Inhibition of ERK5 Elicits Cellular Senescence in Melanoma via the Cyclin-Dependent Kinase Inhibitor p21.

Melanoma is the deadliest skin cancer with a very poor prognosis in advanced stages. Although targeted and immune therapies have improved survival, not all patients benefit from these treatments. The mitogen-activated protein kinase ERK5 supports the growth of melanoma cells in vitro and in vivo. However, ERK5 inhibition results in cell-cycle arrest rather than appreciable apoptosis. To clarify the role of ERK5 in melanoma growth, we performed transcriptomic analyses following ERK5 knockdown in melanoma cells expressing BRAFV600E and found that cellular senescence was among the most affected processes. In melanoma cells expressing either wild-type or mutant (V600E) BRAF, both genetic and pharmacologic inhibition of ERK5 elicited cellular senescence, as observed by a marked increase in senescence-associated ß-galactosidase activity and p21 expression. In addition, depletion of ERK5 from melanoma cells resulted in increased levels of CXCL1, CXCL8, and CCL20, proteins typically involved in the senescence-associated secretory phenotype. Knockdown of p21 suppressed the induction of cellular senescence by ERK5 blockade, pointing to p21 as a key mediator of this process. In vivo, ERK5 knockdown or inhibition with XMD8-92 in melanoma xenografts promoted cellular senescence. Based on these results, small-molecule compounds targeting ERK5 constitute a rational series of prosenescence drugs that may be exploited for melanoma treatment. SIGNIFICANCE: This study shows that targeting ERK5 induces p21-mediated cellular senescence in melanoma, identifying a prosenescence effect of ERK5 inhibitors that may be exploited for melanoma treatment.

An Anti-inflammatory Fe3 O4 -Porphyrin Nanohybrid Capable of Apoptosis through Upregulation of p21 Kinase Inhibitor Having Immunoprotective Properties under Anticancer PDT Conditions.

We report the influence of Fe3 O4 nanoparticles (NPs) on porphyrins in the development of photosensitizers (PSs) for efficient photodynamic therapy (PDT) and possible post-PDT responses for inflicting cancer cell death. Except for Au, most metal-based nanomaterials are unsuitable for clinical applications. The US Food and Drug Administration and other agencies have approved Feraheme and a few other iron oxide NPs for clinical use, paving the way for novel biocompatible immunoprotective superparamagnetic iron oxide nanohybrids to be developed as nanotherapeutics. A water-soluble nanohybrid, referred to here as E-NP, comprising superparamagnetic Fe3 O4 NPs functionalised with tripyridyl porphyrin PS was introduced through a rigid 4-carboxyphenyl linker. As a PDT agent, the efficacy of E-NP toward the AGS cancer cell line showed enhanced photosensitising ability as determined through in vitro photobiological assays. The cellular uptake of E-NPs by AGS cells led to apoptosis by upregulating ROS through cell-cycle arrest and loss of mitochondrial membrane potential. The subcellular localisation of the PSs in mitochondria stimulated apoptosis through upregulation of p21, a proliferation inhibitor capable of preventing tumour development. Under both PDT and non-PDT conditions, this nanohybrid can act as an anti-inflammatory agent by decreasing the production of NO and superoxide ions in murine macrophages, thus minimising collateral damage to healthy cells.

The NUCKS1-SKP2-p21/p27 axis controls S phase entry.

Efficient entry into S phase of the cell cycle is necessary for embryonic development and tissue homoeostasis. However, unscheduled S phase entry triggers DNA damage and promotes oncogenesis, underlining the requirement for strict control. Here, we identify the NUCKS1-SKP2-p21/p27 axis as a checkpoint pathway for the G1/S transition. In response to mitogenic stimulation, NUCKS1, a transcription factor, is recruited to chromatin to activate expression of SKP2, the F-box component of the SCFSKP2 ubiquitin ligase, leading to degradation of p21 and p27 and promoting progression into S phase. In contrast, DNA damage induces p53-dependent transcriptional repression of NUCKS1, leading to SKP2 downregulation, p21/p27 upregulation, and cell cycle arrest. We propose that the NUCKS1-SKP2-p21/p27 axis integrates mitogenic and DNA damage signalling to control S phase entry. The Cancer Genome Atlas (TCGA) data reveal that this mechanism is hijacked in many cancers, potentially allowing cancer cells to sustain uncontrolled proliferation.

