormal tissue remodeling and extracellular matrix dysfunction, triggering a senescence-associated secretory phenotype (SASP). A novel therapeutic approach to prevent skin aging is to specifically eliminate senescent dermal fibroblasts; this requires the identification of specific protein markers for senescent cells. Apolipoprotein D (ApoD) is involved in lipid metabolism and antioxidant responses and is abundantly expressed in tissues affected by age-related diseases such as Alzheimer's disease and atherosclerosis. However, its behavior and role in skin aging remain unclear. In this study, we examined whether ApoD functions as a marker of aging using human dermal fibroblast aging models. In cellular senescence models induced through replicative aging and ionizing radiation exposure, ApoD expression was upregulated at the gene and protein levels and correlated with senescence-associated ß-galactosidase activity and the decreased uptake of the proliferation marker bromodeoxyuridine, which was concomitant with the upregulation of SASP genes. Furthermore, ApoD-positive cells were found to be more abundant in the aging human dermis using fluorescence flow cytometry. These results suggest that ApoD is a potential clinical marker for identifying aging dermal fibroblasts.

Cathepsin F is a potential marker for senescent human skin fibroblasts and keratinocytes associated with skin aging.

Cellular senescence is characterized by cell cycle arrest and the senescence-associated secretory phenotype (SASP) and can be triggered by a variety of stimuli, including deoxyribonucleic acid (DNA) damage, oxidative stress, and telomere exhaustion. Cellular senescence is associated with skin aging, and identification of specific markers of senescent cells is essential for development of targeted therapies. Cathepsin F (CTSF) has been implicated in dermatitis and various cancers and participates in cell immortalization through its association with Bcl family proteins. It is a candidate therapeutic target to specifically label and eliminate human skin fibroblasts and keratinocytes immortalized by aging and achieve skin rejuvenation. In this study, we investigated whether CTSF is associated with senescence in human fibroblasts and keratinocytes. In senescence models, created using replicative aging, ionizing radiation exposure, and the anticancer drug doxorubicin, various senescence markers were observed, such as senescence-associated ß-galactosidase (SA-ß-gal) activity, increased SASP gene expression, and decreased uptake of the proliferation marker BrdU. Furthermore, CTSF expression was elevated at the gene and protein levels. In addition, CTSF-positive cells were abundant in aged human epidermis and in some parts of the dermis. In the population of senescent cells with arrested division, the number of CTSF-positive cells was significantly higher than that in the proliferating cell population. These results suggest that CTSF is a candidate for therapeutic modalities targeting aging fibroblasts and keratinocytes.

Aging Fibroblasts Adversely Affect Extracellular Matrix Formation via the Senescent Humoral Factor Ependymin-Related Protein 1.

Skin senescence is characterized by a decrease in extracellular matrix and the accumulation of senescent fibroblasts in the dermis, and their secretion of humoral factors. Ependymin-related protein 1 (EPDR1) is involved in abnormal fibroblast metabolism and collagen deposition, however, its relation to skin aging is unclear. We investigated whether and how EPDR1 is involved in age-related dermal deterioration. When young dermal fibroblasts and senescent cells were co-cultured in a semipermeable membrane separation system, the young fibroblasts showed decreased gene expression of collagen type I α1 chain (COL1A1) and elastin, and increased expression of matrix metalloproteinase (MMP)1 and MMP3. Senescence marker expression and EPDR1 production were increased in the culture medium of senescent cells. Treatment of young fibroblasts with recombinant EPDR1, enhanced matrix-related gene expression and suppressed COL1A1 expression, whereas EPDR1 knockdown had the opposite effects. EPDR1 gene and protein expression were increased in aged skin, compared to young skin. These results suggest that senescent cells affect nearby fibroblasts, in part through EPDR1 secretion, and exert negative effects on matrix production in the dermis. These results may lead to the discovery of potential candidate targets in the development of skin anti-aging therapies.

Glutaminase inhibitors rejuvenate human skin via clearance of senescent cells: a study using a mouse/human chimeric model.

