p16INK4a Protein Expression Research Articles

Metallophosphoesterase-Domain-Containing Protein 2 (MPPED2) Expression in High-Risk Human Papilloma Virus-Induced Cervical Carcinoma and Its Correlation With p16INK4A Protein.

Introduction Recently, the expression of metallophosphoesterase-domain-containing protein 2 (MPPED2) was identified in cervical cancer. However, its precise role and correlation with other tumor suppressor proteins, such as p16INK4A, is not well studied in high-risk human papillomavirus (HPV) integrated human cervical carcinoma. Hence, in the present study, we try to see the expression of MPPED2 in human cervical carcinoma and its correlation with age and p16INK4A protein expression level. Methods The prospective study consists of 200 samples of 150 known cervical carcinoma and 50 controls. Histopathological evaluation, immunohistochemical staining, and semi-quantitative scoring of the intensity of proteins were performed. Statistical analysis was performed with the Shapiro-Wilk test, Spearman's rho correlation sig. (two-tailed), and Student's t-test. Results The data show that among the 150 cases, 136 (68.0%) cervical carcinoma tissues express the presence of high-risk HPV viral genome integration in the host cell. The expression of p16INK4A protein is higher in those tissues identified with high-risk HPV viral genomes. In contrast, the expression of MPPED2 protein is lesser or absent in those cervical tissues that have the higher expression of p16INK4A protein and vice versa. There is a significant correlation (p=0.000) between age and p16INK4A protein expression but not with MPPED2. A significant linear correlation (p=0.000) is found between the p16INK4A and MPPED2 proteins. Conclusion It may support the therapeutic application of MPPED2 protein to prevent cervical carcinoma progression in the near future.

Abstract A063: What can we learn from a retrospective evaluation of HER2, P53 and P16INK4A protein expression in rare ovarian tumors?

Abstract A063: What can we learn from a retrospective evaluation of HER2, P53 and P16INK4A protein expression in rare ovarian tumors?

FISH analysis reveals CDKN2A and IFNA14 co-deletion is heterogeneous and is a prominent feature of glioblastoma.

Deletion of CDKN2A occurs in 50% of glioblastomas (GBM), and IFNA locus deletion in 25%. These genes reside closely on chromosome 9. We investigated whether CDKN2A and IFNA were co-deleted within the same heterogeneous tumour and their prognostic implications. We assessed CDKN2A and IFNA14 deletions in 45 glioma samples using an in-house three-colour FISH probe. We examined the correlation between p16INK4a protein expression (via IHC) and CDKN2A deletion along with the impact of these genomic events on patient survival. FISH analyses demonstrated that grades II and III had either wildtype (wt) or amplified CDKN2A/IFNA14, whilst 44% of GBMs harboured homozygous deletions of both genes. Cores with CDKN2A homozygous deletion (n = 11) were negative for p16INK4a. Twenty p16INK4a positive samples lacked CDKN2A deletion with some of cells showing negative p16INK4a. There was heterogeneity in IFNA14/CDKN2A ploidy within each GBM. Survival analyses of primary GBMs suggested a positive association between increased p16INK4a and longer survival; this persisted when considering CDKN2A/IFNA14 status. Furthermore, wt (intact) CDKN2A/IFNA14 were found to be associated with longer survival in recurrent GBMs. Our data suggest that co-deletion of CDKN2A/IFNA14 in GBM negatively correlates with survival and CDKN2A-wt status correlated with longer survival, and with second surgery, itself a marker for improved patient outcomes.

THE STUDY OF PROGNOSTIC VALUE OF microRNAs (miR-10b AND -155) AND CDKN2A/P16INK4A IN ORAL SQUAMOUS CELL CARCINOMA.