EZH2-mediated long-chain non-coding RNA LINC00963 promotes proliferation and invasion of glioma cells through inhibiting p21 expression.

PURPOSE: In the early stage, bioinformatics analysis revealed that the expression of long-chain non-coding RNA LINC00963 in glioma tissues was remarkably increased, but its biological effects on glioma and the potential molecular mechanisms have not been reported. This study aimed to conduct a preliminary discussion on the impact of LINC00963 on glioma, so as to provide new ideas for the treatment of this cancer. METHODS: GEPIA database was consulted to determine the expression level of LINC00963 in gliomas. In addition, the interplay between LINC00963 expression and the prognosis of glioma patients was analyzed by Kaplan-Meier method. Effects of LINC00963 on the proliferation and migration of glioma cells were determined using Cell Counting Kit (CCK-8) and Transwell assay. The subcellular localization of LINC00963 was determined by nuclear separation experiments. At the same time, the regulation of LINC00963 on p21 expression was verified through qRT-PCR and Western blot experiments. RESULTS: By analyzing the GEPIA database, we found that LINC00963 was highly expressed in glioma tissues. Meanwhile, qRT-PCR results revealed that LINC00963 level in glioma tissues and cell lines was remarkably higher than that in the normal control group. Kaplan-Meier and log-rank test revealed that there was no statistically significant association between the expression level of LINC00963 and the prognosis of patients with glioma. In addition, in vitro cell assay results indicated that downregulation of LINC00963 markedly suppressed the proliferation and invasiveness of glioma cells. Finally, the related mechanism analysis revealed that LINC00963 may inhibit p21 expression through modulation of EZH2. CONCLUSIONS: LINC00963 can inhibit p21 expression through EZH2 and thus enhance the proliferative and invasive capacities of glioma cells. Consequently, LINC00963 may be a potential therapeutic target for gliomas.

WIPI1 promotes osteosarcoma cell proliferation by inhibiting CDKN1A.

Detection of TCGA data revealed that WIPI1 is highly expressed in osteosarcoma cells. So we explore the mechanisms of WIPI1 affecting the proliferation of osteosarcoma cells through Affymetrix microarray analysis. Functional analysis of differentially expressed genes shows that the classical signaling pathways affecting tumor formation and development have changed significantly. By fitting analysis, it is speculated that the WIPI1 may function in the direction of osteosarcoma by regulating the expression of multiple cell cycle-related genes such as CDKN1A, CDK4 and CCND1. Therefore, the key genes are selected for RT-PCR and Western-blot verification. Combined with flow and other means, WIPI1 may affect the cell cycle and the osteosarcoma by regulating the expression of CDKN1A, CDK4 and CCND1. To verify the results, the effect of WIPI1 on cell proliferation was quantified by MTT, cell counts and nude mouse tumorigenicity assay. The results showed that WIPI1 promotes osteosarcoma cell proliferation.

Sequential targeting of YAP1 and p21 enhances the elimination of senescent cells induced by the BET inhibitor JQ1.

Chondrosarcoma (CHS) is the second most common bone malignancy with limited therapeutic approaches. Our previous study has found that Yes associated protein 1 (YAP1) is downregulated in CHS cells treated with bromodomain and extraterminal domain (BET) inhibitor JQ1. However, the precise role of YAP1 in CHS is largely unknown. Herein, we found that YAP1 expression was upregulated in CHS tissues, and positively correlated with its grading score. Loss of YAP1 inhibited CHS proliferation and induced cellular senescence, while expression of YAP1 mutants revealed YAP1/TEA domain family member (TEAD)-dependent negative regulation of p21 and subsequent cellular senescence. These results were validated by in vivo experiments using stable shYAP1 cell lines. Mechanistically, negative regulation of p21 by YAP1 occurred post-transcriptionally via Dicer-regulated miRNA networks, specifically, the miR-17 family. Furthermore, we demonstrated that sequential targeting of YAP1 and p21 enhanced the elimination of JQ1-induced senescent cells in a Bcl-2-like 1 (Bcl-XL)/Caspase-3 dependent manner. Altogether, we unveil a novel role of YAP1 signaling in mediating CHS cell senescence and propose a one-two punch approach that sequentially targets the YAP1/p21 axis to eliminate senescent cells.

