Levels Of Telomerase Activity Research Articles

Telomerase activity in breast cancer patients: association with poor prognosis and more aggressive phenotype.

Telomerase expression is an important mechanism of tumor unlimited replicative potential. The aim of this study was to evaluate prognostic impact of telomerase activity in breast cancer patients and to correlate telomerase activity with established prognostic factors. We analyzed tissue of 102 malignant breast lesions and 20 healthy breast tissues. Telomerase activity was determined by telomeric repeat amplification protocol assay. Telomerase activity was present in 77 (75.49%) of 102 breast cancers. Telomerase activity in breast cancers was statistically significantly higher in comparison with the activity in normal breast tissue. The levels of telomerase activity were significantly positively correlated with tumor size, axillary nodal status, histological grade, HER-2/neu protein expression in tumor tissue and expression of the nuclear antigen Ki-67. A statistically significant negative correlation was found between the presence of ER and telomerase activity. There was no correlation between telomerase activity and concentration of PR or the age of patients. Kaplan-Meier analysis showed that patients with higher telomerase activity had significantly shorter 10-year disease-free survival (p<0.0001) and 10-year overall survival (p<0.0001) than those with lower telomerase activity. These results were confirmed by logistic regression analysis. Our results support the prognostic role of telomerase activity and its relationship with the more aggressive phenotype of breast cancer.

Telomerase activity in non-small cell lung cancer.

IntroductionHigh telomerase activity has been detected in the majority of malignant neoplasms including lung cancer. The purpose of the study was to attempt to use telomerase activity as a prognostic factor in patients with non-small cell lung cancer (NSCLC).Material and methodsTelomerase activity was analyzed in 47 tissue specimens taken from patients with NSCLC. The control group consisted of 30 specimens of non-cancerous lung parenchyma. Telomerase activity was measured by means of the telomeric repeat amplification protocol (TRAP).ResultsTelomerase activity in the neoplastic tissue was significantly higher than in the lung parenchyma that was free from neoplastic infiltration. There was no significant association between telomerase activity and age, gender, tobacco smoking, histological type of the tumor, or staging (pTNM). No association was found between the level of telomerase activity in NSCLC specimens and the two-year survival rate of patients (p = 0.326). A higher level of telomerase activity in poorly differentiated tumors (G3) as compared to moderately differentiated tumors (G2) was detected (p = 0.008). A positive association was identified between telomerase activity in pulmonary parenchyma free from tumor infiltration and the presence of leukocyte infiltration (p = 0.0001).ConclusionsNo association was found between the level of telomerase activity in NSCLC specimens and the two-year survival rate of patients. The study has revealed a positive association between telomerase activity and the grade of differentiation (G) in NSCLC.

Abstract B1-46: How collection, processing and storage of PBL samples impact the measurement of telomerase activity

Abstract Introduction: Activation of telomerase and telomere lengthening in patient peripheral blood leukocytes (PBL) is associated with longer survival in cancer patients. There are few studies to date that have measured telomerase activity in clinical PBL samples, and it is unclear how initial collection and processing of PBL samples impact the measurement of telomerase activity. In this study, we assessed if telomerase activity measurement varies based on the method of blood collection, processing and storage. Methods: We first collected whole blood drawn in EDTA tubes from two healthy patients that were then processed three different ways. The first whole blood sample was spun down and the buffy coat was pipetted from the top of the red blood cells, and then placed directly into the -80°C freezer. The next two whole blood samples were isolated using an ACCUSPIN™ tube, which utilizes density gradient separation to recover the PBL without contamination of red blood cells. The resulting PBL from these two samples were suspended in freezing media containing RPMI and DMSO, with one sample also containing penicillin streptomycin glutamine, and then both samples were stored in liquid nitrogen following a slow freeze. We then measured enzymatic telomerase activity in each of the three PBL samples from the two different patients. Results: We detected a comparable level of telomerase activity in all three PBL samples. These findings indicate that telomerase activity can be measured in buffy coat samples from patients independent of how the samples were processed and stored. Conclusions: These findings are important because many biorepositories may not have collected or processed PBL samples by identical processes, therefore not allowing direct comparison across samples. Additionally, our finding that telomerase activity levels are comparable between PBL samples containing red blood cells and those processed using an ACCUSPIN™ tube is valuable because it eliminates the necessity for the extra cost of the ACCUSPIN™ tube, freezing media, antibiotics and use of liquid nitrogen storage tanks. Future studies will focus on the determining the relationship of telomerase activity between the PBL, normal colon and cancer tissue in colorectal cancer patients. Citation Format: Ruth A. Johnson, Brooke R. Druliner, Jill Washechek Alleto, Donna Felmlee Devine, Xiaoyang Ruan, Russell Vanderboom, Lisa A. Boardman. How collection, processing and storage of PBL samples impact the measurement of telomerase activity. [abstract]. In: Proceedings of the AACR Special Conference on Computational and Systems Biology of Cancer; Feb 8-11 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 2):Abstract nr B1-46.