Oral squamous cell carcinoma (OSCC) is one of the most common types of cancer worldwide. Disco- vering novel prognostic markers for OSCC can improve treatment outcomes by allowing for more effective therapy strategies. To identify the prognostic value of CDKN2A (p16INK4a) and miRNAs involved in its regulation as markers of OSCC. The work is based on the results of the examination and treatment of 70 patients with stage II-IV OSCC. miR-10b, -155, and CDKN2A mRNA expression in tumor samples was ana- lyzed by real-time reverse transcription polymerase chain reaction. The expression of p16INK4a and Ki-67 proteins was determined immunohistochemically. No association of CDKN2A mRNA and p16INK4a protein with Ki-67 expression in tumor tissue and clinical pathological parameters of OSCC patients was found. Most of the p16INK4a-positive cases were characterized by a high Ki-67 expression. We found a strong correlation of the studied miRNAs expression levels with lymph node metastasis (r = 0.56 for miR-10b and r = 0.59 for miR-155). Also, there was no difference in miR-10b and -155 expression between p16INK4a+ and p16INK4a- samples. The association of both miRNAs with lymph node metastases was not affected by p16INK4a status. The results indicate the relationship between miR-10b and -155 and the presence of lymph node metastases in OSCC patients, so these miRNAs can be considered as prognostic markers of the disease.

FISH-BASED INVESTIGATION OF CDKN2A AND IFNA14 IN GLIOBLASTOMA

Abstract AIMS Glioblastoma (GBM) tumours have a dismal prognosis despite aggressive anticancer therapy. Deletion of CDKN2A is among the most common genetic changes in GBM (~50%) and has been strongly associated with worse prognosis. Next generation DNA sequencing had shown that deletion of IFNA genes (located proximal to CDKN2A) is also a major genetic event in GBM (~25%). Clinical Identification of in vivo state of these two genes might facilitate improved individualized therapeutics. METHOD Glioma tissue microarray consisting of 45 samples was used to assess the co-deletion of CDKN2A and IFNA14 genes simultaneously, using a novel three-color fluorescence in situ hybridization (FISH) probe. We examined the correlation between CDKN2A and p16INK4a protein expression detected using immunohistochemistry (IHC). RESULTS FISH analysis showed that 44% of the primary GBMs harboured homozygous deletions of both CDKN2a and INFA14. By contrast, all grade II and III gliomas, had either wild-type or amplified CDKN2A and INFA14. All samples that showed CDKN2A homozygous deletion (n=11) were negative for p16INK4a staining, while 20 cores that were positive for p16INK4a expression did not harbour CDKN2A deletion. Survival curves of primary GBMs suggested that both the co-deletion of CDKN2A/INFA14 and negative p16INK4a expression were associated with shorter survival time. CONCLUSIONS Our FISH/IHC analyses indicate a strong correlation between CDKN2A and p16INK4a protein expression in grade III/II glioma, suggesting that amplification of CDKN2A might act as a gatekeeper to reduce the incidence of grade IV. Our data also highlight the high frequency of CDKN2A/INFA14 co-deletion in GBM tumours that might together impact on survival time.

Programmed death-ligand 1 and p53 as a biomarker in predicting oropharyngeal squamous cell carcinoma

Introduction. Currently, the pathogenetic role of the human papillomavirus (HPV) in carcinogenesis is well studied, that most cases of oropharyngeal squamous cell carcinoma are associated with the persistence of this infection. In addition to HPV infection, the involvement of programmed death-ligand 1 (PD-L1) and p53 proteins in the initiation and progression of oropharyngeal squamous cell carcinoma provides their formation as additional biomarkers in predicting the disease.Aim. To study the prognostic role of PD-L1 and p53 expression in oropharyngeal squamous cell carcinoma depending on HPV status.Materials and methods. The study included 62 patients with oropharyngeal squamous cell carcinoma T1–4N0–3M0 (Tumor, Nodus and Metastasis, (TNM), 7th edition) treated in 2015–2020 at the Republican Specialized Scientific and Practical Medical Center of Oncology and Radiology and in the Tashkent and Samarkand city branches. All patients underwent immunohistochemical analysis for the presence of p16INK4a, PD-L1 and p53 protein expression in formalin-fixed and paraffin-embedded tumor tissue samples.Results. Based on the results of the assessment, it was found that positive HPV status, low and medium levels of PD-L1 expression and positive regulation of wild-type p53 are associated with favorable outcomes in patients with oropharyngeal squamous cell carcinoma. Negative HPV status, high and very high levels of PD-L1 expression, loss of wild type p53 function with upregulation of p53 mutant type leads to worse disease outcomes.Conclusion. Thus, the assessment of the expression of the onco-suppressor protein p53 helps to determine the biology of cancer cells in patients with oropharyngeal squamous cell carcinoma and suggest sensitivity to ongoing therapy, and by studying the regulation of the PD-L1 protein, antitumor immune regulation can be suggested in these patients.

