EZH2-dependent epigenetic modulation of histone H3 lysine-27 contributes to psoriasis by promoting keratinocyte proliferation.
ABSTRACT: Psoriasis is characterized by keratinocyte hyperproliferation. While significant progress has been made in understanding the molecular mechanism regulating the proliferation of keratinocytes, little is known about the epigenetic factors that control this process. EZH2 and EZH2 mediated trimethylation of histone H3 lysine 27 (H3K27me3) was previously shown ectopically expressed in carcinoma and mediated proliferation, thereby we sought to clarify the role of EZH2-H3K27me3 in the proliferation of psoriatic keratinocyte. Interestingly, we found that EZH2 and H3K27me3 were both overexpressed in the epidermis of psoriatic lesional skin compared to normal skin. In vitro, the expression of EZH2 and H3K27me3 was stimulated in human keratinocytes treated with mixture of psoriasis-related cytokines pool (TNF-?, IFN-?, IL-17A, and IL-22). Knockdown of EZH2 significantly reduced keratinocyte proliferative activity. Results from mRNA microarray analysis suggested that Kallikrein-8 (KLK8) might be the target gene of EZH2 in psoriatic keratinocytes. Overexpression or knockdown KLK8 could partially reverse the abnormal proliferation of keratinocytes caused by knockdown or overexpression of EZH2. In vivo, the inhibitor of EZH2, GSK126 could ameliorate the imiquimod-induced psoriasiform lesion. These results suggest that EZH2 might be a therapeutic target for the treatment of psoriasis.
Project description:Psoriasis is characterized by keratinocyte proliferation and immune cell infiltration. M2 isoform of pyruvate kinase (PKM2) was reported to have an important role in cell proliferation, which is a rate-limiting enzyme that regulates the final step of glycolysis. However, how PKM2 regulates cell metabolism and proliferation in psoriatic keratinocytes is still poorly understood. Interestingly, we found that PKM2 was highly expressed in psoriatic epidermis from patients and mouse models. PKM2 overexpression promoted keratinocyte glycolytic metabolism while knockdown inhibited keratinocyte proliferation and glycolysis. Mice lacking PKM2 specifically in keratinocytes, pharmacological inhibition of PKM2 or glycolysis inhibited keratinocyte proliferation and showed obvious remission in an imiquimod-induced psoriatic mouse model. Moreover, the inhibitor of the EGF-receptor blocked EGF-stimulated PKM2 expression and glycolysis in keratinocytes. We identify PKM2 as an upregulated gene in psoriasis. PKM2 is essential in keratinocyte over-proliferation and may represent a therapeutic target for psoriasis.
Project description:Psoriasis is a frequent, inflammatory skin disease characterized by keratinocyte hyperproliferation and a disease-related infiltration of immune cells. Here, we identified a novel proinflammatory signaling pathway driven by cyclin-dependent kinase 4 (CDK4) and CDK6 and the methyltransferase EZH2 as a valid target for psoriasis therapy. Delineation of the pathway revealed that CDK4/6 phosphorylated EZH2 in keratinocytes, thereby triggering a methylation-induced activation of STAT3. Subsequently, active STAT3 resulted in the induction of I?B?, which is a key proinflammatory transcription factor required for cytokine synthesis in psoriasis. Pharmacological or genetic inhibition of CDK4/6 or EZH2 abrogated psoriasis-related proinflammatory gene expression by suppressing I?B? induction in keratinocytes. Importantly, topical application of CDK4/6 or EZH2 inhibitors on the skin was sufficient to fully prevent the development of psoriasis in various mouse models by suppressing STAT3-mediated I?B? expression. Moreover, we found a hyperactivation of the CDK4/6-EZH2 pathway in human and mouse psoriatic skin lesions. Thus, this study not only identifies a novel psoriasis-relevant proinflammatory pathway, but also proposes the repurposing of CDK4/6 or EZH2 inhibitors as a new therapeutic option for patients with psoriasis.
Project description:Epithelial keratinocyte proliferation is an essential element of wound repair, and abnormal epithelial proliferation is an intrinsic element in the skin disorder psoriasis. The factors that trigger epithelial proliferation in these inflammatory processes are incompletely understood. Here we have shown that regenerating islet-derived protein 3-alpha (REG3A) is highly expressed in keratinocytes during psoriasis and wound repair and in imiquimod-induced psoriatic skin lesions. The expression of REG3A by keratinocytes is induced by interleukin-17 (IL-17) via activation of keratinocyte-encoded IL-17 receptor A (IL-17RA) and feeds back on keratinocytes to inhibit terminal differentiation and increase cell proliferation by binding to exostosin-like 3 (EXTL3) followed by activation of phosphatidylinositol 3 kinase (PI3K) and the kinase AKT. These findings reveal that REG3A, a secreted intestinal antimicrobial protein, can promote skin keratinocyte proliferation and can be induced by IL-17. This observation suggests that REG3A may mediate the epidermal hyperproliferation observed in normal wound repair and in psoriasis.