Multiple modes of cell death in neuroendocrine tumors induced by artesunate.

BACKGROUND: The paucity of effective treatment in neuroendocrine tumors (NETs) encouraged us to investigate the therapeutic value of artesunate (ART) promised by its inhibitory effect against various tumors and broad safety profile. METHODS: We evaluated the impact of ART on three NET cell lines, BON-1, QGP-1 and NCI-H727 on cellular and molecular levels. RESULTS: Our results showed that ART induced endoplasmic reticulum (ER) stress through phosphorylation of eIF2α, which further gave rise to autophagy in all three NET cell lines. Specifically, apoptosis and ferroptosis were also observed in BON-1 cells, which made BON-1 cell line more vulnerable upon ART treatment. The different sensitivities presented on the three cell lines also associated with a differential regulation of p21 on the long run. Co-treatment with p21 inhibitor UC2288 showed an additive effect on QGP-1 and NCI-H727 cell lines indicating p21 upregulation in these two cell lines might confer resistance towards ART treatment. CONCLUSIONS: It is possible to include ART in the treatment of NETs in the future.

CDKN1A Gene Expression in Two Multiple Myeloma Cell Lines With Different P53 Functionality.

BACKGROUND/AIM: Multiple myeloma is a highly heterogeneous disease of clonal plasma cells. Histone deacetylase (HDAC) inhibitors are promising anticancer drugs but their precise mechanisms of actions are not well understood. MATERIALS AND METHODS: Cell-cycle regulation and pro-apoptotic effects of two histone deacetylase inhibitors, suberohydroxamic acid (SAHA) and suberoylanilide hydroxamic acid (SBHA), were analyzed in multiple myeloma cell lines RPMI8226 and U266 with differing TP53 status using gene-expression analysis. RESULTS: Enhanced expression of cyclin-dependent kinase inhibitor 1A (CDKN1A/p21WAF/CIP1) detected in the TP53-deleted U266 cell line after SAHA treatment indicates the P53-independent mode of transcriptional activation of CDKN1A gene. In contrast, CDKN1A gene expression was significantly increased by both SBHA and SAHA treatment of TP53-mutated RPMI8226 cells. CONCLUSION: SAHA appears to be a potentially effective pro-apoptotic and anticancer drug with universal application in the treatment of heterogeneous populations of multiple myeloma cells.

Identification of nagilactone E as a protein synthesis inhibitor with anticancer activity.

Norditerpenoids and dinorditerpenoids represent diterpenoids widely distributed in the genus Podocarpus with notable chemical structures and biological activities. We previously reported that nagilactone E (NLE), a dinorditerpenoid isolated from Podocarpus nagi, possessed anticancer effects against lung cancer cells in vitro. In this study we investigated the in vivo effect of NLE against lung cancer as well as the underlying mechanisms. We administered NLE (10 mg·kg-1·d-1, ip) to CB-17/SCID mice bearing human lung cancer cell line A549 xenograft for 3 weeks. We found that NLE administration significantly suppressed the tumor growth without obvious adverse effects. Thereafter, RNA sequencing (RNA-seq) analysis was performed to study the mechanisms of NLE. The effects of NLE on A549 cells have been illustrated by GO and pathway enrichment analyses. CMap dataset analysis supported NLE to be a potential protein synthesis inhibitor. The inhibitory effect of NLE on synthesis of total de novo protein was confirmed in Click-iT assay. Using the pcDNA3-RLUC-POLIRES-FLUC luciferase assay we further demonstrated that NLE inhibited both cap-dependent and cap-independent translation. Finally, molecular docking revealed the low-energy binding conformations of NLE and its potential target RIOK2. In conclusion, NLE is a protein synthesis inhibitor with anticancer activity.