Senescent renal tubular epithelial cells activate fibroblasts by secreting Shh to promote the progression of diabetic kidney disease.

Diabetic kidney disease (DKD) is one of the complications of diabetes; however, the pathogenesis is not yet clear. A recent study has shown that senescence is associated with the course of DKD. In the present study, we explored whether senescent renal tubular cells promote renal tubulointerstitial fibrosis by secreting Sonic hedgehog (Shh) which mediates fibroblast activation and proliferation in DKD. A 36-week-old db/db mice model and the renal tubular epithelial cells were cultured in high glucose (HG, 60 mmol/L) medium for in vivo and in vitro experiments. Compared to db/m mice, blood glucose, microalbuminuria, serum creatinine, urea nitrogen, and UACR (microalbuminuria/urine creatinine) were markedly increased in db/db mice. Collagen III, monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) were also increased in db/db mice kidneys, suggesting fibrosis and inflammation in the organ. Moreover, the detection of SA-β-galactosidase (SA-β-Gal) showed that the activity of SA-β-Gal in the cytoplasm of renal tubular epithelial cells increased, and the cell cycle inhibition of the expression of senescence-related gene cell cycle inhibitor p16 INK4A protein and p21 protein increased, indicating that renal fibrosis in db/db mice was accompanied by cell senescence. Furthermore, Shh is highly expressed in the injured renal tubules and in the kidney tissue of db/db mice, as detected by enzyme-linked immunosorbent assay (ELISA). The results of immunofluorescence staining showed increased positive staining for Shh in renal tubular epithelial cells of db/db mice and decreased positive staining for Lamin B1, but increased positive staining for γH2A.X in cells with high Shh expression; similar results were obtained in vitro. In addition, HG stimulated renal tubular epithelial cells to secrete Shh in the supernatant of the medium. D-gal treatment of renal tubular epithelial cells increased the protein levels of Shh and p21. We also found enhanced activation and proliferation of fibroblasts cultured with the supernatant of renal tubular epithelial cells stimulated by HG medium but the proliferative effect was significantly diminished when co-cultured with cyclopamine (CPN), an inhibitor of the Shh pathway. In conclusion, HG induces renal tubular epithelial cell senescence, and the secretion of senescence-associated proteins and Shh mediates inflammatory responses and fibroblast activation and proliferation, ultimately leading to renal fibrosis.

Bioactive Oligopeptides from Ginseng (Panax ginseng Meyer) Suppress Oxidative Stress-Induced Senescence in Fibroblasts via NAD+/SIRT1/PGC-1α Signaling Pathway.

The physicochemical properties and multiple bioactive effects of ginseng oligopeptides (GOPs), plant-derived small molecule bioactive peptides, suggest a positive influence on health span and longevity. Given this, cellular senescence is the initiating factor and key mechanism of aging in the organism, and thus the current study sought to explore the effects of GOPs on H2O2-induced cellular senescence and its potential mechanisms. Senescence was induced in mouse embryonic fibroblasts NIH/3T3 by 4 h of exposure to 200 µM H2O2 and confirmed using CCK-8 assay and Western blot analyses of p16INK4A and p21Waf1/Cip1 after 24 h of growth medium administration with or without GOPs supplementation (25, 50, and 100 µg/mL). We found that GOPs delayed oxidative stress-induced NIH/3T3 senescence by inhibiting the G1 phase arrest, increasing DNA synthesis in the S phase, decreasing the relative protein expression of p16INK4A and p21Waf1/Cip1, promoting cell viability, protecting DNA, and enhancing telomerase (TE) activity. Further investigation revealed that the increase in antioxidative capacity and anti-inflammation capacity might form the basis for the retarding of the senescence effects of GOPs. Furthermore, GOPs supplementation significantly improved mitochondrial function and mitochondrial biogenesis via the NAD+/SIRT1/PGC-1𝛼 pathway. These findings indicate that GOPs may have a positive effect on health span and lifespan extension via combating cellular senescence, oxidative stress, and inflammation, as well as modulating longevity regulating pathway NAD+/SIRT1/PGC-1𝛼.