Skin aging caused by various endogenous and exogenous factors results in structural and functional changes to skin components. However, the role of senescent cells in skin aging has not been clarified. To elucidate the function of senescent cells in skin aging, we evaluated the effects of the glutaminase inhibitor BPTES (bis-2-(5-phenylacetamido-1, 3, 4-thiadiazol-2-yl)ethyl sulfide) on human senescent dermal fibroblasts and aged human skin. Here, primary human dermal fibroblasts (HDFs) were induced to senescence by long-term passaging, ionizing radiation, and treatment with doxorubicin, an anticancer drug. Cell viability of HDFs was assessed after BPTES treatment. A mouse/human chimeric model was created by subcutaneously transplanting whole skin grafts from aged humans into nude mice. The model was treated intraperitoneally with BPTES or vehicle for 30 days. Skin samples were collected and subjected to reverse transcription-quantitative polymerase chain reaction (RT-qPCR), western blotting, and histological analysis. BPTES selectively eliminated senescent dermal fibroblasts regardless of the method used to induce senescence; aged human skin grafts treated with BPTES exhibited increased collagen density, increased cell proliferation in the dermis, and decreased aging-related secretory phenotypes, such as matrix metalloprotease and interleukin. These effects were maintained in the grafts 1 month after termination of the treatment. In conclusion, selective removal of senescent dermal fibroblasts can improve the skin aging phenotype, indicating that BPTES may be an effective novel therapeutic agent for skin aging.

Effect of All-trans Retinoic Acid on Panniculus Carnosus Muscle Regeneration in Fetal Mouse Wound Healing.

The dermal panniculus carnosus (PC) muscle is critical for wound contraction in lower mammals and is a useful model of muscle regeneration owing to its high cellular metabolic turnover. During wound healing in mice, skin structures, including PC, are completely regenerated up to embryonic day (E) 13, but PC is only partially regenerated in fetuses or adult animals after E14. Nevertheless, the mechanisms underlying wound repair for complete regeneration in PC have not been fully elucidated. We hypothesized that retinoic acid (RA) signaling, which is involved in muscle differentiation, regulates PC regeneration. Methods: Surgical injury was induced in ICR mice on E13 and E14. RA receptor alpha (RARα) expression in tissue samples from embryos was evaluated using immunohistochemistry and reverse transcription-quantitative polymerase chain reaction. To evaluate the effects of RA on PC regeneration, beads soaked in all-trans RA (ATRA) were implanted in E13 wounds, and tissues were observed. The effects of RA on myoblast migration were evaluated using a cell migration assay. Results: During wound healing, RARα expression was enhanced at the cut surface in PCs of E13 wounds but was attenuated at the cut edge of E14 PCs. Implantation of ATRA-containing beads inhibited PC regeneration on E13 in a concentration-dependent manner. Treatment of myoblasts with ATRA inhibited cell migration. Conclusions: ATRA inhibits PC regeneration, and decreased RARα expression in wounds after E14 inhibits myoblast migration. Our findings may contribute to the development of therapies to promote complete wound regeneration, even in the muscle.

Downregulation of senescence-associated secretory phenotype by knockdown of secreted frizzled-related protein 4 contributes to the prevention of skin aging.

There is growing evidence that the appearance and texture of the skin that is altered during the aging process are considerably enhanced by the accumulation of senescent dermal fibroblasts. These senescent cells magnify aging via an inflammatory, histolytic, and senescence-associated secretory phenotype (SASP). Secreted frizzled-related protein 4 (SFRP4) was previously determined to be expressed in dermal fibroblasts of aging skin, and its increased expression has been shown to promote cellular senescence. However, its role in the SASP remains unknown. We found that SFRP4 was significantly expressed in p16ink4a-positive human skin fibroblasts and that treatment with recombinant SFRP4 promoted SASP and senescence, whereas siRNA knockdown of SFRP4 suppressed SASP. Furthermore, we found that knockdown of SFRP4 in mouse skin ameliorates age-related reduction of subcutaneous adipose tissue, panniculus carnosus muscle layer, and thinning and dispersion of collagen fibers. These findings suggest a potential candidate for the development of new skin rejuvenation therapies that suppress SASP.

Role of Wnt Signaling in Mouse Fetal Skin Wound Healing.

Wnt proteins secrete glycoproteins that are involved in various cellular processes to maintain homeostasis during development and adulthood. However, the expression and role of Wnt in wound healing have not been fully documented. Our previous studies have shown that, in an early-stage mouse fetus, no scarring occurred after cutaneous wounding, and complete regeneration was achieved. In this study, the expression and localization of Wnt proteins in a mouse fetal-wound-healing model and their associations with scar formation were analyzed. Wnt-related molecules were detected by in-situ hybridization, immunostaining, and real-time polymerase chain reaction. The results showed altered expression of Wnt-related molecules during the wound-healing process. Moreover, scar formation was suppressed by Wnt inhibitors, suggesting that Wnt signaling may be involved in wound healing and scar formation. Thus, regulation of Wnt signaling may be a possible mechanism to control scar formation.