LncRNA ROR1-AS1 promotes colon cancer cell proliferation by suppressing the expression of DUSP5/CDKN1A.

OBJECTIVE: The purpose of this study was to explore the possible role of ROR1-AS1 in the pathogenesis of colon cancer and the underlying mechanism. PATIENTS AND METHODS: The expression levels of ROR1-AS1 in 75 colon cancer tissue samples and adjacent ones, as well as in cell lines were examined by quantitative Polymerase Chain Reaction (qPCR). Then, ROR1-AS1 overexpression plasmid and siRNA were transfected into colon cancer cells using liposome method. After that, Cell Counting Kit-8 (CCK-8) and plate colony formation assays were conducted to analyze cell proliferation, while flow cytometry was applied for the analysis of cell cycle and apoptosis. At last, the mechanism of action of ROR1-AS1 was further explored by nuclear separation, RNA binding protein immunoprecipitation (RIP) and chromatin immunoprecipitation (CHIP) assays. RESULTS: ROR1-AS1 level in colon cancer tissues was remarkably higher than that in normal tissues, and the expression in tumors of stage III and IV was remarkably higher than those of stage I and II. Meanwhile, tumors with diameters more than 5 cm had a higher ROR1-AS1 expression than those less than 5 cm. After transfection with ROR1-AS1 overexpression plasmid, the cell proliferation ability was enhanced, the G0/G1 phase time of cell cycle was shortened, and the apoptosis was suppressed. However, the opposite result was observed after ROR1-AS1 was downregulated. Furthermore, RIP showed that ROR1-AS1 can bind to enhancer of zeste homolog 2 (EZH2) and inhibit the expression of DUSP5, and thus be engaged in the proliferation and apoptosis of colon cancer cells. CONCLUSIONS: ROR1-AS1 is highly expressed either in colon cancer tissues or in cell lines, which is able to enhance cell proliferation, accelerate cell cycle, and inhibit cell apoptosis. The mechanism of ROR1-AS1 to participate in the development of colon cancer may be the downregulation of DUSP5 via combination with EZH2.

linc01088 promotes cell proliferation by scaffolding EZH2 and repressing p21 in human non-small cell lung cancer.

AIMS: Non-small cell lung cancer (NSCLC), characterized by extensive metastasis and poor prognosis, is the most common type of lung cancer. Dysregulation of certain lncRNAs is known to be linked to the tumorigenesis of NSCLC. However, the specific roles in NSCLC for many other lncRNAs, such as linc01088, remain largely unknown. MATERIALS AND METHODS: The expression patterns of linc01088, p21 and EZH2 were examined both in NSCLC tissues and cell lines using RT-qPCR assay. CCK-8, colony formation, immunofluorescence staining, and flow cytometry assays were employed to evaluate the effects of linc01088 on NSCLC cell proliferation properties. RNA immunoprecipitation (RIP) assay was performed to determine the direct binding relationship between linc01088 and zeste homolog 2 (EZH2). Western blot and RT-qPCR analysis were performed to assess p21 level within knockdown of either linc01088 or EZH2. Nude mouse subcutaneous NSCLC models were constructed for further validating the effects and mechanisms of linc01088 in vivo. KEY FINDINGS: linc01088 and EZH2 were highly expressed both in NSCLC tissues and cell lines. Knockdown of linc01088 suppressed the proliferation of NSCLC cells, and prolonged the G1 phase while shortened S and G2-M phases. RIP assay revealed the direct binding relationship between linc01088 and EZH2. Knockdown of either linc01088 or EZH2 induced up-regulation of p21 expression, which subsequently inhibited the tumor growth. SIGNIFICANCE: We demonstrated that linc01088 could promote cell proliferation via binding with EZH2 to repress p21, which aggravates the tumorigenesis of NSCLC. Therefore, linc01088 might be a potential oncogene and target for novel anti-tumor therapies.