Virus-associated chronic endometritis: treatment options

Aim. To evaluate the effectiveness of Alloferon (Allokin-alfa) in the complex treatment of virus-associated chronic endometritis (CE) in patients with infertility, papillomavirus infection (PVI) persisting in the uterine cavity, and recurrent herpes-virus infection localized in the genital area.
 Materials and methods. A prospective (n=33) open randomized (2:1) study was conducted to assess the efficacy of Alloferon (Allokin-alfa) in the complex treatment of CE in patients with infertility, PVI, and recurrent herpes-virus infection, aged 25 to 37 years (median age 31 [29; 32.5] years). All patients received valacyclovir therapy at 500 mg once daily for 30 days from the day of randomization. Patients in the main group (n=21) simultaneously with the start of antiviral therapy received Allokin-alfa as 9 subcutaneous injections once every two days (one injection every other day). The uterine cavity microbiota of the patients was assessed 3 months after treatment, and histological and immunohistochemical studies of endometrial biopsy specimens were performed.
 Results. The microbiological data analysis showed HPV elimination in 71.4% vs 16.7% of patients in the alloferon and control groups, respectively (2 7.102, p=0.008). Also, in the main group, a significant decrease in the severity of CE (2 27.586, p0.001) and p16ink4a protein expression levels (2 6.17, p=0.013) were observed.
 Conclusion. In the treatment of virus-associated CE, the addition of alloferon to virus-suppressive therapy leads to higher rates of HPV elimination from the uterine cavity and significantly reduces the severity of CE.

Upregulation of p16INK4A in Peripheral White Blood Cells as a Novel Screening Marker for Colorectal Carcinoma.

Screening of colorectal cancer (CRC) is important for the early detection. CRC is relating to aging and immuno-senescence. One such senescent marker is p16INK4A expression in immune cells. The objective of the study is to investigate the protein expression of p16INK4A in peripheral white blood cells as a screening marker for colorectal cancer. A case-control studies were conducted. Cases were patients with colorectal cancer and controls were matched with cases based on age and sex. Peripheral blood was collected from patients and controls and the protein p16INK4A was measured with immunofluorescent techniques. The p16INK4A levels from cases and controls were evaluated using ROC analysis to be used as a screening marker in CRC patients. Mean fluorescent intensity of p16INK4A of cases and controls were analyzed in CD45+, CD3+ or CD14+ cells. The p16INK4A levels of cases were also correlated with clinical data. Statistically significant increased expression of p16INK4A levels were found in cases compared to controls. p16INK4A in peripheral immune cells had 78% sensitivity and 71% specificity which can possibly be used as a diagnosis tool for colorectal cancer. P16INK4A-positive cell percentage and mean florescent intensity were significantly higher in CD45+ cells, CD3 positive cells and CD14 positive cells. No significant correlation was observed with the clinical data and p16INK4A level of CRC patients. The significant increase of p16 INK4A expression level in peripheral immune cells represents potential for use as a CRC screening marker.

Delayed effects of radiation in adipose tissue reflect progenitor damage and not cellular senescence.

The pathogenesis of many age-related diseases is linked to cellular senescence, a state of inflammation-inducing, irreversible cell cycle arrest. The consequences and mechanisms of age-associated cellular senescence are often studied using in vivo models of radiation exposure. However, it is unknown whether radiation induces persistent senescence, like that observed in ageing. We performed analogous studies in mice and monkeys, where young mice and rhesus macaques received sub-lethal doses of ionizing radiation and were observed for ~ 15% of their expected lifespan. Assessments of 8-hydroxy-2' -deoxyguanosine (8-OHdG), senescence-associated beta-galactosidase (SAβ-gal), and p16Ink4a and p21 were performed on mitotic and post-mitotic tissues - liver and adipose tissue - 6months and 3years post-exposure for the mice and monkeys, respectively. No elevations in 8-OHdG, SA-βgal staining, or p16 Ink4a or p21 gene or protein expression were found in mouse and monkey liver or adipose tissue compared to control animals. Despite no evidence of senescence, progenitor cell dysfunction persisted after radiation exposure, as indicated by lower in situ CD34+ adipose cells (p = 0.03), and deficient adipose stromal vascular cell proliferation (p < 0.05) and differentiation (p = 0.04) ex vivo. Our investigation cautions that employing radiation to study senescence-related processes should be limited to the acute post-exposure period and that stem cell damage likely underpins the dysfunction associated with delayed effects of radiation.

