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Insufficient telomerase activity, stemming from low telomerase reverse transcriptase (TERT) gene transcription, contributes to telomere dysfunction and aging pathologies. Besides its traditional function in telomere synthesis, TERT acts as a transcriptional co-regulator of genes pivotal in aging and age-associated diseases. Here, we report the identification of a TERT activator compound (TAC) that upregulates TERT transcription via the MEK/ERK/AP-1 cascade. In primary human cells and naturally aged mice, TAC-induced elevation of TERT levels promotes telomere synthesis, blunts tissue aging hallmarks with reduced cellular senescence and inflammatory cytokines, and silences p16INK4a expression via upregulation of DNMT3B-mediated promoter hypermethylation. In the brain, TAC alleviates neuroinflammation, increases neurotrophic factors, stimulates adult neurogenesis, and preserves cognitive function without evident toxicity, including cancer risk. Together, these findings underscore TERT's critical role in aging processes and provide preclinical proof of concept for physiological TERT activation as a strategy to mitigate multiple aging hallmarks and associated pathologies.
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Envelhecimento , Metilação de DNA , Telomerase , Telomerase/metabolismo , Telomerase/genética , Humanos , Animais , Camundongos , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA (Citosina-5-)-Metiltransferases/genética , Senescência Celular , Regiões Promotoras Genéticas , DNA Metiltransferase 3B , Encéfalo/metabolismo , Telômero/metabolismo , Camundongos Endogâmicos C57BL , Masculino , Fator de Transcrição AP-1/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , NeurogêneseRESUMO
The human tumor suppressor p16INK4a is a small monomeric protein that can form amyloid structures. Formation of p16INK4a amyloid fibrils is induced by oxidation which creates an intermolecular disulfide bond. The conversion into amyloid is associated with a change from an all α-helical structure into ß-sheet fibrils. Currently, structural insights into p16INK4a amyloid fibrils are lacking. Here, we investigate the amyloid-forming regions of this tumor suppressor using isotope-labeling limited-digestion mass spectrometry analysis. We discover two key regions that likely form the structured core of the amyloid. Further investigations using thioflavin-T fluorescence assays, electron microscopy, and solution nuclear magnetic resonance spectroscopy of shorter peptide regions confirm the self-assembly of the identified sequences that include methionine and leucine repeat regions. This work describes a simple approach for studying protein motifs involved in the conversion of monomeric species into aggregated fibril structures. It provides insight into the polypeptide sequence underlying the core structure of amyloid p16INK4a formed after a unique oxidation-driven structural transition.
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Anterior cruciate ligament (ACL) injuries pose a significant challenge due to their limited healing potential, often resulting in premature arthritis. The factors and mechanisms contributing to this inadequate healing process remain elusive. During the acute phase of injury, ACL tissues express elevated periostin levels that decline over time. The functional significance of periostin in ligament biology remains understudied. In this study, we investigated the functional and mechanistic implications of periostin deficiency in ACL biology, utilizing ligament fibroblasts derived from patients and a murine model of ACL rupture. Our investigations unveiled that periostin knockdown compromised fibroblast growth characteristics, hindered the egress of progenitor cells from explants, and arrested cell-cycle progression, resulting in the accumulation of cells in the G0/G1 phase and moderate apoptosis. Concurrently, a significant reduction in the expression of cell-cycle and matrix-related genes was observed. Moreover, periostin deficiency triggered apoptosis through STAT3Y705/p38MAPK signaling and induced cellular senescence through increased production of reactive oxygen species (ROS). Mechanistically, inhibition of ROS production mitigated cell senescence in these cells. Notably, in vivo data revealed that ACL in Postn-/- mice exhibited a higher tearing frequency than wild-type mice under equivalent loading conditions. Furthermore, injured ACL with silenced periostin expression, achieved through nanoparticle-siRNA complex delivery, displayed an elevated propensity for apoptosis and senescence compared to intact ACL in C57BL/6 mice. Together, our findings underscore the pivotal role of periostin in ACL health, injury, and potential for healing.