Activity of telomerase and telomeric length in Apis mellifera.

Telomerase is an enzyme that adds repeats of DNA sequences to the ends of chromosomes, thereby preventing their shortening. Telomerase activity is associated with proliferative status of cells, organismal development, and aging. We report an analysis of telomerase activity and telomere length in the honeybee, Apis mellifera. Telomerase activity was found to be regulated in a development and caste-specific manner. During the development of somatic tissues of larval drones and workers, telomerase activity declined to 10% of its level in embryos and remained low during pupal and adult stages but was upregulated in testes of late pupae, where it reached 70% of the embryo level. Upregulation of telomerase activity was observed in the ovaries of late pupal queens, reaching 160% of the level in embryos. Compared to workers and drones, queens displayed higher levels of telomerase activity. In the third larval instar of queens, telomerase activity reached the embryo level, and an enormous increase was observed in adult brains of queens, showing a 70-fold increase compared to a brain of an adult worker. Southern hybridization of terminal TTAGG fragments revealed a high variability of telomeric length between different individuals, although the same pattern of hybridization signals was observed in different tissues of each individual.

Telomerase Activity and Telomere Length in Human Benign Prostatic Hyperplasia Stem-like Cells and Their Progeny Implies the Existence of Distinct Basal and Luminal Cell Lineages

Benign prostatic hyperplasia (BPH) treatments have changed little over many years and do not directly address the underlying cause. Because BPH is characterised by uncontrolled cell growth, the chromosomal telomeres should be eroded in the reported absence or low levels of telomerase activity, but this is not observed. We investigated the telomere biology of cell subpopulations from BPH patients undergoing transurethral resection of prostate (TURP). Measurement of TERC, TERT, and telomerase activity revealed that only the epithelial stem-like and progenitor fractions expressed high levels of telomerase activity (p<0.01) and individual enzyme components (p<0.01). Telomerase activity and TERT expression were not detected in stromal cells. Telomere length measurements reflected this activity, although the average telomere length of (telomerase-negative) luminal cells was equivalent to that of telomerase-expressing stem/progenitor cells. Immunohistochemical analysis of patient-derived BPH arrays identified distinct areas of luminal hyperproliferation, basal hyperproliferation, and basal–luminal hyperproliferation, suggesting that basal and luminal cells can proliferate independently of each other. We propose a separate lineage for the luminal and basal cell components in BPH. Patient summaryWe unexpectedly found an enzyme called telomerase in the cells that maintain benign prostatic hyperplasia (BPH), suggesting that telomerase inhibitors could be used to alleviate BPH symptoms.