Fibroblast Growth Factor 7 Suppresses Fibrosis and Promotes Epithelialization during Wound Healing in Mouse Fetuses.

Adult mammalian wounds leave visible scars, whereas skin wounds in developing mouse fetuses are scarless until a certain point in development when complete regeneration occurs, including the structure of the dermis and skin appendages. Analysis of the molecular mechanisms at this transition will provide clues for achieving scarless wound healing. The fibroblast growth factor (FGF) family is a key regulator of inflammation and fibrosis during wound healing. We aimed to determine the expression and role of FGF family members in fetal wound healing. ICR mouse fetuses were surgically wounded at embryonic day 13 (E13), E15, and E17. Expression of FGF family members and FGF receptor (FGFR) in tissue samples from these fetuses was evaluated using in situ hybridization and reverse transcription-quantitative polymerase chain reaction. Fgfr1 was downregulated in E15 and E17 wounds, and its ligand Fgf7 was upregulated in E13 and downregulated in E15 and E17. Recombinant FGF7 administration in E15 wounds suppressed fibrosis and promoted epithelialization at the wound site. Therefore, the expression level of Fgf7 may correlate with scar formation in late mouse embryos, and external administration of FGF7 may represent a therapeutic option to suppress fibrosis and reduce scarring.

Single-Cell RNA-seq Analysis Reveals Cellular Functional Heterogeneity in Dermis Between Fibrotic and Regenerative Wound Healing Fates.

Background: Fibrotic scars are common in both human and mouse skin wounds. However, wound-induced hair neogenesis in the murine wounding models often results in regenerative repair response. Herein, we aimed to uncover cellular functional heterogeneity in dermis between fibrotic and regenerative wound healing fates. Methods: The expression matrix of single-cell RNA sequencing (scRNA-seq) data of fibrotic and regenerative wound dermal cells was filtered, normalized, and scaled; underwent principal components analysis; and further analyzed by Uniform Manifold Approximation and Projection (UMAP) for dimension reduction with the Seurat package. Cell types were annotated, and cell-cell communications were analyzed. The core cell population myofibroblast was identified and the biological functions of ligand and receptor genes between myofibroblast and macrophage were evaluated. Specific genes between fibrotic and regenerative myofibroblast and macrophage were identified. Temporal dynamics of myofibroblast and macrophage were reconstructed with the Monocle tool. Results: Across dermal cells, there were six cell types, namely, EN1-negative myofibroblasts, EN1-positive myofibroblasts, hematopoietic cells, macrophages, pericytes, and endothelial cells. Ligand and receptor genes between myofibroblasts and macrophages mainly modulated cell proliferation and migration, tube development, and the TGF-ß pathway. Specific genes that were differentially expressed in fibrotic compared to regenerative myofibroblasts or macrophages were separately identified. Specific genes between fibrotic and regenerative myofibroblasts were involved in the mRNA metabolic process and organelle organization. Specific genes between fibrotic and regenerative macrophages participated in regulating immunity and phagocytosis. We then observed the underlying evolution of myofibroblasts or macrophages. Conclusion: Collectively, our findings reveal that myofibroblasts and macrophages may alter the skin wound healing fate through modulating critical signaling pathways.

Selective Elimination of Senescent Fibroblasts by Targeting the Cell Surface Protein ACKR3.

The accumulation of senescent cells in aging tissues is associated with age-related diseases and functional decline. Thus, senolysis, a therapy aimed at rejuvenation by removing senescent cells from the body, is being developed. However, this therapy requires the identification of membrane surface antigens that are specifically expressed on senescent cells for their selective elimination. We showed that atypical chemokine receptor 3 (ACKR3), a receptor of the CXC motif chemokine 12 (CXCL12) implicated in cancer, inflammation, and cardiovascular disorders, is selectively expressed on the surface of senescent human fibroblasts but not on proliferating cells. Importantly, the differential presence of ACKR3 enabled the isolation of senescent cells by flow cytometry using anti-ACKR3 antibodies. Furthermore, antibody-dependent cellular cytotoxicity assays revealed that cell surface ACKR3 preferentially sensitizes senescent but not dividing fibroblasts to cell injury by natural killer cells. Conclusively, the selective expression of ACKR3 on the surface of senescent cells allows the preferential elimination of senescent cells. These results might contribute to the future development of novel senolysis approaches.