Local and transient inhibition of p21 expression ameliorates age-related delayed wound healing.

Nonhealing chronic wounds in the constantly growing elderly population represent a major public health problem with high socioeconomic burden. Yet, the underlying mechanism of age-related impairment of wound healing remains elusive. Here, we show that the number of dermal cells expressing cyclin-dependent kinase inhibitor p21 was elevated upon skin injury, particularly in aged population, in both man and mouse. The nuclear expression of p21 in activated wound fibroblasts delayed the onset of the proliferation phase of wound healing in a p53-independent manner. Further, the local and transient inhibition of p21 expression by in vivo delivered p21-targeting siRNA ameliorated the delayed wound healing in aged mice. Our results suggest that the increased number of p21+ wound fibroblasts enforces the age-related compromised healing, and targeting p21 creates potential clinical avenues to promote wound healing in aged population.

A potential role of cyclin-dependent kinase inhibitor 1 (p21/WAF1) in the pathogenesis of endometriosis: Directions for future research.

Endometriosis is a common gynecological disorder that affects approximately 6-10% of the female population impairing the quality of life of patients. Several pathophysiologic pathways have been proposed as potential regulators of its severity; however, to date, the processes that trigger the onset and that influence the severity of the disease are not fully understood; hence, leading to disease recurrence in approximately 10-67% of cases. Cyclin-dependent kinase inhibitor 1 (p21/WAF1) is a protein that is a major target of p53 and is related to cell cycle arrest (it regulates transition from the G1 to the S phase) when DNA damage is detected. Its activity has been also linked to the angiogenic potential of tumors as it promotes the expression of various kinases that are responsible for endothelial development and function. Although several articles have underlined the importance of this protein in cancer cell development and tumor growth, there are no relevant data in the field of endometriosis. Indirect evidence suggests, however, that it may be involved in the pathogenesis of endometriosis as it inhibits the activity of various kinases which have been correlated with the course and severity of the disease. The present article investigates the background theory that implies the potential role of cyclin-dependent kinase inhibitor 1 (p21/WAF1) in the pathogenesis of endometriosis. Implications for future research are also provided given that indirect evidence seem to associate downregulation of p21 with decreased growth and invasiveness of human endometrial stromal cells.

Patterns of Early p21 Dynamics Determine Proliferation-Senescence Cell Fate after Chemotherapy.

Chemotherapy is designed to induce cell death. However, at non-lethal doses, cancer cells can choose to remain proliferative or become senescent. The slow development of senescence makes studying this decision challenging. Here, by analyzing single-cell p21 dynamics before, during, and days after drug treatment, we link three distinct patterns of early p21 dynamics to final cell fate. Surprisingly, while high p21 expression is classically associated with senescence, we find the opposite at early times during drug treatment: most senescence-fated cells express much lower p21 levels than proliferation-fated cells. We demonstrate that these dynamics lead to a p21 "Goldilocks zone" for proliferation, in which modest increases of p21 expression can lead to an undesirable increase of cancer cell proliferation. Our study identifies a counter-intuitive role for early p21 dynamics in the cell-fate decision and pinpoints a source of proliferative cancer cells that can emerge after exposure to non-lethal doses of chemotherapy.

MDM2-Mediated p21 Proteasomal Degradation Promotes Fluoride Toxicity in Ameloblasts.

Fluoride overexposure is an environmental health hazard and can cause enamel and skeletal fluorosis. Previously we demonstrated that fluoride increased acetylated-p53 and its downstream target p21 in ameloblast-derived LS8 cells. However, p21 function in fluoride toxicity is not well characterized. This study seeks to gain a better understanding of how p53 down-stream mediators, p21 and MDM2, respond to fluoride toxicity. LS8 cells were treated with NaF with/without MG-132 (proteasome inhibitor) or Nutlin-3a (MDM2 antagonist). NaF treatment for 2-6 h increased phospho-p21, which can inhibit apoptosis. However, phospho-p21 and p21 were decreased by NaF at 24 h, even though p21 mRNA was significantly increased at this time point. MG-132 reversed the fluoride-mediated p21 decrease, indicating that fluoride facilitates p21 proteasomal degradation. MG-132 suppressed fluoride-induced caspase-3 cleavage, suggesting that the proteasome plays a pro-apoptotic role in fluoride toxicity. NaF increased phospho-MDM2 in vitro and in mouse ameloblasts in vivo. Nutlin-3a suppressed NaF-mediated MDM2-p21 binding to reverse p21 degradation which increased phospho-p21. This suppressed apoptosis after 24 h NaF treatment. These results suggest that MDM2-mediated p21 proteasomal degradation with subsequent phospho-p21 attenuation contributes to fluoride-induced apoptosis. Inhibition of MDM2-mediated p21 degradation may be a potential therapeutic target to mitigate fluoride toxicity.