Project description:Proliferating cells, compared with quiescent cells, are more dependent on glucose for their growth. Although glucose transport in keratinocytes is mediated largely by the Glut1 facilitative transporter, we found that keratinocyte-specific ablation of Glut1 did not compromise mouse skin development and homeostasis. Ex vivo metabolic profiling revealed altered sphingolipid, hexose, amino acid, and nucleotide metabolism in Glut1-deficient keratinocytes, thus suggesting metabolic adaptation. However, cultured Glut1-deficient keratinocytes displayed metabolic and oxidative stress and impaired proliferation. Similarly, Glut1 deficiency impaired in vivo keratinocyte proliferation and migration within wounded or UV-damaged mouse skin. Notably, both genetic and pharmacological Glut1 inactivation decreased hyperplasia in mouse models of psoriasis-like disease. Topical application of a Glut1 inhibitor also decreased inflammation in these models. Glut1 inhibition decreased the expression of pathology-associated genes in human psoriatic skin organoids. Thus, Glut1 is selectively required for injury- and inflammation-associated keratinocyte proliferation, and its inhibition offers a novel treatment strategy for psoriasis.
Project description:Psoriasis is characterized by uncontrolled proliferation and poor differentiation. Sirtuin1 (SIRT1) a class III deacetylase, crucial for differentiation in normal keratinocytes, is reduced in psoriasis. Down regulated SIRT1 levels may contribute to poor differentiation in psoriasis. In addition, the levels of early differentiation factors Keratin1 (K1) and Keratin10 (K10) are depleted in psoriasis. We attempted to study a possible effect of fructose, a SIRT1 upregulator and Propylthiouracil (PTU) to augment differentiation in psoriatic keratinocytes. Keratinocytes were cultured from lesional biopsies obtained from psoriatic patients and control cells were obtained from patients undergoing abdominoplasty. Cells were treated with fructose and PTU individually. K1 and K10 transcript levels were measured to evaluate early differentiation; SIRT1 protein expression was also studied to decipher its role in the mechanism of differentiation. The K1, K10 transcript levels, SIRT1 protein and transcript levels in fructose treated psoriatic keratinocytes were improved. This suggests keratinocyte differentiation was induced by fructose through SIRT1 upregulation. Whereas PTU induced differentiation, as confirmed by improved K1, K10 transcript levels followed a non-SIRT1 mechanism. We conclude that the use of fructose and PTU may be an adjunct to the existing therapies for psoriasis.
Project description:Psoriasis is a characteristic inflammatory and scaly skin condition with typical histopathological features including increased proliferation and hampered differentiation of keratinocytes. The activation of innate and adaptive inflammatory cellular immune responses is considered to be the main trigger factor of the epidermal changes in psoriatic skin. However, the molecular players that are involved in enhanced proliferation and impaired differentiation of psoriatic keratinocytes are only partly understood. One important factor that regulates differentiation on the cellular level is Ca(2+). In normal epidermis, a Ca(2+) gradient exists that is disturbed in psoriatic plaques, favoring impaired keratinocyte proliferation. Several TRPC channels such as TRPC1, TRPC4, or TRPC6 are key proteins in the regulation of high [Ca(2+)](ex) induced differentiation. Here, we investigated if TRPC channel function is impaired in psoriasis using calcium imaging, RT-PCR, western blot analysis and immunohistochemical staining of skin biopsies. We demonstrated substantial defects in Ca(2+) influx in psoriatic keratinocytes in response to high extracellular Ca(2+) levels, associated with a downregulation of all TRPC channels investigated, including TRPC6 channels. As TRPC6 channel activation can partially overcome this Ca(2+) entry defect, specific TRPC channel activators may be potential new drug candidates for the topical treatment of psoriasis.
Project description:<h4>Background</h4>Keratinocytes of psoriasis have anti-apoptotic properties including delayed apoptosis process, accelerated proliferation metabolism and postponed differentiation process. However, the specific mechanism leading to the abnormal biological behavior of keratinocytes remains unclear.<h4>Objectives</h4>We investigated the role of increased RPL22 expression in regulating the abnormal biological behavior of keratinocytes and the mechanism of regulation of RPL22 expression in skin lesions of psoriatic patients.<h4>Methods</h4>We examined clinical samples and utilized cytokine-induced cell and IMQ-treated mouse models. We determined the expression and functions of RPL22 <i>in vitro</i> and <i>in vivo</i>.<h4>Results</h4>We showed that RPL22 expression was significantly increased in the skin lesions of psoriasis patients and IMQ-treated psoriatic-like mice. Such increased expression is attributed to hyperacetylation of histone H3K27 in the promoter region of RPL22. Interestingly, overexpression of RPL22 enhanced keratinocyte proliferation by increasing cyclinD1 expression and accelerated CD4<sup>+</sup>T cells recruitment <i>via</i> upregulating CXCL10 expression. Finally, we demonstrated that RPL22 overexpression promoted psoriasiform phenotypes in IMQ-induced mouse skins.<h4>Conclusions</h4>These findings suggested that RPL22 regulates keratinocytes abnormal biological behavior and contributes to the development of psoriatic phenotypes. Thus, RPL22 might be a novel potential molecular target for treatment of psoriasis.