Comparisons of Development Potential in Bovine SCNT Embryos using Donor Cells treated with Different Demethylating Inhibitors

To improve the developmental potential of bovine somatic cell nuclear transfer (SCNT) embryos, this study compared the developmental rates to blastocyst stage in the SCNT embryos using donor fibroblasts treated with 5-azacytidine (5AC) and S-adenosylhomocysteine (SAH) at different concentrations. Their reprogramming efficiency level was investigated with level of telomerase activity. Donor fibroblasts isolated from adult ear skin of a cow were exposed to 5AC and SAH at different concentrations during 2 passages. After nuclear transfer into enucleated recipient oocytes, the cleavage and developmental rates were significantly (p<0.05) decreased in the SCNT embryos using 5AC-treated fibroblasts (5AC-SCNT embryos), compared with those of non-treated control (control-SCNT embryos) and SAH-treated fibroblasts (SAH-SCNT embryos). The developmental rates to blastocyst stage tended to be slightly increased in the SAH-SCNT embryos at each of the concentrations, and especially, the developmental rates in the SCNT embryos using 1.0 mM SAH-treated fibroblasts were significantly (p<0.05) higher than that of control SCNT embryos. The mean numbers of total and ICM cell in blastocysts were also significantly (p<0.05) decreased in the 5AC-SCNT embryos, compared with those of other SCNT blastocysts. Further, the level of telomerase activity was also significantly (p< 0.05) decreased in the 5AC-SCNT embryos than those of control and SAH-SCNT embryos. Whereas, a significantly (p<0.05) up-regulated telomerase activity was observed in SAH-SCNT embryos, compare with that of control-SCNT embryos. In conclusion, SCNT embryos using hypomethylated donor cells with SAH, not 5AC, may improve the developmental potential and reprogramming efficiency.

Inhibition of cell growth and down-regulation of telomerase activity by amygdalin in human cancer cell lines

The purpose of this study was to examine the effect of amygdalin on cell growth and telomerase activity in human cancer and MRC-5 fibroblast cell lines. The level of β-glucosidase activity for releasing cyanide was significantly (P < .05) higher in cancer cell lines (A-549, MDA-MB-231, MCF-7 and U87-MG) than in MRC-5 fibroblasts. Growth rate of cancer cells was apparently inhibited in concentrations above 10 mg/ml amygdalin with senescent-like abnormal morphology. Whereas the effects were absent or marginally detected in MRC-5 fibroblasts. High incidence of β-galactosidase activity was observed in amygdalin-treated cancer cells, compared with that of untreated control while no difference was observed between the control and amygdalin-treated MRC-5 fibroblasts. Furthermore, level of telomerase activity was significantly (P < .05) higher (∼8–13 fold) in cancer cell lines along with high expression of telomerase reverse transcriptase (TERT) and telomerase RNA component (TERC) than in MRC-5 fibroblasts which did not expressed TERT and TERC. However, telomerase activity was significantly (P < .05) down-regulated in amygdalin-treated cancer cells with the decreased expression of TERT and TERC compared with control cancer cells. There were no difference in the telomerase activity between control and amygdalin-treated MRC-5 fibroblasts. Based on these observations, we concluded that amygdalin treatment offers a valuable option for the cancer treatment, causing inhibition of cell growth and down-regulation of telomerase activity in human cancer cell lines by increasing β-glucosidase activity.

Telomerase inhibition improves tumor response to radiotherapy in a murine orthotopic model of human glioblastoma.

BackgroundGlioblastoma (GBM) is the most frequent and aggressive type of adult brain tumor. Most GBMs express telomerase; a high level of intra-tumoral telomerase activity (TA) is predictive of poor prognosis. Thus, telomerase inhibitors are promising options to treat GBM. These inhibitors increase the response to radiotherapy (RT), in vitro as well as in vivo. Since typical treatments for GBM include RT, our objective was to evaluate the efficiency of Imetelstat (TA inhibitor) combined with RT.FindingsWe used a murine orthotopic model of human GBM (N = 8 to11 mice per group) and μMRI imaging to evaluate the efficacy of Imetelstat (delivered by intra-peritoneal injection) alone and combined with RT. Using a clinically established protocol, we demonstrated that Imetelstat significantly: (i) inhibited the TA in the very center of the tumor, (ii) reduced tumor volume as a proportion of TA inhibition, and (iii) increased the response to RT, in terms of tumor volume regression and survival increase.ConclusionsImetelstat is currently evaluated in refractory brain tumors in young patients (without RT). Our results support its clinical evaluation combined with RT to treat GBM.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0376-3) contains supplementary material, which is available to authorized users.