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Lesões do Ligamento Cruzado Anterior , Ligamento Cruzado Anterior , Moléculas de Adesão Celular , Senescência Celular , Fibroblastos , Espécies Reativas de Oxigênio , Moléculas de Adesão Celular/metabolismo , Moléculas de Adesão Celular/genética , Animais , Camundongos , Fibroblastos/metabolismo , Senescência Celular/fisiologia , Humanos , Espécies Reativas de Oxigênio/metabolismo , Ligamento Cruzado Anterior/metabolismo , Lesões do Ligamento Cruzado Anterior/metabolismo , Lesões do Ligamento Cruzado Anterior/patologia , Apoptose , Camundongos Endogâmicos C57BL , Masculino , Fator de Transcrição STAT3/metabolismo , Feminino , Células Cultivadas , PeriostinaRESUMO
Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders in women of reproductive age, but its pathology has not been fully characterized and the optimal treatment strategy remains unclear. Cellular senescence is a permanent state of cell-cycle arrest that can be induced by multiple stresses. Senescent cells contribute to the pathogenesis of various diseases, owing to an alteration in secretory profile, termed 'senescence-associated secretory phenotype' (SASP), including with respect to pro-inflammatory cytokines. Senolytics, a class of drugs that selectively eliminate senescent cells, are now being used clinically, and a combination of dasatinib and quercetin (DQ) has been extensively used as a senolytic. We aimed to investigate whether cellular senescence is involved in the pathology of PCOS and whether DQ treatment has beneficial effects in patients with PCOS. We obtained ovaries from patients with or without PCOS, and established a mouse model of PCOS by injecting dehydroepiandrosterone. The expression of the senescence markers p16INK4a, p21, p53, γH2AX, and senescence-associated ß-galactosidase and the SASP-related factor interleukin-6 was significantly higher in the ovaries of patients with PCOS and PCOS mice than in controls. To evaluate the effects of hyperandrogenism and DQ on cellular senescence in vitro, we stimulated cultured human granulosa cells (GCs) with testosterone and treated them with DQ. The expression of markers of senescence and a SASP-related factor was increased by testosterone, and DQ reduced this increase. DQ reduced the expression of markers of senescence and a SASP-related factor in the ovaries of PCOS mice and improved their morphology. These results indicate that cellular senescence occurs in PCOS. Hyperandrogenism causes cellular senescence in GCs in PCOS, and senolytic treatment reduces the accumulation of senescent GCs and improves ovarian morphology under hyperandrogenism. Thus, DQ might represent a novel therapy for PCOS.
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Senescência Celular , Células da Granulosa , Síndrome do Ovário Policístico , Quercetina , Síndrome do Ovário Policístico/metabolismo , Síndrome do Ovário Policístico/patologia , Feminino , Senescência Celular/efeitos dos fármacos , Humanos , Animais , Células da Granulosa/metabolismo , Células da Granulosa/efeitos dos fármacos , Células da Granulosa/patologia , Quercetina/farmacologia , Camundongos , Fenótipo Secretor Associado à Senescência , Adulto , Dasatinibe/farmacologia , Modelos Animais de Doenças , Senoterapia/farmacologia , Hiperandrogenismo/patologia , Hiperandrogenismo/metabolismo , Interleucina-6/metabolismo , Desidroepiandrosterona/farmacologiaRESUMO
Tumor-associated macrophages (TAMs) are an important component of the tumor microenvironment (TME) and the most abundant population of immune cells infiltrating a tumor. TAMs can largely determine direction of anti-tumor immune response by promoting it or, conversely, contribute to formation of an immunosuppressive TME that allows tumors to evade immune control. Through interactions with tumor cells or other cells in the microenvironment and, as a result of action of anti-cancer therapy, macrophages can enter senescence. In this review, we have attempted to summarize information available in the literature on the role of senescent macrophages in tumors. With the recent development of senolytic therapeutic strategies aimed at removing senescent cells from an organism, it seems important to discuss functions of the senescent macrophages and potential role of the senolytic drugs in reprogramming TAMs to enhance anti-tumor immune response and improve efficacy of cancer treatment.