Oridonin protects against cardiac hypertrophy by promoting P21-related autophagy.

Autophagy is an endogenous protective process; the loss of autophagy could destabilize proteostasis and elevate intracellular oxidative stress, which is critically involved in the development of cardiac hypertrophy and heart failure. Oridonin, a natural tetracycline diterpenoid from the Chinese herb Rabdosia, has autophagy activation properties. In this study, we tested whether oridonin protects against cardiac hypertrophy in mice and cardiomyocytes. We implemented aortic banding to induce a cardiac hypertrophy mouse model, and oridonin was given by gavage for 4 weeks. Neonatal rat cardiomyocytes were stimulated with angiotensin II to simulate neurohumoural stress. Both in vivo and in vitro studies suggested that oridonin treatment mitigated pressure overload-induced cardiac hypertrophy and fibrosis, and also preserved heart function. Mice that received oridonin exhibited increased antioxidase activities and suppressed oxidative injury compared with the aortic banding group. Moreover, oridonin enhanced myocardial autophagy in pressure-overloaded hearts and angiotensin II-stimulated cardiomyocytes. Mechanistically, we discovered that oridonin administration regulated myocardial P21, and cytoplasmic P21 activated autophagy via regulating Akt and AMPK phosphorylation. These findings were further corroborated in a P21 knockout mouse model. Collectively, pressure overload-induced autophagy dysfunction causes intracellular protein accumulation, resulting in ROS injury while aggravating cardiac hypertrophy. Thus, our data show that oridonin promoted P21-related autophagic lysosomal degradation, hence attenuating oxidative injury and cardiac hypertrophy.

Sirtuin 2 expression suppresses oxidative stress and senescence of nucleus pulposus cells through inhibition of the p53/p21 pathway.

Intervertebral disc degeneration (IDD) is a kind of disease associated with nucleus pulposus (NP) cell senescence. Previous studies have shown that the sirtuin family plays an extremely important role in the progress of cell aging. However, whether sirtuin2 (Sirt2) protects against IDD remains unknown. The aim of this study was to determine whether Sirt2 protected NP from degradation in IDD. The expression of Sirt2 in different degree of degenerate disc tissues was determined by reverse transcription-polymerase chain reaction. Interleukin 1 beta (IL-1ß) was used to stimulate the degeneration of NP cells. Subsequently, lentivirus transfection was performed to increase Sirt2 expression in vitro. Meanwhile, the function of Sirt2 overexpression in the progress of NP cell degeneration was evaluated. Our study showed that the expression of Sirt2 markedly decreased in severe degenerated disc tissues. IL-1ß significantly promoted the progress of IDD. Meanwhile, overexpression of Sirt2 could reverse the effects of IL-1ß. The data also revealed that Sirt2 overexpression obviously increased the production of antioxidant SOD1/2 and suppressed oxidative stress in the disc. Moreover, p53 and p21 could be significantly suppressed by Sirt2 overexpression. These results suggested that Sirt2 prevented NP degradation via restraining oxidative stress and cell senescence through inhibition of the p53/p21 pathway. Furthermore, Sirt2 might become a novel target for IDD therapy in the future.

Ironing out the role of the cyclin-dependent kinase inhibitor, p21 in cancer: Novel iron chelating agents to target p21 expression and activity.