Anti-aging effect of estrogen on telomerase activity in ovariectomised rats – animal model for menopause

Objective: The aim of this study was to investigate the anti-aging effects of exogenous estrogen on telomerase activity in ovariectomized female Sprague-Dawley rats.Methods: Thirty-three 12-week-old female rats were divided into three groups: the ovariectomized-Treated group (Treated, n = 11), the ovariectomized control group (OVX, n = 11) and the Sham-operated group (Sham, n = 11). The rats in the Treated group were given 0.21 mg/kg estradiol valerate intragastric administration while other two groups were given the amount of physiological saline daily. All of the animals were euthanized 12 weeks after treatment, and abdominal aortic blood samples were taken to assess the level of estradiol (E2), follicle stimulating hormone (FSH). Telomerase activity and telomerase reverse transcriptase (TERT) mRNA expression in the heart, liver, brain tissues of all rats were measured by reverse transcriptional polymerase chain reaction (RT-PCR) and competitive enzyme-linked immunosorbent assay (ELISA).Results: Compared to the OVX and Sham group, telomerase activity and TERT mRNA levels were significantly increased in the heart, liver and brain tissues of rats in the Treated group (p < 0.05). The telomerase expression was significantly higher in the heart than in liver and brain tissues in the Treated group (p < 0.05).Conclusion: Based on the findings in the ovariectomized female rats model, exogenous estrogen can significantly up-regulate telomerase activity and TERT mRNA expression to exert the effects of anti-aging.

Angelica sinensis polysaccharides inhibit endothelial progenitor cell senescence through the reduction of oxidative stress and activation of the Akt/hTERT pathway

Context: Angelica sinensis (Oliv.) Diels (Apiaceae) polysaccharides (ASP) may play a key role in anti-ischemic activity. However, the anti-atherosclerotic activity and mechanism are unknown.Objective: This study investigated the protective effects of ASP against ox-LDL-induced senescence of EPCs and explored its underlying molecular mechanisms.Materials and methods: Mononuclear cells were isolated from bone marrow (BM) of SD rats and differentiated to EPCs. EPCs were exposed to oxidized low-density lipoprotein (ox-LDL, 10 µg/mL, 24 h) and incubated with or without high-dose (100 µg/mL, 48 h) or low-dose (20 µg/mL, 48 h) ASP. Another group of EPCs was pre-treated with Wortmannin (100 nM, 45 min), a PI3K/Akt inhibitor. EPC senescence, telomerase activity, and superoxide anion levels were assessed using SA-β-galactosidase staining, telomerase PCR-ELISA analysis, and DHE staining, respectively. The expression of related proteins, including Akt, p-Akt, hTERT, p-hTERT, and gp91phox, were detected using western blot.Results: EPCs (47.3%) were SA-β-gal positive after treatment by ox-LDL, additionally, ox-LDL significantly increased superoxide anion levels (375% versus 100%), and inhibited telomerase activity (42% versus 100%). However, the pro-senescent effect of ox-LDL was attenuated about three-fold (16.7%), superoxide anion levels were decreased more than two-fold (148%), and telomerase activity was recovered partly (88% versus 42%) in the EPCs when treated with ASP (100 µg/mL). The immunoblotting confirmed that ASP attenuated inhibition of phosphorylation of Akt and hTERT induced by ox-LDL and down-regulated increased the expression of gp91-phox. Moreover, some effects of ASP were partially abrogated in the presence of Wortmannin.Discussion: Ox-LDL induced senescence of EPCs via inhibition of telomerase activity, which was influenced by oxidative stress and the Akt/hTERT pathway. The inhibition of EPC senescence by ASP could be important for potential therapeutics.Conclusion: Treatment of EPCs with ASP remarkably attenuates the harmful effects of ox-LDL via augmentation of Akt/hTERT phosphorylation and inhibition of oxidative stress.