Project description:NF-?B is constitutively activated in psoriatic epidermis. However, how activated NF-?B promotes keratinocyte hyperproliferation in psoriasis is largely unknown. Here we report that the NF-?B activation triggered by inflammatory cytokines induces the transcription of microRNA (miRNA) miR-31, one of the most dynamic miRNAs identified in the skin of psoriatic patients and mouse models. The genetic deficiency of miR-31 in keratinocytes inhibits their hyperproliferation, decreases acanthosis and reduces the disease severity in psoriasis mouse models. Furthermore, protein phosphatase 6 (ppp6c), a negative regulator that restricts the G1 to S phase progression, is diminished in human psoriatic epidermis and is directly targeted by miR-31. The inhibition of ppp6c is functionally important for miR-31-mediated biological effects. Moreover, NF-?B activation inhibits ppp6c expression directly through the induction of miR-31, and enhances keratinocyte proliferation. Thus, our data identify NF-?B-induced miR-31 and its target, ppp6c, as critical factors for the hyperproliferation of epidermis in psoriasis.
Project description:Psoriasis is a chronic Th1/Th17 lymphocytes-mediated inflammatory skin disease, in which epidermal keratinocytes exhibit a peculiar senescent state, resistance to apoptosis and the acquisition of senescence-associated secretory phenotype (SASP). SASP consists of the release of soluble factors, including IGFBPs, that exert extracellular and intracellular functions in IGF-dependent or independent manner.In this report, we investigated the expression and function of IGFBP2 in senescent keratinocytes isolated from the skin of patients with plaque psoriasis. We found that IGFBP2 is aberrantly expressed and released by these cells in vivo, as well as in vitro in keratinocyte cultures undergoing progressive senescence, and it associates with the cyclin-dependent kinase inhibitors p21 and p16 expression. For the first time, we provide evidence for a dual action of IGFBP2 in psoriatic keratinocytes during growth and senescence processes. While extracellular IGFBP2 counter-regulates IGF-induced keratinocyte hyper-proliferation, intracellular IGFBP2 inhibits apoptosis by interacting with p21 and protecting it from ubiquitin-dependent degradation. Indeed, we found that cytoplasmic p21 sustains anti-apoptotic processes, by inhibiting pro-caspase 3 cleavage and JNK phosphorylation in senescent psoriatic keratinocytes. As a consequence, abrogation of p21, as well as that of IGFBP2, found to stabilize cytoplasmic p21 levels, lead to the restoration of apoptosis mechanisms in psoriatic keratinocytes, commonly observed in healthy cells.
Project description:Psoriasis is a chronic inflammatory skin disease characterized by exaggerated keratinocyte proliferation. Current opinion indicates that psoriasis is driven by T cell-mediated immune responses targeting keratinocytes. However, psoriasis cannot be explained solely on the basis of T-cell activation, and it is likely that an intrinsic alteration in epidermal keratinocytes plays a very important role in disease expression. Syndecans comprise a major family of cell surface heparan sulfate proteoglycans. Several studies indicate their role in adhesion, cell-extracellular matrix interactions, migration, keratinocyte proliferation and differentiation, inflammation, and wound healing. To determine the expression of syndecan-1 in psoriasis, skin samples from 29 patients with fully developed psoriasis and skin samples from 14 healthy volunteer persons with no personal or family history of psoriasis were immunohistochemically examined using monoclonal antibody against syndecan-1. The expression of syndecan-1 was analyzed in whole mount section of psoriatic and non-psoriatic skin biopsies under high magnification (400x). In addition, the intensity and topography of reaction in the cell, as well as localization of positive cells in the epidermis were evaluated. Strong syndecan-1 reactivity in epidermal cells in all non-psoriatic and psoriatic samples was observed. Statistical analysis showed no significant differences between two analyzed groups (P > 0.05). In normal skin syndecan-1 was expressed in full thickness of the epidermis. The strongest reaction was observed in membranes and intercellular junctions of spinous and granular layer while basal cells showed weaker expression that was confined to cytoplasm. In psoriatic skin syndecan-1 was expressed in the membrane and intercellular junction of cells located in thickened and elongated rete ridges of the epidermis. The strongest reaction was in basal and suprabasal layers and expression diminished through spinous layer. Cells in spinous layer lose syndecan-1 expression, which is opposite pattern to normal skin. Our results suggest that aberrant skin expression of syndecan-1 may be involved in the development of psoriasis.