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Senescência Celular , Neoplasias , Microambiente Tumoral , Macrófagos Associados a Tumor , Microambiente Tumoral/imunologia , Humanos , Neoplasias/imunologia , Neoplasias/patologia , Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismo , Animais , Macrófagos/imunologia , Macrófagos/metabolismo , Biomarcadores Tumorais/metabolismoRESUMO
BACKGROUND: The association between p16INK4a and p21, a marker of cellular senescence, and the Immunoscore, an immunological prognostic indicator, in rectal cancer patients undergoing curative surgery were investigated. METHODS: A total of 82 patients who underwent curative surgery for rectal cancer were evaluated. The resected specimens were analyzed for p16INK4a, p21, CD3 and CD8 expression by immunohistochemistry. Immunoscore was calculated on the basis of CD3 and CD8 expressions. The clinicopathological characteristics and long-term outcomes were evaluated. RESULTS: Among the 82 patients, 24 (29.3%) were p16INK4a-positive and 11 (13.4%) were p21-positive. The patients were classified into the following five Immunoscore groups (IS0-5). IS0, IS1 and IS2 were classified as the low Immunoscore group (45 patients, 54.9%) and IS3 and IS4 as the high Immunoscore group (37 patients, 45.1%). There was no significant difference in age, sex, body mass index, American Society of Anesthesiologists physical status, depth of invasion of the tumor, lymph node metastasis and histological classification of the tumor with p16INK4a or p21 expression or Immunoscore. p16INK4a-positive expression and low Immunoscore each showed a tendency to indicate poor prognosis of disease-free survival (DFS). Patients with the combination of p16INK4a and p21 positivity and with p16INK4a positivity and low Immunoscore showed significantly poor prognosis of DFS. Patients with p21 positive positivity and low Immunoscore tended to have worse DFS. CONCLUSIONS: p16INK4a, p21 and Immunoscore may be prognostic indicators of rectal cancer. The combination of them may provide more accurate prognostic prediction than either factor alone.
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Biomarcadores Tumorais , Inibidor p16 de Quinase Dependente de Ciclina , Inibidor de Quinase Dependente de Ciclina p21 , Neoplasias Retais , Humanos , Masculino , Neoplasias Retais/patologia , Neoplasias Retais/imunologia , Neoplasias Retais/cirurgia , Feminino , Idoso , Pessoa de Meia-Idade , Prognóstico , Adulto , Idoso de 80 Anos ou mais , Intervalo Livre de Doença , Complexo CD3/metabolismo , Antígenos CD8/análise , Imuno-HistoquímicaRESUMO
Background and Objectives: Human papillomavirus (HPV) was previously investigated in lung cancer with wide inter-geographic discrepancies. p16INK4a has been used as a surrogate for detecting high-risk HPV (HR-HPV) in some cancer types. This study assessed the evidence of HPV in non-small-cell lung cancer (NSCLC) among Jordanian patients, investigated the expression of p16INK4a, and evaluated its prognostic value and association with HPV status. Materials and Methods: The archived samples of 100 patients were used. HPV DNA detection was performed by real-time polymerase chain reaction (RT-PCR). p16INK4a expression was assessed by immunohistochemistry (IHC). The Eighth American Joint Committee on Cancer protocol (AJCC) of head and neck cancer criteria were applied to evaluate p16INK4a positivity considering a moderate/strong nuclear/cytoplasmic expression intensity with a distribution in ≥75% of cells as positive. Results: HPV DNA was detected in 5% of NSCLC cases. Three positive cases showed HR-HPV subtypes (16, 18, 52), and two cases showed the probable HR-HPV 26 subtype. p16INK4a expression was positive in 20 (20%) NSCLC cases. None of the HPV-positive tumors were positive for p16INK4a expression. A statistically significant association was identified between p16INK4a expression and the pathological stage (p = 0.029) but not with other variables. No survival impact of p16INK4a expression was detected in NSCLC cases as a group; however, it showed a statistically significant association with overall survival (OS) in squamous cell carcinoma (SqCC) cases (p = 0.033). Conclusions: This is the first study to assess HPV and p16INK4a expression in a Jordanian population. HPV positivity is rare in NSCLC among a Jordanian subpopulation. P16 INK4a reliability as a surrogate marker for HPV infection in lung cancer must be revisited.