Significance of Combined Analysis of High-Risk Human Papillomaviruses Polymerized Chain Reaction Analysis and Immunohistochemical Expression of p16INK4A in Cervical Cancer in a Cohort of South-Indian Population.

IntroductionCervical cancer is the fourth most frequent cancer in women worldwide, and it continues to be a big issue in developing countries. The current case-control study sought to determine the presence of high-risk human papillomaviruses (hr-HPV) in the development of cervical cancer, as well as their relationship with the cell cycle inhibitor gene p16INK4A in cervical cancer.MethodsThe association between p16INK4A protein and the presence of hr-HPV DNA in cervical lesions was explored in this study, which included 150 cervical cancer patients and 100 normal cervix samples. The immunohistochemistry approach was used to identify the expression of the p16INK4A protein, while the semi-quantitative polymerized chain reaction (PCR) method was used to identify the genomic identity of hr-HPV.ResultsAbout 90.67% (n=136) of the 150 case samples were found to be hr-HPV positive. Within the 136 HPV-positive samples, 45 (33.08%) show moderate expression of the p16INK4A protein, whereas 91 (66.91%) show overexpression, which is statistically significant (0.05). Among the 136 HPV-positive samples, 22.08% (N=30) were classified as having cervical intraepithelial neoplasia (CIN), with 56.66% (n=17) having CIN3, 36.66% (n=11) having CIN2, and 6.67% (n=2) having CIN1.ConclusionBased on the semi-quantitative immune staining scoring method of p16INK4A protein, genomic expression of HPV demonstrates that the expression of p16INK4A protein increases with the infectious load of the hr-HPV genome in the host cell. The result directly shows that immunostaining of the p16INK4A protein, in conjunction with the assessment of high-risk HPV in the host genome, will aid in the identification of cervical cancer in the cervix.

Stress-Induced Premature Senescence Related to Oxidative Stress in the Developmental Programming of Nonalcoholic Fatty Liver Disease in a Rat Model of Intrauterine Growth Restriction

Metabolic syndrome (MetS) refers to cardiometabolic risk factors, such as visceral obesity, dyslipidemia, hyperglycemia/insulin resistance, arterial hypertension and non-alcoholic fatty liver disease (NAFLD). Individuals born after intrauterine growth restriction (IUGR) are particularly at risk of developing metabolic/hepatic disorders later in life. Oxidative stress and cellular senescence have been associated with MetS and are observed in infants born following IUGR. However, whether these mechanisms could be particularly associated with the development of NAFLD in these individuals is still unknown. IUGR was induced in rats by a maternal low-protein diet during gestation versus. a control (CTRL) diet. In six-month-old offspring, we observed an increased visceral fat mass, glucose intolerance, and hepatic alterations (increased transaminase levels, triglyceride and neutral lipid deposit) in male rats with induced IUGR compared with the CTRL males; no differences were found in females. In IUGR male livers, we identified some markers of stress-induced premature senescence (SIPS) (lipofuscin deposit, increased protein expression of p21WAF, p16INK4a and Acp53, but decreased pRb/Rb ratio, foxo-1 and sirtuin-1 protein and mRNA expression) associated with oxidative stress (higher superoxide anion levels, DNA damages, decreased Cu/Zn SOD, increased catalase protein expression, increased nfe2 and decreased keap1 mRNA expression). Impaired lipogenesis pathways (decreased pAMPK/AMPK ratio, increased pAKT/AKT ratio, SREBP1 and PPARγ protein expression) were also observed in IUGR male livers. At birth, no differences were observed in liver histology, markers of SIPS and oxidative stress between CTRL and IUGR males. These data demonstrate that the livers of IUGR males at adulthood display SIPS and impaired liver structure and function related to oxidative stress and allow the identification of specific therapeutic strategies to limit or prevent adverse consequences of IUGR, particularly metabolic and hepatic disorders.