Widespread telomere instability in prostatic lesions.

A critical function of the telomere is to disguise chromosome ends from cellular recognition as double strand breaks, thereby preventing aberrant chromosome fusion events. Such chromosome end-to-end fusions are known to initiate genomic instability via breakage-fusion-bridge cycles. Telomere dysfunction and other forms of genomic assault likely result in misregulation of genes involved in growth control, cell death, and senescence pathways, lowering the threshold to malignancy and likely drive disease progression. Shortened telomeres and anaphase bridges have been reported in a wide variety of early precursor and malignant cancer lesions including those of the prostate. These findings are being extended using methods for the analysis of telomere fusions (decisive genetic markers for telomere dysfunction) specifically within human tissue DNA. Here we report that benign prostatic hyperplasia (BPH), high-grade prostatic intraepithelial neoplasia (PIN), and prostate cancer (PCa) prostate lesions all contain similarly high frequencies of telomere fusions and anaphase bridges. Tumor-adjacent, histologically normal prostate tissue generally did not contain telomere fusions or anaphase bridges as compared to matched PCa tissues. However, we found relatively high levels of telomerase activity in this histologically normal tumor-adjacent tissue that was reduced but closely correlated with telomerase levels in corresponding PCa samples. Thus, we present evidence of high levels of telomere dysfunction in BPH, an established early precursor (PIN) and prostate cancer lesions but not generally in tumor adjacent normal tissue. Our results suggest that telomere dysfunction may be a common gateway event leading to genomic instability in prostate tumorigenesis. .

Dose-dependent cytotoxic effects of boldine in HepG-2 cells-telomerase inhibition and apoptosis induction.

Plant metabolites are valuable sources of novel therapeutic compounds. In an anti-telomerase screening study of plant secondary metabolites, the aporphine alkaloid boldine (1,10-dimethoxy-2,9-dihydroxyaporphine) exhibited a dose and time dependent cytotoxicity against hepatocarcinoma HepG-2 cells. Here we focus on the modes and mechanisms of the growth-limiting effects of this compound. Telomerase activity and expression level of some related genes were estimated by real-time PCR. Modes of cell death also were examined by microscopic inspection, staining methods and by evaluating the expression level of some critically relevant genes. The growth inhibition was correlated with down-regulation of the catalytic subunit of telomerase (hTERT) gene (p < 0.01) and the corresponding reduction of telomerase activity in sub-cytotoxic concentrations of boldine (p < 0.002). However, various modes of cell death were stimulated, depending on the concentration of boldine. Very low concentrations of boldine over a few passages resulted in an accumulation of senescent cells so that HepG-2 cells lost their immortality. Moreover, boldine induced apoptosis concomitantly with increasing the expression of bax/bcl2 (p < 0.02) and p21 (p < 0.01) genes. Boldine might thus be an interesting candidate as a potential natural compound that suppresses telomerase activity in non-toxic concentrations.

Is there a connection between inflammation, telomerase activity and the transcriptional status of telomerase reverse transcriptase in renal failure?

Telomerase is involved in the elongation of telomeres. It remains active in very few types of cell in mature organisms. One such cell type is the lymphocytes. In this study, we investigated the activity and expression of telomerase in lymphocytes from renal failure patients and compared it to that for normal controls. Inflammation status was determined at the same time. The enzyme activity was measured using PCR-ELISA with peripheral blood mononuclear cells (PBMCs) from three groups: 53 healthy individuals, 50 patients with chronic kidney disease (CKD) and 50 dialysis patients. In the same cell populations, the expression of the reverse transcriptase of the human telomerase gene (hTERT) was measured via real-time PCR. The inflammationstatus of these individuals was determined by calculating the interleukin 6 (IL-6), IL-10, C-reactive protein (CRP) and tumor necrosis factor alpha (TNF-a) serum concentrations via ELISA. The lowest levels of telomerase activity were detected in CKD, and this group had the highest IL-6 and CRP values and the lowest hTERT expression. The dialysis group showed significant differences in comparison to the normal subjects and to the CKD patients. Further studies are warranted in order to explore the way inflammation influences telomerase activity and hTERT expression.