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Carcinoma Pulmonar de Células não Pequenas , Inibidor p16 de Quinase Dependente de Ciclina , Neoplasias Pulmonares , Infecções por Papillomavirus , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/virologia , Inibidor p16 de Quinase Dependente de Ciclina/genética , DNA Viral/análise , Papillomavirus Humano , Imuno-Histoquímica , Jordânia/epidemiologia , Neoplasias Pulmonares/virologia , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/genética , Prognóstico , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Laryngeal squamous cell carcinoma (LSCC) is a common malignancy that, despite scientific advancements, has not seen an improvement in its prognosis in the last decades. Few promising predictive markers have been found and none are relevant in clinical practice. p16ink4a, an oncosuppressor protein involved in cell cycle arrest, with a prognostic impact on other cancers, has been widely used in the head and neck region as a surrogate marker of HPV infection. Published papers and recent meta-analyses seem to minimize the biological role of HPV in the context of LSCC's cancerogenesis, and to disprove the reliability of p16ink4a as a surrogate prognostic marker in this context, while still highlighting its potential role as an independent predictor of survival. Unfortunately, the available literature, in particular during the last two decades, is often not focused on its potential role as an independent biomarker and few relevant data are found in papers mainly focused on HPV. The available data suggest that future research should focus specifically on p16ink4a, taking into account both its potential inactivation and overexpression, different patterns of staining, and immunohistochemistry cutoffs, and should focus not on its potential role as a surrogate marker but on its independent role as a predictor of survival.
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Age-dependent accumulation of amyloid plaques in patients with sporadic Alzheimer's disease (AD) is associated with reduced amyloid clearance. Older microglia have a reduced ability to phagocytose amyloid, so phagocytosis of amyloid plaques by microglia could be regulated to prevent amyloid accumulation. Furthermore, considering the aging-related disruption of cell cycle machinery in old microglia, we hypothesize that regulating their cell cycle could rejuvenate them and enhance their ability to promote more efficient amyloid clearance. First, we used gene ontology analysis of microglia from young and old mice to identify differential expression of cyclin-dependent kinase inhibitor 2A (p16ink4a), a cell cycle factor related to aging. We found that p16ink4a expression was increased in microglia near amyloid plaques in brain tissue from patients with AD and 5XFAD mice, a model of AD. In BV2 microglia, small interfering RNA (siRNA)-mediated p16ink4a downregulation transformed microglia with enhanced amyloid phagocytic capacity through regulated the cell cycle and increased cell proliferation. To regulate microglial phagocytosis by gene transduction, we used poly (D,L-lactic-co-glycolic acid) (PLGA) nanoparticles, which predominantly target microglia, to deliver the siRNA and to control microglial reactivity. Nanoparticle-based delivery of p16ink4a siRNA reduced amyloid plaque formation and the number of aged microglia surrounding the plaque and reversed learning deterioration and spatial memory deficits. We propose that downregulation of p16ink4a in microglia is a promising strategy for the treatment of Alzheimer's disease.
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Doença de Alzheimer , Idoso , Animais , Humanos , Camundongos , Doença de Alzheimer/metabolismo , Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Modelos Animais de Doenças , Camundongos Transgênicos , Microglia/metabolismo , Placa Amiloide/metabolismo , RNA Interferente PequenoRESUMO
PURPOSE: The ocular surface microenvironment changes with aging. However, it remains unclear if cellular senescence influences the ocular surface. We investigated the presence of p16INK4a-expressing senescent cells in healthy human conjunctiva. STUDY DESIGN: Clinical and experimental. METHODS: Healthy conjunctival tissue samples were obtained from middle-aged and elderly subjects. RT-qPCR was performed to assess the expression of senescence markers CDKN2A (p16INK4a) and CDKN1A (p21CIP1/WAF1) and immunostaining was performed to examine the expression of the senescence marker p16INK4a, stem cell markers Ki67 and p63, tight-junction marker ZO-1. RESULTS: Our study involved 19 conjunctival tissue samples (10 elderly and 9 middle-aged), mean age [elderly: 75.8 ± 3.7 years (72-81), middle-aged: 52.7 ± 7 years (38-59)], sex (elderly: 3 men, 7 women; middle-aged: 3 men, 6 women). The expression of p16INK4a was significantly increased at the RNA level in the elderly compared to middle-aged (p < 0.05). Positivity rate of p16INK4a was significantly elevated in the elderly (15.0 ± 7.8%) compared to middle-aged (0.2 ± 0.6%) (p < 0.05). Positivity rate of Ki67and p63 was significantly reduced in the elderly (1.7 ± 1.7% and 16.5 ± 9.5%) compared to middle-aged (3.9 ± 1.8% and 24.7 ± 5.7%) (p < 0.05). ZO-1 expression was reduced in tissue samples showing p16INK4a-positivity but retained in tissue samples in which p16INK4a was undetectable. CONCLUSIONS: Senescent cells accumulate with age in the conjunctival epithelium, accompanied by a decrease in Ki67, p63 and ZO-1 expressing cells.