Manageable risk factors for progression of HPV-associated cervical intraepithelial neoplasia

Objective: to identify informative markers of the progression of cervical intraepithelial lesions of low degree.Materials and methods: the dynamics of the results of the cytological study of cervical epithelium, genotyping of human papilloma viruses of high carcinogenic risk (HPV) with an assessment of the viral load, an immunocytochemical study of the expression of protein p16 (p16INK4α), serum folic acid in 90 HPV-positive patients of reproductive age was analyzed. 43 of the 90 patients (47.8%) who made up group I had regression of the disease. Group II included 30 of 90 patients (33.3%) with disease persistence - no significant changes in the results of laboratory and instrumental research methods. Group III was 17 out of 90 patients (18.9%) with disease progression. Statistical processing of the results was carried out using parametric and non-parametric analysis methods using IBM SPSS Statistics 28.0.1.1 (developed by IBM Corporation), STATISTICA 13.5.0.17 (developed by StatSoft.Inc) and MedCalc 20.027. Results: factors that significantly distinguish patients with persistence or progression of low grade cervical intraepithelial lesions from women with regression of the disease are: earlier age of sexual onset, detection of atypical changes in low grade cervical epithelium, clinically significant HPV viral load, detection of p16INK4a protein expression, serum folic acid below 3.1 ng/ml. Conclusions: deficiency of serum folic acid in HPV of positive patients should be considered as a pathogenetically significant risk factor for the implementation of infection into the clinical form of the disease, which is confirmed by its correlation with the cytological detection of LSIL, progressive increase in the copy of HPV, and p16INKα expression. The patient management strategy, which provides a personalized assessment of the risk of progression of cervical intraepithelial lesions, expands the monitoring of patients with cervical neoplasias associated with HPV.

Altered p16Ink4a, IL-1β, and Lamin b1 Protein Expression Suggest Cellular Senescence in Deep Endometriotic Lesions.

Endometriosis causes immunological and cellular alterations. Endometriosis lesions have lower levels of lamin b1 than the endometrium. Moreover, high levels of pro-inflammatory markers are observed in the peritoneal fluid, follicular fluid, and serum in endometriosis lesions. Thus, we hypothesized that the accumulation of senescent cells in endometriosis tissues would facilitate endometriosis maintenance in an inflammatory microenvironment. To study senescent cell markers and the senescence-associated secretory phenotype (SASP) in endometriosis lesions, we conducted a cross-sectional study with 27 patients undergoing video laparoscopy for endometriosis resection and 19 patients without endometriosis. Endometriosis lesions were collected from patients with endometriosis, while eutopic endometrium was collected from patients both with and without endometriosis. Tissues were evaluated for senescence markers (p16Ink4a, lamin b1, and IL-1β) and interleukin concentrations. The expression of p16Ink4a increased in lesions compared to that in eutopic endometrium from endometriosis patients in the secretory phase. In the proliferative phase, lesions exhibited lower lamin b1 expression but higher IL-4 expression than the eutopic endometrium. Further, IL-1β levels were higher in the lesions than in the eutopic endometrium in both the secretory and proliferative phases. We believe that our findings may provide targets for better therapeutic interventions to alleviate the symptoms of endometriosis.

Generation of mice for evaluating endogenous p16Ink4a protein expression

The cyclin-dependent kinase inhibitor p16Ink4a plays a central role in cellular senescence in vitro. Although previous studies suggested cellular senescence is integrated in the systemic mechanisms of organismal aging, the localization and the dynamics of p16Ink4a in tissues remain poorly understood, which hinders uncovering the role of p16Ink4a under the in vivo context. One of the reasons is due to the lack of reliable reagents; as we also demonstrate here that commonly used antibodies raised against human p16INK4A barely recognize its murine ortholog. Here we generated a mouse model, in which the endogenous p16Ink4a is HA-tagged at its N-terminus, to explore the protein expression of p16Ink4a at the organismal level. p16Ink4a was induced at the protein level along the course of senescence in primary embryonic fibroblasts derived from the mice, consistently to its transcriptional level. Remarkably, however, p16Ink4a was not detected in the tissues of the mice exposed to pro-senescence conditions including genotoxic stress and activation of oncogenic signaling pathways, indicating that there is only subtle p16Ink4a proteins induced. These results in our mouse model highlight the need for caution in evaluating p16Ink4a protein expression in vivo.