Impact of chemotherapy on telomere length in sporadic and familial breast cancer patients.

Recently, we observed that telomeres of BRCA1/2 mutation carriers were shorter than those of controls or sporadic breast cancer patients, suggesting that mutations in these genes might be responsible for this event. Given the contradictory results reported in the literature, we tested whether other parameters, such as chemotherapy, could be modifying telomere length (TL). We performed a cross-sectional study measuring leukocyte TL of 266 sporadic breasts cancer patients treated with first-line chemotherapy, with a median follow-up of 240 days. Additionally, we performed both cross-sectional and longitudinal studies in a series of 236 familial breast cancer patients that included affected and non-affected BRCA1/2 mutation carriers. We have measured in leukocytes from peripheral blood: the TL, percentage of short telomeres (<3 kb), telomerase activity levels and the annual telomere shortening speed. In sporadic cases we found that chemotherapy exerts a transient telomere shortening effect (around 2 years) that varies depending on the drug combination. In familial cases, only patients receiving treatment were associated with telomere shortening but they recovered normal TL after a period of 2 years. Chemotherapy affects TL and should be considered in the studies that correlate TL with disease susceptibility.

Downregulation of Dyskerin Impairs Expansion of Normal Hematopoietic Progenitors and Erythroid Colony Formation in the Absence of Apparent Telomere Shortening

Dyskerin is an RNA binding protein that functions as a core component of the telomerase holoenzyme as well as in ribonuclear protein complexes involved in ribosome biogenesis and RNA processing. Within the telomerase holoenzyme, dyskerin stabilizes the telomerase RNA component (hTR) that functions as a template for synthesis of telomeric DNA at chromosome ends. Telomerase is active in malignant cells as well as normal hematopoietic stem and progenitor cells where it enables long-term cell replication.Mutation of the gene encoding dyskerin (DKC1) is the underlying cause of the inherited disease dyskeratosis congenita (DC), which is characterized by low levels of telomerase activity and shortened telomeres. Bone marrow failure is the most common cause of mortality in DC, however since dyskerin has telomerase-independent roles in fundamental cellular processes, it is not clear whether telomerase insufficiency is the sole cause of hematopoietic dysfunction in this disorder.To gain further insight to the consequences of deregulated expression of dyskerin in haematopoietic cells, we transduced normal human cord blood CD34+ progenitor cells with retroviral vectors encoding two different shRNAs targeting dyskerin. Transduced cells from seven independent experiments were FACS sorted then assayed for expansion and differentiation in vitro, clonogenic potential, telomerase activity and telomere length maintenance. Results from the functional assays showed that depletion of dyskerin impaired cell expansion and compromised the formation of BFU-E colonies. In contrast to BFU-E, CFU-GM colonies did not appear to be altered by dyskerin depletion. Analysis of non-transuduced cells subject to ex vivo expansion along erythroid and myelomonocytic lineages revealed that erythroid precursor cells expressed disproportionately high levels of dyskerin, and correspondingly high telomerase enzyme activity. shRNA-mediated downregulation of dyskerin lowered hTR levels and quenched telomerase activity in the erythroid precursors without altering expression of the telomerase catalytic component, hTERT. These results suggest that upregulation of dyskerin bolsters telomerase activity and renders erythroid precursors vulnerable to dyskerin depletion. However in spite of the reduction in telomerase activity induced by dyskerin shRNA, Southern blot analysis showed that the rate of telomere length shortening was not significantly altered by dyskerin depletion over the time course of these experiments. Together the data show that dyskerin is upregulated during erythroid differentiation, and has an essential function in the erythroid lineage that appears to be independent of its role in telomere length maintenance. DisclosuresNo relevant conflicts of interest to declare.

Human cells lacking coilin and Cajal bodies are proficient in telomerase assembly, trafficking and telomere maintenance.