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Envelhecimento , Inibidor p16 de Quinase Dependente de Ciclina , Idoso , Pessoa de Meia-Idade , Masculino , Humanos , Feminino , Idoso de 80 Anos ou mais , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/análise , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Antígeno Ki-67 , Senescência Celular , Epitélio/química , Epitélio/metabolismoRESUMO
NIPP1 is a ubiquitously expressed regulatory subunit of PP1. Its embryonic deletion in keratinocytes causes chronic sterile skin inflammation, epidermal hyperproliferation, and resistance to mutagens in adult mice. To explore the primary effects of NIPP1 deletion, we first examined hair cycle progression of NIPP1 skin knockouts (SKOs). The entry of the first hair cycle in the SKOs was delayed owing to prolonged quiescence of hair follicle stem cells. In contrast, the entry of the second hair cycle in the SKOs was advanced as a result of precocious activation of hair follicle stem cells. The epidermis of SKOs progressively accumulated senescent cells, and this cell-fate switch was accelerated by DNA damage. Primary keratinocytes from SKO neonates and human NIPP1-depleted HaCaT keratinocytes failed to proliferate and showed an increase in the expression of cell cycle inhibitors (p21, p16/Ink4a, and/or p19/Arf) and senescence-associated-secretory-phenotype factors as well as in DNA damage (γH2AX and 53BP1). Our data demonstrate that the primary effect of NIPP1 deletion in keratinocytes is a cell cycle arrest and premature senescence that gradually progresse to chronic senescence and likely contribute to the decreased sensitivity of SKOs to mutagens.
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Cathepsin L (CTSL) has been implicated in aging and age-related diseases, such as cardiovascular diseases, specifically atherosclerosis. However, the underlying mechanism(s) is not well documented. Recently, we demonstrated a role of CUT-like homeobox 1 (CUX1) in regulating the p16INK4a-dependent cellular senescence in human endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) via its binding to an atherosclerosis-associated functional SNP (fSNP) rs1537371 on the CDKN2A/B locus. In this study, to determine if CTSL, which was reported to proteolytically activate CUX1, regulates cellular senescence via CUX1, we measured the expression of CTSL, together with CUX1 and p16INK4a, in human ECs and VSMCs undergoing senescence. We discovered that CUX1 is not a substrate that is cleaved by CTSL. Instead, CTSL is an upstream regulator that activates CUX1 transcription indirectly in a process that requires the proteolytic activity of CTSL. Our findings suggest that there is a transcription factor in between CTSL and CUX1, and cleavage of this factor by CTSL can activate CUX1 transcription, inducing endothelial senescence. Thus, our findings provide new insights into the signal transduction pathway that leads to atherosclerosis-associated cellular senescence.
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Catepsina L , Senescência Celular , Inibidor p16 de Quinase Dependente de Ciclina , Proteínas de Homeodomínio , Músculo Liso Vascular , Proteínas Repressoras , Humanos , Senescência Celular/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Catepsina L/metabolismo , Catepsina L/genética , Proteínas Repressoras/metabolismo , Proteínas Repressoras/genética , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/citologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Regulação para Cima , Células Endoteliais/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Miócitos de Músculo Liso/metabolismo , Aterosclerose/metabolismo , Aterosclerose/genética , Aterosclerose/patologia , Células Endoteliais da Veia Umbilical Humana , Células CultivadasRESUMO
Introduction: Tissue expression of P16ink4A is correlated with cervical lesions. In this study we determined the association between serum P16ink4A concentrations and cervical lesions among women attending the cervical cancer clinic at Mbarara Regional Hospital (MRRH) South Western Uganda. Material and Methods: We recruited 90 cervical intraepithelial neoplasia (CIN) cases, 90 cervical cancer (CC) cases before treatment and 90 controls. Clinical and demographic data were recorded. Serum P16ink4A concentrations were measured by quantitative Elisa. Cases were confirmed with cytology and/or histology. Descriptive statistics and logistic regression were done with STATA 17 and P-values of <0.05 were considered statistically significant. Results: The mean serum P16ink4A concentration among CIN cases, CC cases and controls was 1.11(+/-0.66) ng/ml, 1.45(+/-1.11) ng/ml and 1.13(+/-0.61) ng/ml respectively (p = 0.008). 50 % of CIN cases and controls as well as 60 % of CC cases had P16ink4A concentration above 0.946 ng/ml. There were increased odds of CIN for serum P16ink4A though statistically insignificant (AOR: 1.11, p-value: 0.70). There was also a statistically significant reduction in odds of CC for serum P16ink4A (AOR: 0.55, p-value: 0.01). Conclusion: Serum P16ink4A may likely be associated with cervical lesions especially CC in our study population and this may aid detection of such lesions. Diagnostic utility studies for circulating P16ink4A in detection of cervical cancer are recommended.