Prognostic value of high-risk human papillomavirus DNA and p16INK4a immunohistochemistry in patients with anal cancer: An individual patient data meta-analysis

BackgroundHigh-risk human papillomavirus (hrHPV) types represent the aetiological agents in a major proportion of anal squamous cell carcinomas (ASCC). Several studies have suggested a prognostic relevance of HPV-related markers, particularly hrHPV DNA and p16INK4a (p16) protein expression, in patients with ASCC. However, broader evaluation of these prognostic marker candidates has been hampered by small cohort sizes and heterogeneous survival data among the individual studies. We conducted an individual patient data (IPD) meta-analysis to determine the prognostic value of hrHPV DNA and p16 in patients with ASCC while controlling for major clinical and tumour covariates. Patients and methodsA systematic literature search was conducted to identify all published studies analysing p16 alone or in combination with hrHPV DNA and reporting survival data in patients with ASCC. Clinical and tumour-related IPD were requested from authors of potentially eligible studies. Survival analyses were performed with a proportional hazard Cox model stratified by study and adjusted for relevant covariates. The study-specific hazard ratios (HRs) for the exposures were pooled using a random-effects model. Kaplan-Meier curves from different studies were pooled per exposure group and weighted by the study's total sample size. ResultsSeven studies providing IPD from 693 patients with ASCC could be included in the meta-analysis. Seventy-six percent of patients were p16+/hrHPV DNA+, whereas 11% were negative for both markers. A discordant marker status was observed in 13% of cases. Patients with p16+/hrHPV DNA+ ASCC showed significantly superior overall survival (OS) compared with patients with p16-/hrHPV DNA- tumours (pooled adjusted HR = 0.26 [95% confidence interval {CI}, 0.14–0.50]) with pooled three-year OS rates of 86% (95% CI, 82–90%) versus 39% (95% CI, 24–54%). Patients with discordant p16 and hrHPV DNA status showed intermediate three-year OS rates (75% [95% CI, 56–86%] for p16+/hrHPV DNA- and 55% [95% CI, 35–71%] for p16-/hrHPV DNA+ ASCC). ConclusionThis first IPD meta-analysis controlling for confounding variables shows that patients with p16+/hrHPV DNA+ ASCC have a significantly better survival than patients with p16-/hrHPV DNA- tumours.

Increased Placental Cell Senescence and Oxidative Stress in Women with Pre-Eclampsia and Normotensive Post-Term Pregnancies.

Up to 11% of pregnancies extend to post-term with adverse obstetric events linked to pregnancies over 42 weeks. Oxidative stress and senescence (cells stop growing and dividing by irreversibly arresting their cell cycle and gradually ageing) can result in diminished cell function. There are no detailed studies of placental cell senescence markers across a range of gestational ages, although increased levels have been linked to pre-eclampsia before full term. This study aimed to determine placental senescence and oxidative markers across a range of gestational ages in women with uncomplicated pregnancies and those with a diagnosis of pre-eclampsia. Placentae were obtained from 37 women with uncomplicated pregnancies of 37–42 weeks and from 13 cases of pre-eclampsia of 31+2–41+2 weeks. The expression of markers of senescence, oxidative stress, and antioxidant defence (tumour suppressor protein p16INK4a, kinase inhibitor p21, interleukin-6 (IL-6), NADPH oxidase 4 (NOX4), glutathione peroxidases 1, 3, and 4 (GPx1, GPx3, and GPx4), placental growth factor (PlGF), and soluble fms-like tyrosine kinase-1 (sFlt-1)) genes was measured (quantitative real-time PCR). Protein abundance of p16INK4a, IL-6, NOX4, 8-hydroxy-2′-deoxy-guanosine (8-OHdG), and PlGF was assessed by immunocytochemistry. Placental NOX4 protein was higher in post-term than term deliveries and further increased by pre-eclampsia (p < 0.05 for all). P21 expression was higher in post-term placentae (p = 0.012) and in pre-eclampsia (p = 0.04), compared to term. Placental P16INK4a protein expression was increased post-term, compared to term (p = 0.01). In normotensive women, gestational age at delivery was negatively associated with GPx4 and PlGF (mRNA and protein) (p < 0.05 for all), whereas a positive correlation was seen with placental P21, NOX4, and P16INK4a (p < 0.05 for all) expression. Markers of placental oxidative stress and senescence appear to increase as gestational age increases, with antioxidant defences diminishing concomitantly. These observations increase our understanding of placental health and may contribute to assessment of the optimal gestational age for delivery.