The RNA component of human telomerase (hTR) localizes to Cajal bodies, and it has been proposed that Cajal bodies play a role in the assembly of telomerase holoenzyme and telomerase trafficking. Here, the role of Cajal bodies was examined in Human cells deficient of coilin (i.e. coilin-knockout (KO) cells), in which no Cajal bodies are detected. In coilin-KO cells, a normal level of telomerase activity is detected and interactions between core factors of holoenzyme are preserved, indicating that telomerase assembly occurs in the absence of Cajal bodies. Moreover, dispersed hTR aggregates and forms foci specifically during S and G2 phase in coilin-KO cells. Colocalization of these hTR foci with telomeres implies proper telomerase trafficking, independent of Cajal bodies. Therefore, telomerase adds similar numbers of TTAGGG repeats to telomeres in coilin-KO and controls cells. Overexpression of TPP1-OB-fold blocks cell cycle-dependent formation of hTR foci and inhibits telomere extension. These findings suggest that telomerase assembly, trafficking and extension occur with normal efficiency in Cajal bodies deficient human cells. Thus, Cajal bodies, as such, are not essential in these processes, although it remains possible that non-coilin components of Cajal bodies and/or telomere binding proteins (e.g. TPP1) do play roles in telomerase biogenesis and telomere homeostasis.

Predicting clinical outcome in feline oral squamous cell carcinoma: tumour initiating cells, telomeres and telomerase.

Feline oral squamous cell carcinoma (SCC) has very poor prognosis. Here, a retrospective pilot study was conducted on 20 feline oral SCC patients who underwent stereotactic radiation therapy (SRT), to evaluate: (1) the value of putative tumour initiating cell (TIC) markers of human head and neck SCC (CD44, Bmi-1); (2) telomere length (TL) specifically in putative TICs; and (3) tumour relative telomerase activity (TA). Significant inverse correlations were found between treatment outcomes and Bmi-1 expression, supporting the predictive value of Bmi-1 as a negative prognostic indicator. While TL exhibited a wide range of variability, particularly in very short fractions, many tumours possessed high levels of TA, which correlated with high levels of Bmi-1, Ki67 and EGFR. Taken together, our results imply that Bmi-1 and telomerase may represent novel therapeutic targets in feline oral SCC, as their inhibition-in combination with SRT-would be expected to have beneficial treatment outcome.

Effect of a qigong intervention program on telomerase activity and psychological stress in abused Chinese women: a randomized, wait-list controlled trial

BackgroundAbused women, who suffer from chronic psychological stress, have been shown to have shorter telomeres than never abused women. Telomere shortening is associated with increased risk of cell death, and it is believed that adopting health-promoting behaviors can help to increase the activity of telomerase, an enzyme that counters telomere shortening. Qigong is an ancient Chinese mind-body integration, health-oriented practice designed to enhance the function of qi, an energy that sustains well-being. Therefore, an assessor-blind, randomized, wait-list controlled trial was developed to evaluate the effect of a qigong intervention on telomerase activity (primary objective) and proinflammatory cytokines, perceived stress, perceived coping, and depressive symptoms (secondary objectives) in abused Chinese women.Methods/DesignA total of 240 Chinese women, aged ≥18 years, who have been abused by an intimate partner within the past three years will be recruited from a community setting in Hong Kong and randomized to receive either a qigong intervention or wait-list control condition as follows: the qigong intervention will comprise (i) a 2-hour group qigong training session twice a week for 6 weeks, (ii) a 1-hour follow-up group qigong exercise session once a week for 4 months, and (iii) a 30-minute self-practice qigong exercise session once a day for 5.5 months. The wait-list control group will receive qigong training after the intervention group completes the program. Upon completion of the qigong intervention program, it is expected that abused Chinese women in the intervention group will have higher levels of telomerase activity and perceived coping and lower levels of proinflammatory cytokines, perceived stress, and depressive symptoms than will abused Chinese women in the wait-list control group.DiscussionThis study will provide information about the effect of qigong exercise on telomerase activity and chronic psychological stress in abused Chinese women. The findings will inform the design of interventions to relieve the effects of IPV-related psychological stress on health. Also, the concept that health-promoting behaviors could slow down cellular aging might even motivate abused women to change their lifestyles.Trial registrationCurrent Controlled Trials NCT02060123. Registered February 6, 2014.