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Contrast-induced acute kidney injury (CI-AKI) is the third leading cause of hospital-acquired acute kidney injury. Cellular senescence is associated with CI-AKI. P16INK4a (p16) is a cell cycle regulator and link to aging and senescence. We found that the expression of p16 was elevated in CI-AKI renal tissues, however its role in CI-AKI remains insufficiently understood. In this study, we used p16 knockout (p16KO) mice and wild-type (WT) littermates to establish CI-AKI mice model to elucidate the impact of p16 on CI-AKI. The results showed that serum creatinine (SCr), blood urea nitrogen (BUN), and serum neutrophil gelatinase-associated lipocalin (NGAL) levels were markedly reduced in p16KO CI-AKI mice. Both immunohistochemistry and western blot analyses confirmed that p16 knockout alleviated renal cell apoptosis. Furthermore, interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α (TNF-α) were attenuated by downregulating NLRP3 and NF-κB inflammasomes. Additionally, ROS levels were diminished via activating Nrf2/Keap-1 pathway in p16KO CI-AKI mice. Collectively, our findings suggest that p16 deletion exerts protective effects against apoptosis, inflammation, and oxidative stress in CI-AKI mice model, p16 deletion might be a potential therapeutic strategy for ameliorating CI-AKI.
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Injúria Renal Aguda , Meios de Contraste , Inibidor p16 de Quinase Dependente de Ciclina , Animais , Camundongos , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/genética , Injúria Renal Aguda/prevenção & controle , Apoptose , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inflamação/metabolismo , Rim/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo , Meios de Contraste/efeitos adversosRESUMO
Higher intensity exercise, despite causing more tissue damage, improved aging conditions. We previously observed decreased p16INK4a mRNA in human skeletal muscle after high-intensity interval exercise (HIIE), with no change following equivalent work in moderate-intensity continuous exercise. This raises the question of whether the observed senolytic effect of exercise is mediated by inflammation, an immune response induced by muscle damage. In this study, inflammation was blocked using a multiple dose of ibuprofen (total dose: 1200 mg), a commonly consumed nonsteroidal anti-inflammatory drug (NSAID), in a placebo-controlled, counterbalanced crossover trial. Twelve men aged 20-26 consumed ibuprofen or placebo before and after HIIE at 120% maximum aerobic power. Multiple muscle biopsies were taken for tissue analysis before and after HIIE. p16INK4a+ cells were located surrounding myofibers in muscle tissues. The maximum decrease in p16INK4a mRNA levels within muscle tissues occurred at 3 h post-exercise (-82%, p < 0.01), gradually recovering over the next 3-24 h. A concurrent reduction pattern in CD11b mRNA (-87%, p < 0.01) was also found within the same time frame. Ibuprofen treatment attenuated the post-exercise reduction in both p16INK4a mRNA and CD11b mRNA. The strong correlation (r = 0.88, p < 0.01) between p16INK4a mRNA and CD11b mRNA in muscle tissues suggests a connection between the markers of tissue aging and pro-inflammatory myeloid differentiation. In conclusion, our results suggest that the senolytic effect of high-intensity exercise on human skeletal muscle is mediated by acute inflammation.