Ginsenoside Rg1 induces senescence of leukemic stem cells by upregulating p16INK4a and downregulating hTERT expression.

Leukemic stem cells (LSCs) play an important role in the pathogenesis of leukemia. This research attempted to clarify effects of the telomere system on ginsenoside Rg1-induced senescence of LSCs. This research attempted to clarify effects of the telomere system on ginsenoside Rg1-induced senescence of LSCs. CD34+CD38- LSCs were isolated, sorted, and divided into a control group and a Rg1 group (treated with 40 μmol/L Rg1). Cell Counting Kit-8 (CCK-8) was used to evaluate cell proliferation, and flow cytometry was used to assess the cell cycle of CD34+CD38- LSCs. The senescence-associated β-galactosidase (SA-β-Gal) staining and CFU-Mix assay were conducted to measure senescence of CD34+CD38- LSCs. The mRNA transcription and protein expression of p16INK4a and human telomerase reverse transcriptase (hTERT) were determined using a real-time polymerase chain reaction (RT-PCR) and western blot assay, respectively. The Rg1 treatment significantly attenuated proliferative activity and decreased the proliferative index (PI) of CD34+CD38- LSCs compared to those of the control group (p < 0.05). It remarkably increased positive SA-β-Gal staining rate, and suppressed formation of the CFU-Mix of CD34+CD38- LSCs compared with those of the control group (p < 0.05). The Rg1 treatment markedly boosted telomere effector, p16INK4a, in CD34+CD38- LSCs compared with that of control group (p < 0.05). Such treatment obviously reduced telomere regulator, hTERT, in CD34+CD38- LSCs compared with the control group (p < 0.05). Ginsenoside Rg1-induced senescence of CD34+CD38- LSCs through upregulating p16INK4a and downregulating hTERT expression, both of which are associated with telomere systems. The present study would be beneficial for the treatment of leukemia by providing a promising strategy to induce senescence of CD34+CD38- LSCs.

Cellular senescence as a driver of cognitive decline triggered by chronic unpredictable stress

When an individual is under stress, the undesired effect on the brain often exceeds expectations. Additionally, when stress persists for a long time, it can trigger serious health problems, particularly depression. Recent studies have revealed that depressed patients have a higher rate of brain aging than healthy subjects and that depression increases dementia risk later in life. However, it remains unknown which factors are involved in brain aging triggered by chronic stress. The most critical change during brain aging is the decline in cognitive function. In addition, cellular senescence is a stable state of cell cycle arrest that occurs because of damage and/or stress and is considered a sign of aging. We used the chronic unpredictable stress (CUS) model to mimic stressful life situations and found that, compared with nonstressed control mice, CUS-treated C57BL/6 mice exhibited depression-like behaviors and cognitive decline. Additionally, the protein expression of the senescence marker p16INK4a was increased in the hippocampus, and senescence-associated β-galactosidase (SA-β-gal)-positive cells were found in the hippocampal dentate gyrus (DG) in CUS-treated mice. Furthermore, the levels of SA-β-gal or p16INK4a were strongly correlated with the severity of memory impairment in CUS-treated mice, whereas clearing senescent cells using the pharmacological senolytic cocktail dasatinib plus quercetin (D + Q) alleviated CUS-induced cognitive deficits, suggesting that targeting senescent cells may be a promising candidate approach to study chronic stress-induced cognitive decline. Our findings open new avenues for stress-related research and provide new insight into the association of chronic stress-induced cellular senescence with cognitive deficits.