A short-term treatment with tumor necrosis factor-alpha enhances stem cell phenotype of human dental pulp cells

IntroductionDuring normal pulp tissue healing, inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) or interleukins, act in the initial 48 hours (inflammatory phase) and play important roles not only as chemo-attractants of inflammatory cells and stem/progenitor cells but also in inducing a cascade of reactions toward tissue regeneration or reparative dentin formation or both. Previous reports have shown that inflammatory cytokines regulate the differentiation capacity of dental pulp stem/progenitor cells (DPCs), but none has interrogated the impact of these cytokines on the stem cell phenotype of stem/progenitor cells. This study investigated the effects of a short-term treatment with TNF-α on the stem cell phenotype and differentiation ability of human DPCs.MethodsAn in vivo mouse model of pulp exposure was performed for analysis of expression of the mesenchymal stem cell marker CD146 in DPCs during the initial stage of inflammatory response. For in vitro studies, human DPCs were isolated and incubated with TNF-α for 2 days and passaged to eliminate TNF-α completely. Analysis of stem cell phenotype was performed by quantification of cells positive for mesenchymal stem cell markers SSEA-4 (stage-specific embryonic antigen 4) and CD146 by flow cytometry as well as by quantitative analysis of telomerase activity and mRNA levels of OCT-4 and NANOG. Cell migration, colony-forming ability, and differentiation toward odontogenesis and adipogenesis were also investigated.ResultsThe pulp exposure model revealed a strong staining for CD146 during the initial inflammatory response, at 2 days after pulp exposure. In vitro experiments demonstrated that a short-term (2-day) treatment of TNF-α increased by twofold the percentage of SSEA-4+ cells. Accordingly, STRO-1, CD146, and SSEA-4 protein levels as well as OCT-4 and NANOG mRNA levels were also significantly upregulated upon TNF-α treatment. A short-term TNF-α treatment also enhanced DPC function, including the ability to form cell colonies, to migrate, and to differentiate into odontogenic and adipogenic lineages.ConclusionsA short-term treatment with TNF-α enhanced the stem cell phenotype, migration, and differentiation ability of DPCs.

Poly (ADP-ribose) polymerase 3 (PARP3), a potential repressor of telomerase activity

BackgroundConsidering previous result in Non-Small Cell Lung Cancer (NSCLC), we investigated in human cancer cells the role of PARP3 in the regulation of telomerase activity.MethodsWe selected A549 (lung adenocarcinoma cell line) and Saos-2 (osteosarcoma cell line), with high and low telomerase activity levels, respectively. The first one was transfected using a plasmid construction containing a PARP3 sequence, whereas the Saos-2 cells were submitted to shRNA transfection to get PARP3 depletion. PARP3 expression on both cell systems was evaluated by real-time quantitative PCR and PARP3 protein levels, by Western-blot. Telomerase activity was determined by TRAP assay.ResultsIn A549 cells, after PARP3 transient transfection, data obtained indicated that twenty-four hours after transfection, up to 100-fold increased gene expression levels were found in the transfected cells with pcDNA/GW-53/PARP3 in comparison to transfected cells with the empty vector. Moreover, 48 hours post-transfection, telomerase activity decreased around 33%, and around 27%, 96 hours post-transfection. Telomerase activity average ratio was 0.67 ± 0.05, and 0.73 ± 0.06, respectively, with significant differences. In Saos-2 cells, after shRNA-mediated PARP3 silencing, a 2.3-fold increase in telomerase activity was detected in relation to the control.ConclusionOur data indicated that, at least in some cancer cells, repression of PARP3 could be responsible for an increased telomerase activity, this fact contributing to telomere maintenance and, therefore, avoiding genome instability.