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Anti-Inflamatórios não Esteroides , Estudos Cross-Over , Ibuprofeno , Inflamação , Músculo Esquelético , Humanos , Masculino , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Adulto , Ibuprofeno/farmacologia , Inflamação/metabolismo , Adulto Jovem , Anti-Inflamatórios não Esteroides/farmacologia , Exercício Físico/fisiologia , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Antígeno CD11b/metabolismo , Antígeno CD11b/genética , RNA Mensageiro/metabolismo , Treinamento Intervalado de Alta IntensidadeRESUMO
DNA methylation is one of the most important epigenetic mark involved in many physiologic cellular processes and pathologies. During mitosis, the transmission of DNA methylation patterns from a mother to the daughter cells is ensured through the action of the Ubiquitin-like, containing PHD and RING domains, 1/DNA methyltransferase 1 (UHRF1/DNMT1) tandem. UHRF1 is involved in the silencing of many tumor suppressor genes (TSGs) via mechanisms that remain largely to be deciphered. The present study investigated the role and the regulation of UHRF1 poly-ubiquitination induced by thymoquinone, a natural anti-cancer drug, known to enhance or re-activate the expression of TSGs. We found that the auto-ubiquitination of UHRF1, induced by TQ, is mediated by reactive oxygen species, and occurs following DNA damage. We demonstrated that the poly-ubiquitinated form of UHRF1 is K63-linked and can still silence the tumor suppressor gene p16INK4A/CDKN2A. We further showed that TQ-induced auto-ubiquitination is mediated via the activity of Tip60. Since this latter is known as a nuclear receptor co-factor, we investigated if the glucocorticoid receptor (GR) might be involved in the regulation of UHRF1 ubiquitination. Activation of the GR, with dexamethasone, did not influence auto-ubiquitination of UHRF1. However, we could observe that TQ induced a K48-linked poly-ubiquitination of GR, probably involved in the proteosomal degradation pathway. Mass-spectrometry analysis of FLAG-HA-tagged UHRF1 identified UHRF1 partners involved in DNA repair and showed that TQ increased their association with UHRF1, suggesting that poly-ubiquitination of UHRF1 is involved in the DNA repair process. We propose that poly-ubiquitination of UHRF1 serves as a scaffold to recruit the DNA repair machinery at DNA damage sites.
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Benzoquinonas , Proteínas Estimuladoras de Ligação a CCAAT , Reparo do DNA , Ubiquitina-Proteína Ligases , Ubiquitinação , Humanos , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/genética , Ubiquitinação/efeitos dos fármacos , Benzoquinonas/farmacologia , Reparo do DNA/efeitos dos fármacos , Antineoplásicos/farmacologia , Dano ao DNA/efeitos dos fármacosRESUMO
Aging progresses through the interaction of metabolic processes, including changes in the immune and endocrine systems. Glucocorticoids (GCs), which are regulated by the hypothalamic-pituitary-adrenal (HPA) axis, play an important role in regulating metabolism and immune responses. However, the age-related changes in the secretion mechanisms of GCs remain elusive. Here, we found that corticosterone (CORT) secretion follows a circadian rhythm in young mice, whereas it oversecreted throughout the day in aged mice >18 months old, resulting in the disappearance of diurnal variation. Furthermore, senescent cells progressively accumulated in the zF of the adrenal gland as mice aged beyond 18 months. This accumulation was accompanied by an increase in the number of Ad4BP/SF1 (SF1), a key transcription factor, strongly expressing cells (SF1-high positive: HP). Removal of senescent cells with senolytics, dasatinib, and quercetin resulted in the reduction of the number of SF1-HP cells and recovery of CORT diurnal oscillation in 24-month-old mice. Similarly, administration of a neutralizing antibody against IL1ß, which was found to be strongly expressed in the adrenocortical cells of the zF, resulted in a marked decrease in SF1-HP cells and restoration of the CORT circadian rhythm. Our findings suggest that the disappearance of CORT diurnal oscillation is a characteristic of aging individuals and is caused by the secretion of IL1ß, one of the SASPs, from senescent cells that accumulate in the zF of the adrenal cortex. These findings provide a novel insight into aging. Age-related hypersecretory GCs could be a potential therapeutic target for aging-related diseases.