Iron (Fe) has become an important target for the development of anti-cancer therapeutics with a number of Fe chelators entering human clinical trials for advanced and resistant cancer. An important aspect of the activity of these compounds is their multiple molecular targets, including those that play roles in arresting the cell cycle, such as the cyclin-dependent kinase inhibitor, p21. At present, the exact mechanism by which Fe chelators regulate p21 expression remains unclear. However, recent studies indicate the ability of chelators to up-regulate p21 at the mRNA level was dependent on the chelator and cell-type investigated. Analysis of the p21 promoter identified that the Sp1-3-binding site played a significant role in the activation of p21 transcription by Fe chelators. Furthermore, there was increased Sp1/ER-α and Sp1/c-Jun complex formation in melanoma cells, suggesting these complexes were involved in p21 promoter activation. Elucidating the mechanisms involved in the regulation of p21 expression in response to Fe chelator treatment in neoplastic cells will further clarify how these agents achieve their anti-tumor activity. It will also enhance our understanding of the complex roles p21 may play in neoplastic cells and lead to the development of more effective and specific anti-cancer therapies.

miR-93-5p suppresses cellular senescence by directly targeting Bcl-w and p21.

Cellular senescence, a distinctive type of irreversible growth arrest, develops in response to various stimuli. Bcl-w, an oncogene and member of the Bcl-2 family, has been reported to promote tumorigenicity in various cancer cells. Here, we sought to explore the potential role of Bcl-w in premature senescence, which has received relatively little research attention. Our findings demonstrate that Bcl-w enhances the activity of senescence-associated ß-galactosidase (SA-ß-gal) and promotes histone H3 tri-methylation at lysine 9 (H3K9me3) and expressions of p53, Notch2, p21, and p16-hallmarks of the senescent phenotype-in human U251 glioblastoma and H460 lung carcinoma cells. It is also known that microRNAs (miRNAs) regulate processes related to tumor development, such as cell proliferation, differentiation, survival, metabolism, inflammation, invasion, angiogenesis, and senescence. In this context, we found that miR-93-5p inhibited premature cellular senescence by directly suppressing Bcl-w and p21 expressions. Collectively, these findings suggest that targeting miR-93-5p-regulated Bcl-w may be a useful strategy for preventing premature senescence.

Upregulation of microRNA-17-5p contributes to hypoxia-induced proliferation in human pulmonary artery smooth muscle cells through modulation of p21 and PTEN.

BACKGROUND: Pulmonary arterial smooth muscle cell (PASMC) proliferation in response to hypoxia plays an important role in the vascular remodelling that occurs in hypoxic pulmonary hypertension. MicroRNAs (miRs) are emerging as important regulators in the progression of pulmonary hypertension. In this study, we investigated whether the expression of miR-17-5p is modulated by hypoxia and is involved in the hypoxia-induced proliferation of PASMCs. METHODS: Human PASMCs were cultured under hypoxic conditions. miR-17-5p expression was determined by real-time RT-PCR. A BrdU incorporation assay and time-lapse recording were utilized to determine cell proliferation and migration. RESULTS: PASMC proliferation was increased by moderate hypoxia (3% oxygen) but was reduced by severe hypoxia (0.1% oxygen) after 48 h. Moderate hypoxia induced miR-17-5p expression. Overexpression of miR-17-5p by transfection with miR-17-5p enhanced cell proliferation and migration in normoxia, whereas knockdown of miR-17-5p with anti-miR-17-5p inhibitors significantly reduced cell proliferation and migration. The expression of miR-17-5p target genes, specifically phosphatase and tensin homologue (PTEN) and cyclin-dependent kinase inhibitor 1 (p21WAF1/Cip1, p21), was reduced under moderate hypoxia in PASMCs. Under normoxia, overexpression of miR-17-5p in PASMCs reduced the expression of PTEN and p21. CONCLUSION: Our data indicate that miR-17-5p might play a significant role in hypoxia-induced pulmonary vascular smooth muscle cell proliferation by regulating multiple gene targets, including PTEN and p21, and that miR-17-5p could be a novel therapeutic target for the management of hypoxia-induced PH.