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Bladder cancer (BlCa) is an extensively heterogeneous disease that leads to great variability in tumor evolution scenarios and lifelong patient surveillance, emphasizing the need for modern, minimally invasive precision medicine. Here, we explored the clinical significance of copy number alterations (CNAs) in BlCa. CNA profiling was performed in 15 patient-derived xenografts (PDXs) and validated in The Cancer Genome Atlas BlCa (TCGA-BLCA; n = 408) and Lindgren et al. (n = 143) cohorts. CDKN2A copy number loss was identified as the most frequent CNA in bladder tumors, associated with reduced CDKN2A expression, tumors of a papillary phenotype, and prolonged PDX survival. The study's screening cohort consisted of 243 BlCa patients, and CDKN2A copy number was assessed in genomic DNA and cell-free DNA (cfDNA) from 217 tumors and 189 pre-treatment serum samples, respectively. CDKN2A copy number loss was correlated with superior disease-free and progression-free survival of non-muscle-invasive BlCa (NMIBC) patients. Moreover, a higher CDKN2A index (CDKN2A/LEP ratio) in pre-treatment cfDNA was associated with advanced tumor stage and grade and short-term NMIBC progression to invasive disease, while multivariate models fitted for CDKN2A index in pre-treatment cfDNA offered superior risk stratification of T1/high-grade and EORTC high-risk patients, enhancing prediction of treatment outcome. CDKN2A copy number status could serve as a minimally invasive tool to improve risk stratification and support personalized prognosis in BlCa.
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Accumulation of senescent cells is a recognized feature in hepatocellular carcinoma (HCC), but their specific types and prognostic implications remain under investigation. This study aimed to delineate senescent cell types and their senescent patterns in HCC using publicly available bulk and single-cell mRNA sequencing data. Through gene expression and gene set enrichment analysis, we identified distinct senescent patterns within HCC samples. Notably, unconventional T cells, specifically natural killer T cells and γδT cells, were found to be the predominant senescent cell types. These cells exhibited enriched pathways related to DNA damage, senescence and the negative regulation of lymphocyte activation. Furthermore, we observed upregulation of the mTOR signaling pathway, which correlated positively with the expression of senescence-associated genes. This suggests a potential regulatory role for mTOR in the senescence of HCC. Strikingly, patients with elevated expression of senescence markers, including p16INK4A, p21, and GLB1, demonstrated significantly reduced overall survival rates. Our findings indicate that immunosenescence in unconventional T cells may play a role in HCC progression. The potential therapeutic implications of targeting the mTOR pathway or eliminating senescent unconventional T cells warrant further exploration to improve HCC patient outcomes.
RESUMO
p16INK4a and p21WAF1/Cip1 are cyclin-dependent kinase inhibitors involved in cell cycle control, which can function as oncogenes or tumor suppressors, depending on the context of various extracellular and intracellular signals, and cell type. In human papillomavirus-induced cervical cancer, p16 INK4a shows oncogenic activity and functions as a diagnostic marker of cervical neoplasia, whereas p21 WAF1/Cip1 acts as a tumor suppressor and its downregulation is associated with the progression of malignant transformation. Several histone deacetylase (HDAC) inhibitors promote the positive and negative regulation of a number of genes, including p16 INK4a and p21 WAF1/Cip1; however, the effects of sodium valproate (VPA) on these genes and on the proteins they encode remain uncertain in HeLa cervical cancer cells. In the present study, these effects were investigated in HeLa cells treated with 0.5 or 2 mM VPA for 24 h, using reverse transcription-quantitative PCR, confocal microscopy and western blotting. The results revealed a decrease in the mRNA expression levels of p16 INK4a and a tendency for p16INK4a protein abundance to decrease in the presence of 2 mM VPA. By contrast, an increase in the protein expression levels of p21WAF1/Cip1 was detected in the presence of 0.5 and 2 mM VPA. Furthermore, VPA was confirmed to inhibit HDAC activity and induce global hyperacetylation of histone H3. Notably, VPA was shown to suppress p16 INK4a, a biomarker gene of cervical carcinoma, and to increase the abundance of the tumor suppressor protein p21WAF1/Cip1, thus contributing to the basic knowledge regarding the antitumorigenic potential of VPA. Exploration of epigenetic changes associated with the promoters of p16 INK4a and p21 WAF1/Cip1, such as histone H3 methylation, may provide further information and improve the understanding of these findings.